Monitoring SARS-CoV-2 variant transitions using differences in diagnostic cycle threshold values of target genes.

Monitoring the emergence of new SARS-CoV-2 variants is important to detect potential risks of increased transmission or disease severity. We investigated the identification of SARS-CoV-2 variants from real-time reverse transcriptase polymerase chain reaction (RT-PCR) routine diagnostics data. Cycle threshold (Ct) values of positive samples were collected from April 2021 to January 2022 in the Northern Metropolitan Area of Barcelona (n = 15,254). Viral lineage identification from whole genome sequencing (WGS) was available for 4618 (30.3%) of these samples. Pairwise differences in the Ct values between gene targets (ΔCt) were analyzed for variants of concern or interest circulating in our area. A specific delay in the Ct of the N-gene compared to the RdRp-gene (ΔCtNR) was observed for Alpha, Delta, Eta and Omicron. Temporal differences in ΔCtNR correlated with the dynamics of viral replacement of Alpha by Delta and of Delta by Omicron according to WGS results. Using ΔCtNR, prediction of new variants of concern at early stages of circulation was achieved with high sensitivity and specificity (91.1% and 97.8% for Delta; 98.5% and 90.8% for Omicron). Thus, tracking population-wide trends in ΔCt values obtained from routine diagnostics testing in combination with WGS could be useful for real-time management and response to local epidemics.

Commercial real time PCR implementation for rapid diagnosis of onychomycosis: A new workflow in a clinical laboratory.

INTRODUCTION: Onychomycosis is a frequent and underdiagnosed condition. Approximately 90% of toenail onychomycosis infections are caused by dermatophytes, but classical diagnosis based on culture and microscopy observation is slow and has low sensitivity. Both limitations can be solved incorporating molecular techniques to routine diagnosis of onychomycosis. OBJECTIVE: Prospective evaluation of the utility of incorporating in the clinical laboratory workflow a commercial real time PCR (qPCR) for dermatophytes detection in nails after potassium hydroxide direct observation screening. MATERIALS AND METHODS: 152 nail samples were included (34 KOH negative and 118 KOH positive) and processed by culture and qPCR. RESULTS: In the negative KOH group, only one dermatophyte grew in culture and three were detected by qPCR. In the group of positive KOH, 57 dermatophytes grew in culture and 81 were detected by qPCR. In this group, 25% of diagnosed dermatophytes were detected only by qPCR. The sensitivity of qPCR compared to culture is 92.8% and time of response decreases from days to hours. CONCLUSION: Based in our results, we propose a workflow algorithm for a clinical laboratory that eliminates culture for qPCR positive samples.

Pre-Treatment with Grape Seed Extract Reduces Inflammatory Response and Oxidative Stress Induced by Helicobacter pylori Infection in Human Gastric Epithelial Cells.

Helicobacter pylori (H. pylori) is a pathogenic bacteria identified as a potential risk factor for gastritis, gastric ulcers and gastric cancer. During the stomach colonization, H. pylori triggers a strong inflammatory response and subsequent oxidative stress, which are associated with tissue damage. For this reason, it is of particular interest to develop alternative natural tools that enable modulation of the associated damaging immune response. With this purpose, we obtained grape seed extract (GSE) from sweet (not fermented) food grade seeds. The aim of our study was to investigate the effect of GSE and its two enriched procyanidins fractions (OPC and PPC) on the inflammatory process and oxidative stress produced by different H. pylori strains in human gastric epithelial cells (AGS). Anti-inflammatory activity was evaluated by measuring the level of interleukin-8 (IL-8) secretion. IL-8 production was significantly reduced in H. pylori-infected human gastric epithelial cells pre-treated with GSE or its enriched fractions when compared with non-pre-treated infected cells (from 21.6% to 87.8%). Pre-treatment with GSE or its fractions significantly decreased intracellular reactive oxygen species (ROS) production in AGS cells after infection, depending on the H. pylori strain. Our results also showed that GSE and its fractions demonstrate antibacterial activity against all strains of H. pylori used in the study. This work demonstrates the effectiveness of GSE enriched in procyanidins against the main events associated with H. pylori infection.