Progress in understanding preferential detection of live cells using viability dyes in combination with DNA amplification

Optimization of Propidium Monoazide Quantitative PCR for Evaluating Performances of Bioaerosol Samplers for Sampling AirborneStaphylococcus aureus

“…These two studies applied 50 mM PMA (corresponding to 25 mg/ml) to distinguish between intact and dead S. aureus in suspension, but we found that this concentration of PMA may penetrate intact cell membranes. In contrast to most previous studies, which used PMA concentrations ranging from 25 to 50 mg/ml (Fittipaldi et al 2012), a recent study recommend a lower PMA concentration of 10 mM (corresponding to 5 mg/ml) combined with a longer incubation time (30 min) to quantify another gram-positive species, Listeria monocytogenes (Nkuipou-Kenfack et al 2013). Although making a direct comparison is difficult due to the variations in the methodology, these results indicated that the optimized PMA concentration may be affected by many parameters such as microbial species, cell concentration, the ratio between live and dead cells, and the dye incubation time (Fittipaldi et al 2012).…”

“…Exposure to light causes PMA to covalently bind to DNA, while unbound PMA is inactivated (Fittipaldi et al 2012). Insufficient light exposure may affect the efficiency of PMA 5 CFU/ml of (a) viable MSSA cells and (b) dead MSSA cells.…”

“…However, a complex set of parameters that may affect the efficiency of PMA-qPCR must be taken into account before applying PMA-qPCR to detect different bioaerosols. The optimal conditions for PMA-qPCR may vary with microbe species (Fittipaldi et al 2012). Therefore, there is a need to separately investigate the efficiency of the same technology for different microbe species.…”

Viable quantitative PCR for assessing the response of Candida albicans to antifungal treatment

“…Selective nucleic acid intercalating dyes, such as EMA and PMA, represent one of the most successful recent approaches to detect viable cells (as defined by an intact cell membrane) by PCR (Fittipaldi et al 2012) and have been effectively evaluated in different microorganisms Rudi et al 2005;van Frankenhuyzen et al 2011). As both dyes have similar structures, comparable results might be expected; however, some studies have showed differences (Andorrà et al 2010;Cawthorn et al 2008;Chang et al 2010;Nam et al 2011;Nocker and Camper 2009;Pan and Breidt 2007;Wagner et al 2008).…”

“…In this bound state, DNA cannot be amplified by PCR (Nocker and Camper 2009;Rudi et al 2005). This promising analytical approach is still in development and needs to be investigated further (Fittipaldi et al 2012). …”