Solid-liquid distribution of SARS-CoV-2 in primary effluent of a wastewater treatment plant

Femi F Oloye; Yuwei Xie; Mohsen Asadi; Jonathan K Challis; Charles A Osunla; Pu Xia; Jenna Cantin; Oluwabunmi P Femi-Oloye; Markus Brinkmann; Kerry N McPhedran; Mike Sadowski; Sudhir Pandey; Paul D Jones; Chand Mangat; Mark R Servos; John P Giesy

doi:10.1016/j.mex.2024.102645

Solid-liquid distribution of SARS-CoV-2 in primary effluent of a wastewater treatment plant

MethodsX. 2024 Mar 11:12:102645. doi: 10.1016/j.mex.2024.102645. eCollection 2024 Jun.

Authors

Femi F Oloye¹, Yuwei Xie^{1

2

3}, Mohsen Asadi⁴, Jonathan K Challis¹, Charles A Osunla¹, Pu Xia¹, Jenna Cantin¹, Oluwabunmi P Femi-Oloye¹, Markus Brinkmann^{1

5

6}, Kerry N McPhedran⁴, Mike Sadowski⁷, Sudhir Pandey⁷, Paul D Jones^{1

5}, Chand Mangat⁸, Mark R Servos⁹, John P Giesy^{1

10

11

12}

Affiliations

¹ Toxicology Centre, University of Saskatchewan, Saskatoon, SK, Canada.
² Nanjing Institute of Environmental Science, Ministry of Ecology and Environment of China, Nanjing 210042, China.
³ Key Laboratory of Pesticide Environmental Assessment and Pollution Control, Ministry of Ecology and Environment of China, Nanjing 210042, China.
⁴ Department of Civil, Geological and Environmental Engineering, College of Engineering, University of Saskatchewan, Saskatoon, SK, Canada.
⁵ School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK, Canada.
⁶ Global Institute for Water Security, University of Saskatchewan, Saskatoon, SK, Canada.
⁷ Saskatoon Water Department, Wastewater Treatment Plant, City of Saskatoon, Saskatoon, SK, Canada.
⁸ Antimicrobial Resistance and Nosocomial Infections, National Microbiology Laboratory / Public Health Agency of Canada, Canada.
⁹ Department of Biology, University of Waterloo, Waterloo, ON, Canada.
¹⁰ Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, SK, Canada.
¹¹ Department of Environmental Sciences, Baylor University, Waco, TX, USA.
¹² Department of Zoology and Center for Integrative Toxicology, Michigan State University, East Lansing, MI, USA.

Abstract

Distributions of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and fecal viral biomarkers between solid and liquid phases of wastewater are largely unknown. Herein, distributions of SARS-CoV-2, Pepper Mild Mottle Virus (PMMoV), and F-RNA bacteriophage group II (FRNAPH-II) were determined by viral RNA RT-qPCR. Comparison of viral recovery using three conventional fractionation methods included membrane filtration, a combination of mid-speed centrifugation and membrane filtration, and high-speed centrifugation. SARS-CoV-2 partitioned to the solids fraction in greater abundance compared to liquid fractions in a combination of mid-speed centrifugation and membrane filtration and high-speed centrifugation, but not in membrane filtration method in a particular assay, while fecal biomarkers (PMMoV and FRNAPH-II) exhibited the reciprocal relationship. The wastewater fractionation method had minimal effects on the solids-liquids distribution for all viral and phage markers tested; however, viral RNA load was significantly greater in solid-liquid fractions viral RNA loads compared with the than whole-wastewater PEG precipitation. A RNeasy PowerWater Kit with PCR inhibitor removal resulted in greater viral RNA loads and lesser PCR inhibition compared to a QIAamp Viral RNA Mini Kit without PCR inhibitor removal. These results support the development of improved methods and interpretation of WBE of SARS-CoV-2. •Distribution of SARS-CoV-2 to liquid and solid portions was addressed.•Addressing PCR inhibition is important in wastewater-based epidemiology.•Fraction methods have minimal effect.

Keywords: COVID-19; Environmental RNA; Method development for SARS-CoV-2 RNA partitioning in wastewater; PCR inhibitors; Surveillance; Wastewater-based epidemiology.