Current monitoring and regulatory approaches for microbial water quality, relying on the Fecal Indicator Bacteria (FIB), insufficiently protect public health due to a poor representation of viruses. Direct measurement of viral pathogens by conventional means is often incomplete due to the diversity of viral pathogens (>200) that may be in the environment. In addition, due to a historical reliance on FIB, there is relatively little knowledge regarding viral pathogen presence and diversity in wastewater. The PI plans to develop quantitative viral metagenomics, sequencing of viral DNA/RNA, for viral water quality monitoring. The proposed technical developments will significantly advance not only water quality monitoring but also viral pathogen detection in other settings (e.g., food safety). The proposed efforts in viral water quality assessment will also offer an educational development opportunity for students (high school through graduate school), water quality professionals, and the general public.
Specific technical goals include developing a rapid viral isolation and concentration approach and an improved methodology to remove free nucleic acids from non-viable viruses, an open-source bioinformatics pipeline to quantitatively identify viral pathogens from DNA sequence data, and a national survey of viral pathogen diversity in sewage. These developments may lead to new methods for monitoring microbial water quality as well as a methodology for viral water quality monitoring to elucidate the presence and diversity of viral pathogens in the water environment. Specifically, current metagenomics methods are applied non-quantitatively (e.g., on a relative abundance basis) and annotation accuracy is unverified; viral concentration methods are too slow for practical application and current methods for free DNA removal are inconsistent. Finally, little is known about viral pathogen diversity and dynamics in wastewater impacted systems. Application of viral metagenomics will help to inform investigations into the ecology of disease. Each task represents a discrete advance with applications beyond water quality in public health, microbiology, and bioinformatics. Additionally, bioinformatics tasks will be applicable to any sequencing platform and will be released as an open-access bioinformatic tool for use by researchers worldwide. Enabling quantitative and target-independent virus detection will facilitate improved water quality management, reducing overall human health impacts and informing infrastructure upgrades. Specific educational goals include a hands-on educational module for high school students to characterize microbial water quality, research experiences for high school and undergraduate students, expanding a current interactive exhibit established by the project PI at the Carnegie Science Center, and annual viral water quality webinars to be conducted through the Water Environment and Reuse Foundation. This effort will result in the release of an open-source bioinformatics viral annotation tool and an educational module for the high school classroom, extending the impact of technical and educational developments beyond the immediate project scope.
