Question

Briefly describe the procedure for isolating and testing a suspected E. coli 0157:H7 .

Answer 1

Escherichia coli O157:H7 is a primary cause of severe and bloody diarrhea, especially bloody diarrhea or hemolytic uremic syndrome (HUS). Stool specimens (whole stools, swabs prepared from whole stools or rectal swabs with visible fecal staining) should be collected. Ideally, specimens should be collected as close to the time of onset of diarrhea as possible; however, specimens taken even weeks after the onset of symptoms are sometimes positive

Inoculate stool specimens onto Sorbitol-MacConkey (SMAC) agar and incubate 18-24 hours at 35-37⁰C. Sorbitol-negative colonies will appear colorless on Sorbitol-MacConkey (SMAC) agar.

Test sorbitol- negative colonies selected from Sorbitol-MacConkey with E. coli O157 antiserum or latex reagents (O157 antibody-coated latex and control latex) according to the procedures recommended by the manufacturer. Colonies may be tested with antisera directly from the plate, or subcultured to another nonselective medium (blood agar, for example) and tested the next day. If colonies are tested directly from the plate, O157- positive colonies should also be transferred to another medium for subsequent testing. Although it is more labor-intensive and delays results by a day, subculturmg to another medium and testing the next day offers the advantage of providing more bacterial growth on which to perform the O157 agglutination assay. The extra growth makes it easier to observe agglutination and allows repeat testing of the isolate. if necessary. Once one colony from a plate as been identified as O157-positive, no further colonies from the same plate need to be tested.

Isolates agglutinating in O157 antiserum or O157 latex reagent should be identified biochemically as E. coli, since strains of several species cross-react with O157 antiserum. However, because biochemical confirmation may take 24 hours or longer, an oral report of presumptive E. coli O 157 may be given before biochemical identification is completed.

Specimens from which sorbitol-negative colonies have been isolated that agglutinate in O157 antiserum or O157 latex reagent, and are biochemically E. coli, may be reported as presumptively positive for E. coli O157:H7.