Bacteroides fragilis Case File

Bacteroides fragilis Case File

Eugene C.Toy, MD, Cynthia Debord, PHD, Audrey Wanger, PHD, Gilbert Castro, PHD, James D. Kettering, PHD, Donald Briscoe, MD

CASE 2

A 60-year-old man presents to the emergency room with severe abdominal pain. He has had mild, left lower abdominal cramping pain for about 3 days, which has worsened in the past 8 hours. He has also had nausea, fever, and chills. On examination, he is in obvious pain, has a fever of 38.6°C (101.5°F) and has an elevated heart rate (tachycardia). His abdominal examination is notable for absent bowel sounds, diffuse tenderness, and rigidity when palpated. An x-ray reveals the presence of free air in the abdominal cavity. He is taken for emergency surgery and found to have severe diverticulitis with a perforated colon. Cloudy peritoneal fluid is collected. An anaerobic culture grows Bacteroides fragilis.

◆ What characteristics are noted on Gram staining of B. fragilis?

◆ What are its primary mechanisms for resisting phagocytosis?

ANSWERS TO CASE 2: Bacteroides fragilis

Summary: A 60-year-old man has a ruptured diverticulitis leading to peritonitis. The cultures of the purulent drainage reveal B. fragilis.

◆ Characteristics on gram staining: Bacteroides fragilis appears encapsulated, with irregular staining, pleomorphism, and vacuolization.

◆ Primary mechanisms of resisting phagocytosis: The capsular polysaccharide and succinic acid production.

CLINICAL CORRELATION

Bacteroides fragilis is one of the most clinically significant anaerobic organisms. It is part of the normal flora of the gastrointestinal (GI) tract and causes clinical infections when it escapes from this environment following surgery, traumatic bowel perforation, or other diseases, such as diverticulitis. Although many anaerobes are part of the normal gastrointestinal flora, B. fragilis is the most common cause of intraabdominal infections. Bacteroides fragilis is also associated with respiratory tract infections (sinusitis, otitis), genital tract infections, brain, skin, and soft tissue infections.

Diverticulitis is an inflammation of a small food and particle collecting sac in the large intestine; it may lead to colonic rupture and therefore allow the organisms normally present in the GI tract to penetrate the peritoneal cavity and possibly the bloodstream. These infections usually involve a mixture of both aerobes and anaerobes.

APPROACH TO Bacteroides fragilis

Objectives

1. Know the microbiologic characteristics of B. fragilis and other Bacteroides species.
2. Know the virulence factors associated with B. fragilis.

Definitions

Anaerobes: Organisms that do not require oxygen for growth and may die in its presence.

Bacteroides bile esculin (BBE) agar: Media selective for B. fragilis on which the colonies appear black.

Diverticulitis: Inflammation of a diverticulum, which is a small bulging sac in the colon wall which can trap food particles and become inflamed and painful.

DISCUSSION

Characteristics of Bacteroides Species

Bacteroides species include the B. fragilis group as well as many other species. Two new genera were recently created, Prevotella and Porphyromas, to remove the pigmented, bile-sensitive anaerobes previously in the genus Bacteroides. All are small, anaerobic, gram-negative bacilli and many strains are encapsulated. Vacuolization, irregular staining, and pleomorphism are common.

Bacteroides fragilis has a distinct capsule composed of two polysaccharides, which appears to inhibit phagocytosis and allow adherence to peritoneal surfaces. Other virulence factors for this bacterium include the presence of pili, which promote adherence to epithelial cells and the production of succinic acid, which inhibits phagocytosis. Bacteroides fragilis produces an endotoxin that has little biologic activity. It also produces superoxide dismutase, an enzyme, which allows the organism to survive in the presence of small amounts of oxygen.

Diagnosis

Anaerobes are not usually the primary cause of an infection, but are involved in a mixed aerobic, anaerobic infection. Often diagnosis of anaerobic infections is based on clinical features including a foul smelling wound with the presence of gas in the involved tissue usually located in close proximity to a mucosal surface. Infections that involve spillage of GI material into the peritoneum are likely to involve aerobes and anaerobes. The most commonly associated anaerobe is B. fragilis. Patients with severe diverticulitis, appendicitis, or colonic injury often develop B. fragilis peritonitis.

Bacteroides species produce small colonies on anaerobic blood agar medium within 24 hours. Selective media such as kanamycin/gentamicin laked blood agar will support growth of gram-negative anaerobes only. Presumptive identification of B. fragilis can be made by growth of black pigmented colonies on Bacteroides bile esculin agar, and resistance to kanamycin, colistin, and vancomycin special potency antimicrobial disks. Definitive identification of anaerobes or B. fragilis is made with commercial identification systems that are based on the presence of preformed enzymes or in reference laboratories using gas liquid chromatography to determine the specific gases produced by the organism.

Treatment and Prevention

Surgical debridement is usually necessary at least in part for the treatment of anaerobic infections. β-Lactamase activity is common in Bacteroides species, especially B. fragilis, which results in resistance to penicillin and cephalosporin antibiotics. Drugs of choice for Bacteroides species include β-lactam–β-lactamase inhibitor combinations, such as piperacillin/tazobactam, metronidazole, and imipenem.

COMPREHENSION QUESTIONS

[2.1] During an emergency surgery, a 60-year-old male is found to have severe peritonitis and a perforated colon. Foul-smelling cloudy peritoneal fluid is collected. Subsequent analysis reveals the growth of black pigmented colonies on Bacteroides bile esculin agar. No growth is detected in the presence of kanamycin, colistin, or vancomycin. Which of the following microorganisms is most likely involved in this case?

A. Actinomyces israelii

B. Bacteroides fragilis

C. Clostridium difficile

D. Enterococcus faecalis

E. Porphyromonas gingivalis

F. Prevotella melaninogenica

[2.2] Which of the following is the treatment of choice to control this infection in this patient (described in Question [2.1])?

A. Cephalothin

B. Erythromycin

C. Metronidazole

D. Penicillin

[2.3] Among the many virulence factors produced, B. fragilis produces an enzyme that allows the organism to survive in the presence of small amounts of oxygen. Which of the enzymes listed below catalyzes the following reaction?

2O₂- + 2H+ → H₂O₂ + O₂

A. β-Lactamase

B. Myeloperoxidase

C. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase

D. NO synthase

E. Oxidase

F. Superoxide dismutase

[2.4] A foul-smelling specimen was obtained from a 26-year-old woman with a pelvic abscess. Culture grew both aerobic and anaerobic gram-negative bacteria. The most likely organisms are which of the following?

A. Actinomyces israelii and Escherichia coli

B. Bacteroides fragilis and Listeria monocytogenes

C. Bacteroides fragilis and Neisseria gonorrhoeae

D. Clostridium perfringens and Bacteroides fragilis

E. Escherichia coli and Peptostreptococcus

Answers

[2.1] B. Bacteroides species are normal inhabitants of the bowel and other sites. Normal stools contain large numbers of B. fragilis (1011 organisms per gram). As a result, they are very important anaerobes that can cause human infection. Members of the B. fragilis group are most commonly isolated from infections associated with contamination by the contents of the colon, where they may cause suppuration, for example, peritonitis after bowel injury. Classification is based on colonial and biochemical features and on characteristic short-chain fatty acid patterns in gas chromatography. These short-chain fatty acids also contribute to the foul-smelling odor emanating from the wound in the above case.

[2.2] C. Metronidazole, mainly used as an antiprotozoal agent, is also highly effective against anaerobic bacterial infections, such as those infections caused by Bacteroides species. It is the drug of first choice for GI strains of Bacteroides. Two other effective antibiotics are imipenem and piperacillin/tazobactam. Bacteroides species, such as B. fragilis, commonly possess β-lactamase activity resulting in resistance to penicillin and cephalosporin (e.g., cephalothin) antibiotics. Erythromycin is not indicated in the treatment of Bacteroides species.

[2.3] F. A key feature of obligate anaerobes such as Clostridium, Bacteroides, and Actinomyces is that they lack catalase and/or super-oxide dismutase (SOD) and are therefore susceptible to oxidative damage. Bacteroides fragilis, however, is able to survive (not grow) in environments with low oxygen content because of its ability to produce small amounts of both SOD and catalase. Anaerobes that possess SOD and/or catalase are able to negate the toxic effects of oxygen radicals and hydrogen peroxide and thus tolerate oxygen. Other common enzymes listed above catalyze the following reactions:

Catalase/superoxide dismutase catalyzes: 2H₂O₂ → 2H₂O + O₂

Myeloperoxidase catalyzes: Cl- + H₂O₂ → ClO- + H₂O

NADPH oxidase catalyzes: NADPH + 2O₂ → 2O₂- + H+ NADP+

NO synthase catalyzes: ½O₂ + arginine → NO + citrulline

Oxidase catalyzes: 2H+ + 2e- + ½O₂ → H₂O

[2.4] C. In infections, such as intra-abdominal abscesses, Bacteroides species are often associated with other organisms. The only other organism in the list above that is solely aerobic and gram-negative is N. gonorrhoeae. Clostridium and Listeria are both gram-positive. E. coli is gram-negative and a facultative anaerobe.

MICROBIOLOGY PEARLS ❖ The treatment of choice of B. fragilis is surgical debridement in addition to metronidazole, imipenem, or piperacillin/tazobactam. ❖ Most anaerobes are part of mixed infections at mucosal surfaces. ❖ Bacteroides fragilis is the most common anaerobe in the human GI tract. ❖ Bacteroides fragilis usually express superoxide dismutase, an enzyme, which allows the organism to survive in the presence of small amounts of oxygen.

REFERENCES

Brook I, Frazier EH. Aerobic and anaerobic microbiology in intraabdominal infections associated with diverticulitis. J Med Microbiol 2000;49:827–30. 

Engelkirk PG, Duben-Engelkirk JD, Dowell VR. Principles and Practice of Clinical Anaerobic Bacteriology. Belmont, CA: Star Publishing, 1992. 

Murray PR, Rosenthal KS, Pfaller MA. Anaerobic gram negative bacilli. In: Murray PR, Rosenthal KS, Pfaller MA. Medical Microbiology, 5th ed. St. Louis, MO: Mosby, 2005:421–26.

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