title: Development of a monoclonal antibody-based colony blot immunoassay for detection of thermotolerant Campylobacter species.
abstract: Campylobacter species, particularly thermotolerant Campylobacter spp., such as C. jejuni, are major human foodborne pathogens. Culture methods have been routinely used for the detection of this organism in various types of samples. An alternative, simple and rapid confirmation test(s) without further tedious biochemical tests would be useful. Meanwhile, Campylobacter-like colonies can be difficult to identify on agar plates overgrown with competitive bacteria, which can lead to false-negative results. This study was to develop a simple colony blot immunoassay using a new monoclonal antibody (Mab) produced in the present study for rapid screening, confirmation and quantification of campylobacters on culture agar plates. The procedure developed in this study was able to specifically detect thermotolerant Campylobacter spp., but not other non-thermotolerant Campylobacter and non-Campylobacter reference strains tested. This assay could detect 105cells in a single dot. This assay showed 100% correlation with the culture method for the blotted membranes from 21 either chicken meat or vegetable samples experimentally inoculated with thermotolerant campylobacters. Among 101 natural samples of chicken meat (n=44), chicken feces (n=20) and vegetables (n=37), this assay also showed positive for 23 chicken meat and 14 fecal samples that were positive for thermotolerant campylobacters by culture method, and identified four additional suspects that were culture negative. Membranes stored at 4°C for at least 4years could also be used for this assay. The assay developed in this study can be used in quantitative study for immediate or archival usage, and for diagnostic test to preliminarily confirm the presence of thermotolerant Campylobacter on agar plates.
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada. Electronic address: hongsheng.huang@inspection.gc.ca.
authors.forename: Hongsheng
authors.initials: H
authors.lastname: Huang
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada.
authors.forename: Beverley
authors.initials: B
authors.lastname: Phipps-Todd
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada.
authors.forename: Tanis
authors.initials: T
authors.lastname: McMahon
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada.
authors.forename: Catherine L
authors.initials: CL
authors.lastname: Elmgren
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada.
authors.forename: Cheryl
authors.initials: C
authors.lastname: Lutze-Wallace
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada.
authors.forename: Zoe A
authors.initials: ZA
authors.lastname: Todd
authors.affiliation: Ottawa Laboratory - Fallowfield (OLF), Canadian Food Inspection Agency (CFIA), Ottawa K2H 8P9, ON, Canada.
authors.forename: Manuel M
authors.initials: MM
authors.lastname: Garcia
mesh_terms: D000362:Agar; D000818:Animals; D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D015373:Bacterial Typing Techniques / Q000379:methods*; D015153:Blotting, Western; D002167:Campylobacter / Q000166:cytology / Q000254:growth & development / Q000276:immunology / Q000302:isolation & purification*; D002645:Chickens / Q000382:microbiology; D015169:Colony Count, Microbial / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay; D005243:Feces / Q000382:microbiology; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D007159:Immunologic Tests / Q000379:methods; D018189:Immunomagnetic Separation / Q000379:methods; D008460:Meat / Q000382:microbiology; D008855:Microscopy, Electron, Scanning; D012680:Sensitivity and Specificity; D000071436:Thermotolerance; D014675:Vegetables / Q000382:microbiology
chemical_list: D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000362:Agar
keywords: Colony blot immunoassay; Electron microscopy; Immunomagnetic beads separation; Monoclonal antibody; Thermotolerant and non-thermotolerant Campylobacter spp.