Hospital acquired infections : Molecular study and infection control guidlinesSupervisorsProf.Dr.Lotfy Abdel-Naby MahmoudProf. of Clinical PathologyFaculty of Medicine-Mansoura University

Dr.Wafaa Mohamed Mohamed ElemshatyAssociate Professor of Clinical PathologyFaculty of Medicine -Mansoura University

Essay By Abd Alla Ibrahim Ahmed Shady Resident in Clinical PathologyDepartmentFaculty of Medicine-Mansoura University

Acknowledgement

Thanks, first and last, now and forever to "ALLAH", for great care,

guidance, and help in every step of my life and for a l l the countless

gifts I have been offered. Of these gifts, those people who were assigned

to give me a precious hand to be able to fulfill this essay.

I would like to express my sincere gratitude, deepest thanks and

appreciation to Prof .Dr. Lotfy Abdel-Naby Mahmoud

, Professor of Clinical Pathology for his very kind supervision,

stimulating suggestions, continous encouragement, valuable advice and

endless effort in all the time of the research and writing of this essay.

I am deeply indebted to Dr . Wafaa Mohamed Mohamed

Elemshaty, Associate Professor of Clinical Pathology, Mansoura

University, for her kind cooperation, valuable aid, and advise, I am

deeply touched by their endless support and concern.

Abd Alla Ibrahim Ahmed Shady

2012

1Introduction and aim of work*

4

4

Review

Hospital acquired infections

Definitions

*

5Frequency of HAIs 8 Impact of HAIs

9 Routes of transmission

13 Predisposing factor for HAIs

1818

19

22

2425

27

2929

30

31

32

Common HAIs 1-Urinary Tract Infection

2- Respiratory tract infections

3-Surgical site infections

4- Burn infections

5- Blood stream infections

6-Device associated infection

7-Ocular Infections

8-Central Nervous System Infections

9-Ear & Nose infection

10-Gastrointestinal Tract Infections

11 -Obstetrics and neonatal Infections

35 Diagnosis of HAIs

41

4141

43

44

4648

49

52

5556

56

57

6264

65

67

68

A - Phenotypic methods

1-Biotyping

2-Antimicrobial susceptibility testing

3-Serotyping

4-Bacteriophage and bacteriocin typing

B-Genotypic methods

1- Plasmid Analysis

2- Pulsed-field gel electrophoresis

3-Southern Blot Analysis-Ribotyping

4-Northern blotting

5-Heteroduplex Migration Analysis

6-Single Strand Conformation Polymorphism

Analysis

7-Typing Methods Using PCR

8 -DNA arrays

9 -Pyrosequencing

10 –Spectroscopy

11-Proteomics and metabolomics

12- Nucleotide sequence- based analysis

70

74

Infection control Guidelines

1-Hand hygiene

80 2 -Environmental Decontamination &Cleaning

85 3-Nutrition 87 4-Surveillance & Outbreak control92 5-Preventing urinary tract infection and

urinary catheter infections

94 6-Preventing central venous line infection

and Blood stream infections 987-Preventing respiratory tract infections 1008-Control of infections related to surgery

and surgical equipment 104 9- Prevention of Gastrointestinal Tract

and waterborne hospital infections 106 10-Preventing burn infections 10911-Isolation and precautions 112Summary & Conclusion *118References*

1Arabic Summary *

Amplified fragment length polymorphismAFLP

Acquired immunodeficiency syndromeAIDS

Arbitrarily primed PCRAP-PCR

Catheter Related Blood stream infectionsCABSIs

Catheter-associated urinary tract infectionsCAUTIs

Clostridium difficile-associated diseaseCDAD

Centers for Disease Control and PreventionCDC

Contour-clamped homogenous electric fieldCHEF

Coagulase-negative staphylococciCo NS

Catheter Related Blood stream infectionsCRBSI

Central venous catheterCVC

Deoxynucleoside triphosphatesDNTPs

Group A β-hemolytic streptococci(Streptococcus pyogenes)

GABHS

Hospital acquired infectionHAI

Hospital Acquired PneumoniaHAP

Hepatitis B virusHBV

Health care associated infectionHCAI

Hepatitis C virusHCV

Healthcare workersHCWs

High Efficiency Particulate AirHEPA

High-intensity narrow-spectrum light environmental decontamination system

HINS-light EDS

Human immunodeficiency virusHIV

Heteroduplex Migration AnalysisHMA

Intensive care unitICU

Multidrug-resistant organismMDRO

Multidrug-resistant tuberculosisMDR-TB

Multi dose vialsMDV

Multi-locus sequence typingMLST

Multi Locus Variable copy Numbers of Tandem Repeats Analysis

MLVA

Methicillin-resistant Staphylococcus aureusMRSA

Mass spectrometryMS

Nosocomial infectionNI

Neonatal intensive care unitNICU

Needle stick and sharps injuryNSI

Polymerase chain reactionPCR

Pulsed-field gel electrophoresisPFGE

Postoperative endophthalmitisPOE

Post tympanostomy tube otorrheaPTTO

Repetitive element PCRRep-PCR

Restriction fragment length polymorphismsRFLP

Respiratory Syncetial VirusRSV

Severe acute respiratory syndromeSARS

Single-locus sequence typingSLST

Single Strand Conformation Polymorphism AnalysisSSCP

Surgical site infectionsSSIs

Urinary tract infectionsUTIs

Ventilator-associated pneumoniaVAP

Vancomycin-resistant enterococciVRE

World health organizationWHO

Figure 1 Studies on general HAIs rates from developing and developed countries 1995-2008

7

Figure 2 Recent evolution of bacterial strain identification for epidemiological purpose

40

Figure 3 Disk diffusion testing of a hyper b-lactamase producing Staphylococcus aureus.

43

Figure 4 Phage typing is the identification of bacterial species and strains by determining their susceptibility to various phages

45

Figure 5 Flow chart comparison of the different procedural steps used for various molecular typing techniques

47

Figure 6 Schematic diagram of Plasmid Analysis 49

Figure 7 Diagram of pulsed-field gel electrophoresis 51

Figure 8 Schematic diagram of Southern Blot Analysis-Ribotyping

54

Figure 9 Schematic diagram of Northern blotting 55

Figure 10 Schematic diagram of the amplified fragment length polymorphism (AFLP) technique

60

Figure 11 Schematic diagram of Restriction fragment length polymorphisms

61

Figure 12 Schematic diagram of DNA arrays 64

Figure 13 Overview of Raman procedure and spectrometer

66

Figure 14 Schematic diagram of hand washing 79

Table 1 Infection control program structure 71Table 2 Infection control program element 72Table 3 Non- pharmacological hospital infection

control strategies73

Table 4 Immediate control measures for outbreak management

91

Table 5 Colour coding for the disposal of clinical waste

97

Table 6 Major headings and some factors in prevention of surgical site infection

100

Introduction

Hospital-acquired infections (HAIs) or nosocomial infections (NI)

are a major challenge to patient safety and contribute significantly to

morbidity and mortality, as well as to excess costs for hospital stay

(French and Cheng , 1991) .They affect both developed and resource-

poor countries and constitute a significant burden both for the patient and

for the health care system (WHO, 2002) .

A prevalence survey was conducted, under the authority of the World

Heath Organization (WHO), in 55 hospitals from 14 countries

representing 4 regions (Europe, Eastern Mediterranean, Southeast Asia,

and Western Pacific).This survey revealed that an average of 8.7% of

hospitalized patients developed HAIs, with the highest frequencies of

such infections occurring among hospitals in the Eastern Mediterranean

and South east Asian regions “11.8%and10% respectively”(Talaat , et

al., 2006) .

Several risk factors for acquiring an infection have been commonly

cited, including the presence of underlying conditions (such as diabetes,

renal failure, or malignancies), long term hospitalizations, surgical

procedures, receipt of prior antimicrobial therapy, and the presence of

indwelling catheters (Singh et al., 2006) .

Understanding pathogen distribution and relatedness is essential for

determining the epidemiology of NI and aiding in the design of rational

pathogen control methods. The role of pathogen typing is to determine if

epidemiologically related isolates are also genetically related .

Historically, this analysis of nosocomial pathogens has relied on a

comparison of phenotypic characteristics such as biotypes ,

serotypes ,bacteriophage or bacteriocin types, and antimicrobial

susceptibility profiles. This approach has begun to change over the past 2

decades, with the development and implementation of new technologies

based on DNA, or molecular, analysis (Arbeit , 1999 & Cockerill ,et

al.,2004 ) .

Most studies related to HAI were conducted in the developed countries

and demonstrated the efficacy of HAI surveillance and its significant

incidence concerning patient morbidity and mortality (Cooke , 2000 &

Barrett , 2002 & Gastmeier ,2006) .Conversely, in the developing

countries, few studies provide data of device associated infection rates

using the standardized definitions of HAI rates per 1000 days (Rosenthal

, 2006 & Leblebicioglu , 2007) .

More over, much of the recent research on NI has dealt with the

need for new antibiotics ,better antibiotic management and better

diagnostic techniques to detect infections earlier .Better drug treatment

and earlier infection diagnosis can certainly play a major role in reducing

morbidity and mortality from HAIs . However, there are many non

pharmacological interventions that can significantly reduce the incidence

of HAIs, but these are often overlooked in practice (Curtis, 2008) .

Aim of work

The aim of this essay is to review morbidity, mortality , infections

routes and medical cost associated with hospital acquired infections ,

with stress on molecular studies of these infections along with infection

control guidelines.

Hospital acquired infections

Definitions

Nosocomial infection (NI) or hospital acquired infection (HAI) can

be defined as an infection acquired in hospital by a patient who was

admitted for a reason other than that infection . This includes infections

acquired in the hospital but appearing after discharge, and also

occupational infections among staff of the facility (WHO, 2002) .

Endemic is the usual level or presence of an agent or disease in a

defined population during a given period. Epidemic is an unusual,

higher-than-expected level of infection or disease by an agent in a defined

population in a given period. This definition assumes previous knowledge

of the usual, or endemic, levels. Pandemic is an epidemic that spreads

over several countries or continents and affects many people

(Gordis ,2000).

An outbreak is defined as an unusual or unexpected increase of cases

of a known nosocomial infection or the emergence of cases of a new

infection. Outbreaks of nosocomial infection should be identified and

promptly investigated because of their importance in terms of morbidity,

costs and institutional image. Outbreak investigation may also lead to

sustained improvement in patient care practices (Gordis , 1996) .

Incidence rate is defined as the ratio of the number of new infections

or disease in a defined population in a given period to the number of

individuals at risk in the population. At risk is frequently defined as the

number of potentially exposed susceptibles. The rate is usually expressed

as numbers of new cases per thousands (1,000, 10,000, or 100,000) per

year . Prevalence rate is defined as the ratio of the number of individuals

measurably affected or diseased by an agent in a defined population at a

particular point in time, or over a specified time period, without regard to

when the process or disease began (Gordis ,2000).

Frequency of HAIs

Among the more industrialized and developed nations, the World

Health Organization found 8.7% of all hospitalized patients to have

nosocomial infections. While HAI are an important health care concern

worldwide.They are especially troublesome in developing nations.

Nosocomial infection rates range from 1% in Northern Europe, especially

the Netherlands, which introduced extremely aggressive infection control

measures, to 40% in some parts of Asia, South America, and sub-Saharan

Africa (Starakis et al .,2002& Eriksen et al .,2005& Klevens et

al .,2007).

Compared with average prevalence of health-care-associated infection

in Europe (reported as 7.1 per 100 patients by the European Centre for

Disease Prevention and Control) and estimated incidence in the USA (4.5

per 100 patients ), prevalence of health-care-associated infection in

resource-limited settings is substantially higher, particularly in high-

quality studies (15.5 per 100 patients). The difference between

developing and developed countries is even more striking when

considering incidence of ICU-acquired infection (pooled density 47.9 per

1000 patient-days in developing countries), which is estimated to be 13.6

per 1000 patient-days in the USA (Klevens et al ., 2007).

Importantly, very high rates of health-care-associated infection in

neonatal and paediatric populations were noted not only in ICUs but also

in some paediatric wards and children’s hospitals. (Cavalcante et al .,

2006) . Previous findings indicate that surgical-site infection is both the

most frequently studied and the leading health-care-associated infection

hospital-wide in the developing world (Haynes et al ., 2009). In a nation

wide study undertaken in the USA, the cumulative incidence of surgical-

site infection was 2.6 per 100 surgical procedures; similarly, it was 2.9

per 100 surgical procedures in different European countries, and 1.6 per

100 procedures in Germany . ( Bhutta et al ., 2005).

Several studies have observed a high frequency of bloodstream

infections and high mortality among patients admitted to pediatric or

neonatal care units in Egypt. In a study of patients admitted to a pediatric

intensive care unit (ICU), investigators reported a high frequency of

sepsis and an overall mortality rate of 50%. Similarly, a study of 115

infants admitted to a neonatal intensive care unit (NICU) in a major

university hospital over 3-month period found that 77% had sepsis, with

hospital-acquired pathogens isolated from bloodstream, and mortality

rates exceeding 51%. The results of this investigation prompted a cross-

sectional survey of 22 NICUs, from various Ministry of Health hospitals

throughout Egypt, to assess the magnitude of this problem. Among the

180 infants admitted to these 22 units, 97 (54%) had a clinical diagnosis

of sepsis, and 86 (69%) had positive blood cultures at the time of the

survey (El-Nawawy , 2003). There are paucity of national reports accords

with findings of a survey done by WHO, in which only 23 of 147

developing countries (16%) reported a functioning national surveillance

system (figure 1) (WHO , 2008).

Figure 1 . Studies on general HAIs rates from developing and

developed countries 1995-2008 (WHO 2008).

Impact of HAIs .

Nosocomial infections (NI) contribute significantly to morbidity and

mortality, as well as to excess costs for hospitalized patients. According

to the available evidence, the impact of Health care associated infection

(HCAI) implies prolonged hospital stay, long-term disability, increased

resistance of microorganisms to antimicrobials, massive additional

financial burden for health systems, high costs for patients and their

family, and unnecessary deaths . The increased length of stay for infected

patients is the greatest contributor to cost (Burke , 2004 &Klevens et

al .,2007& Edwards et al .,2007) .

The overall increase in the duration of hospitalization for patients

with surgical wound infections was 8.2 days, ranging from 3 days for

gynaecology to 9.9 for general surgery and 19.8 for orthopaedic surgery.

Prolonged stay not only increases direct costs to patients or payers but

also indirect costs due to lost work (Coello et al ., 1993). The increased

use of drugs, the need for isolation, and the use of additional laboratory

and other diagnostic studies also contribute to costs (Wenzel ,1995).

NI is the fourth commonest cause of in-hospital deaths, after

cardiovascular disease, cancers and community-acquired infections.

Lower respiratory tract and bloodstream infections were the main sites of

NI which contributed to death, surgical-site infections were the third most

common factor contributing to death. Conversely, urinary tract infections

rarely contribute to death, although some of them could be considered to

play a role in fatal outcomes, particularly when they were associated with

a bloodstream infection . Most cases of peritonitis (included in

gastrointestinal tract infections) were not healthcare-associated or of

iatrogenic origin, but were associated with underlying diseases, such as

cancer or immunocompromised status (Astagneau et al .,2001) .

Routes of transmission

For infection to take place, microorganisms must be transferred from

a reservoir to an acceptable entry site on a susceptible host in sufficient

numbers (the infecting dose) for multiplication of the agent to take place.

The infecting dose of a microorganism may depend in varying degrees on

the infectivity, pathogenicity, and virulence of the microorganism itself.

The entire transmission process constitutes the chain of infection. Within

the healthcare setting, the reservoir of an agent may include patients

themselves, healthcare workers, tap water, soap dispensers, mechanical

ventilators, intravenous devices and infusates, multidose vials, and other

factors in the environment (Johnson and Gerding ,1998). .

1-Direct transmission

Direct transmission from another host (healthy or ill) or from an

environmental reservoir or surface by direct contact or direct large-

droplet spread of infectious secretions is the simplest route of agent

spread. Examples of direct-contact transmission routes include kissing

(infectious mononucleosis), shaking hands [common cold (rhinovirus)],

or other skin contact (e.g., contamination of a wound with Staphylococci

or Enterococcus spp. during trauma, surgical procedures or dressing

changes) (Johnson and Gerding ,1998).

Droplet of saliva are expelled from respiratory tract by coughing .

sneezing and talking . These droplets may contain a small number of

pathogenic organism from respiratory tract . The large droplets (more

than 0.1 mm in diameter ) fall to the ground within a few seconds and are

not inhaled. Small droplet (less than 0.1 mm in diameter ) evaporate

rapidly and these very small particle , called droplet nuclei , can remain

airbrone for hours and be inhaled in the same way as small dust particles

and carried deep into alveoli of the lungs ( Goldmann et al , 1996).

Transmission of Neisseria meningitidis, group A streptococcus, or

the respiratory syncytial virus (an important cause of respiratory infection

in young children worldwide) by large respiratory droplets that travel

only a few feet is regarded as a special case of direct-contact

transmission. Vertical transmission of infection from mother to fetus is

another form of direct-contact transmission that may occur through the

placenta during pregnancy (e.g., HIV, rubella virus, hepatitis B virus, or

parvovirus) ,by direct contact of the infant with the birth canal during

childbirth (group B streptococci) or via breast milk (HIV) (Lennox et

al ,2004).

Numerous studies document the pivotal role of healthcare workers’

(HCWs) hands in the propagation of micro-organisms within the

healthcare environment and ultimately to patients (WHO,2006). Patients’

skin can be colonized by transient pathogens that are subsequently shed

onto surfaces in the immediate patient surroundings, thus leading to

environmental contamination.As a consequence, HCWs contaminate their

hands by touching the environment or patients’ skin during routine care

activities, sometimes even despite glove use (Pittet et al .,2006) .

Failure to perform appropriate hand hygiene is a leading cause of

health care associated infections and the spread of multidrug-resistant

organisms and contributes to outbreaks (CDC,2002). It has been shown

that organisms are capable of surviving on HCWs’ hands for at least

several minutes following contamination. (Karadeniz et al .,2001 ) .

2- Indirect transmission

Potentially pathogenic micro-organisms can colonize environmental

surfaces in the hospital environment and so act as a source for outbreaks

of nosocomial infection. Studies have presented evidence that the

majority of Gram-positive bacteria, including Staphylococcus aureus and

Enterococcus spp., are able to survive for months on dry surfaces. Gram-

negative bacteria, such as Klebsiella spp., Escherichia coli, and

Acinetobacter spp. can also survive for a relatively long time on

inanimate surfaces, while common fungi such as Candida spp. have

similar properties. Environmental conditions such as low temperature or

humidity appear to be crucial for the persistence of these organisms on

inanimate surfaces (Kramer et al ., 2006) .

The airborne transfer of droplet nuclei is the principal route of

transmission of Mycobacterium tuberculosis, varicella (chicken pox), and

measles ( Sepkowitz ,1996), while the transmission of Legionella spp.

through the air in droplet nuclei from cooling tower emissions, and from

environmental water sites, such as air-conditioning systems, central

humidifiers, and respiratory humidification devices, is another recent

important example of this type of spread ( CDC,1997).

Intrinsic contamination of commercially manufactured products, e.g.,

intravenous fluids, may also occur, e.g., as in 1970/1971 when 150 cases

of hospital-acquired bacteraemia were noticed during a large outbreak in

eight hospitals in the USA. Contamination of blood products and

heparine sodium chloride solutions may also occur . The most frequent

pathogens were Yersinia enterocolitica, and Serratia spp. for blood

products and Burkholderia cepacia and Enterobacter spp. for substances

other than blood products. (Vonberg and Gastmeier.,2006) . Another

frequent route of contamination is by the use of multi dose vials (MDV)

that had been manufactured for single use only (O’Grady et al .,2002 ).

Nosocomial Infections are frequently caused by environmental

organisms and have been linked to a wide variety of contaminated

hospital equipment, suggesting that the risk of NI following contact with

equipment is high . Equipment used in the non-critical setting is less

likely to have standard cleaning protocols than equipment used in the

critical setting, making it more likely to carry large numbers of micro-

organisms. The majority of bacterial species isolated were those found in

normal skin and environmental flora; coagulase-negative staphylococci

were isolated most frequently ( Karadeniz et al .,2001).

The high levels of contamination ,on stethoscope membranes -

diagnostic ultrasound - stethoscope ear tips – stethoscopes - otoscopes

and auriscopes . A higher proportion of death was found in patients who

received insertions of arterial lines, chest tubes, central venous catheters,

urinary catheters, Port-A devices, and Swan-Ganz catheters. Likewise,

increased risk of mortality was evident in patients receiving hemodialysis,

endoscopic examination, total parenteral nutrition, and

immunosuppressive steroid or chemotherapy ( Sheng et al . ,2007) .

Vector-borne transmission by arthropods or other insects is the final

type of indirect transmission, and may be mechanical or biologic. In

mechanical vector-borne transmission, the agent does not multiply or

undergo physiologic changes in the vector; in biologic vector-borne

transmission, the agent is modified within the host before being

transmitted. In tropical countries with endemic vector-borne disease,

including dengue, yellow fever, and malaria, this type of transmission is

more important, requiring screening and other controls not required of

medical structures in colder climates (Daniel et al .,1992) .

Predisposing factors for HAIs

The highest prevalence of HAI occurred in ICUs and acute care

surgical and orthopedic settings. Old age, multiple morbidities or disease

severity, and decreased immunity increase patient susceptibility. Poor

infection control measures , invasive procedures including central venous

or urinary catheter placements ,and antimicrobial misuse are another risk

factors (Wenzel , 2007 & Klevens et al .,2007) .

1-Microbial factor

The agents causing healthcare-associated infectious diseases are

microorganisms ranging in size and complexity from viruses and bacteria

to protozoa and helminths. Bacteria, fungi, and certain viruses have been

the agents most recognized and studied as causes of healthcare-associated

infections (Emori and Gaynes ,1993).

For transmission to take place, the microorganism must remain

viable in the environment until contact with the host has been sufficient to

allow infection. Reservoirs that allow the agent to survive or multiply

may be animate, as a healthcare worker carriage of staphylococci in the

anterior nares, or inanimate in the environment, as demonstrated by

Pseudomonas species or Legionella in air-conditioning humidification

systems (Alary ,1992), Clostridium difficile spores on patient surfaces, or

Serratia marcescens growing in contaminated soap or hand lotion

preparations (Archibald et al .,1997).

Certain intrinsic and genetically determined properties of a

microorganism are important for it to survive in the environment. These

include the ability to resist the effects of heat, drying, ultraviolet light,

and chemical agents, including antimicrobials; the ability to compete with

other microorganisms; and the ability to independently multiply in the

environment or to develop and multiply within another (vector) host

Cohen et al .,1991) .

Intrinsic factors important to the production of disease include

infectivity, pathogenicity, virulence, the infecting dose, the agent's ability

to produce toxins, its immunogenicity and ability to resist or overcome

the human immune defense system, its ability to replicate only in certain

types of cells, tissues, or hosts (vectors), its ability to persist or cause

chronic infection, and its interaction with other host mechanisms,

including the ability to cause immunosuppression (e.g., HIV) (Cohen et

al .,1991) .

Many patients receive antimicrobial drugs. Through selection and

exchange of genetic resistance elements, antibiotics promote the

emergence of multi drug resistant strains of bacteria; microorganisms in

the normal human flora sensitive to the given drug are suppressed, while

resistant strains persist and may become endemic in the hospital

(Ducel ,1995) .

2-Host factor

Normally, the patient has three principal defenses against infection:

physical defenses, nonspecific immune response and a specific immune

response. Changes in these defenses determine the patient’s susceptibility

to infection (Mele et al .,1998). The host immune defenses attempt to

prevent infection. Thus, any reduction in host defenses may allow

infection to take place with a smaller dose of microorganisms and/or at a

body site that is not usually susceptible to infection . Host factors

important to the development and severity of infection or disease may be

categorized as intrinsic or extrinsic (Scrimshaw ,1989) .

Intrinsic factors ; include the age at infection; birth weight; sex; race;

nutritional status ; comorbid conditions (including anatomic anomalies)

and diseases; genetically determined immune status; immunosuppression

associated with other infections, diseases, or therapy; vaccination or

immunization status; previous experience with this or similar agents; and

the psychologic state of the host ( Cohen et al .,1991 ) .

Comorbid condition also include malignancies, recent chemotherapy

or radiotherapy, chronic renal diseases, chronic heart diseases, chronic

liver diseases, and chronic lung diseases had higher proportion of deaths.

In contrast, patients with neurologic diseases had a lower risk for death

( Sheng et al .,2007) .

Very young children and the elderly have an increased risk of poor

clinical outcome than patients in other age groups (Pruitt ,2002). Obese

adults and those who have underlying medical conditions, such as

diabetes, have also been shown to have higher morbidity and mortality.

AIDS patients seem to have more complications because of infection,

delayed wound healing and increased mortality (McCampbell et

al .,2002& Memmel et al . ,2004).

Extrinsic factors include invasive medical or surgical procedures;

medical devices, such as intravenous catheters , mechanical ventilators or

flexible bronchoscope . The epidemiologist discovered important failures

in processing and storage of the flexible bronchoscope . In this report,

cross infection was also a significant risk factor in the development of

infection, probably via the hands of healthcare workers (McNeil et

al .,2001& Zawacki et al .,2004).

3-Environmental factor

The potential for contaminated environmental surfaces to contribute

to transmission of healthcare-associated pathogens depends on a number

of factors, including the ability of pathogens to remain viable on a variety

of dry environmental surfaces, the frequency with which they

contaminate surfaces commonly touched by patients and healthcare

workers, and whether or not levels of contamination are sufficiently high

to result in transmission to patients (Duckworth and Jordens ,1990).

Pathogens such as Methicillin-resistant Staphylococcus aureus ,

Vancomycin-resistant enterococci and C. difficile have the ability to

remain viable on dry surfaces for days, weeks or even months. The

proportion of hospital surfaces contaminated with MRSA has varied

considerably in published reports, ranging from 1% to 27% of surfaces in

patient rooms on regular hospital wards, and from a few percent to 64%

of surfaces in burn units with MRSA patients. MRSA was recovered most

frequently from bedside rails (100% of those cultured), followed by blood

pressure cuffs (88%), television remote control devices (75%), bedside

tables and toilet seats (63% each), toilet rails and dressers (50% each),

door handles (38%) and intravenous pumps (25%) (Bhalla et al .,2004 &

Johnston et al .,2006).

Device-associated infections , such as catheter-associated urinary tract

infections (CAUTIs), central line-associated blood stream infections

(CABSIs), and ventilator-associated pneumonia(VAP) pose the greatest

threat to patient safety in intensive care units(ICUs). Multivariate analysis

revealed that ICU location, major teaching hospitals, university hospitals,

the type of ICU, the number of hospital beds, and beds per infection .

CABSI rates were higher in medical ICUs and VAP rates were greater in

surgical ICUs (Tambyah et al ., 2002).

Special units for intensive medical or surgical care for extensive

burns, trauma, transplantation, and cancer chemotherapy frequently house

patients with little resistance to infection and infectious diseases (Pittet et

al .,1998). In these patients, reduced inocula of pathogens are required to

cause infection, infection may take place at unusual sites, and usually

nonpathogenic agents may cause serious disease and death. Frequent

opportunistic infections in these patients require repeated, broad, and

extended therapy with multiple antimicrobials, leading to increasingly

resistant resident microbial populations (Goldmann et al .,1996 &

Archibald et al .,1997) .

Common HAIs

Urinary tract, respiratory tract, surgical site and bloodstream

infections are currently recognized as the major nosocomial infections.

Surveillance for these infections is well developed. However, it is

becoming increasingly clear that gastroenteritis outbreaks are also a

major burden on the health services of industrialized nations (Chadwick

et al.,2000) .

1-Urinary Tract Infection

Hospital-acquired urinary tract infections (UTIs) are the most

frequent nosocomial infections and are responsible for 20—30% of

nosocomial infections in medical or surgical intensive care units (ICUs) .

Patients aged over 50 years, diabetic patients, or patients who are

immunocompromised are more likely to develope a hospital-acquired

UTI (Mojtahedzadeh et al ., 2008) .

Urinary catheters are characterized by site of insertion (e.g., urethral,

suprapubic, or nephrostomy) and by duration of use (e.g., intermittent or

indwelling). Modern catheters are typically manufactured of latex rubber,

silicone- or Teflon-coated latex rubber, or solid silicone, and come in a

variety of types and sizes (Rosser et al .,1999) .

For the diagnosis of catheter-associated urinary tract infection

(CAUTI), the patient must meet 1 of 2 criteria. The first criterion is when

a patient with a urinary catheter has 1 or more of the following symptoms

with no other recognized cause: fever (temperature > 38°C), urgency, or

suprapubic tenderness or when the urine culture is positive for 105 colony

forming units per mL or more, with no more than 2 microorganisms

isolated. The second criterion is when a patient with a urinary catheter

has at least 2 of the following criteria with no other recognized cause

positive dipstick analysis for leukocyte esterase or nitrate, pyuria (≥ 10

leukocytes per mL of urine), organisms seen on Gram stain, physician

diagnosis of urinary tract infection, or physician initiates appropriate

therapy for a urinary tract infection ( Madani et al .,2009) .

The overall rates of CAUTIs in the 4 ICUs in Alexandria University

hospitals (15.7/1000 catheter-days) were higher than the US rates

described by the National Nosocomial Infections Study (3.9/1000

catheter-days) and the overall CAUTI rates described in 8 Latin

American countries as presented by the International Infection Control

Consortium (8.9/1000 catheter-days) (Talaat et al .,2009).

Several microbial agents have been found to be responsible for ICU-

acquired UTIs such as Escherichia coli, Pseudomonas spp, Proteus

mirabilis, Klebsiella spp, Enterobacter spp, Staphylococcus spp,

Enterococcus faecalis, Candida spp, and Enterococcus spp. E. coli is the

leading pathogen, implicated in 24% of all nosocomial UTIs . K.

pneumoniae is the fifth leading cause of nosocomial UTIs and is

recovered from 8% of cases. Enterobacter spp. and P. mirabilis are

ranked sixth, and are recovered from 5% of cases (Laupland et

al .,2005& Wagenlehner et al .,2006) .

2- Respiratory tract infections

Hospital Acquired Pneumonia (HAP) is defined as an inflammatory

condition of the lung parenchyma caused by infectious agents not present

or incubating at the time of hospital admission; that develop more than

48h after admission (Tablan et al .,2003). Ventilator-Associated

Pneumonia (VAP), on the other hand, is a subset of HAP and includes all

patients receiving mechanical ventilation at the time of infection. VAP

occurs almost exclusively in the ICU and represents approximately 86%

of all ICU HAP (Rotstein et al ., 2008).

HAP is the second most common nosocomial infection with a crude

overall rate of 6.1 per 1000 discharges . By comparison, the infection rate

for nosocomial urinary tract infection, the most common hospital-

acquired infection, is 11 per 1000 discharges. The incidence of HAP

varies depending on the hospital environment. Death from bacteremic

HAP occurred in 20% of patients within one week of their first positive

blood culture, and Pseudomonas aeruginosa bacteremia was associated

with the highest mortality rate (45%) (Rotstein et al ., 2008).

Ventilator-associated pneumonia is the diagnosis in a mechanically

ventilated patient with a chest radiograph that shows new or progressive

infiltrates, consolidation, cavitation, or pleural effusion. The patient must

also have at least 1 of the following criteria: new onset of purulent

sputum or change in character of sputum; organism cultured from blood;

or isolation of an etiologic agent from a specimen obtained by tracheal

aspirate, bronchial brushing , bronchoalveolar lavage, or biopsy ( Madani

et al .,2009). In general, the bacteriology of patients with HAP or VAP

is similar, although Stenotrophomonas maltophilia and Acinetobacter

species are found predominantly in VAP (Cross and Campbell .,2001&

Hanes et al .,2002&Vincent ,2004) .

Due to the predominance of certain virulent pathogens in HAP and

VAP, the concept of ‘core’ pathogens was developed . Core pathogens

should be considered as potential causes of HAP or VAP in all patients.

Core pathogens include Streptococcus pneumoniae, Streptococcus

species, Haemophilus influenzae, Enterobacteriaceae such as Escherichia

coli, Klebsiella species, Enterobacter species, Proteus species and

Serratia marcescens, as well as methicillin-susceptible Staphylococcus

aureus . Unusual pathogens such as Aspergillus species, Candida species,

Legionella pneumophila, Pneumocystis jiroveci (previously Pneumocystis

carinii), Nocardia species and viruses such as cytomegalovirus are causes

of HAP and VAP in patients who are immunosuppressed (Lynch ,

2001& Diaz et al .,2003).

In studies using expectorated sputum or endotracheal secretions

from intubated patients, up to 30% of the pathogens involved in

nosocomial pneumonia are Enterobacteriaceae . Enterobacter spp. and K.

pneumoniae are the third and fourth (11% and 8%) leading causes of

nosocomial pneumonia after S. aureus and P. aeruginosa (19% and 17%)

(CDC,1997) .

Multidrug resistant P. aeruginosa isolates were defined as isolates

demonstrating resistance to antimicrobials from 2 or more different

classes. P. aeruginosa and other nonfermentative Gram-negative bacilli

are resistant to many common antibiotics, including first- and second-

generation cephalosporins. Only advanced-generation cephalosporins,

extended-spectrum penicillins, carbapenems, aminoglycosides, and

fluoroquinolones generally offer useful activity (Jones et

al .,1997&Visalli et al .,1998& Quinn ,1998).

Mycobacteria distinct from the Mycobacterium tuberculosis complex

(M. tuberculosis, Mycobacterium microti, Mycobacterium bovis and

Mycobacterium africanum) and Mycobacterium leprae occasionally

cause disease in humans. Referred to by various names such as ‘atypical

mycobacteria’, ‘mycobacteria other than M. tuberculosis complex’ and

‘non tuberculous mycobacteria’, these mycobacteria have been

responsible for increasing worldwide reports of hospital outbreaks and

isolated cases of atypical or non-tuberculous mycobacterial infections

( Kotach ,2004).M. tuberculosis is by far the most frequent and most

important pathogen in this complex . Pulmonary TB is the most common

form of the disease, and the most important from the perspective of

hospital infection control (Huard et al .,2003).

The infectiousness of a TB patient correlates with the number of

microorganisms expelled into the air; this correlates with the site of

disease (pulmonary, laryngeal, tracheal, or endobronchial TB being the

most infectious), the presence of cough (or performance of cough

inducing procedures), the presence of acid-fast bacilli on sputum smears,

the presence of cavitation on chest radiograph, the duration of adequate

chemotherapy, and the ability or willingness of the patient to cover his or

her mouth when coughing (Kelly et al .,2004).

Multidrug-resistant TB (MDR-TB), defined as disease caused by

strains with resistance to at least isoniazid and rifampin,Transmissibility

continues for at least one week after the start of TB treatment, and this

period may be even longer for those with extensive disease or MDR-TB.

The ‘potential time of transmission’ of TB within the hospital is defined

as the infectious time span of pulmonary TB inpatients from date of

admission to seven days after starting anti-TB therapy (Phillips et

al .,2004&Andrews et al .,2007).

3-Surgical site infections

Surgical site infections(SSIs) are defined as infections occurring

within 30 days after a surgical operation (or within one year if an implant

is left in place after the procedure) and affecting either the incision or

deep tissue at the operation site. These infections may be superficial or

deep incisional infections (Owens and Stoessel ,2008 ) .

The increasing use of minimally invasive (laparoscopic) surgery has

resulted in a decrease in the incidence of SSIs. For example, in patients

undergoing cholecystectomy, the SSI rate following laparoscopic

procedures has been reported to be 1.1%, compared with 4% following

open procedures. Similarly, in patients with acute appendicitis, the SSI

rate has been reported to be 2% with minimally invasive procedures and

8% with open procedures (Boni et al . ,2006).

Possible reasons for the lower incidence of SSIs with minimally

invasive procedures include the smaller incision, earlier mobilization,

better preservation of immune system function, and decreased use of

central venous catheters . SSIs impose a substantial clinical burden.

Patients with SSIs are more likely to require readmission to hospital or

ICU treatment, and are at higher risk of death, than those without such

infections (Boni et al .,2006).

In most SSIs, the responsible pathogens originate from the patient’s

endogenous flora. The most commonly isolated organisms are S. aureus,

coagulase-negative staphylococci, Enterococcus spp. and Escherichia

coli; however, the pathogens isolated depend on the procedure . An

increasing number of SSIs are attributable to antibiotic-resistant

pathogens such as meticillin-resistant S. aureus (MRSA) or Candida

albicans. This development may reflect the increasing number of severely

ill or immunocompromised surgical patients, and the widespread use of

broad-spectrum antibiotics (Dohmen ,2006).

Pathogens may also originate from preoperative infections at sites

remote from the operative site, particularly in patients undergoing

insertion of a prosthesis or other implant. In addition to the patient’s

endogenous flora, SSI pathogens may originate from exogenous sources

such as members of the surgical team, the operating theatre environment,

and instruments and materials brought within the sterile field during the

procedure. Such pathogens are predominantly aerobes, particularly Gram-

positive organisms such as staphylococci and streptococci (Owens and

Stoessel ,2008 ).

4-Burn infections

Burns are one of the most common and devastating forms of trauma.

Patients with serious thermal injury require immediate specialized care to

minimize morbidity and mortality. Burn wound infections are one of the

most important and potentially serious complications that occur in the

acute period following injury (Appelgren et al .,2002).

Risk factors identified in patients colonized with drug-resistant

organisms include prior use of third-generation cephalosporins and

antibiotics active against anaerobes, critically ill patients with severe

underlying disease or immunosuppression and prolonged hospital stay

(Tredget et al .,2004).

Sources of organisms may be endogenous (patient’s own normal flora)

or exogenous (environmental or from health care personnel). Organisms

associated with infection in burn patients include Gram-positive, Gram-

negative and yeast or fungal organisms . Organisms of particular concern

include MRSA , enterococci, group A B-hemolytic streptococcus, Gram-

negative rods such as Pseudomonas aeruginosa and Escherichia coli

(Tredget et al .,1992).

The major streptococcal species encountered are group A β-

hemolytic streptococci (Streptococcus pyogenes) (GABHS), group Bα -

hemolytic streptococci (Streptococcus agalactiae) and Streptococcus

pneumoniae . GABHS were important pathogens in burn units before the

introduction of routine penicillin prophylaxis for patients with thermal

injuries. They continue to cause episodic burn wound infections and

occasional outbreaks (Stanley et al .,1997).

Fungal organisms, especially Candida (yeast) species and molds such

as aspergillus, mucor and rhizopus, have been associated with serious

infections in burn patients . Candida colonization seems to be primarily

from endogenous sources, whereas molds are ubiquitous in the

environment and can be found in air handling and ventilation systems,

plants and soil ( Rafla and Tredget , 2010).

5-Blood stream infections

Bloodstream infections are an important cause of morbidity and

mortality in immunocompromised population .In patients with

haematological disorders, infection may lead to delayed administration of

chemotherapy, prolonged hospitalisation, and additional costs associated

with directed antimicrobial therapy ( Cherif et al .,2003).

Staphylococcus aureus bacteraemia is not uncommon in patients with

haematological malignancy. One 10-year study estimated that 7% of all

bacteraemic episodes in neutropenic patients with malignancy were

caused by this organism . As with non-neutropenic patients, S. aureus

bacteraemia is associated with skin and soft-tissue infections, or with

intravascular devices ( Gonzalez-Barca et al .,2001).

The Coagulase-negative staphylococci (CoNS) are responsible for

the largest proportion of central venous catheter-related bloodstream

infections in inpatients with post-operative endophthalmitis , prosthetic

valve endocarditis , native valve endocarditis , shunt-related central

nervous system infections , pneumonia, osteomyelitis and wound

infection (O’Grady et al .,2002 ).

Coagulase-negative staphylococci are frequently identified as skin

commensals in this population, and are often regarded as a contaminant

when isolated in blood cultures ( Favre et al .,2005). However CoNS

may also be responsible for clinically significant nosocomial bacteraemia,

especially in relation to intravascular devices and underlying

immunocompromised states (Beekmann et al .,2005). This group of

organisms is the single largest cause of bloodstream and central venous

catheter (CVC)-related bloodstream infections (Velasco et al .,2004&

Ortega et al .,2005 ).

Methicillin-resistant Staphylococcus aureus (MRSA) has been

recognised as an important and universal hospital-acquired pathogen

causing endemic and epidemic infections in healthcare centres

worldwide. MRSA bacteraemia is usually hospital acquired, has a high

incidence in ICU, and is commonly associated with intravenous devices.

Several different genotypes of MRSA rather than a specific type are

associated with invasive infections in a hospital . Mortality associated

with MRSA bacteraemia has been reported to be significantly higher than

Methicillin-susptible Staphylococcus aureus (MSSA) bacteraemia

(Whitby et al .,2001).

Streptococcus pneumoniae bacteraemia may be associated with

pneumonia or CVC-related infection. Risk factors for pneumococcal

bloodstream infection include autologous transplantation with total body

irradiation conditioning, allogeneic bone marrow transplantation , and

chronic Graft versus host disease , with significant mortality

(approximately 20%) in both early and late transplant periods (Engelhard

et al .,2002).

Outbreaks of Pseudomonas spp. bacteraemia have been reported in

relation to disinfectant fluids. Pseudomonal bloodstream infections may

be associated with high mortality ( Vianelli et al .,2006& Siebor et

al .,2007).

In adult Bone marrow transplant recipients, anaerobic bloodstream

infections have been found to be associated with severe mucositis.

Anaerobic bacteraemia in the haematology population has also been

associated with neutropenic enterocolitis ( Lark et al .,2001). In patients

with haematological malignancy, colonization with Candida and previous

glycopeptide therapy have been associated with increased risk for

development of candidaemia (Worth and Slavin , 2009).

Candida species were isolated as a causative agent in 10—15% of all

nosocomial infections, 70—80% of all nosocomial fungal infections and

of 8—10% of nosocomial bloodstream infections within the last 10—15

years. Candidemia is the most frequently encountered clinical form of

invasive candidiasis (Diekema et al .,2004& Puzniak et al .,2004) .

C. albicans (40—60%), C. glabrata, C. tropicalis and C.

parapsilosis are the most frequently encountered causative agents in

candidemia (Viudes et al .,2002&Pfaller et al .,2006). Cutaneous lesions

may develop in patients with candidemia, especially those with acute

leukemia. Although these lesions may be extremely variable in number

and appearance, they are usually described as firm, erythematous, raised

nodules (Darmstadt et al .,2000).

The use of total parenteral nutrition , candida colonization (or

candiduria) and surgery (especially abdominal surgery) were the most

frequently encountered independent risk factors for candidemia.

However, the type of ICU, the severity of the underlying diseases and

antibiotic use policies can also have an important effect on the risk factors

found in all ICU candidemia studies (Gürcüog et al .,2010).

6-Device associated infection

Central venous catheter-associated blood stream infection is

laboratory-confirmed when a patient with a CVC has a recognized

pathogen that is isolated from 1 or more percutaneous blood cultures after

48 hours of vascular catheterization and is not related to an infection at

another site. The patient also has at least 1 of the following signs or

symptoms: fever (temperature > 38°C), chills, or hypotension. With skin

commensals for example, Diphtheroids, Bacillus spp., Propionibacterium

spp., Coagulase negative staphylococci, or micrococci) ( Madani et

al .,2009).

There are two main pathways leading to catheter related bloodstream

infections . The first is contact between skin surface organisms either at

the time of insertion or thereafter, leading to migration of organisms

down, and colonization of, the external catheter surface. It is thought to

be the dominant mechanism associated with short-term catheters. The

second involves transfer of organisms to the catheter hubs from patient

skin or healthcare workers’ hands, usually leading to colonisation of the

internal catheter surface, and is more common in long-term

catheters(Maki et al .,1997).

In developing and transitional economy countries, nosocomial

transmission of hepatitis C virus (HCV) through the re-use of

contaminated or inadequately sterilised syringes and needles used in

medical, paramedical and dental procedures remains a major source of

HCV infection and puts the public in these areas at high-risk (Simonsen

et al .,1999& Kane et al .,1999).

In Egypt, treatment of endemic schistosomiasis in mass programmes

(discontinued in the 1980s) that frequently used unsterilised needles and

syringes has lead to a national HCV prevalence of more than 14%, with

rates of 20–30% in young male adults (Meky et al .,2006).HCV

nosocomial transmission continues to be a problem in many countries

because of ongoing reuse of unsterilised needles and syringes in health

care facilities and programmes .Chronic HCV infection is common in

patients on chronic haemodialysis, with prevalences of 10–33% (Medhat

et al .,2002& Stoszek et al .,2006).

Intravascular catheters may not only serve as the portal of entry

for Candida species and be an important primary source of candidemia

but also provide a secondary site of attachment for Candida species that

invade the bloodstream from other sites, most frequently the

gastrointestinal tract (Walsh et al ., 1993) .

7-Ocular Infections

Postoperative endophthalmitis (POE) is defined as a severe

inflammation involving both the anterior and posterior segments of the

eye secondary to an infectious agent. Bacterial and fungal

endophthalmitis following penetrating keratoplasty is unusual, but the

frequency is likely higher than the rate of endophthalmitis following

cataract surgery or pars plana vitrectomy (Taban et al .,2005&Wilhelmus

and Hassan ,2007).

Complications of POE may be devastating. Despite appropriate

therapy, POE results in severe visual loss in at least 30% of patients, and

retinal detachment in 8–10% of patients ( Doft et al .,2000). Blindness

secondary to POE has been reported in up to 18% of patients (Yamada et

al .,2002). Common pathogens associated with POE are Staphylococcus

aureus (24%), coagulase negative Staphylococci (23%), Pseudomonas

aeruginosa (13%), Streptococcal species (8%), and Escherichia coli

(7%). Extrinsic contamination of a new intravenous anesthetic agent

without a preservative was also responsible for an outbreak of post

surgical C. albicans fungemia and endophthalmitis ( David et al .,2004) .

8-Central Nervous System Infections

Bacterial meningitis is a significant problem among hospitalized

patients. Risk factors for nosocomial meningitis were categorized as:

(1) a history of neurosurgery, CSF leakage or recent head trauma

(2) a distant focus of infection (e.g. otitis/sinusitis or pneumonia) or

(3) an immunocompromised state (Weisfelt et al ., 2007).

Isolated micro-organisms were classified as cutaneous

(Staphylococcus aureus, coagulase-negative Staphylococci,

Propionibacterium acnes), or non cuteaneous (Enterobacteriaceae,

Pseudomonas aeruginosa, Acinetobacter spp., Streptococci,

Pneumococci, enterococci)( Korinek et al .,2006).Factors associated with

a nosocomial Acinetobacter sp. meningitis include intraventricular

haemorrhage and invasive neurosurgical procedures (Wang et al .,2005).

9-Ear & Nose infection

Posttympanostomy tube otorrhea (PTTO), defined as active otorrhea

from the middle ear cavity through the patient tympanostomy tube, is the

most common complication of tube insertion. PTTO can be classified

according to time of its onset, with early PTTO occurring within 2 weeks

of tube insertion, and late (delayed) PTTO occurring after 2 weeks. PTTO

persisting for over 8 weeks can be classified as chronic PTTO. the

incidence of early PTTO has been reported to range from 5 to 38%. A

recent meta analysis found that late PTTO occurred in 26% of patients,

transient PTTO in 16%, and chronic PTTO in 3.8% ( Jung et al .,2009).

PTTO was Previously found to be caused by the same strains of

bacteria that give rise to acute otitis media , most typically Haemophilus

influenzae, Moraxella catarrhalis and Streptococcus pneumoniae.

Recently, however, atypical strains, including Staphylococcus aureus and

Pseudomonas aeruginosa, have also been isolated from PTTO. These

include antibiotic-resistant strains, including MRSA ( Coticchia and

Dohar ,2005).

Sinusitis, one of the complications of intubation or mechanical

ventilation, can lead to severe sepsis in critically ill patients. Several

studies have shown that prolonged naso-gastric or naso-tracheal

intubation, prolonged mechanical ventilation, administration of high-dose

corticosteroids and prolonged antibiotic therapy are risk factors for

sinusitis . In addition traumatic head injury may have a higher risk in

terms of maxillary sinusitis than mechanically ventilation, because of the

possible existence of bleeding in maxillary sinuses (Cengiz et al .,2009).

10-Gastrointestinal Tract Infections

Nosocomial outbreaks of gastroenteritis are a major disruption to

health services in many countries. Noroviruses are the predominant

pathogen detected in such outbreaks (Billgren et al .,2002& Green et

al .,2002&Lopman et al .,2004).Nosocomial Rotavirus gastroenteritis

(RVGE) is a major component of hospital-acquired infections in children

(Gleizes et al .,2006& Waisbourd-Zinman et al .,2009).

Prolonged hospitalisation, age above 65 years, antibiotic usage,

underlying medical conditions, neoplastic disease, gastrointestinal

surgery, nasogastric tubes, and gastrointestinal disorders including

inflammatory bowel diseases are the most important risk factors for the

development of nosocomial C. difficile-associated disease ( CDAD)

(McFarland et al .,2007& Issa et al .,2007) .

Studies have found that the prevalence of asymptomatic colonization

with C. difficile is 7%–26% among adult inpatients in acute care facilities

and is 5%– 7% among elderly patients in long-term care facilities. Other

studies, however, indicate that the prevalence of asymptomatic

colonization may be more on the order of 20%–50% in facilities where

clostridium difficile infection is endemic. The risk of colonization

increases at a steady rate during hospitalization, suggesting a cumulative

daily risk of exposure to C. difficile spores in the healthcare setting

( Rivera and Woods , 2003& Riggs et al .,2007) .

The disease has been reported in children aged from 5 days to 17

years. Risk factors for children include disrupted normal microflora of the

gastrointestinal tract (antibiotic-associated and non-antibiotic associated),

age, immune response, diet, underlying conditions, concurrent infections,

and cancer (McFarland et al .,2000) .

Ciprofloxacin and other oral quinolones are active against many

intestinal bacteria and are highly concentrated in the stool. Patients with

CDAD are at least five times more likely to have been exposed to

ciprofloxacin than control patients (Angel et al ., 2004).

Over the last two decades, candida has emerged as an important

healthcare-associated pathogen in the world .There are few studies on the

source of candidaemia. The gastrointestinal tract has been considered the

main endogenous reservoir of Candida spp., but there is increasing

evidence of exogenous acquision (Nucci and Anaissie ,2001& Aragao et

al .,2001).

11- Obstetrics and neonatal Infections

Pregnant women are more susceptible to certain infections due to

reduced cell-mediated immunity and raised corticosteroid levels. The

onset of life-threatening sepsis in pregnant women can be insidious, with

rapid clinical deterioration, and pyrexia is not always present. One reason

that the prognosis is still more favourable than the nonobstetric

population, is that the source of infection is usually the pelvis, and

potentially more amenable to intervention (Farmer et al .,2005).

Conditions associated with an increased risk of sepsis are prolonged

rupture of membranes, emergency Cesarean Section , instrumentation of

the genital tract, and retained products of conception or placenta.

Endotoxin producing aerobic Gram negative bacilli are the most common

cause (60–80%), but sometimes Gram-positive bacteria, mixed (usually

anaerobes such as Bacteroides or Clostridium) or fungal infections are

implicated (Nelson-Piercy ,2006).

Septic shock with Disseminated Intravascular Coagulopathy is an

ominous sign if it develop . Sepsis is defined as Systemic inflammatory

response syndrome secondary to an infection, and severe sepsis when

there are features of organ dysfunction such as hypotension or oliguria.

Septic shock has developed when hypotension persists despite adequate

fluid resuscitation. Sepsis is the leading cause of multiple organ failure,

acute renal failure, and Acute Respiratory Distress Syndrome , and carries

a mortality of 40–60% (James et al .,2005) .

Vaginal flora is a dynamic ecosystem, with some differences between

vaginal and cervical flora. Anaerobic bacteria usually outnumber aerobes,

with anaerobic and facultative lactobacilli predominating. Other

anaerobes include Peptostreptococcus species, Bacteroides species, and

Prevotella species. The aerobic gram-positive flora include coagulase-

negative staphylococci, with varying amounts of streptococci,

enterococci, and Staphylococcus aureus, and the gram-negative flora

include Escherichia coli, Gardnerella vaginalis, Enterobacter species,

Klebsiella pneumoniae, and Proteus mirabilis . Both Mycoplasma and

Ureaplasma are also found in the vagina. Vaginal flora may change

during pregnancy. Some studies suggested that lactobacilli increase in

pregnancy and that other anaerobes decrease . Antibiotics also change the

flora, and the use of multiple doses of cephalosporins for prophylaxis has

been reported to increase enterococci and perhaps Enterobacter

species .C. albicans is a part of the normal microbial flora of the human

respiratory, enteric, and female genital tracts (Casey and Cox ,1997).

Group B streptococcus (GBS) is the leading cause of neonatal sepsis

and meningitis. Early-onset (EO) GBS disease is usually defined as

infection presenting in the first 6 days of life (some definitions use the

first 2, 3 or 5 days as the cut-off) and accounts for approximately 60–70%

of all GBS disease in the first 3 months of life. Maternal carriage of GBS

in the gastrointestinal and/or genital tracts is a prerequisite for EO

disease, vertical transmission occurring prior to or during birth. An

estimated 20–30% of pregnant women are colonized by GBS(Jones et

al .,2006), 50% of babies become colonized perinatally, and 1% becomes

infected. Disease occurs rapidly, being evident at birth or within 12 hours

in over 90% of cases, and presenting with overwhelming sepsis (60%) or

pneumonia (25%) (Heath et al .,2004).

Late-onset disease appears to have a different pathophysiology. It is

predominantly caused by serotype III, is acquired perinatally,

nosocomially, or from community sources, and in up to 50% of cases

presents with meningitis (Weisner et al .,2004).GBS disease is associated

with significant morbidity and mortality. GBS meningitis leaves half of

those infected with long-term neurodevelopmental effects at 5-year

follow-up. Neurodevelopmental impairment is also associated with

clinical (i.e. culture-negative) neonatal sepsis (Stoll et al .,2004).

Another GBS disease that is difficult to quantify is prematurity. The

association between the two is complex, but GBS is recognized as one of

the infections that may cause prematurity. The incidence of invasive

disease is certainly higher among preterm infants than among those born

at term (Malek et al .,1996). An important contributory factor may be the

lack of maternal antibody, as materno-fetal transport of IgG is inversely

related to gestational age. Thus, preterm infants born to GBS-colonized

mothers will have low or absent concentrations of protective maternal

antibody. Premature rupture of membranes (i.e. rupture before

spontaneous onset of labour) is strongly associated with early-onset GBS

disease (Oddie and Embleton ,2002).

Diagnosis of HAIs

Sampling

Examples of biological materials that are analyzed in clinical

laboratories include whole blood , serum , plasma , urine , feces , saliva ,

spinal and other body fluid . All these specimens also used for molecular

testing . Specimens devoid of patient cells are useful in detection of

infectious agents (CLSI .,2006) .

1-Blood

Special blood-collecting equipment is used . Sterile collecting

bottles containing the proper nutrient broth media , blood collecting sets

with needles and tubing that allow the blood to flow into the collecting

bottle and the proper skin-cleaning supplies are necessary to ensure a

properly collected blood specimen for culture . Special care must be taken

to clean the venipuncture site carefully before puncture to avoid possible

contamination of blood sample with skin contaminants . One method uses

an initial cleaning with a 70% solution of alcohol to remove dirts and

lipids . A circular motion moving from puncture site out is used . A 1-

10% povidine-iodine solution is used , followed by an alcohol rinse

(CLSI ., 2003).

2-Cerebospinal fluid

Physician collects Cerebospinal fluid (CSF) through a lumbar

puncture. Rapid handling of CSF samples in the laboratory is important

because of serious nature of meningitis and organism in meningitis are

sensitive to temperature change ,so refrigeration not done . CSF is

typically placed in sterile tubes . The tubes are sent to laboratory

immediately for testing , including culture and Gram stain (Finn ,2007) .

3-Stool

Stool specimens contain large numbers of bacteria (normal flora) ,

and a stool specimen is usually cultured only to isolate certain types of

pathogenic enteric organism . Stool specimen should be cultured within 2

hours . If this cannot be done , transport media special for stool samples

can be used . Swabs of the rectal area can be used for infant , but this is

not the preferred method of collection for other age groups (Finn,2007) .

4-Sputum

When a specimen of sputum is collected , the patient must cooperate

fully to ensure that a proper specimen is obtained . Sputum is usually

collected in the morning and it should be sent to the laboratory and

processed immediately . Deep coughing will usually bring up a good

sputum specimen . It is necessary to avoid collecting saliva . A wide

mouthed sterile container is best used for collecting this type of specimen.

An acceptable sputum specimen that free from contamination with saliva

can be Gram-stained and checked microscopically for the presence of

squamous epithelial cells . Finding an average of more than 10 squamous

epithelial cells per low power field indicate that the specimen is saliva

and is not acceptable specimen to culture ( Muarry et al ., 2003) .

5-Swab of various fluids

Swabs are used to collect cultures from various openings of the body

, such as the nose ,throat , mouth , vagina , anus and wound . These swabs

must be collected carefully and placed in the proper transport media

before they are taken to the laboratory for processing . If swabs are not

properly handled , the micro-organism may dry out , or their numbers

may be insufficient for culture (Finn , 2007) .

6-Urine

The collection of urine for microbiological studies also requires the

cooperation . A clean –catch midstream sample , usually the first morning

specimen , is suitable for culture , provided care has been taken to clean

the uretheral area before the collection . A sterile container must be used

for the collection of the urine for the culture . When the patient is too ill

or cannot void properly , aspecimen is obtained by catheterization . After

collection , specimens should be sent to the laboratory for the immidate

processing or refrigeration , or a preservative can be used to maintain

bacterial counts (CLSI ., 2001) .

Methods :

Historically, this analysis of nosocomial pathogens has relied on a

comparison of phenotypic characteristics such as biotypes, serotypes,

bacteriophage or bacteriocin types, and antimicrobial susceptibility

profiles. This approach has begun to change over the past 2 decades, with

the development and implementation of new technologies based on DNA,

or molecular, analysis. These DNA-based molecular methodologies ,

include pulsed-field gel electrophoresis (PFGE) and other restriction-

based methods, plasmid analysis, and PCR-based typing methods. The

incorporation of molecular methods for typing of nosocomial patho-gens

has assisted in efforts to obtain a more fundamental assessment of strain

interrelationship (Cockerill and Smith, 2004).

A number of nosocomial infections are endemic, epidemic or

sporadic infections. Most nosocomial infections are endemic and are the

focus of most infection control efforts. The earliest methods that were

used to identify and type organisms were based upon their phenotypic

characteristics. This is followed by a series of genotypic methods (Figure

2) , one of the most widely utilized techniques is biotyping, or the

differentiation of strains based on properties such as differences in

biochemical reactions, morphology, and environmental tolerances (Singh

et al . ,2006 ).

There are a number of important attributes for successful typing

schemes : the methodologies should be standardized, sensitive, specific,

objective, and subject to critical appraisal. All typing systems can be

characterized in terms of typeability, reproducibility, discriminatory

power, ease of performance and interpretation, and cost (in terms of time

and money) . Typeability refers to the ability of a technique to assign an

unambiguous result (type) to each isolate. Non typeable isolates are more

common with phenotypic methods but can also occur with genotypic

methods. The reproducibility of a method refers to the ability to yield the

same result upon repeat testing of a bacterial strain. The discriminatory

power of a technique refers to its ability to differentiate among

epidemiologically unrelated isolates, ideally assigning each to a different

type . In general, phenotypic methods have lower discriminatory power

than genotypic methods (Olive and Bean ,1999).

Figure 2. Recent evolution of bacterial strain identification for

epidemiological purpose ( Belkum , 2007)

A - Phenotypic methods

1-Biotyping

Biotyping is often used to determine the species of microorganisms

based upon their abilities to utilize components in different growth media

and carry out certain chemical reactions, but it can also be used to

separate members of a particular species due to biochemical differences

among the organisms . Biotyping often lacks discriminatory power

because of variations in gene expression and random mutations that may

alter biologic properties of microorganisms. Biotyping cannot

differentiate among strains where biochemical diversity is uncommon,

such as the enterococci, and therefore the utility of biotyping in

epidemiologic studies is quite limited (Struelens ,2002) .

2-Antimicrobial susceptibility testing

Strains defined by this method should always be confirmed by

genomic typing, because unrelated clones can undergo evolutionary

convergence to the same resistance phenotype under antibiotic selective

pressure, through mutations and genetic exchanges. Antimicrobial

susceptibility testing is a common practice in the clinical microbiology

laboratory. The resultant antibiogram indicates the pattern of in vitro

resistance or susceptibility of an organism to a panel of antimicrobial

agents . Antimicrobial susceptibility testing is typically performed using

either automated broth microdilution or disk diffusion methods. Disk

diffusion methods are not used as commonly as they once were because

of the lack of automation for testing. Microdilution testing provides a

quantitative measure of the minimum bactericidal concentration (MIC),

which is defined as the lowest concentration of the antimicrobial agent

that inhibits the growth of the organism (Barenfanger et al .,1999).

Bactericidal activity can be measured by one of three methods:

(1) calculation of the minimum bactericidal concentration;

(2) performance of time-kill studies; or

(3) serum bactericidal assay . The minimum bactericidal concentration is

obtained by subculturing the tubes (macrodilution) or wells

(microdilution) that do not demonstrate growth at 24 hours and is defined

as the lowest concentration of antibiotic at which a 99.9% reduction of

viable organisms occurs.A bactericidal drug achieves this within two

dilutions of the MIC (Bates et al .,1991).

Agar dilution is a well-established technique for obtaining

quantitative susceptibility results and is the reference method commonly

performed in Europe. Mueller-Hinton agar (MHA) is the recommended

medium for the testing of most commonly encountered aerobic and

facultative anaerobic bacteria and is standardized such that calcium and

magnesium supplementation is not indicated. For predictable diffusion,

the depth of the agar should be between 3 and 4 mm. The recommended

final inoculum is 104 CFU per spot. The method is labor-intensive and

costly, but is recommended for fastidious organisms that do not grow

well in broth media (Neisseria gonorrhoeae, anaerobes)( Jorgensen and

Turnidge , 2007).

The disk diffusion method has been standardized for the testing of

common, rapidly growing organisms and allows for a qualitative

categorization of isolates as susceptible, intermediate, or resistant. An

antibiotic-impregnated filter paper disk is placed on the surface of an agar

plate inoculated with a ‘‘lawn’’ of organism at a known turbidity (0.5

McFarland). The inoculum may be prepared by either log phase growth

or direct suspension from colonies on the agar plate. As with dilution

methods, direct suspension is preferred for fastidious organisms and for

detection of methicillin resistance in staphylococci. The antibiotic

diffuses through the agar almost instantaneously after placement on the

agar and creates a zone of inhibition that can be measured in millimeters

(edge to edge, including the disk) (Figure 3) ( Jorgensen and

Ferraro ,2000) .Interpretation of the test is based on the inverse

correlation of the zone diameter with MIC for each combination of

antimicrobial and organism (CLSI,2008).

Figure 3. Disk diffusion testing of a hyper b-lactamase producing

Staphylococcus aureus. The zone of inhibition (clear halo) around the

cefoxitin disk (FOX) is sharp. In contrast, the zone of inhibition around

the oxacillin disk (OX) is blurred by the presence of individual colonies

(Malhotra-Kumar et al .,2008).

3-Serotyping

Serotyping uses a series of antibodies to detect antigens on the

surface of bacteria that have been shown to demonstrate antigenic

variability . Serotyping methods have been used for decades for the

taxonomic grouping of a number of bacterial pathogen species and

remain important for typing Salmonella, Legionella, Shigella, and

Streptococcus pneumoniae isolates.Serotyping also has been shown to

have epidemiologic value in differentiating strains within species of

nosocomial pathogens such as Klebsiella and Pseudomonas ( Reis et

al .,2000) .

There are a number of different ways in which serotyping can be

performed; each varies in the way in which the antibody-antigen reactions

are detected. Often direct antibody-antigen agglutination is used, in which

a bacterial cell suspension is mixed with panels of antibodies. Based upon

agglutination profiles, the serotype is determined. Additionally, for

organisms such as S. pneumoniae the quellung test is used, in which test

antibodies bind to the corresponding capsular antigens and induce

swelling of the capsule, which can be observed with microscopy.

Limitations of serotyping include a lack of availability of certain antisera

and problems with standardization of different methods(Babl et

al .,2001).

4-Bacteriophage and bacteriocin typing

Bacteriophage and bacteriocin typing as epidemiologic tools are

limited to bacteria. Bacteriophage (phage) (figure 4) typing classifies

bacteria based on the pattern of resistance or susceptibility to a certain set

of phages. Bacteriophages are viruses that are able to attach to the cell

walls of certain bacteria, enter, multiply, and lyse the cells. The

differential ability of phages to infect certain cells is based upon the

availability of corresponding receptors on the cell surface for the phage to

bind. Often different strains of pathogens have a different cohort of

receptors, leading to variable lysis profiles ( Hopkins et al .,2004).

Bacteriophage typing has some drawbacks due to a lack of

widespread availability of biologically active phages and the technical

difficulty of performing the technique, but the method has been applied to

a number of bacteria associated with nosocomial infections, such as S.

aureus, P. aeruginosa and Salmonella species. Additionally strains can be

typed based on their susceptibility to a set of heterogeneous substances

(generally proteins) that are produced by other bacteria. These inhibitory

compounds, or bacteriocins, often limit the growth of closely related

species . Bacteriocin typing has had limited utility because of drawbacks

similar to those of phage typing, but it has been used for typing P.

aeruginosa . Additionally, an analogous approach has been developed for

Candida species (particularly C. albicans) (Singh et al .,2006 ).

Figure 4 . Phage typing is the identification of bacterial species and

strains by determining their susceptibility to various phages

(pearson , 2004) .

B-Genotypic methods.

Molecular techniques can be very effective in tracing the spread of

nososcomial infections due to genetically related pathogens, which would

allow infection control personnel to more rationally identify potential

sources of pathogens and aid infectious disease physicians in the

development of treatment regimens to manage patients affected by related

organisms. Therefore, the use of molecular tests is essential in many

circumstances for establishing disease epidemiology, which leads to

improved patient health and economic benefits through the reduction of

nosocomial infections (figure 5) (Singh et al .,2006 ).

The isolates involved in a nosocomial outbreak are genetically

related and thus originate from the same strain. Therefore, the use of

strain typing in infection control decisions is based on several

assumptions: (i) isolates associated with the outbreak are recent progeny

of a single (common) precursor or clone, (ii) such isolates will have the

same genotype, and (iii) epidemiologically unrelated isolates will have

different genotypes (Singh et al .,2006 ).

Figure 5. Flow chart comparison of the different procedural steps

used for various molecular typing techniques (Chang and

Chui ,1998).

1- Plasmid Analysis

Among genotyping methods, plasmid analysis is essential for the

epidemiologic analysis of HAIs with multiple antibiotic- resistant

organisms and for tracing dissemination of mobile antibiotic-resistance

genes. Plasmid typing was the first molecular method to be used as a

bacterial typing tool . Plasmids are self-replicating, often-transferable

extrachromosomal DNA elements in the prokaryote cytoplasm. Typing is

performed through the isolation of plasmid DNA and comparison of the

numbers and sizes of the plasmids by agarose gel

electrophoresis(Eisgruber et al .,1995).

Some bacteria have large plasmids (Figure 6) in the range of 100 to

150 kb, making their separation difficult; for these strains, the addition of

a restriction endonuclease digestion step following plasmid isolation will

often aid in typing because multiple fragments are generated, which

makes interpretation of strain relatedness more feasible. Plasmid

restriction is also commonly used for the analysis of staphylococci and

enterococci, whose plasmids are typically less than 50 kb in size. The

inclusion of restriction enzyme analysis increases the discriminatory

power of plasmid analysis (Liu et al .,1996).

Plasmids are not generally helpful in differentiation between endemic

and epidemic strains, because plasmids are often mobile

extrachromosomal DNA fragments that can be acquired and deleted. A

consequence of this plasmid mobility is that epidemiologically related

isolates can exhibit different plasmid profiles. Many plasmids carry

antibiotic resistance determinants that are contained within mobile

genetic elements (transposons) that can move in or out of plasmids and

the chromosome, allowing for the DNA composition of a plasmid

potentially to change rapidly (Figure 6) (Feil et al ., 2003).

A transposon epidemic is suggested when isolates from different

species, or isolates of the same species with differing PFGE and profiles

plasmid contents, have similar resistance genes. Further analysis by PCR

and DNA sequencing of transposon content by insertion sequence

evaluation have been useful to establish the potential of a transposon

epidemic (Thal et al .,1998).

Figure 6 ; Schematic diagram of Plasmid Analysis (Access

Excellence , 2009)

2- Pulsed-field gel electrophoresis

The chromosome is the most fundamental component of identity of

the cell and therefore represents a preferred measure for assessing strain

interrelatedness. One approach has been to digest chromosomal DNA

with restriction enzymes, resulting in a series of fragments of different

sizes that form different patterns when analyzed by agarose gel

electrophoresis (Figure 7) . Enzymes used to cleave DNA often recognize

numerous sites within the bacterial chromosome, resulting in too many

band fragments to efficiently and accurately compare following

conventional agarose gel electrophoresis. More recently, restriction

enzymes that cleave chromosomal DNA less frequently (<30) recognition

sites have been utilized for analysis. The resulting DNA fragments are too

large to be separated by conventional agarose gel electrophoresis . A

number of alternative methods, generally classified as PFGE, are capable

of separating these large DNA fragments (Stephenson , 2004 & Roberts

et al ., 2010). It has excellent discriminatory power and is broadly

applicable to bacteria and yeasts. Consensus rules are available for

interpretation of patterns in outbreak investigations ( Struelens et al .,

2001).

In conventional agarose gel electrophoresis, DNA molecules that are

more than 40 to 50 kb in size fail to migrate efficiently. By periodically

changing the direction of the electrical field (Figure 7) in which the DNA

is separated, PFGE allows the separation of DNA molecules of over

1,000 kbp in length (often referred to as megabase-sized DNA). PFGE

methods differ in the way the pulsed electric field is delivered to the

agarose gel. Two of the most commonly utilized approaches are contour-

clamped homogenous electric field (CHEF) and field inversion gel

electrophoresis (Goering , 2010).

Field inversion gel electrophoresis utilizes a conventional

electrophoresis chamber in which the orientation of the electric field is

periodically inverted by 180o. CHEF uses a more complex electrophoresis

chamber with multiple electrodes to achieve highly efficient electric field

conditions for separation; typically the electrophoresis apparatus reorients

the DNA molecules by switching the electric fields at 120o angles. CHEF

has been used to evaluate the spread of various antimicrobial resistant

bacteria. The finding of isolates that have identical or related restriction

endonuclease patterns suggests spread from single strains (Finney, 1993).

Figure 7 . Diagram of pulsed-field gel electrophoresis (Davidson ,

2002) .

Ideally, the PFGE isolates representing an outbreak strain will be

indistinguishable from each other and distinctly different from those of

epidemiologically unrelated strains. If this occurs, the outbreak is

relatively easy to identify. Alternatively, random genetic events, such as

point mutations or insertions and deletions of DNA, that can alter the

restriction profile obtained during the course of an outbreak can occur

(Quintiliani and Courvalin, 1996& Thal et al .,1997).

The purpose of interpretative criteria is to establish a guide for

distinguishing true differences in strains from random genetic

polymorphisms that may occur over the time of a given nosocomial

outbreak. Appropriate interpretative criteria provide consistent, objective

guidelines for correlating restriction pattern variations observed between

individual isolates and the putative outbreak strain and provide an

estimate of the likelihood that the isolate is part of the outbreak

( Goering, 1998).

This correlation focuses on the number of genetic events required to

generate the observed pattern variation. Because only a small portion of

the organism’s genetic component is undergoing analysis, isolates that

give identical results are classified as “indistinguishable,” not “identical.”

With the detection of two genetic variation events by differences in

fragment patterns compared to the outbreak strain, the determination of

relatedness to an outbreak falls into a gray zone. The results may indicate

that these isolates are related (especially if isolates were collected over a

long period of time, such as 3 to 6 months), but there is also a possibility

that strains are unrelated and not part of the outbreak (Struelens , 1996 &

Tenover et al .,1997).

3-Southern Blot Analysis-Ribotyping

The bacterial DNA is digested using a frequent cutting restriction

enzyme, the DNA fragments are separated by agarose gel electrophoresis,

and then the fragments are transferred (blotted) onto a nitrocellulose or

nylon membrane. Next, a labeled (colorimetric or radioactive) piece of

homologous DNA is used to probe the membrane. Under the appropriate

conditions, the probe hybridizes to a complementary base pair, and the

banding patterns are resolved through the detection of the probe label.

The discriminatory power of this method is related to the copy numbers

of the targeted genetic elements in the bacterial genome and their

distribution among the restriction fragments following electrophoresis.

Variations in the number and sizes of fragments detected are used to type

the microorganisms (Singh et al , 2006).

One of the most common targets for Southern blotting is the gene for

the rRNA, and the targeting of the rRNA gene is referred to as ribotyping.

Typically, the discriminatory power of ribotyping has been shown to be

less that of PFGE or some PCR-based methods ; however, a variety of

organisms have been studied using this method . Most bacterial species

have several ribosomal operons per chromosome and produce ribotype

patterns of 5–15 bands. Ribotyping exhibits excellent reproducibility but

only moderate discriminatory power for most bacterial species. An

automated ribotyping system is commercially available (figure 8) (

Maiden et al ., 1998).

Figure 8 . schematic diagram of Southern Blot Analysis (pearson ,

2004) .

4-Northern blotting

Northern blotting is a technique that uses RNA rather than DNA .

RNA is transferred from the gel after electrophoresis onto a solid support

followed by hybridization with specific labeled probe . Because RNA

molecules have defined lengths and are much shorter than genomic

DNA ,it is not necessary to cleave RNA before

electrophoresis .However , because of the secondary structure of RNA , it

is necessary to perform electrophoresis under denaturing condition,

usually with formaldehyde or fomamide buffer in agarose gels. RNA

extracted from cells consists primarily of ribosomal and transfer

RNA .The mRNA comprises only 1% to 2% of total cellular RNA . After

electrophoresis and staining ,intact RNA reveals two clearly visible bands

of ribosomal RNA (figure 9) (Benson et al .,2002).

Figure 9 . Schematic diagram of Northern blotting (Alberts etal .,

2008).

5-Heteroduplex Migration Analysis

Heteroduplex migration analysis reveals the presence of mutation by

the altered electrophoretic mobility of a dsDNA fragment that contains

one or more mismatched bases (heteroduplex ) versus one that is

perfectly matched ( homoduplex ). Originally described as a PCR

artifact , heteroduplex migration has become a popular mutation –

scanning technique ,primarily because of its technical simplicity .With

this technique , the dsDNA generated by PCR is denatured and then

allowed to reanneal , followed by electrophoresis under slightly

denaturing conditions (e.g. 15% urea ,40 0 c ) on polyacrylamide

gels .Detection is performed by silver staining of the gel or by

fluorescence detection if one of the PCR primers is labeled .

Heteroduplexes usually tend to migrate more slowly than

homoduplexes during electrophoresis . Whereas mutant alleles are often

present as heterozygotes in a clinical specimen , homozygous mutation

require mixing with wild –type DNA for the mutation to be detected

(Schouten et al .,2002).

6-Single Strand Conformation Polymorphism Analysis

Single strand conformation polymorphism analysis is a technique

used to scan for unknown variants in nucleic acid sequence . Similar to

heteroduplex analysis , it first requires PCR amplification . The

amplification is then diluted , denatured with heat and formamide , and

the resulting ssDNA is separated by non denaturing polyacrylamide

electrophoresis (usually run at 40 c ) . During electrophoresis the single –

stranded molecules fold into three-dimensional structures according to

their primary sequence . Electorphoretic mobility then becomes a

function of size and shape of the folded single –stranded molecules . If

the sequence of a reference sample differs from that of the fragment being

tested , even by only a single nucleotide , often at least one of the strands,

if not both , will adopt different conformations and exhibit a unique

banding pattern . Results are visualized by silver staining of the gel or by

fluorescent detection using labeled primers during PCR (Reed and

Wittwer , 2004) .

7-Typing Methods Using PCR

a-Multiplex PCR

In order to increase the efficiency of PCR typing and reduce reagent

costs, multiple sets of primers can be included in a single reaction tube in

a process termed multiplex PCR. A key strategy in the development of a

multiplex PCR assay is the design of the primers. Primers must be

designed such that all of the primers have very close annealing

temperature optimum, and the amplification products that they produce

need to be of notice different sizes to facilitate interpretation. If the

amplification products were too close in size, it would be difficult to

determine the identity of the amplification product (Focucault et

al .,2005). An additional concern with multiplex PCR is that the mixing

of different primers can potentially cause interference in the amplification

process, thus making optimization of the reaction difficult, especially as

the number of primer pairs in the reaction mixture increases (Francois et

al .,2004).

b-Nested PCR

When there is an extreme need for sensitivity and specificity in PCR,

the process of nested PCR can be carried out. Nested PCR involves the

sequential use of two PCR primer sets. The first primer set is used to

amplify a target sequence , the amplicon generated then serves as the

template for a second amplification using primers internal to those of the

first amplicon. This secondary amplification proceeds only if the intended

target was initially amplified; if the primary amplification was

nonspecific, the secondary amplification would not occur . A major

drawback of nested PCR is that the reaction vessel needs to be opened in

order to add the second primer set, increasing the potential for

contamination of the work environment with amplified DNA (Eribe and

Olsen , 2000).

c- Arbitrarily primed PCR

Arbitrarily primed PCR (AP-PCR) typing and random amplified

polymorphic DNA analysis are based on low-stringency PCR

amplification of genomic DNA with arbitrary sequence primer. Segments

of DNA lying between closely spaced annealing sites are amplified to

produce a strain-specific array of DNA fragments (Struelens et al,2004).

This simple and rapid technique can be used for strain typing of bacteria,

fungi and protozoa. AP-PCR typing exhibits variable discriminatory

power according to the number and sequence of arbitrary primers; its

performance is limited by a poor reproducibility and the lack of

consensus rules for interpretation of results. The key to the random

priming is that low annealing temperatures are used (at least initially)

during amplification, allowing imperfect hybridization at multiple

random chromosomal locations to occur and initiate DNA synthesis

(Louie et al .,1996).

Amplification will continue if two of the primers bind in close enough

proximity to one another on the complementary strands to allow synthesis

of the DNA fragment. Although the method is much faster than many of

the other typing methods for nosocomial pathogens, it is much more

susceptible to technical variation than most other methods. Slight

variations in the reaction conditions or reagents can lead to difficulty in

reproducibility of results and to differences in the band patterns

generated. Therefore, trying to make comparisons among potential

outbreak strains can be very problematic. When tightly controlled, AP-

PCR can provide a high level of discrimination, especially when multiple

amplifications with different primers are performed (Samore et

al .,1996).

d- Amplified fragment length polymorphism

Amplified fragment length polymorphism (AFLP) is a typing

method that utilizes a combination of restriction enzyme digestion and

PCR . In the AFLP procedure,( Figure 10) the DNA is digested with two

different restriction endonucleases, usually chosen so that one cuts more

frequently than the other. This restriction strategy generates a large

number of fragments. In order to make the interpretation of the results

more feasible, only a specific subset is used for isolate comparison. The

subset is generated by linking adapter sequences to the ends of the

restriction fragments extending the length of the known end sequences.

PCR primers are designed to hybridize to the adapter sequence, the

remaining restriction site sequence, and an additional one or two

nucleotides of the unknown template sequence (Neeleman et al .,2004).

The addition of each nucleotide, chosen at random, to the end of the

primer reduces the number of fragments that will be amplified by a factor

of four. Following PCR, the reaction products are separated by gel

electrophoresis and their banding patterns are resolved. ( Szczuka and

Kaznowski , 2004& Whatmore et al .,2005).

Figure 10 . Schematic diagram of the amplified fragment length

polymorphism (AFLP) technique. (Jiang et al ., 2000) .

e- Restriction fragment length polymorphisms

As alternative, several PCR-based typing methods have been

developed. In PCR-RFLP typing, a polymorphic DNA target sequence is

PCR-amplified and digested with restriction endonucleases, separated by

electrophoresis, and isolates are compared by RFLP pattern(Figure 11).

This simple technique is well reproducible, with moderate discrimination.

Genome RFLP analysis can use different types of probes, including

ribosomal RNA or cloned ribosomal DNA sequences. The latter

application, called ribotyping, is the most universal RFLP typing strategy

(Leeuwen et al .,2002).

Figure 11 . Schematic diagram of Restriction fragment length polymorphisms (Freeman , 2002) .

f-Repetitive element PCR

Repetitive element PCR (rep-PCR) typing targets spacer fragments

lying between repeat motifs of the genome. It shows moderate

discriminatory power but a better reproducibility than AP-PCR analysis.

These PCR typing methods are often used as a first-pass method to assess

the clonality of organisms during outbreak investigations. This rep-PCR

typing is based on the presence of multiple copies of short repetitive

sequences found in microbial genomes. These sequences are interspersed

throughout the genome and are usually located in non-coding regions of

DNA (Leeuwen et al .,2002) .

The number of repetitive elements and their respective genomic

locations are used to genotype isolates and to differentiate highly related

strains. In rep-PCR, primers are designed to amplify regions between

repetitive sequences, resulting in products of various lengths and

sequences. Electrophoretic resolution of these fragments generates a

profile, or “fingerprint,” that contains multiple bands of different sizes.

The profiles are compared to a library database, and cluster analysis is

performed to identify matching or similar patterns. (Clarridge,2004).

g- Multi Locus Variable copy Numbers of Tandem Repeats Analysis

Multi Locus Variable copy Numbers of Tandem Repeats (VNTR)

Analysis (MLVA) is a method for high -resolution typing of microbial

isolates based upon VNTR) . MLVA is based on the detection of short

sequence repeats that vary in copy number in the microbial genome at

various loci. MLVA detects polymorphisms at five different sites in the

genome. Four regions of detection are on the bacterial chromosome and

one is located on the serotype specific plasmid . MLVA has high

discriminatory power within clonal species and appears to be more rapid

and more amenable to standardization than pulse-field gel electrophoresis

for both surveillance and outbreak investigations (Torpdahl et al .,2007) .

8 -DNA arrays

Conventional DNA micro arrays consist of nucleic acid probes

deposited on a planar glass surface. The surface is usually coated with

chemically reactive groups (epoxy, poly-L-Lysine or aldehyde) to ensure

efficient binding of nucleotidic probes on the surface. To assess the

presence of target genes, nucleic acid samples are labeled, either

chemically or by an enzymatic reaction. Labeled samples are then

hybridized onto the array, and washed using different stringency buffers.

The remaining signal resulting from specific interactions between probes

and target nucleic acids is measured using a confocal micro array scanner.

Only probes hybridized to a labeled target will yield signal, thus revealing

the presence of the cognate nucleic acid motif in the sample (Wang et al .

, 2003).

High-density DNA micro arrays have been used in a broad variety of

applications such as transcriptomics, comparative genome hybridization

(CGH), resequencing, drug discovery, microbial community

characterization or single nucleotide polymorphism (SNP) analysis. The

field of application depends primarily on the strategy and the marker

genes used to design the probes. A variety of genes (virulence factors,

phylogenetic markers, antibiotic resistance genes, etc.) have been

employed on microbial diagnostic microarrays, depending on the

question raised by the researcher. Microarrays have thus been

instrumental in the detection of known pathogens as well as in the

discovery of novel infectious agents, such as severe acute respiratory

distress syndrome (SARS) (Lin et al ., 2007).

Comparative genome hybridization, called comparative phylo-

genomics by some, is the technology used to assess differences among

bacterial strains by hybridizing labeled genomic DNA fragments to

synthetic DNA arrays. The components of these arrays share homology

with all of the genes identified to be present in the organism on the basis

of the species’ genome sequence . Binary probe typing systems detect

polymorphism in the bacterial genome by solid-phase hybridisation of

total DNA with a panel of sequence-variant specific probes immobilized

on a solid substrate . This highly reproducible method is currently

developed on DNA micro-array technology (Figure 12). These

approaches are suitable to combine genotyping with resistance and

virulence gene profiling, thereby providing increased information for the

epidemiologic monitoring of pathogens. ( Stabler , 2006).

These methods identify the genome complements of strains for

which no genome sequence is available . The outcome of such analyses

can lead to the development of high-throughput and sensitive typing

methods by selecting for only those genes that show differential absence

or presence in a black-and-white strain-specific fashion . For example,

recent studies with S. aureus revealed that approximately 20% of the

staphylococcal genome consists of core-variable and variable sequences

Obviously, the later category of nucleic acid sequences is best suited for

identification of potentially useful typing targets (Lindsay ,2006).

Figure 12 . Schematic diagram of DNA arrays (Affymetrix .,2002) .

9 -Pyrosequencing

Pyrosequencing is an alternative sequencing method that generates

short sequence read lengths of less than 200 base pairs. It does not require

labeled nucleotides, capillary electrophoresis, or post-reaction

purification. In pyrosequencing, or sequencing by synthesis, the sequence

is read as the nucleotides are incorporated. The chemistry differs from

modified Sanger sequencing in that it uses a combination of enzymes,

including DNA polymerase, ATP sulfurylase, luciferase, and apyrase,

along with adenosine 5' phosphosulfate and luciferin substrates. The

polymerase incorporates nucleotides (deoxynucleoside triphosphates

[dNTPs]) as they are added one at a time to extend the complementary

strand from the primer sequence. Incorporation of the nucleotide is

accompanied by eqimolar release of pyrophosphate, which is

subsequently converted to ATP by ATP sulfurylase (Ronaghi , 2001) .

ATP is used to drive the luciferase mediated conversion of luciferin

to oxyluciferin. This reaction produces light, and a charge-coupled-device

camera detects the chemiluminescent signal. The sequence data are

interpreted as a peak in the pyrogram, and the peak height is proportional

to the number of incorporated nucleotides. Unincorporated nucleotides

and ATP are degraded by apyrase, and the entire process begins again

with the addition of a different dNTP. Today, with increased automation

and decreasing costs of DNA sequencing technology, determination of

nucleotide sequence is emerging as the reference method for comparing

microbial and virus types based on localised genomic polymorphism. It is

the most accurate and informative method. Moreover, sequence data are

completely portable between laboratories. For typing bacterial pathogens

of nosocomial importance, several methods are used (Tavanti et

al .,2003) .

10 -Spectroscopy

Various types of mass spectrometry (MS) are being developed to

facilitate bacterial genotyping. MS was initially focused toward species

identification, also in complex genera, such as the staphylococci .The

mass spectra of bacterial whole cells can be measured, compared, and put

into frameworks of molecular diversity. Alternatively, sequence-variable

PCR products or RNA transcripts there of can be analyzed by MS, which

can be very helpful in the generation of strain-specific MS fingerprints

(Carbonnelle , 2007).

Using this technique,( Figure 13 ) pure bacterial isolates are mixed

with a chemical matrix, spotted onto a metal plate, and then ionized by a

laser. Ionized molecules corresponding to fragmented bacterial peptides

are released from the matrix (desorption) and move in an electric field

toward a detector. The “time of flight“ corresponds to the time required

for charged ions to reach the detector and depends on the mass to charge

(m/z) ratio of the individual molecules. The resulting signal profiles of

unknown samples provide proteomic patterns, or “spectral fingerprints,”

that can be used for identification when compared to a database. The

spectra are specific to genera, species, or even strains of bacteria.

Although the spectral profiles correspond to peptides derived from

multiple bacterial proteins, the majority of signals are due to highly

abundant proteins, such as ribosomes (Sauer and Kliem , 2010 ).

Figure 13. Overview of Raman procedure and spectrometer. Biomass

from a bacterial culture (a) on Trypticase soy agar (TSA) medium is

collected using a inoculation loop and suspended in of demineralized

water (b). After a brief centrifugation step to remove air bubbles, the wet

pellet is transferred onto a fused silica slide (c), where it is allowed to dry

(a typical slide holds 24 samples). The slide with the dried biomass is

placed in the measurement stage (d), where the samples are illuminated

with laser light (e). The Raman signal generated is collected along the

same optical path and separated from the laser light using an optical filter

(f) that only reflects light of a higher wavelength than the laser. The laser

light is passed through. The wavelength of the Raman signal is dispersed

on an optical grating (g) and collected using a near-infrared-optimized

charge-coupled device detector (h). The Raman spectra are gathered,

stored, and analyzed on a personal computer (i) (Scholtes-Timmerman et

al .,2009).

11 -Proteomics and metabolomics

Proteomics combines all technologies that can be used for protein

separation and molecular characterization. Large numbers of individual

proteins expressed in living organisms can thus be identified down to the

amino acid sequence; such panels of proteins generate strains pecific

polypeptide fingerprints. Clinical proteomics is a branch of classical

proteomics focusing on the identification of clinically relevant protein

expression . Major efforts to standardize this technology are ongoing,

which may be instrumental in the introduction of proteomics in bacterial

typing (Mischak , 2007 ).

Metabolites are the intermediate or final products of chemical

reactions occurring in living creatures. The full spectrum of usually

small-molecular size metabolites in a biological system is called the

metabolome, and the discipline studying the full composition of the

metabolome is called metabolomics. This discipline assesses the

intracellular complexity of compounds, such as amino acids, glucosides,

hormones, and others. Since there is extensive variability in the intra-

cellular presence and concentration of small molecules, the specific

composition of a strain’s metabolites when grown under defined

conditions can be highly characteristic (Raman ,2005).

12- Nucleotide sequence- based analysis

* Single-locus sequence typing

Sequence data for specific loci (genes for virulence, pathogenicity,

drug resistance, etc.) from different strains of the same species have

revealed variability in a specific gene, such as single-nucleotide

polymorphisms and areas with repetitive sequence that demonstrate

potential for epidemiologic application (Koreen et al .,2004). At present,

the single-locus sequence typing (SLST) approach with most promise

involves analysis of a particular region of the staphylococcal protein A

gene (spa) which is polymorphic due to 24-bp repeat sequences that may

vary in both the number of repeats and the overall sequence in the

polymorphic X or short sequence repeat region . Although it is applicable

only to S. aureus, spa typing appears to be very robust, with benefits in

throughput, ease of use, and interpretation that tend to balance a lower

level of epidemiologic discrimination than that of established genotypic

methods such as PFGE ( Belkum et al .,2006).

* Multi-locus sequence typing

Multi-locus sequence typing (MLST) is a powerful molecular tool

used for characterizing relationships among bacterial isolates for

epidemiological purposes. MLST utilizes sequence analysis of internal

fragments (~500 bp) for six or seven housekeeping genes. Multiple genes

are used because each gene alone does not provide adequate

discriminatory power . Gene targets selected for MLST analysis should

not be under selective evolutionary pressure (i.e., virulence genes),

present in multiple copies, subject to recombination, or closely linked

with other genes (Gevers , 2005).

When designing primers for MLST studies, it is important to note

that there are no sets of primers that are universally applicable to all

bacteria, and alternative sets of primers may need to be designed for each

gene, since there may be sequence variation at the primer-annealing site

among different strains of the same species (Maiden, 2006).

Although MLST generates an important frame work for the

assessment of microbial (non)clonality and ecological spread

(Ogden ,2007). Derived, in principle, from multilocus enzyme

electrophoresis, MLST utilizes a larger, and potentially more

representative, portion of the genome than SLST. MLST compares the

nucleotide sequences of internal 400- to 500-bp regions of a series of

housekeeping genes (typically seven or more) which are present in all

isolates of a particular species. For each gene fragment, genetic

polymorphisms in sequences are considered distinct alleles. Each isolate

is defined by the alleles at each of the sequenced housekeeping loci,

which together comprise the allelic profile or sequence type. Because

there are many potential alleles at each of the loci, it is unlikely that

identical allelic profiles will occur by chance. Thus, isolates with the

same allelic profile are assigned as members of the same clone (Lemee et

al .,2004).

MLST was originally employed to identify hypervirulent lineages of

Neisseria meningitides .However, the approach has now been applied to a

variety of other pathogens, including S. aureus and enterococci , for the

assignment of S. pneumoniae strains to major hypervirulent, penicillin-

resistant, and multiple-antibiotic resistant clones and to a large number of

other organisms (Stampone et al .,2005).

Infection control Guidelines

Healthcare-associated infection affects hundreds of millions of

people world wide and is a major global issue for patient safety. It

complicates between 5 and 10% of admissions in acute care hospitals in

industrialized countries. In developing countries, the risk is two to twenty

times higher and the proportion of infected patients frequently exceeds

25% . IC activities are still developing in many health institutions in

Egypt. The national infection control program was started in 2003 by the

Ministry of Health and Population. The national IC strategic plan entailed

instituting IC programs in all hospitals in Egypt by 2010 (Yassin etal .,

2003).

The structure and components of an infection control program are

shown in Tables 1, 2 and 3 . Several authors have discussed the

components of an infection control program in the Long-term care

facilities (LTCF) . Most authors feel that an infection control program

should include some form of surveillance for infections, an epidemic

control program, education of employees in infection control methods,

policy and procedure formation and review, an employee health program,

a resident health program, and monitoring of resident care practices. The

program also may be involved in quality improvement, patient safety,

environmental review, antibiotic monitoring, product review and

evaluation, resident safety, prepareness planning, and reporting of

diseases to public health authorities (Ouslander et al.,2006) .

Table 1 ; Infection control program structure .

(ICP) Infection control professional . (ICC) infection control committee

(Philip et al ., 2008).

Table 2 ; Infection control program element .

(LTCFs) Long-term care facilities . (PI) Professionals in Infection

Control (Philip et al ., 2008).

Table 3 ; Non- pharmacological hospital infection control strategies

(Curtis et al ., 2008)

1-Hand hygiene

The extreme importance of hand washing has been known since at

least 1847, when Dr Ignaz Semmelweis discovered that washing hands

before performing obstetric exams on pregnant women reduced

childbirth-related infectious mortality from more than 10% to less than

1% (Bjerke ,2004). However, rates of hand washing among healthcare

providers usually range from only about 20 to 50% per hospital patient

encounter, although some studies have reported hand-washing rates as

high as 81%. Viable pathogens are often found on hands of healthcare

providers (Rosenthal et al .,2005).

The skin is an inhospitable environment for most micro-organism as

it is dry , acidic and poor in nutrients. However , some micro-organism

have adapted to these conditions and exist in stable population known as

the resident or normal flora . These organism live in deep crevices in the

skin , in hair follicle and sebaceous glands . The micro-organism present

in largest numbers are Gram –positive bacteria , mainly coagulase

negative staphylococci, micrococci and coryneform . Viruses are also

easily acquired (Sattar et al .,2002).

The ‘‘My Five Moments for Hand Hygiene’’ program included in the

WHO guidelines document deserves emphasis because of its success rate

in 400 hospitals worldwide in 2006 to 2008. The evidence-based program

is easy to follow and reminds the health care worker to practice good

hand hygiene:

1. before touching a patient

2. before a clean/aseptic procedure

3. after body fluid exposure risk

4. after touching a patient

5. after touching patient surroundings .

Indications for hand washing and hand antisepsis:

* When hands are visibly dirty or contaminated with proteinaceous

material or are visibly soiled with blood or other body fluids, wash hands

with either a non antimicrobial soap and water or an antimicrobial soap

and water.

- Proper hand washing technique is essential for best results:

Wet hands with water

Apply enough soap to cover all surfaces

Rub hands palm to palm

Right palm over left dorsum with interlaced finger and vice versa

Palm to palm with fingers interlaced

Backs of fingers to opposing palms with fingers interlocked

Rotational rubbing of left thumb clasped in right palm and vice

versa

Rotational rubbing, backwards and forwards with clasped fingers

of right hand in left palm and vice versa

Rinse hands with water

Dry thoroughly with a single-use towel

Use towel to turn off faucet/tap

Duration of entire procedure: 40 to 60 sec and your hands are safe.

- Paper towels, warm air dryers, and cloth towels were no different in the

efficiency to dry wet hands.

- When using towels, pat dry instead of rubbing dry to avoid cracking

(Lederer et al .,2009) .

* If hands are not visibly soiled, use an alcohol-based hand rub for

routinely decontaminating hands or wash hands with an antimicrobial

soap and water.

- Proper hand rubbing with alcohol-based hand rub is:

Apply a palm full of the product in a cupped hand and cover all

surfaces

Rub hands palm to palm

Right palm over left dorsum with interlaced finger and vice versa

Palm to palm with fingers interlaced

Backs of fingers to opposing palms with fingers interlocked

Rotational rubbing of left thumb clasped in right palm and vice

versa

Rotational rubbing, backwards and forwards with clasped fingers

of right hand in left palm and vice versa

Duration of the entire procedure: 20 to 30 sec and, once dry, your

hands are safe (figure 14) (WHO , 2009) .

It is estimated that hand washing with plain soap for 30 second (s)

removes most soil and dirt, eliminates about 90% of transient hand flora

but a low percentage of resident hand flora. Hand washing for 15 s with a

soap containing chlorhexidine or triclosan removes most soil and dirt and

about 99.9% of transient flora and about 50% of resident flora. Hand

rubbing for 15 s with an alcohol-based gel does not remove soil or dirt,

but kills about 99.9% of transient flora and about 99% of resident flora

(Simon ,2004).

Alcohol-based hand-washing solutions are generally considered to be

more effective than soap and water. Compared with plain soap and water,

some studies have reported significantly lower rates of nosocomial

infections when alcohol-based solutions or chlorhexidine- or triclosan-

based hand-washing agents are used (Brown et al .,2003).

Many healthcare providers prefer using alcohol based solutions

instead of soap and water, and compliance rates are generally higher

when alcohol- based hand-washing solutions are used. Use of alcohol-

based cleaners saves time and these generally abrade and irritate the skin

less than antiseptic soaps. However, some people complain that alcohol-

based cleaners dry out and crack their skin. Hospitals and healthcare

providers may want to experiment with several alcohol or chlorhexidine-

based hand cleaners. Soap and water may still have to be used in cases

when hands are visibly soiled. In that case, staff and visitors should wash

hands carefully for at least 15 s with soap and water (Widmer ,2000).

Gloves are used to prevent contamination of healthcare personnel

hands when;

1(anticipating direct contact with blood or body fluids, mucous

membranes, non intact skin and other potentially infectious material.

2 (having direct contact with patients who are colonized or infected with

pathogens transmitted by the contact route e.g., vancomycin-resistant

enterococci (VRE), meticillin-resistant Staphylococcus aureus

) MRSA ,(Respiratory Syncential Virus (RSV) .

3(handling or touching visibly or potentially contaminated patient care

equipment and environmental surfaces(CDC ,2002) .

Gloves can protect both patients and healthcare personnel from

exposure to infectious material that may be carried on hands . The extent

to which gloves will protect healthcare personnel from transmission of

blood borne pathogens (e.g., HIV, HBV, HCV) following a needle stick or

other pucture that penetrates the glove barrier has not been determined.

Although gloves may reduce the volume of blood on the external surface

of a sharp by 46 to 86% , the residual blood in the lumen of a hollow bore

needle would not be affected; therefore, the effect on transmission risk is

unknown (Duckro et al .,2005) .

It is not certain what type of glove provides the best protection for

infection control. Some studies have suggested that latex gloves are

somewhat better in preventing penetration of water and virus than vinyl

gloves. However, about 3 to 16% of healthcare workers are sensitive to

latex and sometimes experience severe respiratory reactions to it. If latex

gloves are used in the healthcare setting, only the powder-free gloves

should be used since these release much lower levels of latex allergens

than the powdered latex gloves. Nitrile gloves also have good barrier

penetration but are more expensive and heavier than either latex or vinyl

gloves (Yip ,2004) .

Shoe and head covers are often recommended for use in areas

containing immunocompromised or surgical patients. Although bacterial

pathogens have been collected from shoes, research on the use of shoe

covers and/or separate hospital shoes and spread of pathogens has been

meager (Santos et al ., 2005) .

Proper cleaning techniques and proper cleaning chemicals can also

significantly reduce hospital pathogen levels and risk of nosocomial

infections . Terminal room cleaning after patient discharge was able to

adequately clean only a mean of 49% of the standardised surfaces,

including less than 30% for toilet hand holds, bedpan cleaners, room door

knobs and bathroom light switches. It is recommended that hospitals

monitor performance of cleaning personnel and provide feedback and

training as needed to optimise cleaning effectiveness (Carling et

al .,2008).

Figure 14 . Schematic diagram of hand wahing (WHO , 2007).

2-Environmental Decontamination &Cleaning

Greater awareness of the hospital environment as a source of

nosocomial pathogens has led to calls for enhanced investment in more

effective conventional cleaning, as well as encouraging the development

of a range of new cleaning and decontamination technologies(Casey et

al .,2010) . Sites that are frequently touched by hands are thought to

provide the greatest risk for patients, and those situated right beside

patients provide the biggest risk of all. The responsibility for cleaning

near-patient hand-touch sites does not always rest with the ward cleaners

(White et al .,2008) .

The microbial pathogens that cause HAI have two special properties:

first, they are recognized as hospital pathogens; second, they have an

innate ability to survive on surfaces in the hospital environment for long

periods of time . They include organisms such as MRSA, Clostridium

difficile, VRE , Acinetobacter spp. and norovirus (Dancer , 2008).

The potential for contaminated environmental surfaces to contribute

to transmission of healthcare-associated pathogens depends on a number

of factors, including the ability of pathogens to remain viable on a variety

of dry environmental surfaces, the frequency with which they

contaminate surfaces commonly touched by patients and healthcare

workers, and whether or not levels of contamination are sufficiently high

to result in transmission to patients . Any isolation of pathogens, or

pathogen indicators, causes concern and warrants immediate action. By

contrast, environmental surface sampling in hospitals usually only takes

place in response to an outbreak and then only if the infection control

team responsible has the motive, means and interest to initiate

environmental screening (Dancer , 2009) .

Disinfection is one of the cornerstones of infection prevention and

control, defined as the antimicrobial reduction of micro-organisms to a

level specified as appropriate. This definition is intentionally broad to

cover a variety of applications, including medical/ surgical device

reprocessing, liquid/gas treatment and general environmental surface

disinfection . It encompasses other processes, such as pasteurization,

sanitization, antisepsis, fumigation and preservation. Disinfection

methods can be classified as being physical or chemical in antimicrobial

activity . Physical methods include radiation and heat, while chemical

methods are based on the use of biocides such as alcohols, aldehydes,

halogens and quaternary ammonium compounds (McDonnell and

Burke ., 2011) .

Given the range of disinfection methods available and their clinical

applications, classification systems are used to aid healthcare workers to

choose the correct method to safely reduce patient risks. One such system

is the Spaulding classification for surgical or medical devices, which has

been in use since 1957.Spaulding defined the minimum levels of

disinfection to be employed according to the infection risk associated

with a device when used with a patient. Critical devices present the

highest risk as they enter a normally ‘sterile’ area of the body, such as the

bloodstream. Sterilization of these devices is recommended . Sterilization

is distinct from but encompasses disinfection, being defined as a process

used to render a surface or product free from viable micro-organisms,

including bacterial spores. A sterile device is free from viable organisms,

while disinfected devices or surfaces can only be presumed to have

reduced microbial levels. Typical sterilization processes use steam,

ethylene oxide, liquid peracetic acid and hydrogen peroxide gas (Song et

al .,2009) .

Semi-critical devices pose a lower risk as they may only contact

mucous membranes or non-intact (broken) skin. In the past, many of

these devices (such as flexible endoscopes) could not be sterilized in a

reasonable time frame for practical clinical use. The compromise was to

recommend high-level disinfection, thereby inactivating most pathogenic

micro-organisms such as viruses, bacteria (including mycobacteria),

fungi and, if possible, bacterial spores (in these cases, generally requiring

longer exposure times). High-level disinfectants, such as those based on

heat (hot water for some devices), glutaraldehyde, ortho-phthaldehyde ,

hydrogen peroxide and peracetic acid, could provide rapid turn around

times for these devices (Antonnucci et al .,2008) .

Non-critical devices present the lowest risk to patients, as they may

only contact intact skin. In these cases, low- or intermediate level

disinfection is often recommended, encompassing certain types of viruses

[especially enveloped viruses such as influenza and human

immunodeficiency virus (HIV)], most bacteria and some fungi.

Intermediate-level disinfectants should also provide efficacy against a

broader group of viruses (non-enveloped) and some mycobacteria.

Examples include alcohol-, aldehyde-, phenolic- and quaternary

ammonium- compound-based disinfectants (Beguma et al .,2009) .

* Laboratory-acquired bacterial infection

Laboratory-acquired bacterial infection is a documented occupational

hazard for staff working in microbiology laboratories. Bacterial

contamination of door handles, telephones and computer keyboards has

been demonstrated in the clinical setting but there are few comparable

data for microbiology laboratories. Two studies documented the presence

of environmental contamination with vancomycin-resistant enterococci

but did not look at hand acquisition ( Ng et al ., 2011).

Other than enterococci , staphylococci are common laboratory

isolates and documented to survive well on environmental surfaces in a

clinical setting; Enterobacteriaceae and P. aeruginosa also represent

potential bacterial pathogens. Further indirect evidence of laboratory

environmental contamination with bacteria comes from studies reporting

evidence of cross-contamination of specimens with enterococci and

Salmonella spp. It is demonstrated that the density of commensal bacteria

on the hands of laboratory workers was increased by the use of jewellery

or wrist watches (Dancer ,2008& Lappe et al .,2009) .

Some studies demonstrate that the use of gloves is protective against

hand acquisition of MRSA by laboratory technicians. MRSA is the

commonest bacterial pathogen isolated from laboratory surfaces,

particularly from frequently handled surfaces such as telephone keypads

and computer keyboards. Hand washing at the end of work sessions was

effective at removing pathogenic bacteria (Lappe et al .,2009) .

The primary method of recovery of bacteria from surfaces was the

swab-elution method without enrichment, and this method is subject to

various limitations, including the underestimation of bacteria

density .Some studies prove to perform enrichment of environmental

swabs so that an estimation of relative bacterial density from different

surfaces could be performed. Similarly, it would not have detected

contamination of hands with low numbers of pathogenic bacteria, as the

lower limit of detection was 200 cfu for each tested hand (Landers et

al .,2010) .

* Air filtration and air handling

High-Efficiency Particulate Air (HEPA) filtration is relatively

inexpensive and probably should be used for all hospital rooms. Various

studies have found that the HEPA filtration in hospitals can significantly

reduce airborne levels and/or infection rates for several aerosolized

pathogens. Many studies have reported that the HEPA filters in patient

rooms can significantly reduce both airborne aspergillus concentrations

and rates of human aspergillus infections .Use of HEPA filters has been

found to significantly reduce airborne levels of MRSA and P. aeruginosa

in hospitals and reduce airborne concentrations of droplet nuclei (which

transport tuberculosis) by 90% (Boswell et al .,2006).

Hospital air filtration system filters air at 60 air changes per hour and

uses a ‘cold plasma’ system to destroy microbes. Early tests have

indicated that such a system has a more than 99% single-pass efficiency

in destroying bacteria, viruses and moulds such as Aspergillus (Poirot et

al .,2007). UltraViolet (UV) light machines in rooms or in ventilation

systems can effectively kill mycobacteria, legionella and many viruses,

but UV light is not effective in killing many species of bacteria and

moulds (Leung and Chan , 2006).

3-Nutrition

Better nutrition can also play a critical role in reducing nosocomial

infections. Malnutrition is very common in hospitalised patients.

Malnutrition was measured by such parameters as weight loss ( body

mass index, grip strength) respiratory function, nutritional intake and

blood levels of albumin, and prealbumin. Many nutrients play a key role

in maintaining immunity including protein, omega-3 fatty acids, vitamins

A, B6, B12, C, D, and E; selenium, zinc, copper and iron. Most of these

nutrients become depleted following acute illness. Malnutrition is a major

risk factor for infection (Wintergerst et al .,2007) .

The liquid nutrients provide a favourable medium in which bacteria

can grow and are easily contaminated during assembly and manipulation

of the administration sets . Many types of bacteria ,including

Salmonella ,Klebsiella , Enterobacter , E.coli and S.aureus have been

found in high concentrations in enteral feeds . These may cause

gastroenteritis and, through colonization of the gut , may also result in

septicaemia and pneumonia (Howell , 2002) .

Manipulation of enteral feeding systems increases the risk of feed

contamination . Administration sets should be designed to require

minimal manipulation and recessed connection may help to reduce the

risk of contamination . Hands must be washed before handling enteral

feeds or the feed administration systems and a rigorous no-touch

technique must be used when assembling the administration sets and

handling the feed . The use of clean ,disposale gloves has been

recommended to minimize the risk of contamination (pellowe et

al .,2003).

Sterile ready-to-use feeds should be administered over a maximum

of 24h . The feeding tube should be flushed with fresh tape water before

and after aspiration , feed changing or drug administration to minimize

the risk of micro-organism adhering to the internal surface and the

administration reservoir and tubing should discarded after a maximum of

24 h in use (Howell, 2002) .

4-Surveillance & Outbreak control

Infection surveillance may either include all residents in a facility or

be targeted at specific subpopulations. Although facility-wide

surveillance is useful for calculating baseline rates and detecting

outbreaks, a more focused analysis could include examination of

infection rates in residents who are at risk for certain kinds of infection

(such as aspiration pneumonia in residents receiving tube feedings or

bloodstream infection among residents with indwelling vascular

catheters). These surveillance data are used primarily to guide control

activities, to plan educational programs, and to detect epidemics, but

surveillance also may detect infections that require therapeutic action

(McGeer et al .,1991).

Surveillance requires objective, valid definitions of infections. Most

hospital surveillance definitions are based on the National Nosocomial

Infections Surveillance System (NNIS) criteria, Prevalence studies detect

the number of existing (old and new) cases in a population at a given

time, whereas incidence studies find new cases during a defined time

period. The latter is preferred because more concurrent information can

be collected by an incidence study if data are collected with regularity

(Satterfield ,1993).

Surveillance systems must be extremely flexible; effective infection

control teams will not use a “one size fits all” approach for surveillance.

Some hospitals should focus on patients at high risk, such as those

hospitalized in intensive care and neonatal units (Pottinger et al .,1997).

After defining the priorities of the institution, the focus could also be on

specific problems, such as bacteremia or surgical site infection. Focusing

on neonatal bacteremia pays high because extrinsic contamination of IV

fluids seems to be a common problem in many settings (Avila-Figueroa

et al .,2000).

Automated surveillance (AS) is the process of obtaining useful

information from infection control data through the systematic

application of medical informatics and computer science technologies.

(Wright , 2008) .

* Surveillance Methods

1- Case-finding Issues

First, should infections be sought by passive or active means . In

passive surveillance, persons who do not have a primary surveillance

role, that is, persons other than ICPs , are relied on for identification and

reporting of infections. Active surveillance is the process of vigorously

looking for nosocomial infections using trained personnel, nearly always

ICPs. ICPs seek out nosocomial infections by using various data sources

to accumulate information and decide whether or not a nosocomial

infection has occurred (Bates et al ., 2003) .

Second, should infection detection be patient- or laboratory-based .

Patient-based surveillance includes counting nosocomial infections,

assessing risk factors, and monitoring patient care procedures and

practices for adherence to infection control principles. It requires ward

rounds and discussions with caregivers. In laboratory-based surveillance,

detection is based solely on the findings of laboratory studies of clinical

specimens. Third, should infections be detected prospectively or

retrospectively . Prospective surveillance refers to monitoring patients

while they are still hospitalized and, for SSIs, includes the postdischarge

period. Retrospective surveillance uses chart review after patient

discharge as the sole means of identifying infections (Burke , 2004).

2- Incidence Versus Prevalence in Hospital-Wide Surveillance

Incidence surveillance is continual monitoring of all patients for

new nosocomial infections of all kinds on all wards. It has also been

termed ongoing, total, housewide, or comprehensive surveillance .

Hospital-wide surveillance has the advantage of providing a global view

of what is happening in the hospital so that potential clusters of infection

or antibiotic resistance can be detected anywhere . The advantage of

prevalence surveillance is that it is a rapid inexpensive way to estimate

the magnitude of nosocomial infection problems in a hospital . There are

two major disadvantages of prevalence surveillance. First, in small

hospitals, the number of patients surveyed is insufficient to detect

important differences among patient populations . Second, patients' risk

of infection is overestimated with the prevalence rate, which is calculated

as the number of active infections on the day of the visit divided by the

number of beds visited (Gaynes et al ., 2001) .

3- Targeted Surveillance

These strategies focused or targeted efforts on certain areas in the

hospitals (e.g., ICUs), patient groups (e.g., surgical patients), or infection

sites (e.g., bloodstream infections). These targeted efforts have become

increasingly common in this decade not only because of their positive

impact on resource management but because they have the potential for

yielding more meaningful results than hospital-wide surveillance. A

disadvantage of these limited strategies is that clusters of infection in

areas not under surveillance may be missed ( Emori et al ., 1998).

4- Objective/Priority-Directed Surveillance

Accordingly, SSIs and pneumonias would be allocated the most

surveillance resources (one half and one third, respectively), with much

less for bloodstream and urinary tract infections. Objectives for

surveillance should be evaluated annually and adjusted as necessary. The

obvious advantage of this method is that specific measurable objectives

are set and attainment is carefully evaluated. Therefore, ICP time and

effort are directed in a very productive manner. A potential disadvantage

is undetected outbreaks, although some studies recommended that the

ICP train other hospital staff to be alert for and report unusual clustering

(Leape , 2002).

5- Limited Periodic Surveillance

This method is a combination of hospital-wide and site-specific

targeted surveillance. Some studies used total surveillance for 1 month

per quarter and targeted bloodstream infection surveillance during the

other 8 months. Although the potential for missing clusters is less than for

targeted methods, it still exists during two thirds of the year (Platt ,

2002).

6- Postdischarge Surveillance

Because of the shorter postoperative stay, it is estimated that as

many as 50% of SSIs may be missed if a formal postdischarge

surveillance system is not in place . Significant methodologic problems

with postdischarge surveillance include reliance on physicians to return

information on patients to the ICP in a timely manner, patients' inability

to accurately diagnose infection , and determination of how to handle

patients lost to follow-up (Sands et al ., 1999) .

* Outbreak control

In fact, we are living in a worldwide pandemic in many hospitals

where sophisticated protocols are performed with no attention to the

risks and no control programs; under such conditions, a substantial

proportion of NIs occur as outbreaks . Hospitals having outbreaks (Table

4) as main causes of NIs must implement programs focused on their

prevention as a first step towards consolidation of their programs

(Ostrosky-Zeichner et al .,2000).

Table 4 ; Immediate control measures for outbreak management

(Fletcher ,1996).

Type of transmission suspected Suggested action

Cross-transmission (transmission between Patient isolation and barrier

individuals) precautions determined by

infectious agent(s)

Hand transmission Improvements in

Hand washing; cohorting

Airborne agent Patient isolation with

appropriate ventilation

Waterborne agent Checking of water supply

and all liquid containers

Use of disposable devices

Food borne agent Elimination of the food at

Risk

5-Prevention of urinary tract and urinary catheter

infections

About 80 to 95% of hospital-acquired urinary tract infections

originate from urinary catheters. Guide for CAUTIs outlines interventions

for avoiding and/or minimizing the duration of indwelling urinary

catheter use, including the following:

(1) Using catheters only when medically necessary

(2) assessing patients daily for the need for catheterization and

documenting a continued need

(3) using reminder systems for health care workers aimed at removing

catheters

(4) using external catheters in men when feasible

(5) considering intermittent catheterization instead of indwelling catheter

insertion

(6) promptly removing unnecessary urinary catheters. About 15% of

urinary HAIs have been linked to improper hand washing and poor

aseptic techniques in cleaning the urinary meatus area and inserting and

maintaining the urinary catheters (Rebmann and Linda., 2010).

Other interventions aimed at preventing CAUTIs focus on

maintaining a sterile, closed drainage system to prevent microorganism

colonization of the catheter. Examples include ;

(1) using aseptic technique during catheter insertion

(2) allowing only trained health care professionals to insert urinary

catheters

(3) securing catheters to prevent movement and urethral traction

(4) keeping the drainage bag below the level of the bladder

(5) changing the indwelling catheter or urinary drainage bag only when

necessary (ie, do not routinely change the catheter or drainage system at

arbitrary intervals)

(6) Other recommended measures include avoiding irrigation unless the

catheter is obstructed

(7) scanning the bladder for residual urine amounts

(8) administering anti infective therapy only when an infection is

suspected rather than after colonization ( CDC .,2009).

Bacteria enter the drainage system in the drainage bag or at the

junction between the catheter and the drainage bag . These bacteria reach

the bladder along the tubing after a few days . The drainage system

should not be opened to take specimens which should instead be obtained

aseptically from sampling port with a needle . The drainage bag should be

emptied when necessary to avoid reflux of urine . A clean pair of gloves

should be worn for emptying the drainage bag and discarded on

completion of the procedure . When the bag is emptied ,care should be

taken to ensure that micro-organism are not introduced on to the tap by

contact with a contaminated container or other surface . Containers

should be decontaminated in a bedpan washer or autoclaved after each

use (Pratt et al ., 2001).

Another CAUTI prevention intervention is the use of antimicrobial-

or silver-coated catheters. The use silver-tipped catheters (both short-

and long-term users) was associated with a 57% reduction in urinary tract

infections (Rupp et al .,2004).

6-Prevention of central venous line and Blood stream

infections

The prevalence of Catheter Related Blood stream infections

( CRBSI) increases linearly with the duration of catheterization. Thus, the

continued need for CVCs , and other short-term invasive devices, should

be assessed each day and the devices should be removed as soon as they

are no longer required for patient care (Mermel , 2007).

Use of maximal barrier precautions for CVC insertion (i.e. use of

long-sleeved gown, sterile gloves, mask, cap and large sterile sheet drape)

reduces catheter-related bloodstream infections compared to minimal

(sterile gloves and small drape) precautions and should be the standard of

care. Placement of CVCs in the femoral vein compared with the

subclavian vein increases the risk of CVC colonization , with increased

risk of CRBSI. Femoral CVC placement also increases the risk of

thrombosis compared with subclavian vein insertion . Thus, CVC

placement in femoral veins should be avoided and if such placement is

necessary in an emergent situation, such catheters should be replaced in

another anatomical site as soon as it can be safely done (Rijnders et

al .,2002).

A meta-analysis of prospective, randomized studies has found that

use of chlorhexidine-containing antiseptics for preparing the catheter

insertion site reduces risk of CRBSI compared with povidone iodine and

the former antiseptics should be the standard of care for cleaning catheter

insertion sites.Use of a chlorhexidine containing dressing directly over

the CVC insertion site has been demonstrated to reduce the risk of

catheter colonization in paediatric patients , especially in neonates.

However, this dressing can cause skin irritation in very low birth-weight

neonates. Thus, such dressings should be considered for use over CVC

insertion sites in paediatric patients, apart from very low birth-weight

neonates, and considered for high-risk adult patient populations (Levy et

al .,2005).

Doubling the number of hours per ICU shift in which float nurses

work instead of nurses who regularly staff ICUs leads to a nearly 4-fold

independent increased risk of primary bloodstream infections, most of

which are due to CRBSI (Edwards et al .,2003).

One prospective, randomized trial found that continuous infusion of

heparin (100 units/kg/d) in haematology/oncology patients with a CVC in

place reduced the incidence density of CRBSI compared to control

patients continuously infused with saline .The incidence of CVC-related

deep venous thrombosis was similarly reduced (Abd el kefi et

al .,2005 ) .

Use of vancomycin-containing catheter lock solutions has been shown

to reduce the incidence of CRBSI .Other antimicrobial agents have also

been used in catheter lock solutions to prevent CRBSI. Concerns about

the potential risk of antibiotic resistance have limited the widespread use

of this preventative strategy. Nevertheless, certain high-risk patient

populations, such as patients who require a long term CVC but have

limited vascular access and/or recurrent CRBSI despite other

interventions, are good candidates for such an intervention (Safdar and

Maki , 2006).

Needleless catheter connectors have come into widespread use in

some countries in an effort to reduce sharps injuries . A prospective,

randomized trial found that use of a needleless catheter connector was

independently associated with reduced catheter colonization. Despite

these results, a more recent study found that the incidence of CRBSI

increased more than 3-fold in a pediatric ICU when one type of

needleless catheter connector was substituted by another type.

(Maragakis et al .,2006).

* Safe Disposal Of Sharp Instrument and Waste

Injection drug use remains the single most important vector for the

spread of blood-borne disease in the United States, accounting for

approximately one third of all AIDS cases and 60% of new hepatitis C

infections. Access to sterile syringes helps prevent the spread of blood-

borne infections among injection drug users (IDUs) ( Jarlais et al .,2005

& Talaat et al .,2006).

Clinical wastes predominantly comprise wound dressings and swabs

together with infusion and irrigation equipment, catheters, blades,

syringes and needles. Also included are tissue and postmortem wastes,

waste from clinical laboratories, sanitary wastes including incontinence

pads and nappies etc., and waste pharmaceuticals. Most wastes will be

disposed to yellow bags or rigid yellow bins, including sharps bins where

appropriate, and removed from clinical areas for destruction by high

temperature incineration, or made safe using one of a number of alternate

treatment technologies including autoclave, microwave or hot oil auger

treatments (Table 5) (Rushbrook et al .,1999) .

Table 5 ; Colour coding for the disposal of clinical waste (Health

Services Advisory Committee , 1999 ) .

Colour of bag Type of waste .

Black Household waste ,treated clinical waste

(e.g .paper ,food ,flower ,etc)

Yellow Clinical waste (e.g. material contaminated

with blood or body fluid ,human or

animal tissue )

Yellow sharps container Needle syringes ,broken glass and any

other contaminated sharp item

Blue or transparent with Waste for autoclaving (e.g. pathology

blue inscription specimens)

Yellow black stripes Non-infectious human waste (e.g. sanitary

towels , incontinence pad )

Multihazardous waste includes waste that is infectious and that

contains radionuclides and/or hazardous chemicals. An example is waste

contaminated with blood or body fluids and with a chemotherapy drug.

Multihazardous waste is best managed and treated separately from other

infectious waste . Low-level radioactive infectious waste (e.g. swabs,

syringes for diagnostic or therapeutic use) may be collected in yellow

bags or containers for infectious waste if these are destined for

incineration. It should be noted that mercury thermometers are not

infectious waste, and they should not be classified and managed as such.

All unwanted or broken mercury thermometers should be managed and

disposed of as hazardous chemical waste. They should never be placed in

sharps containers (Denys , 2000) .

7-Preventing respiratory tract infections

Preventive strategies to reduce the colonization of the upper

respiratory tract are the selective decontamination of the digestive tract

(SDD), oropharyngeal decontamination and combinations of these with

or without the use of systemic antibiotics. Oropharyngeal

decontamination can be achieved using topical antibiotics, which may

increase the risk of antibiotic resistance, or using topical antiseptics.

Though decolonization of the oropharynx using topical antimicrobial

agents like chlorhexidine (CHX) should prove useful in the prevention of

nosocomial respiratory tract infection, scientific evidence to recommend

routine oral decolonization is lacking, as several randomized clinical

trials have failed to demonstrate a significant reduction in the incidence

of respiratory tract infections in the CHX group (Fourrier et al .,2005).

Supine patient positioning facilitates aspiration; semirecumbent

positioning decreases it . Infection in patients in the supine position was

associated with the simultaneous administration of enteral nutrition and

an increased risk of aspiration of gastric contents (Tablan et al .,2004).

Gastroesophagal reflux occurs less frequently in the semirecumbent

position. Thus, it is recommended that intubated patients should be

managed in a semirecumbent position, particularly during feeding .

Kinetic beds or continuous lateral rotational therapy is a technique using

a continuous movement of the bed along its longitudinal axis within a

certain range (–40° to +40°) . This movement improves secretion

drainage, and thus may decrease the risk of VAP (Collard et al .,2003 &

Dodek et al .,2004).

The use of endotracheal tubes bypasses the natural defense

mechanisms of the upper respiratory tract and impairs the host’s

capability to fend off infection . Endotracheal tubes also favour the

development of bacterial biofilms that may contribute to the occurrence

of VAP. Noninvasive positive pressure ventilation (NIV) using a face

mask is an alternative to intubation (Esteban et al .,2004). There are two

types of suctioning catheters: open single-use and closed multiuse. There

is no significant difference in the incidence of VAP. A closed suction

catheter changed between each patient is more advantageous in terms of

cost and maintenance (Hubmayr et al .,2002).

The CDC guideline for prevention of pneumonia is oriented toward

acute care hospitals , including respiratory therapy equipment, suctioning

techniques, tracheostomy care, prevention of aspiration with enteral

feedings, and immunizations. Examples of relevant recommendations

include hand hygiene after contact with respiratory secretions, wearing

gloves for suctioning, elevating the head of the bed 30 to 45 degrees

during tube feeding and for at least 1 hour after to decrease aspiration,

and vaccination of high-risk residents with pneumococcal vaccine

( CDC ,1997).

The evidence for the efficacy of pneumococcal vaccine in high-risk

populations, including the elderly population, is debated. However, the

vaccine is safe, relatively inexpensive, and recommended for routine use

in individuals over the age of 65 years (Watson et al .,2002).

Bacterial colonization of condensates in ventilatory circuits plays a

role in the pathogenesis of VAP (Cook et al .,1998). Frequent changes of

ventilator circuits should theoretically lower the risk of initial bacterial

colonization .The upper airways filter, heat and moisten the inspired air

so that it reaches the lower airways at body temperature with added

water. In mechanically ventilated patients the ventilator tube by passes

the upper airways and inspired gases require artificial conditioning by

heating and humdification to prevent mucosal injury and ventilator-

associated pneumonia (VAP) (Kollef et al .,1995).

8-Control of infections related to surgery and surgical

equipment

Surgical site infections (SSIs) are among the most common and

serious complications for patients who undergo operative procedures.

SSIs account for 14% to 17% of all hospital-acquired infections and 38%

of nosocomial infections in surgical patients. The Centers for Disease

Control and Prevention (CDC) (Table 6) estimates that SSIs complicate

approximately 5% of the nearly 30 million surgeries performed each year

(John et al .,2010).

Table 6 ; Major headings in prevention of surgical site infection

(National Institute for Health and Clinical Excellence ,2008) .

Preoperative strategies

About 2 to 5% of all surgical patients develop a significant infection

at the wound site . Higher rates of surgical infections are associated with

operations of two or more hours, a contaminated or dirty procedure, or

inadequate scrubbing procedures (Cheadle ,2006). Preoperative

strategies focus on controlling patient related risk factors and appropriate

hand/forearm antisepsis for surgical team members . Pre-existing

infections at sites remote from the operation site should be identified and

treated, and if practicable elective surgery should be delayed until such

infections have resolved. Obese patients should be encouraged to lose

weight before surgical operations and smokers should be encouraged to

stop smoking (although such lifestyle modifications may be unrealistic

for many patients) (Mangram et al .,1999).

On the night before the operation, the patient can wash or shower

with an antiseptic agent, and immediately before the operation the skin

should be adequately cleaned with an antiseptic solution. However, hair

removal should be avoided unless it is likely to interfere with the

operation. If hair removal is necessary, clippers should be used rather

than shaving, since there is evidence that shaving can result in

microscopic skin cuts that can act as foci for subsequent colonisation and

infection (Tanner et al .,2006).

Short courses of antimicrobial prophylaxis are widely used to reduce

SSI risk. The aim of this approach is not to sterilise tissue, but to reduce

intraoperative contamination to levels where it does not overwhelm the

patient’s defences. Antimicrobial prophylaxis is primarily indicated in

elective procedures in which skin incisions are closed in the operating

theatre. The choice of agent should be based on the pathogens most

commonly associated with the procedure being performed (Mangram et

al .,1999) .

In practice, broad-spectrum beta-lactam agents (particularly

cephalosporins) are most widely used, with an agent such as

metronidazole being added if necessary to provide cover against

anaerobes; vancomycin is not recommended for routine prophylaxis . The

first dose should be timed to ensure that bactericidal concentrations are

achieved in serum and tissue at the time of the incision, and these

concentrations should then be maintained for up to a few hours after

wound closure in the operating theatre. Recently, a statement for

urological surgery incorporated important principles of appropriate

surgical prophylaxis and offered sensible options in terms of the choice of

antibiotics for a variety of urological procedures. (Nicolas et al .,2007).

Perioperative strategies

The CDC guidelines emphasise the importance of good surgical

technique and aseptic precautions for the prevention of SSIs. Good

surgical technique requires attention to the maintenance of haemostasis,

removal of devitalised tissue and foreign bodies as completely as

possible, and elimination of dead space at the surgical site. Gloves,

facemasks, caps, gowns and sterile drapes should be used to minimise

transmission of potential pathogens to the wound. Surgical instruments

should be adequately sterilised according to published guidelines; flash

sterilization should be reserved only for instruments intended for

immediate use . It should be noted that despite precautions such as these,

some contamination of the surgical site is inevitable because some

endogenous bacteria remain even after excellent preoperative preparation

of the site (Owens and Stoessel , 2008 ).

Surgical personnel should undertake a thorough surgical scrub before

donning surgical gowns and gloves. Personnel who are colonised or

infected with potential pathogens should be encouraged to report their

condition, and procedures developed to prevent transmission of

pathogens from colonised personnel to the patient . Hand hygiene is

regarded as one of the key components in any infection prevention

strategy (Humphreys ,2009).

Warming the patient before or during surgery has also been shown to

significantly reduce rates of surgical infection .Warming may reduce

surgical infection rates by improving blood circulation and immune

function in the surgical areas (Melling et al .,2001).

Tissue hypoxia leads to necrosis and is often followed by infection.

High blood glucose levels, such that occur in patients with diabetes

mellitus for example, are also associated with increased risks of infection.

It is logical to assume that normalizing these will assist in the prevention

of SSI, and recent studies have supported this . It is believed that the

mechanism of action is by more effective neutrophil killing of potential

pathogens (Belda et al .,2005).

Postoperative strategies

The risk of SSI can persist for up to 30 days after a surgical operation

or for as long as one year after an operation in which the patient is given

an implant; indeed, a significant proportion (12 to 84%) of SSIs are first

detected after the patient has been discharged from hospital . The CDC

guidelines recommend that incisions that have been closed by primary

intention should be protected by sterile dressings for 24 48 h, and that

personnel should use sterile technique when changing dressings on any

kind of skin incision . However, two recent systematic reviews and meta-

analyses strongly suggest that prophylactic intranasal mupirocin

significantly reduces the rate of postoperative infections including that

caused by MRSA and meticillin-susceptible S. aureus (MSSA) ( Rijen et

al ., 2008) .

One of the major advances in surgical practice in recent decades has

been the development of laparoscopic or minimally invasive surgery.

This offers the potential for reducing infection as the incision site is much

smaller, but it is unclear whether all other practices and the environment

setting associated with open surgery should be replicated (Smyth et

al .,2005). Meta-analysis of laparoscopic compared with open repair of a

perforated peptic ulcer suggests that the risk of postoperative SSI is

reduced when carried out laparoscopically, and similar findings have

been associated with colon surgery (Poon et al .,2009).

9-Prevention of Gastrointestinal Tract and

waterborne hospital infections

A number of interventions have been proven effective in reducing

rates of hospital waterborne infections. Numerous studies have found that

replacing tap water with sterile water for drinking, bathing and

procedures can significantly reduce rates of many hospital infections

including crytosporidium, legionella, aeromonas and stenotrophomonas.

Sterile sponges can be used for bathing. Boiling and water filtration in

hospital water systems can also sterilise water, but these systems need to

be monitored closely because many problems can develop which cause

these systems to fail . Daily cleaning of patient shower areas with a

detergent and phenolic compound has been shown to significantly

decrease airborne levels of moulds including aspergillus (Anaisie et

al .,2002).

Heating water to more than 50 0 C has been shown to significantly

reduce levels of Legionella spp. in storage tanks and hospital water

systems; however, water heating alone will not usually eliminate all

legionella in a contaminated hospital water system. Some studies have

found that the UV-light water treatment can greatly reduce levels of

legionella in hospital water systems (Modol et al .,2007). Copper silver-

based ionisation systems can also significantly reduce waterborne

concentrations of legionella, moulds and Gram-negative bacteria such as

P. aeruginosa and Actinetobacter baumannii. Routine surveillance of

hospital water supplies for legionella is highly recommended in cases of

confirmed legionella infections; however, it is controversial as to whether

such routine testing is needed in hospitals with no legionella infection

history (Huang et al .,2008).

All water leaks and water damage should be repaired and remediated

within 24 h to prevent growth of pathogenic bacteria and moulds.

Hospitals should avoid using indoor decorative fountains since they

encourage legionella and the splashing water facilitates ready

aerosolisation of the organism (O’Neill and Humphreys ,2005).

10-Preventing burn infections

Prevention of burn wound infection involves assessment of the

wound at each dressing change for changes in the character, odor or

amount of wound drainage. Strict aseptic technique should be used when

handling the open wound and dressing materials and frequency of

dressing should be based on wound condition. If the wound has necrotic

material present, a debriding dressing should be chosen, whereas a

protective dressing is preferable for clean healing wounds. Treatment of

an existing wound infection includes considering changes to the topical

agent being used along with changing the frequency of the dressing

changes. In those cases where invasive infection is present, surgical

excision of the infected wound and appropriate systemic antimicrobial

therapy may be required (Weber et al .,2002).

The open burn wound increases the environmental contamination

present around the patient, which is the major difference in burn versus

non burn patients. The larger the burn size the more vulnerable it is to

contamination (Weber et al .,1998).

Patients with larger burn injuries (>25–30% Total Body Surface Area

(TBSA) are also immuno compromised, because of the larger size of their

injury , loss of physical defenses and need for invasive devices . These

patients also represent a significant risk for contamination of their

surrounding environment with multiple resistant organisms . For these

reasons, it is recommended that patients with larger burn injuries be

isolated in private rooms or other enclosed bed spaces (Kates et

al .,1991).

Special attention is also required for patients with smaller burn

injuries who are colonized or infected with multiple drug-resistant

organisms (i.e., methicillin-resistant S aureus, vancomycin-resistant

enterococci, multiple drug-resistant gram negative organisms). This is

especially true for patients with wound drainage that cannot be

adequately contained in dry, occlusive dressings or pediatric patients who

cannot comply with hand washing or other precautions. Patients

transferred to the burn unit after treatment in another hospital should also

be included in this group until the results of their admission cultures are

known. These patients are frequently colonized with resistant organisms

and may serve as an unsuspected reservoir for transmission to other

patients unless they are isolated. Isolation for this group of patients

generally includes placement in a private room and contact precautions,

with the addition of droplet precautions in some circumstances (Weber et

al .,1993).

Patients colonized with multiple drug resistant organisms must

frequently have their need for isolation balanced with their need for

rehabilitation and psychosocial needs. In general, if the patient’s dressing

cannot be occlusive, the patient should not be taken to the rehabilitation

department for therapy when other patients are present in the same area.

If rehabilitation needs cannot be met in the patient’s room, then sufficient

time should be scheduled in the rehabilitation department to allow for the

patient’s treatment followed by thorough cleaning of all equipment and

surfaces before the area is used by other patients. The rehabilitation

therapy staff should wear appropriate attire during therapy (Rutala and

Weber , 2004).

Pediatric burn patients should also have policies restricting the

presence of non-washable toys, such as stuffed animals and cloth objects.

These can harbor large numbers of bacteria and are difficult to disinfect.

Routine cleaning, disposal of waste and gathering of soiled linen is

essential to reduce the load of organisms and ensure that the unit is as

clean as possible. Routine environmental surveillance culturing is

generally limited to the hydrotherapy room and common treatment room

used in burn wound care; however, its scope needs to be extended

(Rutala and Weber , 2004).

Plants and flowers should not be allowed in units with burn patients

because they harbor gram-negative organisms, such as Pseudomonas

species, other enteric gram-negative organisms and fungi. Many of these

organisms are intrinsically resistant to multiple antibiotics, which may

serve as reservoirs to colonize the burn wound .Modern burn centers have

a contained perimeter that is designed to minimize unnecessary traffic of

healthcare workers and visitors through the unit . Cross-contamination is

further diminished within the burn unit by housing burn patients in

individual nursing units composed of individual isolation rooms, each

with its own laminar airflow (Herndon and Spies ,2001).

11-Isolation and precautions

Isolation precautions are an important aspect of hospital infection

control programs and are particularly important in pediatric settings given

the high admission rates for viral respiratory (VRI) and gastrointestinal

(VGI) infections. Factors such as diapering of patients and inability of

young patients to adhere to proper respiratory etiquette help facilitate

transmission of infectious pathogens. Education and periodic evaluation

of adherence to precautions are recommended administrative controls to

optimize isolation practices in hospitals (Page ,2005).

Although the neonatal intensive care unit (NICU) had the lowest rate

of isolation overall, it had the highest rate of isolation for health care-

associated infections/ multidrug-resistant organism (MDRO)

colonization, with colonizations with MDROs accounting for the majority

of isolations. The hematology/oncology/ hematopoeitic stem cell

transplantation ward followed, with a high rate of patients isolated for

health care associated infections/MDRO colonization, in this case,

primarily contact isolation for gastrointestinal infection including VGI

and Clostridium difficile infection. Isolation for MDROs accounted for

12.3% of patients in isolation. Few studies have reported on actual

compliance with isolation precautions (Chatterjee et al .,2004) .

Options for patient placement include single patient rooms, two

patient rooms, and multi-bed wards. Of these, single patient rooms are

prefered when there is a concern about transmission of an infectious

agent (Cepeda et al .,2005) .

In the absence of obvious infectious diseases that require specified

airborne infection isolation rooms (e.g., tuberculosis, SARS, chickenpox),

the risk of transmission of infectious agents is not always considered

when making placement decisions. When there are only a limited number

of single-patient rooms, it is prudent to prioritize them for those patients

who have conditions that facilitate transmission of infectious material to

other patients (e.g., draining wounds, stool incontinence, uncontained

secretions) and for those who are at increased risk of acquisition and

adverse outcomes resulting from HAI (e.g., immunosuppression, open

wounds, indwelling catheters, anticipated prolonged length of stay, total

dependence on HCWs for activities of daily living)(Krause et al .,2003) .

During a suspected or proven outbreak caused by a pathogen whose

reservoir is the gastrointestinal tract, use of single patient rooms with

private bathrooms limits opportunities for transmission, especially when

the colonized or infected patient has poor personal hygiene habits, fecal

incontinence, or cannot be expected to assist in maintaining procedures

that prevent transmission of microorganisms (e.g., infants, children, and

patients with altered mental status or developmental delay) (Bridges et

al .,2003) .

High-intensity narrow-spectrum light (HINS-light) is a new light-

based disinfection method that has been shown to inactivate a wide range

of bacterial pathogens including those that are commonly associated with

hospital-acquired infections. The HINS-light method utilises a narrow

bandwidth of high-intensity visible violet light, with peak output at 405

nm. Inactivation of bacteria by exposure to high-intensity 405 nm light is

thought to be associated with the photo-excitation of molecules such as

porphyrins within the bacteria, a process that results in the production of

highly reactive compounds such as singlet oxygen which are strongly

bactericidal (Maclean et al .,2009) .

Although HINS-light EDS (high-intensity narrow spectrum light

environmental decontamination system) has wide spectrum bactericidal

activity, it is concentrated on its effectiveness in reducing mainly

staphylococcal bacteria. Staphylococcus aureus is one of the most

commonly isolated pathogens from infected burns patients .In addition,

the inanimate environment surrounding burns patients has been shown to

be a reservoir for pathogens including MRSA, and various studies have

shown an association between S. aureus dispersal from burns patients and

the size of the burn wound (Dettenkofer and Spencer ,2007) .

* Special interventions for control of tuberculosis

Tuberculosis (TB) remains a serious health problem in both the

developed and developing world. Recent CDC guidelines have

recommended a number of administrative, engineering and personal

protection measures to control TB spread in healthcare settings.

Recommended administrative controls include TB testing for all patients

at risk of TB, implementing a written TB control plan in the hospital and

housing infected patients in separate rooms. All rooms housing TB

patients should have at least 12 outdoor air changes per hour (ACH) ,

have a negative pressure of at least 0.01 inch water, and the rooms of

patients with actual or suspected TB should be checked visually with tests

such as smoke tests. HEPA air filters in patient rooms and UV irradiation

in the ventilation systems or upper part of rooms is also strongly

recommended to reduce airborne TB levels. HEPA masks or other

respiratory protection need to be worn by healthcare workers and visitors

to rooms of infectious TB patients. Proper cleaning and disinfecting of

instruments used by TB patients are also essential (Humphreys ,2007) .

Summary & Conclusion

Nosocomial infection (NI) or hospital acquired infection (HAI) can

be defined as an infection acquired in hospital by a patient who was

admitted for a reason other than that infection . This includes infections

acquired in the hospital but appearing after discharge, and also

occupational infections among staff of the facility .

Among the more industrialized and developed nations, the World

Health Organization found 8.7 % of all hospitalized patients to have

nosocomial infections. While HAI are an important health care concern

worldwide , they are especially troublesome in developing nations.

Nosocomial infection rates range from 1% in Northern Europe, especially

the Netherlands, which introduced extremely aggressive infection control

measures, to 40% in some parts of Asia, South America, and sub-Saharan

Africa .

Nosocomial infections (NI) contribute significantly to morbidity and

mortality, as well as to excess costs for hospitalized patients. According

to the available evidence, the impact of Health care associated infection

(HCAI) implies prolonged hospital stay, long-term disability, increased

resistance of microorganisms to antimicrobials, massive additional

financial burden for health systems, high costs for patients and their

family, and unnecessary deaths .The increased length of stay for infected

patients is the greatest contributor to cost .

Direct transmission from another host (healthy or ill) or from an

environmental reservoir or surface by direct contact or direct large-

droplet spread of infectious secretions is the simplest route of agent

spread. Examples of direct-contact transmission routes include kissing

(infectious mononucleosis), shaking hands [common cold (rhinovirus)],

or other skin contact (e.g., contamination of a wound with Staphylococci

or Enterococcus spp. during trauma, surgical procedures or dressing

changes) .

Potentially pathogenic micro-organisms can colonize environmental

surfaces in the hospital environment and so act as a source for outbreaks

of nosocomial infection. Studies have presented evidence that the

majority of Gram-positive bacteria, including Staphylococcus aureus and

Enterococcus spp., are able to survive for months on dry surfaces. Gram-

negative bacteria, such as Klebsiella spp., Escherichia coli, and

Acinetobacter spp. can also survive for a relatively long time on

inanimate surfaces, while common fungi such as Candida spp. have

similar properties. Environmental conditions such as low temperature or

humidity appear to be crucial for the persistence of these organisms on

inanimate surfaces .

The highest prevalence of HAI occurred in ICUs and acute care

surgical and orthopedic settings. Old age, multiple morbidities or disease

severity, and decreased immunity increase patient susceptibility. Poor

infection control measures are an overall risk factor as are certain

invasive procedures including central venous or urinary catheter

placements. Antimicrobial misuse is associated with drug-resistant HAI .

Urinary tract, respiratory tract, surgical site, skin and bloodstream

infections are currently recognized as the major nosocomial infections.

However, it is becoming increasingly clear that gastroenteritis outbreaks

are also a major burden on the health services of industrialized nations .

Analysis of nosocomial pathogens has relied on a comparison of

phenotypic characteristics such as biotypes, serotypes, bacteriophage or

bacteriocin types, and antimicrobial susceptibility profiles. This approach

has begun to change over the past 2 decades, with the development and

implementation of new technologies based on DNA, or molecular

analysis. These DNA-based molecular methodologies, include pulsed-

field gel electrophoresis (PFGE) and other restriction-based methods,

plasmid analysis, and PCR-based typing methods.

There are a number important attributes for successful typing

schemes: the methodologies should be standardized, sensitive, specific,

objective, and subject to critical appraisal. All typing systems can be

characterized in terms of typeability, reproducibility, discriminatory

power, ease of performance and interpretation, and cost (in terms of time

and money) . The use of strain typing in infection control decisions is

based on several assumptions: (i) isolates associated with the outbreak are

recent progeny of a single (common) precursor or clone, (ii) such isolates

will have the same genotype, and (iii) epidemiologically unrelated

isolates will have different genotypes .

Molecular techniques can be very effective in tracing the spread of

nososcomial infections due to genetically related pathogens, which would

allow infection control personnel to more rationally identify potential

sources of pathogens and aid infectious disease physicians in the

development of treatment regimens to manage patients affected by related

organisms. Therefore, the use of molecular tests is essential in many

circumstances for establishing disease epidemiology, which leads to

improved patient health and economic benefits through the reduction of

nosocomial infections .

Infection control (IC) activities are still developing in many health

institutions in Egypt. The national infection control program was started

in 2003 by the Ministry of Health and Population. The national IC

strategic plan entailed instituting IC programs in all hospitals in Egypt by

2010 .

The components of an infection control program are drawn from

regulatory requirements, current nursing home practices, and

extrapolations from hospital programs. The limited resources affect the

type and extent of programs developed . The infection control program

should include some form of surveillance for infections, an epidemic

control program, education of employees in infection control methods,

policy and procedure formation and review, an employee health program,

a resident health program, and monitoring of resident care practices. The

program also may be involved in quality improvement, patient safety,

environmental review, antibiotic monitoring, product review and

evaluation, resident safety, prepareness planning, and reporting of

diseases to public health authorities .

Conclusion

There are issues of concern about the emergence of nosocomial

infections, and the increase in morbidity, mortality, and costs

associated with these infections will drive the need for refinement

of molecular approaches to aid in the diagnosis and epidemiologic

analysis of nosocomial infections.

The evaluation of hospital-associated infections will continue to

rely on clinical infection surveillance as the first step to

understanding disease epidemiology and management of

infections.

Molecular testing will continue to be an essential tool, for tracing

of the source of infection .

Outbreak Control—A system for detection, investigation, and

control of epidemic infectious diseases is an important component

of infection control program.

Isolation—An isolation and precautions system to reduce the risk

of transmission of infectious agents

Continuing education in infection prevention and control ,Resident

health program , Employee health program , Disease reporting to

public health authorities , Facility management, including

environmental control, waste management, product evaluation and

disinfection, sterilization and asepsis are integrated component of

infection control program.

Recommendations

Many non-pharmacological interventions have been shown to

significantly reduce rates of HAIs, but are often overlooked in clinical

practice so this article recommend ;

Proper hand washing

Better nutrition

Housing patients in separate rooms

Sufficient numbers of nursing staff

Coated urinary and CVCs

Lower overall antibiotic use which will reduce risk of antibiotic-

resistant organisms and improve efficacy of antibiotics given to

patients who acquire nosocomial infections.

Molecular technique can be very effective in tracing the spread of

nosocomial infection .

References

Abdelkefi A, Torjman L, Ladeb S, Othman TB, Achour W,

Lakhal A, Hsairi M, Kammoun L, Hassen AB, Abdeladhim AB

(2005) : Randomized trial of prevention of catheter-related

bloodstream infectionby continuous infusion of low-dose

unfractionated heparin in patients with hematologic and oncologic

disease. J Clin Oncol ,7864 -7870.

Access Excellence (2009) : Schematic diagram of Plasmid

Analysis . Access Excellence @ National health museum .

Affymetrix (2002) : . Schematic diagram of DNA arrays . http://

www.affymetrix .com .

Alary M and Joly TR (1992) : Factors contributing to the

contamination of hospital water distribution systems by legionellae.

J Infect Dis; 165:565 - 569.

Alexopoulou K, Foka A, Petinaki E, Jelastopulu E,

Dimitracopoulos G, Spiliopoulou I (2006) : Comparison of two

commercial methods with PCR restriction fragment length

polymorphism of the tuf gene in the identification of coagulase-

negative staphylococci. Lett Appl Microbiol 43:450–454.

Alberts B., Johnson A., Lewis J. Raff M., Roberts, K., et al

(2008) : Molecular Biology of the Cell, 5th ed. Garland Science,

Taylor & Francis Group, NY, pp 538-539.

Anaisie EJ, Stratton SL, Dignani MC, Lee CK, Mahfouz TH, Rex

JH, Summerbell RC, Walsh TJ (2002) : Cleaning patient shower

facilities: a novel approach to reducing patient exposure to

aerosolized Aspergillus species and other opportunistic molds. Clin

Infect Dis ;35:86-88.

Andrews JR, Shah NS, Gandhi N ,Moll T, Friedland G (2007) :

Multidrug-resistant and extensively drug-resistant tuberculosis:

implications for the HIV epidemic and antiretroviral therapy rollout

in South Africa. J Infect Dis;196(3):482–490.

Angel CA, Green J, Swischuk L , Patel J (2004) : Severe

ciprofloxacin-associated pseudomembranous colitis in an eight-

year-old child. J Pediatr Surg ; 39(10):1590-1592.

Antonnucci TA, Williams N, Robinson NA. et al (2008) :

Reprocessing flexible endoscopes: origin or standards, overview of

structure/function, and review of recent outbreaks. In:

Manivannann G, editor. Disinfection and decontamination:

principles, applications and related issues. Boca Raton, FL: CRC

Press; . 177-214.

Appelgren P, Bjornhagen V, Bragderyd K, Jonsson CE, Ransjö U.

(2002) : A prospective study of infections in burn patients.

Burns;28(1):39–46.

Aragao PA, Oshiro IC, Manrique EI, Gomes CC, Matsuo LL,

Leone C, Moretti-Branchini ML, Levin AS (2001) : Pichia

anomala outbreak in a nursery exogenous source? Pediatr Infect

Dis J ;20:843-848.

Arbeit, R. D (1999): Laboratory procedures for the epidemiologic

analysis of microorganisms . In P. R. Murray, E. J. Baron, M. A.

Pfaller,F. C. Tenover, and R. H. Yolken (ed.), Manual of clinical

microbiology, 6thed. American Society for Microbiology,

Washington, D.C. 190-208 .

Archibald LK, Corl A, Shah B , Schulte M, Arduino MJ, Aguero

S, Fisher DJ, Stechenberg BW, Banerjee SN, Jarvis WR

(1997) : Serratia marcescens outbreak associated with extrinsic

contamination of 1% chlorxylenol soap. Infect Control Hosp

Epidemiol ;18:704-709.

Astagneau P, Rioux C, Golliot F, Brücker G (2001) : Morbidity and

mortality associated with surgical site infections: results from the

1997-1999 INCISO surveillance. J Hosp Infect ; 48:267—274..

Avila-Figueroa C , Herna´ndez-Ramos I, Gaita´n-Meza J,et al.,

León-Ramírez AR, Justiniani-Cedeño N (2000) : Extrinsic

contamination of intravenous infusates administered to hospitalized

children in Mexico. Pediatr Infect Dis;19:888–890.

Babl, F. E., Pelton S. I. , Theodore S. , and Klein J. O (2001) :

Constancy of distribution of serogroups of invasive pneumococcal

isolates among children: experience during 4 decades. Clin. Infect.

Dis. 32:1155–1161.

Barenfanger, J., Drake C. , and Kacich. G (1999) : Clinical and

financial benefits of rapid bacterial identification and antimicrobial

susceptibility testing. J. Clin. Microbiol. 37:1415–1418.

Bartlett JG (2002) : Clinical practice. Antibiotic-associated diarrhea.

N Engl J Med;346:334–349.

Bates DW, Evans RS, Murff H, Stetson PD, Pizziferri L, Hripcsak

G (2003) : Detecting adverse events using information technology.

J Am Med Inform Assoc;10(2):115-128.

Bates DW, Goldman L, Lee TH (1991) : Contaminant blood

cultures and resource utilization: the true consequences of false-

positive results. JAMA; ;265(3):365- 369.

Becker WK, Cioffi Jr WG, McManus AT, Kim SH, McManus

WF, Mason AD, Pruitt BA Jr (1991) :Fungal burn wound

infection. A 10- year experience. Arch Surg ;126(1):44–8.

Beekmann SE, Diekema DJ, Doern GV (2005) : Determining the

clinical significance of coagulase-negative staphylococci isolated

from blood cultures. Infect Control Hosp Epidemiol;26 (6):559–

566.

Beguma M, Hocking AD, Miskelly D (2009) : Inactivation of food

spoilage fungi by ultra violet (UVC) irradiation. Int J Food

Microbiol ;129 (1) :74-77.

Belda FJ, Aguilera L, Garcı´a de la, Asuncio´n J, Alberti J,

Vicente R, Ferrándiz L, Rodríguez R, Company R, Sessler DI,

Aguilar G, Botello SG, Ortí R (2005) : Supplemental

perioperative oxygen and the risk of surgical wound infection. A

randomised controlled trial. J Am Med Assoc ;294:2035-2042.

Belkum V, A., Riewerts Eriksen N., Sijmons M., W , Leeuwen W,

VandenBergh M, Kluytmans J, Espersen F, Verbrugh H (2006)

: Are variable repeats in the spa gene suitable targets for

epidemiological studies., MOLECULAR TECHNIQUES IN

HOSPITAL EPIDEMIOLOGY 529 of methicillin-resistant

Staphylococcus aureus strains? Eur. J. Clin. Microbiol. Infect. Dis.

19(15):768–770.

Belkum V , A (2007) : Guidelines for the validation and application

of typing methods for use in bacterial epidemiology. Clin.

Microbiol. Infect. 13(Suppl. 3):1-46.

Benson DA , Karsch-Mizrachi I ,Lipman DJ,Ostell J,Rapp BA,

Wheeler DL (2002) : GenBank . Nucleic Acids Res ; 30 ;17-20 .

Bhalla A, Pultz NJ, Gries DM, Ray AJ, Eckstein EC, Aron DC,

Donskey CJ. (2004) : Acquisition of nosocomial pathogens on

hands after contact with environmental surfaces near hospitalized

patients. Infect Control Hosp Epidemiol ;25(2):164-167.

Bhutta ZA, Okeke IN, Laxminarayan R, , Duse AG, Jenkins P,

O'Brien TF, Pablos-Mendez A, Klugman KP (2005):

Antimicrobial resistance in developing countries: part I—recent

trends and current status. Lancet Infect Dis; 5: 481–93.

Billgren M, Christenson B, Hedlund KO, Vinje J (2002) :

Epidemiology of Norwalk-like human caliciviruses in hospital

outbreaks of acute gastroenteritis in the Stockholm area in 1996. J

Infect ;44:26—32.

Bjerke NB (2004) : The evolution: handwashing to hand hygiene

guidance. Crit Care Nurs Q ;27:295-307.

Boni L, Benevento A, Rovera F, Dionigi G, Di Giuseppe M,

Bertoglio C, Dionigi R (2006) : Infective complications in

laparoscopic surgery. Surg Infect (Larchmt);7( 2):109 -111.

Boswell T, Machin K, Meredith K, Soo SS,et al (2006) : Airborne

dissemination of epidemic Pseudomonas aeruginosa in the

Nottingham Cystic Fibrosis population: a role for portable HEPA

filtration? Abstract presented at the Federation of Infection

Societies Meeting in Cardiff, UK; .

Boyce JM , Havill NL, Otter JA, McDonald LC, Adams NM,

Cooper T, Thompson A, Wiggs L, Killgore G, Tauman A,

Noble-Wang J (2006) : Impact of hydrogen peroxide vapor room

bio-decontamination on environmental contamination and

nosocomial transmission by Clostridium difficile. Presented at the

16th Annual Scientific Meeting of the Society for Healthcare

Epidemiology of America, Chicago, IL,; 155.

Bridges CB, Kuehnert MJ, Hall CB (2003) : Transmission of

influenza: implications for control in health care settings. Clin

Infect Dis ;37(8):1094-1101.

Brown SM, Lubimova AV, Khrustalyeva NM, Shulaeva SV,

Tekhova I, Zueva LP, Goldmann D, O'Rourke EJ (2003) : Use

of an alcohol-based hand rub and quality improvement

interventions to improve hand hygiene in a Russian neonatal

intensive care unit. Infect Control Hosp Epidemiol; 24:172 - 179.

Burke JP (2004) : Infection control—A problem for patient safety. N

Engl J Med ;348:651- 656.

Carbonnelle, E (2007) : Rapid identification of staphylococci

isolated in clinical microbiology laboratories by matrix-assisted

laser desorption ionization time of flight mass spectrometry. J Clin.

Microbiol. 45:2156-2216.

Carling PC, Parry MF, Von Beheren SM (2008) : Identifying

opportunities to enhance environmental cleaning in 23 acute care

hospitals. Infect Control Hosp Epidemiol ;29:1- 7.

Carling PC, Von Beheren S, Kim P, Woods C (2008) : Intensive

care unit environmental cleaning: an evaluation in sixteen hospitals

using a novel assessment tool. J Hosp Infect ;68:39 - 44.

Casey AL, Adams D, Karpanen TJ, Lambert PA, Cookson BD,

Nightingale P, Miruszenko L, Shillam R, Christian P, Elliott

TS (2010) : Role of copper in reducing hospital environment

contamination. J Hosp Infect;74:72 - 77.

Casey BM and Cox SM (1997) : Chorioamnionitis and

endometritis. Infect Dis Clin North Am;11:203-222.

Cavalcante SS, Mota E, Silva LR, Teixeira LF, Cavalcante LB

(2006) : Risk factors for developing nosocomial infections among

pediatric patients. Pediatr Infect Dis J 2006; 25: 438–45.

CDC (1997) : Prevention of pneumococcal disease: recommendations

of the Advisory Committee on Immunization Practices (ACIP).

MMWR Recomm Rep ;46:1-24.

CDC (1997) : Sustained transmission of nosocomial Legionnaires

disease. MMWR; 46:416-421.

CDC (2002) : Guideline for Hand Hygiene in Health-Care Settings:

Recommendations of the Healthcare Infection Control Practices

Advisory Committee and the HICPAC/SHEA/APIC/IDSA Hand

Hygiene Task Force. MMWR ;51(16):1-44.

CDC (2009) : Gould CV, Umscheid CA, Agarwal RK, Kuntz G,

Pegues DA, and the Healthcare Infection Control Practices

Advisory Committee (HICPAC). Guidelines for prevention of

catheter-associated urinary tract infections,.

Cengiz M , Celikbilek G , Andic, Dosemeci L, Yilmaz M, Karaali

K, Ramazanoglu A (2009 ) : Maxillary sinusitis in patients

ventilated for a severe head injury and with nostrils free of any

foreign body . ; Injury . 42(1):33-37.

Cepeda JA, White house T, Cooper B , Hails J, Jones K, Kwaku F,

Taylor L, Hayman S, Cookson B, Shaw S, Kibbler C, Singer M,

Bellingan G, Wilson AP (2005) : Isolation of patients in single

rooms or cohorts to reduce spread of MRSA in intensive-care units:

prospective two-centre study. Lancet ;365(9456):295-304 .

Chadwick PR, Beards G, Brown D, Caul EO, Cheesbrough J,

Clarke I, Curry A, O'Brien S, Quigley K, Sellwood J,

Westmoreland D (2000) : Management of hospital outbreaks of

gastro-enteritis due to small round structured viruses. J Hosp Infect

;45:1—10.

Champlin R (2000) : Control of nosocomial Clostridium difficile

transmission in bone marrow transplant patients. Infect Control

Hosp Epidemiol ;21:226–228.

Chang, N., and Chui. L (1998) : A standardized protocol for the

rapid preparation of bacterial DNA for pulsed-field gel

electrophoresis. Diagn. Microbiol. Infect. Dis. 31:275–279.

Chatterjee A, Heybrock B, Plummer S , Eischen K. (2004) :

Impact of surveillance rounds on adherence to infection control

policies and procedures at a children’s hospital. Infect Control

Hosp Epidemiol ;25:786-788.

Cheadle WG ( 2006) : Risk factors for surgical site infections. Surg

Infect (Larchmt) ;7(1):7 - 11.

Cherif H, Kronvall G, Bjorkholm M, Kalin M (2003) : Bacteraemia

in hospitalized patients with malignant blood disorders: a

retrospective study of causative agents and their resistance profiles

during a 14-year period without antibacterial prophylaxis. Hematol

J;4:420–426.

Church DL (2005) : Intensive care unit-acquired urinary tract

infections .

Clarridge, J.E (2004) : Impact of 16S rRNA gene sequence

analysis for identification of bacteria on clinical microbiology and

infectious diseases. Clin. Microbiol. Rev. 17:840-862.

CLSI (2001) : Routine urine analysis and collection , transportation

and preservation of urine specimens . 2 ed Wayne .

CLSI (2003) : Procedure for the collection of diagnostic blood

specimen by venipuncture ; approved standard . 5 ed Wayne.

CLSI (2006) : Collection , transport , preparation and storage of

specimens for molecular methods . Wayne (PA): CLSI.

CLSI (2008) : Performance standards for antimicrobial susceptibility

testing; eighteenth informational supplement. Wayne (PA): CLSI.

Cockerill, F. R., and Smith, T. F (2004): Response of the clinical

microbiology laboratory to emerging (new) and reemerging

infectious diseases. J. Clin. Microbiol.42:2359–2365.

Cohen S, Tyrell AJ, Smith AP (1991) : Psychological stress and

susceptibility to the common cold. N Engl J Med;325:606-612.

Coello R , Glenister H, Fereres J, Bartlett C, Leigh D, Sedgwick J,

Cooke EM (1993) : The cost of infection in surgical patients: a

case study. J Hosp Infect, 25:239– 250.

Collard HR, Saint S, Matthay MA (2003) : Prevention of ventilator

associated pneumonia: An evidence-based systematic review. Ann

Intern Med;138:494-501.

Cook DJ, Walter SD, Cook RJ, Griffith LE, Guyatt GH, Leasa D,

Jaeschke RZ, Brun-Buisson C (1998) : Incidence of and risk

factors for ventilator-associated pneumonia in critically ill patients.

Ann Intern Med;129:433-440.

Cooke EM, Coello R, Sedgwick J, Ward V, Wilson J, Charlett A,

Pearson (2000) :A national surveillance scheme for hospital

associated infections in England. Team of the Nosocomial

Infection National Surveillance Scheme. J Hosp Infect, 46:1-3.

Coticchia JM and Dohar JE (2005) : Methicillin-resistant

Staphylococcus aureus otorrhea after tympanostomy tube

placement. Arch Otolaryngol Head Neck Surg;131:868-873.

Cross JT Jr, Campbell GD Jr (2001) : Therapy of nosocomial

pneumonia. Med Clin North Am ;85:1583-1594.

Curtis LT (2008): Prevention of hospital-acquired infections: review

of non- pharmacological interventions. Journal of Hospital

Infection 69, 204-219.

Dancer SJ (2008) : Importance of the environment in

meticillinresistant Staphylococcus aureus acquisition: the case for

hospital cleaning. Lancet Infect Dis ;8:101-113.

Dancer S.J (2009) : The role of environmental cleaning in the

control of hospital-acquired infection , Journal of Hospital

Infection 73, 378-385 .

Daniel M, Sramova V, Absolonova D , Dĕdicová D, Lhotová H,

Masková L, Petrás P (1992) : Arthropods in a hospital and their

potential significance in the epidemiology of hospital infections.

Folia Parasitol ;39:159-170.

Darmstadt GL, Dinulos JG, Miller Z (2000) : Congenital cutaneous

candidiasis: clinical presentation, pathogenesis, and management

guidelines. Pediatrics;105:438-444.

David J. Weber William A. Rutala Mayhall, et al (2004) :

Nosocomial Ocular Infections Title: Hospital Epidemiology and

Infection Control, 3rd Edition . Lippincott Williams & Wilkins .

Davidson (2002) : Diagram of pulsed-field gel electrophoresis .

http:// www. biochDavidson . com .

Dembry, Clewell D. B., Alshab D., and Zervos. M. J (1998) :

Molecular analysis of glycopeptide-resistant Enterococcus faecium

isolates collected from Michigan hospitals over a 6-year period. J.

Clin. Microbiol. 36:3303– 3308.

Denys GA (2000) ; Infectious waste management. In: Lederberg J, ed.

Encyclopedia of microbiology, vol 2, 2nd ed. Orlando, FL:

Academic Press,:782 - 796.

Dettenkofer M and Spencer RC (2007) : Importance of

environmental decontamination e a critical view. J Hosp

Infect ;65:55-57.

Dia NM, Ka R, Dieng C, Dia ML, Fortes L, Diop BM, Sow AI,

Sow PS (2008) : Prevalence of nosocomial infections in a

university hospital (Dakar, Senegal). Med Mal Infect; 38: 270–274.

Diaz O, Diaz E, Rello J (2003) : Risk factors for pneumonia in the

intubated patient. Infect Dis Clin North Am ;17:697-705.

Diekema DJ, Messer RJ, Hollis R, Jones N, Pfaller MA (2004) :

Nosocomial candidemia: an ounce of prevention is better than a

pound of cure. Infect Cont Hosp Epidemiol;624—626.

Dodek P, Keenan S, Cook D, Heyland D, Jacka M, Hand L,

Muscedere J, Foster D, Mehta N, Hall R, Brun-Buisson C

(2004) : Evidence-based clinical practice guideline for the

prevention of ventilator-associated pneumonia. Ann Intern

Med ;141:305-313.

Doft BM, Kelsey SF, Wisniewski SR (2000) : Retinal detachment in

the endophthalmitis vitrectomy study. Arch Ophthalmol 118:1661–

1665.

Dohar J (2005) : Microbiology of otorrhea in children with

tympanostomy tubes: implications for therapy. Int J Pediatr

Otorhinolaryngol ;67:1317-1323.

Dohmen PM (2006) : Influence of skin flora and preventive

measures on surgical site infection during cardiac surgery. Surg

Infect (Larchmt);7(1):13 - 17.

Ducel G (1995) : Les nouveaux risques infectieux. Futuribles, ,

203:5–32.

Duckro AN, Blom DW, Lyle EA, Weinstein RA, Hayden MK

(2005) : Transfer of vancomycin-resistant enterococci via health

care worker hands. Arch Intern Med ;165(3):302-307 .

Duckworth GJ and Jordens JZ (1990) : Adherence and survival

properties of an epidemic methicillin-resistant strain of

Staphylococcus aureus compared with those of methicillin-

sensitive strains. J Med Microbiol ;32: 195- 200.

Duffy LM, Cleary J, Ahern S, Kuskowski MA, West M, Wheeler

L, Mortimer JA (1995) : Clean intermittent catheterization: safe,

cost-effective bladder management for male residents of VA

nursing homes. J Am Geriatr Soc ;43:865-870.

Edwards JR, Alonso-Echanove J, Richards MJ, Brennan P,

Venezia RA, Keen J, Ashline V, Kirkland K, Chou E, Hupert

M, Veeder AV, Speas J, Kaye J, Sharma K, Martin A, Moroz

VD, Gaynes RP (2003) : Effect of nurse staffing and

antimicrobial-impregnated central venous catheters on the risk for

bloodstream infections in intensive care units. Infect Control Hosp

Epidemiol ;24:916 - 925.

Edwards JR, Peterson KD, Andrus ML, Dudeck MA, Pollock DA,

Horan TC (2007) : National Healthcare Safety Network (NHSN)

Report. Data summary for 2006, . Am J Infect Cont 2007;35:290-

301.

Eisgruber H., Wiedmann M., and Stolle A (1995) : Use of plasmid

profiling as a typing method for epidemiologically related

Clostridium perfringens isolates from food poisoning cases and

outbreaks. Lett. Appl. Microbiol. 20:290–294.

El Sayed NM, Gomatos PJ, Beck-Sague CM, Dietrich U, von

Briesen H, Osmanov S, Esparza J, Arthur RR, Wahdan MH,

Jarvis WR. (2000) Epidemic transmission of human immunodefi

ciency virus in renal dialysis centers in Egypt. J Infect Dis; 181:

91–97.

El-Nawawy A (2003) : Evaluation of the outcome of patients

admitted to the pediatric intensive care unit in Alexandria using the

pediatric risk of mortality (PRISM) score. J Trop Pediatr;49:109-

114.

Emori TG and Gaynes R (1993) : An overview of nosocomial

infections, including the role of the microbiology laboratory. Clin

Microbiol Rev;6:428-442.

Emori TG, Edwards JR, Culver DH, Sartor C, Stroud LA, Gaunt

EE, Horan TC, Gaynes RP (1998) : Accuracy of reporting

nosocomial infections in intensive care unit patients to the National

Nosocomial Infections Surveillance (NNIS) system: a pilot study.

Infect Control Hosp Epidemiol;19:308- 316.

Engelhard D, Cordonnier C, Shaw PJ, Parkalli T, Guenther C,

Martino R, Dekker AW, Prentice HG, Gustavsson A,

Nurnberger W, Ljungman P (2002) : Early and late invasive

pneumococcal infection following stem cell transplantation: a

European bone marrow transplantation survey. Br J

Haematol;117:444–450.

Eribe, E. R., and Olsen. I (2000) : Strain differentiation in

Bacteroides fragilis by ribotyping and computer-assisted gel

analysis. APMIS 108:429–438.

Eriksen HM, Iversen BG, Aavitsland P (2005) : Prevalence of

nosocomial infections in hospitals in Norway, 2002 and 2003. J

Hosp Infect;60:40-45

Esteban A, Frutos-Vivar F, Ferguson ND, Arabi Y, Apezteguía C,

González M, Epstein SK, Hill NS, Nava S, Soares MA,

D'Empaire G, Alía I, Anzueto A (2004) : Non invasive positive-

pressure ventilation for respiratory failure after extubation. N Engl

J Med ;350:2452-2460.

Farmer JC, Guntupalli KK, Baldisseri M, Gilstrap L (2005) :

Critical illness of pregnancy. Crit Care Med ;33:247–397.

Favre B, Hugonnet S, Correa L, Sax H, Rohner P, Pittet D (2005) :

Nosocomial bacteremia: clinical significance of a single blood

culture positive for coagulase-negative staphylococci. Infect

Control Hosp Epidemiol 26: 697–702.

Feil E. J., Cooper H. , Grundmann D. A., Robinson DA, Enright

MC, Berendt T, Peacock SJ, Smith JM, Murphy M, Spratt BG,

Moore CE, Day NP (2003) : How clonal is Staphylococcus

aureus? J. Bacteriol. 185:3307–3316.

Finn C (2007) : Introduction to microbiology . in Mary L . cClinical

Laboratory Science . 5 ed . Mosby . 455-457 .

Finney, M (1993) : Pulsed-field gel electrophoresis, In F. M.

Ausubel, R. Brent, R. E. Kingston, D. D. Moore, J. G. Seidman, J.

A.Smith, and K. Struhl (ed.), Current protocols in molecular

biology,. Greene-Wiley, New York, N.Y. 1 (7) 251–259.

Fletcher RH (1996) : Clinical epidemiology, the essentials.

Baltimore, Williams & Wilkins .

Focucault, C., LaScola B., Lindroos H., Andersson S. G. E., and

Raoult. D (2005) : Multispacer typing technique for sequence-

based typing of Bartonella quintana. J. Clin. Microbiol. 43:41–48.

Fourrier F, Dubois D, Pronnier P, Herbecq P, Leroy O, Desmettre

T, Pottier-Cau E, Boutigny H, Di Pompéo C, Durocher A,

Roussel-Delvallez M (2005) : Effect of gingival and dental plaque

antiseptic decontamination on nosocomial infections acquired in

the intensive care unit: a double-blind placebo-controlled

multicenter study. Crit Care Med ; 33:1728-1735.

Francois, P., G. Renzi, D. Pittet, M. Bento, Lew D, Harbarth S,

Vaudaux P, Schrenzel J (2004) : A novel multiplex real-time

PCR assay for rapid typing of major staphylococcal cassette

chromosome mec elements. J. Clin. Microbiol. 42:3309–3312.

Freeman s ( 2002) : Schematic diagram of Restriction fragment

length polymorphisms http://biology .arizona.edu /sciconn/lessons2

/lessons. Html .

French GL and Cheng AF (1991): Measurement of the costs of

hospital infection by prevalence surveys. J Hosp Infect 18(Suppl.

A):65—72.

Gaia V, Fry NK, Afshar B, Lück PC, Meugnier H, Etienne J,

Peduzzi R, Harrison TG (2005) : Consensus sequence-based

scheme for epidemiological typing of clinical and environmental

isolates of Legionella pneumophila. J Clin Microbiol;43:2047-

2052.

Gallinella G., Venturoli S., Manaresi E., Musiani M., Zerbini M

(2003) : B19 virus genome diversity: epidemiological and clinical

correlations. J. Clin. Virol. 28, 1–13.

Gastmeier P, Geffers C, Brandt C, Zuschneid I, Sohr D, Schwab

F,Behnke M, Daschner F, Rüden H (2006): Effectiveness of a

nationwide nosocomial infection surveillance system for reducing

nosocomial infections. J Hosp Infect, 64:16 - 22 .

Gaynes RP, Culver DH, Horan TC, Edwards JR, Richards C,

Tolson JS (2001) : Surgical site infection (SSI) rates in the

United States, 1992-1998: the National Nosocomial Infections

Surveillance System basic risk index. Clin Infect Dis ;33(2):69 -

77.

Gevers D ( 2005) : Opinion: reevaluating prokaryotic species. Nat.

Rev. Microbiol. 3:733-739.

Gleizes O, Desselberger U, Tatochenko V, Rodrigo C, Salman N,

Mezner Z, Giaquinto C, Grimprel E (2006) : Nosocomial

rotavirus infection in European countries: a reviewof the

epidemiology, severity and economic burden of hospitalacquired

rotavirus disease. Pediatr Infect Dis J ;25: 12-21.

Goering, R. V (1998) : The molecular epidemiology of nosocomial

infection: an overview of principles, application, and interpretation,

. In S. Specter, M. Bendinelli, and H. Friedman (ed.), Rapid

detection of infectious agents. Plenum Press, New York, N.Y. 131–

157.

Goering RV (2010) : Pulsed field gel electrophoresis: A review of

application and interpretation in the molecular epidemiology of

infectious disease Infection, Genetics and Evolution ,10, 866–875 .

Goldmann DA, Weinstein RA, Wenzel RP, Tablan OC, Duma RJ,

Gaynes RP, Schlosser J, Martone WJ (1996) : Strategies to

prevent and control the emergence and spread of antimicrobial-

resistant microorganisms in hospitals: a challenge to hospital

leadership. JAMA;275:234-240.

Gonzalez-Barca E, Carratala J, Mykietiuk A, Fernandez-Sevilla

A, Gudiol F (2001) : Predisposing factors and outcome of

Staphylococcus aureus bacteremia in neutropenic patients with

cancer. Eur J Clin Microbiol Infect Dis;20:117–119.

Gordis L (1996) : Epidemiology. Philadelphia, W.B. Saunders

Company,

Gordis L (2000) : Epidemiology, 2nd ed. Philadelphia: WB

Saunders,.

Gray J and Hobbs PM (2002) : Management of outbreaks of

Gramnegative bacteria in neonatal units. J Hosp Infect ;52: 317—

318.

Green KY, Belliot G, Taylor JL, Valdesuso J, Lew JF, Kapikian

AZ, Lin FY (2002) : A predominant role for Norwalk-like viruses

as agents of epidemic gastroenteritis in Maryland nursing homes

for the elderly. J Infect Dis ; 185:133—146.

Grimm V, Ezaki S, Susa M, Knabbe C, Schmid RD, Bachmann

TT (2004) : Use of DNA microarrays for rapid genotyping of

TEM beta-lactamases that confer resistance. J Clin Microbiol

42:3766–3774.

Gürcüog˘lu , Akalın H., Ener B. , Ocakog˘lu G. , Sınırtas M. ¸ ,

Sevim Akçag˘lar , Yılmaz E. , Evci C. B (2010) : Epidemiology

of nosocomial candidaemia in a university hospital: a 12-year

study. ;138(9):1328-1335.

Hanes SD, Demirkan K, Tolley E, Boucher BA, Croce MA, Wood

GC, Fabian TC (2002) : Risk factors for late onset nosocomial

pneumonia caused by Stenotrophomonas maltophilia in critically

ill trauma patients. Clin Infect Dis ;35:228-235.

Haynes AB, Weiser TG, Berry WR, , Lipsitz SR, Breizat AH,

Dellinger EP, Herbosa T, Joseph S, Kibatala PL, Lapitan MC,

Merry AF, Moorthy K, Reznick RK, Taylor B, Gawande AA

(2009): A surgical safety checklist to reduce morbidity and

mortality in a global population. N Engl J Med ; 360: 491–99.

Health Services Advisory Committee ( 1999 ) : Safe disposal of

clinical waste HSE Books ,Sudbury.

Heath PT, Balfour G, Weisner AM, Efstratiou A, Lamagni TL,

Tighe H, O'Connell LA, Cafferkey M, Verlander NQ, Nicoll A,

McCartney AC (2004) : Group B streptococcal disease in UK

and Irish infants younger than 90 days. Lancet Jan 24; 363(9405):

292–294.

Herndon DN and Spies M ( 2001) : Modern burn care. Semin Pediatr

Surg ;10(1):28–31.

Hopkins K. L., Desai M., Frost J. A., Stanley J., and. Logan. J. M.

J (2004) : Fluorescent amplified fragment length polymorphism

genotyping of Campylobacter jejuni and Campylobacter coli

strains and its relationship with host specificity, serotyping and

phage typing. J. Clin. Microbiol. 42:229–235.

Hornga Yu-Tze , Po-Chi S, Bin-Jon S , Hung YL, Lo KY, Su HP,

Wei JR, Hsieh SC, Hsueh PR, Lai HC (2006 ): Development of

an improved PCR–ICT hybrid assay for direct detection of

Legionellae and Legionella pneumophila from cooling towerwater

specimens ;http://dx. doi.org/10.1016 /j.watres.03.033 .

Howell M (2002) : Do nurses know enough about percutaneous

endoscopic gastrostomy ? Nursing Times , 98 (17) 40-42 .

Huang HI, Shih HY, Lee CM, Yang TC, Lay JJ, Lin YE (2008) :

In vitro efficacy of copper and silver ions in eradicating

Pseudomonas aeruginosa, Stenophomonas maltophilia and

Acinetobacter baumannii: implications for on-site disinfection for

hospital infection control. Water Res ;42:73- 80.

Huard RC, Lazzarini LC, Butler WR , van Soolingen D, Ho JL

(2003) : PCR-based method to differentiate the subspecies of the

Mycobacterium tuberculosis complex on the basis of genomic

deletions. J Clin Microbiol;41:1637-1650.

Hubmayr RD, Burchardi H, Elliot M , Fessler H, Georgopoulos D,

Jubran A, Limper A, Pesenti A, Rubenfeld G, Stewart T, Villar

J (2002) : Statement of the 4th International Consensus

Conference in Critical Care on ICU-Acquired Pneumonia –

Chicago, Illinois, May 2002. Intensive Care Med ;28:1521-1536.

Humphreys H (2007) : Control and prevention of

healthcareassociated tuberculosis: the role of respiratory isolation

and personal respiratory protection. J Hosp Infect ; 66:1-5.

Humphreys H (2009) : Preventing surgical site infection. Where

now? Journal of Hospital Infection 73, 316-322 .

Issa M, Vijayapal A, Graham MB, Beaulieu DB, Otterson MF,

Lundeen S, Skaros S, Weber LR, Komorowski RA, Knox JF,

Emmons J, Bajaj JS, Binion DG (2007) : Impact of Clostridium

difficile on inflammatory bowel disease. Clin Gastroenterol

Hepatol ;5:345 351.

James DK, Steer PJ, Weiner CP, Gonik B (2005 ) : High risk

pregnancy. 3rd ed. London: WB Saunders.

Jarlais D , D. C., Perlis, T., et al. (2005) : HIV incidence among

injection drug users in New York City, 1990 to 2002. American

Journal of Public Health, 95, 1444– 1493.

Jiang S. C., Matte M., Matte G., Huq A., and Colwell. R. R.

(2000) : Genetic diversity of clinical and environmental isolates of

Vibrio cholerae determined by amplified fragment length

polymorphism fingerprinting. Appl. Environ. Microb. 66:148-153.

John A. W, Benjamin A., Lipsky, Tabak Y P., , Derby KG., Kim

M , and Gupta V (2010) Surgical site infections: Causative

pathogens and associated outcomes Am J Infect Control;38:112-20.

Johnson S and Gerding DN (1998) : Clostridium difficile-associated

diarrhea. Clin Infect Dis;26:1027-1034.

Johnston CP, Cooper L, Ruby W, Carroll KC, Cosgrove SE, Perl

TM (2006) : Epidemiology of communityacquired methicillin-

resistant Staphylococcus aureus skin infections among healthcare

workers in an outpatient clinic. Infect Control Hosp Epidemiol ;

27:1133- 1136.

Jones N, Oliver K, Jones Y, Haines A, Crook D (2006) : Carriage

of group B streptococcus in pregnant women from Oxford, UK. J

Clin Pathol Apr; 59(4): 363–366.

Jones RN, Pfaller MA, Marshall SA , Hollis RJ, Wilke WW

(1997) : Antimicrobial activity of 12 broad spectrum-spectrum

agents tested against 270 nosocomial blood stream infection

isolates caused by non-enteric gram-negative bacilli: occurrence of

resistance, molecular epidemiology, and screening for metallo-

enzymes. Diagn Microbiol Infect Dis;29:187-192.

Jorgensen JH and Ferraro MJ (2000) : Antimicrobial

susceptibility testing: special needs for fastidious organisms and

difficult-to-detect resistance mechanisms. Clin Infect Dis;30:799–

808.

Jorgensen JH and Turnidge JD (2007) : Susceptibility test

methods: dilution and disk diffusion methods. In: Murray PR,

Baron EJ, Jorgensen JH, et al, editors. Manual of clinical

microbiology. 9th edition. Washington (DC): American Society for

Microbiology;. 1152–1172.

Jung H, Lee S K , Cha S , Byun J Y , Park M S , Yeo S G

(2009) : Current bacteriology of chronic otitis media with

effusion: High rate of nosocomial infection and decreased

antibiotic sensitivity* Journal of Infection 59, 308-316 .

Kam K.M., Luey C.K., Parsons M.B., Cooper KL, Nair GB, Alam

M, Islam MA, Cheung DT, Chu YW, Ramamurthy T, Pazhani

GP, Bhattacharya SK, Watanabe H, Terajima J, Arakawa E,

Ratchtrachenchai OA, Huttayananont S, Ribot EM, Gerner-

Smidt P, Swaminathan B (2008) : Evaluation and validation of a

PulseNet standardized pulsed-field gel electrophoresis protocol for

subtyping Vibrio parahaemolyticus: an international multicenter

collaborative study. J. Clin. Microbiol. 46, 2766–2773.

Kane A, Lloyd J, Zaffran M , Simonsen L, Kane M (1999) :

Transmission of hepatitis B, hepatitis C and human

immunodeficiency viruses through unsafe injections in the

developing world: model-based regional estimates. Bull World

Health Organ ; 77: 801–807.

Karadeniz Y, Kilic D, Altan S, Altinok D, Güney S (2001) :

Evaluation of the role of ultrasound machines as a source of

nosocomial and cross infection. Invest Radiol;36:554-558.

Kates SG, McGinley KJ, Larson EL, Leyden JJ (1991) :

Indigenous multiresistant bacteria from flowers in hospital and

nonhospital environments. Am J Infect Control ;19(3):156–161.

Kelly L. Moore Samuel W. Dooley William R. et al (2004) :

Mycobacterium Tuberculosis . In Mayhall, C. Glen . Hospital

Epidemiology and Infection Control, 3rd Edition . Lippincott

Williams & Wilkins .

Klevens RM, Edwards JR, Richards CL Jr, Horan TC, Gaynes

RP, Pollock DA, Cardo DM (2007) : Estimating health care-

associated infections and deaths in US hospitals, . Public Health

Rep ; 122: 160–66.

Koff MD, Loftus RW, Burchman CC, Schwartzman JD, Read

ME, Henry ES, Beach ML (2009) : Reduction in intraoperative

bacterial contamination of peripheral intravenous tubing through

the use of a novel device. Anesthesiology.;110: 978-985.

Kollef MH, Shapiro SD, Fraser VJ , Silver P, Murphy DM,

Trovillion E, Hearns ML, Richards RD, Cracchilo L, Hossin L

(1995) : Mechanical ventilation with or without 7-day circuit

changes. A randomized controlled trial. Ann Intern Med ;123:168-

74.

Koreen, L., Ramaswamy S. V., Graviss E. A., Naidich S., Musser

J. M., and Kreiswirth. B. N (2004) : Spa typing method for

discriminating among Staphylococcus aureus isolates: implications

for use of a single marker to detect genetic micro- and

macrovariation. J. Clin. Microbiol. 42:792–799.

Korinek AM, Baugnon T, Golmard JL, van Effenterre R, Coriat

P, Puybasset L (2006) ; Risk factors for adult nosocomial

meningitis after craniotomy: role of antibiotic prophylaxis.

Neurosurgery;59:126-133 .

Kotach VM (2004) : Infections due to nontuberculous mycobacteria

(NTM). Indian J Med Res;120:290-304.

Kramer A, Schwebke I, Kampf G (2006) : How long do

nosocomial pathogens persist on inanimate surfaces? A systematic

review. BMC Infect Dis;6:130.

Krause P, Drinka PJ, Nest L, Goodman BM, Gravenstein S

(2003) : Risk of acquiring influenza A in a nursing home from a

culture-positive roommate. Infect Control Hosp

Epidemiol ;24(11):872-874.

Kristo´f K, Szabo´ D, Marsh JW , Cser V, Janik L, Rozgonyi F,

Nobilis A, Nagy K, Paterson DL (2007) : Extended-spectrum

beta-lactamase-producing Klebsiella spp. in a neonatal intensive

care unit: risk factors for the infection and the dynamics of the

molecular epidemiology. Eur J Clin Microbiol Infect Dis ;26:563-

570.

Kroenke DM (2006) : Database processing. Upper Saddle River,

NJ: Pearson Prentice Hall; 549-554.

Landers TF, Hoet A, Wittum TE (2010) : Swab type, moistening,

and preenrichment for Staphylococcus aureus on environmental

surfaces. J Clin Microbiol ;48: 2235-2236.

Lappe N D, Connor JO, Doran G, Devane G, Cormican M.

(2009) : Role of subtyping in detecting Salmonella cross

contamination in the laboratory. BMC Microbiol ;9:155.

Lark RL, McNeil SA, VanderHyde K, Noorani Z, Uberti J,

Chenoweth C (2001) : Risk factors for anaerobic bloodstream

infections in bone marrow transplant recipients. Clin Infect

Dis ;33:338–343.

Laupland KB, Bagshaw SM, Gregson , Kirkpatrick AW, Ross T,

Church DL (2005) : Intensive care unit-acquired urinary tract

infections in a regional critical care system. Crit Care;9: 60—65 .

Leape LL (2002) : Reporting of adverse events. N Engl J

Med ;347:1633-1638.

Leblebicioglu H, Rosenthal VD, Arikan OA, Ozgültekin A, Yalcin

AN, Koksal I, Usluer G, Sardan YC, Ulusoy S ( 2007): Turkish

Branch of INICC: Device associated hospital-acquired infection

rates in intensive care units. Findings of the International

Nosocomial Infection Control Consortium J Hosp Infect 65:251-

257 .

Lederer JW Jr, Best D, Hendrix V (2009) : A comprehensive hand

hygiene approach to reducing MRSA health care-associated

infections. Jt Comm J Qual Patient Saf. ;35:180-184.

Lemee, L., Dhalluin A., Pestel-Caron M., Lemeland J. F., and.

Pons J. L (2004) : Multilocus sequence typing analysis of human

and animal Clostridium difficile isolates of various toxigenic types.

J. Clin. Microbiol. 42:2609–2617.

Lennox K. Archibald Walter J. Hierholzer Jr , et al (2004) :

Principles of Infectious Diseases Epidemiology. In Mayhall, C.

Glen . Hospital Epidemiology and Infection Control, 3rd Edition .

Lippincott Williams & Wilkins .

Leon S (2000) : Epidemiology of nosocomial outbreaks: 14-year

experience at a tertiary-care center. Infect Control Hosp

Epidemiol ;21:527–529.

Leung M and Chan AHS (2006) : Control and management of

hospital water quality. Med Sci Monit ;12:17-23.

Leeuwen W.B. , Snoeijers S. , Werken-Libregts C, Tuip A, van

der Zee A, Egberink D, de Proost M, Bik E, Lunter B,

Kluytmans J, Gits E, van Duyn I, Heck M, van der Zwaluw K,

Wannet W, Noordhoek GT, Mulder S, Renders N, Boers M,

Zaat S, van der Riet D, Kooistra M, Talens A, Dijkshoorn L,

van der Reyden T, Veenendaal D, Bakker N, Cookson B, Lynch

A, Witte W, Cuny C, Blanc D, Vernez I, Hryniewicz W, Fiett J,

Struelens M, Deplano A, Landegent J, Verbrugh HA, van

Belkum A (2002) : Intercenter reproducibility of binary typing

for Staphylococcus aureus, J. Microbiol. Methods 51 (1) 19–28.

Levy I, Katz J, Solter E, Samra Z, Vidne B, Birk E, Ashkenazi S,

Dagan O (2005) : Chlorhexidine impregnated dressing for

prevention of colonization of central venous catheters in infants

and children: a randomized controlled study. Pediatr Infect Dis

J ;24:676 - 679.

Lin B., Malanoski A.P., Wang Z., Blaney K.M., Ligler A.G.,

Rowley R.K., Hanson E.H.,von Rosenvinge E., Ligler F.S.,

Kusterbeck A.W., Metzgar D., Barrozo C.P. Russell K.L.,

Tibbetts C., Schnur J.M., Stenger D.A (2007). Application of

broadspectrum, sequence-based pathogen identification in an urban

population. PLoS ONE 2, 419 – 423 .

Lindsay J.A (2006) : Microarrays reveal that each of the ten

dominant lineages of Staphylococcus aureus has a unique

combination of surfaceassociated and regulatory genes. J.

Bacteriol. 188:669-676.

Liu P. Y. F., Shi Z. Y., Lan Y. J., Hu B. S., Shyr J. M., Tsai W. S.,.

Lin Y. H, and Tseng. C. Y (1996 ) : Use of restriction

endonuclease analysis of plasmids and pulsed-field gel

electrophoresis to investigate outbreaks of methicillinresistant

Staphylococcus aureus infection. Clin. Infect. Dis. 22:86

Lopman BA, Reacher MH, Vipond IB, Hill D, Perry C, Halladay

T, Brown DW, Edmunds WJ, Sarangi J (2004) : Epidemiology

and cost of nosocomial gastroenteritis, Avon, England, 2002–

2003. Emerg Infect Dis;10:1827-1834.

Louie, M., Jayaratne P. Luchsinger I., Devenish J., Yao J.,

Schlech W. and Simor. A (1996) : Comparison of ribotyping,

arbitrarily primed PCR, and pulsed-field gel electrophoresis for

molecular typing of Listeria monocytogenes. J. Clin. Microbiol.

34:15.

Lynch JP (2001) : Hospital-acquired pneumonia: Risk factors,

microbiology, and treatment. Chest ;119:373-384.

Maclean M, MacGregor SJ, Anderson JG, Woolsey GA (2009) :

Inactivation of bacterial pathogens following exposure to light

from a 405-nm LED array. Appl Environ Microbiol ;75:1932-1937.

Madani N, Rosenthal V D , Dendane T , Abidi K, Zeggwagh AA,

Abouqal R (2009) : Health-care associated infections rates, length

of stay, and bacterial resistance in an intensive care unit of

Morocco: Findings of the International Nosocomial Infection

Control Consortium (INICC) Int Arch Med. 2(1):29 .

Mahdi U and Koopmans M (2000) : Emerging group-A rotavirus

and a nosocomial outbreak of diarrhoea. Lancet ;356:1161–1162.

Maiden M.C.,. Bygraves J.A, Feil E. , Morelli G., Russell J.E.,

Urwin R , Zhang Q, Zhou J, Zurth K, Caugant DA, Feavers

IM, Achtman M, Spratt BG (1998) ; Multilocus sequence

typing: a portable approach to the identification of clones within

populations of pathogenic microorganisms, Proc. Natl Acad. Sci.

USA 95 (6) 3140–3145.

Maiden, M (2006) : Multilocus sequence typing of bacteria. Annu.

Rev. Microbiol. 60:561-588.

Maki DG, Stolz SM, Wheeler S, Mermel LA (1997) : Prevention

of central venous catheter-related bloodstream infection by use of

an antiseptic-impregnated catheter. A randomized, controlled trial.

Ann Intern Med;127:257-266.

Malek A, Sager R, Kuhn P , Nicolaides KH, Schneider H (1996) :

Evolution of maternofetal transport of immunoglobulins during

human pregnancy. Am J Reprod Immunol Nov; 36(5): 248–255.

Malhotra-Kumar S, Haccuria K, Michiels M , Ieven M, Poyart C,

Hryniewicz W, Goossens H (2008) : Current trends in rapid

diagnostics for methicillin-resistant Staphylococcus aureus and

glycopeptide-resistant enterococcus species. J Clin

Microbiol;46(5):1577–1587.

Mandell LA and Campbell GD Jr (1998) : Nosocomial pneumonia

guidelines: an international perspective. Chest;113:188-193.

Mangram AJ, Horan TC, Pearson ML, Silver LC, Jarvis WR

(1999) : Hospital Infection Control Practice Advisory Committee.

Guideline for prevention of surgical site infection,. Infect Control

Hosp Epidemiol 1999;20:247- 278.

Maragakis LL, Bradley KL, Song X, Beers C, Miller MR,

Cosgrove SE, Perl TM ( 2006): Increased catheter-related

bloodstream infection rates after the introduction of a new

mechanical valve intravenous access port. Infect Control Hosp

Epidemiol;27: 67- 70.

McCampbell B, Wasif N, Rabbitts A, Staiano-Coico L, Yurt RW,

Schwartz S (2002) : Diabetes and burns: retrospective cohort

study. J Burn Care Rehabil ;23(3):157–66.

McDonnell G. and Burke P (2011) : Disinfection: is it time to

reconsider Spaulding? Journal of Hospital Infection 78 , 163-170 .

McFarland LV, Beneda HW, Clarridge JE, Raugi GJ (2007) :

Implications of the changing face of Clostridium difficile disease

for health care practitioners. Am J Infect Control ;35:237 -253.

McFarland LV, Brandmarker SA, Guandalini S (2000) : Paediatric

Clostridium difficile: a phantom menace or clinical reality? J

Pediatr Gastroenterol Nutr ;31: 220-231.

McGeer A, Campbell B, Emori TG, Hierholzer WJ, Jackson MM,

Nicolle LE, Peppler C, Rivera A, Schollenberger DG, Simor AE

(1991) : Definitions of infection for surveillance in long-term care

facilities. Am J Infect Control;19:1-7.

McKibben L, Horan TC, Tokars JI, Fowler G, Cardo DM,

Pearson ML, Brennan PJ; Heathcare Infection Control

Practices Advisory Committee (2005) : Guidance on public

reporting of health care-associated infections: recommendations of

the Healthcare Infection Control Practices Advisory Committee.

Infect Control Hosp Epidemiol;26:580-587.

McNeil MM, Lasker BA, Lott TJ, Jarvis WR (1999) : Postsurgical

Candida albicans infections associated with an extrinsically

contaminated intravenous anesthetic agent. J Clin

Microbiol;37:1398-1403.

McNeil SA, Nordstrom-Lerner L, Malani PN, Zervos M,

Kauffman CA (2001) : Outbreak of sternal surgical site infections

due to Pseudomonas aeruginosa traced to a scrub nurse with

onychomycosis. Clin Infect Dis;33:317-323.

Medhat A, Shehata M, Magder LS , Mikhail N, Abdel-Baki L,

Nafeh M, Abdel-Hamid M, Strickland GT, Fix AD (2002) :

Hepatitis C in a community in Upper Egypt: risk factors for

infection. Am J Trop Med Hyg ; 66: 633–638.

Meky FA, Stoszek SK, Abdel-Hamid M , Selim S, Abdel-Wahab

A, Mikhail N, El-Kafrawy S, El-Daly M, Abdel-Aziz F, Sharaf

S, Mohamed MK, Engle RE, Emerson SU, Purcell RH, Fix AD,

Strickland GT (2006) : Active surveillance for acute viral

hepatitis in rural villages in the Nile Delta. Clin Infect Dis; 42:

628–633.

Mele JA, Linder SA, Calabria R, Ikeda CJ (1998) : HIV

seropositivity in a burn center’s population. J Burn Care

Rehabil;19(2):138–141.

Melling AC, Ali B, Scott EM, Leaper DJ ( 2001) : Effects of

preoperative warming on the incidence of wound infection after

clean surgery: a randomised controlled trial. Lancet ;35(8):876-

880.

Memmel H, Kowal-Vern A, Latenser BA (2004) : Infections in

diabetic burn patients. Diabetes Care ;27(1):229–33.

Mermel LA (2007) : Prevention of central venous catheter-related

infections: what works other than impregnated or coated catheters?

J Hosp Infect ;65:30-33.

Mischak, H (2007) : Clinical proteomics: a need to define the field

and to begin to set adequate standards. Proteomics Clin. Appl.

1:148-156.

Modol J, Sabria M, Reynaga E, Pedro-Botet ML, Sopena N,

Tudela P, Casas I, Rey-Joly C. (2007) : Hospital acquired

legionnaires disease in a university hospital: impact of the coppere

silver ionization system. Clin Infect Dis ;44:263-265.

Mojtahedzadeh M , Panahi Y , Fazeli MR , Najafi A , Pazouki

M, Navehsi BM , Bazzaz A , Naghizadeh MM , Beiraghdar F

(2008) : Intensive care unit-acquired urinary tract infections in

patients admitted with sepsis: etiology, risk factors, and patterns of

antimicrobial resistance International Journal of Infectious

Diseases 12, 312—318 .

Molina JM, Leport C, Bure A, Wolff M, Michon C, Vilde JL

(1991) : Clinical and bacterial features of infections caused by

Streptococcus milleri. Scand J Infect Dis;23:659-666.

Moore KL, Kainer MA, Badrawi N, et al (2001) : Early-onset

sepsis among neonatal intensive care unit patients-Cairo, Egypt,.

Abstract in Epidemic Intelligence Service. 51st Annual

Conference. 24(7):590-594.

Muarry PR, Baron EJ, Jorgensen JH et al (2003): Manual clinical

microbiology , ed 8 , Washington , 129 , 290-304 .

National Institute for Health and Clinical Excellence (2008) :

Surgical site infection, prevention and treatment of surgical site

infection. National Collaborating Centre for Women’s and

Children’s Health. 337.

Neeleman, C., Klaassen C. H. W., Valk H. A. de, de Ruiter M. T.,

and Mouton. J. W (2004) : Amplified fragment length

polymorphism fingerprinting is an effective technique to

distinguish Streptococcus pneumoniae from other streptococci and

efficient alternative to pulsed-field gel electrophoresis for

molecular typing of pneumococci. J. Clin. Microbiol. 42:369–371.

Nelson-Piercy C (2006) : Handbook of obstetric medicine, 3rd ed.

London: Taylor Francis.

Ng L.S.Y., The W.T. , Ng S.K., Eng L.C., Tan T.Y (2011) :

Bacterial contamination of hands and the environment in a

microbiology Laboratory / Journal of Hospital Infection 78 231-

233 .

Nicolas P, Yazdanpanah Y, Marie-Pierre C, et al (2007) :

Prevention of surgical site infection after breast cancer surgery by

targeted prophylaxis antibiotic in patients at high risk of surgical

site infection. J Surg Oncol ;96:124-129.

Nucci M and Anaissie E (2001) : Revisiting the source of

candidemia: skin or gut? Clin Infect Dis;33:1959-1967.

O’Grady NP, Alexander M, Dellinger EP, Gerberding JL, Heard

SO, Maki DG, Masur H, McCormick RD, Mermel LA, Pearson

ML, Raad II, Randolph A, Weinstein RA (2002) : Guidelines

for the prevention of intravascular catheter-related infections.

Centers for Disease Control and Prevention. MMWR Recomm

Rep;51:1-29.

O’Neill EO and Humphreys H (2005) : Surveillance of hospital

water and primary prevention of nosocomial legionellosis: what is

the evidence? J Hosp Infect ;59:273-279.

Oddie S and Embleton ND (2002) : Risk factors for early onset

neonatal group B streptococcal sepsis: casecontrol study. BMJ .

325(7359): 308.

Ogden, I.D (2007) : Use of multilocus sequence typing to

investigate the association between the presence of Campylobacter

spp. in broiler drinking water and Campylobacter colonization in

broilers. Appl. Environ. Microbiol. 73:5125-5129.

Olive, D. M., and Bean. P (1999) : Principles and applications of

methods for DNA-based typing of microbial organisms. J. Clin.

Microbiol. 37:1661– 1669.

Ortega M, Rovira M, Almela M, Marco F, de la Bellacasa JP,

Martínez JA, Carreras E, Mensa J (2005) : Bacterial and fungal

bloodstream isolates from 796 hematopoietic stem cell transplant

recipients between 1991 and 2000. Ann Hematol;84:40–46.

Ostrosky-Zeichner L, Baez-Martı´nez R, Rangel-Frausto S, Ponce

de Leon S (2000) : Epidemiology of nosocomial outbreaks: 14-

year experience at a tertiary-care center. Infect Control Hosp

Epidemiol ;21:527–529.

Ouslander JG, Yoshikawa TT, Mody L (2006) : Establishing an

infection control program. Infection management for geriatrics in

longterm care facilities. 2nd ed. New York, NY: Marcel Dekker,

Inc; . 115-130.

Owens C.D. and Stoessel K (2008) : Surgical site infections:

epidemiology, microbiology and prevention Kimberly-Clark

Healthcare, Atlanta, GA, USA Journal of Hospital Infection 70(2)

3–10 .

Page L (2005) : Coming soon: state reports on infection rates. Mater

Manag Health Care ;14:16-20.

Paterson DL (2006) : Resistance in gram-negative bacteria:

Enterobacteriaceae . Am J Med;119:20-28. discussion 62-70.

Pearson (2004) : Phage typing is the identification of bacterial

Species and strains by determining their susceptibility to various

phages www . Pearson education .com .

Pearson (2004) : schematic diagram of Southern Blot Analysis

www . Pearson education .com.

Pellowe CM ,Pratt RJ , Harper , Loveday HP, Robinson N, Jones

SR, MacRae ED, Mulhall A, Smith GW, Bray J, Carroll A,

Chieveley Williams S, Colpman D, Cooper L, McInnes E,

McQuarrie I, Newey JA, Peters J, Pratelli N, Richardson G,

Shah PJ, Silk D, Wheatley C (2003) : Evidence-based guidelines

for preventing health care associated infections in primary and

community care in England . J.Hosp.Infection ; 55(2) ; 61 -85 .

Pfaller MA, Pappas PG, Wingard JR (2006) : Invasive fungal

pathogens: current epidemiological trends. Clin Infect Dis;3-14.

Phillips L, Carlile J, Smith D (2004) : Epidemiology of a

tuberculosis outbreak in a rural Missouri high school.

Pediatrics ;113:514–519.

Philip W. Bennett G, Bradley S , Drinka P , Lautenbach E ,

Marx J , Mody L , Nicolle L , and Stevenson K (2008):

SHEA/APIC Guideline: Infection prevention and control in the

long-term care facility , Am J Infect Control ;36:504-35.

Pierce (2005) : Benjamin. Genetics: A Conceptual Approach, 2nd

ed. (New York: W. H. Freeman and Company), W. H. Freeman 271.

Pittet D, Allegranzi B, Sax H, Dharan S, Pessoa-Silva CL,

Donaldson L, Boyce JM (2006) : Evidence-based model for hand

transmission during patient care and the role of improved

practices. Lancet Infect Dis;6:641-652.

Pittet D, Harbarth SJ (1998) : The intensive care unit. In: Bennett

JV, Brachman PS, eds. Hospital infections, 4th ed. Philadelphia:

Lippincott-Raven,:381-402.

Platt R (2002) : Progress in surgical site infection surveillance.

Infect Control Hosp Epidemiol ;22:361-363.

Poirot JL, Gangneux JP, Fischer A , Malbernard M, Challier S,

Laudinet N, Bergeron V (2007) : Evaluation of a new mobile

system for protecting immune-suppressed patients against airborne

contamination. Am J Infect Control ;35:460-466.

Poon JT, Law W-L, Wong IW, Ching PT, Wong LM, Fan JK, Lo

OS (2009) : Impact of laparoscopic colorectal resection on

surgical site infection. Ann Surg ;249:77-81.

Pottinger JM, Herwaldt LA, Perl TM (1997) : Basics of

surveillance—an overview. Infect Control Hosp

Epidemiol;18:513–527.

Pratt RA , Pellowe CM ,Loveday HP, Robinson N, Smith GW,

Barrett S, Davey P, Harper P, Loveday C, McDougall C,

Mulhall A, Privett S, Smales C, Taylor L, Weller B, Wilcox M

(2001) : The Epic project ; developing national evidence based

guidelines for preventing health associated infection . J.

Hosp .Infect;47 . 3-82 .

Pruitt Jr BA (2002) : Epidemiological, demographic and outcome

characteristics of bur injury. In: Herndon DN, editor. Total burn

care.. London, UK: Saunders; . 16–30.

Puzniak L, Teutsch S, Powderly W, Polish L (2004) : Has the

epidemiology of nosocomial candidemia changed? Infect Control

Hosp Epidemiol;628-633.

Quinn JP (1998) : Clinical problems posed by multiresistant

nonfermenting gram-negative pathogens. Clin Infect

Dis;27(1):117-124.

Quintiliani, R., Jr., and P. Courvalin (1996) : Conjugal transfer of

the vancomycin resistance determinant vanB between enterococci

involves the movement of large genetic elements from

chromosome to chromosome. FEMS Microbiol. Lett. 119:359–364.

Rafla K,. Tredget E E (2010) : Infection control in the burn unit .

Burns 1- 11 .

Raman, R (2005) : Glycomics: an integrated systems approach to

structure- function relationships of glycans. Nat. Methods 2:817-

824.

Rebmann T and Linda R (2010): Preventing catheter-associated

urinary tract infections: An executive summary of the Association

for Professionals in Infection Control and Epidemiology, Inc,

Elimination Guide . Am J Infect Control ;38: 644-646.

Reed GH and Witter CT ( 2004) : Sensitivity and specificity of

single – nucleotide polymorphism scanning by high –resolution

melting analysis . Clin Chem ; 50 ;1748-1754 .

Reis J. N. , Coppola S. J.,. Gouveia E. L, Ko, A. I , Cordeiro SM,

Lôbo TS, Pinheiro RM, Salgado K, Ribeiro Dourado CM,

Tavares-Neto J, Rocha H, Galvão Reis M, Johnson WD Jr,

Riley LW (2000) : Clonally related penicillin-nonsusceptible

Streptococcus pneumoniae serotype 14 from cases of meningitis in

Salvador, Brazil. Clin. Infect. Dis. 30:78–86.

Riggs MM, Sethi AK, Zabarsky TF, Eckstein EC, Jump RL,

Donskey CJ (2007) : Asymptomatic carriers are a potential

source for transmission of epidemic and nonepidemic Clostridium

difficile strains among long-term care facility residents. Clin Infect

Dis ;45:992–998.

Rijen M V, Bonten M, Wenzel RP, Kluytmans (2008) : Intranasal

mupirocin for reduction of Staphylococcus aureus infections in

surgical patients with nasal carriage: a systematic review. J

Antimicrob Chemother ;61: 254-261.

Rijnders BJ, Van Wijngaerden E, Peetermans WE (2002) :

Catheter-tip colonization as a surrogate end point in clinical studies

on catheter-related bloodstream infection: how strong is the

evidence? Clin Infect Dis;35:1053 - 1058.

Rivera EV and Woods S (2003) : Prevalence of asymptomatic

Clostridium difficile colonization in a nursing home population: a

cross-sectional study. J Gend Specif Med ;6:27–30.

Roberts, R.J., Vincze, T., Posfai, J., Macelis, D (2010) : a database

for DNA restriction and modification: enzymes, genes and

genomes. Nucleic Acids Res. 38, 234–236.

Ronaghi, M (2001) : Pyrosequencing sheds light on DNA

sequencing. Genome Res. 11:3-11.

Rosenthal VD, Guzman S, Safdar N (2005) : Reduction in

nosocomial infection with improved hand hygiene in intensive care

units of a tertiary care hospital in Argentina. Am J Infect

Control;33:392-397.

Rosenthal VD, Maki DG, Salomao R, Moreno CA, Mehta Y,

Higuera F, Cuellar LE, Arikan OA, Abouqal R, Leblebicioglu

H (2006): International Nosocomial Infection Control

Consortium: Device- Associated nosocomial infections in 55

intensive care units of 8 developing countries. Ann Intern Med,

145:582-591.

Rosser CJ, Bare RL, Meredith JW (1999) : Urinary tract infections

in the critically ill patient with a urinary catheter. Am J Surg;177:

287-290.

Rotstein C , Evans G, Born A , Grossman R, Light R B , Magder

S , McTaggart B , Weiss K , Zhanel G (2008): Clinical

practice guidelines for hospital-acquired pneumonia and ventilator-

associated pneumonia in adults Can J Infect Dis Med Microbiol 19

(1) 19-53 .

Rupp ME, Fitzgerald T, Marion N, Helget V, Puumala S,

Anderson JR, Fey PD (2004) : Effect of silver coated urinary

catheters: efficacy, cost-effectiveness, and antimicrobial resistance.

Am J Infect Control ; 32:445- 450.

Rushbrook P , Pru¨ss A, Giroult E (1999) : Safe management of

wastes from health-care activities. Geneva: 2 .16-20 .

Rutala MWA and Weber DJ (2004) ; Disinfection and sterilization

in health care facilities: what clinicians need to know. Clin Infect

Dis ;39:702-709.

Safdar N and Maki DG (2006) : Use of vancomycin-containing

lock or flush solutions for prevention of bloodstream infection

associated with central venous access devices: a metaanalysis of

prospective, randomized trials. Clin Infect Dis ;43:474 -484.

Sands K, Vineyard G, Livingston J, Christiansen C, Platt R

(1999) : Efficient identification of postdischarge surgical site

infections using automated medical records. J Infect Dis;179;434 -

441.

Samore M. H., Kristjansson M., Venkataraman L., DeGirolami P.

C., and Arbeit. R. D (1996) : Comparison of arbitrarily primed

polymerase chain reaction, restriction enzyme analysis and pulsed-

field gel electrophoresis for typing Clostridium difficile. J.

Microbiol. Methods 25:215.

Santos AML, Lacerda RA, Graziano KU (2005) : Evidence of

control and prevention of surgical site infection by shoe covers and

private shoes e a systemic literature review [in Portuguese]. Rev

Lat Am Enfermagem;13:86-92.

Sattar SA , Springthorpe VS , TetroJ , Vashon R, Keswick B

(2002) : Hygienic hand antiseptics should they not have activity

and label claims against viruses? ; Am.J.Contr ., 30;355-372.

Satterfield N (1993) : Infection control in long-term care facilities:

the hospitalbased practitioner’s role. Infect Control Hosp

Epidemiol;14:40-47.

Sauer, S. and M. Kliem (2010) : Mass spectrometry tools for the

classification and identification of bacteria. Nat. Rev. Microbiol.

8:74-82.

Scholtes-Timmerman, Lin, B., Malanoski, A.P., et al (2009): A

novel approach to correct specific variations in Raman spectra due

to bleachable cellular components. Analyst 134(2):387-93..

Schouten JP , McElgunn CJ , Waaijer R , Zwijnenburg D ,

Diepvens F , Pals G (2002) : Relative quantification of 40

nucleic acids sequences by multiplex ligation-dependent probe

amplification . Nucleic Acids Res ; 30 ; 57 .

Scrimshaw NS (1989) : Malnutrition and nosocomial infection Infect

Control Hosp Epidemiol 10:191-193.

Sepkowitz KA ( 1996) : Occupationally acquired infections in

healthcare workers. Part I. Ann Intern Med;125:826-834.

Sheng W S , Wang JT , Lin MS , Chang SC (2007) : Risk Factors

Affecting In-hospital Mortality in Patients with Nosocomial

Infections Formos Med Assoc 106 (2) .110-118 .

Siebor E, Llanes C, Lafon I, Ogier-Desserrey A, Duez JM,

Pechinot A, Caillot D, Grandjean M, Sixt N, Neuwirth C (2007)

: Presumed pseudobacteremia outbreak resulting from

contamination of proportional disinfectant dispenser. Eur J Clin

Microbiol Infect Dis;26:195–198.

Simon AC (2004) : Hand hygiene, the crusade of the infection

control specialist. Alcohol-based handrub: the solution! Acta Clin

Belg;59:189-193.

Simonsen L, Kane A, Lloyd J, Zaffran M, Kane M. (1999) :

Unsafe injections in the developing world and transmission of

bloodborne pathogens: a review. Bull World Health Organ ; 77:

789–800.

Smyth ETM, Humphreys H, Stacey A, Taylor EW, Hoffman P,

Bannister G (2005) : Survey of operating theatre ventilation

facilities for minimally invasive surgery in Great Britain and

Northern Ireland: current practice and considerations for the future.

J Hosp Infect ;61:112-122.

Singh A, Goering RV , Simjee S ,Foley S, Zervos MJ (2006):

Application of Molecular Techniques to the Study of Hospital

Infection Clin. Microbiol. Rev American Society for Microbiology

19 : 512–530 .

Song S, Itawi EA, Saber AA (2009) : Natural orifice translumenal

endoscopic surgery (NOTES). J Invest Surg ;22:214-217.

Stabler, R.A (2006) : Comparative phylogenomics of Clostridium

difficile reveals clade specificity and microevolution of

hypervirulent strains. J. Bacteriol. 188:7297-7305.

Stampone, L., M. Del Grosso, D. Boccia, and A. Pantosti (2005) :

Clonal spread of a vancomycin-resistant Enterococcus faecium

strain among bloodstream- infecting isolates in Italy. J. Clin.

Microbiol. 43:1575–1580.

Stanley J, Desai M, Efstratiou A, George R. (1997) : High-

resolution genotyping of Streptococcus pyrogenes: Application to

outbreak studies and population genetics. In: Horaud T, Bouvet A,

Leclercq R, de Montclos H, Sicard M, eds. Streptococci and the

host. New York: Plenum Press, 37(6):1948-1952.

Starakis J, Marangos M, Gikas A, Pediaditis I, Bassaris H.

(2002) : Repeated point prevalence survey of nosocomial

infections in a Greek University Hospital. J Chemother;14:272-

278.

Stephenson, F.H (2004) : Calculations in Molecular Biology and

Biotechnology: A Guide to Mathematics. Academic Press,

Burlington, MA.

Stoll BJ, Hansen NI, Adams-Chapman I , Fanaroff AA, Hintz SR,

Vohr B, Higgins RD (2004) : Neurodevelopmental and growth

impairment among extremely low-birth-weight infants with

neonatal infection. JAMA Nov 17; 292(19): 2357–2365.

Stoszek SK, Abdel-Hamid M, Narooz S , Narooz S, El Daly M,

Saleh DA, Mikhail N, Kassem E, Hawash Y, El Kafrawy S,

Said A, El Batanony M, Shebl FM, Sayed M, Sharaf S, Fix AD,

Strickland GT (2006) : Prevalence of and risk factors for hepatitis

C in rural pregnant Egyptian women. Trans R Soc Trop Med Hyg ;

100: 102–107.

Struelens MJ (1996) : Consensus guidelines for appropriate use and

evaluation of microbial epidemiological typing systems. Clin.

Microbiol. Infect. 2:1–10.

Struelens, M (2002) : Molecular typing: a key tool for the

surveillance and control of nosocomial infection, Curr. Opin.

Infect. Dis. 15 (4) 383–385.

Struelens, M.J., De Ryck, R., Deplano, A (2001) : Analysis of

microbial genomic macrorestriction patterns by pulsed-field gel

electrophoresis (PFGE) typing. In: Dijkshoorn, L., Towner, K.J.,

Struelens, M. (Eds.), New Approaches for the Generation and

Analysis of Microbial Typing Data. Elsevier, Amsterdam, 159-

176.

Struelens M J , Denis O , Rodriguez-Villalobos H (2004) :

Microbiology of nosocomial infections: progress and challenges .

6(11):1043-1048.

Szczuka, E., and Kaznowski. A (2004) : Typing of clinical and

environmental Aeromonas sp. strains by random amplified

polymorphic DNA PCR, repetitive extragenic palindromic PCR,

and enterobacterial repetitive intergenic consensus sequence PCR.

J. Clin. Microbiol. 42:220-228.

Taban M, Behrens A, Newcomb RL, Nobe MY, McDonnell PJ

(2005) : Incidence of acute endophthalmitis following penetrating

keratoplasty: a systematic review. Arch Ophthalmol ;123(5):605-

609.

Tablan OC, Anderson LJ, Besser R, Bridges C, Hajjeh R, CDC ;

Healthcare Infection Control Practices Advisory Committee

(2003) : Guidelines for preventing health-care-associated

pneumonia : recommendations of CDC and the Healthcare

Infection Control Practices Advisory Committee. MMWR

Recomm Rep 2004;53:1-36.

Talaat, M,a Kandeel A, , Rasslan O, Hajjeh R ,, Hallaj Z , El-

Sayed N, and Mahoney F (2006) : Evolution of infection control

in Egypt: Achievements and challenges Am J Infect

Control;34:193-200.

Talaat M, Hafez S , Saied T , Elfeky R, El-Shoubary W, and

Pimentel G (2009) : Surveillance of catheter-associated urinary

tract infection in 4 intensive care units at Alexandria university

hospitals in Egypt . 38(3):222-228.

TambyahPA, KnasinskiV, Maki DG (2002) : The direct costs of

nosocomial catheter- associated urinary tract infection in the era of

managed care. Infect Control Hosp Epidemiol ;23:27–31.

Tanner J, Woodings D, Moncaster K (2006) : Preoperative hair

removal to reduce surgical site infection. Cochrane Database Syst.

19;(2):4122.

Tavanti .A, Gow N.A. , Senesi S. , Maiden MC, Odds FC (2003) :

Optimization and validation of multilocus sequence typing for

Candida albicans, J. Clin. Microbiol. 41 (8) 3765–3776.

Tenover, F. C., Arbeit R. D., Goering R. V (1997) : How to select

and interpret molecular strain typing methods for epidemiological

studies of bacterial infections: a review for healthcare

epidemiologists. Infect. Control Hosp. Epidemiol. 18:426–439.

Thal, L. A., Silverman J., Donabedian S., and. Zervos. M. J (1997)

: The effect of Tn916 insertions on contour-clamped homogenous

electrophoresis patterns of the Enterococcus faecalis. J. Clin.

Microbiol. 35:969–972.

Thal, L. A., Donabedian S. M., Robinson-Dunn B., J. W. Chow, L.

Dembry L, Clewell DB, Alshab D, Zervos MJ (1998) :

Molecular analysis of glycopeptide-resistant Enterococcus faecium

isolates collected from Michigan hospitals over a 6-year period. J.

Clin. Microbiol. 36:3303– 3308.

Torpdahl M, Sorensen G, Lindstedt B, Moller Nielsen E

(2007) :Tandem repeat analysis for surveillance of human

Salmonella Typhimurium infections. Emerg Infect Dis;13(3):388–

395.

Tredget EE, Shankowsky HA, Joffe AM, Moskal TI, Volpel K,

Paranchych W, Kibsey PC, Alton JD, Burke JF (1992) :

Epidemiology of infections with Pseudomonas aeruginosa in burn

patients: the role of hydrotherapy. Clin Infect Dis;15(6):941–949.

Tredget EE, Shankowsky HA, Rennie R, Burrell RE, Logsetty

(2004) : S. Pseudomonas infections in the thermally injured

patient. Burns;30(1):3–26.

Velasco E, Byington R, Martins CS, Schirmer M, Dias LC,

Goncalves VM (2004) : Bloodstream infection surveillance in a

cancer centre: a prospective look at clinical microbiology aspects.

Clin Microbiol Infect;10:542–549.

Vianelli N, Giannini MB, Quarti C, Bucci Sabattini MA, Fiacchini

M, de Vivo A, Graldi P, Galli S, Nanetti A, Baccarani M, Ricci

P (2006) : Resolution of a Pseudomonas aeruginosa outbreak in a

hematology unit with the use of disposable sterile water filters.

Haematologica.;91:983–985.

Vincent JL (2004) : Ventilator-associated pneumonia. J Hosp

Infect ;57:272-280.

Visalli MA, Jacobs MR, Appelbaum PC (1998) : Activities of

three quinolones, alone and in combination with extended-

spectrum cephalosporins or gentamicin, against Stenotrophomonas

maltophilia. Antimicrob Agents Chemother ;42:2002-2005.

Viudes A, Peman J, Canton E, Ubeda P (2002) : Candidemia at a

tertiary-care hospital: epidemiology, treatment, clinical outcome

and risk factors for death. Eur J Clin Microbiol Infect Dis;767—74.

Vonberg R.-P. , and Gastmeier P (2006) : Hospital-acquired

infections related to contaminated substances . 65(1):15-23.

Wagenlehner FM, Loibl E, Vogel H, Naber KG (2006) : Incidence

of nosocomial urinary tract infections on a surgical intensive care

unit and implications for management. Int J Antimicrob

Agents;28:86-90.

Waisbourd-Zinman O, Ben-Ziony S, Solter E, Scherf E, Samra Z,

Ashkenazi S (2009) : Hospitalizations for nosocomial rotavirus

gastroenteritis in a tertiary pediatric center: a 4-year prospective

study. Am J Infect Control;37:465-469 .

Walsh TJ, Pizzo PA. Bodey GP, et al (1993) : Candidiasis:

pathogenesis, diagnosis, and treatment, 2nd ed. New York: Raven

Press, : Nosocomial candidemia in adults: Risk and prognostic

factors 109-135.

Wang KW, Chang WN, Huang CR, Tsai NW, Tsui HW (2005) :

Postneurosurgical nosocomial bacterial meningitis in adults:

microbiology, clinical features, and outcomes. J Clin

Neurosci;12:647-650.

Wang, D., Urisman, A., Liu, Y.T., Springer M, Ksiazek TG,

Erdman DD, Mardis ER, Hickenbotham M, Magrini V, Eldred

J, Latreille JP, Wilson RK, Ganem D, DeRisi JL (2003) : Viral

discovery and sequence recovery using DNA microarrays. PLoS

Biol. 1, 2.

Watson L, Wilson BJ, Waugh N (2002) : Pneumococcal

polysaccharide vaccine: a systematic review of clinical

effectiveness in adults. Vaccine;20:2166-2173.

Weber JM, Sheridan RL, Schulz JT, Tompkins RG, Ryan CM

( 2002) : Effectiveness of bacteria-controlled nursing units in

preventing cross-colonization with resistant bacteria in severely

burned children. Infect Control Hosp Epidemiol ;23(9):549–551.

Weber JM and Tompkins DM (1993) : Improving survival:

infection control and burns. AACN Clin Issues Crit Care

Nurs ;4(2):414–423.

Weber JM (1998) : Epidemiology of infections and strategies for

control. In: Carrougher GJ, editor. Burn care therapy. St. Louis,

MO: Mosby Inc.; . 185–211.

Weisfelt M. , Beek D. , Spanjaard L (2007) : a Nosocomial

bacterial meningitis in adults: a prospective series of 50 cases

Journal of Hospital Infection 66, 71-78 .

Weisner AM, Johnson AP, Lamagni TL , Arnold E, Warner M,

Heath PT, Efstratiou A (2004) : Characterization of group B

streptococci recovered from infants with invasive disease in

England and Wales. Clin Infect Dis 38(9): 1203–1208.

Wenzel RP (1995) : The economics of nosocomial infections. J

Hosp Infect, 31:79–87.

Wenzel RP (2007) ; Health care-associated infections: major issues

in the early years of the 21st century. Clin Infect Dis ; 45:85-88.

Whatmore, A. M., Murphy T. J., Shankster S., Young E., Cutler

S. J., and Macmillan A. P (2005) : Use of amplified fragment

length polymorphism to identify and type Brucella isolates of

medical and veterinary interest. J. Clin. Microbiol. 43:761–769.

Whitby M, McLaws ML, Berry G (2001) : Risk of death from

methicillin resistant Staphylococcus aureus bacteraemia: a

metaanalysis. Med J Aust ;1 75:264-267.

White L, Dancer SJ, Robertson C, McDonald J (2008) : Are

hygiene standards useful in assessing infection risk? Am J Infect

Control ;36:381-384.

WHO (2002): Prevention of hospital acquired infections. a practical

guide.2nd edition. WHO/CDS/CSR/ EPH/2002.12.

WHO (2006) : guidelines for hand hygiene in health care (Advanced

draft). Geneva: World Health Organization .

WHO (2007) : guidelines for hand hygiene in health care . Geneva:

World Health Organization.

WHO (2008) : The global burden of health care-associated infections

World Health Organization .

WHO (2009) : Guidelines on Hand Hygiene

inHealthCare ,May2009.Availableat .

Widmer AF (2000) : Replace hand washing with use of a waterless

alcohol hand rub? Clin Infect Dis;31:136-143.

Wilhelmus KR and Hassan SS (2007) : The prognostic role of

donor corneoscleral rim cultures in corneal transplantation.

Ophthalmology;114(3):440–445 .

Wintergerst ES, Maggini R, Hornig DH (2007) : Contribution of

selected vitamins and trace elements to immune function. Ann Nutr

Metab;51:301-323.

Worth LJ. , Slavin M A (2009) : Bloodstream infections in

haematology: Risks and new challenges for prevention Blood

Reviews 23 113–122 .

Wright M O (2008) , Automated surveillance and infection control:

Toward a better tomorrow , Am J Infect Control , 36 (3) 2-6 .

Yamada M, Mochizuki H, Yamada K (2002) : Aqueous humor

levels of topically applied levofloxacin in human eyes. Curr Eye

Res 24:403–406,

Yassin S, Fahmi N, Kandeel A, Mansour E, Mohsen L, Gipson R,

et al (2003) : Evaluation and monitoring of infection control

training in neonatal intensive care units in Egypt. Abstract. Fourth

Congress of the International Federation of Infection Control

Malta, 9-12, .

Yip E (2004) : Consideration of barrier protection and latex protein

allergy in the evaluation of medical gloves. J Infus Nurs ;27:227-

231.

Zawacki A, O’Rourke E, Potter-Bynoe G, et al (2004) : An

outbreak of Pseudomonas aeruginosa pneumonia and bloodstream

infection associated with intermittent otitis externa in a healthcare

worker. Infect Control Hosp Epidemiol ; 25:1083-1089.