A rapid and simple protocol for concentration of SARS-CoV-2 from sewage

Abstract

The aim of this study was to set up a simple protocol to concentrate SARS-CoV-2 from sewage, which can be implemented in laboratories with minimal equipment resources. The method avoids the need for extensive purification steps and reduces the concentration of potential inhibitors of RT-qPCR contained in sewage. The concentration method consists of a single step, in which a small volume of sewage sample is incubated with polyaluminum chloride (PAC). Virus particles adsorbed to the precipitate are collected by low-speed centrifugation, after which the recovered pellet is resuspended with a saline buffer. The PAC concentration method produced an average shift of 4.4-units in Cq values compared to non-concentrated samples, indicating a 25-fold increase in detection sensitivity. The lower detection limit corresponded approximately to 100 copies per ml. Kappa index indicated substantial agreement between PAC and PEG precipitation protocols (k=0.688, CI 0.457-0.919). PAC concentrated samples can be processed immediately for RNA purification and qPCR or sent refrigerated to a diagnosis center, where SARS-CoV-2 detection should be performed in the same way as for clinical samples. This low cost protocol could be useful to aid in the monitoring of community circulation of SARS-CoV-2, especially in low- and middle-income countries, which do not have massive access to support from specialized labs for sewage surveillance.

Monitoring COVID‐19 through SARS‐CoV‐2 quantification in wastewater: progress, challenges and prospects

“…Thus, PEG is being substituted for polyaluminium chloride (PAC) followed by low‐speed centrifugation, which is not so time‐consuming and requires simpler equipment commonly used in WWTP laboratories. PAC concentrated samples are stable for one week at 4°C, and the elimination of PCR inhibitors is similar than in PEG concentrates (Wehrendt et al ., 2021).…”