Performance characteristics of qPCR assays targeting human- and ruminant-associated bacteroidetes for microbial source tracking across sixteen countries on six continents.

Numerous quantitative PCR assays for microbial fecal source tracking (MST) have been developed and evaluated in recent years. Widespread application has been hindered by a lack of knowledge regarding the geographical stability and hence applicability of such methods beyond the regional level. This study assessed the performance of five previously reported quantitative PCR assays targeting human-, cattle-, or ruminant-associated Bacteroidetes populations on 280 human and animal fecal samples from 16 countries across six continents. The tested cattle-associated markers were shown to be ruminant-associated. The quantitative distributions of marker concentrations in target and nontarget samples proved to be essential for the assessment of assay performance and were used to establish a new metric for quantitative source-specificity. In general, this study demonstrates that stable target populations required for marker-based MST occur around the globe. Ruminant-associated marker concentrations were strongly correlated with total intestinal Bacteroidetes populations and with each other, indicating that the detected ruminant-associated populations seem to be part of the intestinal core microbiome of ruminants worldwide. Consequently tested ruminant-targeted assays appear to be suitable quantitative MST tools beyond the regional level while the targeted human-associated populations seem to be less prevalent and stable, suggesting potential for improvements in human-targeted methods.

Evaluation of Commercial qPCR Kits for Detection of SARS-CoV-2 in Pooled Samples.

Due to the current pandemic, a global shortage of reagents has drawn interest in developing alternatives to increase the number of coronavirus tests. One such alternative is sample pooling. We compared commercial kits that are used in COVID-19 diagnostics in terms of their sensitivity and feasibility for use in pooling. In this preliminary study, we showed that pooling of up to 80 samples did not affect the efficacy of the kits. Additionally, the RNA-dependent RNA polymerase (RdRp) gene is a more suitable target in pooled samples than the envelope (E) gene. This approach could provide an easy method of screening a large number of samples and help adjust different governmental regulations.