anti-infective activities of lactobacillus strains in the ... · minally and intracellularly in the...

Anti-Infective Activities of Lactobacillus Strains in the HumanIntestinal Microbiota: from Probiotics to Gastrointestinal Anti-Infectious Biotherapeutic Agents

Vanessa Liévin-Le Moal,a,b,c Alain L. Servina,b,c

CNRS, UMR 8076 BioCIS, Faculty of Pharmacy, Châtenay-Malabry, Francea; Laboratory of Excellence in Research on Medication and Innovative Therapeutics, Châtenay-Malabry, Franceb; University Paris-Sud, Faculty of Pharmacy, Châtenay-Malabry, Francec

SUMMARY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .167INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .168GASTROINTESTINAL MICROBIOTA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .168PROBIOTIC LACTOBACILLUS STRAINS ISOLATED FROM THE HUMAN INTESTINAL MICROBIOTA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .170IN VITRO ANTIBACTERIAL ACTIVITIES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .171

Direct Activities against Enterovirulent Bacteria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .171Bacteriostatic activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .171Bactericidal activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .171Activity on the expression or functionality of virulence factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .175

Direct Activities against H. pylori . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .175Bactericidal activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .175Activity against the expression and functionality of virulence factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .175

ACTIVITIES AGAINST THE DELETERIOUS EFFECTS INDUCED BY INFECTIOUS AGENTS AT THE INTESTINAL EPITHELIAL BARRIER . . . . . . . . . . . . . . . . . . . . .176Activities against Enterovirulent Bacteria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .176

Effects at the brush border . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .176Effects at the epithelial junctional domain. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .177Activation of host epithelial defense responses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .177

Effects against Rotavirus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .179Effects against H. pylori . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .179

ACTIVITIES IN ANIMAL INFECTION MODELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .179Bacterium-Infected Animals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .179Rotavirus-Infected Animals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .181

CLINICAL STUDIES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .181Therapeutic Effects against Various Forms of Acute Diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .181

Acute infectious diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .184Nosocomial infections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .184Traveler’s diarrhea. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .184

Therapeutic Effects against C. difficile-Associated Diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .185Meta-Analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .185Therapeutic Effects against Infectious Gastritis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .185

CONCLUDING REMARKS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .186ACKNOWLEDGMENTS. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .187REFERENCES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .187AUTHOR BIOS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .199

SUMMARY

A vast and diverse array of microbial species displaying great phy-logenic, genomic, and metabolic diversity have colonized the gas-trointestinal tract. Resident microbes play a beneficial role by reg-ulating the intestinal immune system, stimulating the maturationof host tissues, and playing a variety of roles in nutrition and inhost resistance to gastric and enteric bacterial pathogens. Themechanisms by which the resident microbial species combat gas-trointestinal pathogens are complex and include competitive met-abolic interactions and the production of antimicrobial mole-cules. The human intestinal microbiota is a source from whichLactobacillus probiotic strains have often been isolated. Only sixprobiotic Lactobacillus strains isolated from human intestinal mi-crobiota, i.e., L. rhamnosus GG, L. casei Shirota YIT9029, L. caseiDN-114 001, L. johnsonii NCC 533, L. acidophilus LB, and L. reu-teri DSM 17938, have been well characterized with regard to their

potential antimicrobial effects against the major gastric and en-teric bacterial pathogens and rotavirus. In this review, we describethe current knowledge concerning the experimental antibacterialactivities, including antibiotic-like and cell-regulating activities,and therapeutic effects demonstrated in well-conducted, placebo-controlled, randomized clinical trials of these probiotic Lactoba-cillus strains. What is known about the antimicrobial activitiessupported by the molecules secreted by such probiotic Lactobacil-

Address correspondence to Alain L. Servin, [email protected].

This article is dedicated to the memory of Jean-Richard Neeser (Nestec ResearchCenter, Lausanne, Switzerland).

Copyright © 2014, American Society for Microbiology. All Rights Reserved.

doi:10.1128/CMR.00080-13

April 2014 Volume 27 Number 2 Clinical Microbiology Reviews p. 167–199 cmr.asm.org 167

on Novem

ber 7, 2020 by guesthttp://cm

r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1128/CMR.00080-13

http://cmr.asm.org

http://cmr.asm.org/

lus strains suggests that they constitute a promising new source forthe development of innovative anti-infectious agents that act lu-minally and intracellularly in the gastrointestinal tract.

INTRODUCTION

The gastrointestinal (GI) tract forms complex ecosystem thatfunctions in concert with the resident microbiota as a struc-

tural and functional barrier that protects the host from attack byunwanted, harmful enterovirulent microorganisms (1). The mu-cosal surface of the gastrointestinal tract faces the external envi-ronment (2, 3). The stomach is a muscular organ that secretes theacid and enzymes involved in digesting food. Histologically, thehuman stomach can be divided into three regions: the cardia,the fundus/corpus, and the antrum (3). Specialized secretory cellphenotypes are present: acid-secreting parietal cells, mucus neckcells, and pepsinogen-secreting zymogenic cells in the fundus andcorpus and gastrin-secreting cells and gland cells in the antrum(4). These different cell types are localized in glandular invagina-tions, which are known as oxyntic glands in the corpus/fundusregion and as pyloric glands in the antrum. Members of the firstline of defense of the gastrointestinal tract against the unwantedintrusion of pathogenic bacteria are present in the stomach (3).Mucus-producing surface mucus cells cover the whole gastric mu-cosa, creating a mucus barrier composed of mucin glycoproteins:the membrane-associated MUC1 and the secreted MUC5AC andMUC6. A large number of bacteria are present in the outer mucuslayer, whereas the inner mucus layer is virtually free of bacteria. Inthe gastric epithelium several antimicrobial peptides (AMPs) arepresent, such as �-defensin 1, �-defensin 2, and cathelicidin LL37.In addition, hepcidin has been identified as a major regulator ofiron homeostasis and links the iron metabolism and host responseto infection (5). One of the main functions of the stomach is toachieve digestion, and the harsh gastric environment contributesto inactivating ingested microorganisms, including pathogens, toprevent them from reaching the intestine. The pyloric sphincterconnects the stomach with the intestine and controls passage ofthe digested food into the intestine.

Anatomically, the intestine is formed by four segments: theduodenum, jejunum, ileum, and colon. The gastrointestinal epi-thelium consists of a single layer of fully differentiated, polarizedepithelial cells of various phenotypes that create an impermeable,regulated epithelial barrier separating the external and internalenvironments. Four highly specialized cell phenotypes composethe intestinal epithelium: enterocytes (also known as fluid-trans-porting cells), neuroendocrine cells, mucin-secreting cells (alsoknown as goblet cells), and Paneth cells (6). Tight junctions posi-tioned most apically in the junctional domain of intestinal epithe-lial cells are the primary cellular determinant ensuring the closureof the intestinal epithelial barrier. There are several lines of chem-ical defenses in the intestine that function to prevent the passage ofluminal enteric bacterial pathogens across the epithelium. Theintestinal mucosa is coated by secreted mucus delivered by mucin-secreting cells (7, 8). The outer mucus layer favors the growth ofthe mucosa-associated resident microbiota and of enterovirulentbacteria by providing nutrients, and the density of the inner mu-cus layer limits the contact between luminal enterovirulent bacte-ria and the intestinal epithelial cells. The thinness of the mucuslayer in parts of the small intestine renders efficient specializedimmune anti-infective mechanisms. In contrast, the roles ofmembrane-bound mucins are poorly documented. The Paneth

cells are pyramid-shaped, columnar, exocrine cells located in thecrypts of the epithelium that provide AMPs, including defensins,C-type lectins, and cathelicidins (9, 10). AMPs are retained by themucus overlying the intestinal epithelium and act rapidly to kill orinactivate pathogenic microorganisms. For example, �-defensinsand cathelicidins kill both Gram-negative and Gram-positionpathogens, whereas C-type lectins kill only Gram-positive bacte-ria. Nevertheless, several enterovirulent bacteria can partiallyevade the action of AMPs by altering the anionic charge of theirown surface molecules. Intestinal host defense systems againstpathogenic enteric bacteria include both adaptive and innate im-munity. Adaptive immune responses are induced mainly in thefollicle-associated epithelium that overlies the organized gut-as-sociated lymphoid tissue through the interaction of intestinal ep-ithelial cells with antigen-presenting cells and lymphoid cells. Theintestinal epithelium senses the microbial environment and pro-duces strong cellular defense responses, including the release ofcytokines and chemokines which trigger the recruitment of leu-kocytes and others immune cells (11–13). Pathogen recognitionreceptors (PRRs), including Toll-like receptors, retinoic acid-in-ducible gene 1-like receptors, NOD-like receptors, and DNA re-ceptors, recognize pathogens expressing various signature mole-cules called pathogen-associated molecular patterns (PAMPs)and rapidly trigger a panel of antimicrobial immune responsesand also some adaptive immune responses. In addition, au-tophagy, which is an evolutionarily conserved process by whichcell constituents are recycled, acts as a cell defense mechanismagainst intracellular pathogenic bacteria (14).

GASTROINTESTINAL MICROBIOTA

Knowledge about the composition of the microbiota of the hu-man stomach is currently increasing (3). A low level of residentbacteria (101 to 103/ml) colonize the human stomach. The fivemain phyla that have been identified are the Firmicutes, Actinobac-teria, Proteobacteria, Fusobacteria, and Bacteroidetes. The domi-nant genera in Firmicutes are Lactobacillus, Streptococcus, Veillo-nella, Staphylococcus, and Bacillus. Lactobacillus species that havebeen identified in the gastric microbiota are L. antri, L. gastricus, L.kalixensis, L. reuteri, L. ultunensis, L. plantarum, L. salivarius, L.fermentum, and L. gasseri.

The adult human intestine has been estimated to contain tril-lions of microbes, including hundreds of species and thousands ofsubspecies, which display age- and geography-related differencesand are distributed as a function of the intestinal site; they have apredominantly symbiotic relationship with their host (15, 16).The intestinal microbiota of healthy adults expresses two domi-nant phyla, the Gram-negative Bacteroidetes and the Gram-posi-tive Firmicutes. Less abundant are the Proteobacteria, the Verruco-microbia, the Tenericutes, the Deferribacteres, and the Fusobacteria.Microbial colonization in the human GI tract evolves along thelength of the intestinal tract. A low level of 101 to 103 bacteria pergram of content is present in the duodenum, progressing to 104 to107 bacteria per gram of content in the jejunum and ileum andreaching 1011 to 1012 bacteria per gram of content in the colon. Inthe colon, most of the bacteria present are anaerobes, with about1,000-fold fewer facultative anaerobes. Infants acquire their com-mensal bacteria from other human beings, in particular from themother. Microbial colonization of the gut begins during birth andearly infancy and then proceeds under the influence of breastfeed-ing and skin-to-skin contact with the mother and other people

Liévin-Le Moal and Servin

168 cmr.asm.org Clinical Microbiology Reviews

on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

(17, 18). Many of these resident intestinal bacteria are adapted tothe intestinal environment and develop complex ecological net-works with other bacteria to acquire nutrients. Supported by bothphylogenetic and functional analyses conducted by the interna-tional MetaHIT consortium, data obtained from examining dif-ferent metagenomes indicate that the microbial communities in-clude three distinct clusters known as “enterotypes” (19). Eachenterotype displays specific species and functional compositionsthat differ from those of the other two.

The intestinal microbiota fulfills three functions (20). First, itassists host nutrition. The species that make up the resident mi-crobiota ferment components of the host’s diet that the host isunable to digest and synthesize low-molecular-weight metabo-lites, amino acids, and vitamins. After being absorbed from theintestinal lumen, these low-molecular-weight metabolites aretransported into the systemic circulation, where they play roles inboth health and disease. The functions of low-molecular-weightmetabolites produced by the intestinal microbiota that constitutethe intestinal metabolome remain largely unknown, although theroles of some of them, such as short-chain fatty acids and poly-amines, have been investigated (21). It is noteworthy that the an-tibiotic treatment induces modifications in the composition of theintestinal microbiota that have an impact on the intestinal ho-meostasis as a result of changes in the intestinal metabolome (22).Second, the intestinal microbiota also participates in the struc-tural and functional maturation of the epithelial cells lining theepithelial barrier and influences intestinal immune development(23). The third function relates to the intestinal host defensemechanisms, since several of the species that make up the residentmicrobiota inhibit the unwanted intrusion of harmful microor-ganisms by competing for nutrients and blocking the deleteriouseffects of bacterial virulence factors on the host cell by enhancingthe mechanisms of the host epithelial and immune defenses andproducing organic acids and antibacterial compounds (24, 25). Inaddition, the regulation of several host defense mechanisms re-sults in activation of innate immune receptors by microbial mol-ecules produced by the resident intestinal microbiota (26).

The resident intestinal microbiota has recently been shown toconstitute an innovative therapeutic strategy in the treatment ofseveral intestinal disorders. Indeed, fecal bacteriotherapy, alsocalled fecal microbiota transplantation, has recently been success-fully used as a therapy to correct the dysbiosis that characterizeschronic Clostridium difficile infection (27). Moreover, owing tothe role of resident intestinal microbiota in inflammatory boweldisease (IBD) (28) and the presence of an “activated intestinalmicrobiota” containing an elevated number of normally under-represented potentially harmful bacteria resulting from dysbiosis(20, 29), microbiota transplantation also appears to be potentiallyuseful to treat IBD and irritable bowel syndrome (27, 30, 31).Interestingly, an emerging recently described role of members ofthe microbiota is to provide protection against pathobionts, i.e.,normally harmless microorganisms that can become pathogensunder certain environmental conditions (32, 33).

Pathogenic bacteria infect specific regions of the gastrointesti-nal tract. Helicobacter pylori, which is never found in the small orlarge intestine, infects the stomach mucosa and to do this hasdeveloped special adaptation properties that enable this condi-tional pathogen to survive in the harsh gastric environment (34,35). Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC)target the polarized epithelial cells lining the small intestine,

whereas C. difficile, Shigella spp., enterohemorrhagic E. coli(EHEC), and Campylobacter spp. target the cells lining the colonepithelium, and Yersinia spp. and Salmonella spp. can affect boththe small intestine and the colon (36–40). The high level of inter-dependency within intestinal microbiota bacterial communitiesmakes it particularly difficult to study the cooperation betweenthe various intestinal bacterial species in producing the chemicalbarrier effect of the intestinal microbiota against the unwantedintrusion of bacterial pathogens. Various intestinal microbiotabacterial species that produce antimicrobial molecules triggeringa pivotal role in the chemical barrier effect of the intestinal micro-biota against the unwanted intrusion of bacterial pathogens havebeen identified. By producing bacteriocins, resident Gram-posi-tive bacteria in the intestinal microbiota have bacteriostatic orbactericidal effects against closely related Gram-positive species(41–43). Based on biochemical characteristics, bacteriocins of in-testinal microbiota Gram-positive bacteria are classified in twogroups, i.e., lantibiotics and heat-stable proteins not containinglanthionine residues, each of which is subclassified into multiplesubgroups (44–46). Bacteriocin-associated antibacterial activitieshave been observed both in vitro and in vivo at concentrations inthe nanomolar range and develop against Gram-positive bacteriaclosely related to the producing strain. Several bacteriocins actupon the cell envelopes of target pathogens, and others are activewithin the cell and affect gene expression (47, 48). Pores and ionchannels formed in the cytoplasmic membrane of the target mi-crobial cell and the subsequent leakage of intracellular compo-nents and protein production are the typical modes of bacterio-static and bactericidal activities, despite the structural andphysicochemical differences observed between the differentclasses of bacteriocins. In addition, resident Escherichia coli strainsexert potent bactericidal activity against closely related Gram-neg-ative species by also producing bacteriocins (43, 49), known as thelow-molecular-weight microcins (50) and the larger colicins (51),which are typically plasmid encoded. Microcins range from 1 to 10kDa, are subclassified based on the presence and localization ofposttranslational modifications and the organization of the genecluster and the leader sequence, and have MICs in the nanomolarrange. They disrupt a wide range of functions in the target cell,including ATP synthetase and DNA gyrase. Colicins range in sizefrom 30 to 80 kDa and have MICs in the picomolar to nanomolarrange. They have varied killing mechanisms, which include poreformation, DNase or RNase activity, or inhibition of peptidoglycanbiosynthesis. In bacteria, an intercellular communication processcalled quorum sensing (QS) is based on the synthesis and secretion ofsmall hormone-like molecules, termed autoinducers, coordinatedmainly in response to the bacterial population density (52, 53). Inbacterial pathogens, QS molecules bind to cognate receptors which,after activation, directly or indirectly control expression of targetgenes coding for virulence factors (54). A QS mechanism(s) regulatesthe production of bacteriocins by lactic acid bacteria via secreted bac-teriocin-like peptide pheromones (55–57). Interestingly, Lactobacil-lus QS molecules controlling bacteriocin production have beenfound to be activated in response to infection (58). Moreover, ithas been reported that Lactobacillus QS-quenching compoundshave been found to be involved in the control of virulence ofbacterial pathogens in vitro and in vivo (59–65).

Elie Metchnikoff, the Russian-born Nobel Prize winner whoworked at the Pasteur Institute in Paris, reported the first obser-vation of a beneficial role played by some bacteria of the human

From Probiotic Lactobacillus to Anti-Infectious Drugs

April 2014 Volume 27 Number 2 cmr.asm.org 169

on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

intestinal flora and suggested that “the dependence of the intesti-nal microbes on food makes it possible to adopt measures to mod-ify the flora in our bodies and to replace the harmful microbes byuseful microbes” (66). Henry Tissier (67), observing that “bifid”bacteria are abundant in the stools of healthy children but presentat only low levels in the stools of children with diarrhea, postulatedthat these bacteria administered to patients with diarrhea couldrestore a healthy gut flora. For Fuller (68), the natural balance ofthe gut microflora could be restored by administering probiotics.In this review, we highlight the key traits of six human intestinalprobiotic strains of Lactobacillus that have displayed experimen-tally demonstrated antibacterial activities against gastric or en-terovirulent bacterial pathogens and have clinically demonstratedtherapeutic efficacy against enterovirulent bacterium- and rotavi-rus-induced diarrhea and H. pylori-induced gastritis. Experimen-tal and clinical data indicate that the Lactobacillus-produced mol-ecules underpin activities that could be a potential source of newanti-infectious molecules offering alternatives to antibiotics totreat gastrointestinal infections.

PROBIOTIC LACTOBACILLUS STRAINS ISOLATED FROM THEHUMAN INTESTINAL MICROBIOTA

The FAO/WHO Expert Committee has defined probiotic strainsas “live microorganisms which, when consumed in appropriateamounts in food, confer a health benefit on the host” (69, 70). Inaddition, a set of major criteria has been defined for probioticspecies, among which may be cited proper taxonomic identifica-tion by molecular techniques, deposition in an internationallyrecognized culture collection, a lack of transmissible antibioticresistance genes, persistence in a viable state in the gastrointestinaltract, experimentally and clinically demonstrated health benefits,safety for human use, and resistance to technological processes asshown by, e.g., the preservation of cell viability and probiotic ac-tivities throughout the processing, handling, and storage of thefood product containing the probiotic strain (69). The probioticactivity that has undergone the most experimental and clinicalinvestigation is that exerted against gastric or enterovirulent bac-terial pathogens and rotaviruses, both of which constitute majorhuman health problems. Lactobacillus strains isolated from thehuman intestinal microbiota display antibacterial activities as aresult of producing metabolites such as lactic acid, bacteriocins,nonbacteriocin compounds (defined as sensitive to proteolyticenzymes but unable to be precipitated with 80% saturated ammo-nium sulfate), and nonproteinaceous molecules that exercise adirect bactericidal effect (41–43, 71) and/or downregulate the ex-pression of the virulence factors of enterovirulent pathogens ormodulate the deleterious effects of these factors on the host’s in-testinal cell structure, machinery, and functions (72–75). It isnoteworthy that a given effect of a probiotic Lactobacillus is strainspecific and cannot be extrapolated to other strains of the Lacto-bacillus genus or even to other strains belonging to the same spe-cies and subspecies. In addition, substances produced by probioticLactobacillus strains, including cell wall components and secretedmolecules, have been reported to display immunomodulatory ac-tivities, mainly in in vitro experiments (76, 77). The mechanisticstudies showing the immunomodulatory effects of probiotics arebased principally on in vitro cell culture models (78). Positiveresults have recently been reported in in vivo models, but currentlythere has been no convincing clinical demonstration of a probi-otic-induced immunostimulatory effect in human patients.

A large number of Lactobacillus strains have been isolated fromhuman and animal intestinal microbiota, and the properties ofthese strains, including their adhesion to cultured epithelial cellsor mucus and their inhibitory activities, have been reportedmainly from in vitro experiments. Only six Lactobacillus strainsisolated from the human intestinal microbiota have been clearlydemonstrated to have probiotic antimicrobial and antirotavirusproperties in a comprehensive set of in vitro and in vivo experi-ments and randomized controlled trials (RCTs). These probioticstrains are L. rhamnosus strain GG (ATCC 53103) (Valio Ltd.,Finland) (79–82), L. casei strain Shirota YIT9029 (Yakult HonshaCo., Ltd., Japan) (83, 84), L. acidophilus strain LB (Forest Labora-tories, Inc., New York, NY) (85), L. johnsonii NCC 533 (first de-signed La1) (CNCM I-1225) (Nestlé, Switzerland) (86), L. caseiDN-114 001 (CNCM I-1518) (Danone, France) (87), and L. reu-teri DSM 17938 (ATCC 55730 cured of the pLR581 and pLR585antibiotic resistance plasmids, also designed SD2112, ING1, andMM53) (BioGaia AB, Sweden) (88–90). (L. acidophilus strain LBwas historically identified as such on the basis of its biochemicaland metabolic activities; subsequent molecular investigation re-classified the identity of this strain as a symbiotic culture of L.fermentum [L. fermentum LB-f] [CNCM I-2998] [91, 92] and L.delbrueckii strains in a 95:5 ratio.)

Other probiotic Lactobacillus strains have been isolated fromthe human intestinal microbiota; these include L. acidophilusstrain NCFM (also designated RL8K/NCK45/NCK56/N2) (ATCC700396) (Danisco A/S, Denmark) (93), L. plantarum 299v (DSM6595) (Probi AB, Sweden), and L. fermentum ME-3 (DSM 14241)(University of Tartu and Tere AS, Estonia) (94). It is noteworthythat these probiotic strains have been experimentally tested onlyin vitro against gastrointestinal pathogens or in animal infectionmodels or have been experimentally tested for other probioticeffects. It was noted that there have been no RCTs for these strainsin human infection situations, but several of these strains havebeen therapeutically tested in humans for other probiotic effects.In addition, it was noted that the probiotic L. rhamnosus R0011(CNCM I-1720) and L. helveticus R0052 (CNCM I-1722) strainsisolated from dairy cultures (Institut Rosell-Lallemand Inc., Can-ada) (95, 96) have been shown to exhibit both experimental andclinical anti-infectious effects (97).

According to a U.S. Food and Drug Administration (FDA)working definition, probiotics are classified as “live biotherapeu-tics”: “live microorganisms with an intended therapeutic effect inhumans” (98, 99). Guidelines for the clinical use of probioticstrains were published after a Yale University workshop in 2005and were updated in 2007 (100). The advice is graded as “A,” “B,”“C,” or no category. Classified in the A-grade advice are probioticLactobacillus strains used to treat acute childhood diarrhea (101)and C. difficile-associated diarrhea (102). Probiotic Lactobacillusstrains used to treat chronic disorders of the gut, including IBD,are classified as having B-grade advice because there have beensome negative studies (103). It was noted that the C grade relatesto results that were significant but unable to receive strongergrades because of the numbers of patients enrolled in studies. Theprobiotic strains are considered to be safe, or “generally recog-nized as safe” (GRAS) (104–106). However, as Sanders et al. (105)point out and as stated in the report of the Agency for HealthcareResearch and Quality (AHRQ) (104), there was a small number ofstudies specifically designed to assess probiotic safety, contrastingwith the long history of safe use of foods containing probiotic



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

bacteria. The information that is required for the development ofprobiotic products containing live or lyophilized dormant probi-otic Lactobacillus in food or dietary supplements includes the step-by-step description of the manufacturing process, which is carriedout under aseptic conditions, and the quality control process, in-cluding the process-input parameters and the expected outputresults that ensure that the product remains stable and pharma-cologically effective over the indicated conservation time (69, 99).Indeed, Fayol-Messaoudi et al. (107) have found that the bacteri-cidal activity against Salmonella enterica serovar Typhimurium of24 h-cultures of L. rhamnosus GG, L. johnsonii NCC 533, L. caseiShirota, and L. casei DN-114 001 rapidly decreased by 4 logCFU/ml after 1 day in storage at 4°C. Grzeskowiak et al. (80) haveinvestigated the probiotic antibacterial effect of L. rhamnosus GGisolated from 10 probiotic products, including capsules, commercialinfant foods, and freeze-dried powders from several different coun-tries. Compared to parental L. rhamnosus GG, the antibacterial activ-ities of the product-isolated strains varied significantly. Moreover, theresuscitation of dormant probiotic strains (108, 109) needs particularattention during processing. Indeed, as demonstrated by Muller et al.(110), the resuscitation of dried probiotic strains, including L. john-sonii NCC 533, is a critical aspect for obtaining active and effectiveprobiotic strains, as multivariate factors, including the pH, thediluent, and the reconstitution time, can all have a strong influ-ence on the content, viability, and activity of the restituted probi-otic bacteria. As a consequence, the authors stress that it is neces-sary for the resuscitation conditions to be optimized for each ofthe lyophilized probiotic strains used. These experimental obser-vations are in line with what was previously defined by the FAO/WHO expert group (69) and by Sanders (111), i.e., the need toconduct appropriate quality control measures for all foods con-taining probiotic strains to ascertain whether the original proper-ties of each of the probiotic strains have been preserved after themanufacturing process undergone by the food.

IN VITRO ANTIBACTERIAL ACTIVITIES

Direct Activities against Enterovirulent Bacteria

Bacteriostatic activity. Lactobacillus strains exert direct antago-nistic activities by a bacteriostatic activity that blocks the growth ofenterovirulent bacteria as a result of the strain-specific productionof bacteriocins. Several bacteriocins are used as biopreservatives infood and food products to inhibit or control the growth of food-borne bacterial pathogens (41–43). The generally held view is thatbacteriocins exhibit less potential as chemotherapeutics for infec-tions with Gram-negative pathogens. However, it should be notedthat several bacteriocins and bacteriocin-like molecules producedby probiotic Lactobacillus strains have been also found to be activeagainst the growth of several Gram-negative gastric or enteroviru-lent bacterial pathogens, including H. pylori, EHEC, Shigella, Sal-monella, and Campylobacter (112–116).

Bactericidal activity. Certain probiotic Lactobacillus strainsdisplay bactericidal activity against Gram-negative or Gram-pos-itive gastric or enterovirulent bacteria after direct contact in vitro.Bactericidal activities of probiotic Lactobacillus strains have beengenerally explored using probiotic cultures. For some of the sixprobiotic Lactobacillus strains examined here, it has been shownthat bacteria isolated from a culture do not have a bactericidaleffect after direct contact with pathogens and that the bactericidalactivities of cultures are reproduced with the cell-free spent cul-

ture supernatants (CFCSs) (Fig. 1 to 3 and Table 1). It is importantto note that this activity develops within the specified limits for anantibiotic acting against a pathogenic microorganism, i.e., thebactericidal activity producing greater than a 3-log reduction of aviable cell count of a test microorganism after incubation for afixed length of time under controlled conditions (117). A loss of�4 log CFU/ml of Shigella viability has been observed to be trig-gered by the L. rhamnosus GG (118, 119), L. johnsonii NCC 533(120), L. reuteri ATCC 55730 (121), and L. acidophilus LB (122)strains after 4 h of direct contact. The viability of Listeria wasaffected by 3 to 4 log CFU/ml after 4 h of exposure to the L.johnsonii NCC 533 (120) and L. acidophilus LB (122) strains. TheL. rhamnosus GG (119, 123), L. casei Shirota (124), L. reuteriATCC 55730 (121), and L. acidophilus LB (122, 125) strains de-creased the viability of enterovirulent E. coli by 3 to 4 log CFU/mlafter 4 h of direct contact. The viability of S. Typhimurium wasdramatically lowered, by �5 log CFU/ml, after 4 h of exposure tothe L. rhamnosus GG (107, 118, 119, 126–131), L. johnsonii NCC533 (107, 120, 128, 129, 132–134), L. casei Shirota (107, 128, 129,135), L. casei DN-114 001 (107), L. reuteri ATCC 55730 (121), andL. acidophilus LB (122, 136, 137) strains. Antagonistic activityagainst Vibrio cholerae has been reported only for L. reuteri ATCC

FIG 1 Bactericidal effect of probiotic L. acidophilus strain LB against gastric orenterovirulent bacterial pathogens. (A) Time course of the bactericidal effectof L. acidophilus strain LB against wild-type ETEC H10407 expressing coloni-zation factor CFA/I, EPEC E2348/69, Afa/Dr DAEC C1845, S. TyphimuriumSL1344, and H. pylori 1101 after direct contact. (B) Scanning electron micros-copy micrographs showing the transformation of the helical form of H. pylorito the U-shaped form after treatment with L. acidophilus strain LB. For bothpanels A and B, pathogens were placed in direct contact with L. acidophilus LBCFCS. (The time course of the bactericidal effect in panel A is based on dataextracted from reference 182 with permission of the publisher and from ref-erences 122 and 139. The two micrographs in panel B are reprinted fromreference 139.)



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

55730 (121). It is noteworthy that the bactericidal activity mustdevelop when a bacterial pathogen attaches to human intestinalcells, since diffusely adhering E. coli strains (DEAC) expressingAfa/Dr adhesins (Afa/Dr DAEC) attached within brush borders ofenterocyte-like Caco-2/TC7 cells are killed after treatment of the in-fected cells with L. acidophilus LB CFCS (125) (Fig. 2). A problemin intestinal antibiotic therapy is the ineffectiveness of antibioticsagainst enteroinvasive bacterial pathogens that have already en-

tered host intestinal cells and taken up residence within the cellcytoplasm or intracellular vacuoles. Consequently, the observa-tion that a compound(s) secreted by L. acidophilus LB was able tokill S. Typhimurium resident in the intracellular vacuoles locatedwithin the enterocyte-like Caco-2/TC7 cells is particularly inter-esting (136).

The bactericidal activity of L. rhamnosus GG, L. johnsonii NCC533, L. casei Shirota, L. casei DN-114 001, and L. acidophilus LB has

FIG 2 Bactericidal effect of the probiotic L. acidophilus strain LB against gastric or enterovirulent bacterial pathogens adhering to infected cultured humanintestinal cells. (A) Time course of the bactericidal effect of L. acidophilus strain LB against the wild-type Afa/Dr DAEC C1845 strain adhering to the brush bordersof enterocyte-like Caco-2/TC7 cells. (B and C) Scanning electron micrographs showing the L. acidophilus strain LB-induced changes in the wild-type Afa/DrDAEC C1845 strain adhering to the brush borders of enterocyte-like Caco-2/TC7 cells and in H. pylori strain 1101 adhering to cultured human mucus-secretingHT29-MTX cells, respectively. For panels A to C, preinfected cells were treated with L. acidophilus LB CFCS. The drawing on the right indicates the localizationin enterocytes of the effects reported in panels A to C. (The time course of bactericidal effects in panel A and the two micrographs in panel B are reproduced fromreference 125 with permission from BMJ Publishing Group, Ltd. The two micrographs in panel C are reproduced from reference 139.)



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

been shown to be entirely triggered by secreted compounds pres-ent in CFCSs (107, 122, 125, 129, 130, 138). The activity exertedseems to result from various molecules acting alone or synergisti-cally (Table 1). In vitro, the bactericidal activity of probiotic Lac-tobacillus cultures has been proposed to result from the acidic pH.However, Fayol-Messaoudi et al. (107), investigating how the invitro bactericidal activity of L. rhamnosus GG, L. casei Shirota, L.casei DN-114-001, and L. johnsonii NCC 533 CFCSs against S.Typhimurium develops, observed that the acidic pH contributedonly a small part. During fermentative metabolism, lactobacilliproduce organic acids as terminal products, triggering antagonis-tic activities against bacterial pathogens through intracellularacidification and membrane permeabilization. The main meta-bolic compound secreted, i.e., lactic acid, has been suspected toplay a pivotal role in the bactericidal activity of probiotic Lactoba-cillus strains. It has been observed that the in vitro bactericidalactivity of lactic acid against S. Typhimurium increased linearlywith increasing lactic acid concentrations (107, 119, 129), but atthe concentrations present in the probiotic Lactobacillus cultures(50 to 80 mM), it displayed no bactericidal activity (107, 122, 136,139). The antimicrobial effect of lactic acid is not just due to thelowering of the intracellular pH. Indeed, hydrogen peroxide pro-

duced by L. johnsonii NCC 533 and other L. johnsonii strains invitro kills S. Typhimurium (133), an effect enhanced in the pres-ence of the membrane permeabilizer lactic acid (132). As deducedfrom in vitro experiments that have tested the sensitivities of se-creted compounds present in L. rhamnosus GG, L. johnsonii NCC533, and L. acidophilus LB CFCSs to a set of physical and chemicaltreatments and from partial isolation experiments, bactericidalactivity against S. Typhimurium results from small (dialysis cut-off, 1,000 Da), nonproteinaceous compounds (120, 122, 123). Thecompound(s) supporting the bactericidal activity present in L.acidophilus LB CFCS is heat resistant (122). Recently, five low-molecular-weight, nonproteinaceous bactericidal compoundsthat are heat stable and active at acidic pH values have been foundin L. rhamnosus GG CFCS, which either with or without lactic aciddisplay antimicrobial activity against S. Typhimurium (140).Moreover, Lu et al. (141) have reported that seven heat-resistantsmall peptides, two of which have NPSRQERR and PDENK se-quences, are present in L. rhamnosus GG CFCS and display anti-bacterial activity against enteroaggregative E. coli (EAEC) strain042 and Salmonella enterica serovar Typhi. It was noted thatLactobacillus strains that produce bacteriocins that are active tokill Salmonella, Campylobacter, and E. coli have been found

FIG 3 Bactericidal effect of probiotic L. acidophilus strain LB against wild-type S. Typhimurium SL1344 residing within intracellularly localized vacuoles inpreinfected cultured human intestinal Caco-2/TC7 cells. (A) Time course of the bacterial effect. (B) Confocal laser microscopy scanning examination offluorescein-labeled S. Typhimurium SL1344 cells showing the altered morphology of bacteria residing within a typical intracellular vacuole in L. acidophilusstrain LB-treated cells compared to untreated cells. For panels A and B, preinfected cells were treated with L. acidophilus LB CFCS. The drawing on the rightindicates the localization in enterocytes of the effects reported in panels A and B. (The time course of the bactericidal effects in panel A and the two micrographsin panel B are reproduced from reference 136.)



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

TABLE 1 Overviews of in vitro antibacterial effects of probiotic Lactobacillus strains isolated from human intestinal microbiota against gastric orenterovirulent bacterial pathogens

Pathogen Probiotic strain Experimental conditions Observed effect(s) Reference(s)

Shigella L. rhamnosus GG Direct contact Bactericidal 118, 119L. johnsonii NCC 533 Direct contact with culture or CFCS Bactericidal 120L. casei DN-114 001 Direct contact Inhibition of upregulation of proinflammatory genes 224L. reuteri ATCC 55730 Direct contact Bactericidal 121L. acidophilus LB Direct contact with culture or CFCS Bactericidal 122, 125

Listeria L. johnsonii NCC 533 Direct contact with culture or CFCS Bactericidal 120L. acidophilus LB Direct contact with culture or CFCS Bactericidal 122, 125

Enterovirulent E. coli L. rhamnosus GG Direct contact Bactericidal 119, 123Peptides with NPSRQERR and

PDENK sequences isolated fromCFCS

Bactericidal 141

Direct contact Decrease of Shiga toxin Stx2A mRNA 145Direct contact Inhibition of adhesion onto cultured epithelial cells 177, 178, 179Direct contact Inhibition of TJ lesions in cultured enterocyte-like cells 215.Direct contact Inhibition of IL-8, CCL, and CXCL production 218, 219, 220,

222, 223Direct contact Increased MUC2 and MUC3 mRNA, which in turn

inhibited the adhesion of EPEC and EHEC228

L. johnsonii NCC 533 Direct contact Inhibition of adhesion onto cultured epithelial cells 179, 180L. casei Shirota Direct contact Bactericidal 124

Direct contact Inhibition of adhesion onto cultured epithelial cells 177, 178L. reuteri ATCC 55730 Direct contact Bactericidal 121

Direct contact Repression of A/E locus 146Direct contact Inhibition of adhesion onto cultured epithelial cells 179

L. acidophilus LB Direct contact with culture or CFCS Bactericidal 122, 125Direct contact with heat-treated CFCS Conservation of bactericidal 122, 125Direct contact with culture or CFCS Inhibition of adhesion onto cultured epithelial cells 125, 181, 182,

183, 187Direct contact with heat-treated CFCS Conservation of inhibitory effect against adhesion onto

cultured epithelial cells125, 181, 182,

183, 187, 188Direct contact with untreated or heat-

treated CFCSInhibition of structural and functional injuries at the brush

borders of cultured enterocyte-like cells125

Direct contact with CFCS Inhibition of TJ lesions in cultured enterocyte-like cells 207L. casei DN-114 001 Direct contact Inhibition of adhesion onto cultured epithelial cells 185

Direct contact Inhibition of TJ lesions in cultured enterocyte-like cells 186

Vibrio cholerae L. reuteri ATCC 55730 Direct contact Bactericidal 121

S. Typhimurium L. rhamnosus GG Direct contact with culture or CFCS Bactericidal 107, 118, 119,126–131

Peptides with NPSRQERR andPDENK sequences isolated fromCFCS

Bactericidal 141

Direct contact Inhibition of adhesion onto cultured epithelial cells 178Direct contact with culture or CFCS Inhibition of cell-entry into cultured enterocyte-like cells 126, 128–130Direct contact Inhibition of IL-8 production 219, 220

L. johnsonii NCC 533 Direct contact with culture or CFCS Bactericidal 107, 120, 128,129, 132–134

Direct contact with culture or CFCS Bactericidal effect of the produced hydrogen peroxide andcooperation with lactic acid

132, 133

Direct contact with culture or CFCS Inhibition of cell entry into cultured enterocyte-like cells 128–130, 180L. casei Shirota Direct contact with culture or CFCS Bactericidal 107, 128, 129, 135

Direct contact Inhibition of adhesion onto cultured epithelial cells 178Direct contact with culture or CFCS Inhibition of cell-entry into cultured enterocyte-like cells 128, 129Direct contact with CFCS Inhibition of flagellum swimming motility 148Direct contact with CFCS Inhibition of IL-8 production 221

L. casei DN-114 001 Direct contact with culture or CFCS Bactericidal 107L. acidophilus LB Direct contact with culture or CFCS Bactericidal 122, 125

Direct contact with CFCS Inhibition of flagellum swimming motility 147Direct contact with culture or CFCS Inhibition of cell entry into cultured enterocyte-like cells 136, 137, 139,

147, 182, 183Direct contact with CFCS Bactericidal effect against intracellular vacuole-localized S.

Typhimurium136

Direct contact with heat-treated CFCS Conservation of the bactericidal effect, inhibitory effectagainst flagellum motility, inhibitory effect against cellentry into cultured enterocyte-like cells and againstintracellular vacuole-localized S. Typhimurium

136, 137, 139,147, 182, 183

(Continued on following page)



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

(142–144), but none of the six Lactobacillus strains described hereseems to produce bacteriocin.

The bacterial membrane damage that accompanies the bacte-ricidal effects produced by the CFCSs of several human intestinalmicrobiota Lactobacillus strains resembles the bacterial cell dam-age produced by antibiotics and bacteriocins (41–43) and by sev-eral intestinal epithelial AMPs (9). For example, before L. acidoph-ilus LB CFCS-induced S. Typhimurium cell death is achieved,there is a change in the cell membrane (122) accompanied by therelease of lipopolysaccharide, an increase in membrane permea-bility, and a loss of intracellular ATP (137).

Activity on the expression or functionality of virulence fac-tors. Molecules produced by probiotic Lactobacillus strains canalso directly affect the expression or functionality of virulence fac-tors of enterovirulent bacteria without affecting cell viability (Ta-ble 1). In Shiga toxin-producing E. coli O157:H7, L. rhamnosusGG has been shown to reduce the levels of Shiga toxin stx2A mRNA(145). L. reuteri ATCC 55730 repressed the expression of the locusof the enterocyte effacement-encoded regulator involved in theattachment/effacement (A/E) lesion of enterocyte microvilli byEHEC (146). Impairment of S. Typhimurium swimming motilityhas been observed after treatment with CFCSs of L. acidophilus LBand L. casei Shirota as a result of membrane depolarization thataffects the functionality of the flagellar motor but not flagellumexpression (147, 148). The inhibition of the swimming motilityresults from the secretion of a small compound(s) (dialysis cutoff,1,000 Da), which in the case of the L. acidophilus LB compound(s)was heat and trypsin resistant whereas in that of the L. casei Shirotacompound(s) was heat and trypsin sensitive (147, 148).

Direct Activities against H. pylori

Bactericidal activity. H. pylori is a member of the class Epsilonpro-teobacteria, composed almost exclusively of helical and curvedorganisms (34, 35). H. pylori colonizes the gastric tissue and causesserious disorders, ranging from mild gastritis to the onset of

chronic gastric inflammation that can lead to ulcers and gastriccancer, although most infected individuals are asymptomatic. Ithas been documented that the human probiotic intestinal micro-biota L. johnsonii NCC 533, L. casei Shirota, and L. acidophilus LBdisplay direct antagonistic activity in vitro against the gastritis-associated bacterium H. pylori (Fig. 1 and Table 1). pH-dependentbactericidal activity against H. pylori was exhibited by L. casei Shi-rota CFCS (149). Similarly, direct contact with the CFCSs of L.johnsonii NCC 533 (150) or L. acidophilus LB (139) resulted in therapid and dramatic loss (�6 log) of H. pylori viability. Moreover,H. pylori cells were morphologically affected by treatment with L.johnsonii NCC 533 (149), L. acidophilus (139), or L. casei Shirota(148) CFCS, shifting from their characteristic helical form to U-shaped and coccoid forms (Fig. 1). Avonts and De Vuyst (151)have also shown that L. johnsonii NCC 533 and L. casei Shirota canproduce bacteriocins which are active against H. pylori. It is note-worthy that other Lactobacillus strains producing bacteriocinsthat are also active against H. pylori have been identified (113,152–154). The molecule(s) present in L. acidophilus LB CFCS thatexerts bactericidal activity against H. pylori remain to be identi-fied. The appearance of coccoid forms after treatment with theabove-mentioned Lactobacillus strains is important since it resem-bles the morphological changes produced in H. pylori by antibiot-ics (155, 156) or colloidal bismuth subcitrate (157). Moreover, thetransformation into coccoid forms is interesting in terms ofpathogenesis, since even though these forms conserve the genesthat code for virulence factors found in the spiral form, they areknown to be less likely to colonize and induce inflammation thanthe corresponding spiral forms (158).

Activity against the expression and functionality of virulencefactors. Secreted compounds produced by human probiotic in-testinal microbiota Lactobacillus strains have a direct effect on theexpression or function of H. pylori virulence factors (Table 1).Urease is a surface protein component of H. pylori which allows it

TABLE 1 (Continued)

Pathogen Probiotic strain Experimental conditions Observed effect(s) Reference(s)

H. pylori L. rhamnosus GG Direct contact with CFCS Low bactericidal activity and loss of activity by CFCS heattreatment

139

Direct contact with a producedbacteriocin

Bactericidal activity 151

Direct contact Inhibition of adhesion onto gastric cells 235Direct contact with CFCS Absence of inhibitory effect against adhesion onto mucus-

secreting cells139

Direct contact Inhibition of IL-8 production 235, 236L. johnsonii NCC 533 Direct contact with a produced

bacteriocinBactericidal activity 139, 150, 151

GroEL protein GroEL protein-dependent aggregation of bound cells ontocultured epithelial cells

160, 234

Direct contact with CFCS Inhibition of IL-8 production 149Direct contact with CFCS Inhibition of flagellum swimming motility 160

L. casei Shirota Direct contact with CFCS Bactericidal activity with formation of coccoid forms 149Direct contact with CFCS Inhibition of urease activity 149Direct contact with CFCS Inhibition of flagellum swimming motility with formation

of U-shaped forms148

L. acidophilus LB Direct contact with culture or CFCS Bactericidal with formation of coccoid forms 139Direct contact with culture or CFCS Inhibition of urease activity 139Direct contact with CFCS Bactericidal effect against preadhering cells onto mucus-

secreting cells139

Direct contact with CFCS Inhibition of adhesion onto mucus-secreting cells 139Direct contact with heat-treated CFCS Conservation of bactericidal activity and inhibition of

urease activity139



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

to survive within the stomach by neutralizing the gastric acidicenvironment (35). Both L. acidophilus LB (139) and L. casei Shi-rota (149) CFCSs have been shown to induce a dramatic loss of H.pylori urease activity. In contrast, L. rhamnosus GG CFCS has alower effect on H. pylori urease activity (139). Flagellar motilitytogether with the helical cell shape of the pathogen has beenshown to be essential for the ability of H. pylori to colonize thestomach (35). H. pylori cells swim within the gastric mucus layer,propelled by a bundle of rotating flagella (159). L. johnsonii NCC533 secretes nonproteinaceous compounds of �1,000 Da that in-hibit the swimming motility of H. pylori (160). L. casei ShirotaCFCS irreversibly inhibits the swimming motility of H. pylori bychanging the morphology of the pathogen from its characteristichelical form with polar flagella to U-shaped and coccoid formsdevoid of flagella (148). Irreversible inhibition of H. pylori swim-ming motility by L. casei Shirota results from the secretion of asmall compound(s) (dialysis cutoff, 1,000 Da) that are heat andtrypsin sensitive (148).

ACTIVITIES AGAINST THE DELETERIOUS EFFECTS INDUCEDBY INFECTIOUS AGENTS AT THE INTESTINAL EPITHELIALBARRIER

Activities against Enterovirulent Bacteria

Enterovirulent bacteria initiate their infectious process by attach-ing themselves to the target epithelial cells that line the intestinalepithelium by use of specialized adhesive factors (36, 161–164).For example, for interacting with the polarized host epithelial cellsthat line the intestinal barrier (165–167), many bacterial speciesmove in the luminal intestinal compartment by rotating their fla-gella (168, 169). The adhesion of enterovirulent bacteria to thebrush border of the enterocyte involves much more than just sim-ple attachment (161). In the case of ETEC, it permits the optimaldelivery of cytotoxic toxins in the vicinity of their membrane-associated receptors, followed by signaling events affecting elec-trolytes and fluid secretion (164). For EAEC, attachment opti-mizes the delivery of autotransporter toxins at the brush border(170). Intestinal bacterial pathogens are equipped with a variety ofweapons that provide them with a variety of mechanisms for sub-verting the cellular machinery and circumventing host defenses.Pathogens have developed sophisticated ways to secrete proteinsfrom the cytoplasmic compartment outside the bacterial cell. Thisallows the type III secretion systems (T3SS) of enteropathogenic E.coli (EPEC) and EHEC to insert a translocated intimin receptorinto the host cell membrane, thus triggering the recruitment ofactin directly underneath the attached bacteria to form pedestalstructures leading to intimate attachment of the bacterium, result-ing in characteristic A/E lesions on the brush border microvilli andin dramatic defects in the absorption/secretion functions (36, 162,171, 172). Some enterovirulent bacteria have developed sophisti-cated strategies for altering and opening the junctional domain ofthe intestinal epithelial barrier (6, 173). For enteroinvasive patho-gens such as Shigella (40) and Salmonella (163), adhesion initiatesan orderly series of T3SS-dependent, bacterial effector-controlledmolecular events within a defined area on the host cell membrane,which facilitate the formation of the dramatic actin-rich cell sur-face ruffles that are pivotal to the successful completion of bacte-rial invasion, after which the internalized bacteria adopt specificintracellular lifestyles. Some of these pathogens, such as Shigella(163) and Listeria (174), live in the cell cytoplasm, within which

the bacteria move by means of actin-based motility, and thencespread into the neighboring cells via cellular membrane invagina-tions known as transpodia. Once other invasive pathogens, suchas Salmonella, have been internalized, they take up residencewithin the cell cytoplasm inside large vesicles where they replicate(163, 175). In addition, emerging evidence indicates that theseeffectors are mimetic proteins expressing functional domains ormotifs by which bacteria activate cell signaling pathways thatmodify signaling-regulated cell functions. Antagonistic activitiesof probiotic Lactobacillus strains against the structural and func-tional cell injuries promoted by enterovirulent pathogens at theintestinal barrier have been investigated mainly using culturedfully differentiated colon carcinoma cells that structurally andfunctionally mimic the human intestinal barrier and are used ex-tensively to dissect the mechanisms of virulence of the major en-terovirulent pathogens (176). Data have demonstrated that regu-latory molecules produced by human intestinal microbiotaLactobacillus strains act by modulating several receptor signalingcascades that are known to have a pivotal role in the deleteriouseffects of enterovirulent bacteria on the structural organizationand functionality of cell types lining the intestinal epithelial bar-rier (Table 1).

Effects at the brush border. Inhibition of the intestinal cell as-sociation of enterovirulent bacteria involved in acute infantile andtraveler’s diarrhea by human probiotic Lactobacillus strains hasbeen reported (Table 1). Inhibition (�5 to 6 log CFU/ml) of theadhesion of ETEC expressing colonization factors CFA/I andCFA-II, EPEC, EHEC, and Afa/Dr DAEC to the brush border ofcultured human enterocyte-like Caco-2 cells (176) develops in thepresence of adhering L. rhamnosus GG (177–179), L. johnsoniiNCC 533 (179, 180), L. casei Shirota (177, 178), L. reuteri ATCC55730 (179), and L. acidophilus LB (125, 181–183). The interac-tion of S. Typhimurium with enterocyte-like Caco-2 cells was in-hibited by �6 to 7 log CFU/ml in the presence of L. rhamnosus GG(128–130, 178), L. johnsonii NCC 533 (128, 129, 180), and L. caseiShirota (128, 129, 178) cultures and CFCSs. In contrast, L. rham-nosus GG did not affect the adhesion of S. Typhimurium to hu-man colonic tissue specimens (184). L. casei DN-114 001 inhibitedby �5 log CFU/ml the interaction of adherent-invasive E. coliisolated from Crohn’s disease patients with cultured human intes-tinal epithelial cells (185) but failed to block the adhesion of EPEC(186). It is noteworthy that heat-treated L. acidophilus LB cultureor CFCS conserved the antagonistic activity of the live strainagainst the attachment at the brush borders of Caco-2 cells ofETEC (181, 183, 187, 188), EPEC (182, 183), and Afa/Dr DAEC(125). Moreover, L. rhamnosus GG and L. casei Shirota inhibitedthe adhesion of enterovirulent bacteria onto mucus (177, 178,189). The presence of carbohydrate-binding specificities in L.johnsonii NCC 533 mimicking cell surface adhesins of enteric bac-terial pathogens (190) and the recently evidenced presence of piliinvolved in adhesion of L. rhamnosus GG (191–197) may explainthe competitive inhibition exerted by these two probiotic strainsagainst adhesion of enterovirulent bacteria.

The entry of S. Typhimurium into enterocyte-like Caco-2 cells(198, 199) was entirely abolished (decrease of �7 to 8 log CFU/ml) in the presence of cultures or CFCSs of L. rhamnosus GG, L.casei Shirota, L. johnsonii NCC 533, L. acidophilus LB, and L. caseiDN-114 001 (120, 126, 128–130, 136, 137, 139, 182, 183). ForLehto and Salminen (131), the inhibition of internalization of S.Typhimurium into Caco-2 cells by L. rhamnosus GG resulted from



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

the acidic pH. In contrast, experiments using noncultivated Lac-tobacillus culture medium acidified to pH 4.5 have demonstratedthat the inhibitory activity of CFCSs of L. rhamnosus GG, L. john-sonii NCC 533, L. casei Shirota, L. acidophilus LB, and L. caseiDN-114 001 did not result solely from the acidic pH (128, 129,136, 137, 139, 182, 183). It has recently been demonstrated that acompound(s) secreted by Lactobacillus strains blocked the swim-ming motility of S. Typhimurium, which plays a pivotal role inenabling the pathogen to swim into the intestinal contents andinteract with the intestinal epithelial cells (165, 167, 200). A heat-stable and trypsin-resistant secreted compound(s) present in theL. acidophilus LB CFCS (147) and a heat- and trypsin-sensitivecompound(s) present in the L. casei Shirota CFCS (148) by theblockade of S. Typhimurium delayed the penetration of the inva-sive bacteria into enterocyte-like Caco-2/TC7 cells.

The entry of Salmonella into intestinal cells is followed by acharacteristic intracellular lifestyle, including the presence of livebacteria within large intracellular vacuoles (201). S. Typhimuriumis a typical invasive enteropathogen that develops a mechanism ofmicrobial “nutritional virulence” which, through gaining accessto host nutrients in infected tissues, controls its growth, virulence,disease progression, and infection (202). Internalized S. Typhi-murium cells live in intracellular vacuoles where they haveadapted to the host cell environment by expressing versatile cata-bolic pathways to exploit multiple host nutrients for bacterialgrowth (175). In Caco-2/TC7 cells preinfected with S. Typhimu-rium, the treatment of the cells with L. acidophilus LB CFCS re-sulted in a dramatic and irreversible decrease in the intracellularlevel of S. Typhimurium (a decrease of �4 to 5 log CFU/ml) (136)(Fig. 3). Moreover, transmission electron microscopy observationof the S. Typhimurium cells that remained in the intracellularvacuoles revealed cells displaying the morphology typical of deadcells (136) (Fig. 3). This observation is of interest, because intra-cellular enterovirulent bacteria are known to be resistant to anti-biotics and can constitute a population of persistent and dormantpathogens (203).

The brush border of enterocytes is constituted by a regulararray of microvilli, the membrane of which is endowed with hy-drolases, such as sucrase-isomaltase (SI), alkaline phosphatase(AP), lactase-phloridzin hydrolase, maltase-glucoamylase amino-peptidase N, and dipeptidylpeptidase IV (DPP IV), and transport-ers, such as sodium/glucose cotransporter 1, GLUT1, GLUT2,GLUT3, and GLUT5 hexose transporters, peptide transporter 1,H�-coupled dipeptide transporter, cholesteryl ester transfer pro-tein, and Na�/H� exchanger isoforms (204). The protection ofbrush border-associated intestinal functions has been demon-strated only for L. acidophilus LB. The enterovirulent Afa/DrDAEC, which promotes the destruction of the enterocyte brushborder and, in turn, a dramatic loss of brush border-associatedfunctions (205), has been used to examine the protective effect ofL. acidophilus strain LB. The killing of Afa/Dr DAEC adhering toenterocyte-like Caco-2/TC7 cells, seen after treatment with L. ac-idophilus LB CFCS, resulted in the maintenance of a normal F-ac-tin brush border cytoskeleton (125). Moreover, when Afa/DrDAEC-infected intestinal cells were treated with L. acidophilus LBCFCS at a concentration that did not affect the viability of Afa/DrDAEC, the expression of brush border-associated functional hy-drolases SI, DPP IV, and AP and fructose transporter GLUT5 wasnormal, in contrast to the dramatic loss of expression observed inuntreated Afa/Dr DAEC-infected cells (125) (Fig. 4). The secreted

autotransporter toxin, Sat, belonging to the subfamily of SPATEtoxins (170), expressed by Afa/Dr DAEC strain C1845 (206) pro-motes an increase of fluid dome formation in Caco-2/TC7 cellmonolayers by modifying the transcellular passage of fluids. L.acidophilus LB CFCS treatment of Caco-2 cell monolayers resultsin the disappearance of the Sat-induced fluid dome formation,indicating a regulatory effect on the intestinal transcellular path-way of fluids (207).

It has been clearly demonstrated that the structural and func-tional lesions triggered by some enterovirulent bacteria resultfrom virulence factors, including bacterial effectors secreted bythe T3SS, and toxins, which hijack the cell signaling pathways thatcontrol the polarized organization of the cell cytoskeleton andderegulate the activities of functional membrane-associated pro-teins that have specific intestinal functions (162–164, 205, 208).Lactobacillus secreted compounds have been found to downregu-late the expression of virulence factors acting at the brush border(145, 146). Other antagonistic activities observed using entero-cyte-like cellular models also certainly result from regulatory ef-fects by these secreted compounds at the cellular signaling path-way level. It is noteworthy that similar regulatory effects have beenobserved experimentally for several probiotic Lactobacillus strainswhen intestinal functions were impaired in a noninfectious con-text (209–213).

Effects at the epithelial junctional domain. The intestinal bar-rier is kept closed by three intercellular junctional complexes: tightjunctions (TJs), adherent junctions, and desmosomes (214). Inparticular, the TJs form a highly regulated structure that acts as a“fence” separating the apical and basolateral membrane domainsof polarized cells, thereby segregating the various functional pro-teins in each of the domains. The TJs also function as a “gate”which allows paracellular vectorial transport to occur across theepithelial cell barrier. The junctional domain of the intestinal ep-ithelium is targeted by enterovirulent bacteria (173). Only L. rh-amnosus GG and L. casei DN-114 001 have been shown to havecytoprotective effects against the enterovirulent bacterium-in-duced structural and functional injuries at the intestinal junc-tional domain produced by enteric pathogens (Table 1). In T84cell monolayers, Johnson-Henry et al. (215) observed that theEHEC-induced changes in electrical resistance, dextran permea-bility, and distribution and expression of claudin 1 and zonulaoccludens 1 (ZO-1) are antagonized by live L. rhamnosus GG butnot by the heat-inactivated bacteria. In T84 cells, L. casei DN-114001 abrogates the EPEC strain E2348/69-induced increase in para-cellular permeability and rearrangements of ZO-1 in a dose-de-pendent manner (186). The antagonistic activity of L. acidophilusLB CFCS against the Sat-induced increase in paracellular perme-ability and formation of fluid-formed domes in enterocyte-likeCaco-2/TC7 cell monolayers (207) probably includes an effect atthe TJs, since the toxin affects the structural TJ organization (206).Consistent with this hypothesis, it has been observed that L. aci-dophilus LB culture protects TJs of cultured human intestinalHT-29 cells by counteracting the aspirin-induced delocalizationof structural TJ-associated ZO-1 protein (211).

Activation of host epithelial defense responses. Enteroviru-lent bacteria have the capacity to induce a host-controlled inflam-matory response which includes release of inflammatory cyto-kines such as interleukin-6 (IL-6), IL-8, IL-1�, tumor necrosisfactor alpha (TNF-�), and TNF-�. IL-8 is involved in the trans-migration of polymorphonuclear leukocytes across the intestinal



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

barrier, which initiates deleterious inflammatory cellular lesions(216). Other pathogen-induced dramatic proinflammatory re-sponses are more deleterious for the host; for example, the highlyharmful Shigella, causing bacillary dysentery in humans, has thecapacity to cause the inflammatory destruction of the intestinalepithelium (40). Probiotic Lactobacillus strains have been shownto antagonize the pathogen-induced production of proinflamma-tory cytokines (75). In addition, probiotic Lactobacillus strains areable to increase the production of molecules generated by hostintestinal epithelial cells which are major players in the first line ofhost defenses against enterovirulent bacteria, such as AMPs andmucins (217).

In enterocyte-like Caco-2 cells, the NF-�B-dependent flagel-lin-induced increase in IL-8 production was antagonized by both

live and UV-inactivated L. rhamnosus GG (218) (Table 1). In cul-tured human enterocyte-like HT-29 and crypt colonic T84 cells, L.rhamnosus GG decreased the production of IL-8 after V. cholerae,Salmonella, and EHEC infection but, surprisingly, did not signif-icantly alter the Shigella-induced IL-8 production (219, 220). L.casei Shirota CFCS decreased Salmonella enterica serovar Enterit-idis-induced IL-8 production and, in addition, promoted the ex-pression of cytoprotective heat shock protein (Hsp) 70 in Caco-2cells (221). L. rhamnosus GG suppressed the expression of CCL20and CXCL10 triggered in Caco-2 cells by effector molecules, pep-tidoglycan or flagellin, of enterovirulent E. coli (222). In Caco-2BBe cells, in which EHEC infection induces the upregulation ofproinflammatory genes, preincubation of the pathogen with L.rhamnosus GG prior to infection reduced the EHEC-induced up-

FIG 4 Normal expression of structural or functional brush border-associated proteins in L. acidophilus LB-treated, Afa/Dr DAEC strain C1845-infected culturedhuman enterocyte-like Caco-2/TC7 cells. (A) Transmission electron micrographs show the well-ordered microvilli in uninfected cells and the disappearance ofthe brush border in C1845-infected cells. (B) Confocal laser microscopy scanning examination of immunolabeling of brush border-associated structural F actinand functional sucrase-isomaltase (SI) in uninfected, C1845-infected, and L. acidophilus strain LB-treated C1845-infected Caco-2/TC7 cells. Micrographs on theleft show the normal mosaic pattern of distribution of fluorescein isothiocyanate (FITC)-labeled F actin (green) and rhodamine isothiocyanate (RITC)-labeledSI (red) in uninfected Caco-2/TC7 cells. Central micrographs show the disappearance of FITC-labeled F actin (green) and RITC-labeled SI (red) located centrallyin the cells and the persistence of the proteins at the cell-to-cell contacts. The micrographs on the right show that L. acidophilus strain LB-treated C1845-infectedcells conserve the normal mosaic pattern of the FITC-labeled F-actin (green) and RITC-labeled SI (red) distribution. For panel B, cells were apically infected withC1845 alone or in the presence of L. acidophilus strain LB CFCS (0.5-fold concentrate, which does not induce a bactericidal effect). (The micrographs in panelsA and B are reproduced from reference 125 with permission from BMJ Publishing Group, Ltd.)



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

regulation of the CXCL1, CXCL8, and NF-�B1A genes (223). Taoet al. (210) identified a heat-stable, low-molecular-weight peptideas the L. rhamnosus GG factor inducing the p38 and Jun N-termi-nal protein kinase (JNK) mitogen-activated protein kinase(MAPK)-dependent expression of cell regulatory Hsps 25 and 72in cultured mouse colonic YAMC cells. L. casei DN-114 001 triggeredan NF-�-dependent downregulation of the transcription of genes en-coding proinflammatory effectors and adherence molecules in Shi-gella flexneri-infected Caco-2 cells (224). Moreover, a DNA mi-croarray analysis conducted with distal duodenal mucosa biopsysamples from human patients receiving L. rhamnosus GG revealedthat 334 genes were upregulated, including genes that are involvedin immune and infectious responses, and that 92 genes weredownregulated (225). In mice, L. casei Shirota enhanced the ex-pression of genes acting in defense and immune responses (226).Altogether, these results indicated that these Lactobacillus strainsproduced effectors having regulatory activities on the host signal-ing pathways activated in response to enterovirulent infection andon the host signaling pathways involved in intestinal mucosal de-fenses.

Only L. rhamnosus GG has been shown to promote the produc-tion of intestinal mucus. L. rhamnosus GG mediates the upregulationof epithelial mucin MUC2 and MUC3 mRNAs or proteins in Caco-2cells and HT-29 cells (227, 228), which is accompanied by a concom-itant inhibition of adhesion of EPEC and EHEC (228).

Effects against Rotavirus

Human rotavirus is associated with 450,000 deaths each year,mainly in developing countries and among children under 5 yearsof age (229). Infection is characterized by a spectrum of responsesthat can vary from asymptomatic to mild or severe symptoms andcan result in a lethal dehydrating illness. Via either the entire virusor specific structural and nonstructural proteins, including theenterotoxin NSP4, the rotavirus induces substantial structuraland functional intestinal mucosa lesions that lead to secretorydiarrhea (229). Antirotavirus activities of probiotic Lactobacillusstrains have been poorly investigated in vitro. Secreted com-pounds produced by the probiotic strains L. rhamnosus GG, L.johnsonii NCC 533, and L. acidophilus LB do not directly affect therotavirus, since the pretreated rhesus rotavirus (RRV) strain rep-licates normally, in the same way as untreated RRV, in the Africangreen monkey MA104 epithelial cells used classically to maintaininfective rotavirus strains in culture (unpublished data). It hasbeen reported that soluble factors released by L. casei DN-114 001block the infection of cultured human mucus-secreting HT29-MTX cells by rotavirus RF and WA strains (230). In cultured pigand human epithelial cells infected with rotavirus, Maragkoudakiset al. (231) observed that the rotavirus-induced release of reactiveoxygen species was decreased in the presence of either L. rhamno-sus GG or L. casei Shirota. In a nontransformed porcine jejunumepithelial cell line, the presence of L. rhamnosus GG resulted in areduced rotavirus-induced IL-6 response (232). Rotavirus in-duces signaling-dependent structural and functional lesions at thebrush border and junctional domains of human enterocyte-likeCaco-2 cells similar to those observed in intestinal biopsy speci-mens from children with rotavirus-associated acute diarrhea(233). It has been difficult to investigate experimentally the impactof probiotic Lactobacillus strains with known antirotavirus thera-peutic effects on the rotavirus-induced structural and functionallesions in Caco-2 cells because of the cellular toxicity of Lactoba-

cillus CFCSs when the cells are infected long enough for all thereplications and stages of the intracellular lifestyle of the rotavirusto occur (unpublished data). To overcome this problem, furtherinvestigations can be conducted using fractionated LactobacillusCFCSs.

Effects against H. pylori

Probiotic Lactobacillus strains display antagonistic activitiesagainst the H. pylori cell association or cell responses accompany-ing H. pylori infection (Table 1). Secreted GroEL of L. johnsoniiNCC 533 has the capacity to generate the aggregation of H. pylori(234), a phenomenon observed on the surface of H. pylori-in-fected cultured human gastric epithelial cells (160). L. rhamnosusGG inhibits the adhesion of H. pylori onto AGS gastric cells, aneffect that is abolished by heat treatment of the Lactobacillus strain(235). When human mucus-secreting HT29-MTX cells were in-fected with H. pylori, the adhesion of the pathogen was inhibitedin a dose-dependent manner by L. acidophilus LB CFCS (139).Moreover, L. acidophilus LB CFCS treatment resulted in the deathof the adhering H. pylori, and any remaining adhering H. pyloricells displayed lower urease activity and, when observed by scan-ning electron microscopy, appeared to have undergone lysis asdetermined by cell morphology (139) (Fig. 2). In contrast, L. rh-amnosus GG CFCS-treated H. pylori adheres normally to HT29-MTX cells and displays normal urease activity (139). Conversely,L. rhamnosus GG enhanced the H. pylori-induced barrier injuryfollowing prolonged incubation (236). In human adenocarci-noma AGS cells, L. johnsonii NCC 533 (149) and L. rhamnosus GG(235, 236) decreased the H. pylori-induced IL-8 production.

ACTIVITIES IN ANIMAL INFECTION MODELS

Several antibacterial activities of probiotic Lactobacillus strainsobserved in vitro have been also observed in animal infectiousmodels (Table 2). In addition, activities of certain probiotic Lac-tobacillus strains against rotavirus infection have been examinedin rotavirus-infected animals. These studies have been conductedin classical rodent infectious models, including conventional oraxenic rodents. It was interesting to note that some other animalmodels are promising to study probiotic activities. Citrobacter ro-dentium-infected conventional mice mimicked the deleteriousstructural and functional cellular lesions of the human diarrhea-associated EPEC and EHEC (237, 238). The conventional strep-tomycin-pretreated mice infected with S. Typhimurium estab-lished by Hardt and coworkers (239, 240) mimicked the S.Typhimurium-induced cell deleterious effects. The impact of pro-biotics on the pathogen-induced deleterious effects on intestinalepithelial cells and immune responses has rarely been investigatedin these two models (241, 242). Caenorhabditis elegans is anemerging model to study microbial pathogenesis (243) and alsofor studying microorganisms that have implications for humanhealth (244). This model seems useful for the elucidation of themechanisms by which probiotics combat enteric pathogens (245–248) and may influence quality of life.

Bacterium-Infected Animals

Once human intestinal microbiota Lactobacillus probiotic strainsare established in the gastrointestinal tracts of axenic animals, theyreduce the association of gastric or enterovirulent bacterial patho-gens with the gastric and intestinal epithelia, inhibit the translo-cation of pathogens, lower cytopathic effects on epithelial cells,



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

stimulate cellular defense responses, and increase the survival ofinfected animals. Administration of heat-treated L. acidophilus LBcultures increases the survival of ETEC-infected suckling micecompared to that of their untreated counterparts (249). In new-born rat pups, L. rhamnosus GG decreases intestinal colonizationby enteroinvasive E. coli (250). In rabbits, administration of L.casei Shirota reduces the severity of diarrhea, lowers Shiga toxin-producing E. coli intestinal colonization, and decreases the intes-tinal concentrations of the Stx1 and Stx2 toxins and lowers theassociated histological damage (251). In pigs infected with anETEC strain, administration of L. rhamnosus GG reduces the du-ration of diarrhea (252).

Establishment of L. rhamnosus GG in the guts of axenic micechallenged with S. Typhimurium strain C5 results in lower cecalcolonization levels, a reduced translocation rate of the pathogen,and survival rates higher than those of noncolonized mice (130).L. johnsonii NCC 533 displayed its antibacterial activity in axenicmice orally infected by S. Typhimurium, since the level of Salmo-nella was lower in the feces of the treated conventional mice, andthe establishment of L. johnsonii NCC 533 in the guts of axenicmice resulted in improved survival (120). Moreover, decreasedintestinal colonization and translocation of C. jejuni caused by aheat-treated L. acidophilus LB culture has been observed in axenicmice (253). It is possible but not demonstrated that when probi-otic Lactobacillus cells colonize the intestinal epithelia of axenicmice, the observed in vivo antagonistic effect against Salmonellaresults in a local production of antibacterial molecules having abactericidal activity observed in vitro (120, 130).

In conventional mice orally infected with S. Typhimuriumstrain C5, oral administration of L. rhamnosus GG leads to a lowerlevel of the pathogen in the feces than in that of untreated mice

(130). In conventional rats orally receiving L. casei Shirota, there isa reduction in the number of L. monocytogenes organisms found inthe stomach, cecum, and feces, accompanied by reduced translo-cation from the intestine (254). In conventional mice, L. caseiShirota colonizing epithelial cells lining the stomach, small intes-tine, and colon and present in the intestinal contents reduced thelevels of S. Typhimurium associated with the epithelial cells of theduodenum and jejunum or present in the intestinal contents(128). It can be deduced from these experiments that when pro-biotic Lactobacillus cells colonize epithelial cells in different partsof the conventional mouse intestine (128), the adherent bacteriacreate a barrier effect against Salmonella similar to what is ob-served in vitro when probiotic bacteria attached to the brush bor-ders of enterocyte-like Caco-2 cells in culture inhibit attachmentand entry of Salmonella into cells (107). L. acidophilus LB CFCStreatment displayed antibacterial activity in the conventionalC3H/He/Oujco mouse infected with S. Typhimurium strain C5,since lower levels of fecal excretion of S. Typhimurium were ob-served than in untreated mice (122). This suggests that moleculespresent in L. acidophilus LB CFCS that exert in vitro bactericidalactivity against this pathogen (122, 136, 137) are able to producetheir bactericidal effect in vivo. It is noteworthy that in conven-tional mice infected with S. Typhimurium there was an absence ofan antagonistic effect when the infected mice were treated withuncultivated deMan-Rogosa-Sharpe broth acidified at pH 4.5(107, 122), demonstrating that the pH effect observed in vitro(131) is irrelevant for probiotic activity in vivo.

L. casei Shirota displays an antagonistic activity against the mul-tidrug-resistant S. Typhimurium strain DT104 in a fosfomycin-treated murine model, since the intestinal growth of the pathogenand its subsequent lethal extraintestinal translocation were inhib-

TABLE 2 Overview of antibacterial effects of probiotic Lactobacillus strains isolated from human intestinal microbiota against gastric orenterovirulent bacterial pathogens in animal infection models

Pathogen Probiotic strain Animal model Observed effect(s) Reference

Listeria L. casei Shirota Conventional rats Decrease of translocation from the intestine 254

Enterovirulent E. coli L. acidophilus LB Suckling mice Increase of survival rate 249Campylobacter jejuni L. acidophilus LB Axenic mice Decrease of mucosa-associated bacteria and bacterial

translocation253

S. Typhimurium L. rhamnosus GG Conventional mice Decrease of viable bacteria in feces 130Axenic mice Decrease of intestinal colonization and increase of

survival rate130

Newborn rat pups Decrease of intestinal colonization 250L. johnsonii NCC 533 Conventional mice Decrease of viable bacteria in feces 120

Axenic mice Decrease of intestinal colonization and increase ofsurvival rate

120

Conventional mice Decrease of bacteria associated with the intestinaltissues or present in the intestinal contents

128

L. casei Shirota Conventional mice Decrease of bacteria associated with the intestinaltissues or present in the intestinal contents

128

Fosfomycin-treated mice Decrease of intestinal colonization and translocationof a multiresistant strain

135

L. acidophilus LB Conventional mice Decrease of fecal excretion 122

H. pylori L. johnsonii NCC 533 Conventional mice Decrease of stomach colonization 149Mongolian gerbils Decrease of stomach colonization and gastritis 160

L. casei Shirota Conventional mice Decrease of stomach colonization and gastritis 149L. acidophilus LB Conventional mice Decrease of stomach colonization, urease activity in

stomach, and gastritis139



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

ited in a manner correlated with intestinal colonization by theLactobacillus strain (135). This observation is interesting in thelight of the currently increasing public health problem of multi-drug-resistant enterovirulent bacteria (255, 256) and persistercells (257, 258).

Administration of L. johnsonii NCC 533 to H. pylori-infectedmice reduced the inflammatory infiltration into the stomach lam-ina propria, although the level of colonizing H. pylori was notdecreased (149). Similarly, H. pylori colonization and gastritiswere significantly less intense in L. johnsonii NCC 533-treatedMongolian gerbils than in untreated animals (160). Oral admin-istration of L. casei Shirota in mice previously infected with H.pylori resulted in significantly lower levels of H. pylori colonizationin the antrum and body mucosa and a reduction in the gastricmucosal inflammation compared with those in the H. pylori-in-fected untreated group (149). Oral treatment of conventionalmice with concentrated L. acidophilus LB CFCS antagonized He-licobacter felis infection in mice by inhibiting gastric colonizationand decreasing H. felis urease activity, which in turn prevented thedevelopment of gastric inflammation. These effects were not sup-pressed by subjecting L. acidophilus LB CFCS to heat treatment(139). L. rhamnosus GG has been found to enhance gastric ulcerhealing in a rat model by decreasing cell apoptosis and increasingangiogenesis (259).

The exact mechanism(s) by which the probiotic Lactobacillusstrains exerted their antagonistic activities in vivo against infectinggastric or enterovirulent pathogens remains to be identified. Tworeports have presented evidence for specific mechanisms devel-oped by a probiotic Lactobacillus strain and a probiotic E. colistrain. It has been postulated that probiotic strains may competeagainst bacterial pathogens in intestinal ecological niches. It is welldemonstrated that bacteriocins produced by Gram-positive andGram-negative probiotic bacteria inhibit strains closely related tothe producer strain (43). Consequently, it is thought that thesepeptides may assist the producers to compete within their specificintestinal ecological niches. This has recently been demonstratedfor a Gram-negative probiotic strain. Deriu et al. (260) haveshown that when the probiotic Gram-negative E. coli Nissle strain(261) was administered to mice with S. Typhimurium-inducedcolitis, there was an impressive reduction in S. Typhimurium col-onization as a result of successful competition of the probioticstrain with Salmonella for iron acquisition via non-enterobactin-mediated pathways. Moreover, the production of a bacteriocin invivo by L. salivarius UCC118 can efficiently protect mice againstinfection by the invasive L. monocytogenes (262). Such ecologicalcompetition between the six Lactobacillus probiotic strains exam-ined and Gram-negative enteric bacterial pathogens remains to bedemonstrated.

Rotavirus-Infected Animals

There are few studies that have analyzed the protective effect of thesix probiotic Lactobacillus strains against rotavirus infection inanimal models. Neonatal mice and rats provide reliable animalmodels for studying the kinetics of viremia, spread, and pathologyof rotavirus and also immune responses during a primary rotavi-rus infection (229). In rotavirus-infected rodent models, L. rham-nosus GG reversed the rotavirus-induced increase in intestinalbarrier permeability (263), reduced both the duration and theseverity of the resulting diarrhea and the histopathologicalchanges and virus load in the intestine in combination with anti-

rotavirus antibodies (264), and shortened the duration of diarrheaand decreased epithelium vacuolation in the jejunum (265). Ingermfree suckling rats receiving milk fermented by the L. caseistrain DN-114 001 and infected with rotavirus, the frequency ofstools and severity of diarrhea were reduced, as were intestinal celllesions, including cell vacuolation and changes in the morphologyof the intestinal villi (266). In rotavirus-infected mice, oral admin-istration of L. reuteri DSM 17938 reduced the duration of diarrheaand decreased the accompanying intestinal cell lesions (267).

CLINICAL STUDIES

The major forms of the six probiotic Lactobacillus strains availableto consumers consist principally of yogurt or bottled fermentedmilk, but capsules or sachets containing the lyophilized strain inpowder form for rehydration prior to consumption and chewabletablets are also available as dietary supplements (268). In addition,several probiotic strains are now included in dietetic products forchildren (269). The RCTs conducted in children and adults usinghuman probiotic Lactobacillus strains have been intervention stud-ies in which various vehicles containing the probiotic Lactobacillusstrains have been used (Tables 3 and 4). Those conducted to in-vestigate the anti-infectious therapeutic efficacy of L. rhamnosusGG have been carried out mainly using lyophilized powders ofLactobacillus cells or fermented milk containing the probioticstrain. RCTs designed to evaluate the anti-infectious clinical effi-cacy of the probiotic strains L. casei Shirota and L. casei DN-114001 have been conducted using Lactobacillus-containing milkdrinks. Drinkable whey-based or commercial dairy products con-taining L. johnsonii NCC 533 have been used to evaluate in RCTsthe anti-H. pylori therapeutic efficacy of this probiotic strain. Forthe L. acidophilus LB, the forms used in RCTs are sachets or cap-sules containing a combination of 10 billion heat-treated and ly-ophilized L. acidophilus LB cells and 160 mg of 2-fold-concen-trated CFCS (Lacteol).

Human probiotic Lactobacillus strains survived after oral ad-ministration in laboratory animals and were present at apprecia-ble levels in the gastrointestinal tract. For example, in conven-tional or germfree mice, L. rhamnosus GG cells have been found inthe different segments of the gut (130), and L. johnsonii NCC 553and L. casei Shirota cells efficiently colonize the different parts ofthe conventional mouse intestine (128). In human adults and in-fants receiving L. rhamnosus GG, the strain has been found in fecaland/or intestinal mucosa biopsy samples (270–275). L. casei Shi-rota cells have been found in the feces of human volunteers receiv-ing fermented milk containing L. casei Shirota (276–278). L. caseiDN-114 001 cells have been found in ileal and fecal samples ofhuman volunteers receiving fermented milk containing L. caseiDN-114 001 or the derived L. casei strain DN-114 001Rif (279,280). In human adult volunteers and infants, L. reuteri ATCC55730 cells have been found in fecal samples (275, 281, 282). It isnoteworthy that probiotic Lactobacillus strains do not colonize thegut and that the persistence of a strain is limited to the duration ofthe administration of the probiotic preparation. Indeed, as soon asthe probiotic strain is no longer consumed, probiotic cells disap-pear from the intestinal tract of the consumer.

Therapeutic Effects against Various Forms of AcuteDiarrhea

The World Health Organization (WHO) has defined diarrhea asthe occurrence of three or more loose or watery stools within a



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

24-h period. Diarrhea is described as acute when it began less than14 days before and as persistent when its duration is 14 days ormore. Rotavirus is worldwide a major cause of severe diarrhea andof diarrhea leading to mortality in children. Other diarrhea-asso-ciated viruses include astroviruses, noroviruses, sapoviruses, andenteric adenoviruses. The major bacterial pathogens involved indiarrhea are ETEC, Salmonella, Shigella, Yersinia, Campylobacter,and Vibrio cholerae. The major causes of acute infectious diarrheadue to enteric viruses or enterovirulent bacteria vary from place toplace, and there are relatively few vaccines available to preventhuman mucosal infections (283). Rotavirus vaccines (284–286)are effective in preventing rotavirus-associated acute diarrhea in

all countries (287–289) and particularly in those in which highmortality is attributable to rotavirus-induced diarrhea (290, 291).Several interventions, including improved sanitation and handhygiene and the promotion of breast-feeding, have been clinicallyused for the prevention of rotavirus-induced acute diarrhea in chil-dren (292). Such treatment is intended to prevent the dehydrationand nutritional damage and to reduce the length and severity of acutediarrhea. To prevent dehydration and nutritional damage in childrenand infants with acute diarrhea, the therapeutic regimen recom-mended by the WHO, the European Society for Pediatric InfectiousDiseases (ESPID), and the European Society for Pediatric Gastroen-terology, Hepatology, and Nutrition (ESPGHAN) is to provide an

TABLE 3 Overview of therapeutic effects against acute or persistent diarrhea in RCTs of probiotic Lactobacillus strains isolated from humanintestinal microbiota

Probioticstrain Cause of infection

No. (age group)of patients Treatment Reported clinical effect(s) (control/treated)a Reference

L. rhamnosusGG

Not documented 71 (children) Fermented milk, 109 CFU/ml for 5days

Shortened duration of acute diarrhea (2.4/1.4 days) 296

Rotavirus 42 (children) Freeze-dried powder, 1010 CFU/mltwice daily for 5 days

Lowering of no. of patients with acute diarrhea (43/10 at day 3 of treatment)

297

Rotavirus 49 (children) Freeze-dried powder, 1010-1011 CFU/ml twice daily for 5 days

Shortened duration of acute diarrhea (2.7/1.8 days)accompanied by stimulation of rotavirus-specificIgA antibody responses

299


Lowering of no. of patients with acute diarrhea (75/31% at day 2 of treatment)

300


Shortened duration of acute diarrhea (3.3/1.9 days) 301

Rotavirus (28%) and enterovirulentbacteria (21%)

123 (children) Freeze-dried powder, 5 � 109 CFU/ml twice daily for 5 days with ORS

Shortened duration of acute diarrhea (3.7/2.7 dayswith a decrease of frequency of stools

302

Rotavirus 123 (children) Freeze-dried powder, 5 � 109 CFU/ml twice daily with ORS

Shortened duration of acute diarrhea (30.4/17.7 h) 303

Rotavirus and enterovirulent bacteria 204 (children) 3.7 � 1010 CFU/ml once daily for 6days/wk for 15 mo

Prophylactic effect on the incidence of diarrhea 304

Rotavirus 287 (children) Freeze-dried powder, 1010 CFU/mlwith ORS until diarrhea stopped

Shortened duration of acute diarrhea (76.6/57.2 h) 305

Rotavirus 81 (children) 6 � 109 CFU/ml twice daily forduration of hospital stay

Prophylactic effect on the incidence of diarrhea 331

Rotavirus (27.5%) and enterovirulentbacteria (36%)

179 (infants) Fermented milk, 109 CFU/ml per daywith ORS

No significant differences in duration of diarrhea,rate of treatment failure, and proportion ofunresolved diarrhea

314

Not documented 192 (children) 6 � 109 CFU/ml per day with ORS Shortened duration of acute diarrhea (115.5/78.5 h) 306Not documented 662 (children) 6 � 1010 CFU/ml per day with ORS No effect on duration of diarrhea (6.8/6.6 days) 313Rotavirus and enterovirulent bacteria 235 (children) 6 � 1010 CFU/ml per day with ORS Shortened duration of persistent diarrhea

(9.2/5.3 days)307

Not documented 559 (children) 6 � 1010-1012 CFU/ml per day withORS

Decrease of frequency of stools and duration ofdiarrhea

308

Not documented 229 (infants) 109 CFU/ml per day for 10 days No effect on duration of diarrhea or numbers ofstools

315

Rotavirus and enterovirulent bacteria 64 (children) 5 � 109 CFU/ml three times per dayfor 3 days with ORS

No change in duration of diarrhea, total stools, ordiarrhea score

316

L. casei DN-114 001

Not documented 287 (children) Fermented milk, 108 CFU/ml daily Shortened duration of acute diarrhea (8.0/4.3 days) 338Not documented 928 (children) Fermented milk, 108 CFU/ml daily Decrease of no. of patients with acute diarrhea

(22.0/15.9%)337

L. reuteri DSM17938

Rotavirus 66 (children) 1010-1011 CFU/ml daily for up 5 days Shortened duration of acute diarrhea (2.9/1.7 days) 317Rotavirus 40 (children) 107-1010 CFU/ml daily for up to 5

daysShortened duration of acute diarrhea (2.5/1.7 days)

and no. of patients with acute diarrhea (80/48%)282

Rotavirus 74 (children) 4 � 108 CFU/ml daily Decrease of no. of patients with acute diarrhea(82/45%)

319

Heat-treated L.acidophilusLB cultureb

Rotavirus 73 (children) 6 sachets with ORSc Shortened duration of acute diarrhea (74.0/42.9 h) 325Enterovirulent bacterium-induced

acute diarrhea80 (children) 6 sachets during 35 h with ORSc Shortened duration of acute diarrhea (30.4/8.2 h) 327

Enterovirulent bacterium-inducedacute diarrhea

80 (children) 8 sachets during 96 h with ORSc Shortened duration of acute diarrhea (63.4/39.5 h) 147

a Number of days of acute diarrhea or duration of diarrhea or number or percentage of patients with acute diarrhea.b Lacteol.c One sachet consisted of 10 billion heat-treated and lyophilized L. acidophilus LB cells and 160 mg of 2-fold-concentrated neutralized CFCS.



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

oral rehydration solution (ORS) and to continue feeding (293). Al-though ORS administration effectively mitigates dehydration, it hasno effect on the length and severity of diarrheal episodes. A variety ofadjuvant therapies have been used to treat the infectious origin ofacute diarrhea (294). Luminally acting antidiarrheic drugs are con-fined mainly to the gastrointestinal tract, but several of these agentscan cause adverse effects outside the gastrointestinal tract. Moreover,

several problems resulting from the use of antibiotic therapy to treatdiarrhea associated with enterovirulent bacteria have been observed,including the emergence of resistance to single antibiotics, followedby the emergence of multiple resistance and, if broad-spectrum anti-biotics are used, a adverse effect on the intestinal microbiota, whichcan in turn favor the emergence of C. difficile-associated diarrhea(295).

TABLE 4 Overview of therapeutic effects against H. pylori infection in RCTs of probiotic Lactobacillus strains isolated from human intestinalmicrobiota

Probiotic strain

No. (agegroup) ofpatients Treatment Associated treatment Reported clinical effects (control/treated) Reference

L. rhamnosus GG 120 (adults) 109 CFU/ml twice daily for 14days in conjunction withtriple therapy

Pantoprazole, 40 mg; clarithromycin, 500 mg;tinidazole, 500 mg

No improvement of H. pylorieradication rate

382

43 (adults) 6 � 109 CFU/ml twice daily for 7days after 1 wk of tripletherapy

Rabeprazole, 20 mg; clarithromycin, 500 mg;tinidazole, 500 mg (daily)


383

47 (adults) Probiotic prepn including L.rhamnosus GG (109 CFU/ml)twice daily during tripletherapy and once a dayduring 3 wk after cessation oftriple therapy

Lansoprazole, 30 mg; clarithromycin, 500 mg;amoxicillin, 1,000 mg (daily)


384

83 (children) 109 CFU/ml twice daily for 7days in conjunction withtriple therapy

Amoxicillin, 25 mg; clarithromycin, 10 mg;omeprazole, 0.5 mg (twice daily)


385

L. johnsonii NCC533 (La1)

20 (adults) 50 ml of drinkable, whey-based,CFCS four times daily for 14days in conjunction withmonotherapy

Omeprazole, 20 mg four times daily Decrease in breath test values 150

53 (adults) 180 ml fermented milk twicedaily for 3 wk

Clarithromycin, 500 mg during the last 2 wkof milk therapy

Improvement of clarithromycin-inducedH. pylori eradication by decreasing H.pylori density and gastritis in stomach.

373

50 (adults) Fermented milk (LC1a) twicedaily for 16 weeks

No anti-H. pylori therapy No cure of H. pylori infection, decreaseof inflammatory score (6.3/5.3)

374

12 (adults) 80 ml fermented milk (LC1)(107 CFU/ml) 8 times dailyfor 2 weeks

No anti-H. pylori therapy Decrease in breath test values 375

100 (children) 2 � 80 ml fermented milk (LC1)(107 CFU/ml) for 4 wk


136 (children) 80 ml fermented milk (LC1)(107 CFU/ml) daily for 2weeks


L. casei Shirota 14 (adults) Drinkable fermented milkb (108

CFU/ml) for 3 wkNo anti-H. pylori therapy Decrease in breath test values 378

64 (adults) Drinkable fermented milkb (108

CFU/ml) for 8 wkClarithromycin, amoxicillin, and omeprazole Improvement of triple-therapy-induced

H. pylori eradication379

L. casei DN-114001

86 (children) Drinkable fermented milkc for14 days

Omeprazole, amoxicillin, and clarithromycin Improvement of triple-therapy-inducedH. pylori eradication (57.5/84.6%)

380

L. reuteri DSM17938

40 (adults) Chewable tabletsd (108 CFU/ml)daily for 28 days

After probiotic treatment: rabeprazole (20 mgtwice a day) plus amoxicillin (1 g, twicedaily) for 5 days followed by rabeprazole(20 mg twice a day), clarithromycin (500mg, twice daily), and tinidazole (500 mg,twice daily) for the next 5 days

Decrease in breath test values at the endof probiotic treatment, no differencein H. pylori eradication rates aftertriple therapy or not

386

Heat-treated L.acidophilus LBculturee

120 (adults) 1 sachetf daily for 7 days and 3days after cessation of tripletherapy

Rabeprazole (20 mg), clarithromycin (250mg), and amoxicillin (500 mg) for 7 days

Improvement of triple-therapy-inducedH. pylori eradication (72/88%)

381

84 (adults) 2 sachets daily for 4 wk Omeprazole (20 mg) and amoxicillin(1,000 mg)

No improvement of the low H. pylorieradication rate of double therapy

387

120 (children) 1 sachet daily for 8 wk No anti-H. pylori therapy No decrease in breath test values 388

a Fermented milk LC1 (Nestlé Company).b Yakult drink (Yakult Company).c Actimel drink (Danone Company).d BioGaia AB.e Lacteol.f One sachet consisted of 10 billion heat-treated and lyophilized L. acidophilus LB cells and 160 mg of 2-fold-concentrated L. acidophilus LB spent culture medium.



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

Acute infectious diarrhea. Human probiotic Lactobacillusstrains and drugs derived from several of these strains have beenclinically investigated to treat acute infectious diarrhea (Table 3).In RCTs investigating their therapeutic efficacy, they have beengenerally used in association with ORS (293). In RTCs, L. rham-nosus GG alone or in combination with ORS has demonstratedtherapeutic efficacy in significantly reducing by at least half theduration of acute diarrhea mainly resulting from rotavirus infec-tion in children (296–310). Two meta-analyses have been con-ducted to specifically examine the RCTs conducted with L. rham-nosus GG (311, 312). The conclusion is that L. rhamnosus GG hadno impact on the total stool volume but did reduce the duration ofdiarrhea, particularly in the context of a rotavirus etiology. Incontrast, four trials in children and infants with enterovirulentbacterium- or rotavirus- and enterovirulent bacterium-induceddiarrhea showed no decrease in the daily number of stools orduration of diarrhea after treatment with L rhamnosus GG (313–316). Multicenter and other RCTs in children and infants haveshown that the administration of L. reuteri DSM 17938 (L. reuteriATCC 55730) shortened the duration of acute diarrhea and low-ered the diarrhea relapse rate (282, 317–319).

Heat treatment of probiotic Lactobacillus strains results in amarked decrease of antimicrobial activity (320) compared to thatof the parental live strains (135, 139, 215, 298). Heat-treated Lac-tobacillus strains cannot be considered to be probiotics (321), be-cause the definition of probiotics states that the strains are alive(69). However, a heat-treated Lactobacillus strain may retain thepharmacological “probiotic” activities of the parental live strainwhen the substances produced by the strain that underpin thepharmacological activities are able to resist the treatments to whichthey are subjected during the pharmaceutical manufacturing process,including heat treatment (322). L. acidophilus strain LB is one of thefew probiotic Lactobacillus strains which display this characteristicsince, as we have already mentioned, it produces heat-stableantimicrobial compounds. Lacteol has been authorized for useas an antidiarrheal agent by the French Safety of MedicinesAgency (marketing authorizations AMM 3400933073602, AMM3400934847837, and AMM 3400934840224) (Forest Laborato-ries, Inc., New York, NY). In three RCTs conducted with Lacteolin combination with ORS in infants and young children with ro-tavirus-associated diarrhea, the treatment shortened by at leasthalf the duration of diarrhea and accelerated the reappearance ofthe first normal consistent and formed stool compared to the pla-cebo combined with ORS (323–325).

While the numerous experimental in vitro and in vivo datareported above demonstrated the bactericidal activity of L. rham-nosus GG against enterovirulent bacteria, it is both surprising andintriguing to note that therapeutic efficacy was absent in RCTsconducted with this probiotic strain in children with establishedbacterially induced diarrhea (302, 305, 314, 326). Consistent withthe in vitro and in vivo antibacterial activities reported above forlive and heat-treated L. acidophilus strain LB against enteroadher-ent and enteroinvasive bacteria (122, 137, 147, 181, 182), twoRCTs have demonstrated the therapeutic efficacy of Lacteol incombination with ORS in decreasing the frequency and duration(reduced by at least half) of enterovirulent bacterium-associatedacute watery diarrhea in children between the ages of 1 and 4 years(207, 327). Moreover, in adults over 16 years of age with chronicbacterium- or parasite-associated diarrhea, a prospective, ran-domized, multicenter clinical trial has demonstrated that admin-

istering Lacteol led to the recovery of consistent stools in 81% ofthe treated patients (328).

Nosocomial infections. It is noteworthy that the few preven-tion clinical studies conducted with probiotic Lactobacillus strainshave given disappointing results, even though the regular con-sumption of probiotic food products is often recommended onhealth grounds, including to prevent intestinal infections and toimprove quality of life (69, 111, 329). Contrasting results havebeen obtained in trials investigating the effect of probiotic Lacto-bacillus strains in preventing nosocomial infections. Hojsak et al.(330) observed a reduced risk of gastrointestinal infections in chil-dren after L. rhamnosus GG treatment. L. rhamnosus GG signifi-cantly reduced the occurrence of nosocomial diarrhea in infants,particularly in cases resulting from rotavirus infection (331). Incontrast, two RCTs in children have shown that the consumptionof L. rhamnosus GG was ineffective in preventing nosocomial di-arrheal episodes and also failed to reduce the number of days withacute diarrhea (332, 333). The occurrence of the primary episodesof acute diarrhea was significantly retarded in children receiving aprobiotic drink containing L. casei Shirota compared to untreatedcontrol children (334). In two multicenter RCTs investigating theeffect of L. casei DN-114 001 on overall common infectious dis-eases (CIDs), including gastrointestinal tract infections, it hasbeen observed that the consumption of a fermented dairy productcontaining L. casei DN-114 001 was associated with a shorter du-ration of CIDs in aging patients (335, 336). In contrast, a trial inhealthy children receiving yogurt containing L. casei strain DN-114 001 found that there was no reduction of the incidence ofacute diarrhea but that the period of acute diarrhea was lower thanthat in children receiving traditional yogurt (337, 338). A study inchildren found no difference between the probiotic L. reuteri DSM17938 group and the placebo group with regard to the incidence ofrotavirus infection, the incidence and duration of acute diarrhea,the incidence of chronic diarrhea, the duration of hospital stay indays, and the frequency of the need for rehydration (339).

Traveler’s diarrhea. Acute diarrhea induced by enterovirulentbacteria is common among travelers and tourists. Three RCTsinvestigating the efficacy of human probiotic Lactobacillus strainsin preventing intestinal infectious episodes in travelers gave neg-ative results. Two trials were conducted in adults and infants trav-eling in southern Turkey (given 2 � 109 L. rhamnosus GG cellsdaily for 1 to 2 weeks of travel) (340) and in adult American trav-elers visiting South America, the subcontinent of India, CentralAmerica, the Middle East, West Africa, and North Africa (givenone capsule [2 � 109 L. rhamnosus GG cells] once daily, starting 2days prior to departure and continuing throughout the travel)(273). The results show that the occurrence of acute diarrhea wasquite similar in the placebo group and the L. rhamnosus GG group(273, 340). Similarly, another study demonstrated a lack of pre-vention of infectious diarrhea by Lacteol (one capsule consistingof 10 billion heat-treated and lyophilized L. acidophilus LB cellsand 160 mg of 2-fold-concentrated L. acidophilus LB spent culturemedium twice daily from 1 day before their departure to 3 daysafter their return) compared to placebo in adult travelers visitingWest, East, Central and North Africa, Oceania, South America,Asia, Central America/the Caribbean, and the Middle East from2001 to 2004 (341). These studies do not prejudge the efficacy ofother probiotic Lactobacillus strains to prevent traveler’s diarrhea.



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

Therapeutic Effects against C. difficile-Associated Diarrhea

C. difficile is a Gram-positive anaerobic bacterium that is one ofthe most important causes of antibiotic-associated diarrhea in thedeveloped world, and discontinuation of antibiotics generallypromotes a cure of mild infections (342). The incidence and se-verity of C. difficile infection appear to be on the increase, leadingto significant morbidity and mortality and placing a considerableeconomic burden on health care systems, particularly in Europeand North America (343, 344). Recent severe C. difficile diseaseand higher mortality rates have been associated with the emer-gence of strains of increased virulence, or “hypervirulent” isolates,belonging to the BI/NAP1/027 category and which are fluoro-quinolone resistant (345). C. difficile produces a number of puta-tive virulence factors, including two members of the large clostrid-ial cytotoxin family, known as toxin A and toxin B, both of whichinduce intense colonic inflammation and dramatic structural andfunctional epithelial tissue damage followed by a rapid fluid lossinto the intestinal lumen, leading to diarrhea and an adaptive im-mune response (342, 346).

The antagonistic activity exerted by probiotic Lactobacillusstrains against C. difficile has received little experimental investi-gation (118, 347). Despite this lack of experimental evidence, theuse of probiotic Lactobacillus strains to treat C. difficile-induceddiarrhea has been extensively promoted in a high number of re-views (348–352), but it remains controversial (353). Meta-analy-ses have evidenced that probiotic strains are associated with areduction in C. difficile-associated diarrhea (102, 354–357). Therewere a small number of reports showing persuasive therapeuticeffects against C. difficile-induced diarrhea after the administra-tion of a single probiotic Lactobacillus strain. RCTs have showntherapeutic efficacy of L. rhamnosus GG to treat C. difficile-in-duced diarrhea (102, 358–361). However, several reports haveshown a lack of therapeutic efficacy against C. difficile-induceddiarrhea for L. rhamnosus GG (362). There is a lack of reportsdocumenting the experimental and therapeutic effects of the pro-biotic L. casei Shirota YIT9029, L. acidophilus LB, L. johnsonii NCC533, L. casei DN-114 001, and L. reuteri DSM 17938 strains againstC. difficile infection.

Hell et al. (363) have suggested that a combination of differentprobiotic strains may be more useful to reduce C. difficile infec-tion. It is noteworthy that fecal microbiota transplantation (FMT)is an emerging treatment to restore the normal microbial homeo-stasis after antibiotic agents have disrupted the intestinal micro-biota, leading to C. difficile-associated diarrhea. Interestingly,Russell et al. (364) recently reported the success of FMT in a youngchild showing infection caused by C. difficile strain BI/NAP1/O27.This pathogenic strain is refractory to metronidazole, vancomy-cin, rifaximin, and nitazoxanide antibiotic treatments or treat-ment with L. rhamnosus GG. Kassam et al. (365) conducted ameta-analysis of 11 studies concerning patients with C. difficileinfection and treated with FMT and concluded that “FMT holdsconsiderable promise as a therapy for recurrent C. difficile infec-tion, but well-designed, RCTs and long-term follow-up registriesare still required.”

Meta-Analyses

The clinical trials discussed above all have methodological flaws,including having groups of patients that were extremely differentin size and with regard to age, differences in the concentrations of

probiotic bacteria administered and in the duration of adminis-tration, and differences in the vehicles containing the probioticstrains. Moreover, different outcomes to evaluate or quantita-tively measure the therapeutic efficacy have been used, such as theperiod of acute diarrhea or the time to the reappearance of a con-sistent stool. To mitigate these problems, meta-analyses have at-tempted to examine RCTs conducted with a wide variety of pro-biotic lactic acid-producing strains, such as L. rhamnosus strainGG, strains of L. acidophilus, L. casei, L. plantarum, L. fermentum,and L. bulgaricus, Bifidobacterium strains BB12 and BB536, B. in-fantis and B. lactis strains, and VSL3 (viable lyophilized bacteria ofL. paracasei, L. plantarum, L. acidophilus, L. delbrueckii subsp. bul-garicus, B. longum, B. breve, B. infantis, and a Streptococcus sali-varius subsp.), and other, non-lactic acid-producing probioticstrains (355, 366–368). Moreover, Allen et al. (369) have exam-ined the Cochrane Infectious Diseases Group’s trials register(2010), the Cochrane Controlled Trials Register (2010),MEDLINE (1966 to 2010), EMBASE (1988 to 2010), and lists ofreferences in reviews for analyzing RCTs and quasi-RCTs thatcompare probiotic treatment with no probiotic administration ortreatments with or without placebo administration in patientswith acute diarrhea. The authors of these meta-analyses concludethat “probiotic strains used alongside rehydration therapy appearto have clear beneficial effects in shortening the duration and re-ducing stool frequency in acute infectious diarrhea, although thesize of the effect varied considerably between studies.” Othermeta-analyses concluded that probiotic Lactobacillus strains sig-nificantly reduce C. difficile-associated diarrhea and acute diar-rhea of diverse causes (102, 357, 370). In contrast, two meta-anal-yses showed that probiotic Lactobacillus strains have no effect ontraveler’s diarrhea (355, 369). Moreover, there is an absence ofconvincing evidence that probiotics are effective for treating per-sistent diarrhea in children (371).

Therapeutic Effects against Infectious Gastritis

Combinations of drugs, including a proton pump inhibitor (PPI)plus three antibiotics or a PPI plus bismuth plus two antibiotics,provide the best therapeutic efficacy for treating gastric H. pyloriinfection (372). However, the effectiveness of many of these treat-ments has been compromised by an increase in the resistance of H.pylori to antibiotic treatment. Several human probiotic Lactoba-cillus strains have been shown to improve the efficacy of the anti-H. pylori triple therapy in well-conducted RCTs (Table 4). Oraladministration of L. johnsonii NCC 533 CFCS in omeprazole-treated patients induced a decrease in breath test values (150).Oral administration of acidified milk containing L. johnsonii NCC533 in clarithromycin-treated patients induced a decrease in H.pylori levels and reduced inflammation in the antrum and corpus(373). The intake of fermented milk containing L. johnsonii NCC533 without taking antibiotics resulted in the persistence of H.pylori but with a decreased inflammatory score (374) or breath testvalues (375). It was noted that heat treatment of L. johnsonii NCC533 affects its therapeutic efficacy (376, 377). Patients receiving afermented milk drink containing L. casei Shirota displayed lowerurease activity (378). Eradication of H. pylori after triple treatmentwith clarithromycin, amoxicillin, and omeprazole was betterwhen a fermented milk drink containing L. casei Shirota was com-bined than with the triple therapy (379). A multicenter RCTshowed that with adjunct of a drinkable fermented milk contain-ing L. casei DN-114 001 improved the rate of eradication of H.



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

pylori in children treated with amoxicillin, clarithromycin, andomeprazole (380). An RCT carried out by Canducci et al. (381)has shown that in patients with H. pylori infection and receiving atriple therapy based on rabeprazole, clarithromycin, and amoxi-cillin for 7 days, supplementation with Lacteol significantly in-creased the eradication of H. pylori (87% of patients) compared tothe triple therapy alone (72% of patients).

Negative results have been reported in four RCTs investigatingthe therapeutic efficacy of L. rhamnosus GG against H. pylori-induced infection in conjunction with pantoprazole, clarithromy-cin, and tinidazole (382), rabeprazole, clarithromycin, and tinida-zole (383), lanzoprazole, clarithromycin, and amoxicillin (384),or amoxicillin, clarithromycin, and omeprazole (385). An RCT inadults has shown that the administration of tablets containing L.reuteri DSM 17938 reduced the occurrence of dyspeptic symp-toms but did not improve H. pylori eradication (386). De Fran-cesco et al. (387) showed that Lacteol did not ameliorate the loweradication rate of an anti-H. pylori therapy composed of a protonpump inhibitor and amoxicillin. Gotteland et al. (388) evaluatedLacteol treatment alone versus triple therapy in an open, prospec-tive, randomized trial and observed that the treatment was rela-tively ineffective in eradicating H. pylori in contrast to the tripletherapy (lanzoprazole, clarithromycin, and amoxicillin).

CONCLUDING REMARKS

Despite the fact that a large number of Lactobacillus strains havebeen isolated from human microbiota and tested in vitro for theirprobiotic activities, such as their adhesiveness to intestinal cellsand mucus and antagonistic activities against gastroenteropatho-gens, only a few of these strains have displayed the propertiesneeded for industrial development to generate biotherapeutic an-ti-infectious agents (389). The experimental data and RCTs ana-lyzed above provide evidence of the following: (i) L. rhamnosusstrain GG in fermented milk and lyophilized forms, fermentedmilk containing L. casei strain DN-114 001, the lyophilized formof L. reuteri DSM 17938, and Lacteol containing lyophilized, heat-treated L. acidophilus LB cells and concentrated spent culture me-dium are therapeutically effective against rotavirus-associatedacute diarrhea when used in addition to ORS; (ii) only Lacteol hasbeen shown in RCTs to demonstrate therapeutic efficacy againstenterovirulent bacterium-induced acute diarrhea when used inaddition to ORS; and (iii) fermented milk containing L. johnsoniiNCC 533, L. casei Shirota, or L. casei DN-114 001 and Lacteol aretherapeutically effective in antagonizing gastritis-associated H.pylori or improving the eradication rate of the triple therapy. Incontrast, there is no clinical evidence that consumption of theseprobiotic strains prevents the occurrence of intestinal infectiousepisodes. The mechanism underlying these anti-infectious effectsappears to be multifaceted. Indeed, a large set of mechanisms havebeen identified, including (i) inhibition of the growth of patho-gens or a direct bactericidal effect exerted by secreted molecules,(ii) inhibition of expression of virulence genes coding for viru-lence factors or interference with the cell membrane expression orsecretion of virulence factors, including toxins, (iii) competitiveexclusion of pathogenic bacteria by competition for binding sites,(iv) inhibition of the signaling-dependent structural and func-tional deleterious effects triggered by virulence factors in host in-testinal cells, and (v) stimulation of antimicrobial host intestinalcell responses. It was noticed, interestingly, that experimental dataand RCTs have provided evidence that Lactobacillus species iso-

lated from the human vaginal microbiota developed anti-infec-tious properties against vaginosis-associated bacterial pathogens,including Prevotella bivia and Gardnerella vaginalis, by mecha-nisms similar to that of human intestinal microbiota Lactobacillusstrains (390). On the basis of the experimental and clinical evi-dence of the production of strain-specific derived bioactivemolecules by human microbiota Lactobacillus strains havingconcentration-dependent pharmacological activities such as anti-biotic-like activities and host cell regulatory activities, Shahanan etal. (391) have proposed that these bacterial molecules can be col-lectively termed “pharmabiotics.”

The increasing occurrence of drug-resistant bacterial patho-gens is currently one of the major public health challenges. Al-though medical practice has limited the development and spreadof pathogens, the rapid global emergence of pathogens resistant tomost conventional antibiotics and the recent emergence of mul-tidrug-resistant pathogens constitute an increasing major publichealth threat (256, 392). Moreover, antibiotic resistance resultsalso from the presence of dormant bacteria and persisters exhib-iting an extraordinary tolerance to antibiotics resulting from tran-sient growth inhibition and inactivity of essential cell functionscontrolled by a multiplicity of bacterially regulated mechanisms(258, 393). It is becoming increasingly evident that in order toreduce the rate of appearance of antibiotic-resistant strains, bettermanagement and a more reasonable use of antibiotics both inhumans and in animal husbandry are necessary. However, the lackof new antibiotics coming onto the market highlights the need todiscover new antimicrobial agents and, to do this, to develop in-novative strategies. Host intestinal antimicrobial molecules, in-cluding epithelial cell-produced AMPs and bacteriocin and non-bacteriocin molecules generated by microbiota Gram-positiveand Gram-negative bacteria, are potential sources of new antimi-crobial molecules. AMPs such as cationic AMPs, which have theadvantages of broad-spectrum activity and a lower propensity toselect for drug resistance phenotypes, have been intensely investi-gated as a potential source of complementary antimicrobial agents(9, 10, 394, 395). However, many pathogens are able to overcomehost AMPs. They often kill bacteria by membrane disruption,which is a highly effective and rapid mechanism (396). Impor-tantly, since human and bacterial membranes are similar, it isoften difficult to target the bacteria and avoid damaging the pa-tient’s cells. To make it possible to use membrane-disruptingAMPs in antibacterial chemotherapy in a viable fashion, theirsafety needs to be investigated. In a recent commentary on thedevelopment and spread of antibiotic resistance in bacteria, Bushet al. (397) said, “The use of probiotics is likely to become moreimportant in years to come as microbiological studies of the rolesof gastrointestinal bacterial populations (the human microbiome)lead to the identification of those bacterial genera and species thathave key roles in human health and disease. Such advances may welllead to the use of bacteria and their products as specific therapeutics.”It has recently been convincingly argued that bacteriocins are poten-tial alternatives to traditional antibiotics; however, the anti-infectioustherapeutic efficacy of bacteriocins in human beings remains to bedemonstrated in properly designed clinical studies (41–43). More-over, combinations of bacteriocins with old antibiotics, which haveunfortunately displayed toxic side effects in human beings, are cur-rently being investigated with a view to reducing their toxicity bylowering the concentrations generally used, accompanied by an en-hancement of their therapeutic efficacies.



on Novem


r.asm.org/

Dow

nloaded from

http://cmr.asm.org

http://cmr.asm.org/

So far, attempts to purify the antimicrobial nonbacteriocincompounds secreted by most probiotic Lactobacillus strains havefailed, and their structures remain unknown. It is essential to pu-rify these compounds in order to identify their exact mechanismsof action and to resolve the structures of these new, natural anti-microbial agents before there can be any hope of developing aninnovative drug design strategy involving them. However, thisdoes not seem to be an easy task, because the antimicrobial activityis exercised by compounds that become very unstable once theyare extracted from the culture medium. In addition, it has beendifficult in some cases to purify these molecules because bacteri-cidal activity disappears in the final stages of the purification pro-cess, as the active molecules probably act synergistically with lacticacid exerting a bacterial membrane permeabilization activity (un-published data). Genomic analyses (398, 399) have recently beenconducted in order to understand the core mechanisms that con-trol and regulate probiotic Lactobacillus bacterial growth, survival,signaling, and fermentative processes and, in some cases, poten-tially underlying probiotic activities (74, 400, 401). Genomic anal-yses have been published for only four of the six probiotic strainsexamined here: L. rhamnosus GG (402, 403), L. johnsonii NCC 533(404, 405), L. casei Shirota (403), and L. casei DN-114 001 (403).Genes coding for several strain-specific properties have been iden-tified. Genes of L. johnsonii NCC 533 code for the cell factors thataccount for the particular properties of adhesion/colonization(406, 407). A cluster of genes in L. casei Shirota code for the syn-thesis of the high-molecular-mass components that can be rele-vant for the bacterial whole-cell-induced, cell contact-dependentimmune modulation (408). Genes of L. rhamnosus GG code forthe pili involved in adhesion (191–194), for the formation of bio-film (409), and for enzymes involved in the biosynthesis of extra-cellular polysaccharides (410). Genes of L. casei DN-114 001 codefor adaptation of metabolic properties involved in intestinal col-onization (411). Currently, the comparative genomic studies havenot given pertinent information about the antibacterial moleculesproduced by the probiotic Lactobacillus strains. A strategy involv-ing mutant strains has been successfully developed to identifystructures, antibacterial activities, genetic systems, and biosynthe-ses, as well as the mechanisms of action, of bacteriocins of Gram-negative E. coli, i.e., microcins and colicins (49–51). The establish-ment of mutant strains of Lactobacillus in order to identify thegenes coding for anti-infectious molecules has rarely been at-tempted. Mutants of L. reuteri strains have been used for examin-ing the activity of the antibiotic 3-hydroxypropionaldehyde, alsoknown as reuterin (412), against enteric bacterial pathogens(146). Recently, using wild-type L. salivarius strain UCC118 and astable mutant in which the gene coding for the production of thebroad-spectrum class IIb bacteriocin Abp118 had been deleted,the probiotic activities of the strain have been characterized, in-cluding the protection of mice against L. monocytogenes infection(262) and changes in the composition of mouse and pig intestinalmicrobiota (413). A major shortcoming in the field of anti-infec-tious Lactobacillus molecules is the absence of comparativegenomic analyses coupled with a strategy involving mutants.

ACKNOWLEDGMENTS

Sincere thanks to Fabrice Atassi, Marie-Françoise Bernet-Camard, GillesChauvière, Marie-Hélène Coconnier-Polter, and Domitille Fayol-Mes-saoudi for their outstanding contributions to the understanding of thecellular and molecular anti-infectious mechanisms of human probiotic

strains (Inserm and University Paris-Sud Research Teams CJF 94.07 andUnits 510 and 756 at the University Paris-Sud, Faculty of Pharmacy,Châtenay-Malabry, France).

A.L.S. was a regulatory consultant to pharmaceutical companies, with-out holding any shares or equity. He does not have any ownership role orserve on governing boards for any company. A.L.S. has received industrialresearch contracts as Principal Investigator from Nestec (Vers-chez-les-Blancs, Switzerland), Laboratoire du Lactéol (Houdan, France), AxcanPharma (Mont Saint-Hilaire, Quebec, Canada), and Aptalis Pharma (Bir-mingham, AL). A.L.S. has received lecture fees from Laboratoire du Lac-téol, Aptalis Pharma, and Yakult (Seoul, South Korea). A.L.S. andV.L.-L.M. both hold U.S. and European patents for probiotic human Lac-tobacillus strains (Laboratoire du Lactéol and Aptalis Pharma), and A.L.S.holds U.S. and European patents for probiotic human Bifidobacteriumand Lactobacillus strains (Nestlé, Vevey, Switzerland), without holdingshares or equity. A.L.S. and V.L.-L.M. currently have no conflict of inter-est.

REFERENCES1. Lievin-Le Moal V, Servin AL. 2006. The front line of enteric host defense

against unwelcome intrusion of harmful microorganisms: mucins, anti-microbial peptides, and microbiota. Clin. Microbiol. Rev. 19:315–337.http://dx.doi.org/10.1128/CMR.19.2.315-337.2006.

2. Turner HL, Turner JR. 2010. Good fences make good neighbors: gas-trointestinal mucosal structure. Gut Microbes 1:22–29. http://dx.doi.org/10.4161/gmic.1.1.11427.

3. Yang I, Nell S, Suerbaum S. 2013. Survival in hostile territory: themicrobiota of the stomach. FEMS Microbiol. Rev. 37:736 –761. http://dx.doi.org/10.1111/1574-6976.12027.

4. Kouznetsova I, Kalinski T, Meyer F, Hoffmann W. 2011. Self-renewalof the human gastric epithelium: new insights from expression profilingusing laser microdissection. Mol. Biosyst. 7:1105–1112. http://dx.doi.org/10.1039/c0mb00233j.

5. Wessling-Resnick M. 2010. Iron homeostasis and the inflammatory re-sponse. Annu. Rev. Nutr. 30:105–122. http://dx.doi.org/10.1146/annurev.nutr.012809.104804.

6. Turner JR. 2009. Intestinal mucosal barrier function in health and disease.Nat. Rev. Immunol. 9:799–809. http://dx.doi.org/10.1038/nri2653.

7. McGuckin MA, Linden SK, Sutton P, Florin TH. 2011. Mucin dynam-ics and enteric pathogens. Nat. Rev. Microbiol. 9:265–278. http://dx.doi.org/10.1038/nrmicro2538.

8. Kim YS, Ho SB. 2010. Intestinal goblet cells and mucins in health anddisease: recent insights and progress. Curr. Gastroenterol. Rep. 12:319 –330. http://dx.doi.org/10.1007/s11894-010-0131-2.

9. Bevins CL, Salzman NH. 2011. Paneth cells, antimicrobial peptides andmaintenance of intestinal homeostasis. Nat. Rev. Microbiol. 9:356 –368.http://dx.doi.org/10.1038/nrmicro2546.

10. Melo MN, Ferre R, Castanho MA. 2009. Antimicrobial peptides: link-ing partition, activity and high membrane-bound concentrations. Nat.Rev. Microbiol. 7:245–250. http://dx.doi.org/10.1038/nrmicro2095.

11. Kumar H, Kawai T, Akira S. 2011. Pathogen recognition by the innateimmune system. Int. Rev. Immunol. 30:16 –34. http://dx.doi.org/10.3109/08830185.2010.529976.

12. O’Neill LA, Golenbock D, Bowie AG. 2013. The history of Toll-likereceptors—redefining innate immunity. Nat. Rev. Immunol. 13:453–460. http://dx.doi.org/10.1038/nri3446.

13. Chu H, Mazmanian SK. 2013. Innate immune recognition of the mi-crobiota promotes host-microbial symbiosis. Nat. Immunol. 14:668 –675. http://dx.doi.org/10.1038/ni.2635.

14. Deretic V. 2011. Autophagy in immunity and cell-autonomous defenseagainst intracellular microbes. Immunol. Rev. 240:92–104. http://dx.doi.org/10.1111/j.1600-065X.2010.00995.x.

15. Human Microbiome Project Consortium. 2012. Structure, functionand diversity of the healthy human microbiome. Nature 486:207–214.http://dx.doi.org/10.1038/nature11234.

16. Yatsunenko T, Rey FE, Manary MJ, Trehan I, Dominguez-Bello MG,Contreras M, Magris M, Hidalgo G, Baldassano RN, Anokhin AP,Heath AC, Warner B, Reeder J, Kuczynski J, Caporaso JG, LozuponeCA, Lauber C, Clemente JC, Knights D, Knight R, Gordon JI. 2012.Human gut microbiome viewed across age and geography. Nature 486:222–227.



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1128/CMR.19.2.315-337.2006

http://dx.doi.org/10.4161/gmic.1.1.11427


http://dx.doi.org/10.1111/1574-6976.12027

http://dx.doi.org/10.1111/1574-6976.12027

http://dx.doi.org/10.1039/c0mb00233j

http://dx.doi.org/10.1039/c0mb00233j

http://dx.doi.org/10.1146/annurev.nutr.012809.104804

http://dx.doi.org/10.1146/annurev.nutr.012809.104804

http://dx.doi.org/10.1038/nri2653

http://dx.doi.org/10.1038/nrmicro2538


http://dx.doi.org/10.1007/s11894-010-0131-2



http://dx.doi.org/10.3109/08830185.2010.529976

http://dx.doi.org/10.3109/08830185.2010.529976


http://dx.doi.org/10.1038/ni.2635

http://dx.doi.org/10.1111/j.1600-065X.2010.00995.x


http://dx.doi.org/10.1038/nature11234

http://cmr.asm.org

http://cmr.asm.org/

17. Rautava S, Luoto R, Salminen S, Isolauri E. 2012. Microbial contactduring pregnancy, intestinal colonization and human disease. Nat. Rev.Gastroenterol. Hepatol. 9:565–576. http://dx.doi.org/10.1038/nrgastro.2012.144.

18. Kamada N, Chen GY, Inohara N, Nunez G. 2013. Control of pathogensand pathobionts by the gut microbiota. Nat. Immunol. 14:685– 690.http://dx.doi.org/10.1038/ni.2608.

19. Arumugam M, Raes J, Pelletier E, Le Paslier D, Yamada T, Mende DR,Fernandes GR, Tap J, Bruls T, Batto JM, Bertalan M, Borruel N,Casellas F, Fernandez L, Gautier L, Hansen T, Hattori M, Hayashi T,Kleerebezem M, Kurokawa K, Leclerc M, Levenez F, Manichanh C,Nielsen HB, Nielsen T, Pons N, Poulain J, Qin J, Sicheritz-Ponten T,Tims S, Torrents D, Ugarte E, Zoetendal EG, Wang J, Guarner F,Pedersen O, de Vos WM, Brunak S, Dore J, Meta HITC, Antolin M,Artiguenave F, Blottiere HM, Almeida M, Brechot C, Cara C, Cher-vaux C, Cultrone A, Delorme C, Denariaz G, Dervyn R, et al. 2011.Enterotypes of the human gut microbiome. Nature 473:174 –180. http://dx.doi.org/10.1038/nature09944.

20. Sommer F, Backhed F. 2013. The gut microbiota—masters of hostdevelopment and physiology. Nat. Rev. Microbiol. 11:227–238. http://dx.doi.org/10.1038/nrmicro2974.

21. Matsumoto M, Kibe R, Ooga T, Aiba Y, Kurihara S, Sawaki E, KogaY, Benno Y. 2012. Impact of intestinal microbiota on intestinal luminalmetabolome. Sci. Rep. 2:233. http://dx.doi.org/10.1038/srep00233.

22. Antunes LC, Han J, Ferreira RB, Lolic P, Borchers CH, Finlay BB.2011. Effect of antibiotic treatment on the intestinal metabolome. Anti-microb. Agents Chemother. 55:1494 –1503. http://dx.doi.org/10.1128/AAC.01664-10.

23. Kamada N, Nunez G. 2013. Role of the gut microbiota in the develop-ment and function of lymphoid cells. J. Immunol. 190:1389 –1395. http://dx.doi.org/10.4049/jimmunol.1203100.

24. Brown EM, Sadarangani M, Finlay BB. 2013. The role of the immunesystem in governing host-microbe interactions in the intestine. Nat. Im-munol. 14:660 – 667. http://dx.doi.org/10.1038/ni.2611.

25. Buffie CG, Pamer EG. 2013. Microbiota-mediated colonization resis-tance against intestinal pathogens. Nat. Rev. Immunol. 13:790 – 801.http://dx.doi.org/10.1038/nri3535.

26. Kinnebrew MA, Pamer EG. 2012. Innate immune signaling in defenseagainst intestinal microbes. Immunol. Rev. 245:113–131. http://dx.doi.org/10.1111/j.1600-065X.2011.01081.x.

27. Aroniadis OC, Brandt LJ. 2013. Fecal microbiota transplantation: past,present and future. Curr. Opin. Gastroenterol. 29:79 – 84. http://dx.doi.org/10.1097/MOG.0b013e32835a4b3e.

28. Nell S, Suerbaum S, Josenhans C. 2010. The impact of the microbiotaon the pathogenesis of IBD: lessons from mouse infection models. Nat.Rev. Microbiol. 8:564 –577. http://dx.doi.org/10.1038/nrmicro2403.

29. Kamada N, Seo SU, Chen GY, Nunez G. 2013. Role of the gut micro-biota in immunity and inflammatory disease. Nat. Rev. Immunol. 13:321–335. http://dx.doi.org/10.1038/nri3430.

30. Damman CJ, Miller SI, Surawicz CM, Zisman TL. 2012. The micro-biome and inflammatory bowel disease: is there a therapeutic role forfecal microbiota transplantation? Am. J. Gastroenterol. 107:1452–1459.http://dx.doi.org/10.1038/ajg.2012.93.

31. Walker AW, Lawley TD. 2013. Therapeutic modulation of intestinaldysbiosis. Pharmacol. Res. 69:75– 86. http://dx.doi.org/10.1016/j.phrs.2012.09.008.

32. Stecher B, Maier L, Hardt WD. 2013. ‘Blooming’ in the gut: howdysbiosis might contribute to pathogen evolution. Nat. Rev. Microbiol.11:277–284. http://dx.doi.org/10.1038/nrmicro2989.

33. Chow J, Tang H, Mazmanian SK. 2011. Pathobionts of the gastrointes-tinal microbiota and inflammatory disease. Curr. Opin. Immunol. 23:473– 480. http://dx.doi.org/10.1016/j.coi.2011.07.010.

34. Yamaoka Y. 2010. Mechanisms of disease: Helicobacter pylori virulencefactors. Nat. Rev. Gastroenterol. Hepatol. 7:629 – 641. http://dx.doi.org/10.1038/nrgastro.2010.154.

35. Cover TL, Blaser MJ. 2009. Helicobacter pylori in health and disease.Gastroenterology 136:1863–1873. http://dx.doi.org/10.1053/j.gastro.2009.01.073.

36. Clements A, Young JC, Constantinou N, Frankel G. 2012. Infectionstrategies of enteric pathogenic Escherichia coli. Gut Microbes 3:71– 87.http://dx.doi.org/10.4161/gmic.19182.

37. Croxen MA, Law RJ, Scholz R, Keeney KM, Wlodarska M, Finlay BB.2013. Recent advances in understanding enteric pathogenic Escherichia

coli. Clin. Microbiol. Rev. 26:822– 880. http://dx.doi.org/10.1128/CMR.00022-13.

38. Coburn B, Grassl GA, Finlay BB. 2007. Salmonella, the host and disease:a brief review. Immunol. Cell Biol. 85:112–118. http://dx.doi.org/10.1038/sj.icb.7100007.

39. Nelson EJ, Harris JB, Morris JG, Jr, Calderwood SB, Camilli A. 2009.Cholera transmission: the host, pathogen and bacteriophage dynamic. Nat.Rev. Microbiol. 7:693–702. http://dx.doi.org/10.1038/nrmicro2204.

40. Marteyn B, Gazi A, Sansonetti P. 2012. Shigella: a model of virulenceregulation in vivo. Gut Microbes 3:104 –120. http://dx.doi.org/10.4161/gmic.19325.

41. Cotter PD, Ross RP, Hill C. 2013. Bacteriocins—a viable alternative toantibiotics? Nat. Rev. Microbiol. 11:95–105. http://dx.doi.org/10.1038/nrmicro2937.

42. Dobson A, Cotter PD, Ross RP, Hill C. 2012. Bacteriocin production:a probiotic trait? Appl. Environ. Microbiol. 78:1– 6. http://dx.doi.org/10.1128/AEM.05576-11.

43. O’Shea EF, Cotter PD, Stanton C, Ross RP, Hill C. 2012. Productionof bioactive substances by intestinal bacteria as a basis for explainingprobiotic mechanisms: bacteriocins and conjugated linoleic acid. Int. J.Food Microbiol. 152:189 –205. http://dx.doi.org/10.1016/j.ijfoodmicro.2011.05.025.

44. Cotter PD, Hill C, Ross RP. 2005. Bacteriocins: developing innateimmunity for food. Nat. Rev. Microbiol. 3:777–788. http://dx.doi.org/10.1038/nrmicro1273.

45. Klaenhammer TR. 1993. Genetics of bacteriocins produced by lacticacid bacteria. FEMS Microbiol. Rev. 12:39 – 85. http://dx.doi.org/10.1111/j.1574-6976.1993.tb00012.x.

46. Nes IF, Holo H. 2000. Class II antimicrobial peptides from lactic acidbacteria. Biopolymers 55:50 – 61. http://dx.doi.org/10.1002/1097-0282(2000)55:150::AID-BIP50�3.0.CO;2-3.

47. Oscariz JC, Pisabarro AG. 2001. Classification and mode of action ofmembrane-active bacteriocins produced by gram-positive bacteria. Int.Microbiol. 4:13–19. http://dx.doi.org/10.1007/s101230100003.

48. McAuliffe O, Ross RP, Hill C. 2001. Lantibiotics: structure, biosynthesisand mode of action. FEMS Microbiol. Rev. 25:285–308. http://dx.doi.org/10.1111/j.1574-6976.2001.tb00579.x.

49. Gordon DM, O’Brien CL. 2006. Bacteriocin diversity and the frequencyof multiple bacteriocin production in Escherichia coli. Microbiology 152:3239 –3244. http://dx.doi.org/10.1099/mic.0.28690-0.

50. Duquesne S, Destoumieux-Garzon D, Peduzzi J, Rebuffat S. 2007.Microcins, gene-encoded antibacterial peptides from enterobacteria.Nat. Prod. Rep. 24:708 –734. http://dx.doi.org/10.1039/b516237h.

51. Cascales E, Buchanan SK, Duche D, Kleanthous C, Lloubes R, PostleK, Riley M, Slatin S, Cavard D. 2007. Colicin biology. Microbiol. Mol.Biol. Rev. 71:158 –229. http://dx.doi.org/10.1128/MMBR.00036-06.

52. Sperandio V. 2009. Deciphering bacterial language. Nat. Chem. Biol.5:870 – 871. http://dx.doi.org/10.1038/nchembio.263.

53. Sperandio V, Torres AG, Jarvis B, Nataro JP, Kaper JB. 2003. Bacteria-host communication: the language of hormones. Proc. Natl. Acad. Sci.U. S. A. 100:8951– 8956. http://dx.doi.org/10.1073/pnas.1537100100.

54. Parker CT, Sperandio V. 2009. Cell-to-cell signalling during pathogen-esis. Cell. Microbiol. 11:363–369. http://dx.doi.org/10.1111/j.1462-5822.2008.01272.x.

55. Risoen PA, Brurberg MB, Eijsink VG, Nes IF. 2000. Functional analysisof promoters involved in quorum sensing-based regulation of bacterio-cin production in Lactobacillus. Mol. Microbiol. 37:619 – 628. http://dx.doi.org/10.1046/j.1365-2958.2000.02029.x.

56. Eijsink VG, Axelsson L, Diep DB, Havarstein LS, Holo H, Nes IF.2002. Production of class II bacteriocins by lactic acid bacteria; an exam-ple of biological warfare and communication. Antonie Van Leeuwen-hoek 81:639 – 654. http://dx.doi.org/10.1023/A:1020582211262.

57. Sturme MH, Francke C, Siezen RJ, de Vos WM, Kleerebezem M. 2007.Making sense of quorum sensing in lactobacilli: a special focus on Lacto-bacillus plantarum WCFS1. Microbiology 153:3939 –3947. http://dx.doi.org/10.1099/mic.0.2007/012831-0.

58. Moslehi-Jenabian S, Vogensen FK, Jespersen L. 2011. The quorumsensing luxS gene is induced in Lactobacillus acidophilus NCFM in re-sponse to Listeria monocytogenes. Int. J. Food Microbiol. 149:269 –273.http://dx.doi.org/10.1016/j.ijfoodmicro.2011.06.011.

59. Valdez JC, Peral MC, Rachid M, Santana M, Perdigon G. 2005.Interference of Lactobacillus plantarum with Pseudomonas aeruginosa invitro and in infected burns: the potential use of probiotics in wound



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1038/nrgastro.2012.144







http://dx.doi.org/10.1038/srep00233

http://dx.doi.org/10.1128/AAC.01664-10


http://dx.doi.org/10.4049/jimmunol.1203100

http://dx.doi.org/10.4049/jimmunol.1203100





http://dx.doi.org/10.1097/MOG.0b013e32835a4b3e

http://dx.doi.org/10.1097/MOG.0b013e32835a4b3e



http://dx.doi.org/10.1038/ajg.2012.93

http://dx.doi.org/10.1016/j.phrs.2012.09.008

http://dx.doi.org/10.1016/j.phrs.2012.09.008


http://dx.doi.org/10.1016/j.coi.2011.07.010



http://dx.doi.org/10.1053/j.gastro.2009.01.073


http://dx.doi.org/10.4161/gmic.19182



http://dx.doi.org/10.1038/sj.icb.7100007

http://dx.doi.org/10.1038/sj.icb.7100007






http://dx.doi.org/10.1128/AEM.05576-11


http://dx.doi.org/10.1016/j.ijfoodmicro.2011.05.025




http://dx.doi.org/10.1111/j.1574-6976.1993.tb00012.x


http://dx.doi.org/10.1002/1097-0282(2000)55:1%3C50::AID-BIP50%3E3.0.CO;2-3

http://dx.doi.org/10.1002/1097-0282(2000)55:1%3C50::AID-BIP50%3E3.0.CO;2-3

http://dx.doi.org/10.1007/s101230100003



http://dx.doi.org/10.1099/mic.0.28690-0

http://dx.doi.org/10.1039/b516237h

http://dx.doi.org/10.1128/MMBR.00036-06

http://dx.doi.org/10.1038/nchembio.263

http://dx.doi.org/10.1073/pnas.1537100100

http://dx.doi.org/10.1111/j.1462-5822.2008.01272.x

http://dx.doi.org/10.1111/j.1462-5822.2008.01272.x

http://dx.doi.org/10.1046/j.1365-2958.2000.02029.x

http://dx.doi.org/10.1046/j.1365-2958.2000.02029.x

http://dx.doi.org/10.1023/A:1020582211262

http://dx.doi.org/10.1099/mic.0.2007/012831-0

http://dx.doi.org/10.1099/mic.0.2007/012831-0


http://cmr.asm.org

http://cmr.asm.org/

treatment. Clin. Microbiol. Infect. 11:472– 479. http://dx.doi.org/10.1111/j.1469-0691.2005.01142.x.

60. Medellin-Pena MJ, Wang H, Johnson R, Anand S, Griffiths MW. 2007.Probiotics affect virulence-related gene expression in Escherichia coliO157:H7. Appl. Environ. Microbiol. 73:4259 – 4267. http://dx.doi.org/10.1128/AEM.00159-07.

61. Medellin-Pena MJ, Griffiths MW. 2009. Effect of molecules secreted byLactobacillus acidophilus strain La-5 on Escherichia coli O157:H7 coloni-zation. Appl. Environ. Microbiol. 75:1165–1172. http://dx.doi.org/10.1128/AEM.01651-08.

62. Chifiriuc MC, Ditu LM, Banu O, Bleotu C, Dracea O, Bucur M, LarionC, Israil AM, Lazar V. 2009. Subinhibitory concentrations of phenyllactic acid interfere with the expression of virulence factors in Staphylo-coccus aureus and Pseudomonas aeruginosa clinical strains. Roum. Arch.Microbiol. Immunol. 68:27–33.

63. Lazar V, Miyazaki Y, Hanawa T, Chifiriuc MC, Ditu LM, MarutescuL, Bleotu C, Kamiya S. 2009. The influence of some probiotic superna-tants on the growth and virulence features expression of several selectedenteroaggregative E. coli clinical strains. Roumanian Arch. Microbiol.Immunol. 68:207–214.

64. Li J, Wang W, Xu SX, Magarvey NA, McCormick JK. 2011. Lactobacillusreuteri-produced cyclic dipeptides quench agr-mediated expression of toxicshock syndrome toxin-1 in staphylococci. Proc. Natl. Acad. Sci. U. S. A.108:3360–3365. http://dx.doi.org/10.1073/pnas.1017431108.

65. Ramos AN, Cabral ME, Noseda D, Bosch A, Yantorno OM, Valdez JC.2012. Antipathogenic properties of Lactobacillus plantarum on Pseu-domonas aeruginosa: the potential use of its supernatants in the treatmentof infected chronic wounds. Wound Rep. Regen. 20:552–562. http://dx.doi.org/10.1111/j.1524-475X.2012.00798.x.

66. Metchnikoff E. 1907. The prolongation of life: optimistic studies, p 161–183. W. Heinemann, London, United Kingdom.

67. Tissier H. 1906. Traitement des infections intestinales par la méthode dela flore bactérienne de l’intestin. C.R. Soc. Biol. 60:359 –361.

68. Fuller R. 1989. Probiotics in man and animals. J. Appl. Bacteriol. 66:365–378. http://dx.doi.org/10.1111/j.1365-2672.1989.tb05105.x.

69. FAO/WHO. 2001. Joint FAO/WHO expert consultation on evaluationof health and nutritional properties of probiotics in food including pow-der milk with live lactic acid bacteria, Cordoba, Argentina, 1 to 4 October2001. http://www.who.int/foodsafety/publications/fs_management/en/probiotics.pdf.

70. FAO/WHO. 2002. Guidelines for the evaluation of probiotics in food.Food and Agriculture Organization of the United Nations and WorldHealth Organization Working Group Report. ftp://ftp.fao.org/es/esn/food/wgreport2.pdf.

71. Servin AL. 2004. Antagonistic activities of lactobacilli and bifidobacteriaagainst microbial pathogens. FEMS Microbiol. Rev. 28:405– 440. http://dx.doi.org/10.1016/j.femsre.2004.01.003.

72. Bron PA, van Baarlen P, Kleerebezem M. 2012. Emerging molecularinsights into the interaction between probiotics and the host intestinalmucosa. Nat. Rev. Microbiol. 10:66 –78. http://dx.doi.org/10.1038/nrmicro2690.

73. Lebeer S, Vanderleyden J, De Keersmaecker SC. 2010. Host interac-tions of probiotic bacterial surface molecules: comparison with com-mensals and pathogens. Nat. Rev. Microbiol. 8:171–184. http://dx.doi.org/10.1038/nrmicro2297.

74. Kleerebezem M, Hols P, Bernard E, Rolain T, Zhou M, Siezen RJ,Bron PA. 2010. The extracellular biology of the lactobacilli. FEMS Mi-crobiol. Rev. 34:199 –230. http://dx.doi.org/10.1111/j.1574-6976.2009.00208.x.

75. van Baarlen P, Wells JM, Kleerebezem M. 2013. Regulation of intestinalhomeostasis and immunity with probiotic lactobacilli. Trends Immunol.34:208 –215. http://dx.doi.org/10.1016/j.it.2013.01.005.

76. Sanchez B, Bressollier P, Urdaci MC. 2008. Exported proteins in pro-biotic bacteria: adhesion to intestinal surfaces, host immunomodulationand molecular cross-talking with the host. FEMS Immunol. Med. Micro-biol. 54:1–17. http://dx.doi.org/10.1111/j.1574-695X.2008.00454.x.

77. Sanchez B, Urdaci MC, Margolles A. 2010. Extracellular proteins se-creted by probiotic bacteria as mediators of effects that promote mucosa-bacteria interactions. Microbiology 156:3232–3242. http://dx.doi.org/10.1099/mic.0.044057-0.

78. Klaenhammer TR, Kleerebezem M, Kopp MV, Rescigno M. 2012. Theimpact of probiotics and prebiotics on the immune system. Nat. Rev.Immunol. 12:728 –734. http://dx.doi.org/10.1038/nri3312.

79. Doron S, Snydman DR, Gorbach SL. 2005. Lactobacillus GG: bacteri-ology and clinical applications. Gastroenterol. Clin. North Am. 34:483–498, ix. http://dx.doi.org/10.1016/j.gtc.2005.05.011.

80. Grzeskowiak L, Isolauri E, Salminen S, Gueimonde M. 2011. Manu-facturing process influences properties of probiotic bacteria. Br. J. Nutr.105:887– 894. http://dx.doi.org/10.1017/S0007114510004496.

81. Goldin BR, Gorbach SL. April 1985. Lactobacillus strains and methodsof selection. US patent 4,839,281.

82. Goldin BR, Gorbach SL. July 1991. L. acidophilus strains. US patent5,032,399.

83. Degeest B. 2001. Applications of probiotics. Meded. Rijksuniv. GentFak. Landbouwkd. Toegep. Biol. Wet. 66:557–561.

84. Yoshinori U, Hiromi S, Akiyoshi I, Taeko H, Yuushiro A, Yukifumi N.September 2004. Japanese patent 2004/244378.

85. Boisseau R, Chauviere G, Coconnier MH, Houlier P, Lievin V, ServinA. May 2000. Antibacterial composition. European patent EP1000625.

86. Brassart D, Donnet A, Link H, Mignot O, Neeser JR, Rochat F,Schiffrin E, Servin, A. February 1997. Lactobacillus johnsonii CNCMI-1225. US patent 5,603,930.

87. Postaire E, Bouley C, Guerin-Danan C, Andrieux C. June 2002.Method and composition for treatment of infant diarrhea. US patent6,399,055.

88. Rosander A, Connolly E, Roos S. 2008. Removal of antibiotic resistancegene-carrying plasmids from Lactobacillus reuteri ATCC 55730 and char-acterization of the resulting daughter strain, L. reuteri DSM 17938. Appl.Environ. Microbiol. 74:6032– 6040. http://dx.doi.org/10.1128/AEM.00991-08.

89. Connolly E, Mollstam B. May 2008. Use of selected lactic acid bacteriafor reducing infantile colic. US patent 7,374,924.

90. Kang HJ, Connolly E. June 2012. Method of improving immune func-tion in mammals using Lactobacillus strains with certain lipids. Europeanpatent EP2468284.

91. Servin A, Chauviere G, Polter M-H, Lievin-Le Moal V, Gastebois B.July 2006. Lactobacillus fermentum strain and uses thereof. US patent0,134,220.

92. Chauviere G, Gastebois B, Lievin-Le Moal V, Polter MH, Servin A.December 2005. Lactobacillus fermentum strain and uses thereof. Euro-pean patent EP1608737.

93. Sanders ME, Klaenhammer TR. 2001. The scientific basis of Lactobacil-lus acidophilus NCFM functionality as a probiotic. J. Dairy Sci. 84:319 –331. http://dx.doi.org/10.3168/jds.S0022-0302(01)74481-5.

94. Mikelsaar M, Zilmer M. 2009. Lactobacillus fermentum ME-3—anantimicrobial and antioxidative probiotic. Microb. Ecol. Health Dis. 21:1–27. http://dx.doi.org/10.1080/08910600902815561.

95. Hagen KE, Tramp CA, Altermann E, Welker DL, Tompkins TA. 2010.Sequence analysis of plasmid pIR52-1 from Lactobacillus helveticusR0052 and investigation of its origin of replication. Plasmid 63:108 –117.http://dx.doi.org/10.1016/j.plasmid.2009.12.004.

96. Tompkins TA, Barreau G, Broadbent JR. 2012. Complete genomesequence of Lactobacillus helveticus R0052, a commercial probioticstrain. J. Bacteriol. 194:6349. http://dx.doi.org/10.1128/JB.01638-12.

97. Foster LM, Tompkins TA, Dahl WJ. 2011. A comprehensive post-market review of studies on a probiotic product containing Lactobacillushelveticus R0052 and Lactobacillus rhamnosus R0011. Benef. Microbes2:319 –334. http://dx.doi.org/10.3920/BM2011.0032.

98. Hoffman FA. 2008. Development of probiotics as biologic drugs. Clin.Infect. Dis. 46(Suppl 2):S125–S127. http://dx.doi.org/10.1086/523326.

99. Sutton A. 2008. Product development of probiotics as biological drugs.Clin. Infect. Dis. 46(Suppl 2):S128 –S132. http://dx.doi.org/10.1086/523325.

100. Floch MH, Walker WA, Guandalini S, Hibberd P, Gorbach S, Sura-wicz C, Sanders ME, Garcia-Tsao G, Quigley EM, Isolauri E, FedorakRN, Dieleman LA. 2008. Recommendations for probiotic use—2008. J.Clin. Gastroenterol. 42(Suppl 2):S104 –S108. http://dx.doi.org/10.1097/MCG.0b013e31816b903f.

101. Goldin BR, Gorbach SL. 2008. Clinical indications for probiotics: anoverview. Clin. Infect. Dis. 46(Suppl 2):S96 –S100. http://dx.doi.org/10.1086/523333.

102. Hempel S, Newberry SJ, Maher AR, Wang Z, Miles JN, Shanman R,Johnsen B, Shekelle PG. 2012. Probiotics for the prevention and treat-ment of antibiotic-associated diarrhea: a systematic review and meta-analysis. JAMA 307:1959 –1969. http://dx.doi.org/10.1001/jama.2012.3507.



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1111/j.1469-0691.2005.01142.x

http://dx.doi.org/10.1111/j.1469-0691.2005.01142.x









http://www.who.int/foodsafety/publications/fs_management/en/probiotics.pdf

http://www.who.int/foodsafety/publications/fs_management/en/probiotics.pdf

ftp://ftp.fao.org/es/esn/food/wgreport2.pdf

ftp://ftp.fao.org/es/esn/food/wgreport2.pdf

http://dx.doi.org/10.1016/j.femsre.2004.01.003

http://dx.doi.org/10.1016/j.femsre.2004.01.003





http://dx.doi.org/10.1111/j.1574-6976.2009.00208.x

http://dx.doi.org/10.1111/j.1574-6976.2009.00208.x

http://dx.doi.org/10.1016/j.it.2013.01.005





http://dx.doi.org/10.1016/j.gtc.2005.05.011

http://dx.doi.org/10.1017/S0007114510004496



http://dx.doi.org/10.3168/jds.S0022-0302(01)74481-5

http://dx.doi.org/10.1080/08910600902815561

http://dx.doi.org/10.1016/j.plasmid.2009.12.004

http://dx.doi.org/10.1128/JB.01638-12

http://dx.doi.org/10.3920/BM2011.0032

http://dx.doi.org/10.1086/523326

http://dx.doi.org/10.1086/523325

http://dx.doi.org/10.1086/523325

http://dx.doi.org/10.1097/MCG.0b013e31816b903f

http://dx.doi.org/10.1097/MCG.0b013e31816b903f

http://dx.doi.org/10.1086/523333

http://dx.doi.org/10.1086/523333

http://dx.doi.org/10.1001/jama.2012.3507

http://dx.doi.org/10.1001/jama.2012.3507

http://cmr.asm.org

http://cmr.asm.org/

103. Jonkers D, Penders J, Masclee A, Pierik M. 2012. Probiotics in themanagement of inflammatory bowel disease: a systematic review of in-tervention studies in adult patients. Drugs 72:803– 823. http://dx.doi.org/10.2165/11632710-000000000-00000.

104. Hempel S, Newberry S, Ruelaz A, Wang Z, Miles J, Suttorp M,Johnsen B, Fu N, Smith A, Roth E, Polak J, Motala A, Shekelle PG.2011. Safety of probiotics used to reduce risk and prevent or treat disease.Report/technology assessment no. 200. AHRQ publication no. 11-E007.Agency for Healthcare Research and Quality, Rockville, MD. http://www.ahrq.gov/clinic/tp/probiotictp.htm.

105. Sanders ME, Akkermans LM, Haller D, Hammerman C, Heimbach J,Hormannsperger G, Huys G, Levy DD, Lutgendorff F, Mack D,Phothirath P, Solano-Aguilar G, Vaughan E. 2010. Safety assessment ofprobiotics for human use. Gut Microbes 1:164 –185. http://dx.doi.org/10.4161/gmic.1.3.12127.

106. Whelan K, Myers CE. 2010. Safety of probiotics in patients receivingnutritional support: a systematic review of case reports, randomized con-trolled trials, and nonrandomized trials. Am. J. Clin. Nutr. 91:687–703.http://dx.doi.org/10.3945/ajcn.2009.28759.

107. Fayol-Messaoudi D, Berger CN, Coconnier-Polter MH, Lievin-LeMoal V, Servin AL. 2005. pH-, lactic acid-, and non-lactic acid-dependent activities of probiotic lactobacilli against Salmonella entericaserovar Typhimurium. Appl. Environ. Microbiol. 71:6008 – 6013. http://dx.doi.org/10.1128/AEM.71.10.6008-6013.2005.

108. Lahtinen SJ, Gueimonde M, Ouwehand AC, Reinikainen JP, SalminenSJ. 2005. Probiotic bacteria may become dormant during storage. Appl.Environ. Microbiol. 71:1662–1663. http://dx.doi.org/10.1128/AEM.71.3.1662-1663.2005.

109. Lahtinen SJ, Ouwehand AC, Reinikainen JP, Korpela JM, Sandholm J,Salminen SJ. 2006. Intrinsic properties of so-called dormant probioticbacteria, determined by flow cytometric viability assays. Appl. Environ.Microbiol. 72:5132–5134. http://dx.doi.org/10.1128/AEM.02897-05.

110. Muller JA, Stanton C, Sybesma W, Fitzgerald GF, Ross RP. 2010.Reconstitution conditions for dried probiotic powders represent a criti-cal step in determining cell viability. J. Appl. Microbiol. 108:1369 –1379.http://dx.doi.org/10.1111/j.1365-2672.2009.04533.x.

111. Sanders ME. 2008. Probiotics: definition, sources, selection, anduses. Clin. Infect. Dis. 46(Suppl 2):S58 –S61. http://dx.doi.org/10.1086/523341.

112. Han KS, Kim Y, Kim SH, Oh S. 2007. Characterization and purificationof acidocin 1B, a bacteriocin produced by Lactobacillus acidophilus GP1B.J. Microbiol. Biotechnol. 17:774 –783.

113. Kim TS, Hur JW, Yu MA, Cheigh CI, Kim KN, Hwang JK, Pyun YR.2003. Antagonism of Helicobacter pylori by bacteriocins of lactic acidbacteria. J. Food Prot. 66:3–12.

114. Messaoudi S, Kergourlay G, Dalgalarrondo M, Choiset Y, Ferchichi M,Prevost H, Pilet MF, Chobert JM, Manai M, Dousset X. 2012. Purifi-cation and characterization of a new bacteriocin active against Campylo-bacter produced by Lactobacillus salivarius SMXD51. Food Microbiol.32:129 –134. http://dx.doi.org/10.1016/j.fm.2012.05.002.

115. Pascual LM, Daniele MB, Giordano W, Pajaro MC, Barberis IL. 2008.Purification and partial characterization of novel bacteriocin L23 pro-duced by Lactobacillus fermentum L23. Curr. Microbiol. 56:397– 402.http://dx.doi.org/10.1007/s00284-007-9094-4.

116. Zamfir M, Callewaert R, Cornea PC, Savu L, Vatafu I, De Vuyst L.1999. Purification and characterization of a bacteriocin produced by Lac-tobacillus acidophilus IBB 801. J. Appl. Microbiol. 87:923–931. http://dx.doi.org/10.1046/j.1365-2672.1999.00950.x.

117. NCCLS. 1999. Methods for determining bactericidal activity of antimi-crobial agents. Approved guideline. NCCLS document M26-A, p 1-29.National Committee for Clinical Laboratory Standards, Villanova, PA.

118. Hutt P, Shchepetova J, Loivukene K, Kullisaar T, Mikelsaar M. 2006.Antagonistic activity of probiotic lactobacilli and bifidobacteria againstentero- and uropathogens. J. Appl. Microbiol. 100:1324 –1332. http://dx.doi.org/10.1111/j.1365-2672.2006.02857.x.

119. Zhang Y, Zhang L, Du M, Yi H, Guo C, Tuo Y, Han X, Li J, Yang L.2011. Antimicrobial activity against Shigella sonnei and probiotic prop-erties of wild lactobacilli from fermented food. Microbiol. Res. 167:27–31. http://dx.doi.org/10.1016/j.micres.2011.02.006.

120. Bernet-Camard MF, Lievin V, Brassart D, Neeser JR, Servin AL,Hudault S. 1997. The human Lactobacillus acidophilus strain LA1 se-cretes a nonbacteriocin antibacterial substance(s) active in vitro and invivo. Appl. Environ. Microbiol. 63:2747–2753.

121. Spinler JK, Taweechotipatr M, Rognerud CL, Ou CN, Tumwasorn S,Versalovic J. 2008. Human-derived probiotic Lactobacillus reuteri dem-onstrate antimicrobial activities targeting diverse enteric bacterial patho-gens. Anaerobe 14:166 –171. http://dx.doi.org/10.1016/j.anaerobe.2008.02.001.

122. Coconnier MH, Lievin V, Bernet-Camard MF, Hudault S, Servin AL.1997. Antibacterial effect of the adhering human Lactobacillus acidophi-lus strain LB. Antimicrob. Agents Chemother. 41:1046 –1052.

123. Silva M, Jacobus NV, Deneke C, Gorbach SL. 1987. Antimicrobialsubstance from a human Lactobacillus strain. Antimicrob. Agents Che-mother. 31:1231–1233. http://dx.doi.org/10.1128/AAC.31.8.1231.

124. Ogawa M, Shimizu K, Nomoto K, Tanaka R, Hamabata T, YamasakiS, Takeda T, Takeda Y. 2001. Inhibition of in vitro growth of Shigatoxin-producing Escherichia coli O157:H7 by probiotic Lactobacillusstrains due to production of lactic acid. Int. J. Food Microbiol. 68:135–140. http://dx.doi.org/10.1016/S0168-1605(01)00465-2.

125. Lievin-Le Moal V, Amsellem R, Servin AL, Coconnier MH. 2002.Lactobacillus acidophilus (strain LB) from the resident adult human gas-trointestinal microflora exerts activity against brush border damage pro-moted by a diarrhoeagenic Escherichia coli in human enterocyte-likecells. Gut 50:803– 811. http://dx.doi.org/10.1136/gut.50.6.803.

126. Burkholder KM, Bhunia AK. 2009. Salmonella enterica serovar Typhi-murium adhesion and cytotoxicity during epithelial cell stress is reducedby Lactobacillus rhamnosus GG. Gut Pathog. 1:14. http://dx.doi.org/10.1186/1757-4749-1-14.

127. Marianelli C, Cifani N, Pasquali P. 2010. Evaluation of antimicrobialactivity of probiotic bacteria against Salmonella enterica subsp. entericaserovar typhimurium 1344 in a common medium under different envi-ronmental conditions. Res. Microbiol. 161:673– 680. http://dx.doi.org/10.1016/j.resmic.2010.06.007.

128. Fayol-Messaoudi D, Coconnier-Polter MH, Moal VL, Atassi F, BergerCN, Servin AL. 2007. The Lactobacillus plantarum strain ACA-DC287isolated from a Greek cheese demonstrates antagonistic activity in vitroand in vivo against Salmonella enterica serovar Typhimurium. J. Appl.Microbiol. 103:657– 665. http://dx.doi.org/10.1111/j.1365-2672.2007.03293.x.

129. Makras L, Triantafyllou V, Fayol-Messaoudi D, Adriany T, Zoumpo-poulou G, Tsakalidou E, Servin A, De Vuyst L. 2006. Kinetic analysisof the antibacterial activity of probiotic lactobacilli towards Salmonellaenterica serovar Typhimurium reveals a role for lactic acid and otherinhibitory compounds. Res. Microbiol. 157:241–247. http://dx.doi.org/10.1016/j.resmic.2005.09.002.

130. Hudault S, Lievin V, Bernet-Camard MF, Servin AL. 1997. Antagonis-tic activity exerted in vitro and in vivo by Lactobacillus casei (strain GG)against Salmonella typhimurium C5 infection. Appl. Environ. Microbiol.63:513–518.

131. Lehto EM, Salminen SJ. 1997. Inhibition of Salmonella typhimuriumadhesion to Caco-2 cell cultures by Lactobacillus strain GG spent culturesupernate: only a pH effect? FEMS Immunol. Med. Microbiol. 18:125–132. http://dx.doi.org/10.1111/j.1574-695X.1997.tb01037.x.

132. Atassi F, Servin AL. 2010. Individual and co-operative roles of lactic acidand hydrogen peroxide in the killing activity of enteric strain Lactobacil-lus johnsonii NCC933 and vaginal strain Lactobacillus gasseri KS120.1against enteric, uropathogenic and vaginosis-associated pathogens.FEMS Microbiol. Lett. 304:29 –38. http://dx.doi.org/10.1111/j.1574-6968.2009.01887.x.

133. Pridmore RD, Pittet AC, Praplan F, Cavadini C. 2008. Hydrogenperoxide production by Lactobacillus johnsonii NCC 533 and its role inanti-Salmonella activity. FEMS Microbiol. Lett. 283:210 –215. http://dx.doi.org/10.1111/j.1574-6968.2008.01176.x.

134. Vizoso Pinto MG, Franz CM, Schillinger U, Holzapfel WH. 2006.Lactobacillus spp. with in vitro probiotic properties from human faecesand traditional fermented products. Int. J. Food Microbiol. 109:205–214.http://dx.doi.org/10.1016/j.ijfoodmicro.2006.01.029.

135. Asahara T, Shimizu K, Takada T, Kado S, Yuki N, Morotomi M,Tanaka R, Nomoto K. 2011. Protective effect of Lactobacillus casei strainShirota against lethal infection with multi-drug resistant Salmonella en-terica serovar Typhimurium DT104 in mice. J. Appl. Microbiol. 110:163–173. http://dx.doi.org/10.1111/j.1365-2672.2010.04884.x.

136. Coconnier MH, Lievin V, Lorrot M, Servin AL. 2000. Antagonisticactivity of Lactobacillus acidophilus LB against intracellular Salmonellaenterica serovar Typhimurium infecting human enterocyte-like Caco-2/



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.2165/11632710-000000000-00000

http://dx.doi.org/10.2165/11632710-000000000-00000

http://www.ahrq.gov/clinic/tp/probiotictp.htm

http://www.ahrq.gov/clinic/tp/probiotictp.htm



http://dx.doi.org/10.3945/ajcn.2009.28759

http://dx.doi.org/10.1128/AEM.71.10.6008-6013.2005





http://dx.doi.org/10.1111/j.1365-2672.2009.04533.x

http://dx.doi.org/10.1086/523341

http://dx.doi.org/10.1086/523341

http://dx.doi.org/10.1016/j.fm.2012.05.002

http://dx.doi.org/10.1007/s00284-007-9094-4

http://dx.doi.org/10.1046/j.1365-2672.1999.00950.x

http://dx.doi.org/10.1046/j.1365-2672.1999.00950.x

http://dx.doi.org/10.1111/j.1365-2672.2006.02857.x

http://dx.doi.org/10.1111/j.1365-2672.2006.02857.x

http://dx.doi.org/10.1016/j.micres.2011.02.006

http://dx.doi.org/10.1016/j.anaerobe.2008.02.001


http://dx.doi.org/10.1128/AAC.31.8.1231

http://dx.doi.org/10.1016/S0168-1605(01)00465-2

http://dx.doi.org/10.1136/gut.50.6.803

http://dx.doi.org/10.1186/1757-4749-1-14

http://dx.doi.org/10.1186/1757-4749-1-14

http://dx.doi.org/10.1016/j.resmic.2010.06.007


http://dx.doi.org/10.1111/j.1365-2672.2007.03293.x

http://dx.doi.org/10.1111/j.1365-2672.2007.03293.x



http://dx.doi.org/10.1111/j.1574-695X.1997.tb01037.x

http://dx.doi.org/10.1111/j.1574-6968.2009.01887.x

http://dx.doi.org/10.1111/j.1574-6968.2009.01887.x

http://dx.doi.org/10.1111/j.1574-6968.2008.01176.x

http://dx.doi.org/10.1111/j.1574-6968.2008.01176.x


http://dx.doi.org/10.1111/j.1365-2672.2010.04884.x

http://cmr.asm.org

http://cmr.asm.org/

TC-7 cells. Appl. Environ. Microbiol. 66:1152–1157. http://dx.doi.org/10.1128/AEM.66.3.1152-1157.2000.

137. Coconnier-Polter MH, Lievin-Le Moal V, Servin AL. 2005. A Lactoba-cillus acidophilus strain of human gastrointestinal microbiota origin elic-its killing of enterovirulent Salmonella enterica serovar Typhimurium bytriggering lethal bacterial membrane damage. Appl. Environ. Microbiol.71:6115– 6120. http://dx.doi.org/10.1128/AEM.71.10.6115-6120.2005.

138. Bernet-Camard MF, Coconnier MH, Hudault S, Servin AL. 1996.Pathogenicity of the diffusely adhering strain Escherichia coli C1845:F1845 adhesin-decay accelerating factor interaction, brush border mi-crovillus injury, and actin disassembly in cultured human intestinal ep-ithelial cells. Infect. Immun. 64:1918 –1928.

139. Coconnier MH, Lievin V, Hemery E, Servin AL. 1998. Antagonisticactivity against Helicobacter infection in vitro and in vivo by the humanLactobacillus acidophilus strain LB. Appl. Environ. Microbiol. 64:4573–4580.

140. De Keersmaecker SC, Verhoeven TL, Desair J, Marchal K, Vanderley-den J, Nagy I. 2006. Strong antimicrobial activity of Lactobacillus rham-nosus GG against Salmonella typhimurium is due to accumulation oflactic acid. FEMS Microbiol. Lett. 259:89 –96. http://dx.doi.org/10.1111/j.1574-6968.2006.00250.x.

141. Lu R, Fasano S, Madayiputhiya N, Morin NP, Nataro J, Fasano A.2009. Isolation, identification, and characterization of small bioactivepeptides from Lactobacillus GG conditional media that exert both anti-Gram-negative and Gram-positive bactericidal activity. J. Pediatr. Gas-troenterol. Nutr. 49:23–30. http://dx.doi.org/10.1097/MPG.0b013e3181924d1e.

142. Gong HS, Meng XC, Wang H. 2010. Mode of action of plantaricin MG,a bacteriocin active against Salmonella typhimurium. J. Basic Microbiol.50(Suppl 1):S37–S45. http://dx.doi.org/10.1002/jobm.201000130.

143. Sharafi H, Maleki H, Ahmadian G, Shahbani Zahiri H, Sajedinejad N,Houshmand B, Vali H, Akbari Noghabi K. 2013. Antibacterial activityand probiotic potential of Lactobacillus plantarum HKN01: a new insightinto the morphological changes of antibacterial compound-treated Esch-erichia coli by electron microscopy. J. Microbiol. Biotechnol. 23:225–236.http://dx.doi.org/10.4014/jmb.1208.08005.

144. Svetoch EA, Eruslanov BV, Levchuk VP, Perelygin VV, Mitsevich EV,Mitsevich IP, Stepanshin J, Dyatlov I, Seal BS, Stern NJ. 2011. Isolationof Lactobacillus salivarius 1077 (NRRL B-50053) and characterization ofits bacteriocin, including the antimicrobial activity spectrum. Appl.Environ. Microbiol. 77:2749 –2754. http://dx.doi.org/10.1128/AEM.02481-10.

145. Carey CM, Kostrzynska M, Ojha S, Thompson S. 2008. The effect ofprobiotics and organic acids on Shiga-toxin 2 gene expression in entero-hemorrhagic Escherichia coli O157:H7. J. Microbiol. Methods 73:125–132. http://dx.doi.org/10.1016/j.mimet.2008.01.014.

146. Jelcic I, Hufner E, Schmidt H, Hertel C. 2008. Repression of the locusof the enterocyte effacement-encoded regulator of gene transcription ofEscherichia coli O157:H7 by Lactobacillus reuteri culture supernatants isLuxS and strain dependent. Appl. Environ. Microbiol. 74:3310 –3314.http://dx.doi.org/10.1128/AEM.00072-08.

147. Lievin-Le Moal V, Amsellem R, Servin AL. 2011. Impairment of swim-ming motility by antidiarrheic Lactobacillus acidophilus strain LB retardsinternalization of Salmonella enterica serovar Typhimurium within hu-man enterocyte-like cells. Antimicrob. Agents Chemother. 55:4810 –4820. http://dx.doi.org/10.1128/AAC.00418-11.

148. Lievin-Le Moal V, Fayol-Messaoudi D, Servin AL. 2013. Compound(s)secreted by Lactobacillus casei strain Shirota YIT9029 irreversibly andreversibly impair the swimming motility of Helicobacter pylori and Sal-monella enterica serovar Typhimurium, respectively. Microbiology 159:1956 –1971. http://dx.doi.org/10.1099/mic.0.067678-0.

149. Sgouras DN, Panayotopoulou EG, Martinez-Gonzalez B, Petraki K,Michopoulos S, Mentis A. 2005. Lactobacillus johnsonii La1 attenuatesHelicobacter pylori-associated gastritis and reduces levels of proinflam-matory chemokines in C57BL/6 mice. Clin. Diagn. Lab. Immunol. 12:1378 –1386. http://dx.doi.org/10.1128/CDLI.12.12.1378-1386.2005.

150. Michetti P, Dorta G, Wiesel PH, Brassart D, Verdu E, Herranz M,Felley C, Porta N, Rouvet M, Blum AL, Corthesy-Theulaz I. 1999.Effect of whey-based culture supernatant of Lactobacillus acidophilus(johnsonii) La1 on Helicobacter pylori infection in humans. Digestion60:203–209. http://dx.doi.org/10.1159/000007660.

151. Avonts L, De Vuyst L. 2001. Antimicrobial potential of probiotic lactic

acid bacteria. Meded. Rijksuniv. Gent Fak. Landbouwkd. Toegep. Biol.Wet. 66:543–550.

152. Morency H, Mota-Meira M, LaPointe G, Lacroix C, Lavoie MC. 2001.Comparison of the activity spectra against pathogens of bacterial strainsproducing a mutacin or a lantibiotic. Can. J. Microbiol. 47:322–331. http://dx.doi.org/10.1139/w01-013.

153. Ryan KA, O’Hara AM, van Pijkeren JP, Douillard FP, O’Toole PW.2009. Lactobacillus salivarius modulates cytokine induction and viru-lence factor gene expression in Helicobacter pylori. J. Med. Microbiol.58:996 –1005. http://dx.doi.org/10.1099/jmm.0.009407-0.

154. Simova ED, Beshkova DB, Dimitrov ZP. 2009. Characterization andantimicrobial spectrum of bacteriocins produced by lactic acid bacteriaisolated from traditional Bulgarian dairy products. J. Appl. Microbiol.106:692–701. http://dx.doi.org/10.1111/j.1365-2672.2008.04052.x.

155. Berry V, Jennings K, Woodnutt G. 1995. Bactericidal and morpholog-ical effects of amoxicillin on Helicobacter pylori. Antimicrob. Agents Che-mother. 39:1859 –1861. http://dx.doi.org/10.1128/AAC.39.8.1859.

156. DeLoney CR, Schiller NL. 1999. Competition of various beta-lactamantibiotics for the major penicillin-binding proteins of Helicobacter py-lori: antibacterial activity and effects on bacterial morphology. Antimi-crob. Agents Chemother. 43:2702–2709.

157. Bland MV, Ismail S, Heinemann JA, Keenan JI. 2004. The action ofbismuth against Helicobacter pylori mimics but is not caused by intracel-lular iron deprivation. Antimicrob. Agents Chemother. 48:1983–1988.http://dx.doi.org/10.1128/AAC.48.6.1983-1988.2004.

158. Sisto F, Brenciaglia MI, Scaltrito MM, Dubini F. 2000. Helicobacterpylori: ureA, cagA and vacA expression during conversion to the coccoidform. Int. J. Antimicrob. Agents 15:277–282. http://dx.doi.org/10.1016/S0924-8579(00)00188-6.

159. Lertsethtakarn P, Ottemann KM, Hendrixson DR. 2011. Motility andchemotaxis in Campylobacter and Helicobacter. Annu. Rev. Microbiol.65:389 – 410. http://dx.doi.org/10.1146/annurev-micro-090110-102908.

160. Isobe H, Nishiyama A, Takano T, Higuchi W, Nakagawa S, Taneike I,Fukushima Y, Yamamoto T. 2012. Reduction of overall Helicobacterpylori colonization levels in the stomach of Mongolian gerbil by Lacto-bacillus johnsonii La1 (LC1) and its in vitro activities against H. pylorimotility and adherence. Biosci. Biotechnol. Biochem. 76:850 – 852. http://dx.doi.org/10.1271/bbb.110921.

161. Boyle EC, Finlay BB. 2003. Bacterial pathogenesis: exploiting cellularadherence. Curr. Opin. Cell Biol. 15:633– 639. http://dx.doi.org/10.1016/S0955-0674(03)00099-1.

162. Croxen MA, Finlay BB. 2010. Molecular mechanisms of Escherichia colipathogenicity. Nat. Rev. Microbiol. 8:26 –38. http://dx.doi.org/10.1038/nrmicro2265.

163. Grassl GA, Finlay BB. 2008. Pathogenesis of enteric Salmonella infec-tions. Curr. Opin. Gastroenterol. 24:22–26. http://dx.doi.org/10.1097/MOG.0b013e3282f21388.

164. Viswanathan VK, Hodges K, Hecht G. 2009. Enteric infection meetsintestinal function: how bacterial pathogens cause diarrhoea. Nat. Rev.Microbiol. 7:110 –119.

165. McCormick BA, Stocker BA, Laux DC, Cohen PS. 1988. Roles ofmotility, chemotaxis, and penetration through and growth in intestinalmucus in the ability of an avirulent strain of Salmonella typhimurium tocolonize the large intestine of streptomycin-treated mice. Infect. Immun.56:2209 –2217.

166. Stecher B, Barthel M, Schlumberger MC, Haberli L, Rabsch W, Kre-mer M, Hardt WD. 2008. Motility allows S. Typhimurium to benefitfrom the mucosal defence. Cell. Microbiol. 10:1166 –1180. http://dx.doi.org/10.1111/j.1462-5822.2008.01118.x.

167. van Asten FJ, Hendriks HG, Koninkx JF, van Dijk JE. 2004. Flagella-mediated bacterial motility accelerates but is not required for Salmonellaserotype Enteritidis invasion of differentiated Caco-2 cells. Int. J. Med.Microbiol. 294:395–399. http://dx.doi.org/10.1016/j.ijmm.2004.07.012.

168. Aldridge P, Hughes KT. 2002. Regulation of flagellar assembly. Curr.Opin. Microbiol. 5:160 –165. http://dx.doi.org/10.1016/S1369-5274(02)00302-8.

169. Chevance FF, Hughes KT. 2008. Coordinating assembly of a bacterialmacromolecular machine. Nat. Rev. Microbiol. 6:455– 465. http://dx.doi.org/10.1038/nrmicro1887.

170. Navarro-Garcia F, Elias WP. 2011. Autotransporters and virulence ofenteroaggregative E. coli. Gut Microbes 2:13–24. http://dx.doi.org/10.4161/gmic.2.1.14933.

171. Law RJ, Gur-Arie L, Rosenshine I, Finlay BB. 2013. In vitro and in vivo



on Novem


r.asm.org/

Dow

nloaded from




http://dx.doi.org/10.1111/j.1574-6968.2006.00250.x

http://dx.doi.org/10.1111/j.1574-6968.2006.00250.x

http://dx.doi.org/10.1097/MPG.0b013e3181924d1e

http://dx.doi.org/10.1097/MPG.0b013e3181924d1e

http://dx.doi.org/10.1002/jobm.201000130

http://dx.doi.org/10.4014/jmb.1208.08005



http://dx.doi.org/10.1016/j.mimet.2008.01.014




http://dx.doi.org/10.1128/CDLI.12.12.1378-1386.2005

http://dx.doi.org/10.1159/000007660

http://dx.doi.org/10.1139/w01-013

http://dx.doi.org/10.1139/w01-013

http://dx.doi.org/10.1099/jmm.0.009407-0

http://dx.doi.org/10.1111/j.1365-2672.2008.04052.x

http://dx.doi.org/10.1128/AAC.39.8.1859

http://dx.doi.org/10.1128/AAC.48.6.1983-1988.2004

http://dx.doi.org/10.1016/S0924-8579(00)00188-6

http://dx.doi.org/10.1016/S0924-8579(00)00188-6

http://dx.doi.org/10.1146/annurev-micro-090110-102908

http://dx.doi.org/10.1271/bbb.110921

http://dx.doi.org/10.1271/bbb.110921

http://dx.doi.org/10.1016/S0955-0674(03)00099-1

http://dx.doi.org/10.1016/S0955-0674(03)00099-1



http://dx.doi.org/10.1097/MOG.0b013e3282f21388

http://dx.doi.org/10.1097/MOG.0b013e3282f21388

http://dx.doi.org/10.1111/j.1462-5822.2008.01118.x

http://dx.doi.org/10.1111/j.1462-5822.2008.01118.x

http://dx.doi.org/10.1016/j.ijmm.2004.07.012

http://dx.doi.org/10.1016/S1369-5274(02)00302-8

http://dx.doi.org/10.1016/S1369-5274(02)00302-8





http://cmr.asm.org

http://cmr.asm.org/

model systems for studying enteropathogenic Escherichia coli infections.Cold Spring Harb. Perspect. Med. 3:a009977. http://dx.doi.org/10.1101/cshperspect.a009977.

172. Navarro-Garcia, F., Serapio-Palacios, A., Ugalde-Silva, P., Tapia-Pastrana, G., Chavez-Duenas, L. 2013. Actin cytoskeleton manipulationby effector proteins secreted by diarrheagenic Escherichia coli pathotypes.BioMed Res. Int. 2013:374395. http://dx.doi.org/10.1155/2013/374395.

173. Guttman JA, Finlay BB. 2009. Tight junctions as targets of infectiousagents. Biochim. Biophys. Acta 1788:832– 841. http://dx.doi.org/10.1016/j.bbamem.2008.10.028.

174. Stavru F, Archambaud C, Cossart P. 2011. Cell biology and immunol-ogy of Listeria monocytogenes infections: novel insights. Immunol. Rev.240:160 –184. http://dx.doi.org/10.1111/j.1600-065X.2010.00993.x.

175. Ramsden AE, Holden DW, Mota LJ. 2007. Membrane dynamics andspatial distribution of Salmonella-containing vacuoles. Trends Micro-biol. 15:516 –524. http://dx.doi.org/10.1016/j.tim.2007.10.002.

176. Lievin-Le Moal V, Servin AL. 2013. Pathogenesis of human enteroviru-lent bacteria: lessons from cultured human fully-differentiated coloncancer cell lines. Microbiol. Mol. Biol. Rev. 77:380 – 439. http://dx.doi.org/10.1128/MMBR.00064-12.

177. Lee YK, Lim CY, Teng WL, Ouwehand AC, Tuomola EM, Salminen S.2000. Quantitative approach in the study of adhesion of lactic acid bac-teria to intestinal cells and their competition with enterobacteria. Appl.Environ. Microbiol. 66:3692–3697. http://dx.doi.org/10.1128/AEM.66.9.3692-3697.2000.

178. Lee YK, Puong KY, Ouwehand AC, Salminen S. 2003. Displacement ofbacterial pathogens from mucus and Caco-2 cell surface by lactobacilli. J.Med. Microbiol. 52:925–930. http://dx.doi.org/10.1099/jmm.0.05009-0.

179. Larsen N, Nissen P, Willats WG. 2007. The effect of calcium ions onadhesion and competitive exclusion of Lactobacillus ssp. and E. coli O138.Int. J. Food Microbiol. 114:113–119. http://dx.doi.org/10.1016/j.ijfoodmicro.2006.10.033.

180. Bernet MF, Brassart D, Neeser JR, Servin AL. 1994. Lactobacillusacidophilus LA 1 binds to cultured human intestinal cell lines and inhibitscell attachment and cell invasion by enterovirulent bacteria. Gut 35:483–489. http://dx.doi.org/10.1136/gut.35.4.483.

181. Chauviere G, Coconnier MH, Kerneis S, Darfeuille-Michaud A, Joly B,Servin AL. 1992. Competitive exclusion of diarrheagenic Escherichia coli(ETEC) from human enterocyte-like Caco-2 cells by heat-killed Lactoba-cillus. FEMS Microbiol. Lett. 70:213–217.

182. Coconnier MH, Bernet MF, Chauviere G, Servin AL. 1993. Adheringheat-killed human Lactobacillus acidophilus, strain LB, inhibits the pro-cess of pathogenicity of diarrhoeagenic bacteria in cultured human in-testinal cells. J. Diarrhoeal Dis. Res. 11:235–242.

183. Coconnier MH, Bernet MF, Kerneis S, Chauviere G, Fourniat J, ServinAL. 1993. Inhibition of adhesion of enteroinvasive pathogens to humanintestinal Caco-2 cells by Lactobacillus acidophilus strain LB decreasesbacterial invasion. FEMS Microbiol. Lett. 110:299 –305. http://dx.doi.org/10.1111/j.1574-6968.1993.tb06339.x.

184. Vesterlund S, Paltta J, Karp M, Ouwehand AC. 2005. Adhesion ofbacteria to resected human colonic tissue: quantitative analysis of bacte-rial adhesion and viability. Res. Microbiol. 156:238 –244. http://dx.doi.org/10.1016/j.resmic.2004.08.012.

185. Ingrassia I, Leplingard A, Darfeuille-Michaud A. 2005. Lactobacilluscasei DN-114 001 inhibits the ability of adherent-invasive Escherichia coliisolated from Crohn’s disease patients to adhere to and to invade intes-tinal epithelial cells. Appl. Environ. Microbiol. 71:2880 –2887. http://dx.doi.org/10.1128/AEM.71.6.2880-2887.2005.

186. Parassol N, Freitas M, Thoreux K, Dalmasso G, Bourdet-Sicard R,Rampal P. 2005. Lactobacillus casei DN-114 001 inhibits the increase inparacellular permeability of enteropathogenic Escherichia coli-infectedT84 cells. Res. Microbiol. 156:256 –262. http://dx.doi.org/10.1016/j.resmic.2004.09.013.

187. Fourniat J, Colomban C, Linxe C, Karam D. 1992. Heat-killed Lacto-bacillus acidophilus inhibits adhesion of Escherichia coli B41 to HeLa cells.Ann. Rech. Vet. 23:361–370.

188. Chauviere G, Coconnier MH, Kerneis S, Fourniat J, Servin AL. 1992.Adhesion of human Lactobacillus acidophilus strain LB to human entero-cyte-like Caco-2 cells. J. Gen. Microbiol. 138:1689 –1696. http://dx.doi.org/10.1099/00221287-138-8-1689.

189. Tuomola EM, Ouwehand AC, Salminen SJ. 1999. The effect of probi-otic bacteria on the adhesion of pathogens to human intestinal mucus.

FEMS Immunol. Med. Microbiol. 26:137–142. http://dx.doi.org/10.1111/j.1574-695X.1999.tb01381.x.

190. Neeser JR, Granato D, Rouvet M, Servin A, Teneberg S, Karlsson KA.2000. Lactobacillus johnsonii La1 shares carbohydrate-binding specifici-ties with several enteropathogenic bacteria. Glycobiology 10:1193–1199.http://dx.doi.org/10.1093/glycob/10.11.1193.

191. Kankainen M, Paulin L, Tynkkynen S, von Ossowski I, Reunanen J,Partanen P, Satokari R, Vesterlund S, Hendrickx AP, Lebeer S, DeKeersmaecker SC, Vanderleyden J, Hamalainen T, Laukkanen S, Sal-ovuori N, Ritari J, Alatalo E, Korpela R, Mattila-Sandholm T, Lassig A,Hatakka K, Kinnunen KT, Karjalainen H, Saxelin M, Laakso K,Surakka A, Palva A, Salusjarvi T, Auvinen P, de Vos WM. 2009.Comparative genomic analysis of Lactobacillus rhamnosus GG reveals pilicontaining a human mucus binding protein. Proc. Natl. Acad. Sci.U. S. A. 106:17193–17198. http://dx.doi.org/10.1073/pnas.0908876106.

192. von Ossowski I, Reunanen J, Satokari R, Vesterlund S, Kankainen M,Huhtinen H, Tynkkynen S, Salminen S, de Vos WM, Palva A. 2010.Mucosal adhesion properties of the probiotic Lactobacillus rhamnosusGG SpaCBA and SpaFED pilin subunits. Appl. Environ. Microbiol. 76:2049 –2057. http://dx.doi.org/10.1128/AEM.01958-09.

193. Reunanen J, von Ossowski I, Hendrickx AP, Palva A, de Vos WM.2012. Characterization of the SpaCBA pilus fibers in the probiotic Lac-tobacillus rhamnosus GG. Appl. Environ. Microbiol. 78:2337–2344. http://dx.doi.org/10.1128/AEM.07047-11.

194. Tripathi P, Beaussart A, Alsteens D, Dupres V, Claes I, von OssowskiI, de Vos WM, Palva A, Lebeer S, Vanderleyden J, Dufrene YF. 2013.Adhesion and nanomechanics of pili from the probiotic Lactobacillusrhamnosus GG. ACS Nano. 7:3685–3697. http://dx.doi.org/10.1021/nn400705u.

195. Lebeer S, Claes I, Tytgat HL, Verhoeven TL, Marien E, von OssowskiI, Reunanen J, Palva A, Vos WM, Keersmaecker SC, Vanderleyden J.2012. Functional analysis of Lactobacillus rhamnosus GG pili in relationto adhesion and immunomodulatory interactions with intestinal epithe-lial cells. Appl. Environ. Microbiol. 78:185–193. http://dx.doi.org/10.1128/AEM.06192-11.

196. Tripathi P, Dupres V, Beaussart A, Lebeer S, Claes IJ, Vanderleyden J,Dufrene YF. 2012. Deciphering the nanometer-scale organization andassembly of Lactobacillus rhamnosus GG pili using atomic force micros-copy. Langmuir 28:2211–2216. http://dx.doi.org/10.1021/la203834d.

197. von Ossowski I, Satokari R, Reunanen J, Lebeer S, De KeersmaeckerSC, Vanderleyden J, de Vos WM, Palva A. 2011. Functional character-ization of a mucus-specific LPXTG surface adhesin from probiotic Lac-tobacillus rhamnosus GG. Appl. Environ. Microbiol. 77:4465– 4472. http://dx.doi.org/10.1128/AEM.02497-10.

198. Finlay BB, Falkow S. 1990. Salmonella interactions with polarized hu-man intestinal Caco-2 epithelial cells. J. Infect. Dis. 162:1096 –1106. http://dx.doi.org/10.1093/infdis/162.5.1096.

199. Finlay BB, Ruschkowski S, Dedhar S. 1991. Cytoskeletal rearrange-ments accompanying Salmonella entry into epithelial cells. J. Cell Sci.99:283–296.

200. Jones BD, Lee CA, Falkow S. 1992. Invasion by Salmonella typhimuriumis affected by the direction of flagellar rotation. Infect. Immun. 60:2475–2480.

201. Watson KG, Holden DW. 2010. Dynamics of growth and disseminationof Salmonella in vivo. Cell. Microbiol. 12:1389 –1397. http://dx.doi.org/10.1111/j.1462-5822.2010.01511.x.

202. Abu Kwaik Y, Bumann D. 2013. Microbial quest for food in vivo:‘nutritional virulence’ as an emerging paradigm. Cell. Microbiol. 15:882–890. http://dx.doi.org/10.1111/cmi.12138.

203. Lewis K. 2007. Persister cells, dormancy and infectious disease. Nat. Rev.Microbiol. 5:48 –56. http://dx.doi.org/10.1038/nrmicro1557.

204. Louvard D, Kedinger M, Hauri HP. 1992. The differentiating intestinalepithelial cell: establishment and maintenance of functions through in-teractions between cellular structures. Annu. Rev. Cell Biol. 8:157–195.http://dx.doi.org/10.1146/annurev.cellbio.8.1.157.

205. Servin AL. 2005. Pathogenesis of Afa/Dr diffusely adhering Escherichiacoli. Clin. Microbiol. Rev. 18:264 –292. http://dx.doi.org/10.1128/CMR.18.2.264-292.2005.

206. Guignot J, Chaplais C, Coconnier-Polter MH, Servin AL. 2007. Thesecreted autotransporter toxin, Sat, functions as a virulence factor inAfa/Dr diffusely adhering Escherichia coli by promoting lesions in tightjunction of polarized epithelial cells. Cell. Microbiol. 9:204 –221. http://dx.doi.org/10.1111/j.1462-5822.2006.00782.x.



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1101/cshperspect.a009977


http://dx.doi.org/10.1155/2013/374395

http://dx.doi.org/10.1016/j.bbamem.2008.10.028

http://dx.doi.org/10.1016/j.bbamem.2008.10.028


http://dx.doi.org/10.1016/j.tim.2007.10.002





http://dx.doi.org/10.1099/jmm.0.05009-0



http://dx.doi.org/10.1136/gut.35.4.483









http://dx.doi.org/10.1099/00221287-138-8-1689

http://dx.doi.org/10.1099/00221287-138-8-1689



http://dx.doi.org/10.1093/glycob/10.11.1193





http://dx.doi.org/10.1021/nn400705u

http://dx.doi.org/10.1021/nn400705u



http://dx.doi.org/10.1021/la203834d



http://dx.doi.org/10.1093/infdis/162.5.1096

http://dx.doi.org/10.1093/infdis/162.5.1096

http://dx.doi.org/10.1111/j.1462-5822.2010.01511.x

http://dx.doi.org/10.1111/j.1462-5822.2010.01511.x

http://dx.doi.org/10.1111/cmi.12138


http://dx.doi.org/10.1146/annurev.cellbio.8.1.157



http://dx.doi.org/10.1111/j.1462-5822.2006.00782.x

http://dx.doi.org/10.1111/j.1462-5822.2006.00782.x

http://cmr.asm.org

http://cmr.asm.org/

207. Lievin-Le Moal V, Sarrazin-Davila LE, Servin AL. 2007. An experimen-tal study and a randomized, double-blind, placebo-controlled clinicaltrial to evaluate the antisecretory activity of Lactobacillus acidophilusstrain LB against nonrotavirus diarrhea. Pediatrics 120:e795– 803. http://dx.doi.org/10.1542/peds.2006-2930.

208. Hodges K, Hecht G. 2012. Interspecies communication in the gut, frombacterial delivery to host-cell response. J. Physiol. 590:433– 440. http://dx.doi.org/10.1113/jphysiol.2011.220822.

209. Ruas-Madiedo P, Medrano M, Salazar N, De Los Reyes-Gavilan CG,Perez PF, Abraham AG. 2010. Exopolysaccharides produced by Lacto-bacillus and Bifidobacterium strains abrogate in vitro the cytotoxic effectof bacterial toxins on eukaryotic cells. J. Appl. Microbiol. 109:2079 –2086. http://dx.doi.org/10.1111/j.1365-2672.2010.04839.x.

210. Tao Y, Drabik KA, Waypa TS, Musch MW, Alverdy JC, SchneewindO, Chang EB, Petrof EO. 2006. Soluble factors from Lactobacillus GGactivate MAPKs and induce cytoprotective heat shock proteins in intes-tinal epithelial cells. Am. J. Physiol. Cell Physiol. 290:C1018 –C1030.http://dx.doi.org/10.1152/ajpcell.00131.2005.

211. Montalto M, Maggiano N, Ricci R, Curigliano V, Santoro L, DiNicuolo F, Vecchio FM, Gasbarrini A, Gasbarrini G. 2004. Lactobacil-lus acidophilus protects tight junctions from aspirin damage in HT-29cells. Digestion 69:225–228. http://dx.doi.org/10.1159/000079152.

212. Seth A, Yan F, Polk DB, Rao RK. 2008. Probiotics ameliorate the hydrogenperoxide-induced epithelial barrier disruption by a PKC- and MAP kinase-dependent mechanism. Am. J. Physiol. Gastrointest. Liver Physiol. 294:G1060–G1069. http://dx.doi.org/10.1152/ajpgi.00202.2007.

213. Yan F, Cao H, Cover TL, Whitehead R, Washington MK, Polk DB.2007. Soluble proteins produced by probiotic bacteria regulate intestinalepithelial cell survival and growth. Gastroenterology 132:562–575. http://dx.doi.org/10.1053/j.gastro.2006.11.022.

214. Anderson JM, Van Itallie CM. 2009. Physiology and function of thetight junction. Cold Spring Harb. Perspect. Biol. 1:a002584. http://dx.doi.org/10.1101/cshperspect.a002584.

215. Johnson-Henry KC, Donato KA, Shen-Tu G, Gordanpour M, Sher-man PM. 2008. Lactobacillus rhamnosus strain GG prevents enterohem-orrhagic Escherichia coli O157:H7-induced changes in epithelial barrierfunction. Infect. Immun. 76:1340 –1348. http://dx.doi.org/10.1128/IAI.00778-07.

216. Hofman PM. 2010. Pathobiology of the neutrophil-intestinal epithelialcell interaction: role in carcinogenesis. World J. Gastroenterol. 16:5790 –5800. http://dx.doi.org/10.3748/wjg.v16.i46.5790.

217. Ohland CL, Macnaughton WK. 2010. Probiotic bacteria and intestinalepithelial barrier function. Am. J. Physiol. Gastrointest. Liver Physiol.298:G807–G819. http://dx.doi.org/10.1152/ajpgi.00243.2009.

218. Lopez M, Li N, Kataria J, Russell M, Neu J. 2008. Live and ultraviolet-inactivated Lactobacillus rhamnosus GG decrease flagellin-induced inter-leukin-8 production in Caco-2 cells. J. Nutr. 138:2264 –2268. http://dx.doi.org/10.3945/jn.108.093658.

219. Nandakumar NS, Pugazhendhi S, Madhu Mohan K, Jayakanthan K,Ramakrishna BS. 2009. Effect of Vibrio cholerae on chemokine geneexpression in HT29 cells and its modulation by Lactobacillus GG. Scand.J. Immunol. 69:181–187. http://dx.doi.org/10.1111/j.1365-3083.2008.02214.x.

220. Nandakumar NS, Pugazhendhi S, Ramakrishna BS. 2009. Effects ofenteropathogenic bacteria and lactobacilli on chemokine secretion andToll like receptor gene expression in two human colonic epithelial celllines. Indian J. Med. Res. 130:170 –178.

221. Malago JJ, Nemeth E, Koninkx JF, Tooten PC, Fajdiga S, van Dijk JE.2010. Microbial products from probiotic bacteria inhibit Salmonella en-teritidis 857-induced IL-8 synthesis in Caco-2 cells. Folia Microbiol.(Praha) 55:401– 408. http://dx.doi.org/10.1007/s12223-010-0068-8.

222. Toki S, Kagaya S, Shinohara M, Wakiguchi H, Matsumoto T, Taka-hata Y, Morimatsu F, Saito H, Matsumoto K. 2009. Lactobacillusrhamnosus GG and Lactobacillus casei suppress Escherichia coli-inducedchemokine expression in intestinal epithelial cells. Int. Arch. Allergy Im-munol. 148:45–58. http://dx.doi.org/10.1159/000151505.

223. Ho NK, Hawley SP, Ossa JC, Mathieu O, Tompkins TA, Johnson-Henry KC, Sherman PM. 2013. Immune signalling responses in intes-tinal epithelial cells exposed to pathogenic Escherichia coli and lactic acid-producing probiotics. Benef. Microbes 4:195–209. http://dx.doi.org/10.3920/BM2012.0038.

224. Tien MT, Girardin SE, Regnault B, Le Bourhis L, Dillies MA, CoppeeJY, Bourdet-Sicard R, Sansonetti PJ, Pedron T. 2006. Anti-

inflammatory effect of Lactobacillus casei on Shigella-infected human in-testinal epithelial cells. J. Immunol. 176:1228 –1237.

225. Di Caro S, Tao H, Grillo A, Elia C, Gasbarrini G, Sepulveda AR,Gasbarrini A. 2005. Effects of Lactobacillus GG on genes expressionpattern in small bowel mucosa. Dig. Liver Dis. 37:320 –329. http://dx.doi.org/10.1016/j.dld.2004.12.008.

226. Shima T, Fukushima K, Setoyama H, Imaoka A, Matsumoto S, HaraT, Suda K, Umesaki Y. 2008. Differential effects of two probiotic strainswith different bacteriological properties on intestinal gene expression,with special reference to indigenous bacteria. FEMS Immunol. Med. Mi-crobiol. 52:69 –77. http://dx.doi.org/10.1111/j.1574-695X.2007.00344.x.

227. Mattar AF, Teitelbaum DH, Drongowski RA, Yongyi F, Harmon CM,Coran AG. 2002. Probiotics up-regulate MUC-2 mucin gene expressionin a Caco-2 cell-culture model. Pediatr. Surg. Int. 18:586 –590. http://dx.doi.org/10.1007/s00383-002-0855-7.

228. Mack DR, Michail S, Wei S, McDougall L, Hollingsworth MA. 1999.Probiotics inhibit enteropathogenic E. coli adherence in vitro by induc-ing intestinal mucin gene expression. Am. J. Physiol. 276:G941–G950.

229. Hagbom M, Sharma S, Lundgren O, Svensson L. 2012. Towards ahuman rotavirus disease model. Curr. Opin. Virol. 2:408 – 418. http://dx.doi.org/10.1016/j.coviro.2012.05.006.

230. Varyukhina S, Freitas M, Bardin S, Robillard E, Tavan E, Sapin C,Grill JP, Trugnan G. 2012. Glycan-modifying bacteria-derived solublefactors from Bacteroides thetaiotaomicron and Lactobacillus casei inhibitrotavirus infection in human intestinal cells. Microbes Infect. 14:273–278. http://dx.doi.org/10.1016/j.micinf.2011.10.007.

231. Maragkoudakis PA, Chingwaru W, Gradisnik L, Tsakalidou E, CencicA. 2010. Lactic acid bacteria efficiently protect human and animal intes-tinal epithelial and immune cells from enteric virus infection. Int. J. FoodMicrobiol. 141(Suppl 1):S91–S97. http://dx.doi.org/10.1016/j.ijfoodmicro.2009.12.024.

232. Liu F, Li G, Wen K, Bui T, Cao D, Zhang Y, Yuan L. 2010. Porcinesmall intestinal epithelial cell line (IPEC-J2) of rotavirus infection as anew model for the study of innate immune responses to rotaviruses andprobiotics. Viral Immunol. 23:135–149. http://dx.doi.org/10.1089/vim.2009.0088.

233. Servin AL. 2002. Effects of rotavirus infection on the structure and func-tions of intestinal cells, p 237–254. In Desselberger U, Gray J (ed), Viralgastroenteritis. Elsevier Science B.V., Amsterdam, The Netherlands.

234. Bergonzelli GE, Granato D, Pridmore RD, Marvin-Guy LF, DonnicolaD, Corthesy-Theulaz IE. 2006. GroEL of Lactobacillus johnsonii La1(NCC 533) is cell surface associated: potential role in interactions withthe host and the gastric pathogen Helicobacter pylori. Infect. Immun.74:425– 434. http://dx.doi.org/10.1128/IAI.74.1.425-434.2006.

235. Rokka S, Myllykangas S, Joutsjoki V. 2008. Effect of specific colostralantibodies and selected lactobacilli on the adhesion of Helicobacter pylorion AGS cells and the Helicobacter-induced IL-8 production. Scand. J.Immunol. 68:280 –286. http://dx.doi.org/10.1111/j.1365-3083.2008.02138.x.

236. Myllyluoma E, Ahonen AM, Korpela R, Vapaatalo H, Kankuri E. 2008.Effects of multispecies probiotic combination on Helicobacter pylori in-fection in vitro. Clin. Vaccine Immunol. 15:1472–1482. http://dx.doi.org/10.1128/CVI.00080-08.

237. Bhinder G, Sham HP, Chan JM, Morampudi V, Jacobson K, VallanceBA. 2013. The Citrobacter rodentium mouse model: studying pathogenand host contributions to infectious colitis. J. Vis. Exp. 2013:e50222.http://dx.doi.org/10.3791/50222.

238. Borenshtein D, McBee ME, Schauer DB. 2008. Utility of the Citrobacterrodentium infection model in laboratory mice. Curr. Opin. Gastroen-terol. 24:32–37. http://dx.doi.org/10.1097/MOG.0b013e3282f2b0fb.

239. Barthel M, Hapfelmeier S, Quintanilla-Martinez L, Kremer M, RohdeM, Hogardt M, Pfeffer K, Russmann H, Hardt WD. 2003. Pretreat-ment of mice with streptomycin provides a Salmonella enterica serovarTyphimurium colitis model that allows analysis of both pathogen andhost. Infect. Immun. 71:2839 –2858. http://dx.doi.org/10.1128/IAI.71.5.2839-2858.2003.

240. Kaiser P, Diard M, Stecher B, Hardt WD. 2012. The streptomycinmouse model for Salmonella diarrhea: functional analysis of the micro-biota, the pathogen’s virulence factors, and the host’s mucosal immuneresponse. Immunol. Rev. 245:56 – 83. http://dx.doi.org/10.1111/j.1600-065X.2011.01070.x.

241. Chen CC, Louie S, Shi HN, Walker WA. 2005. Preinoculation with theprobiotic Lactobacillus acidophilus early in life effectively inhibits murine



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1542/peds.2006-2930


http://dx.doi.org/10.1113/jphysiol.2011.220822

http://dx.doi.org/10.1113/jphysiol.2011.220822

http://dx.doi.org/10.1111/j.1365-2672.2010.04839.x

http://dx.doi.org/10.1152/ajpcell.00131.2005

http://dx.doi.org/10.1159/000079152

http://dx.doi.org/10.1152/ajpgi.00202.2007





http://dx.doi.org/10.1128/IAI.00778-07


http://dx.doi.org/10.3748/wjg.v16.i46.5790

http://dx.doi.org/10.1152/ajpgi.00243.2009

http://dx.doi.org/10.3945/jn.108.093658

http://dx.doi.org/10.3945/jn.108.093658

http://dx.doi.org/10.1111/j.1365-3083.2008.02214.x

http://dx.doi.org/10.1111/j.1365-3083.2008.02214.x

http://dx.doi.org/10.1007/s12223-010-0068-8

http://dx.doi.org/10.1159/000151505

http://dx.doi.org/10.3920/BM2012.0038

http://dx.doi.org/10.3920/BM2012.0038

http://dx.doi.org/10.1016/j.dld.2004.12.008

http://dx.doi.org/10.1016/j.dld.2004.12.008


http://dx.doi.org/10.1007/s00383-002-0855-7

http://dx.doi.org/10.1007/s00383-002-0855-7

http://dx.doi.org/10.1016/j.coviro.2012.05.006


http://dx.doi.org/10.1016/j.micinf.2011.10.007



http://dx.doi.org/10.1089/vim.2009.0088

http://dx.doi.org/10.1089/vim.2009.0088

http://dx.doi.org/10.1128/IAI.74.1.425-434.2006

http://dx.doi.org/10.1111/j.1365-3083.2008.02138.x

http://dx.doi.org/10.1111/j.1365-3083.2008.02138.x

http://dx.doi.org/10.1128/CVI.00080-08

http://dx.doi.org/10.1128/CVI.00080-08

http://dx.doi.org/10.3791/50222

http://dx.doi.org/10.1097/MOG.0b013e3282f2b0fb





http://cmr.asm.org

http://cmr.asm.org/

Citrobacter rodentium colitis. Pediatr. Res. 58:1185–1191. http://dx.doi.org/10.1203/01.pdr.0000183660.39116.83.

242. Mackos AR, Eubank TD, Parry NM, Bailey MT. 2013. Probiotic Lac-tobacillus reuteri attenuates the stressor-enhanced severity of Citrobacterrodentium infection. Infect. Immun. 81:3253–3263. http://dx.doi.org/10.1128/IAI.00278-13.

243. Balla KM, Troemel ER. 2013. Caenorhabditis elegans as a model forintracellular pathogen infection. Cell. Microbiol. 15:1313–1322. http://dx.doi.org/10.1111/cmi.12152.

244. Clark LC, Hodgkin J. 29 October 2013. Commensals, probiotics andpathogens in the Caenorhabditis elegans model. Cell. Microbiol. http://dx.doi.org/10.1111/cmi.12234.

245. Ikeda T, Yasui C, Hoshino K, Arikawa K, Nishikawa Y. 2007. Influenceof lactic acid bacteria on longevity of Caenorhabditis elegans and hostdefense against Salmonella enterica serovar enteritidis. Appl. Environ.Microbiol. 73:6404 – 6409. http://dx.doi.org/10.1128/AEM.00704-07.

246. Kim Y, Mylonakis E. 2012. Caenorhabditis elegans immune condition-ing with the probiotic bacterium Lactobacillus acidophilus strain NCFMenhances gram-positive immune responses. Infect. Immun. 80:2500 –2508. http://dx.doi.org/10.1128/IAI.06350-11.

247. Komura T, Ikeda T, Yasui C, Saeki S, Nishikawa Y. 2013. Mechanismunderlying prolongevity induced by bifidobacteria in Caenorhabditis el-egans. Biogerontology. 14:73– 87. http://dx.doi.org/10.1007/s10522-012-9411-6.

248. Zhao Y, Zhao L, Zheng X, Fu T, Guo H, Ren F. 2013. Lactobacillussalivarius strain FDB89 induced longevity in Caenorhabditis elegans bydietary restriction. J. Microbiol. 51:183–188. http://dx.doi.org/10.1007/s12275-013-2076-2.

249. Fourniat J, Djaballi Z, Maccario J, Bourlioux P, German A. 1986.Effect of the administration of killed Lactobacillus acidophilus on thesurvival of suckling mice infected with a strain of enterotoxigenic Esche-richia coli. Ann. Rech. Vet. 17:401– 407.

250. Sherman MP, Bennett SH, Hwang FF, Yu C. 2004. Neonatal smallbowel epithelia: enhancing anti-bacterial defense with lactoferrin andLactobacillus GG. Biometals 17:285–289. http://dx.doi.org/10.1023/B:BIOM.0000027706.51112.62.

251. Ogawa M, Shimizu K, Nomoto K, Takahashi M, Watanuki M, TanakaR, Tanaka T, Hamabata T, Yamasaki S, Takeda Y. 2001. Protectiveeffect of Lactobacillus casei strain Shirota on Shiga toxin-producing Esch-erichia coli O157:H7 infection in infant rabbits. Infect. Immun. 69:1101–1108. http://dx.doi.org/10.1128/IAI.69.2.1101-1108.2001.

252. Zhang L, Xu YQ, Liu HY, Lai T, Ma JL, Wang JF, Zhu YH. 2010.Evaluation of Lactobacillus rhamnosus GG using an Escherichia coli K88model of piglet diarrhoea: Effects on diarrhoea incidence, faecal micro-flora and immune responses. Vet. Microbiol. 141:142–148. http://dx.doi.org/10.1016/j.vetmic.2009.09.003.

253. Moyen EN, Bonneville F, Fauchere JL. 1986. Modification of intestinalcolonization and translocation of Campylobacter jejuni by erythromycinand an extract of Lactobacillus acidophilus in axenic mice. Ann. Inst.Pasteur Microbiol. 137A:199 –207.

254. de Waard R, Garssen J, Bokken GC, Vos JG. 2002. Antagonistic activityof Lactobacillus casei strain Shirota against gastrointestinal Listeria mono-cytogenes infection in rats. Int. J. Food Microbiol. 73:93–100. http://dx.doi.org/10.1016/S0168-1605(01)00699-7.

255. Depardieu F, Podglajen I, Leclercq R, Collatz E, Courvalin P. 2007.Modes and modulations of antibiotic resistance gene expression. Clin.Microbiol. Rev. 20:79 –114. http://dx.doi.org/10.1128/CMR.00015-06.

256. Fernandez L, Hancock RE. 2012. Adaptive and mutational resistance:role of porins and efflux pumps in drug resistance. Clin. Microbiol. Rev.25:661– 681. http://dx.doi.org/10.1128/CMR.00043-12.

257. Balaban NQ, Merrin J, Chait R, Kowalik L, Leibler S. 2004. Bacterialpersistence as a phenotypic switch. Science 305:1622–1625. http://dx.doi.org/10.1126/science.1099390.

258. Lewis K. 2010. Persister cells. Annu. Rev. Microbiol. 64:357–372. http://dx.doi.org/10.1146/annurev.micro.112408.134306.

259. Lam EK, Yu L, Wong HP, Wu WK, Shin VY, Tai EK, So WH, WooPC, Cho CH. 2007. Probiotic Lactobacillus rhamnosus GG enhancesgastric ulcer healing in rats. Eur. J. Pharmacol. 565:171–179. http://dx.doi.org/10.1016/j.ejphar.2007.02.050.

260. Deriu E, Liu JZ, Pezeshki M, Edwards RA, Ochoa RJ, Contreras H,Libby SJ, Fang FC, Raffatellu M. 2013. Probiotic bacteria reduce Sal-monella typhimurium intestinal colonization by competing for iron. CellHost Microbe 14:26 –37. http://dx.doi.org/10.1016/j.chom.2013.06.007.

261. Jacobi CA, Malfertheiner P. 2011. Escherichia coli Nissle 1917 (Muta-flor): new insights into an old probiotic bacterium. Dig. Dis. 29:600 –607. http://dx.doi.org/10.1159/000333307.

262. Corr SC, Li Y, Riedel CU, O’Toole PW, Hill C, Gahan CG. 2007.Bacteriocin production as a mechanism for the antiinfective activity ofLactobacillus salivarius UCC118. Proc. Natl. Acad. Sci. U. S. A. 104:7617–7621. http://dx.doi.org/10.1073/pnas.0700440104.

263. Isolauri E, Kaila M, Arvola T, Majamaa H, Rantala I, Virtanen E,Arvilommi H. 1993. Diet during rotavirus enteritis affects jejunal per-meability to macromolecules in suckling rats. Pediatr. Res. 33:548 –553.http://dx.doi.org/10.1203/00006450-199306000-00002.

264. Pant N, Marcotte H, Brussow H, Svensson L, Hammarstrom L. 2007.Effective prophylaxis against rotavirus diarrhea using a combination ofLactobacillus rhamnosus GG and antibodies. BMC Microbiol. 7:86. http://dx.doi.org/10.1186/1471-2180-7-86.

265. Zhang Z, Xiang Y, Li N, Wang B, Ai H, Wang X, Huang L, Zheng Y.2013. Protective effects of Lactobacillus rhamnosus GG against humanrotavirus-induced diarrhoea in a neonatal mouse model. Pathog. Dis.67:184 –191. http://dx.doi.org/10.1111/2049-632X.12030.

266. Guerin-Danan C, Meslin JC, Chambard A, Charpilienne A, Relano P,Bouley C, Cohen J, Andrieux C. 2001. Food supplementation with milkfermented by Lactobacillus casei DN-114 001 protects suckling rats fromrotavirus-associated diarrhea. J. Nutr. 131:111–117.

267. Preidis GA, Saulnier DM, Blutt SE, Mistretta TA, Riehle KP, MajorAM, Venable SF, Barrish JP, Finegold MJ, Petrosino JF, Guerrant RL,Conner ME, Versalovic J. 2012. Host response to probiotics determinedby nutritional status of rotavirus-infected neonatal mice. J. Pediatr. Gas-troenterol. Nutr. 55:299 –307. http://dx.doi.org/10.1097/MPG.0b013e31824d2548.

268. Sanders ME, Levy DD. 2011. The science and regulations of probioticfood and supplement product labeling. Ann. N. Y. Acad. Sci. 1219(Suppl1):E1–E23. http://dx.doi.org/10.1111/j.1749-6632.2010.05956.x.

269. Agostoni C, Axelsson I, Braegger C, Goulet O, Koletzko B, MichaelsenKF, Rigo J, Shamir R, Szajewska H, Turck D, Weaver LT, NutritionECo. 2004. Probiotic bacteria in dietetic products for infants: a commen-tary by the ESPGHAN Committee on Nutrition. J. Pediatr. Gastroen-terol. Nutr. 38:365–374. http://dx.doi.org/10.1097/00005176-200404000-00001.

270. Goldin BR, Gorbach SL, Saxelin M, Barakat S, Gualtieri L, SalminenS. 1992. Survival of Lactobacillus species (strain GG) in human gastroin-testinal tract. Dig. Dis. Sci. 37:121–128. http://dx.doi.org/10.1007/BF01308354.

271. Millar MR, Bacon C, Smith SL, Walker V, Hall MA. 1993. Enteralfeeding of premature infants with Lactobacillus GG. Arch. Dis. Child.69:483– 487. http://dx.doi.org/10.1136/adc.69.5_Spec_No.483.

272. Sheen P, Oberhelman RA, Gilman RH, Cabrera L, Verastegui M,Madico G. 1995. A placebo-controlled study of Lactobacillus GG coloni-zation in one-to-three-year-old Peruvian children. Am. J. Trop. Med.Hyg. 52:389 –392.

273. Hilton E, Kolakowski P, Singer C, Smith M. 1997. Efficacy of Lactoba-cillus GG as a diarrheal preventive in travelers. J. Travel Med. 4:41– 43.http://dx.doi.org/10.1111/j.1708-8305.1997.tb00772.x.

274. Alander M, Satokari R, Korpela R, Saxelin M, Vilpponen-Salmela T,Mattila-Sandholm T, von Wright A. 1999. Persistence of colonizationof human colonic mucosa by a probiotic strain, Lactobacillus rhamnosusGG, after oral consumption. Appl. Environ. Microbiol. 65:351–354.

275. Dommels YE, Kemperman RA, Zebregs YE, Draaisma RB, Jol A,Wolvers DA, Vaughan EE, Albers R. 2009. Survival of Lactobacillusreuteri DSM 17938 and Lactobacillus rhamnosus GG in the human gas-trointestinal tract with daily consumption of a low-fat probiotic spread.Appl. Environ. Microbiol. 75:6198 – 6204. http://dx.doi.org/10.1128/AEM.01054-09.

276. Tiengrim S, Leelaporn A, Manatsathit S, Thamlikitkul V. 2012. Via-bility of Lactobacillus casei strain Shirota (LcS) from feces of Thai healthysubjects regularly taking milk product containing LcS. J. Med. Assoc.Thai. 95(Suppl 2):S42–S47.

277. Fujimoto J, Matsuki T, Sasamoto M, Tomii Y, Watanabe K. 2008.Identification and quantification of Lactobacillus casei strain Shirota inhuman feces with strain-specific primers derived from randomly ampli-fied polymorphic DNA. Int. J. Food Microbiol. 126:210 –215. http://dx.doi.org/10.1016/j.ijfoodmicro.2008.05.022.

278. Yuki N, Watanabe K, Mike A, Tagami Y, Tanaka R, Ohwaki M,Morotomi M. 1999. Survival of a probiotic, Lactobacillus casei strain



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1203/01.pdr.0000183660.39116.83

http://dx.doi.org/10.1203/01.pdr.0000183660.39116.83









http://dx.doi.org/10.1007/s10522-012-9411-6

http://dx.doi.org/10.1007/s10522-012-9411-6

http://dx.doi.org/10.1007/s12275-013-2076-2

http://dx.doi.org/10.1007/s12275-013-2076-2

http://dx.doi.org/10.1023/B:BIOM.0000027706.51112.62

http://dx.doi.org/10.1023/B:BIOM.0000027706.51112.62


http://dx.doi.org/10.1016/j.vetmic.2009.09.003

http://dx.doi.org/10.1016/j.vetmic.2009.09.003

http://dx.doi.org/10.1016/S0168-1605(01)00699-7

http://dx.doi.org/10.1016/S0168-1605(01)00699-7



http://dx.doi.org/10.1126/science.1099390

http://dx.doi.org/10.1126/science.1099390

http://dx.doi.org/10.1146/annurev.micro.112408.134306

http://dx.doi.org/10.1146/annurev.micro.112408.134306

http://dx.doi.org/10.1016/j.ejphar.2007.02.050

http://dx.doi.org/10.1016/j.ejphar.2007.02.050

http://dx.doi.org/10.1016/j.chom.2013.06.007

http://dx.doi.org/10.1159/000333307


http://dx.doi.org/10.1203/00006450-199306000-00002

http://dx.doi.org/10.1186/1471-2180-7-86

http://dx.doi.org/10.1186/1471-2180-7-86

http://dx.doi.org/10.1111/2049-632X.12030

http://dx.doi.org/10.1097/MPG.0b013e31824d2548

http://dx.doi.org/10.1097/MPG.0b013e31824d2548

http://dx.doi.org/10.1111/j.1749-6632.2010.05956.x

http://dx.doi.org/10.1097/00005176-200404000-00001

http://dx.doi.org/10.1097/00005176-200404000-00001

http://dx.doi.org/10.1007/BF01308354

http://dx.doi.org/10.1007/BF01308354

http://dx.doi.org/10.1136/adc.69.5_Spec_No.483






http://cmr.asm.org

http://cmr.asm.org/

Shirota, in the gastrointestinal tract: selective isolation from faeces andidentification using monoclonal antibodies. Int. J. Food Microbiol. 48:51–57. http://dx.doi.org/10.1016/S0168-1605(99)00029-X.

279. Rochet V, Rigottier-Gois L, Levenez F, Cadiou J, Marteau P, BressonJL, Goupil-Feillerat N, Dore J. 2008. Modulation of Lactobacillus casei inileal and fecal samples from healthy volunteers after consumption of afermented milk containing Lactobacillus casei DN-114 001Rif. Can. J.Microbiol. 54:660 – 667. http://dx.doi.org/10.1139/W08-050.

280. Oozeer R, Leplingard A, Mater DD, Mogenet A, Michelin R, Seksek I,Marteau P, Dore J, Bresson JL, Corthier G. 2006. Survival of Lactoba-cillus casei in the human digestive tract after consumption of fermentedmilk. Appl. Environ. Microbiol. 72:5615–5617. http://dx.doi.org/10.1128/AEM.00722-06.

281. Abrahamsson TR, Sinkiewicz G, Jakobsson T, Fredrikson M, Bjork-sten B. 2009. Probiotic lactobacilli in breast milk and infant stool inrelation to oral intake during the first year of life. J. Pediatr. Gastroen-terol. Nutr. 49:349 –354. http://dx.doi.org/10.1097/MPG.0b013e31818f091b.

282. Shornikova AV, Casas IA, Mykkanen H, Salo E, Vesikari T. 1997.Bacteriotherapy with Lactobacillus reuteri in rotavirus gastroenteritis. Pe-diatr. Infect. Dis. J. 16:1103–1107. http://dx.doi.org/10.1097/00006454-199712000-00002.

283. Holmgren J, Svennerholm AM. 2012. Vaccines against mucosal infec-tions. Curr. Opin. Immunol. 24:343–353. http://dx.doi.org/10.1016/j.coi.2012.03.014.

284. Shaw AR. 2013. The rotavirus saga revisited. Annu. Rev. Med. 64:165–174. http://dx.doi.org/10.1146/annurev-med-121511-093810.

285. Estes MK, Desselberger U. 2012. Rotaviruses: cause of vaccine prevent-able disease yet many fundamental questions remain to be explored.Curr. Opin. Virol. 2:369 –372. http://dx.doi.org/10.1016/j.coviro.2012.06.002.

286. Greenberg HB, Estes MK. 2009. Rotaviruses: from pathogenesis to vac-cination. Gastroenterology 136:1939 –1951. http://dx.doi.org/10.1053/j.gastro.2009.02.076.

287. Soares-Weiser K, Maclehose H, Bergman H, Ben-Aharon I, Nagpal S,Goldberg E, Pitan F, Cunliffe N. 2012. Vaccines for preventing rotavirusdiarrhoea: vaccines in use. Cochrane Database Syst. Rev. 2:CD008521.http://dx.doi.org/10.1002/14651858.CD008521.

288. Tate JE, Burton AH, Boschi-Pinto C, Steele AD, Duque J, ParasharUD, WHO-Coordinated Global Rotavirus Surveillance Network.2012. 2008 estimate of worldwide rotavirus-associated mortality in chil-dren younger than 5 years before the introduction of universal rotavirusvaccination programmes: a systematic review and meta-analysis. LancetInfect. Dis. 12:136 –141. http://dx.doi.org/10.1016/S1473-3099(11)70253-5.

289. Vesikari T, Van Damme P, Giaquinto C, Gray J, Mrukowicz J, Dagan R,Guarino A, Szajewska H, Usonis V. 2008. European Society for PaediatricInfectious Diseases/European Society for Paediatric Gastroenterology,Hepatology, and Nutrition evidence-based recommendations for rotavirusvaccination in Europe. J. Pediatr. Gastroenterol. Nutr. 46(Suppl 2):S38–S48.http://dx.doi.org/10.1097/MPG.0b013e31816f7a10.

290. WHO. 2009. Rotavirus vaccines: an update. Wkly. Epidemiol. Rec. 84:533–540. http://dx.doi.org/10.1016/j.vaccine.2007.01.102.

291. WHO. 2009. Meeting of the Strategic Advisory Group of Experts onImmunization, October 2009 — conclusions and recommendations.Wkly. Epidemiol. Rec. 84:517–532. http://dx.doi.org/10.1016/j.biologicals.2009.12.007.

292. Mrukowicz J, Szajewska H, Vesikari T. 2008. Options for the preven-tion of rotavirus disease other than vaccination. J. Pediatr. Gastroenterol.Nutr. 46(Suppl 2):S32–S37. http://dx.doi.org/10.1097/MPG.0b013e31816f79b0.

293. Guarino A, Albano F, Ashkenazi S, Gendrel D, Hoekstra JH, ShamirR, Szajewska H, European Society for Paediatric GastroenterologyHepatology, and Nutrition/European Society for Paediatric InfectiousDiseases. 2008. European Society for Paediatric Gastroenterology,Hepatology, and Nutrition/European Society for Paediatric InfectiousDiseases evidence-based guidelines for the management of acute gastro-enteritis in children in Europe. J. Pediatr. Gastroenterol. Nutr. 46(Suppl2):S81–S122. http://dx.doi.org/10.1097/MPG.0b013e31816f7b16.

294. Menees S, Saad R, Chey WD. 2012. Agents that act luminally to treatdiarrhoea and constipation. Nat. Rev. Gastroenterol. Hepatol. 9:661–674. http://dx.doi.org/10.1038/nrgastro.2012.162.

295. Willing BP, Russell SL, Finlay BB. 2011. Shifting the balance: antibiotic

effects on host-microbiota mutualism. Nat. Rev. Microbiol. 9:233–243.http://dx.doi.org/10.1038/nrmicro2536.

296. Isolauri E, Juntunen M, Rautanen T, Sillanaukee P, Koivula T. 1991.A human Lactobacillus strain (Lactobacillus casei sp strain GG) promotesrecovery from acute diarrhea in children. Pediatrics 88:90 –97.

297. Isolauri E, Kaila M, Mykkanen H, Ling WH, Salminen S. 1994. Oralbacteriotherapy for viral gastroenteritis. Dig. Dis. Sci. 39:2595–2600.http://dx.doi.org/10.1007/BF02087695.

298. Kaila M, Isolauri E, Saxelin M, Arvilommi H, Vesikari T. 1995. Viableversus inactivated Lactobacillus strain GG in acute rotavirus diarrhoea.Arch. Dis. Child. 72:51–53. http://dx.doi.org/10.1136/adc.72.1.51.

299. Majamaa H, Isolauri E, Saxelin M, Vesikari T. 1995. Lactic acid bacteriain the treatment of acute rotavirus gastroenteritis. J. Pediatr. Gastroen-terol. Nutr. 20:333–338. http://dx.doi.org/10.1097/00005176-199504000-00012.

300. Raza S, Graham SM, Allen SJ, Sultana S, Cuevas L, Hart CA. 1995.Lactobacillus GG promotes recovery from acute nonbloody diarrhea inPakistan. Pediatr. Infect. Dis. J. 14:107–111. http://dx.doi.org/10.1097/00006454-199502000-00005.

301. Pant AR, Graham SM, Allen SJ, Harikul S, Sabchareon A, Cuevas L,Hart CA. 1996. Lactobacillus GG and acute diarrhoea in young childrenin the tropics. J. Trop. Pediatr. 42:162–165. http://dx.doi.org/10.1093/tropej/42.3.162.

302. Shornikova AV, Isolauri E, Burkanova L, Lukovnikova S, Vesikari T.1997. A trial in the Karelian Republic of oral rehydration and Lactobacil-lus GG for treatment of acute diarrhoea. Acta Paediatr. 86:460 – 465. http://dx.doi.org/10.1111/j.1651-2227.1997.tb08913.x.

303. Rautanen T, Isolauri E, Salo E, Vesikari T. 1998. Management of acutediarrhoea with low osmolarity oral rehydration solutions and Lactobacil-lus strain GG. Arch. Dis. Child. 79:157–160. http://dx.doi.org/10.1136/adc.79.2.157.

304. Oberhelman RA, Gilman RH, Sheen P, Taylor DN, Black RE, CabreraL, Lescano AG, Meza R, Madico G. 1999. A placebo-controlled trial ofLactobacillus GG to prevent diarrhea in undernourished Peruvian chil-dren. J. Pediatr. 134:15–20. http://dx.doi.org/10.1016/S0022-3476(99)70366-5.

305. Guandalini S, Pensabene L, Zikri MA, Dias JA, Casali LG, Hoekstra H,Kolacek S, Massar K, Micetic-Turk D, Papadopoulou A, de Sousa JS,Sandhu B, Szajewska H, Weizman Z. 2000. Lactobacillus GG adminis-tered in oral rehydration solution to children with acute diarrhea: a mul-ticenter European trial. J. Pediatr. Gastroenterol. Nutr. 30:54 – 60. http://dx.doi.org/10.1097/00005176-200001000-00018.

306. Canani RB, Cirillo P, Terrin G, Cesarano L, Spagnuolo MI, DeVincenzo A, Albano F, Passariello A, De Marco G, Manguso F,Guarino A. 2007. Probiotics for treatment of acute diarrhoea in children:randomised clinical trial of five different preparations. BMJ 335:340.http://dx.doi.org/10.1136/bmj.39272.581736.55.

307. Basu S, Chatterjee M, Ganguly S, Chandra PK. 2007. Effect of Lacto-bacillus rhamnosus GG in persistent diarrhea in Indian children: a ran-domized controlled trial. J. Clin. Gastroenterol. 41:756 –760. http://dx.doi.org/10.1097/01.mcg.0000248009.47526.ea.

308. Basu S, Paul DK, Ganguly S, Chatterjee M, Chandra PK. 2009. Efficacyof high-dose Lactobacillus rhamnosus GG in controlling acute waterydiarrhea in Indian children: a randomized controlled trial. J. Clin. Gas-troenterol. 43:208 –213. http://dx.doi.org/10.1097/MCG.0b013e31815a5780.

309. Parker MW, Schaffzin JK, Lo Vecchio A, Yau C, Vonderhaar K, GuiotA, Brinkman WB, White CM, Simmons JM, Gerhardt WE, KotagalUR, Conway PH. 2013. Rapid adoption of Lactobacillus rhamnosus GGfor acute gastroenteritis. Pediatrics 131(Suppl 1):S96 –S102. http://dx.doi.org/10.1542/peds.2012-1427l.

310. Piescik-Lech M, Urbanska M, Szajewska H. 2013. Lactobacillus GG(LGG) and smectite versus LGG alone for acute gastroenteritis: a double-blind, randomized controlled trial. Eur. J. Pediatr. 172:247–253. http://dx.doi.org/10.1007/s00431-012-1878-2.

311. Szajewska H, Wanke M, Patro B. 2011. Meta-analysis: the effects ofLactobacillus rhamnosus GG supplementation for the prevention ofhealthcare-associated diarrhoea in children. Aliment. Pharmacol. Ther.34:1079 –1087. http://dx.doi.org/10.1111/j.1365-2036.2011.04837.x.

312. Szajewska H, Skorka A, Ruszczynski M, Gieruszczak-Bialek D. 2007.Meta-analysis: Lactobacillus GG for treating acute diarrhoea in children.Aliment. Pharmacol. Ther. 25:871– 881. http://dx.doi.org/10.1111/j.1365-2036.2007.03282.x.



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1016/S0168-1605(99)00029-X

http://dx.doi.org/10.1139/W08-050



http://dx.doi.org/10.1097/MPG.0b013e31818f091b

http://dx.doi.org/10.1097/MPG.0b013e31818f091b

http://dx.doi.org/10.1097/00006454-199712000-00002

http://dx.doi.org/10.1097/00006454-199712000-00002



http://dx.doi.org/10.1146/annurev-med-121511-093810





http://dx.doi.org/10.1002/14651858.CD008521

http://dx.doi.org/10.1016/S1473-3099(11)70253-5

http://dx.doi.org/10.1016/S1473-3099(11)70253-5

http://dx.doi.org/10.1097/MPG.0b013e31816f7a10

http://dx.doi.org/10.1016/j.vaccine.2007.01.102

http://dx.doi.org/10.1016/j.biologicals.2009.12.007

http://dx.doi.org/10.1016/j.biologicals.2009.12.007

http://dx.doi.org/10.1097/MPG.0b013e31816f79b0





http://dx.doi.org/10.1007/BF02087695

http://dx.doi.org/10.1136/adc.72.1.51

http://dx.doi.org/10.1097/00005176-199504000-00012

http://dx.doi.org/10.1097/00005176-199504000-00012

http://dx.doi.org/10.1097/00006454-199502000-00005

http://dx.doi.org/10.1097/00006454-199502000-00005

http://dx.doi.org/10.1093/tropej/42.3.162

http://dx.doi.org/10.1093/tropej/42.3.162





http://dx.doi.org/10.1016/S0022-3476(99)70366-5

http://dx.doi.org/10.1016/S0022-3476(99)70366-5

http://dx.doi.org/10.1097/00005176-200001000-00018

http://dx.doi.org/10.1097/00005176-200001000-00018

http://dx.doi.org/10.1136/bmj.39272.581736.55

http://dx.doi.org/10.1097/01.mcg.0000248009.47526.ea

http://dx.doi.org/10.1097/01.mcg.0000248009.47526.ea

http://dx.doi.org/10.1097/MCG.0b013e31815a5780

http://dx.doi.org/10.1097/MCG.0b013e31815a5780

http://dx.doi.org/10.1542/peds.2012-1427l

http://dx.doi.org/10.1542/peds.2012-1427l

http://dx.doi.org/10.1007/s00431-012-1878-2

http://dx.doi.org/10.1007/s00431-012-1878-2

http://dx.doi.org/10.1111/j.1365-2036.2011.04837.x

http://dx.doi.org/10.1111/j.1365-2036.2007.03282.x

http://dx.doi.org/10.1111/j.1365-2036.2007.03282.x

http://cmr.asm.org

http://cmr.asm.org/

313. Basu S, Chatterjee M, Ganguly S, Chandra PK. 2007. Efficacy ofLactobacillus rhamnosus GG in acute watery diarrhoea of Indian children:a randomised controlled trial. J. Paediatr. Child. Health 43:837– 842.http://dx.doi.org/10.1111/j.1440-1754.2007.01201.x.

314. Salazar-Lindo E, Miranda-Langschwager P, Campos-Sanchez M,Chea-Woo E, Sack RB. 2004. Lactobacillus casei strain GG in the treat-ment of infants with acute watery diarrhea: a randomized, double-blind,placebo controlled clinical trial. BMC Pediatr. 4:18. http://dx.doi.org/10.1186/1471-2431-4-18.

315. Misra S, Sabui TK, Pal NK. 2009. A randomized controlled trial toevaluate the efficacy of Lactobacillus GG in infantile diarrhea. J. Pediatr.155:129 –132. http://dx.doi.org/10.1016/j.jpeds.2009.01.060.

316. Ritchie BK, Brewster DR, Tran CD, Davidson GP, McNeil Y, ButlerRN. 2010. Efficacy of Lactobacillus GG in aboriginal children with acutediarrhoeal disease: a randomised clinical trial. J. Pediatr. Gastroenterol.Nutr. 50:619 – 624. http://dx.doi.org/10.1097/MPG.0b013e3181bbf53d.

317. Shornikova AV, Casas IA, Isolauri E, Mykkanen H, Vesikari T. 1997.Lactobacillus reuteri as a therapeutic agent in acute diarrhea in youngchildren. J. Pediatr. Gastroenterol. Nutr. 24:399 – 404. http://dx.doi.org/10.1097/00005176-199704000-00008.

318. Weizman Z, Asli G, Alsheikh A. 2005. Effect of a probiotic infantformula on infections in child care centers: comparison of two probioticagents. Pediatrics 115:5–9.

319. Francavilla R, Lionetti E, Castellaneta S, Ciruzzi F, Indrio F,Masciale A, Fontana C, La Rosa MM, Cavallo L, Francavilla A.2012. Randomised clinical trial: Lactobacillus reuteri DSM 17938 vs.placebo in children with acute diarrhoea—a double-blind study. Al-iment. Pharmacol. Ther. 36:363–369. http://dx.doi.org/10.1111/j.1365-2036.2012.05180.x.

320. Adams CA. 2010. The probiotic paradox: live and dead cells are biolog-ical response modifiers. Nutr. Res. Rev. 23:37– 46. http://dx.doi.org/10.1017/S0954422410000090.

321. Sanders ME, Hamilton J, Reid G, Gibson G. 2007. A nonviable prep-aration of Lactobacillus acidophilus is not a probiotic. Clin. Infect. Dis.44:886. http://dx.doi.org/10.1086/511694.

322. Lahtinen SJ. 18 June 2012. Probiotic viability— does it matter? Microb.Ecol. Health Dis. http://dx.doi.org/10.3402/mehd.v23i0.18567.

323. Boulloche J, Mouterde O, Mallet E. 1994. Management of acute diar-rhoea in infants and young children: controlled study of the anti-diarrhoeal efficacy of killed L. acidophilus (LB Strain) versus a placeboand a reference drug (loperamide). Ann. Pediatr. 41:457– 463.

324. Bin LX. 1995. Controlled clinical trial in infants and children comparingLacteol Fort sachets with two antidiarrhoeal reference drugs Ann. Pedi-atr. 42:396 – 401.

325. Simakachorn N, Pichaipat V, Rithipornpaisarn P, Kongkaew C, Tong-pradit P, Varavithya W. 2000. Clinical evaluation of the addition oflyophilized, heat-killed Lactobacillus acidophilus LB to oral rehydrationtherapy in the treatment of acute diarrhea in children. J. Pediatr. Gastro-enterol. Nutr. 30:68 –72. http://dx.doi.org/10.1097/00005176-200001000-00020.

326. Costa-Ribeiro H, Ribeiro TC, Mattos AP, Valois SS, Neri DA, AlmeidaP, Cerqueira CM, Ramos E, Young RJ, Vanderhoof JA. 2003. Limita-tions of probiotic therapy in acute, severe dehydrating diarrhea. J. Pedi-atr. Gastroenterol. Nutr. 36:112–115. http://dx.doi.org/10.1097/00005176-200301000-00021.

327. Salazar-Lindo E, Figueroa-Quintanilla D, Caciano MI, Reto-ValienteV, Chauviere G, Colin P. 2007. Effectiveness and safety of LactobacillusLB in the treatment of mild acute diarrhea in children. J. Pediatr. Gastro-enterol. Nutr. 44:571–576. http://dx.doi.org/10.1097/MPG.0b013e3180375594.

328. Xiao SD, Zhang DZ, Lu H, Jiang SH, Liu HY, Wang GS, Xu GM,Zhang ZB, Lin GJ, Wang GL. 2003. Multicenter, randomized, con-trolled trial of heat-killed Lactobacillus acidophilus LB in patients withchronic diarrhea. Adv. Ther. 20:253–260. http://dx.doi.org/10.1007/BF02849854.

329. Sanders ME. 2008. Use of probiotics and yogurts in maintenance ofhealth. J. Clin. Gastroenterol. 42(Suppl 2):S71–S74. http://dx.doi.org/10.1097/MCG.0b013e3181621e87.

330. Hojsak I, Abdovic S, Szajewska H, Milosevic M, Krznaric Z, KolacekS. 2010. Lactobacillus GG in the prevention of nosocomial gastrointesti-nal and respiratory tract infections. Pediatrics 125:e1171–1177. http://dx.doi.org/10.1542/peds.2009-2568.

331. Szajewska H, Kotowska M, Mrukowicz JZ, Armanska M, Mikolajczyk

W. 2001. Efficacy of Lactobacillus GG in prevention of nosocomial diar-rhea in infants. J. Pediatr. 138:361–365. http://dx.doi.org/10.1067/mpd.2001.111321.

332. Mastretta E, Longo P, Laccisaglia A, Balbo L, Russo R, MazzaccaraA, Gianino P. 2002. Effect of Lactobacillus GG and breast-feeding in theprevention of rotavirus nosocomial infection. J. Pediatr. Gastroenterol.Nutr. 35:527–531. http://dx.doi.org/10.1097/00005176-200210000-00013.

333. Hojsak I, Snovak N, Abdovic S, Szajewska H, Misak Z, Kolacek S.2010. Lactobacillus GG in the prevention of gastrointestinal and respira-tory tract infections in children who attend day care centers: a random-ized, double-blind, placebo-controlled trial. Clin. Nutr. 29:312–316.http://dx.doi.org/10.1016/j.clnu.2009.09.008.

334. Sur D, Manna B, Niyogi SK, Ramamurthy T, Palit A, Nomoto K,Takahashi T, Shima T, Tsuji H, Kurakawa T, Takeda Y, Nair GB,Bhattacharya SK. 2011. Role of probiotic in preventing acute diarrhoeain children: a community-based, randomized, double-blind placebo-controlled field trial in an urban slum. Epidemiol. Infect. 139:919 –926.http://dx.doi.org/10.1017/S0950268810001780.

335. Guillemard E, Tondu F, Lacoin F, Schrezenmeir J. 2010. Consumptionof a fermented dairy product containing the probiotic Lactobacillus caseiDN-114001 reduces the duration of respiratory infections in the elderlyin a randomised controlled trial. Br. J. Nutr. 103:58 – 68. http://dx.doi.org/10.1017/S0007114509991395.

336. Merenstein D, Murphy M, Fokar A, Hernandez RK, Park H, Nsouli H,Sanders ME, Davis BA, Niborski V, Tondu F, Shara NM. 2010. Use ofa fermented dairy probiotic drink containing Lactobacillus casei (DN-114001) to decrease the rate of illness in kids: the DRINK study. A patient-oriented, double-blind, cluster-randomized, placebo-controlled, clinicaltrial. Eur. J. Clin. Nutr. 64:669 – 677.

337. Pedone CA, Arnaud CC, Postaire ER, Bouley CF, Reinert P. 2000.Multicentric study of the effect of milk fermented by Lactobacillus caseion the incidence of diarrhoea. Int. J. Clin. Pract. 54:568 –571.

338. Pedone CA, Bernabeu AO, Postaire ER, Bouley CF, Reinert P. 1999.The effect of supplementation with milk fermented by Lactobacillus casei(strain DN-114 001) on acute diarrhoea in children attending day carecentres. Int. J. Clin. Pract. 53:179 –184.

339. Wanke M, Szajewska H. 2012. Lack of an effect of Lactobacillus reuteriDSM 17938 in preventing nosocomial diarrhea in children: a random-ized, double-blind, placebo-controlled trial. J. Pediatr. 161:40 – 43 e41.http://dx.doi.org/10.1016/j.jpeds.2011.12.049.

340. Oksanen PJ, Salminen S, Saxelin M, Hamalainen P, Ihantola-Vormisto A, Muurasniemi-Isoviita L, Nikkari S, Oksanen T, Porsti I,Salminen E, et al. 1990. Prevention of travellers’ diarrhoea by Lactoba-cillus GG. Ann. Med. 22:53–56. http://dx.doi.org/10.3109/07853899009147242.

341. Briand V, Buffet P, Genty S, Lacombe K, Godineau N, Salomon J,Vandemelbrouck E, Ralaimazava P, Goujon C, Matheron S, FontanetA, Bouchaud O. 2006. Absence of efficacy of nonviable Lactobacillusacidophilus for the prevention of traveler’s diarrhea: a randomized, dou-ble-blind, controlled study. Clin. Infect. Dis. 43:1170 –1175. http://dx.doi.org/10.1086/508178.

342. Rupnik M, Wilcox MH, Gerding DN. 2009. Clostridium difficile infec-tion: new developments in epidemiology and pathogenesis. Nat. Rev.Microbiol. 7:526 –536. http://dx.doi.org/10.1038/nrmicro2164.

343. Lessa FC, Gould CV, McDonald LC. 2012. Current status of Clostrid-ium difficile infection epidemiology. Clin. Infect. Dis. 55(Suppl 2):S65–S70. http://dx.doi.org/10.1093/cid/cis319.

344. Mitchell BG, Gardner A. 2012. Mortality and Clostridium difficile infec-tion: a review. Antimicrob. Res. Infect. Contr. 1:20 –26. http://dx.doi.org/10.1186/2047-2994-1-20.

345. Freeman J, Bauer MP, Baines SD, Corver J, Fawley WN, Goorhuis B,Kuijper EJ, Wilcox MH. 2010. The changing epidemiology of Clostrid-ium difficile infections. Clin. Microbiol. Rev. 23:529 –549. http://dx.doi.org/10.1128/CMR.00082-09.

346. Carter GP, Rood JI, Lyras D. 2012. The role of toxin A and toxin B in thevirulence of Clostridium difficile. Trends Microbiol. 20:21–29. http://dx.doi.org/10.1016/j.tim.2011.11.003.

347. Wultanska D, Pituch H, Obuch-Woszczatynski P, Meisel-MikolajczykF, Luczak M. 2006. Influence of selected Lactobacillus sp. on Clostridiumdifficile strains with different toxigenicity profile. Med. Dosw. Mikrobiol.58:127–133.

348. McFarland LV, Elmer GW. 1997. Pharmaceutical probiotics for the



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1111/j.1440-1754.2007.01201.x

http://dx.doi.org/10.1186/1471-2431-4-18

http://dx.doi.org/10.1186/1471-2431-4-18

http://dx.doi.org/10.1016/j.jpeds.2009.01.060

http://dx.doi.org/10.1097/MPG.0b013e3181bbf53d

http://dx.doi.org/10.1097/00005176-199704000-00008

http://dx.doi.org/10.1097/00005176-199704000-00008

http://dx.doi.org/10.1111/j.1365-2036.2012.05180.x

http://dx.doi.org/10.1111/j.1365-2036.2012.05180.x

http://dx.doi.org/10.1017/S0954422410000090

http://dx.doi.org/10.1017/S0954422410000090

http://dx.doi.org/10.1086/511694

http://dx.doi.org/10.3402/mehd.v23i0.18567

http://dx.doi.org/10.1097/00005176-200001000-00020

http://dx.doi.org/10.1097/00005176-200001000-00020

http://dx.doi.org/10.1097/00005176-200301000-00021

http://dx.doi.org/10.1097/00005176-200301000-00021

http://dx.doi.org/10.1097/MPG.0b013e3180375594

http://dx.doi.org/10.1097/MPG.0b013e3180375594

http://dx.doi.org/10.1007/BF02849854

http://dx.doi.org/10.1007/BF02849854

http://dx.doi.org/10.1097/MCG.0b013e3181621e87

http://dx.doi.org/10.1097/MCG.0b013e3181621e87



http://dx.doi.org/10.1067/mpd.2001.111321

http://dx.doi.org/10.1067/mpd.2001.111321

http://dx.doi.org/10.1097/00005176-200210000-00013

http://dx.doi.org/10.1097/00005176-200210000-00013

http://dx.doi.org/10.1016/j.clnu.2009.09.008

http://dx.doi.org/10.1017/S0950268810001780

http://dx.doi.org/10.1017/S0007114509991395

http://dx.doi.org/10.1017/S0007114509991395

http://dx.doi.org/10.1016/j.jpeds.2011.12.049

http://dx.doi.org/10.3109/07853899009147242

http://dx.doi.org/10.3109/07853899009147242

http://dx.doi.org/10.1086/508178

http://dx.doi.org/10.1086/508178


http://dx.doi.org/10.1093/cid/cis319

http://dx.doi.org/10.1186/2047-2994-1-20

http://dx.doi.org/10.1186/2047-2994-1-20





http://cmr.asm.org

http://cmr.asm.org/

treatment of anaerobic and other infections. Anaerobe 3:73–78. http://dx.doi.org/10.1006/anae.1996.0062.

349. Elmer GW. 2001. Probiotics: “living drugs.” Am. J. Health Syst. Pharm.58:1101–1109.

350. Surawicz CM. 2003. Probiotics, antibiotic-associated diarrhoea andClostridium difficile diarrhoea in humans. Best. Pract. Res. Clin. Gastro-enterol. 17:775–783. http://dx.doi.org/10.1016/S1521-6918(03)00054-4.

351. Venuto C, Butler M, Ashley ED, Brown J. 2010. Alternative therapiesfor Clostridium difficile infections. Pharmacotherapy 30:1266 –1278.http://dx.doi.org/10.1592/phco.30.12.1266.

352. Johnson S, Maziade PJ, McFarland LV, Trick W, Donskey C, Currie B,Low DE, Goldstein EJ. 2012. Is primary prevention of Clostridium dif-ficile infection possible with specific probiotics? Int. J. Infect. Dis. 16:e786 –792. http://dx.doi.org/10.1016/j.ijid.2012.06.005.

353. Miller M. 2009. The fascination with probiotics for Clostridium difficileinfection: lack of evidence for prophylactic or therapeutic efficacy. An-aerobe 15:281–284. http://dx.doi.org/10.1016/j.anaerobe.2009.08.005.

354. McFarland LV. 2006. Meta-analysis of probiotics for the prevention ofantibiotic associated diarrhea and the treatment of Clostridium difficiledisease. Am. J. Gastroenterol. 101:812– 822. http://dx.doi.org/10.1111/j.1572-0241.2006.00465.x.

355. Ritchie ML, Romanuk TN. 2012. A meta-analysis of probiotic efficacyfor gastrointestinal diseases. PLoS One 7:e34938. http://dx.doi.org/10.1371/journal.pone.0034938.

356. Johnston BC, Ma SS, Goldenberg JZ, Thorlund K, Vandvik PO, LoebM, Guyatt GH. 2012. Probiotics for the prevention of Clostridium diffi-cile-associated diarrhea: a systematic review and meta-analysis. Ann. In-tern. Med. 157:878 – 888. http://dx.doi.org/10.7326/0003-4819-157-12-201212180-00563.

357. Johnston BC, Goldenberg JZ, Vandvik PO, Sun X, Guyatt GH. 2011.Probiotics for the prevention of pediatric antibiotic-associated diarrhea.Cochrane Database Syst. Rev. 2011:CD004827. http://dx.doi.org/10.1002/14651858.CD004827.pub2.

358. Gorbach SL, Chang TW, Goldin B. 1987. Successful treatment of re-lapsing Clostridium difficile colitis with Lactobacillus GG. Lancet ii:1519.

359. Biller JA, Katz AJ, Flores AF, Buie TM, Gorbach SL. 1995. Treatmentof recurrent Clostridium difficile colitis with Lactobacillus GG. J. Pediatr.Gastroenterol. Nutr. 21:224 –226. http://dx.doi.org/10.1097/00005176-199508000-00016.

360. Arvola T, Laiho K, Torkkeli S, Mykkanen H, Salminen S, Maunula L,Isolauri E. 1999. Prophylactic Lactobacillus GG reduces antibiotic-associated diarrhea in children with respiratory infections: a randomizedstudy. Pediatrics 104:e64. http://dx.doi.org/10.1542/peds.104.5.e64.

361. Pochapin M. 2000. The effect of probiotics on Clostridium difficile diar-rhea. Am. J. Gastroenterol. 95:S11–13. http://dx.doi.org/10.1016/S0002-9270(99)00809-6.

362. Thomas MR, Litin SC, Osmon DR, Corr AP, Weaver AL, Lohse CM.2001. Lack of effect of Lactobacillus GG on antibiotic-associated diarrhea:a randomized, placebo-controlled trial. Mayo Clin. Proc. 76:883– 889.http://dx.doi.org/10.4065/76.9.883.

363. Hell M, Bernhofer C, Stalzer P, Kern JM, Claassen E. 2013. Probioticsin Clostridium difficile infection: reviewing the need for a multistrainprobiotic. Benef. Microbes 4:39 –51. http://dx.doi.org/10.3920/BM2012.0049.

364. Russell G, Kaplan J, Ferraro M, Michelow IC. 2010. Fecal bacterio-therapy for relapsing Clostridium difficile infection in a child: a proposedtreatment protocol. Pediatrics 126:e239 –242. http://dx.doi.org/10.1542/peds.2009-3363.

365. Kassam Z, Lee CH, Yuan Y, Hunt RH. 2013. Fecal microbiota trans-plantation for Clostridium difficile infection: systematic review and meta-analysis. Am. J. Gastroenterol. 108:500 –508. http://dx.doi.org/10.1038/ajg.2013.59.

366. Sazawal S, Hiremath G, Dhingra U, Malik P, Deb S, Black RE. 2006.Efficacy of probiotics in prevention of acute diarrhoea: a meta-analysis ofmasked, randomised, placebo-controlled trials. Lancet Infect. Dis.6:374 –382. http://dx.doi.org/10.1016/S1473-3099(06)70495-9.

367. Piescik-Lech M, Shamir R, Guarino A, Szajewska H. 2013. The man-agement of acute gastroenteritis in children. Aliment. Pharmacol. Ther.37:289 –303. http://dx.doi.org/10.1111/apt.12163.

368. Van Niel CW, Feudtner C, Garrison MM, Christakis DA. 2002. Lac-tobacillus therapy for acute infectious diarrhea in children: a meta-analysis. Pediatrics 109:678 – 684. http://dx.doi.org/10.1542/peds.109.4.678.

369. Allen SJ, Martinez EG, Gregorio GV, Dans LF. 2010. Probiotics fortreating acute infectious diarrhoea. Cochrane Database Syst. Rev. 2010:CD003048. http://dx.doi.org/10.1002/14651858.CD003048.pub3.

370. Johnston BC, Supina AL, Vohra S. 2006. Probiotics for pediatric anti-biotic-associated diarrhea: a meta-analysis of randomized placebo-controlled trials. CMAJ 175:377–383. http://dx.doi.org/10.1503/cmaj.051603.

371. Bernaola Aponte G, Bada Mancilla CA, Carreazo Pariasca NY, RojasGalarza RA. 2010. Probiotics for treating persistent diarrhoea in chil-dren. Cochrane Database Syst. Rev. 2010:CD007401. http://dx.doi.org/10.1002/14651858.CD007401.pub2.

372. Rimbara E, Fischbach LA, Graham DY. 2011. Optimal therapy forHelicobacter pylori infections. Nat. Rev. Gastroenterol. Hepatol. 8:79 –88. http://dx.doi.org/10.1038/nrgastro.2010.210.

373. Felley CP, Corthesy-Theulaz I, Rivero JL, Sipponen P, Kaufmann M,Bauerfeind P, Wiesel PH, Brassart D, Pfeifer A, Blum AL, Michetti P.2001. Favourable effect of an acidified milk (LC-1) on Helicobacter pylorigastritis in man. Eur. J. Gastroenterol. Hepatol. 13:25–29. http://dx.doi.org/10.1097/00042737-200101000-00005.

374. Pantoflickova D, Corthesy-Theulaz I, Dorta G, Stolte M, Isler P,Rochat F, Enslen M, Blum AL. 2003. Favourable effect of regular intakeof fermented milk containing Lactobacillus johnsonii on Helicobacter py-lori associated gastritis. Aliment. Pharmacol. Ther. 18:805– 813. http://dx.doi.org/10.1046/j.1365-2036.2003.01675.x.

375. Gotteland M, Cruchet S. 2003. Suppressive effect of frequent ingestionof Lactobacillus johnsonii La1 on Helicobacter pylori colonization inasymptomatic volunteers. J. Antimicrob. Chemother. 51:1317–1319.http://dx.doi.org/10.1093/jac/dkg227.

376. Cruchet S, Obregon MC, Salazar G, Diaz E, Gotteland M. 2003. Effectof the ingestion of a dietary product containing Lactobacillus johnsoniiLa1 on Helicobacter pylori colonization in children. Nutrition 19:716 –721. http://dx.doi.org/10.1016/S0899-9007(03)00109-6.

377. Gotteland M, Andrews M, Toledo M, Munoz L, Caceres P, Anziani A,Wittig E, Speisky H, Salazar G. 2008. Modulation of Helicobacter pyloricolonization with cranberry juice and Lactobacillus johnsonii La1 in chil-dren. Nutrition 24:421– 426. http://dx.doi.org/10.1016/j.nut.2008.01.007.

378. Cats A, Kuipers EJ, Bosschaert MA, Pot RG, Vandenbroucke-GraulsCM, Kusters JG. 2003. Effect of frequent consumption of a Lactobacilluscasei-containing milk drink in Helicobacter pylori-colonized subjects. Al-iment. Pharmacol. Ther. 17:429 – 435. http://dx.doi.org/10.1046/j.1365-2036.2003.01452.x.

379. Sahagun-Flores JE, Lopez-Pena LS, de la Cruz-Ramirez Jaimes J,Garcia-Bravo MS, Peregrina-Gomez R, de Alba-Garcia JE. 2007. Erad-ication of Helicobacter pylori: triple treatment scheme plus Lactobacillusvs. triple treatment alone. Cir. Cir. 75:333–336.

380. Sykora J, Valeckova K, Amlerova J, Siala K, Dedek P, Watkins S,Varvarovska J, Stozicky F, Pazdiora P, Schwarz J. 2005. Effects of aspecially designed fermented milk product containing probiotic Lacto-bacillus casei DN-114 001 and the eradication of H. pylori in children: aprospective randomized double-blind study. J. Clin. Gastroenterol. 39:692– 698. http://dx.doi.org/10.1097/01.mcg.0000173855.77191.44.

381. Canducci F, Armuzzi A, Cremonini F, Cammarota G, Bartolozzi F,Pola P, Gasbarrini G, Gasbarrini A. 2000. A lyophilized and inactivatedculture of Lactobacillus acidophilus increases Helicobacter pylori eradica-tion rates. Aliment. Pharmacol. Ther. 14:1625–1629. http://dx.doi.org/10.1046/j.1365-2036.2000.00885.x.

382. Armuzzi A, Cremonini F, Ojetti V, Bartolozzi F, Canducci F, CandelliM, Santarelli L, Cammarota G, De Lorenzo A, Pola P, Gasbarrini G,Gasbarrini A. 2001. Effect of Lactobacillus GG supplementation on an-tibiotic-associated gastrointestinal side effects during Helicobacter pylorieradication therapy: a pilot study. Digestion 63:1–7. http://dx.doi.org/10.1159/000051865.

383. Cremonini F, Di Caro S, Covino M, Armuzzi A, Gabrielli M, SantarelliL, Nista EC, Cammarota G, Gasbarrini G, Gasbarrini A. 2002. Effect ofdifferent probiotic preparations on anti-Helicobacter pylori therapy-related side effects: a parallel group, triple blind, placebo-controlledstudy. Am. J. Gastroenterol. 97:2744 –2749. http://dx.doi.org/10.1111/j.1572-0241.2002.07063.x.

384. Myllyluoma E, Veijola L, Ahlroos T, Tynkkynen S, Kankuri E,Vapaatalo H, Rautelin H, Korpela R. 2005. Probiotic supplementationimproves tolerance to Helicobacter pylori eradication therapy—a place-bo-controlled, double-blind randomized pilot study. Aliment. Pharma-



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1006/anae.1996.0062

http://dx.doi.org/10.1006/anae.1996.0062

http://dx.doi.org/10.1016/S1521-6918(03)00054-4

http://dx.doi.org/10.1592/phco.30.12.1266

http://dx.doi.org/10.1016/j.ijid.2012.06.005


http://dx.doi.org/10.1111/j.1572-0241.2006.00465.x

http://dx.doi.org/10.1111/j.1572-0241.2006.00465.x

http://dx.doi.org/10.1371/journal.pone.0034938


http://dx.doi.org/10.7326/0003-4819-157-12-201212180-00563

http://dx.doi.org/10.7326/0003-4819-157-12-201212180-00563

http://dx.doi.org/10.1002/14651858.CD004827.pub2


http://dx.doi.org/10.1097/00005176-199508000-00016

http://dx.doi.org/10.1097/00005176-199508000-00016

http://dx.doi.org/10.1542/peds.104.5.e64

http://dx.doi.org/10.1016/S0002-9270(99)00809-6

http://dx.doi.org/10.1016/S0002-9270(99)00809-6

http://dx.doi.org/10.4065/76.9.883

http://dx.doi.org/10.3920/BM2012.0049

http://dx.doi.org/10.3920/BM2012.0049





http://dx.doi.org/10.1016/S1473-3099(06)70495-9

http://dx.doi.org/10.1111/apt.12163

http://dx.doi.org/10.1542/peds.109.4.678

http://dx.doi.org/10.1542/peds.109.4.678


http://dx.doi.org/10.1503/cmaj.051603

http://dx.doi.org/10.1503/cmaj.051603




http://dx.doi.org/10.1097/00042737-200101000-00005

http://dx.doi.org/10.1097/00042737-200101000-00005

http://dx.doi.org/10.1046/j.1365-2036.2003.01675.x

http://dx.doi.org/10.1046/j.1365-2036.2003.01675.x

http://dx.doi.org/10.1093/jac/dkg227

http://dx.doi.org/10.1016/S0899-9007(03)00109-6

http://dx.doi.org/10.1016/j.nut.2008.01.007

http://dx.doi.org/10.1016/j.nut.2008.01.007

http://dx.doi.org/10.1046/j.1365-2036.2003.01452.x

http://dx.doi.org/10.1046/j.1365-2036.2003.01452.x

http://dx.doi.org/10.1097/01.mcg.0000173855.77191.44

http://dx.doi.org/10.1046/j.1365-2036.2000.00885.x

http://dx.doi.org/10.1046/j.1365-2036.2000.00885.x

http://dx.doi.org/10.1159/000051865

http://dx.doi.org/10.1159/000051865

http://dx.doi.org/10.1111/j.1572-0241.2002.07063.x

http://dx.doi.org/10.1111/j.1572-0241.2002.07063.x

http://cmr.asm.org

http://cmr.asm.org/

col. Ther. 21:1263–1272. http://dx.doi.org/10.1111/j.1365-2036.2005.02448.x.

385. Szajewska H, Albrecht P, Topczewska-Cabanek A. 2009. Randomized,double-blind, placebo-controlled trial: effect of Lactobacillus GG supple-mentation on Helicobacter pylori eradication rates and side effects duringtreatment in children. J. Pediatr. Gastroenterol. Nutr. 48:431– 436. http://dx.doi.org/10.1097/MPG.0b013e318182e716.

386. Francavilla R, Lionetti E, Castellaneta SP, Magista AM, Maurogio-vanni G, Bucci N, De Canio A, Indrio F, Cavallo L, Ierardi E, MinielloVL. 2008. Inhibition of Helicobacter pylori infection in humans by Lac-tobacillus reuteri ATCC 55730 and effect on eradication therapy: a pilotstudy. Helicobacter 13:127–134. http://dx.doi.org/10.1111/j.1523-5378.2008.00593.x.

387. De Francesco V, Stoppino V, Sgarro C, Faleo D. 2000. Lactobacillusacidophilus administration added to omeprazole/amoxycillin-baseddouble therapy in Helicobacter pylori eradication. Dig. Liver Dis. 32:746 –747. http://dx.doi.org/10.1016/S1590-8658(00)80343-6.

388. Gotteland M, Poliak L, Cruchet S, Brunser O. 2005. Effect of regularingestion of Saccharomyces boulardii plus inulin or Lactobacillus acidoph-ilus LB in children colonized by Helicobacter pylori. Acta Paediatr. 94:1747–1751. http://dx.doi.org/10.1111/j.1651-2227.2005.tb01848.x.

389. Teusink B, Smid EJ. 2006. Modelling strategies for the industrial exploi-tation of lactic acid bacteria. Nat. Rev. Microbiol. 4:46 –56. http://dx.doi.org/10.1038/nrmicro1319.

390. Spurbeck RR, Arvidson CG. 2011. Lactobacilli at the front line of de-fense against vaginally acquired infections. Future Microbiol. 6:567–582.http://dx.doi.org/10.2217/fmb.11.36.

391. Shanahan F, Stanton C, Ross RP, Hill C. 2009. Pharmabiotics: bioac-tives from mining host-microbe-dietary interactions. Funct. Food Rev.1:20 –25.

392. Beceiro A, Tomas M, Bou G. 2013. Antimicrobial resistance and viru-lence: a successful or deleterious association in the bacterial world? Clin.Microbiol. Rev. 26:185–230. http://dx.doi.org/10.1128/CMR.00059-12.

393. Orman MA, Brynildsen MP. 2013. Dormancy is not necessary or suffi-cient for bacterial persistence. Antimicrob. Agents Chemother. 57:3230 –3239. http://dx.doi.org/10.1128/AAC.00243-13.

394. Seo MD, Won HS, Kim JH, Mishig-Ochir T, Lee BJ. 2012. Antimicro-bial peptides for therapeutic applications: a review. Molecules 17:12276 –12286. http://dx.doi.org/10.3390/molecules171012276.

395. Gallo RL, Hooper LV. 2012. Epithelial antimicrobial defence of the skinand intestine. Nat. Rev. Immunol. 12:503–516. http://dx.doi.org/10.1038/nri3228.

396. Brogden KA. 2005. Antimicrobial peptides: pore formers or metabolicinhibitors in bacteria? Nat. Rev. Microbiol. 3:238 –250. http://dx.doi.org/10.1038/nrmicro1098.

397. Bush K, Courvalin P, Dantas G, Davies J, Eisenstein B, Huovinen P,Jacoby GA, Kishony R, Kreiswirth BN, Kutter E, Lerner SA, Levy S,Lewis K, Lomovskaya O, Miller JH, Mobashery S, Piddock LJ, ProjanS, Thomas CM, Tomasz A, Tulkens PM, Walsh TR, Watson JD,Witkowski J, Witte W, Wright G, Yeh P, Zgurskaya HI. 2011. Tacklingantibiotic resistance. Nat. Rev. Microbiol. 9:894 – 896. http://dx.doi.org/10.1038/nrmicro2693.

398. Klaenhammer TR, Barrangou R, Buck BL, Azcarate-Peril MA, Alter-mann E. 2005. Genomic features of lactic acid bacteria effecting biopro-cessing and health. FEMS Microbiol. Rev. 29:393– 409. http://dx.doi.org/10.1016/j.fmrre.2005.04.007.

399. Ventura M, O’Flaherty S, Claesson MJ, Turroni F, Klaenhammer TR,van Sinderen D, O’Toole PW. 2009. Genome-scale analyses of health-promoting bacteria: probiogenomics. Nat. Rev. Microbiol. 7:61–71. http://dx.doi.org/10.1038/nrmicro2047.

400. Klaenhammer TR, Altermann E, Pfeiler E, Buck BL, Goh YJ,O’Flaherty S, Barrangou R, Duong T. 2008. Functional genomics of

probiotic lactobacilli. J. Clin. Gastroenterol. 42(Suppl 3):S160 –S162.http://dx.doi.org/10.1097/MCG.0b013e31817da140.

401. Lebeer S, Vanderleyden J, De Keersmaecker SC. 2008. Genes andmolecules of lactobacilli supporting probiotic action. Microbiol. Mol.Biol. Rev. 72:728 –764. http://dx.doi.org/10.1128/MMBR.00017-08.

402. Sybesma W, Molenaar D, van Ijcken W, Venema K, Kort R. 2013.Genome instability in Lactobacillus rhamnosus GG. Appl. Environ. Mi-crobiol. 79:2233–2239. http://dx.doi.org/10.1128/AEM.03566-12.

403. Douillard FP, Ribbera A, Jarvinen HM, Kant R, Pietila TE, RandazzoC, Paulin L, Laine PK, Caggia C, von Ossowski I, Reunanen J, SatokariR, Salminen S, Palva A, de Vos WM. 2013. Comparative genomic andfunctional analysis of Lactobacillus casei and Lactobacillus rhamnosusstrains marketed as probiotics. Appl. Environ. Microbiol. 79:1923–1933.http://dx.doi.org/10.1128/AEM.03467-12.

404. Pridmore RD, Berger B, Desiere F, Vilanova D, Barretto C, Pittet AC,Zwahlen MC, Rouvet M, Altermann E, Barrangou R, Mollet B, Mer-cenier A, Klaenhammer T, Arigoni F, Schell MA. 2004. The genomesequence of the probiotic intestinal bacterium Lactobacillus johnsoniiNCC 533. Proc. Natl. Acad. Sci. U. S. A. 101:2512–2517. http://dx.doi.org/10.1073/pnas.0307327101.

405. Boekhorst J, Siezen RJ, Zwahlen MC, Vilanova D, Pridmore RD,Mercenier A, Kleerebezem M, de Vos WM, Brussow H, Desiere F.2004. The complete genomes of Lactobacillus plantarum and Lactobacil-lus johnsonii reveal extensive differences in chromosome organizationand gene content. Microbiology 150:3601–3611. http://dx.doi.org/10.1099/mic.0.27392-0.

406. Denou E, Pridmore RD, Berger B, Panoff JM, Arigoni F, Brussow H.2008. Identification of genes associated with the long-gut-persistencephenotype of the probiotic Lactobacillus johnsonii strain NCC533 using acombination of genomics and transcriptome analysis. J. Bacteriol. 190:3161–3168. http://dx.doi.org/10.1128/JB.01637-07.

407. Denou E, Berger B, Barretto C, Panoff JM, Arigoni F, Brussow H.2007. Gene expression of commensal Lactobacillus johnsonii strainNCC533 during in vitro growth and in the murine gut. J. Bacteriol. 189:8109 – 8119. http://dx.doi.org/10.1128/JB.00991-07.

408. Yasuda E, Serata M, Sako T. 2008. Suppressive effect on activation ofmacrophages by Lactobacillus casei strain Shirota genes determining thesynthesis of cell wall-associated polysaccharides. Appl. Environ. Micro-biol. 74:4746 – 4755. http://dx.doi.org/10.1128/AEM.00412-08.

409. Lebeer S, Verhoeven TL, Perea Velez M, Vanderleyden J, De Keers-maecker SC. 2007. Impact of environmental and genetic factors on bio-film formation by the probiotic strain Lactobacillus rhamnosus GG. Appl.Environ. Microbiol. 73:6768 – 6775. http://dx.doi.org/10.1128/AEM.01393-07.

410. Lebeer S, Verhoeven TL, Francius G, Schoofs G, Lambrichts I, Du-frene Y, Vanderleyden J, De Keersmaecker SC. 2009. Identification ofa gene cluster for the biosynthesis of a long, galactose-rich exopolysac-charide in Lactobacillus rhamnosus GG and functional analysis of thepriming glycosyltransferase. Appl. Environ. Microbiol. 75:3554 –3563.http://dx.doi.org/10.1128/AEM.02919-08.

411. Sonnenburg JL, Chen CT, Gordon JI. 2006. Genomic and metabolicstudies of the impact of probiotics on a model gut symbiont and host.PLoS Biol. 4:e413. http://dx.doi.org/10.1371/journal.pbio.0040413.

412. Ganzle MG, Holtzel A, Walter J, Jung G, Hammes WP. 2000. Char-acterization of reutericyclin produced by Lactobacillus reuteri LTH2584.Appl. Environ. Microbiol. 66:4325– 4333. http://dx.doi.org/10.1128/AEM.66.10.4325-4333.2000.

413. Riboulet-Bisson E, Sturme MH, Jeffery IB, O’Donnell MM, NevilleBA, Forde BM, Claesson MJ, Harris H, Gardiner GE, Casey PG,Lawlor PG, O’Toole PW, Ross RP. 2012. Effect of Lactobacillus sali-varius bacteriocin Abp118 on the mouse and pig intestinal microbiota.PLoS One 7:e31113. http://dx.doi.org/10.1371/journal.pone.0031113.



on Novem


r.asm.org/

Dow

nloaded from

http://dx.doi.org/10.1111/j.1365-2036.2005.02448.x

http://dx.doi.org/10.1111/j.1365-2036.2005.02448.x

http://dx.doi.org/10.1097/MPG.0b013e318182e716

http://dx.doi.org/10.1097/MPG.0b013e318182e716

http://dx.doi.org/10.1111/j.1523-5378.2008.00593.x

http://dx.doi.org/10.1111/j.1523-5378.2008.00593.x

http://dx.doi.org/10.1016/S1590-8658(00)80343-6




http://dx.doi.org/10.2217/fmb.11.36



http://dx.doi.org/10.3390/molecules171012276







http://dx.doi.org/10.1016/j.fmrre.2005.04.007

http://dx.doi.org/10.1016/j.fmrre.2005.04.007



http://dx.doi.org/10.1097/MCG.0b013e31817da140








http://dx.doi.org/10.1128/JB.01637-07

http://dx.doi.org/10.1128/JB.00991-07





http://dx.doi.org/10.1371/journal.pbio.0040413




http://cmr.asm.org

http://cmr.asm.org/

Vanessa Liévin-Le Moal is a microbiologistwho trained at the Faculty of Pharmacy, Uni-versity Paris-Sud at Châtenay-Malabry, France,and received her master’s degree in 2001 andher Ph.D. in microbiology in 2005. She studiedthe role of intestinal host defense mechanismsagainst enterovirulent bacterial infection inINSERM Units 510 and 756. She is currently anassociate professor at the Faculty of Pharmacyat Châtenay-Malabry, working on antiparasitictherapeutics at CNRS Unit 8076 BioCis.

Alain L. Servin received his Doctorat d’Universitéfrom University Orsay Paris-Sud, France, in 1973and his Doctorat d’Etat es Sciences in MolecularPharmacology from University Paris VI in 1987.In 1980 he joined the French National Institute ofHealth and Medical Research (INSERM), work-ing on biophysics and cellular pharmacology atINSERM Institute Saint Antoine Hospital, Paris,and INSERM Unit 178 at Paul Brousse Hospital atVillejuif. He was research director at INSERM andhead of INSERM units between 1990 and 2010 atthe Faculty of Pharmacy Châtenay-Malabry, University Paris-Sud. He began hisinterest in cellular microbiology in 1990. He and his colleagues focus their re-search on the molecular and cellular mechanisms of Afa/Dr DAEC and rotaviruspathogenesis. He also studies the role of intestinal and vaginal microbiota incontrolling enteric and vaginal infections and holds patents in this area. He iscurrently an associate-researcher at CNRS Unit 8076 BioCis, Faculty ofPharmacy Châtenay-Malabry. Dr. Servin’s homepage can be found athttp://cvscience.aviesan.fr/cv/827/alain-l-servin.



on Novem


r.asm.org/

Dow

nloaded from

http://cvscience.aviesan.fr/cv/827/alain-l-servin

http://cmr.asm.org

http://cmr.asm.org/

anti-infective activities of lactobacillus strains in the ... · minally and intracellularly in the...

Documents