A Review of Molecular Methods for Viral Detection in Water and Future Public Health Perspectives
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A Review of Molecular Methods for Viral Detection in Water and Future Public Health Perspectives

Jianing Song 1*
1 University of Florida
*Corresponding author: Song.jianing@ufl.edu
Published on 24 September 2025
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TNS Vol.138
ISSN (Print): 2753-8826
ISSN (Online): 2753-8818
ISBN (Print): 978-1-80590-381-9
ISBN (Online): 978-1-80590-382-6
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Abstract

Safe water is essential for public health, but environmental waters frequently contain viral contaminants. Molecular detection methods, particularly Polymerase Chain Reaction (PCR)-based approaches, have transformed water quality monitoring by providing rapid and sensitive analysis without the need for cultivation. This review focuses on two major technologies: quantitative PCR (qPCR) and digital PCR (dPCR). qPCR remains widely used for virus surveillance, source tracking, and infectivity assessments, while dPCR offers absolute quantification, improved tolerance to inhibitors, and greater precision for low-level targets. Both methods require quantitative criteria for performance evaluation and face some challenges, including matrix inhibition, lack of viability confirmation, and the need for standardized workflows. Future progress is expected through improved robustness, more efficient sample processing, enhanced multiplexing, and integration with risk modeling and viability assays. Continued reductions in cost and gains in throughput may broaden access to dPCR, and the complementary use of qPCR and dPCR can further strengthen water quality surveillance and public health protection.

Keywords:

Viral detection, dPCR, qPCR, water environment

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Song,J. (2025). A Review of Molecular Methods for Viral Detection in Water and Future Public Health Perspectives. Theoretical and Natural Science,138,43-48.

References

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[5]. Bixler, T. S., Collins, M. R., & Mo, W. (2024). Risk-based public health impact assessment for drinking water contamination emergencies. Science of the Total Environment, 931, Article 172966. https: //doi.org/10.1016/j.scitotenv.2024.172966.

[6]. Brown, M. R., Camézuli, S., Davenport, R. J., Petelenz‑Kurdziel, E., Øvreås, L., & Curtis, T. P. (2015). Flow cytometric quantification of viruses in activated sludge. Water Research, 68, 414–422. https: //doi.org/10.1016/j.watres.2014.10.018.

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[9]. Liu, G., Qu, J., Rose, J., & Medema, G. (2022). Roadmap for managing SARS-CoV-2 and other viruses in the water environment for public health. Engineering, 12, 139–144. https: //doi.org/10.1016/j.eng.2020.09.015.

[10]. Straub, T. M., & Chandler, D. P. (2003). Towards a unified system for detecting waterborne pathogens. Journal of Microbiological Methods, 53, 185–197. https: //doi.org/10.1016/S0167-7012(03)00023-X.

[11]. Inkster, T., Caldwell, I., Aird, H., Willis, C., Lai, S., & Mallon, J. (2024). Development and evaluation of test methods for the detection and enumeration of opportunistic waterborne pathogens from the hospital environment. Journal of Hospital Infection, 149, 98–103. https: //doi.org/10.1016/j.jhin.2024.04.008.

[12]. Heijnen, L., de Vries, H. J., van Pelt, G., Stroobach, E., Atsma, A., Vranken, J., De Maeyer, K., Vissers, L., & Medema, G. (2024). Qualitative detection of E. coli in distributed drinking water using real-time reverse transcription PCR targeting 16S rRNA: Validation and practical experiences. Water Research, 259, 121843. https: //doi.org/10.1016/j.watres.2024.121843.

[13]. Tiwari, A., Ahmed, W., Oikarinen, S., Sherchan, S. P., Heikinheimo, A., Jiang, G., Simpson, S. L., Greaves, J., & Bivins, A. (2022). Application of digital PCR for public health-related water quality monitoring. Science of the Total Environment, 837, 155663. https: //doi.org/10.1016/j.scitotenv.2022.155663.

[14]. Sivaganesan, M., Dean, S. A., Willis, J. R., Friedman, S. D., Haugland, R., & Shanks, O. C. (2025). A streamlined qPCR method for characterization of Enterococcus spp. levels in ambient surface water samples. Water Research, 284, 123936. https: //doi.org/10.1016/j.watres.2025.123936.

[15]. Sun, K., Fan, G., Dong, H., Fan, Y., Xie, Y., Liang, K., & Zhang, Y. (2024). Water-cooling-based and low-cost qPCR device for rapid nucleic acid analysis. Sensors and Actuators A: Physical, 375, 115496. https: //doi.org/10.1016/j.sna.2024.115496.

[16]. Zhao, B., Fujita, T., Nihei, Y., Yu, Z., Chen, X., Tanaka, H., & Ihara, M. (2023). Tracking community infection dynamics of COVID-19 by monitoring SARS-CoV-2 RNA in wastewater, counting positive reactions by qPCR. Science of The Total Environment, 904, 166420. https: //doi.org/10.1016/j.scitotenv.2023.166420.

[17]. Vu Duc Canh, Torii, S., Furumai, H., & Katayama, H. (2021). Application of capsid integrity (RT-)qPCR to assessing occurrence of intact viruses in surface water and tap water in Japan. Water Research, 189, 116674. https: //doi.org/10.1016/j.watres.2020.116674.

[18]. National Institute of Standards and Technology (NIST). (2021). Certificate of Analysis: Standard Reference Material 2917, Plasmid DNA for Fecal Indicator Detection and Identification. Gaithersburg, MD: U.S. Department of Commerce. https: //doi.org/10.6028/NIST.SP.260-221.

[19]. Borgolte, S., Menzel, W., & Varrelmann, M. (2025). One-step TaqMan® RT-qPCR detection of sugar beet-infecting poleroviruses in Myzus persicae from yellow water pan traps opens up new possibilities for early risk assessment of virus yellows disease. Journal of Virological Methods, 331, 115052. https: //doi.org/10.1016/j.jviromet.2024.115052.

[20]. Sthapit, N., Thapa, B. R., Limam, T., Maharjan, R., Dhakal, U., Jhun, I., Cho, K. H., Sherchan, S. P., Yu, X., Ouyang, W., & Han, J. (2025). Digital PCR-based assessment of pathogens in wastewater and antibiotic resistance genes in drinking water of the Kathmandu Valley, Nepal. Journal of Environmental Management, 389, 126120. https: //doi.org/10.1016/j.jenvman.2025.126120.

[21]. Roman, V., Jourdain, F., Pele, E., Brottet, E., Guinard, A., Mouly, D., Medragh, S., Cordevant, C., Gassilloud, B., & Dina, J. (2025). Tracking wild-type measles virus in wastewater using multiplex RT-dPCR: A novel tool for measles surveillance. Water Research, 287, 124379. https: //doi.org/10.1016/j.watres.2025.124379.

[22]. Chailler, E., Le Bayon, H., Dairain, A., Riquet, F., Stout, L., Chambouvet, A., & Smits, M. (2025). Digital PCR (dPCR) vs. quantitative PCR (qPCR) approaches for quantification of two Perkinsus species within clam tissue samples. Journal of Invertebrate Pathology, 213, 108417. https: //doi.org/10.1016/j.jip.2025.108417.

References

[1]. Alao, J. O., Otorkpa, O. J., Ayejoto, D. A., & Saqr, A. M. (2025). Assessing the community knowledge on waste management practices, drinking water source systems, and the possible implications on public health systems. Cleaner Waste Systems, 11, Article 100295. https: //doi.org/10.1016/j.clwas.2025.100295.

[2]. USEPA, 2022. Toxics Release Inventory (TRI) Program. US EPA [WWW Document]. URL. https: //www.epa.gov/toxics-release-inventory-tri-program (accessed 6.26.22).

[3]. Haque, M. A., Khatun, B., Jewel, M. A. S., Ara, J., Kazal, M. S. I., & Hasan, J. (2024). Assessment of water quality and heavy metal indices in a tropical freshwater river for aquatic life and public health standard. Ecological Indicators, 169, Article 112862. https: //doi.org/10.1016/j.ecolind.2024.112862.

[4]. Zhang, S., Fu, K., Gao, S., Liang, B., Lu, J., & Fu, G. (2023). Bioaccumulation of heavy metals in the water, sediment, and organisms from the sea ranching areas of Haizhou Bay in China. Water, 15(12), Article 2218. https: //doi.org/10.3390/w15122218.

[5]. Bixler, T. S., Collins, M. R., & Mo, W. (2024). Risk-based public health impact assessment for drinking water contamination emergencies. Science of the Total Environment, 931, Article 172966. https: //doi.org/10.1016/j.scitotenv.2024.172966.

[6]. Brown, M. R., Camézuli, S., Davenport, R. J., Petelenz‑Kurdziel, E., Øvreås, L., & Curtis, T. P. (2015). Flow cytometric quantification of viruses in activated sludge. Water Research, 68, 414–422. https: //doi.org/10.1016/j.watres.2014.10.018.

[7]. Ward, R. L., Bernstein, D. I., Young, E. C., Sherwood, J. R., Knowlton, D. R., & Schiff, G. M. (1986). Human rotavirus studies in volunteers: Determination of infectious dose and serological response to infection. Journal of Infectious Diseases, 154(5), 871–880. https: //doi.org/10.1093/infdis/154.5.871.

[8]. World Health Organization. (2011). Guidelines for drinking-water quality (4th ed.). https: //www.who.int/publications/i/item/9789241548151.

[9]. Liu, G., Qu, J., Rose, J., & Medema, G. (2022). Roadmap for managing SARS-CoV-2 and other viruses in the water environment for public health. Engineering, 12, 139–144. https: //doi.org/10.1016/j.eng.2020.09.015.

[10]. Straub, T. M., & Chandler, D. P. (2003). Towards a unified system for detecting waterborne pathogens. Journal of Microbiological Methods, 53, 185–197. https: //doi.org/10.1016/S0167-7012(03)00023-X.

[11]. Inkster, T., Caldwell, I., Aird, H., Willis, C., Lai, S., & Mallon, J. (2024). Development and evaluation of test methods for the detection and enumeration of opportunistic waterborne pathogens from the hospital environment. Journal of Hospital Infection, 149, 98–103. https: //doi.org/10.1016/j.jhin.2024.04.008.

[12]. Heijnen, L., de Vries, H. J., van Pelt, G., Stroobach, E., Atsma, A., Vranken, J., De Maeyer, K., Vissers, L., & Medema, G. (2024). Qualitative detection of E. coli in distributed drinking water using real-time reverse transcription PCR targeting 16S rRNA: Validation and practical experiences. Water Research, 259, 121843. https: //doi.org/10.1016/j.watres.2024.121843.

[13]. Tiwari, A., Ahmed, W., Oikarinen, S., Sherchan, S. P., Heikinheimo, A., Jiang, G., Simpson, S. L., Greaves, J., & Bivins, A. (2022). Application of digital PCR for public health-related water quality monitoring. Science of the Total Environment, 837, 155663. https: //doi.org/10.1016/j.scitotenv.2022.155663.

[14]. Sivaganesan, M., Dean, S. A., Willis, J. R., Friedman, S. D., Haugland, R., & Shanks, O. C. (2025). A streamlined qPCR method for characterization of Enterococcus spp. levels in ambient surface water samples. Water Research, 284, 123936. https: //doi.org/10.1016/j.watres.2025.123936.

[15]. Sun, K., Fan, G., Dong, H., Fan, Y., Xie, Y., Liang, K., & Zhang, Y. (2024). Water-cooling-based and low-cost qPCR device for rapid nucleic acid analysis. Sensors and Actuators A: Physical, 375, 115496. https: //doi.org/10.1016/j.sna.2024.115496.

[16]. Zhao, B., Fujita, T., Nihei, Y., Yu, Z., Chen, X., Tanaka, H., & Ihara, M. (2023). Tracking community infection dynamics of COVID-19 by monitoring SARS-CoV-2 RNA in wastewater, counting positive reactions by qPCR. Science of The Total Environment, 904, 166420. https: //doi.org/10.1016/j.scitotenv.2023.166420.

[17]. Vu Duc Canh, Torii, S., Furumai, H., & Katayama, H. (2021). Application of capsid integrity (RT-)qPCR to assessing occurrence of intact viruses in surface water and tap water in Japan. Water Research, 189, 116674. https: //doi.org/10.1016/j.watres.2020.116674.

[18]. National Institute of Standards and Technology (NIST). (2021). Certificate of Analysis: Standard Reference Material 2917, Plasmid DNA for Fecal Indicator Detection and Identification. Gaithersburg, MD: U.S. Department of Commerce. https: //doi.org/10.6028/NIST.SP.260-221.

[19]. Borgolte, S., Menzel, W., & Varrelmann, M. (2025). One-step TaqMan® RT-qPCR detection of sugar beet-infecting poleroviruses in Myzus persicae from yellow water pan traps opens up new possibilities for early risk assessment of virus yellows disease. Journal of Virological Methods, 331, 115052. https: //doi.org/10.1016/j.jviromet.2024.115052.

[20]. Sthapit, N., Thapa, B. R., Limam, T., Maharjan, R., Dhakal, U., Jhun, I., Cho, K. H., Sherchan, S. P., Yu, X., Ouyang, W., & Han, J. (2025). Digital PCR-based assessment of pathogens in wastewater and antibiotic resistance genes in drinking water of the Kathmandu Valley, Nepal. Journal of Environmental Management, 389, 126120. https: //doi.org/10.1016/j.jenvman.2025.126120.

[21]. Roman, V., Jourdain, F., Pele, E., Brottet, E., Guinard, A., Mouly, D., Medragh, S., Cordevant, C., Gassilloud, B., & Dina, J. (2025). Tracking wild-type measles virus in wastewater using multiplex RT-dPCR: A novel tool for measles surveillance. Water Research, 287, 124379. https: //doi.org/10.1016/j.watres.2025.124379.

[22]. Chailler, E., Le Bayon, H., Dairain, A., Riquet, F., Stout, L., Chambouvet, A., & Smits, M. (2025). Digital PCR (dPCR) vs. quantitative PCR (qPCR) approaches for quantification of two Perkinsus species within clam tissue samples. Journal of Invertebrate Pathology, 213, 108417. https: //doi.org/10.1016/j.jip.2025.108417.

Cite this article

Song,J. (2025). A Review of Molecular Methods for Viral Detection in Water and Future Public Health Perspectives. Theoretical and Natural Science,138,43-48.

Data availability

The datasets used and/or analyzed during the current study will be available from the authors upon reasonable request.

About volume

Volume title: Proceedings of ICBioMed 2025 Symposium: Computational Modelling and Simulation for Biology and Medicine

ISBN: 978-1-80590-381-9(Print) / 978-1-80590-382-6(Online)
Editor: Alan Wang, Roman Bauer
Conference website: https://2025.icbiomed.org/london.html
Conference date: 19 October 2025
Series: Theoretical and Natural Science
Volume number: Vol.138
ISSN: 2753-8818(Print) / 2753-8826(Online)

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