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

Wehrendt, Diana P.; Massó, Mariana G.; Machuca, Adrián Gonzales; Vargas, Claudia V.; Barrios, Melina E.; Campos, Josefina; Costamagna, Damián; Bruzzone, Luis; Cisterna, Daniel M.; Iglesias, Néstor Gabriel; Mbayed, Viviana A.; Baumeister, Elsa; Centrón, Daniela; Quiroga, María Paula; Erijman, Leonardo

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 (40 mL) of sewage sample is incubated with polyaluminum chloride (PAC)(0.00045 N Al(3+) final concentration). Virus particles adsorbed to the precipitate are collected by low-speed centrifugation, after which the recovered pellet is resuspended with a saline buffer. PAC concentrated samples are stable for at least one week at 4 °C. Therefore, they may be sent refrigerated to a diagnosis center for RNA extraction and RT-qPCR for SARS-CoV-2 RNA detection if the lab does not have such capabilities. The PAC concentration method produced an average shift of 4.5-units in quantification cycle (Cq) values compared to non-concentrated samples, indicating a 25-fold increase in detection sensitivity. The lower detection limit corresponded approximately to 100 viral copies per ml. Kappa index indicated substantial agreement between PAC and polyethylene glycol (PEG) precipitation protocols (k = 0.688, CI 0.457-0.919). This low-cost concentration 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.