Direct immunofluorescence assay for rapid environmental detection of Vibrio cholerae O1.
An immunofluorescence assay for direct detection of V. cholerae O1 was developed using polyclonal antibodies raised against outer membrane proteins (OMPs) of V. cholerae O1. Production of OMPs varied with growth media used; maximum production was found in tryptic soy broth. The detection system was specific because no cross-reactivity was observed with other bacteria including V. cholerae O139, E. coli, S. dysenteriae and Salmonella enterica subsp. enterica serovar Typhi. The technique was able to detect 240 CFU/mL of V. cholerae O1 suspended in phosphate-buffered saline. The assay coupled with bacterial enrichment in APW for 6 h detected as few as 5 CFU of V. cholerae in spiked samples. Moreover, a 2-h incubation of enriched bacterial cells in 0.1% yeast extract with 10 ppm nalidixic acid enhanced the bacterial size and helped in morphological identification of V. cholerae. Among 32 potable water samples from afflicted hand pumps and wells collected from a cholera-plagued area 12 were found to be contaminated with V. cholerae by immunofluorescence assay as well as by conventional culture methods. The proposed method could thus be employed in environmental surveillance of V. cholerae O1.
Goel
A K
AK
Biotechnology Division, Defense Research and Development Establishment, Gwalior 474 002, India.
Tamrakar
A K
AK
Kamboj
D V
DV
Singh
L
L
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial / Q000276:immunology / Q000302:isolation & purification; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology / Q000302:isolation & purification; D003429:Cross Reactions; D003470:Culture Media / Q000737:chemistry; D004926:Escherichia coli / Q000276:immunology; D019085:Fluorescent Antibody Technique, Direct / Q000379:methods*; D005618:Fresh Water / Q000382:microbiology; D011159:Population Surveillance; D012485:Salmonella typhi / Q000276:immunology; D012680:Sensitivity and Specificity; D012762:Shigella dysenteriae / Q000276:immunology; D041241:Vibrio cholerae O1 / Q000276:immunology / Q000302:isolation & purification*; D014871:Water Microbiology
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D003470:Culture Media