title: Rapid identification of Escherichia coli O157:H7 in bovine feces using the antibody-direct epifluorescent filter technique (Ab-DEFT).
abstract: The antibody-direct epifluorescent filter technique (Ab-DEFT) was adapted for direct detection of Escherichia coli O157:H7 in bovine feces. The method involved suspension of bovine feces in buffer, centrifugation for 30 s, treatment of the supernatant with trypsin and Triton X-100 at 50 degrees C for 10 min, pre-filtration through 5 and 1.2 microns pore filters, and filtration through a 0.4 micron pore filter. The final filter was stained with fluorescein-labeled polyclonal antibody specific for the O157 antigen and examined by epifluorescence microscopy. The Ab-DEFT was correlated with viable plate counts for enumeration of the pathogen in artificially inoculated bovine feces (r = 0.96). The limit of detection was approximately 10(4)-10(5) CFU/g feces. The procedure provided a clean background for microscopic visualization of cells; however, cell loss and inaccurate quantitation sometimes resulted. E. coli O157:H7 was detected in feces of an inoculated calf for more than 3 weeks post-inoculation. The Ab-DEFT may be useful for rapid screening of cattle for the presence of the E. coli O157:H7 and as an analytical method in ecological studies of the pathogen.
authors.affiliation: National Center for Food Safety and Technology, U.S. Food and Drug Administration, Summit-Argo, IL 60501, USA. mlt@vm.cfsan.fda.gov
authors.forename: M L
authors.initials: ML
authors.lastname: Tortorello
authors.forename: D S
authors.initials: DS
authors.lastname: Stewart
authors.forename: W C
authors.initials: WC
authors.lastname: Cray
mesh_terms: D000818:Animals; D002417:Cattle; D002418:Cattle Diseases; D004927:Escherichia coli Infections / Q000175:diagnosis / Q000662:veterinary*; D019453:Escherichia coli O157 / Q000302:isolation & purification*; D005243:Feces / Q000382:microbiology*; D005455:Fluorescent Antibody Technique