Fate of the Urinary Tract Virus BK Human Polyomavirus in Source-Separated Urine.

Human polyomaviruses are emerging pathogens that infect a large percentage of the human population and are excreted in urine. Consequently, urine that is collected for fertilizer production often has high concentrations of polyomavirus genes. We studied the fate of infectious double-stranded DNA (dsDNA) BK human polyomavirus (BKPyV) in hydrolyzed source-separated urine with infectivity assays and quantitative PCR (qPCR). Although BKPyV genomes persisted in the hydrolyzed urine for long periods of time (T90 [time required for 90% reduction in infectivity or gene copies] of >3 weeks), the viruses were rapidly inactivated (T90 of 1.1 to 11 h) in most of the tested urine samples. Interestingly, the infectivity of dsDNA bacteriophage surrogate T3 (T90 of 24 to 46 days) was much more persistent than that of BKPyV, highlighting a major shortcoming of using bacteriophages as human virus surrogates. Pasteurization and filtration experiments suggest that BKPyV virus inactivation was due to microorganism activity in the source-separated urine, and SDS-PAGE Western blots showed that BKPyV protein capsid disassembly is concurrent with inactivation. Our results imply that stored urine does not pose a substantial risk of BKPyV transmission, that qPCR and infectivity of the dsDNA surrogate do not accurately depict BKPyV fate, and that microbial inactivation is driven by structural elements of the BKPyV capsid.IMPORTANCE We demonstrate that a common urinary tract virus has a high susceptibility to the conditions in hydrolyzed urine and consequently would not be a substantial exposure route to humans using urine-derived fertilizers. The results have significant implications for understanding virus fate. First, by demonstrating that the dsDNA (double-stranded DNA) genome of the polyomavirus lasts for weeks despite infectivity lasting for hours to days, our work highlights the shortcomings of using qPCR to estimate risks from unculturable viruses. Second, commonly used dsDNA surrogate viruses survived for weeks under the same conditions that BK polyomavirus survived for only hours, highlighting issues with using virus surrogates to predict how human viruses will behave in the environment. Finally, our mechanistic inactivation analysis provides strong evidence that microbial activity drives rapid virus inactivation, likely through capsid disassembly. Overall, our work underlines how subtle structural differences between viruses can greatly impact their environmental fate.

Next generation sequencing approaches to evaluate water and wastewater quality.

The emergence of next generation sequencing (NGS) is revolutionizing the potential to address complex microbiological challenges in the water industry. NGS technologies can provide holistic insight into microbial communities and their functional capacities in water and wastewater systems, thus eliminating the need to develop a new assay for each target organism or gene. However, several barriers have hampered wide-scale adoption of NGS by the water industry, including cost, need for specialized expertise and equipment, challenges with data analysis and interpretation, lack of standardized methods, and the rapid pace of development of new technologies. In this critical review, we provide an overview of the current state of the science of NGS technologies as they apply to water, wastewater, and recycled water. In addition, a systematic literature review was conducted in which we identified over 600 peer-reviewed journal articles on this topic and summarized their contributions to six key areas relevant to the water and wastewater fields: taxonomic classification and pathogen detection, functional and catabolic gene characterization, antimicrobial resistance (AMR) profiling, bacterial toxicity characterization, Cyanobacteria and harmful algal bloom identification, and virus characterization. For each application, we have presented key trends, noteworthy advancements, and proposed future directions. Finally, key needs to advance NGS technologies for broader application in water and wastewater fields are assessed.

Environmental strains potentially contribute to the proliferation and maintenance of antibiotic resistance in drinking water: A case study of Cupriavidus metallidurans.

Fitness costs of antibiotic resistance detrimentally affect the fate of resistance carriers. Intriguingly, numerous antibiotic resistant bacteria (ARB) have been detected despite the low concentration of antibiotics in drinking water. To reveal the causes of this discrepancy, we investigated the fitness cost of antimicrobial resistance in strain Cupriavidus metallidurans CR2 which was isolated from a drinking water filter. Pure culture and 1:1 competitive experiments were established at different nutrient levels. The growth rates of strain C. metallidurans CR2 significantly decreased when pure cultured under poor nutrient conditions, however, the multi-resistance and the resistance megaplasmids were well maintained. Competitiveness costs were observed in C. metallidurans when separately co-cultured with environmentally-isolated Flectobacillus BS1 and Pseudomonas sp. S3, while C. metallidurans was outnumbered by the rivals with a decrease of 1-2 logs. But the majority of C. metallidurans retained the plasmids under oligotrophic conditions even after 144 h (1.99 and 0.199 mg C/L). Additionally, C. metallidurans CR2 has a higher tolerance to chlorine and chloramine, which potentially could become prevalent in the subsequent distribution systems other than drinking water treatment plant. As a potential pathogen, the prevalence of Cupriavidus metallidurans in drinking water would also pose certain threats to human health.