DNA-based real-time detection and quantification of aeromonads from fresh water beaches on Lake Ontario.
The present study was designed to develop a novel, rapid, direct DNA-based protocol to enumerate aeromonads in recreational waters. An Aeromonas genus-specific real-time quantitative polymerase chain reaction (Q-PCR) protocol was developed and optimized using newly designed genus-specific oligonucleotide primers derived from the gyrase B subunit (GyrB) gene. A standard curve was developed based on the PCR protocol with a minimum quantification limit of 10 cell equivalents ml(-1) achieved using an autoclaved water sample from recreational water spiked with known quantities of an Aeromonas ATCC strain. The Q-PCR protocol was validated and applied to detect and quantify the total number of aeromonads in water samples collected from two fresh water beaches on Lake Ontario. The Q-PCR protocol revealed significantly higher numbers of aeromonads in all water samples than a culture-based assay at both beaches. Foreshore sand was found to serve as a reservoir of high concentrations of Aeromonas similar to this phenomenon noted for enteric bacteria like Eschershia coli. The new real-time Q-PCR protocol facilitated the rapid quantification of total numbers of Aeromonas cells present in recreational water samples in <3 hours without culturing.
Khan
Izhar U H
IU
National Water Research Institute, Water Science and Technology Directorate, Environment Canada, Burlington, ON L7R 4A6, Canada.
Loughborough
Alyssa
A
Edge
Thomas A
TA
D000333:Aeromonas / Q000235:genetics* / Q000302:isolation & purification*; D001493:Bathing Beaches; D027081:DNA Gyrase / Q000032:analysis / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000032:analysis*; D004784:Environmental Monitoring / Q000379:methods; D005618:Fresh Water / Q000032:analysis* / Q000382:microbiology*; D016905:Gram-Negative Bacterial Infections / Q000517:prevention & control; D006801:Humans; D009864:Ontario; D016133:Polymerase Chain Reaction / Q000379:methods*; D014871:Water Microbiology
D004269:DNA, Bacterial; D027081:DNA Gyrase