SUCCESS! In Clinical Laboratory Science Bacteriology - Anaerobic Bacteria

An anaerobically incubated blood agar plate shows colonies surrounded by an inner zone of complete red cell lysis and an outer zone of incomplete cell lysis (double zone of hemolysis). The most likely presumptive identification of this isolate would be A. Clostridium perfringens B. Clostridium tetani C. Fusobacterium nucleatum D. Prevotella melaninogenica

A. Isolates of the anaerobic, spore-forming bacillus Clostridium perfringens characteristically produce a pattern of double zone hemolysis on sheep blood agar plates. A Gram stain of such colonies should demonstrate a mediumsized gram-positive bacillus that does not contain spores. For further identification the isolate should be inoculated on an egg yolk agar plate to detect lecithinase production.

Growth on kanamycin-vancomycin laked blood agar incubated anaerobically is primarily used for A. Bacteroides fragilis B. Bifidobacterium dentium C. Clostridium perfringens D. Peptostreptococcus anaerobius

A. Kanamycin-vancomycin laked blood (KVLB) agar is selective for the Prevotella and Bacteroides spp. Presumptive identification of B. fragilis group can be accomplished utilizing its antimicrobial resistance pattern. Bacteroides spp. are resistant to vancomycin and kanamycin, unlike Fusobacterium spp., which are resistant to vancomycin but susceptible to kanamycin. A KVLB agar plate should be part of the primary plating media for anaerobic cultures.

The potentially lethal intoxication type of food poisoning often associated with improperly canned food is caused by A. Bacteroidesfragilis B. Clostridium botulinum C. Clostridium perfringens D. Clostridium septicum

B. Improperly home-canned foods, especially low-acid-content vegetables, cause the majority of the cases of food-borne botulism. The ubiquitous nature of Clostridium botulinum enables the spores to contaminate a variety of foods, Contamination and subsequent germination under anaerobic conditions stimulate toxin formation. The patient becomes ill following the ingestion of food that contains nanograms of preformed toxin.

An infant was seen in the emergency department with symptoms of neuromuscular weakness and constipation. The diagnosis of infant botulism was confirmed by the demonstration of toxin in the child's stool. The child most likely contracted this disease by A. A scratch wound caused by a cat B. Ingestion of spores that germinated in the intestine C. A puncture wound with a contaminated household item D. Ingestion of preformed toxin found in a contaminated jar of pureed vegetables

B. Infant botulism or "floppy infant" syndrome is seen in children up to 6 months of age. This infectious process begins with the ingestion of food contaminated with spores of Clostridium botulinum. Following ingestion, viable spores are carried to the lower bowel, where they germinate and elaborate the powerful neurotoxin that produces the characteristic flaccid paralysis.

To ensure that anaerobic conditions have been achieved in anaerobic jars or chambers, an oxygen-sensitive indicator is employed, such as A. Bromcreosol purple B. Methylene blue C. Methyl red D. Phenol red

B. Methylene blue strips are the most commonly used oxidation-reduction (Eh) indicators. When anaerobic conditions are achieved, the methyleneblue indicator will turn from blue (oxidized) to white, indicating reduction. Resazurin, another Eh indicator, is used in anaerobic transport systems and anaerobic culture media such as the prereduced anaerobically sterilized (PRAS) system. Resazurin when oxidized is pink; when reduced, the color fades to white, indicating anaerobiosis.

Gram-positive bacilli with central spores are seen in the direct Gram stain of a tissue biopsy. After 24 hours of incubation, no growth is seen on the sheep blood agar plate incubated aerobically and the chocolate agar plate incubated in increased CO2. Which of the following is the likely cause of the infection? A. Bacillus B. Clostrdium C. Lactobacillus D. Prevotella

B. Most Clostridium spp. are gram-positve, and they generally form spores. Because they are obligate anaerobes, they will not grow on sheep blood or chocolate agars incubated aerobically. Bacillus spp. also form spores, but they are facultative anaerobes and would therefore grow on media incubated aerobically. Lactobacillus is a non-spore-forming, gram-positive bacillus, and Prevotella is a gram-negative bacillus.

After 72 hours of anaerobic incubation, small olive-green to black colonies are seen. A Gram stain reveals gram-postive cocci. What is the most likely identification of this organism? A. Fingoldia magna B. Peptococcus niger C. Peptostreptococcus anaerobius D. Veillonella parvula

B. Peptostreptococcus niger produces a pigment that begins olive-green and gradually becomes black. This is the only species in the genus. It is a weak pathogen sometimes found in polymicrobial infections.

Aspiration of vomitus can lead to pneumonia. Which of the following would not be a likely causative agent in aspiration pneumonia? A. Bacteroides gracilis B. Mobiluncus sp. C. Porphyromonas sp. D. Prevotella melaninogenica

B. The common agents in cases of aspiration pneumonia are oral anaerobes, such as the blackpigmented Prevotella and Porphyromonas, and Bacteroides, fusobacteria, and anaerobic streptococci. These endogenous organisms, when in an abnormal site, possess virulence factors that enable them to produce disease. Often these are polymicrobic infections mixing anaerobes with aerobic or facultative organisms such as Enterobactericeae or Staphylococcus aureus. Mobiluncus is not associated with aspiration pneumonia.

The characteristic colony morphology of Actinomyces israelii on solid agar resembles A. "Medusa head" B. A molar tooth C. A fried egg D. Ground glass

B. The gram-positive, non-spore-forming, anaerobic bacillus Actinomyces israelii is a slowly growing organism that is considered to be an opportunistic pathogen. Colonies may not be visible before 5 to 7 days or longer. When colonies are seen, they appear white, opaque, lobate, irregular, and shiny and are described as resembling a molar tooth. A. israelii is part of the indigenous flora of the human mouth, and a few Actinomyces spp. have been found to inhabit the vagina. Pathogenesis generally involves trauma to tissues of a mucous membrane and the introduction of this endogenous organism.

Which of the following is described as obligately anaerobic gram-positive cocci? A. Capnocytophaga B. Peptostreptococcus C. Propionibacterium D. Veillonella

B. The second most commonly encountered group of anaerobes in human infections is the anaerobic, gram-positive cocci. They may account for one-fourth of all anaerobes isolated in clinical laboratories. Estimating their clinical significance, however, is often difficult. Important isolates include Fingoldia magna (formerly Peptostreptococcus magnus) and Peptostreptococcus anaerobius.

Which of the following is not considered a zoonotic disease? A. Anthrax B. Botulism C. Brucellosis D. Leptospirosis

B. Zoonotic diseases are diseases of animals that are transmissible to humans. Leptospirosis is primarily a disease of small animals such as rabbits. It is contracted by humans through contact with infected carcasses or contaminated water. Bacillus anthracis is found in the environment. Anthrax is transmitted to humans by exposure to contaminated animal products such as cattle hides, goat hair, or wool. Brucellosis is associated with a variety of animals

Which of the following is an important virulence factor of Bacteroides fragilis? A. Endotoxin B. Exotoxins C. Polysaccharide capsule D. Protease

C. Bacteroides fragilis stimulates abscess formation. The capsule is a contributing factor to the pathology produced by this anaerobe. B. fragilis is the most common anaerobic gram-negative bacillus isolated in the clinical laboratory.

The diagnosis of pseudomembranous colitis (Clostridium difficile associated disease) is often made by A. Serology B. Culturing blood specimens C. Assays to detect toxin in stool D. Acid-fast stain of fecal material

C. C. difficile is an important cause of a hospital acquired infection commonly called pseudomembranous colitis. Hospitalized patients treated with broad-spectrum antimicrobial agents become colonized when their normal intestinal flora is diminished. The most rapid and accurate diagnostic method is detecting toxins A and/or B in stool specimens. Cycloserine-cefoxitin-fructose agar (CCFA) is the recommended selective medium for C. difficile, although few laboratories attempt isolation

Which of the following statements is not characteristic of Clostridium botulinum? A. Infant botulism is the most common clinical form. B. Pathogenicity is related to a potent neuro toxin. C. Oval spores are located terminally. D. Of the seven toxogenic types, types A, B, E, and F are associated with human botulism.

C. Clostridium botulinum is the causative agent of botulism, a disease produced by an exotoxin that acts on the central nervous system. Types A, B, E, and F are causes of human botulism; types C and D and less commonly types A and B are associated with disease in animals and birds. Type G has not been associated with disease in humans oranimals. This anaerobic organism produces oval, central, or subterminal, spores that germinate in food products or less commonly in wounds.

Which of the following is an important cause of food poisoning? A. Bacteroides fragilis B. Bacteroides ureolyticus C. Clostridium perfringens D. Clostridium histolyticum

C. Clostridium perfringens is one of the most important causes of food-borne diseases in the U.S. The bacterial spores can survive cooking (typically found in meats and gravies), and upon cooling they germinate into vegetative cells. When the bacteria are ingested, they sporulate in the intestinal tract. The enterotoxin is a spore coat protein made in excess and released by the bacteria.

Which of the following statements is true regarding Clostridium perfringens ? A. There are five serologic types. B. Spores are terminally located. C. Alpha-toxin is produced by all strains. D. Spores are readily seen in laboratory media.

C. Clostridium perfringens produces spores that are oval and central in location but that are rarely seen in foods or on laboratory cultures. This organism is divided into five types, A to E, based on the quantities and types of exotoxins produced. Type A is responsible for human cases of myonecrosis and food poisoning. Alpha-toxin or lecithinase is produced by all strains of C. perfringens.

Which anaerobic, gram-negative rod can be presumptively identified by its Gram stain morphology, and inhibition by bile and a 1-ug kanamycin disk? A. Bacteroides fragilis group B. Eubacterium lentum C. Fusobacterium nucleatum D. Porphyromonas gingivalis

C. Fusobacterium nucleatum characteristically appears on Gram stain as a gram-negative rod with pointed ends. Its growth is inhibited by a 1-ug kanamycin disk and the presence of bile. The Bacteroides fragilis group and the pigmented species Prevotella and Porphyromonas are not inhibited by kanamycin.

A curved appearance on Gram stain is characteristic of which of the following? A. Actinomyces israelii B. Clostridium septicum C. Fusobacterium nucleatum D. Propionibacterium acnes

C. Fusobacterium nucleatum is a thin gramnegative rod with pointed ends and a slightly curved appearance in fresh isolates. As the bacteria are subcultured, they may lose their curved appearance and appear as thin rods. F. nucleatum is found in human specimens and is considered clinically significant.

The gram-negative, non-spore-forming, anaerobic bacillus frequently implicated in serious clinical infections such as brain and lung abscesses is A. Bacteroides urealyticus B. Eubacterium lentum C. Fusobacterium nucleatum D. Peptostreptococcus anaerobius

C. Fusobacterium nucleatum, a gram-negative, anaerobic bacillus, is part of the indigenous microbial flora of the respiratory, gastrointestinal, and genitourinary tracts. It is frequently implicated as the causative agent in metastatic suppurative infections such as brain abscesses. These pale-staining bacilli characteristically appear as long, thin bacilli with pointed ends.

Which bacterium is part of the normal vaginal flora that helps resist the onset of bacterial vaginosis? A. Peptostreptococcus sp. B. Peptococcus sp. C. Lactobacillus sp. D. Mobiluncus sp.

C. Lactobacillus spp. are found as normal flora in the gastrointestinal and female genital tract. The bacteria produce acids from the metabolism of carbohydrates, resulting in an acid environment in the vagina. If the population of lactobacilli decreases, the vaginal pH will rise toward neutrality. This favors the growth of other bacteria, such as Mobiluncus, that can result in bacterial vaginosis.

Small alpha-hemolytic colonies are seen on a sheep blood agar plate after 48 hours of incubation on a vaginal culture. This describes which of the following? A. Bacteroides B. Clostridium C. Lactobacillus D. Porphyromonas

C. Lactobacillus spp. are normal flora of the vagina and digestive tract and are rarely pathogenic. They are aerotolerant anaerobes and will produce alpha-hemolysis on sheep blood agar plates incubated aerobically. These organisms can also produce a green discoloration on chocolate agar.

Which one of the following is a non-spore-forming, gram-positive, anaerobic bacillus? A. Clostridium B. Fusobacterium C. Propionibacterium D. Veillonella

C. Propionibacterium spp. are non-spore-forming, anaerobic, gram-positive bacilli. Clostridium spp. typically form spores, although it is difficult to induce some species to form spores in vitro. Veillonella is a gram negative coccus, and Fusobacterium is a gram negative bacillus.

Which of the following tests is most appropriate for the identification of Clostridium difficile? A. SPS sensitivity test B. Naglertest C. Cytotoxin assay D. Fluorescence test

C. The symptoms of Clostridium difficile infection are toxin mediated. This organism is known to cause pseudomembranous colitis associated with the use of antimicrobial therapy. Cytotoxins can be directly detected in stools by enzyme immunoassays. These tests have generally replaced the use of cell culture monolayers, which were examined after incubation for evidence of cytotoxicity (i.e., cytopathic effect).

The anaerobic, gram-negative, curved, motile bacilli associated with bacterial vaginosis belong to the genus A. Actinomyces B. Bifidobacterium C. Lactobacillus D. Mobiluncus

D. Although it has a gram-positive-like cell wall, Mobiluncus stains gram-variable to gram-negative. This curved and motile bacillus seems to contribute to the pathology of bacterial vaginosis (BV). A Gram stain of the discharge that is produced in this condition can be used for the detection of these distinctively curved organisms. The presence of "clue cells," gram-variable pleomorphic bacilli on vaginal epithelial cells, is diagnositic of BV.

Purulent material from a cerebral abscess was submitted to the laboratory for smear and culture. On direct Gram stain, grampositive cocci in chains and gram-negative bacilli with pointed ends were seen. Plates incubated aerobically exhibited no growth at 24 hours. On the basis of the organisms seen on the smear, what is the most likely presumptive identification of the etiologic anaerobic agents? A. Veillonella sp. and Clostridium sp. B. Eubacterium sp. and Veillonella sp. C. Peptostreptococcus sp. and Nocardia sp. D. Fusobacterium sp. and Peptostreptococcus sp

D. Anaerobes are a major cause of brain abscess. Peptostreptococcus spp. are associated with human disease, usually in polymicrobial infections, and can be seen on a Gram stain of clinical material. The characteristic Gram stain morphology of Fusobacterium would enable a physician to make a presumptive identification of the presence of anaerobic flora in this clinical case.

Which is a correct statement regarding Clostridium tetani? A. It is proteolytic B. It is lecithinase positive C. It is characteristically nonmotile D. It produces terminal spores

D. Clostridium tetani is a strict anaerobe that is motile and produces terminal round spores. Biochemically it does not utilize carbohydrates, with the rare exception of glucose. C. tetani is gelatinase and indole positive but is nonproteolytic and H2S negative. The clinical manifestations of tetanus are the result of the release of a neurotoxic exotoxin.

Which of the following statements is not true of clostridia? A. Pseudomembranous colitis is due to a toxin produced by C. difficile. B. Clinically significant clostridia are found in the normal flora of the colon and in the soil. C. Botulism is caused by ingesting preformed toxin and can be prevented by boiling food prior to eating. D. C. tetani spores will form in the presence of oxygen; therefore, anaerobiosis in a wound is not required to cause tetanus.

D. Clostridium tetani is an obligate anaerobe. Spores are widespread in the soil. When introduced into a puncture wound, the spores require the reduced oxygen environment produced by the necrotic tissue and poor blood supply in the wound. Cleaning and debridement of the wound is important, as is the administration of a tetanus toxoid booster. Many clostridia require anaerobic conditions for spore formation.

Which of the following organisms is not gram-positive? A. Eubacterium lentum B. Bifidobacterium dentium C. Propionibacterium acnes D. Suttonella wadsworthensis

D. Eubacterium, Bifidobacterium, and Propionibacterium are all anaerobic, gram-positive, non-spore-forming bacilli. This group of anaerobic microorganisms is difficult to identify in the clinical laboratory and often requires the use of gas chromatography. These organisms are rarely isolated. Suttonella wadsworthensis is an anaerobic, gram-negative bacillus.

Which of the following tests is most appropriate for the presumptive identification of Prevotella melaninogenica? A. SPS sensitivity test B. Naglertest C. Cytotoxin assay D. Fluorescence test

D. Prevotella melaninogenica can be rapidly presumptively identified on media containing laked blood with the use of an ultraviolet light source. This important anaerobic pathogen can be differentiated after 5 to 7 days' incubation by its black pigmentation. The use of ultraviolet light enables a more rapid differentiation because of the appearance of a brick red fluorescence before the pigment is demonstrated.

Egg yolk agar showing a precipitate in the medium surrounding the colony is positive for A. Lecithinase production B. Lipase production C. Protease activity D. Starch hydrolysis

A. An area of precipitate in the agar around the colonies indicates that the organism produced lecithinase. Lecithinase (alpha-toxin) cleaves lecithin in the medium, producing an insoluble product. Clostridium perfringens is positive for lecithinase.

Which of the following is the most potent bacterial exotoxin known? A. Botulinum toxin B. Erythrogenic toxin C. C. difficile toxin B D. C. perfringens alpha-toxin

A. Botulinal toxin is the most potent exotoxin known. When absorbed, this exotoxin produces the paralyzing disease botulism. Toxin acts in the body by blocking the release of acetylcholine in the neuromuscular junction of the peripheral nervous system, causing muscle paralysis.

Which one of the following is not true of Clostridium tetani ? A. It produces rapid tissue necrosis. B. It is a gram-positive, spore-forming bacillus. C. Microorganisms in soil contaminate puncture wounds. D. Disease is caused by an exotoxin acting on the central nervous system.

A. Clostridium tetani is an obligate anaerobe. Spores are widespread in nature and cause disease by contaminating puncture wounds. The exotoxin, tetanospasmin, produced by this organism is one of the most powerful bacterial toxins known.

Identify the Fusobacterium sp. considered to be the most frequent isolate recovered from clinical infections. A. F. varium B. F. nucleatum C. F. mortiferum D. F. necrophorum

B. Fusobacterium nucleatum is the most frequent clinical isolate within the genus Fusobacterium. These anaerobes are part of the indigenous flora of human mucous membranes, oral cavity, intestine, and urogenital tract. F. necrophorum is, however, much more virulent.

Obligately anaerobic, gram-negative bacilli, recovered from an abdominal wound, were found to be resistant to penicillin. Growth of this organism was not inhibited by bile. What is the most likely identification of this isolate? A. Bacteroides fragilis group B. Clostridium septicum C. Eubacterium lentum D. Fusobacterium nucleatum

A. Members of the Bacteroides fragilis group, the most commonly isolated anaerobes and a predominant part of the indigenous fecal flora in humans, are not inhibited by the presence of bile. Bile-esculin agar plates are used for the selection and presumptive identification of the B. fragilis group. Although not used as a component of selection media for the Bacteroides group, it is important to note that, in general, gram-negative, non-spore-forming, anaerobic bacilli are susceptible to penicillin. The B. fragilis group is an exception in that it is known to be resistant to penicillin.

Color Plate 29 • shows the filamentous gram-positive rod recovered from an aspirate of a closed chest abscess. It grew only under anaerobic conditions and was not acid-fast. What is the most likely presumptive identification of the isolate seen? A. Actinomyces israelii B. Bacteroides fragilis C. Clostridium septicum D. Propionibacterium acnes

A. The closed chest abscess described is characteristic of human actinomycosis, which is caused by Actinomyces israelii, an anaerobic, gram-positive, non-spore-forming bacillus. The organism is not acid-fast, which helps to differentiate it from Nocardia spp. Actinomycotic pus characteristically shows "sulfur granules" or solid yellow particles made up of masses of the filamentous bacilli seen on the Gram stain in Color Plate 29 •.

Which of the following tests is most appropriate for the presumptive identification of Peptostreptococcus anaerobius? A. SPSdisk B. Colistindisk C. Kanamycin disk D. Vancomycin disk

A. The identification of Peptostreptococcus anaerobius is made easier by the use of the sodium polyethanol sulfonate (SPS) disk. The test is performed by growing the organism in the presence of a disk impregnated with SPS. A zone of inhibition of 12-18 mm around the disk is considered sensitive and a presumptive identification of this organism.

What is the predominant indigenous flora of the colon? A. Anaerobic, gram-negative, non-sporeforming bacteria B. Anaerobic, gram-positive, non-sporeforming bacteria C. Aerobic, gram-negative, non-sporeforming bacteria D. Aerobic, gram-positive, spore-forming bacteria

A. The predominant indigenous flora of the human intestinal tract is anaerobic, gram-negative, non-spore-forming bacilli. The Bacteroides fragilis group, in particular, predominates in the fecal flora. Trauma involving the intestinal area or bowel surgery predisposes patients to an endogenous anaerobic infection. Although these organisms are present in large numbers, their routine identification in fecal cultures is of no diagnostic value.

Which of the following tests is most appropriate for the presumptive identification of Clostridium perfringens? A. SPS sensitivity test B. Reverse CAMP test C. Cytotoxin assay D. Esculin hydrolysis

B. A reverse CAMP test aids in the identification of Clostridium perfringens. In this test, a single straight streak of Streptococcus agalactiae is made down the center of the plate. Suspected C. perfringens isolates are inoculated at right angles to the S. agalactiae inoculum. After anaerobic incubation, C. perfringens will exhibit enhanced hemolysis at the intersection where the two species meet.

Which Clostridium sp. is most commonly recovered from cases of gas gangrene? A. C. bifermentans B. C. perfringens C. C. sordellii D. C. difficile

B. Clostridium perfringens is the species most commonly associated with clostridial myonecrosis or gas gangrene. These soil and water saprophytes most frequently gain entrance to the human body through traumatic wounds. Once they have been introduced into injured tissue, the characteristic syndrome of myonecrosis due to the elaboration of exotoxins may occur. Other species involved with myonecrosis are C. septicum, C. novyi, C. sordellii, and C. histolyticum.

Septicemia caused by which of the following is generally associated with an underlying malignancy? A. Bifidobacterium dentium B. Clostridium septicum C. Eubacterium lentum D. Lactobacillus catenaforme

B. Clostridium septicum is isolated in the clinical laboratory in cases of serious or often fatal infections. Bacteremia is seen in association with an underlying maglignancy. The most common types of cancer are colon or cecum, breast, and leukemia or lymphoma.

A cervical mucosal abscess specimen was sent to the laboratory for bacteriologic examination. The culture of this sample grew an anaerobic gram-negative bacillus that was inhibited by bile, produced a black pigment, and was negative for indole production and positive for glucose, sucrose, and lactose fermentation. This isolate would most likely be A. Bacteroides fragilis group B. Bacteroides ureolyticus C. Porphyromonas gingivalis D. Prevotella melaninogenica

D. Prevotella melaninogenica was isolated from this cervical abscess. This anaerobic organism is part of the indigenous microflora of the respiratory, gastrointestinal, and genitourinary tracts and is considered a significant human pathogen. The black pigment appeal's after several days when growing on laked blood agar plates. Prior to pigmentation, this isolate can be presumptively identified by its brick-red fluorescence under UV light. Pigmented Porphyromonas spp. are asaccharolytic

The majority of the gram positive, nonspore-forming, anaerobic bacilli isolated from clinical material will be A. Bifidobacterium dentium B. Capnocytophagia ochracea C. Eubacterium limosum D. Propionibacterium acnes

D. Propionibacterium acnes is the most frequently isolated of all the gram positive, nonspore-forming, anaerobic bacilli. It is a part of the normal human bacterial flora and predominates on the surface of the body, but may also be recovered from the upper respiratory tract, intestines, and urogenital tract. This organism is a common contaminant of blood cultures because of its presence on the skin. Care in the preparation of the skin before venipuncture helps to eliminate confusion caused by the recovery of this anaerobic isolate

A tube of semisolid medium that contains resazurin appears pink. What does this indicate? A. Acid environment B. Alkaline environment C. Motility D. Presence of oxygen

D. Resazurin is an Eh indicator used in anaerobic culture media. When the oxygen concentration is reduced, the resazurin indicator is colorless. A pink color in the medium indicates aeration and an unsuitable environment for the preservation of obligate anaerobic organisms.

Which of the following is not true of Bacteroides fragilis ? A. Lipase and lecithinase negative B. Anaerobic gram-negative bacillus C. Commonly associated with intraabdominal infections D. Among the most antimicrobialsensitive anaerobic bacteria

D. The Bacteroides fragilis group is among the most antimicrobial-resistant anaerobes. Betalactamase production is responsible for their resistance to the penicillins. These anaerobes are also resistant to first-generation cephalosporins and aminoglycosides.

Which of the following clostridia has a terminal spore that causes the cell to swell? A. C. botulinum B. C. difficile C. C. perfringens D. C. tetani

D. The spore of Clostridium tetani is located terminally and is larger than the sporangium. Characteristically, when seen on Gram stain, the cells of Clostridium tetani resemble a drumstick or tennis racket. Spores can be readily seen in late growth phase cultures incubated at 37°C.