Miscellaneous and Fastidious Gram-Negative Rods Flashcards
A visitor to South America who returned
with diarrhea is suspected of being infected with V. cholerae. Select the best medium for recovery and identification of this organism.
A. MacConkey agar
B. Blood agar
C. TCBS agar
D. XLD agar
C
The growth of yellow or green colonies on the selective TCBS agar (thiosulfate citrate bile salts sucrose) is dependent on whether the organism ferments sucrose (producing yellow colonies). Vibrio also grow well on 5% sheep blood, chocolate, and MacConkey agars. Enrichment with alkaline peptone broth, pH 8.4, helps in recovering Vibrio spp. from stool specimens.
A curved gram-negative rod producing oxidase-positive colonies on blood agar was recovered from a stool culture. Given the following results, what is the most likely identification?
Lysine decarboxylase = +
Indole = +
VP = Neg
Urease = ±
TCBS agar = Green colonies
Arginine decarboxylase = Neg
KIA = Alk/Acid
Lactose = Neg
String test = Neg
A. Vibrio cholerae
B. Vibrio parahaemolyticus
C. Shigella spp.
D. Salmonella spp.
B
V. parahaemolyticus appear as green colonies on TCBS agar, whereas V. cholerae appear as yellow colonies on TCBS. V. cholerae is the only Vibrio species that causes a positive string test. In the test, a loopful of bacterial colonies is suspended in sodium deoxycholate, 0.5%, on a glass slide. After 60 seconds, the inoculating loop is lifted out of the suspension. V. cholerae forms a long string resembling a string of pearls. Salmonella spp. and Shigella spp. are oxidase negative.
A gram-negative S-shaped rod recovered from selective media for Campylobacter species gave the following results:
Catalase = +
Motility = +
Growth at 42°C = +
Grape odor = Neg
Oxidase = +
Hippurate hydrolysis = +
Nalidixic acid = Susceptible
Pigment = Neg
Cephalothin = Resistant
The most likely identification is:
A. Pseudomonas aeruginosa
B. Campylobacter jejuni
C. Campylobacter fetus
D. Pseudomonas putida
B
The only Campylobacter spp. that hydrolyze hippurate are C. jejuni and subsp. doylei. However, some
strains of P. aeruginosa grow on agar selective for Campylobacter at 42°C. C. fetus will not grow at 42°C but will grow at 25°C and 37°C.
Which atmospheric condition is needed to recover Campylobacter spp. from specimens inoculated onto a Campy-selective agar at 35°C–37°C
and 42°C?
A. 5% O2, 10% CO2, and 85% N2
B. 20% O2, 10% CO2, and 70% N2
C. 20% O2, 20% CO2, and 60% N2
D. 20% O2, 5% CO2, and 75% N2
A
Campylobacter spp. are best recovered in a micro-aerophilic atmosphere (reduced O2). The use of a CO2 incubator or candle jar is not recommended because the amount of O2 and CO2 do not permit any but the most aerotolerant Campylobacter to survive. Cultures for Campylobacter should be incubated for 48–72 hours before reporting no growth.
Which group of tests best differentiates Helicobacter pylori from C. jejuni?
A. Catalase, oxidase, and Gram stain
B. Catalase, oxidase, and nalidixic acid sensitivity
C. Catalase, oxidase, and cephalothin sensitivity
D. Urease, nitrate, and hippurate hydrolysis
D
Helicobacter pylori is found in specimens from gastric secretions and biopsies and has been implicated
as a cause of gastric ulcers. It is found only in the mucus-secreting epithelial cells of the stomach. Both H. pylori and C. jejuni are catalase and oxidase positive. However, Helicobacter spp. are urease positive, which differentiates them from Campylobacter spp.
Test H. pylori C. jejuni
Nitrate
reduction Neg +
Hippurate
hydrolysis Neg +
Urease + Neg
Cephalothin
sensitivity Sensitive Resistant
Nalidixic acid
sensitivity Resistant Sensitive
Which of the following tests should be done first in order to differentiate Aeromonas spp. from the Enterobacteriaceae?
A. Urease
B. OF glucose
C. Oxidase
D. Catalase
C
Aeromonas hydrophilia and other Aeromonas spp. have been implicated in acute diarrheal disease as well as cellulitis and wound infections. Infections usually follow exposure to contaminated soil, water, or food. Aeromonas growing on enteric media are differentiated from the Enterobacteriaceae by demonstrating that colonies are oxidase positive. The Aeromonas are sometimes overlooked as pathogens because most strains grow on selective enteric agar as lactose fermenters.
Which is the best rapid test to differentiate Plesiomonas shigelloides from a Shigella species on selective enteric agar?
A. Oxidase
B. Indole
C. TSI
D. Urease
A
P. shigelloides is a lactose nonfermenter that will resemble Shigella spp. on MacConkey agar. Both are TSI Alk/Acid and urease negative. Plesiomonas produces indole and Shigella usually causes delayed production of indole. However, Plesiomonas is oxidase positive, whereas Shigella spp. are oxidase negative.
Which are the best two tests to differentiate A. hydrophilia from P. shigelloides?
A. Oxidase and motility
B. DNase and VP
C. Indole and lysine decarboxylase
D. Growth on MacConkey and blood agar
B
Both of these bacteria cause diarrhea, grow well on enteric agar, and may be confused with Enterobacteriaceae. Both organisms are positive for oxidase, motility, indole, and lysine decarboxylase. The following reactions are differential:
Test A. hydrophilia P. shigelloides
β-Hemolysis + Neg
on sheep
blood agar
DNase + Neg
VP + Neg
Which genus (in which most species are oxidase and catalase positive) of small gram-negative coccobacilli is associated mainly with animals but may cause endocarditis, bacteremia, as well as wound and dental infections in humans?
A. Actinobacillus
B. Pseudomonas
C. Campylobacter
D. Vibrio
A
Actinobacillus spp. (formerly CDC groups HB-3
and HB-4) share many biochemical characteristics of the Haemophilus spp. Infections most often associated with this gram-negative coccobacillus are subacute bacterial endocarditis and periodontal disease (its main habitat is the mouth). The most common human isolate is Actinobacillus actinomycetemcomitans, which grows slowly on chocolate agar. It is positive for catalase, nitrate reduction, and glucose fermentation. It does not grow on MacConkey agar and is negative for oxidase, urease, indole, X, and V requirements.
Which of the following tests may be used to differentiate Cardiobacterium hominis from Actinobacillus spp.?
A. Gram stain
B. Indole
C. Anaerobic incubation
D. Oxidase
B
C. hominis is a gram-negative coccobacillus biochemically similar to Actinobacillus spp. Like Actinobacillus, it is a cause of endocarditis. However, Cardiobacterium spp. are positive for cytochrome oxidase and negative for nitrate reduction, while most Actinobacillus are negative for oxidase and positive for nitrate reduction. C. hominis will grow on blood agar after 48–72 hours in 5% CO2 at 35°C, but Actinobacillus requires chocolate agar.
A mixture of slender gram-negative rods and coccobacilli with rounded ends was recovered from blood cultures following a patient’s root canal surgery. Given the following results after 48 hours, what is the most likely organism?
Catalase = Neg
Urease = Neg
Oxidase = +
Indole = Neg
Growth on blood and chocolate agar = + (with pitting of agar)
Growth on MacConkey agar = Neg
Ornithine decarboxylase = +
Lysine decarboxylase = +
X and V requirement = Neg Carbohydrates = Neg (no acid produced)
A. Eikenella corrodens
B. Actinobacillus spp.
C. Cardiobacterium hominis
D. Proteus spp.
A
E. corrodens is a part of the normal flora of the upper respiratory tract and the mouth. It is often seen after trauma to the head and neck, dental infections, and human bite wounds. It requires blood for growth. The organism causes a pitting of the agar where colonies are located. The smell of bleach may be apparent when the plates are uncovered for examination. Actinobacillus spp. and C. hominis both utilize several carbohydrates, and Proteus spp. are oxidase negative.
Kingella kingae can best be differentiated from Eikenella corrodens using which medium?
A. Sheep blood agar
B. Chocolate agar
C. MacConkey agar
D. XLD agar
A
Both Kingella kingae and E. corrodens are gram- negative rods that are oxidase positive and catalase negative. Both grow well on blood and chocolate agars and cause pitting of the media, and neither grows on MacConkey or XLD agar. However, K. kingae strains produce a narrow zone of β-hemolysis on sheep blood agar similar to that of group B streptococci.
Kingella kingae is usually associated with which type of infection?
A. Middle ear
B. Endocarditis
C. Meningitis
D. Urogenital
B
Kingella spp. are gram-negative coccobacilli or plump-looking rods. They are part of the normal flora of the upper respiratory and urogenital tracts of humans. Infection is seen primarily in patients having underlying heart disease, poor oral hygiene, or iatrogenic mucosal ulcerations (e.g., radiation therapy), in whom the organism is recovered from blood cultures.
Cultures obtained from a dog bite wound produced yellow, tan, and slightly pink colonies on blood and chocolate agar with a margin of fingerlike projections appearing as a film around the colonies. Given the following results at 24 hours, which is the most likely organism?
Oxidase = +
Growth on MacConkey agar = Neg
Catalase = +
Motility = Neg
A. Actinobacillus spp.
B. Eikenella spp.
C. Capnocytophaga spp.
D. Pseudomonas spp.
C
Capnocytophaga gingivalis, C. sputigena, and C. ochracea are part of the normal oropharyngeal flora of humans; however, C. canimorsus and C. cynodegmi (formerly CDC groups DF-2 and DF-2-like bacteria) are associated with infections resulting from dog bite wounds.
. Smoothgraycoloniesshowingnohemolysisare recovered from an infected cat scratch on blood
and chocolate agar but fail to grow on MacConkey agar. The organisms are gram-negative pleomorphic rods that are both catalase and oxidase positive
and strongly indole positive. The most likely organism is:
A. Capnocytophaga spp.
B. Pasteurella spp.
C. Proteus spp.
D. Pseudomonas spp.
B
Pasteurella multocida (P. canis) is part of the normal mouth flora of cats and dogs and is frequently recovered from wounds inflicted by them. It produces large amounts of indole and therefore an odor resembling colonies of E. coli. Pseudomonas spp. are also catalase and oxidase positive but can be ruled out because they grow on MacConkey agar and do not produce indole.
Which media should be used to recover Bordetella pertussis from a nasopharyngeal specimen?
A. Chocolate agar
B. Blood agar
C. MacConkey agar
D. Bordet–Gengou agar
D
B. pertussis is an oxidase-positive, nonmotile gram-negative coccobacillus and appears as small, round colonies resembling droplets of mercury. It is fastidious and does not grow on chocolate or MacConkey agar. However, B. pertussis adapts to blood agar, growing within 3–6 days. This organism is the cause of whooping cough, which can be prevented by immunization with diphtheria, tetanus, pertussis (DPT) vaccine. The DPT vaccine contains diphtheria and tetanus toxoids and killed whole-cell B. pertussis.
Which medium is recommended for the recovery of Brucella spp. from blood and bone marrow specimens?
A. Biphasic Castenada bottles with Brucella broth B. Blood culture bottles with Brucella broth
C. Bordet–Gengou agar plates and THIO broth
D. Blood culture bottles with THIO broth
A
Although blood agar will support the growth of Brucella spp., Castenada bottles are the medium of choice. Castenada bottles contain a slant of enriched agar medium that is partially submerged and surrounded by an enriched broth medium. As the specimen is injected into the bottles and mixed, the agar slant is simultaneously coated with the blood (or bone marrow). Brucella is the cause of undulant fever and is responsible for many cases of fever of unknown origin. Brucella spp. are facultative intracellular organisms and grow very slowly, usually requiring 4–6 weeks for recovery. Brucella melitensis is the most frquently recovered species.
In addition to CO2 requirements and biochemical characteristics, Brucella melitensis and Brucella abortus are differentiated by growth on media containing which two dyes?
A. Basic fuchsin and thionin
B. Methylene blue and crystal violet
C. Carbol fuchsin and iodine
D. Safranin and methylene blue
A
Which of the following amino acids are required for growth of Francisella tularensis?
A. Leucine and ornithine
B. Arginine and lysine
C. Cysteine and cystine
D. Histidine and tryptophan
C
F. tularensis is a fastidious gram-negative rod that is best recovered from lymph node aspirates and tissue biopsies. It is oxidase negative, nonmotile, and inert biochemically. Cysteine blood agar is the medium of choice, but F. tularensis will grow on commercially prepared chocolate agar because it contains X factor and is supplemented with a growth enrichment (IsoVitaleX) that contains cysteine. F. tularensis may not grow well on MacConkey agar.
Which medium is best for recovery of Legionella pneumophila from clinical specimens?
A. Chocolate agar
B. Bordet–Gengou agar
C. New yeast extract agar
D. Buffered charcoal–yeast extract (CYE) agar
D
L. pneumophila should be recovered on buffered
CYE agar. This agar is nonselective, but can be made more selective for Legionella spp. by addition of
the antibiotics cefamandole, polymixin B, and anisomycin. Any small, glistening, convex colonies on buffered CYE agar after 2–3 days of incubation that do not grow on L-cysteine–deficient buffered CYE agar or routine nonselective media should be further tested by the direct fluorescent antibody test (DFA) for confirmation of L. pneumophila.
Haemophilus influenzae causes ocular infections (pinkeye) and requires X and V factors in the primary medium for growth. The subspecies Haemophilus influenza (biogroup) aegyptius can further be identified and differentiated by which two tests?
A. Indole and xylose
B. Glucose and urease
C. Oxidase and catalase
D. ALA test and oxidase
A
H. influenzae and subspecies H. aegyptius are both glucose, urease, oxidase, and catalase positive.
H. influenzae (biotype II) is positive for both indole and xylose, whereas H. aegyptius is negative for both tests. Biotype II encompasses 40%–70% of H. influenzae strains recovered from clinical specimens. H. influenzae subspecies aegyptius is responsible for epidemics of conjunctivitis in children.
Haemophilus species that require the V factor (NAD) are easily recovered on which primary agar plate?
A. Blood agar made with sheep red cells
B. Blood agar made with horse red cells C. Chocolate agar
D. Xylose agar
C
The V factor, NAD, must first be released from RBCs before it can be assimilated by Haemophilus spp. Chocolate agar is made by heating blood agar in order to lyse RBCs. The released NAD is directly available to those Haemophilus species requiring it. Chocolate agar also contains the X factor (hemin). All Haemophilus except H. ducreyi and H. aphrophilus require V factor, while X factor is required by H. influenzae, H. haemolyticus, and H. ducreyi.
Which of the following products is responsible for satellite growth of Haemophilus spp. around colonies of Staphylococcus and Neisseria spp. on sheep blood agar?
A. NAD
B. Hemin
C. Indole
D. Oxidase
A
Colonies growing on sheep blood agar secreting NAD (V factor) or producing β-hemolysins (which lyse the sheep RBCs releasing NAD) allow pinpoint-size colonies of Haemophilus spp. to grow around them. Sheep blood agar alone does not support the growth of Haemophilus spp., which require V factor because of the presence of V factor–inactivating enzymes that are present in the agar.
Which of the following plates should be used in order to identify Haemophilus haemolyticus and Haemophilus parahaemolyticus?
A. Sheep blood agar and chocolate agar
B. Horse blood agar and Mueller–Hinton agar with X and V strips
C. Brain–heart infusion agar with sheep red cells added
D. Chocolate agar and Mueller–Hinton agar with X factor added
B
Production of β-hemolysis is used to distinguish these two species from other Haemophilus with the same X and V requirements. Horse blood agar furnishes X factor and, when supplemented with yeast extract, supports the growth of Haemophilus spp. Sheep blood agar is not used because it contains growth inhibitors for some Haemophilus spp. The chart on the next page summarizes the characteristics of the Haemophilus spp.
