The use of bovine serum albumin to improve the RT-qPCR detection of foodborne viruses rinsed from vegetable surfaces


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Abstract

Aims:To demonstrate that produce rinsates used for RT-qPCR detection of foodborne viruses may cause significant PCR inhibition and propose a means to reduce its impact on sensitivity.Methods and Results:Here, it is shown that rinsing and concentration from spinach and precut lettuce have the potential to generate RNA extracts that are inhibitory to RT-qPCRs assembled from commercial kits for the detection of norovirus GII (NoV GII), hepatitis A virus (HAV), hepatitis E virus (HEV), rotavirus (RV) and feline calicivirus (FCV) as sample process control. It is further shown that the addition of bovine serum albumin (BSA) to those reactions restored a positive signal in all cases. The effect of BSA was dependent upon the primer/probe combination. Moreover, two of the detection systems (FCV and HAV) strongly benefited from the addition of BSA even in the absence of PCR inhibitors.Conclusions:BSA was shown to restore positive signals in five different RT-qPCR systems that were otherwise completely inhibited by produce rinsate extracts. It is therefore suggested to consider the addition of BSA to RT-qPCRs for the detection of foodborne viruses when inhibition is observed.Significance and Impact of the Study:This study clearly demonstrates the potency of PCR inhibitors generated during routine virus concentration from produce and that it can be alleviated by the addition of BSA to the RT-qPCRs. Although used elsewhere, the addition of BSA to PCRs is not a common practice in this growing field of research.

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