1. Anaerobic Bacteria Dr. Yasir. A. Hussein, MD, Microbiology & Pathology Update, September 2016

2. Overview 1.Introduction to anaerobic bacteria. 2.Types of anaerobic bacteria. 3.Inhabitant of anaerobic bacteria. 4.Classification of anaerobic bacteria. 5.Pathogenesis of anaerobic bacteria. 6.Predisposing Factor of anaerobic bacteria. 7.Clinical feature of anaerobic bacteria. 8.Diagnosis & treatment of anaerobic bacteria.

3.  Medically important infections due to anaerobic bacteria are common.  Its major part of indigenous flora in human body.  The infections are often polymicrobial that is the anaerobic bacteria are found in mixed infections with other anaerobes, facultative anaerobes and aerobes.  Several important diseases are caused by anaerobic Clostridium species from the environment or from normal flora. Introduction:

4. Types of Anaerobes  Anaerobes that live outside the body are called “exogenous anaerobes” (Example: Clostridium species)  Anaerobes that live inside the body are called “endogenous anaerobes” ◦ Advantages: barrier to colonization by pathogenic organisms, source of fatty acids, vitamins and cofactors. ◦ Disadvantages: Opportunists for immunocompromised. An absence of intestinal bacteria is associated with reductions in cell mucosal muscle wall thickness, cytokine production, and digestive enzyme activity and with defective cell-mediated immunity.  Most anaerobic infections are from endogenous sources.

5. Types of Anaerobic Infections Usually endogenous ◦ Intestinal anaerobes ◦ Oral anaerobes Usually exogenous ◦ Clostridium tetani (tetanus) ◦ Clostridium botulinum (botulism) ◦ Clostridium difficile (colitis) Either endogenous or exogenous ◦ Other Clostridial infections (e.g., gas gangrene)

6. Inhabitant of anaerobic infections:  Today they are found in soil, fresh and salt water, and in normal flora of humans and animals  Mucosal surfaces such as linings of oral cavity, GI tract, and GU tract  Respiratory Tract – 90% of bacteria in the mouth are anaerobes  If mucosal surfaces are disturbed, infections can occur in the oral cavity and in aspiration pneumonia  Skin – frequently normal skin anaerobes contaminate blood cultures.  GU Tract – anaerobes rarely cause infection.  GI Tract – Approximately 2/3’s of all bacteria are in the stool.

7. Role of Endogenous Anaerobes Role in normal host physiology ◦ Prevent colonization & infection by pathogens ◦ Interference with adhesion ◦ Essential for the development and function of the immune system during early life. ◦ Contributes to host physiology ◦ E. coli synthesizes vitamin K and metabolize bile acids

8. Types of Anaerobes ObligateAero-tolerantFacultative

9. Classification of Medically Important Anaerobes

10. Anaerobic microorganisms Obligate (strict) anaerobes ◦ Grow ONLY in the absence of molecular oxygen ◦ Killed by exposure to air Aerotolerate (moderate) anaerobes Can tolerate exposure to air for several hours but perform metabolic activities Facultative anaerobes ◦ Do not require O 2, but use it if available

11. Obligate anaerobes  Obligate anaerobes need an oxygen-free environment to live. They cannot grow in places with oxygen, which can sometimes damage and destroy them.  Obligate anaerobes are major components of the normal microflora on mucous membranes, especially of the mouth, lower GI tract, and vagina.  These anaerobes cause disease when normal mucosal barriers break down.  Ex. Clostridium perfringens, Bacteroides fragilis

12. Facultative anaerobes  Facultative anaerobes use aerobic respiration in the presence of oxygen; without oxygen, some of them ferment; some use anaerobic respiration.aerobic respiration  Some species in the Staphylococcus genus are facultative, and are a leading cause of blood poisoning.  One example is Staphylococcus aureus, which produces a range of infections from common skin problems like acne, boils, and impetigo to acute conditions like meningitis, endocarditis, and pneumonia.  It also causes toxic shock syndrome (TSS).  Another facultative anaerobe is Escherichia coli, which contains many helpful serotypes as well as some damaging ones, such as those that cause food poisoning.

13. Aero-tolerant bacteria  Aero-tolerant bacteria do not use oxygen to live, but can exist in its presence.  Aero-tolerant organisms are strictly fermentative (lactobacillus).

14. Pathogenesis & Transmission  Endogenous route:  Hematogenous.  Direct invasion  Secretion of mucous in oral cavity  Aspiration of mucus  Penetration of organism to peritoneal cavity.  Exogenous:  Penetration of organism through injury

15. Pathogenesis of anaerobic Attachment and adhesion ◦ Polysaccharide capsules and pili Invasion ◦ Alteration in host tissue (trauma, surgery, disease) Establishment of infection ◦ Polysaccharide capsule (B. fragilis) ◦ Spore formation (Clostridium) Tissue damage ◦ Elaboration of toxins

17. General Features of Anaerobic Infection : Brain abscess. Upper respiratory tract infection. Retropharyngeal abscess. Chronic otitis media Eye infection Dental abscess Gingivitis.

18.  Lower respiratory tract infection:  Pneumonia.  Abdominal infection: Peritonitis Intraabdominal abscess Hepatic abscess Splenic abscess. Appendicitis Irritable bowel syndrome Diverticulitis. Biliary tract infection.

19.  Female Genital tract infections: Perineal abscess Vulvar abscess Endometritis Ovarian abscess Pelvic inflammatory disease  Skin infections: Skin ulcer Cellulitis.  Bone infection: osteomyelitis  Neonatal infection: Conjunctivitis, Aspiration pneumonia

20. Clinical Features of anaerobic infections

21. Diagnosis: Specimens for Anaerobic Culture Aspirates ◦ Should be collected with needle and syringe ◦ Excess air expressed from syringe ◦ Specimen injected into oxygen-free transport tube or vial.

22. Specimens for Anaerobic Culture Tissue ◦ must be placed in an oxygen-free transport bag or vial ◦ Usually ground for best results Blood ◦ aerobic & anaerobic bottles are collected for most blood culture requests ◦ History & physical examination are too important

23. Processing Clinical Samples for Anaerobic Culture Must be placed in an anaerobic chamber or holding device while awaiting processing Procedures ◦ Macroscopic exam of specimen ◦ Foul odor ◦ Presence of “sulfur granules” ◦ Black pigmentation ◦ Gram stain ◦ Distinct morphology

24. Typical Anaerobic Media Anaerobic blood agar (BRU/BA) ◦ Supports growth of all obligate and facultative anaerobes Bacteroides bile esculin agar (BBE) ◦ Supports growth of bile-tolerant anaerobes, such as Bacteroides, Prevotella, Porphyromonas, Fusobacterium species Kanamycin-vancomycin-laked blood agar (KVLB) ◦ Supports growth of Bacteroides and Prevotella spp.; certain facultative gnr will also grow Phenylethyl alcohol agar (PEA) ◦ Supports growth of all obligate and gram positive facultative anaerobes, inhibits enteric gnr ◦ Cycloserine-cefoxitin-fructose agar (CCFA) ◦ Selective for C. difficile Anaerobic broth, such as thioglycollate (THIO) or chopped meat ◦ Supports growth of all types of bacteria; obligate aerobes near the top, obligate anaerobes at the bottom and facultative anaerobes throughout

25. Anaerobic Media Bacteroides fragilis on KVLB (left) and BBE agar (right )

26. Indications of Anaerobes in Cultures  Foul odor when opening anaerobic jar or bag  Colonies on anaerobically incubated media but not on aerobic media  Good growth on BBE  Colonies on KVLB that are pigmented or fluorescent  Double zone of hemolysis on blood agar

27. Interpretation of Cultures  Primary cultures are examined after 48 hours of incubation If no growth, reincubate for up to 5 days before discarding.

28. Treatment Surgical therapy ◦ Draining abscesses, removing dead tissue, eliminating obstructions Hyperbaric oxygen ◦ Oxygen is forced into necrotic tissues, killing anaerobes Antitoxins ◦ Used in cases of tetanus and botulism to neutralize the neurotoxins produced by C. tetani and C. botulinum ◦ Metronidazole. ◦ Antibiotics (Chloramphenicol, Penicillin, Ampicillin). ◦ Ticarcillin plus clavulanate, Amoxicillin plus sulbactam, Piperacillin plus tazobactam) and Carbapenems (imipenem, meropenem and ertapenem). ◦

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