Wastewater-based epidemiological surveillance to monitor the prevalence of SARS-CoV-2 in developing countries with onsite sanitation facilities

Wastewater-based epidemiology (WBE) has emerged as a valuable approach for forecasting disease outbreaks in developed countries with a centralized sewage infrastructure. On the other hand, due to the absence of well-defined and systematic sewage networks, WBE is challenging to implement in developing countries like Bangladesh where most people live in rural areas. Identification of appropriate locations for rural Hotspot Based Sampling (HBS) and urban Drain Based Sampling (DBS) are critical to enable WBE based monitoring system. We investigated the best sampling locations from both urban and rural areas in Bangladesh after evaluating the sanitation infrastructure for forecasting COVID-19 prevalence. A total of 168 wastewater samples were collected from 14 districts of Bangladesh during each of the two peak pandemic seasons. RT-qPCR commercial kits were used to target ORF1ab and N genes. The presence of SARS-CoV-2 genetic materials was found in 98% (165/168) and 95% (160/168) wastewater samples in the first and second round sampling, respectively. Although wastewater effluents from both the marketplace and isolation center drains were found with the highest amount of genetic materials according to the mixed model, quantifiable SARS-CoV-2 RNAs were also identified in the other four sampling sites. Hence, wastewater samples of the marketplace in rural areas and isolation centers in urban areas can be considered the appropriate sampling sites to detect contagion hotspots. This is the first complete study to detect SARS-CoV-2 genetic components in wastewater samples collected from rural and urban areas for monitoring the COVID-19 pandemic. The results based on the study revealed a correlation between viral copy numbers in wastewater samples and SARS-CoV-2 positive cases reported by the Directorate General of Health Services (DGHS) as part of the national surveillance program for COVID-19 prevention. The findings of this study will help in setting strategies and guidelines for the selection of appropriate sampling sites, which will facilitate in development of comprehensive wastewater-based epidemiological systems for surveillance of rural and urban areas of low-income countries with inadequate sewage infrastructure.This research was supported by Water Aid Bangladesh, North South University, Dhaka, COVID-19 Diagnostic Lab, Department of Microbiology, Noakhali Science and Technology University (NSTU), Noakhali, Bangladesh, the International Training Network of Bangladesh University of Engineering and Technology (ITN-BUET) - Centre for Water Supply and Waste Management, and KTH Royal Institute of Technology, Sweden. We acknowledge the sincere help and support of the staff and volunteers of NSTU-COVID-19 Diagnostic Lab, Noakhali Science and Technology University, Bangladesh during the different phases of the study. PB and MTI acknowledge the Life Science Technology Platform, Science for Life Laboratory for the seed funding to initiate the wastewater-based epidemiological studies for SARS-CoV-2 in Bangladesh. We would also like to acknowledge the two anonymous reviewers for their critical comments as well as their thoughtful insights, which has significantly improved the manuscript.Peer reviewe