Co-quantification of crAssphage increases confidence in wastewater-based epidemiology for SARS-CoV-2 in low prevalence areas

Maxwell L Wilder; Frank Middleton; David A Larsen; Qian Du; Ariana Fenty; Teng Zeng; Tabassum Insaf; Pruthvi Kilaru; Mary Collins; Brittany Kmush; Hyatt C Green

doi:10.1016/j.wroa.2021.100100

Co-quantification of crAssphage increases confidence in wastewater-based epidemiology for SARS-CoV-2 in low prevalence areas

Water Res X. 2021 May 1:11:100100. doi: 10.1016/j.wroa.2021.100100. Epub 2021 Apr 6.

Authors

Maxwell L Wilder¹, Frank Middleton², David A Larsen³, Qian Du⁴, Ariana Fenty¹, Teng Zeng⁵, Tabassum Insaf^{6

7}, Pruthvi Kilaru³, Mary Collins⁸, Brittany Kmush³, Hyatt C Green¹

Affiliations

¹ Department of Environmental and Forest Biology, SUNY-ESF, Syracuse, NY 13210.
² Department of Neuroscience and Physiology, Upstate Medical University, Syracuse, NY 13210.
³ Department of Public Health, Syracuse University, Syracuse, NY 13244.
⁴ Quadrant Biosciences, Syracuse, NY 13210.
⁵ Department of Civil & Environmental Engineering, Syracuse University, Syracuse, NY 13244.
⁶ Bureau of Environmental and Occupational Epidemiology, New York State Department of Health, Albany, NY 12337.
⁷ Department of Epidemiology and Biostatistics, University at Albany, Rensselaer, NY 12144.
⁸ Department of Environmental Studies, SUNY-ESF, Syracuse, NY 13210.

Abstract

Wastewater surveillance of SARS-CoV-2 RNA is increasingly being incorporated into public health efforts to respond to the COVID-19 pandemic. In order to obtain the maximum benefit from these efforts, approaches to wastewater monitoring need to be rapid, sensitive, and relatable to relevant epidemiological parameters. In this study, we present an ultracentrifugation-based method for the concentration of SARS-CoV-2 wastewater RNA and use crAssphage, a bacteriophage specific to the human gut, to help account for RNA loss during transit in the wastewater system and sample processing. With these methods, we were able to detect, and sometimes quantify, SARS-CoV-2 RNA from 20 mL wastewater samples within as little as 4.5 hours. Using known concentrations of bovine coronavirus RNA and deactivated SARS-CoV-2, we estimate recovery rates of approximately 7-12% of viral RNA using our method. Results from 24 sewersheds across Upstate New York during the spring and summer of 2020 suggested that stronger signals of SARS-CoV-2 RNA from wastewater may be indicative of greater COVID-19 incidence in the represented service area approximately one week in advance. SARS-CoV-2 wastewater RNA was quantifiable in some service areas with daily positives tests of less than 1 per 10,000 people or when weekly positive test rates within a sewershed were as low as 1.7%. crAssphage DNA concentrations were significantly lower during periods of high flow in almost all areas studied. After accounting for flow rate and population served, crAssphage levels per capita were estimated to be about 1.35 × 10¹¹ and 2.42 × 10⁸ genome copies per day for DNA and RNA, respectively. A negative relationship between per capita crAssphage RNA and service area size was also observed likely reflecting degradation of RNA over long transit times. Our results reinforce the potential for wastewater surveillance to be used as a tool to supplement understanding of infectious disease transmission obtained by traditional testing and highlight the potential for crAssphage co-detection to improve interpretations of wastewater surveillance data.

Keywords: Cross-assembly phage; SARS-CoV-2; Viral concentration; Wastewater-based epidemiology.