Removal of Waterborne Viruses by Tetrahymena pyriformis Is Virus-Specific and Coincides with Changes in Protist Swimming Speed.

Biological treatment of waterborne viruses, specifically grazing of viruses by protists, can enhance microbial water quality while avoiding the production of toxic byproducts and high energy costs. However, tangible applications are limited by the lack of understanding of the underlying mechanisms. Here, we examined the feeding behavior of Tetrahymena pyriformis ciliates on 13 viruses, including bacteriophages, enteric viruses, and respiratory viruses. Significant differences in virus removal by T. pyriformis were observed, ranging from no removal (Qbeta, coxsackievirus B5) to ≥2.7 log10 (JC polyomavirus) after 48 h of co-incubation of the protist with the virus. Removal rates were conserved even when protists were co-incubated with multiple viruses simultaneously. Video analysis revealed that the extent of virus removal was correlated with an increase in the protists' swimming speed, a behavioral trait consistent with the protists' response to the availability of food. Protistan feeding may be driven by a virus' hydrophobicity but was independent of virus size or the presence of a lipid envelope.

Impact of the Heterogeneity in Free Chlorine, UV254, and Ozone Susceptibilities Among Coxsackievirus B5 on the Prediction of the Overall Inactivation Efficiency.

The disinfection susceptibilities of viruses vary even among variants, yet the inactivation efficiency of a certain virus genotype, species, or genus was determined based on the susceptibility of its laboratory strain. The objectives were to evaluate the variability in susceptibilities to free chlorine, UV254, and ozone among 13 variants of coxsackievirus B5 (CVB5) and develop the model allowing for predicting the overall inactivation of heterogeneous CVB5. Our results showed that the susceptibilities differed by up to 3.4-fold, 1.3-fold, and 1.8-fold in free chlorine, UV254, and ozone, respectively. CVB5 in genogroup B exhibited significantly lower susceptibility to free chlorine and ozone than genogroup A, where the laboratory strain, Faulkner, belongs. The capsid protein in genogroup B contained a lower number of sulfur-containing amino acids, readily reactive to oxidants. We reformulated the Chick-Watson model by incorporating the probability distributions of inactivation rate constants to capture the heterogeneity. This expanded Chick-Watson model indicated that up to 4.2-fold larger free chlorine CT is required to achieve 6-log inactivation of CVB5 than the prediction by the Faulkner strain. Therefore, it is recommended to incorporate the variation in disinfection susceptibilities for predicting the overall inactivation of a certain type of viruses.

Historical Discoveries on Viruses in the Environment and Their Impact on Public Health.

BACKGROUND: Transmission of many viruses occurs by direct transmission during a close contact between two hosts, or by an indirect transmission through the environment. Several and often interconnected factors, both abiotic and biotic, determine the persistence of these viruses released in the environment, which can last from a few seconds to several years. Moreover, viruses in the environment are able to travel short to very long distances, especially in the air or in water. SUMMARY: Although well described now, the role of these environments as intermediaries or as reservoirs in virus transmission has been extensively studied and debated in the last century. The majority of these discoveries, such as the pioneer work on bacteria transmission, the progressive discoveries of viruses, as well as the persistence of the influenza virus in the air varying along with droplet sizes, or the role of water in the transmission of poliovirus, have contributed to the improvement of public health. Recent outbreaks of human coronavirus, influenza virus, and Ebola virus have also demonstrated the contemporaneity of these research studies and the need to study virus persistence in the environment. Key Messages: In this review, we discuss historical discoveries that contributed to describe biotic and abiotic factors determining viral persistence in the environment.

Viral inactivation using microwave-enhanced membrane filtration.

Pathogenic viruses (e.g., Enteroviruses, Noroviruses, Rotaviruses, and Adenovirus) present in wastewater, even at low concentrations, can cause serious waterborne diseases. Improving water treatment to enhance viral removal is of paramount significance, especially given the COVID-19 pandemic. This study incorporated microwave-enabled catalysis into membrane filtration and evaluated viral removal using a model bacteriophage (MS2) as a surrogate. Microwave irradiation effectively penetrated the PTFE membrane module and enabled surface oxidation reactions on the membrane-coated catalysts (i.e., BiFeO3), which thus elicited strong germicidal effects via local heating and radical formation as reported previously. A log removal of 2.6 was achieved for MS2 within a contact time as low as 20 s using 125-W microwave irradiation with the initial MS2 concentration of 105 PFU∙mL-1. By contrast, almost no inactivation could be achieved without microwave irradiation. COMSOL simulation indicates that the catalyst surface could be heated up to 305 oC with 125-W microwave irradiation for 20 s and also analyzed microwave penetration into catalyst or water film layers. This research provides new insights to the antiviral mechanisms of this microwave-enabled catalytic membrane filtration.

Assessing the transmission risk of red sea bream iridovirus (RSIV) in environmental water: insights from fish farms and experimental settings.

Aquatic animal viruses are considered to be transmitted via environmental water between fish farms. This study aimed to understand the actual transmission risk of red sea bream iridovirus (RSIV) through environmental water among fish farms. An environmental DNA (eDNA) method using iron-based flocculation coupled with large-pore filtration was used to monitor RSIV DNA copies in seawater from fish farms and from an experimental infection model. RSIV dispersion in seawater from a net pen where the disease outbreak occurred was visualized by the inverse distance weighting method using multiple-sampling data sets from a fish farm. The analysis demonstrated that the center of the net pen had a high viral load, and RSIV seemed to be quickly diluted by the tidal current. To evaluate the transmission risk of RSIV in environmental water, the red sea bream Pagrus major (approximately 10 g) was exposed to RSIV-contained seawater (103, 104, 105, 106, and 107 copies/L) for 3 days, which mimicked field exposure. A probit analysis of the challenge test indicated that the inferred infection rates of seawater containing 105.9 copies/L and 103.1 copies/L of RSIV were 50% and 0.0001%, respectively. In the surveillance for 3 years at 10 fixed points (n = 306), there were only seven samples in which the viral load exceeded 104 copies/L in seawater. These results suggest that the transmission of RSIV among fish farms via seawater is highly associated with the distance between the net pens, and the environmental water is not always an infection source for the transmission of RSIV between fish farms. IMPORTANCE Our surveillance of viral loads for red sea bream iridovirus (RSIV) by monitoring environmental DNA in fish farms suggested that the viral loads in the seawater were low, except for the net pens where RSIV outbreaks occurred. Furthermore, our experimental infection model indicated that the infection risk of RSIV-contained seawater with less than 103 copies/L was extremely low. The limited risk of environmental water for transmission of RSIV gives an insight that RSIV could be partly transmitted between fish farms due to the movement of equipment and/or humans from the fish farm where the disease outbreaks. Since our data suggest that seawater can function as a potential wall to reduce the transmission of RSIV, biosecurity management, such as disinfection of equipment associated with fish farms could be effective, even in the semi-open system aquaculture that the environmental water can be freely transferred, to reduce the risk of RSIV outbreaks.

Waterborne virus transport and the associated risks in a large lake.

Waterborne enteric viruses in lakes, especially at recreational water sites, may have a negative impact on human health. However, their fate and transport in lakes are poorly understood. In this study, we propose a coupled water quality and quantitative microbial risk assessment (QMRA) model to study the transport, fate and infection risk of four common waterborne viruses (adenovirus, enterovirus, norovirus and rotavirus), using Lake Geneva as a study site. The measured virus load in raw sewage entering the lake was used as the source term in the water quality simulations for a hypothetical scenario of discharging raw wastewater at the lake surface. After discharge into the lake, virus inactivation was modeled as a function of water temperature and solar irradiance that varied both spatially and temporally during transport throughout the lake. Finally, the probability of infection, while swimming at a popular beach, was quantified and compared among the four viruses. Norovirus was found to be the most abundant virus that causes an infection probability that is at least 10 times greater than the other viruses studied. Furthermore, environmental inactivation was found to be an essential determinant in the infection risks posed by viruses to recreational water users. We determined that infection risks by enterovirus and rotavirus could be up to 1000 times lower when virus inactivation by environmental stressors was accounted for compared with the scenarios considering hydrodynamic transport only. Finally, the model highlighted the role of the wind field in conveying the contamination plume and hence in determining infection probability. Our simulations revealed that for beaches located west of the sewage discharge, the infection probability under eastward wind was 43% lower than that under westward wind conditions. This study highlights the potential of combining water quality simulation and virus-specific risk assessment for a safe water resources usage and management.

Inactivation efficacy and mechanism of pulsed corona discharge plasma on virus in water.

The presence of viruses in water is a major risk for human and animal health due to their high resistance to disinfection. Pulsed corona discharge plasma (PCDP) efficiently inactivates bacteria by causing damage to biological macromolecules, but its effect on waterborne virus has not been reported. This study evaluated the inactivation efficacy of PCDP to viruses using spring viremia of carp virus (SVCV) as a model. The results showed that 4-log10 reduction of SVCV infectivity in cells was reached after 120 s treatment, and there was no significant difference in survival of fish infected with SVCV inactivated by PCDP for 240 s or more longer compared to the control fish without virus challenge, thus confirming the feasibility of PCDP to waterborne virus inactivation. Moreover, the high input energy density caused by voltage significantly improved the inactivation efficiency. The further research indicated that reactive species (RS) generated by pulsed corona discharge firstly reacted with phosphoprotein (P) and polymerase complex proteins (L) through penetration into the SVCV virions, and then caused the loss of viral infectivity by damage to genome and other structural proteins. This study has significant implications for waterborne virus removal and development of novel disinfection technologies.

Recent advances in biosensors for detecting viruses in water and wastewater.

As there is a considerable number of virus particles in wastewater which cause numerous infectious diseases, it is necessary to eliminate viruses from domestic wastewater before it is released in the environment. In addition, on-site detection of viruses in wastewater can provide information on possible virus exposures in the community of a given wastewater catchment. For this purpose, the pre-detection of different strains of viruses in wastewaters is an essential environmental step. Epidemiological studies illustrate that viruses are the most challenging pathogens to be detected in water samples because of their nano sizes, discrete distribution, and low infective doses. Over the past decades, several methods have been applied for the detection of waterborne viruses which include polymerase chain reaction-based methods (PCR), enzyme-linked immunosorbent assay (ELISA), and nucleic acid sequence-based amplification (NASBA). Although they have shown acceptable performance in virus measurements, their drawbacks such as complicated and time-consuming procedures, low sensitivity, and high analytical cost call for alternatives. Although biosensors are still in an early stage for practical applications, they have shown great potential to become an alternative means for virus detection in water and wastewater. This comprehensive review addresses the different types of viruses found in water and the recent development of biosensors for detecting waterborne viruses.

An integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR) method to simultaneously quantify the infectious concentrations of eight environmentally relevant enterovirus serotypes.

Enterovirus (EV) infectivity is typically measured as a bulk parameter, yet EV serotypes vary in their susceptibility to natural and engineered stressors. Here we developed an integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR) method to simultaneously and specifically quantify the infectious concentrations of eight EV serotypes commonly encountered in sewage (coxsackieviruses A9, B1, B2, B3, B4 and B5, and echoviruses 25 and 30). The method uses two cell lines for virus replication and serotype-specific qPCR primers for quantification. Primers were designed to target multiple environmental strains of a given serotype and displayed high specificity. The ICC-RTqPCR method exhibited a linear calibration range between 50 and 1000 (echoviruses) or 5000 (coxsackieviruses) infectious units per mL. Over this range, measurements were not influenced by the presence of non-target serotypes, and calibration slopes were reproducible for different virus batches and cell ages. The ICC-RTqPCR method was able to accurately quantify the infectious concentration of a virus after inactivation by heat, and the concentration of a virus within a wastewater matrix. This method will be valuable to assess the differing fates of EV serotypes in natural or engineered systems, and to portray the associated changes in EV population composition.