Abstract
Quantitative detection of health-related microorganisms is of importance for risk assessment of environmental context, but quantitative real-time PCR (qPCR) alone meets a difficulty to distinguish viable and dead cells. Propidium monoazide (PMA) treatment followed by qPCR has been introduced to overcome this issue. In this study, a LED lamp, instead of a halogen lamp, was used for cross-linking of PMA and DNA, and the effects of filtration for the enumeration of viable Escherichia coli by PMA-qPCR was investigated. PMA-qPCR was applicable to suspended cells after the treatment with 100μM PMA followed by 15 min exposure to a LED lamp. On the other hand, PMA-qPCR was not able to distinguish viable and dead cells after filtration. Cell treatment with LIVE/ DEAD BacLight Bacterial Viability Kits indicated that the most of cell after a filtration procedure were damaged, suggesting the filtration procedure is one crucial point for PMA-qPCR of filtered samples.
