RapidDetection ofEscherichia coli inUrineSamples bya New Chromogenic P-Glucuronidase Assay

was assessed asasubstrate fortherapid detection ofEscherichia coli inurine. Incorporation ofthis compound into MacConkey agarallowed thedirect differentiation ofE.coli asdeepbluecolonies distinct fromlactose andnonlactose fermenters. Thesensitivity was 88to90%,andthe specificity was 100%. Escherichia coli isthemostcommon gram-negative bacterium isolated inclinical laboratories andistheorganism responsible for up to70to95%ofurinary tractinfections (4, 11). Rapid, sensitive, andspecific identification ofE.coli not ohlyisrequired inclinical laboratories butalsoisa priority inmonitoring sanitation andthemicrobiological quality of foodandwater. Recently, a variety oftestshavebeendeveloped to specifically identify E.colibased on theobservation by Kilian andBulow(6)that E.coli isone ofthefewbacteria thatproduce theenzyme P-glucuronidase. Currently, tests tomeasureP-glucuronidase activity include theliberation of yellow p-nitrophenol following hydrolysis ofp-nitrophenylP-D-glucuronide (7) orthefluorogenic recognition ofmethylumbelliferone (5)following hydrolysis of4-methylumbelliferyl-p-D-gluc uronide (MUG).Problems occurwithbothof thesesubstrates, as p-nitrophenol diffuses intotheagar during bacterial growth (7,10), while MUG requires fluorescentlight forvisualization ofcolonies on agarorgrowth in broth. Recently, Leyetal.(10) synthesized indoxyl-p-D-glucuronide(IBDG), whichissplit bytheP-glucuronidase ofE.coli toinsoluble indigo. Their studies on raw sewageindicated a considerable potential fortheIBDGidentification ofE.coli inenvironmental samples. We extended this observation to establish whetherIBDG couldbeusedas a differential mediumfortherapid identification ofE.coli inurine. MacConkey agar(BBLMicrobiology Systems) was supplemented with0.8gofIBDGperliter (MAC-IBDG). Urine specimens frompatients intheHotelDieuHospital, Kingston,Ontario, Canada, were usedtoinoculate MAC-IBDG plates witha 0.01-ml calibrated loop. Deepbluecolonies produced after 18hat35°Cwere scored aspositive forE. coli. Figure 1 showstheease indifferentiating lactose fermenters, nonlactose fermenters, andIBDG-positive E. coli colonies. Thecolonies were distinct, as diffusion of indigo didnotoccur.Theresults ofthedirect screening of urinesamples are showninTable1.No false-positive reactions were noted. (API20E[Analytab Products] and Foxpanels [Beckman Instruments, Inc.] were usedtoidentify theisolates.) Of152gram-negative organisms screened, 99were E.coli. Eighty-three ofthese were positive for IBDGhydrolysis. Ofthe16false-negative strains (10.5%), 6 were slow(>18h)hydrolyzers. Thesensitivity andspeci