Development of a simple and rapid fluorogenic procedure for identification of vibrionaceae family members.
We describe a simple colony overlay procedure for peptidases (COPP) for the rapid fluorogenic detection and quantification of Vibrionaceae from seawater, shellfish, sewage, and clinical samples. The assay detects phosphoglucose isomerase with a lysyl aminopeptidase activity that is produced by Vibrionaceae family members. Overnight cultures are overlaid for 10 min with membranes containing a synthetic substrate, and the membranes are examined for fluorescent foci under UV illumination. Fluorescent foci were produced by all the Vibrionaceae tested, including Vibrio spp., Aeromonas spp., and Plesiomonas spp. Fluorescence was not produced by non-Vibrionaceae pathogens. Vibrio cholerae strains O1, O139, O22, and O155 were strongly positive. Seawater and oysters were assayed, and 87 of 93 (93.5%) of the positive isolates were identified biochemically as Vibrionaceae, principally Vibrio vulnificus, Vibrio parahaemolyticus, Aeromonas hydrophila, Photobacterium damselae, and Shewanella putrefaciens. None of 50 nonfluorescent isolates were Vibrionaceae. No Vibrionaceae were detected in soil, and only A. hydrophila was detected in sewage. The COPP technique may be particularly valuable in environmental and food-testing laboratories and for monitoring water quality in the aquaculture industry.
USDA, ARS, Delaware State University, 1200 N. DuPont Hwy., James W. W. Baker Center, Dover, DE 19901, USA. grichard@desu.edu
Gary P
GP
Richards
Michael A
MA
Watson
Salina
S
Parveen
D000818:Animals; D015373:Bacterial Typing Techniques; D003374:Coumarins / Q000378:metabolism; D003470:Culture Media; D004151:Dipeptides / Q000378:metabolism; D005453:Fluorescence; D016905:Gram-Negative Bacterial Infections / Q000382:microbiology; D006801:Humans; D010125:Ostreidae / Q000382:microbiology; D010447:Peptide Hydrolases / Q000378:metabolism*; D012623:Seawater / Q000382:microbiology; D012722:Sewage / Q000382:microbiology; D012758:Shellfish / Q000382:microbiology; D013997:Time Factors; D014737:Vibrionaceae / Q000145:classification* / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism
D003374:Coumarins; D003470:Culture Media; D004151:Dipeptides; D012722:Sewage; D010447:Peptide Hydrolases