SARS-CoV-2 RNA is enriched by orders of magnitude in solid relative to liquid wastewater at publicly owned treatment works

Abstract

Wastewater-based epidemiology has gained attention throughout the world for detection of SARS-CoV-2 RNA in wastewater to supplement clinical testing. Methods have been developed using both the liquid and the solid fraction of wastewater, with some studies reporting higher concentrations in solids. To investigate this relationship further, we collaborated with six other laboratories to conduct a study across five publicly owned treatment works (POTWs) where both primary solids and raw wastewater influent samples were collected and quantified for SARS-CoV-2 RNA. Solids and influent samples were processed by participating laboratories using their respective methods and retrospectively paired based on date of collection. SARS-CoV-2 RNA concentrations by mass (gene copies per gram) were higher in solids than in influent by approximately three orders of magnitude. Concentrations in matched solids and influent were positively and significantly correlated at all five POTWs. RNA concentrations in both solids and influent were correlated to COVID-19 incidence rates in the sewershed and thus representative of disease burden; the solids methods appeared to produce a comparable relationship between SARS-CoV-2 RNA concentration measurements and incidence rates across all POTWs. Solids and influent methods showed comparable sensitivity, N gene detection frequency, and calculated empirical incidence rate lower limits. Analysis of solids has the advantage of using less sample volume to achieve similar sensitivity to influent methods.

Persistence of Endogenous SARS-CoV-2 and Pepper Mild Mottle Virus RNA in Wastewater-Settled Solids

“…Previous studies showed that RNA from SARS-CoV-2 is naturally concentrated in primary settled solids at wastewater treatment plants. , Further, researchers have found a significant correlation between SARS-CoV-2 RNA isolated from primary settled solids samples and laboratory-confirmed COVID-19 cases; − thus, settled solids might be an advantageous medium for wastewater-based epidemiology efforts. Laboratory-confirmed COVID-19 incidence may be considerably smaller than actual COVID-19 incidence owing to test-seeking behaviors which are influenced by testing availability and symptom presence and severity. , Some studies aim to infer actual COVID-19 incident cases from wastewater concentrations of SARS-CoV-2 RNA using models. − Such models require the input of various parameters including fecal loads, viral shedding rates, flow to the wastewater treatment plant, the decay rate constant and partition coefficient of SARS-CoV-2 RNA, and the average residence time of wastewater prior to sample collection.…”

“…Previous studies showed that RNA from SARS-CoV-2 is naturally concentrated in primary settled solids at wastewater treatment plants. 1 , 2 Further, researchers have found a significant correlation between SARS-CoV-2 RNA isolated from primary settled solids samples and laboratory-confirmed COVID-19 cases; 3 − 7 thus, settled solids might be an advantageous medium for wastewater-based epidemiology efforts. Laboratory-confirmed COVID-19 incidence may be considerably smaller than actual COVID-19 incidence owing to test-seeking behaviors which are influenced by testing availability and symptom presence and severity.…”

Detection of SARS-CoV-2 Variants Mu, Beta, Gamma, Lambda, Delta, Alpha, and Omicron in Wastewater Settled Solids Using Mutation-Specific Assays Is Associated with Regional Detection of Variants in Clinical Samples

“…In particular, we developed and utilized assays for mutations characteristic of Alpha, Beta and Gamma (Beta/Gamma), Delta, Mu, Lambda, and Omicron and then measured these in wastewater solids from a publicly owned treatment work (POTW) located in the Bay Area of California, USA. We measured concentrations in wastewater settled solids, as concentrations of SARS-CoV-2 RNA are enriched several orders of magnitude in solids relative to those in liquid wastewater ( 20 , 21 ). We subsequently compared the measurements to data on occurrence of those variants in clinical specimens, aggregated at the state level.…”

Persistence of endogenous SARS-CoV-2 and pepper mild mottle virus RNA in wastewater settled solids

“…Previous studies showed that RNA from SARS-CoV-2 is naturally concentrated in primary settled solids at wastewater treatment plants. 1,2 Further, researchers have found a significant correlation between SARS-CoV-2 RNA isolated from primary settled solids samples and laboratoryconfirmed COVID-19 cases [3][4][5][6][7] thus, settled solids might be an advantageous medium for wastewater-based epidemiology efforts. Laboratory-confirmed COVID-19 incidence may be considerably smaller than actual COVID-19 incidence owing to test-seeking behaviors which are influenced by testing availability and symptom presence and severity.…”