PMCCPMCCPMCC

Search tips
Search criteria 

Advanced

 
Logo of aemPermissionsJournals.ASM.orgJournalAEM ArticleJournal InfoAuthorsReviewers
 
Appl Environ Microbiol. Oct 2012; 78(20): 7369–7375.
PMCID: PMC3457089
Rapid and Sensitive Quantification of Vibrio cholerae and Vibrio mimicus Cells in Water Samples by Use of Catalyzed Reporter Deposition Fluorescence In Situ Hybridization Combined with Solid-Phase Cytometry
Sonja Schauer,a Regina Sommer,ac Andreas H. Farnleitner,bc and Alexander K. T. Kirschnercorresponding authorac
aMedical University Vienna, Institute for Hygiene and Applied Immunology, Water Hygiene, Vienna, Austria
bInstitute of Chemical Engineering, Research Group Environmental Microbiology and Molecular Ecology, Vienna University of Technology, Vienna, Austria
cInteruniversity Cooperation Center Water and Health (ICC), Vienna, Austria
corresponding authorCorresponding author.
Address correspondence to Alexander K. T. Kirschner, alexander.kirschner/at/meduniwien.ac.at.
For this virtual institution, see http://www.waterandhealth.at.
Received July 11, 2012; Accepted August 1, 2012.
Abstract
A new protocol for rapid, specific, and sensitive cell-based quantification of Vibrio cholerae/Vibrio mimicus in water samples was developed. The protocol is based on catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) in combination with solid-phase cytometry. For pure cultures, we were able to quantify down to 6 V. cholerae cells on one membrane with a relative precision of 39% and down to 12 cells with a relative precision of 17% after hybridization with the horseradish peroxidase (HRP)-labeled probe Vchomim1276 (specific for V. cholerae and V. mimicus) and signal amplification. The corresponding position of the probe on the 16S rRNA is highly accessible even when labeled with HRP. For the first time, we were also able to successfully quantify V. cholerae/V. mimicus via solid-phase cytometry in extremely turbid environmental water samples collected in Austria. Cell numbers ranged from 4.5 × 101 cells ml−1 in the large saline lake Neusiedler See to 5.6 × 104 cells ml−1 in an extremely turbid shallow soda lake situated nearby. We therefore suggest CARD-FISH in combination with solid-phase cytometry as a powerful tool to quantify V. cholerae/V. mimicus in ecological studies as well as for risk assessment and monitoring programs.
Articles from Applied and Environmental Microbiology are provided here courtesy of
American Society for Microbiology (ASM)