ANAEROBICANAEROBICBACTERIOLOGYBACTERIOLOGY

Dr. Ma. Ellery M. Mendez Dr. Ma. Ellery M. Mendez FPAMS , FPAAM, DPSMAP FPAMS , FPAAM, DPSMAP

Anaerobes generate energy by Anaerobes generate energy by fermentationfermentation

Lack the capacity to utilize O2 as a Lack the capacity to utilize O2 as a terminal hydrogen acceptorterminal hydrogen acceptor

Some are sensitive to O2 concentration Some are sensitive to O2 concentration as low as 0.5% O2as low as 0.5% O2

Most can survive in 3%-5% O2Most can survive in 3%-5% O2 A few can grow poorly in the presence of A few can grow poorly in the presence of

air air aerotolerant anaerobes aerotolerant anaerobes Many are members of the normal floraMany are members of the normal flora

created by presence of facultative created by presence of facultative

anaerobesanaerobes

FACTORS THAT INHIBIT THE FACTORS THAT INHIBIT THE GROWTH OF ANAEROBES BY GROWTH OF ANAEROBES BY OXYGENOXYGEN

1. Toxic compounds are produced1. Toxic compounds are produced e.g. H2O2 , Superoxidese.g. H2O2 , Superoxides 2. Absence of catalase & Superoxide 2. Absence of catalase & Superoxide dismutasedismutase

3. Oxidation of essential sulfhydyl groups in 3. Oxidation of essential sulfhydyl groups in enzymes without sufficient reducing power enzymes without sufficient reducing power to regenerate themto regenerate them

FACTORS RESPONSIBLE FOR THEIR FACTORS RESPONSIBLE FOR THEIR VIRULENCEVIRULENCE

1. Lipopolysaccharide1. Lipopolysaccharide - promotes abscess formation, enhanced coagulation- promotes abscess formation, enhanced coagulation

2. Polysaccharide capsule2. Polysaccharide capsule - correlated with abscess production- correlated with abscess production

3. Enzymes3. Enzymes a. Collagenasea. Collagenase b. Heparinaseb. Heparinase * develop thrombophlebitis & septic emboli* develop thrombophlebitis & septic emboli 4. Short chained fatty acids4. Short chained fatty acids a. Butyrate- a. Butyrate- seen in dental plaque seen in dental plaque

b. succinic acid – b. succinic acid – reduces phagocytic killingreduces phagocytic killing

Multiplication of the opportunistic Multiplication of the opportunistic pathogens is facilitated by:pathogens is facilitated by:

1. 1. inhibition of phagocytosis & intracellular killing inhibition of phagocytosis & intracellular killing by PMN in the presence of Bacteroides by:by PMN in the presence of Bacteroides by: a. competition of opsoninsa. competition of opsonins b. inhibition by capsular materialsb. inhibition by capsular materials 2. protection of antibiotic susceptibility strains in 2. protection of antibiotic susceptibility strains in

mixtures thru destruction by the ß-lactamasesmixtures thru destruction by the ß-lactamases

3. utilization of O2 by facultative species that aids in 3. utilization of O2 by facultative species that aids in producing a suitable environment for growth of producing a suitable environment for growth of anaerobe anaerobe

Anaerobic Bacteria of Medical Anaerobic Bacteria of Medical InterestInterest

MORPHOLOGY GRAM STAIN GENUS

Sporeforming (+) Clostridium

Non-sporeforming bacilli

(+)

(-)

Actinomycetes,Bifidobacterium,Eubacte-rium,Propionibacerium,Mobilncus,Lactobacillus

Bacteroides,FusobacteriumPrevotella,Porphyromonas

Non-sporefoming cocci (+)

(-)

Peptococcus,Pepto-streptococcusStreptococcus

Veilonella

CLINICAL MANIFESTATIONCLINICAL MANIFESTATION

Clinical hints Clinical hints 1. odor1. odor 2. tissue 2. tissue 3. location3. location 4. necrotic tissue4. necrotic tissue 5. endocarditis with (-) blood culture5. endocarditis with (-) blood culture 6. infection associated with malignancy6. infection associated with malignancy 7. black discoloration7. black discoloration 8. blood containing exudates8. blood containing exudates 9. associated with sulfur granules 9. associated with sulfur granules 10. Bacteremic feature with jaundice10. Bacteremic feature with jaundice 11. human bites11. human bites

Sites and Infection produced by Sites and Infection produced by Anaerobes Anaerobes

LABORATORY DIAGNOSISLABORATORY DIAGNOSISA. COLLECTIONA. COLLECTION Anaerobes are endogenous in natureAnaerobes are endogenous in nature

I. Appropriate specimens for anaerobicI. Appropriate specimens for anaerobic

culture :culture :

1. pus 1. pus

2. pleural fluid2. pleural fluid

3. urine3. urine

4. pulmonary secretions4. pulmonary secretions

5. uterine secretions or sinus tract 5. uterine secretions or sinus tract materialmaterial

II. Collection by needle aspiration is II. Collection by needle aspiration is preferrable than swab culture because preferrable than swab culture because ofof

a. better survival of pathogena. better survival of pathogen b. greater quantity of specimenb. greater quantity of specimen c. less contamination with extraneous c. less contamination with extraneous organism are often achievedorganism are often achieved

B. HANDLINGB. HANDLING If a swab must be used, a 2 tube If a swab must be used, a 2 tube

systemsystem

must be usedmust be used 11stst tube contains swab in O2 free CO2 tube contains swab in O2 free CO2

22ndnd tube contains PRAS (pre-reduced tube contains PRAS (pre-reduced

anaerobically steilized culture media)anaerobically steilized culture media)

Specimen should be placed in Specimen should be placed in anaerobic transport device with anaerobic transport device with gas mixturegas mixture

C. IsolationC. Isolation

Gram stain should be done in the Gram stain should be done in the

laboratory :laboratory :

a. choice of appropriate media &a. choice of appropriate media &

methods for culturemethods for culture

b. quality control for the types of b. quality control for the types of

bacteria that laboratory culture bacteria that laboratory culture

revealreveal

A solid or liquid medium maybe used & must provide anA solid or liquid medium maybe used & must provide an anaerobic environmentanaerobic environment Anaerobic Culture SystemAnaerobic Culture System

A.A. ANAEROBIC JARANAEROBIC JAR 1. Candle Jar1. Candle Jar

- reduces O2 environment- reduces O2 environment

- only - only ↑↑ CO2 tension CO2 tension

2. Gas Pak Jar2. Gas Pak Jar

a. Palladium aluminum coated pelletsa. Palladium aluminum coated pellets

- catalyst- catalyst

- chemically reduces O2- chemically reduces O2

- reacts with residual O2 in the presence of H2 to - reacts with residual O2 in the presence of H2 to form form

H2OH2O

b. Gas Pak envelope b. Gas Pak envelope

- generates CO2 & H2 gases- generates CO2 & H2 gases

c. Methylene blue strip c. Methylene blue strip

- indicator- indicator

blue blue → → (+) O2(+) O2

white white →→ (-) O2 (-) O2

II. Anaerobic Glove ChamberII. Anaerobic Glove Chamber - close system- close system

- used for premature babies- used for premature babies

- e.g. incubator - e.g. incubator

III. Roll TubeIII. Roll Tube - has a pedal - has a pedal gas ( CO2 & H2 ) would come out gas ( CO2 & H2 ) would come out

- place test tube directly to the outlet- place test tube directly to the outlet

D. IDENTIFICATIOND. IDENTIFICATION Plates are checked at Plates are checked at

> 18-24 hours for faster growing species like> 18-24 hours for faster growing species like

Cl. Perfringens & B.fragilis Cl. Perfringens & B.fragilis & daily thereafter up to& daily thereafter up to

> > 5-7 days for slowly growing species like5-7 days for slowly growing species like

Actinomyces, Eubacterium & propionibacterium Actinomyces, Eubacterium & propionibacterium

GenusGenus is determined by is determined by

- gram stain, cellular morphology, Gas-liquid - gram stain, cellular morphology, Gas-liquid

chromotographychromotography

Species Species determination is based on fermentation of determination is based on fermentation of sugars & other biochemical determinationsugars & other biochemical determination

TREATMENTTREATMENT

- - Susceptibility testing should be doneSusceptibility testing should be done

- surgical drainage & resection of necrotic tissue- surgical drainage & resection of necrotic tissue

- most are resistant to aminoglycosides- most are resistant to aminoglycosides

- for Bacteroides gr- for Bacteroides gr oup, if resistant to Penicillin & oup, if resistant to Penicillin & Cephalosporin, they may use:Cephalosporin, they may use:

a. Clindamycin a. Clindamycin

b. Metronidazole b. Metronidazole

c. Chloramphenicolc. Chloramphenicol

THE ENDTHE END