Stool cultures may be performed on rectal swabs containing feces or submitted stool samples. Swabs are placed in a tube containing Stuart or other transport medium and then delivered to the laboratory. Cultures for C. difficle are usually collected by swabbing the rectum (whereas watery stool is needed for immunoassay of C. difficle toxin). The swab must be placed immediately into prereduced (oxygen free) transport medium because this organism is a strict anaerobe. To submit a stool specimen for routine culture, the patient or caregiver collects a stool sample in a special container, taking care not to contaminate the specimen with water, urine, or other materials. Some containers include a transport solution to stabilize the specimen. Although some requests are for stool cultures on two or more consecutive days, a single specimen is considered to be sufficient. It is important to return the specimen to the doctor's office or the laboratory in the time specified by the physician or nurse. Laboratories normally do not accept stool specimens that are contaminated or that arrive after the specified time period.
A routine bacterial stool culture involves placing a sample of the stool on several kinds of enriched and selective media containing nutrients that support the growth of certain types of organisms. Routine culture should include a sheep blood agar plate, MacConkey agar plate, MacConkey agar with sorbitol, Hektoen or XLD (xylose lysine desoxycholate) plate, Campy plate, and GN (gram-negative) broth. Blood agar supports the growth of most bacteria including Staphylococcus aureus, Listeria monocytogenes, and yeast, which are infrequently implicated in food poisoning or gastrointestinal infections, but do not grow on the other media. Most intestinal pathogens are gram-negative bacilli. MacConkey agar is selective for these organisms and differentiates those that can ferment lactose from those that cannot. MacConkey sorbitol substitutes sorbitol for lactose. This allows differentiation of nonpathogenic E. coli that ferment sorbitol well from the O57:H7 strain, which does not. Hektoen or XLD enhance the growth of Salmonella and Shigella by suppressing the growth of gram-positive organisms and gram-negative normal flora. They also differentiate lactose and sucrose fermenters such as E. coli from Salmonella and Shigella, which are not. Several drops of the GN broth can be transferred to hecktoen or XLD agar after a four-hour incubation at 36Â°C. This procedure can yield isolated colonies of a pathogen the next day that can be used to perform biochemical identification, serotyping, and antibiotic susceptibility tests. Campy agar contains 10% sheep blood, sodium bisulfite, and three antibiotics. The sodium bisulfite reduces some of the oxygen in the medium which enhances recovery of Campylobacter. The antibiotics prevent other gram-negative bacilli and yeast from growing. All inoculated media except the Campy plate are incubated in air or 5-10% carbon dioxide at 36Â°C and are examined for growth at 24 hours and again the next day. Campy plates must be incubated at 42Â°C. Plates are examined at 24 hours and each day for the next two days. Cultures for Clostridium difficile require CCFA agar and thioglycolate broth. These are incubated in an oxygen free environment at 36Â°C for two days. CCFA is cycloserine-cefoxitin fructose agar and it inhibits the growth of other enteric anaerobes found as normal flora in stool. C. difficle produces large yellow colonies on CCFA agar that will fluoresce yellow-green.
Gram stains are not performed routinely, but may be requested for the semiquantitation of white blood cells. If any suspicious bacterial colonies grow, they are presumptively identified on the basis of colonial growth, physical characteristics, microscopic features, and biochemical tests. The colonies are subcu