Advances and Challenges in Viability Detection of Foodborne Pathogens.

Dexin Zeng,Feng Xue,Yuan Jiang,Zi Chen,Baoguang Li

doi:10.3389/fmicb.2016.01833

Abstract

Foodborne outbreaks are a serious public health and food safety concern worldwide. There is a great demand for rapid, sensitive, specific, and accurate methods to detect microbial pathogens in foods. Conventional methods based on cultivation of pathogens have been the gold standard protocols; however, they take up to a week to complete. Molecular assays such as polymerase chain reaction (PCR), sequencing, microarray technologies have been widely used in detection of foodborne pathogens. Among molecular assays, PCR technology [conventional and real-time PCR (qPCR)] is most commonly used in the foodborne pathogen detection because of its high sensitivity and specificity. However, a major drawback of PCR is its inability to differentiate the DNA from dead and viable cells, and this is a critical factor for the food industry, regulatory agencies and the consumer. To remedy this shortcoming, researchers have used biological dyes such as ethidium monoazide and propidium monoazide (PMA) to pretreat samples before DNA extraction to intercalate the DNA of dead cells in food samples, and then proceed with regular DNA preparation and qPCR. By combining PMA treatment with qPCR (PMA-qPCR), scientists have applied this technology to detect viable cells of various bacterial pathogens in foods. The incorporation of PMA into PCR-based assays for viability detection of pathogens in foods has increased significantly in the last decade. On the other hand, some downsides with this approach have been noted, particularly to achieve complete suppression of signal of DNA from the dead cells present in some particular food matrix. Nowadays, there is a tendency of more and more researchers adapting this approach for viability detection; and a few commercial kits based on PMA are available in the market. As time goes on, more scientists apply this approach to a broader range of pathogen detections, this viability approach (PMA or other chemicals such as platinum compound) may eventually become a common methodology for the rapid, sensitive, and accurate detection of foodborne pathogens. In this review, we summarize the development in the field including progress and challenges and give our perspective in this area.

Highlights

Foodborne pathogens such as Escherichia coli O157:H7, Salmonella spp., Staphylococcus aureus, Listeria monocytogenes, Campylobacter spp., and Vibrio parahaemolyticus have been a public health concern and there is a growing demand for rapid, sensitive, and accurate methods to detect these pathogens (Scallan et al, 2011)
We mainly focused on the application of this biological dye ethidium monoazide (EMA)/propidium monoazide (PMA) in differentiation of viable cells of foodborne pathogens, E. coli O157:H7, Salmonella, S. aureus, L. monocytogenes, Campylobacter, and V. parahaemolyticus in foodborne pathogens and summarize the developments in this area
We reviewed the development in application of the biological dyes, PMA/EMA, in differentiation of viable cells of foodborne pathogens including E. coli O157:H7, Salmonella, S. aureus, L. monocytogenes, Campylobacter, and V. parahaemolyticus; and we discussed the challenges in using this approach and proposed strategies to remedy the drawbacks of this approach

Summary

INTRODUCTION

Foodborne pathogens such as Escherichia coli O157:H7, Salmonella spp., Staphylococcus aureus, Listeria monocytogenes, Campylobacter spp., and Vibrio parahaemolyticus have been a public health concern and there is a growing demand for rapid, sensitive, and accurate methods to detect these pathogens (Scallan et al, 2011). They found that the PMA-qPCR assay inhibited the amplification of DNA from dead bacterial cells, and the qPCR results reflected the number of viable bacteria without being impacted by the presence of the dead bacteria This approach of combining qPCR with PMA treatment has promise to limit false-positive PCR results when used to diagnose infections, but needs to be further validated in clinical samples (Kobayashi et al, 2009). The authors noted that when salad with high concentration of L. monocytogenes, PMA treatment cannot completely exclude the influence of the dead cells Their data demonstrate that PMA-qPCR is a suitable technique for the detection and quantification of viable pathogens in fresh-cut vegetables at the levels normally found in vegetable samples (Elizaquivel et al, 2012). Besides viable cells of bacterial pathogens (Nocker and Camper, 2006, 2009; Nocker et al, 2007a; Pan and Breidt, 2007; Luo et al, 2010; Li and Chen, 2012, 2013), the PMA-qPCR has been applied to detect viable cells of fungi (Vesper et al, 2008), parasites (Brescia et al, 2009; Cancino-Faure et al, 2016), and viruses (Fittipaldi et al, 2010; Parshionikar et al, 2010; Kim and Ko, 2012; Sanchez et al, 2012; Coudray-Meunier et al, 2013; Fuster et al, 2016)

LIMITATIONS

Findings

CONCLUSION AND PERSPECTIVE