This study utilizes old and new Norovirus (NoV) human challenge data to model the dose-response relationship for human NoV infection. The combined data set is used to update estimates from a previously published beta-Poisson dose-response model that includes parameters for virus aggregation and for a beta-distribution that describes variable susceptibility among hosts. The quality of the beta-Poisson model is examined and a simpler model is proposed. The new model (fractional Poisson) characterizes hosts as either perfectly susceptible or perfectly immune, requiring a single parameter (the fraction of perfectly susceptible hosts) in place of the two-parameter beta-distribution. A second parameter is included to account for virus aggregation in the same fashion as it is added to the beta-Poisson model. Infection probability is simply the product of the probability of nonzero exposure (at least one virus or aggregate is ingested) and the fraction of susceptible hosts. The model is computationally simple and appears to be well suited to the data from the NoV human challenge studies. The model's deviance is similar to that of the beta-Poisson, but with one parameter, rather than two. As a result, the Akaike information criterion favors the fractional Poisson over the beta-Poisson model. At low, environmentally relevant exposure levels (<100), estimation error is small for the fractional Poisson model; however, caution is advised because no subjects were challenged at such a low dose. New low-dose data would be of great value to further clarify the NoV dose-response relationship and to support improved risk assessment for environmentally relevant exposures.

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Crassostrea ariakensis oysters are under review for introduction into the Chesapeake Bay. However, the human health implications of the introduction have not been fully addressed. This study evaluated rates of bioaccumulation, retention, and depuration of viruses by Crassostrea virginica and C. ariakensis when the two oyster species were maintained in separate tanks containing synthetic seawater of various salinities (8, 12, or 20 ppt). Oyster bioaccumulation tanks were seeded with 10 3 PFU/ml of hepatitis A virus (HAV), poliovirus, male-specific bacteriophage (MS2), and murine norovirus 1 (MNV-1) and 10 3 PCR units/ml of human norovirus (NoV). After 24 h, depuration commenced as oysters (n ‫؍‬ 255) were placed in pathogen-free seawater under continuous filtration. Oysters (n ‫؍‬ 6) were sampled weekly for 1 month from each tank. Viral RNA was recovered using a modified proteinase K, guanidine, and glassmilk method and analyzed by quantitative reverse transcription-PCR. The odds of C. ariakensis oysters harboring NoV, MNV-1, or HAV were statistically greater than the odds of C. virginica oysters harboring the same viruses (MNV-1 odds ratio [OR], 4.5; P ‫؍‬ 0.01; NoV OR, 8.4; P < 0.001; HAV OR, 11.4; P < 0.001). Unlike C. virginica, C. ariakensis bioaccumulated and retained NoV, MNV-1, and HAV for 1 month at all salinities. Additionally, the odds of an oyster testing positive for NoV was 25.5 times greater (P < 0.001) when the oyster also tested positive for MNV-1. This research helps assess the threat of C. ariakensis as a vehicle for viral pathogens due to the consumption of raw oysters and validates the role for MNV-1 as a surrogate for NoV.

The introduction of nonnative oysters (i.e., Crassostrea ariakensis) into the Chesapeake Bay has been proposed as necessary for the restoration of the oyster industry; however, nothing is known about the public health risks related to contamination of these oysters with human pathogens. Commercial market-size C. ariakensis triploids were maintained in large marine tanks with water of low (8-ppt), medium (12-ppt), and high (20-ppt) salinities spiked with 1.0 ؋ 10 5 transmissive stages of the following human pathogens: Cryptosporidium parvum oocysts, Giardia lamblia cysts, and microsporidian spores (i.e., Encephalitozoon intestinalis, Encephalitozoon hellem, and Enterocytozoon bieneusi). Viable oocysts and spores were still detected in oysters on day 33 post-water inoculation (pwi), and cysts were detected on day 14 pwi. The recovery, bioaccumulation, depuration, and inactivation rates of human waterborne pathogens by C. ariakensis triploids were driven by salinity and were optimal in medium-and high-salinity water. The concentration of human pathogens from ambient water by C. ariakensis and the retention of these pathogens without (or with minimal) inactivation and a very low depuration rate provide evidence that these oysters may present a public health threat upon entering the human food chain, if harvested from polluted water. This conclusion is reinforced by the concentration of waterborne pathogens used in the present study, which was representative of levels of infectious agents in surface waters, including the Chesapeake Bay. Aquacultures of nonnative oysters in the Chesapeake Bay will provide excellent ecological services in regard to efficient cleaning of human-infectious agents from the estuarine waters.

Understanding pathogen risks is a critically important consideration in the design of water treatment, particularly for potable reuse projects. As an extension to our published microbial risk assessment methodology to estimate infection risks associated with Direct Potable Reuse (DPR) treatment train unit process combinations, herein, we (1) provide an updated compilation of pathogen density data in raw wastewater and dose-response models; (2) conduct a series of sensitivity analyses to consider potential risk implications using updated data; (3) evaluate the risks associated with log credit allocations in the United States; and (4) identify reference pathogen reductions needed to consistently meet currently applied benchmark risk levels. Sensitivity analyses illustrated changes in cumulative annual risks estimates, the significance of which depends on the pathogen group driving the risk for a given treatment train. For example, updates to norovirus (NoV) raw wastewater values and use of a NoV dose-response approach, capturing the full range of uncertainty, increased risks associated with one of the treatment trains evaluated, but not the other. Additionally, compared to traditional log-credit allocation approaches, our results indicate that the risk methodology provides more nuanced information about how consistently public health benchmarks are achieved. Our results indicate that viruses need to be reduced by 14 logs or more to consistently achieve currently applied benchmark levels of protection associated with DPR. The refined methodology, updated model inputs, and log credit allocation comparisons will be useful to regulators considering DPR projects and design engineers as they consider which unit treatment processes should be employed for particular projects.

Thermophilic‐anaerobic digestion in a single‐stage, mixed, continuous‐flow reactor is not approved in the United States as a process capable of producing Class A biosolids for land application. This study was designed to evaluate the inactivation of pathogens and indicator organisms in such a reactor followed by batch treatment in a smaller reactor. The combined process was evaluated at 53°C with sludges from three different sources and at 51 and 55°C with sludge from one of the sources. Feed sludge to the continuous‐flow reactor was spiked with the pathogen surrogates Ascaris suum and vaccine‐strain poliovirus. Feed and effluent were analyzed for these organisms and for indigenous Salmonella spp., fecal coliforms, Clostridium perfringens spores, and somatic and male‐specific coliphages. No viable Ascaris eggs were observed in the effluent from the continuous reactor at 53 or 55°C, with greater than 2‐log removals across the digester in all cases. Approximately 2‐log removal was observed at 51°C, but all samples of effluent biosolids contained at least one viable Ascaris egg at 51°C. No viable poliovirus was found in the digester effluent at any of the operating conditions, and viable Salmonella spp. were measured in the digester effluent in only one sample throughout the study. The ability of the continuous reactor to remove fecal coliforms to below the Class A monitoring limit depended on the concentration in the feed sludge. There was no significant removal of Clostridium perfringens across the continuous reactor under any condition, and there also was limited removal of somatic coliphages. The removal of male‐specific coliphages across the continuous reactor appeared to be related to temperature. Overall, at least one of the Class A pathogen criteria or the fecal coliform limit was exceeded in at least one sample in the continuous‐reactor effluent at each temperature. Over the range of temperatures evaluated, the maximum time required to meet the Class A criteria by batch treatment of the continuous‐reactor effluent was 1 hour for Ascaris suum and Salmonella spp. and 2 hours for fecal coliforms.

To determine the burden of norovirus infections in children stools from a longitudinal community cohort were evaluated using reverse transcription polymerase chain reaction. Norovirus was detected in 21.3% of diarrheal and 8.0% of nondiarrheal stools (P < 0.01). Norovirus diarrhea was highly associated with age and the odds ratio for norovirus diarrhea fell by 2.8% per month (OR = 0.97, 95% CI: 0.95-0.99). Norovirus seems to be an important etiology of community acquired diarrhea in this study population.