Wastewater-based monitoring of SARS-CoV-2: Insights from small and low-income Brazilian cities

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of COVID-19, has affected millions of people worldwide and significantly impacted public health. The detection of viral RNA in the feces of infected individuals prompted several countries to adopt wastewater-based epidemiology (WBE) as a tool to monitor SARS-CoV-2 circulation within communities. However, most surveillance studies have focused on large urban centers. This study aimed to analyze the presence and persistence of SARS-CoV-2 in wastewater from two small Brazilian towns with low socioeconomic indicators, limited sewage coverage, and low population density, located in northern Minas Gerais. Wastewater samples were collected biweekly over 12 months, between 2023 and 2024, from three sampling points in Salinas and one in Rubelita. Viral RNA was concentrated using electronegative membranes, and SARS-CoV-2 detection was performed using real-time quantitative polymerase chain reaction. SARS-CoV-2 RNA was detected at all sampling points, with a high frequency of positive samples observed in May, June, October, November 2023, and January 2024. Notably, viral RNA was detected even after the World Health Organization declared the end of the global health emergency on May 5, 2023, and in the absence of officially reported human cases. These findings underscore the value of wastewater surveillance as a tool for monitoring SARS-CoV-2, particularly in areas with limited clinical testing capacity. The study highlights the potential of WBE to detect ongoing viral transmission across diverse urban and socioeconomic contexts, including regions with case underreporting and challenges in mass testing, thereby supporting public health preparedness and response.

Keywords

Coronavirus / Wastewater-based epidemiology / Severe acute respiratory syndrome coronavirus 2 / COVID-19 pandemic

Funding

This work received financial support from Network for monitoring COVID-19 in wastewater (REMONAR) (Ministry of Science, Technology and Innovation [MCTI], grant no. 400284/2022-7, and National Council for Scientific and Technological Development [CNPq], grant no. 401933/2020–2), and Minas Gerais Research Foundation (Fapemig) (grant no. APQ-01403-21). GPB and PAAS were awarded CNPq scholarships (grant nos. 400284/2022-7 and 401933/2020–2).

Conflict of interest

The authors declare they have no competing interests.

References

[1] Ahmed W., Angel N., Edson J., Bibby K., Bivins A., O’Brien J.W., et al. (2020). First confirmed detection of SARS-CoV-2 in untreated wastewater in Australia: A proof of concept for the wastewater surveillance of COVID-19 in the community. Science of The Total Environment, 728:138764. https://doi.org/10.1016/j.scitotenv.2020.138764

[2] Ahmed W., Bertsch P.M., Bibby K., Haramoto E., Hewitt J., Huygens F., et al. (2020). Decay of SARS-CoV-2 and surrogate murine hepatitis virus RNA in untreated wastewater to inform application in wastewater-based epidemiology. Environmental Research, 191:110092. https://doi.org/10.1016/j.envres.2020.110092

[3] Ahmed W., Bivins A., Metcalfe S., Smith W.J.M., Verbyla M.E., Symonds E.M., et al. (2022). Evaluation of process limit of detection and quantification variation of SARS-CoV-2 RT-qPCR and RT-dPCR assays for wastewater surveillance. Water Research, 213:118132. https://doi.org/10.1016/j.watres.2022.118132

[4] Ahmed W., Harwood V.J., Gyawali P., Sidhu J.P.S., & Toze S. (2015). Comparison of concentration methods for quantitative detection of sewage-associated viral markers in environmental waters. Applied and Environmental Microbiology, 81(6):2042-2049. https://doi.org/10.1128/AEM.03851-14

[5] Alvares C.A., Stape J.L., Sentelhas P.C., De Moraes Gonçalves J.L., & Sparovek G. (2013). Köppen’s climate classification map for Brazil. Meteorologische Zeitschrift, 22(6):711-728. https://doi.org/10.1127/0941-2948/2013/0507

[6] Arons M.M., Hatfield K.M., Reddy S.C., Kimball A., James A., Jacobs J.R., et al. (2020). Presymptomatic SARS-CoV-2 infections and transmission in a skilled nursing facility. New England Journal of Medicine, 382(22):2081-2090. https://doi.org/10.1056/NEJMoa2008457

[7] Benksim A., Ait Addi R., & Cherkaoui M. (2020). Vulnerability and fragility expose older adults to the potential dangers of COVID-19 pandemic. Iranian Journal of Public Health, 49(1):122-124. https://doi.org/10.18502/ijph.v49iS1.3682

[8] Berra T.Z., Alves Y.M., Popolin M.A.P., Da Costa F.B.P., Tavares R.B.V., Tártaro A.F., et al. (2024). The COVID-19 pandemic in Brazil: Space-time approach of cases, deaths, and vaccination coverage (February 2020 - April 2024). BMC Infectious Diseases, 24(1):704. https://doi.org/10.1186/s12879-024-09598-1

[9] Bivins A., Greaves J., Fischer R., Yinda K.C., Ahmed W., Kitajima M., et al. (2020). Persistence of SARS-CoV-2 in water and wastewater. Environmental Science and Technology Letters, 7(12):937-942. https://doi.org/10.1021/acs.estlett.0c00730

[10] Chernicharo C.A.L., Araújo J.C., Mota Filho C.R., Bressani- Ribeiro T., Chamhum-Silva L., Leal C.D., et al. (2021). Sewage monitoring as a surveillance epidemiological tool for COVID-19 control: A case study in the city of Belo Horizonte. Sanitary and Environmental Engineering, 26(4):691-699. https://doi.org/10.1590/s1413-415220200243

[11] Claro I.C.M., Cabral A.D., Augusto M.R., Duran A.F.A., Graciosa M.C.P., Fonseca F.L.A., et al. (2021). Long-term monitoring of SARS-COV-2 RNA in wastewater in Brazil: A more responsive and economical approach. Water Research, 203:117534. https://doi.org/10.1016/j.watres.2021.117534

[12] Corman V.M., Landt O., Kaiser M., Molenkamp R., Meijer A., Chu D.K., et al. (2020). Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR. Eurosurveillance, 25(3):2000045. https://doi.org/10.2807/1560-7917.ES.2020.25.3.2000045

[13] Corpuz M.V.A., Buonerba A., Zarra T., Hasan S.W., Korshin G.V., Belgiorno V., et al. (2022). Advances in virus detection methods for wastewater-based epidemiological applications. Case Studies in Chemical and Environmental Engineering, 6:100238. https://doi.org/10.1016/j.cscee.2022.100238

[14] De Freitas Bueno, R., Claro I.C.M., Augusto M.R., Duran A.F.A., Camillo L.M.B., Cabral A.D., et al. (2022). Wastewater-based epidemiology: A Brazilian SARS-COV-2 surveillance experience. Journal of Environmental Chemical Engineering, 10(5):108298. https://doi.org/10.1016/j.jece.2022.108298

[15] Dilson Godinho Foundation. (2024). Units. Available from:https://www.fundacaodilsongodinho.org.br/page/unidades [Last accessed on 2026 Jan 20].

[16] Eales O., Plank M.J., Cowling B.J., Howden B.P., Kucharski A.J., Sullivan S.G., et al. (2024). Key challenges for respiratory virus surveillance while transitioning out of acute phase of COVID-19 pandemic. Emerging Infectious Diseases, 30(2):e230768. https://doi.org/10.3201/eid3002.230768

[17] Ebrille E., Lucciola M.T., Amellone C., Ballocca F., Orlando F., & Giammaria M. (2020). Syncope as the presenting symptom of COVID-19 infection. HeartRhythm Case Reports, 6(7):363-366. https://doi.org/10.1016/j.hrcr.2020.04.015

[18] Ferraro B.G., Veneri C., Mancini P., Iaconelli M., Suffredini E., Bonadonna L., et al. (2022). A state-of-the-art scoping review on SARS-CoV-2 in sewage focusing on the potential of wastewater surveillance for the monitoring of the COVID-19 pandemic. Food and Environmental Virology, 14(4):315-354. https://doi.org/10.1007/s12560-021-09498-6

[19] Gudra D., Dejus S., Bartkevics V., Roga A., Kalnina I., Strods M., et al. (2022). Detection of SARS-CoV-2 RNA in wastewater and importance of population size assessment in smaller cities: An exploratory case study from two municipalities in Latvia. Science of The Total Environment, 823:153775. https://doi.org/10.1016/j.scitotenv.2022.153775

[20] Gunawardana M., Breslin J., Cortez J.M., Rivera S., Webster S., Ibarrondo F.J., et al. (2021). Longitudinal COVID- 19 surveillance and characterization in the workplace with public health and diagnostic endpoints. MSphere, 6(4):e0054221. https://doi.org/10.1128/mSphere.00542-21

[21] Hart O.E., & Halden R.U. (2020). Computational analysis of SARS-CoV-2/COVID-19 surveillance by wastewater-based epidemiology locally and globally: Feasibility, economy, opportunities and challenges. Science of The Total Environment, 730:138875. https://doi.org/10.1016/j.scitotenv.2020.138875

[22] IBGE. (2022). Salinas (MG). Cidades e Estados. IBGE. Available from:https://www.ibge.gov.br/cidades-e-estados/mg/salinas.html [Last accessed on 2026 Jan 20].

[23] Juel M.A.I., Stark N., Nicolosi B., Lontai J., Lambirth K., Schlueter J., et al. (2021). Performance evaluation of virus concentration methods for implementing SARS-CoV-2 wastewater based epidemiology emphasizing quick data turnaround. Science of The Total Environment, 801:149656. https://doi.org/10.1016/j.scitotenv.2021.149656

[24] Kallem P., Hegab H.M., Alsafar H., Hasan S.W., & Banat F. (2023). SARS-CoV-2 detection and inactivation in water and wastewater: Review on analytical methods, limitations and future research recommendations. Emerging Microbes and Infections, 12(2):2222850. https://doi.org/10.1080/22221751.2023.2222850

[25] Korajkic A., McMinn B.R., Pemberton A.C., Kelleher J., & Ahmed W. (2024). The comparison of decay rates of infectious SARS-CoV-2 and viral RNA in environmental waters and wastewater. Science of The Total Environment, 946:174379. https://doi.org/10.1016/j.scitotenv.2024.174379

[26] Lopreite M., Panzarasa P., Puliga M., & Riccaboni M. (2021). Early warnings of COVID-19 outbreaks across Europe from social media. Scientific Reports, 11(1):2147. https://doi.org/10.1038/s41598-021-81333-1

[27] Luiz A., Lopes S., & De Cássia Gusmão G. (2012). A Relação Entre Pobreza e Desigualdade na Região Norte de Minas Gerais (The Relationship between Poverty and Inequality in the Northern Region of Minas Gerais). XV Seminário Sobre Economia Mineira:Anais. Anais Eletrônicos. Belo Horizonte: UFMG/Cedeplar. Available from:https://diamantina.cedeplar.ufmg.br/portal/download/diamantina-2012/a_relacao_entre_pobreza_e_desigualdade.pdf [Last accessed on 2026 Jan 20].

[28] Medema G., Heijnen L., Elsinga G., Italiaander R., & Brouwer A. (2020). Presence of SARS-coronavirus-2 RNA in sewage and correlation with reported COVID-19 prevalence in the early stage of the epidemic in the Netherlands. Environmental Science and Technology Letters, 7(7):511-516. https://doi.org/10.1021/acs.estlett.0c00357

[29] Ministry of Health. (2024). Coronavirus Brazil. Available from:https://covid.saude.gov.br [Last accessed on 2026 Jan 20].

[30] Oliveira W.K., Duarte E., França G.V.A., & Garcia L.P. (2020). How Brazil can hold back COVID-19. Epidemiology and Health Services, 29(2):e2020000200023. https://doi.org/10.5123/S1679-49742020000200023

[31] Prado T., Fumian T.M., Mannarino C.F., Resende P.C., Motta F.C., Eppinghaus A.L.F., et al. (2021). Wastewater-based epidemiology as a useful tool to track SARS-CoV-2 and support public health policies at municipal level in Brazil. Water Research, 191:116810. https://doi.org/10.1016/j.watres.2021.116810

[32] Rodda L.B., Netland J., Shehata L., Pruner K.B., Morawski P.A., Thouvenel C.D., et al. (2021). Functional SARS-CoV-2- specific immune memory persists after mild COVID-19. Cell, 184(1):169-183.e17. https://doi.org/10.1016/j.cell.2020.11.029

[33] Silva C.C.M., Santos C.R., Céleri E.P., Salles D., Fardin J.M., Pussi K.F., et al. (2024). An epidemiological assessment of SARS-CoV-2 in the sewage system of a higher education institution. Annals of Global Health, 90(1):50. https://doi.org/10.5334/aogh.4413

[34] Silva C.L.F., Da Silva M.S., Dos Santos D.S., Braga T.G.M., & De Freitas T.P.M., (2020). Socio-environmental impacts of the SARS-CoV-2 (COVID-19) pandemic in Brazil: How to overcome them? Revista Brasileira de Educação Ambiental (RevBEA), 15(4):220-236. https://doi.org/10.34024/revbea.2020.v15.10846

[35] State Health Department of Minas Gerais. (2024). COVID Monitoramento. Available from:https://info.saude.mg.gov.br/1/paineis/2 [Last accessed on 2026 Jan 20].

[36] State Health Department of Minas Gerais. (2024). Portal de Informações em Saúde. Available from:https://info.saude.mg.gov.br/1/paineis/2 [Last accessed on 2026 Jan 20].

[37] Tateishi S., Hamada K., Emoto N., Abe K., Abe K., Kawasaki Y., et al. (2025). Facility wastewater monitoring as an effective tool for pandemic infection control: An experience in COVID-19 pandemic with long-term monitoring. Journal of Infection and Chemotherapy, 31(1):102499. https://doi.org/10.1016/j.jiac.2024.08.014

[38] World Health Organization. (2023). Statement on the Fifteenth Meeting of the IHR (2005) Emergency Committee on the COVID-19 Pandemic. Available from: https://www.who.int/news/item/05-05-2023-statement-on-the-fifteenth-meeting-of-the-international-health-regulations-(2005)-emergency-committee-regarding-the-coronavirus-disease-(covid-19)-pandemic [Last accessed on 2026 Jan 20].

[39] Pan American Health Organization. (2023). WHO Declares end to Public Health Emergency of International Concern regarding COVID-19. Available from: https://www.paho.org/pt/noticias/5-5-2023-oms-declara-fim-da-emergencia-saude-publica-importancia-internacional-referente [Last accessed on 2026 Jan 20].

[40] Wu Y., Guo C., Tang L., Hong Z., Zhou J., Dong X., et al. (2020). Prolonged presence of SARS-CoV-2 viral RNA in faecal samples. The Lancet Gastroenterology and Hepatology, 5(5):434-435. https://doi.org/10.1016/S2468-1253(20)30083-2