DEVELOPMENT OF A RAPID AND SPECIFIC COLONY-LIFT IMMUNOASSAY FOR DETECTION AND ENUMERATION OF CAMPYLOBACTER-JEJUNI, CAMPYLOBACTER-COLI, AND C-LARI

Contamination of retail poultry by Campylobacter spp, is a significant source of human diarrheal disease, We have developed a colony-lift im munoassay (CLI) for the detection of Campylobacter jejuni, C. call, an d C. lari isolated from such sources and grown on selective agar mediu m or on filter membranes. This technique has been successfully utilize d to quantify Campylobacter colonies within 18 to 28 h after sampling, Hydrophobic, high-protein-binding membranes were prewet with methanol and used to imprint bacterial cells from the agar or filter membrane, while leaving colonies intact and viable, The membranes were air drie d, peroxidase neutralized, blocked with bovine serum albumin in phosph ate-buffered saline, and hybridized for 5 min with an affinity-purifie d, horseradish peroxidase-labeled goat anti-Campylobacter antibody pre paration (Kirkegaard and Ferry Laboratories), The membranes were washe d briefly, exposed to a 3,3',5,5'-tetramethylbenzidine membrane substr ate, rinsed in deionized water, and allowed to dry, Lifted colonies of Campylobacter were identified by a blue color reaction on the membran e, Replicas of the membranes were made by marking the location of the Campylobacter colonies on clear transparencies, which were subsequentl y utilized to locate the original colony on the filter membrane or aga r plate, The specificity of this antibody preparation has been evaluat ed against a wide range of Campylobacter spp., including American Type Culture Collection type and reference strains, retail poultry isolate s, and isolates obtained from cloacal swabs of live commercial broiler chickens, Specificity against numerous non-Campylobacter spp. obtaine d from the same sources was also evaluated. The CLI provided a rapid a nd simple means for detection and enumeration of enteropathogenic Camp ylobacter organisms, We have successfully combined this CLI procedure with methods recently developed in our laboratories for retail meat an d poultry sampling, Potentially, broader applications for use of this technique include detection and enumeration of campylobacters from cli nical, veterinary, and environmental samples.