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The threat of shellfish-borne disease from Vibrio parahaemolyticus (Vp) and Vibrio vulnificus (Vv) is a significant public health concern in the US. Although Vibrio cholerae (Vc) had not been an issue for US seafood for 150 years, an outbreak occurred in 2011 in Florida. Changes in pathogenic Vibrio populations in the Northeast US are likely consequences of changing environmental conditions and their detection is complicated because not all strains are virulent. The presence of a species does not imply disease risk. This is further complicated in the Northeast because concentrations of Vibrios are much lower in cooler water. Genetic virulence markers are also not fail-safe, as disease incidence in the Northwest has been attributed to Vp strains lacking accepted virulence markers. In the Northeast, outbreaks of Vp infections from consumption of Cape Cod Bay oysters in 2011 and Oyster Bay, NY oysters in 2012 caused public health scares and significant losses for regional industry. A simple method for screening pathogenic Vibrio species is essential for industry and regulators to shape locale-specific harvest management plans. Pathogenic Vibrios have a limited range of conditions related to seasonal climatic factors that are conducive to their prevalence. <P>Regional knowledge of these conditions can be used to reproduce more controlled processing and management strategies. Project PIs have demonstrated that transporting or "relaying" shellfish to elevated salinity conditions successfully reduces levels of pathogenic Vibrios. The proposed research will address some fundamental questions related to the ecology of Vibrios that are key to understanding disease transmission via shellfish consumption and why relaying is effective, and to address concerns about global warming effects on disease agents best suited to warm conditions, and the potential for their increased significance in what are presently colder Northeast environments. Work at UNH reported the first detected Vv and Vc in NH & ME, the effectiveness of shellfish relaying to reduce Vibrio levels, and the ecology of Vibrios in NH & ME. Recent research has expanded into studies on population structure, virulence potential and evolution. The proposed research is a logical next-step for bringing previous research into applications to help keep Northeast shellfish safe and harvestable. <P>The objectives are to: 1. Develop, refine and apply a tiered approach for cost-effective detection and enumeration of Vp, Vv and Vc and their virulence genes in shellfish and overlying waters in the Northeast U.S. 2. Develop an integrated approach for assessing environmental and biological factors that affect Vibrio-related shellfish safety. 3. Determine optimal conditions for reducing pathogenic Vibrio levels in shellfish under post-harvest treatment conditions. <P>Expected outputs include conducting experiments, mentoring and trainings, and presentations at conferences and workshops. Products include Vibrio incidence and genetic sequencing data, and detection and relaying methods. Information will be disseminated through existing networks for the extension agents, shellfish industry and public health agencies.
Non-Technical Summary:<br/>
Shellfish harvesting and public health are being threatened by the emerging incidence of vibirosis in consumers of shellfish from the Northeastern U.S. Where more northerly states with colder coastal ecosystems had rare disease occurrence in the past, recent increases in incidence has triggered costly harvest management and monitoring needs for industry and public health agencies alike. Continued development of improved Vibrio detection methods and post-harvest treatments for reducing Vibrio levels in shellfish will address growing regional concerns and reduce costs for all involved. New methods for understanding microbial dynamics in ecosystems will be used to address long-outstanding scientific questions about shellfish safety problems and for reducing concerns by shellfish consumers about the safety of what they eat.
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Approach:<br/>
We will develop a tiered approach for monitoring pathogenic species and their virulent strains in oysters and water from the Northeast. Sampling will occur in spring-fall to test detection methods and determine differences in Vibrio stress conditions: temperature, pH, salinity etc. We will use existing method for detecting total and virulent Vv,Vp and Vc using a most probable number enrichment and realtime polymerase chain reaction/qPCR detection of genetic markers. We will also evaluate the DuPont BAX Vibrio kit for detecting Vv,Vp and Vc using a qPCR-based multiplex assay for confirmation, and the use of chromogenic agars that discriminate species based upon enzymatic cleavage of sugar substrates. An overall monitoring approach with a tiered, hierarchical set of detection methods based mostly on water temperature effects on Vibrio incidence will be developed to minimize costs and promote easier methods for adoption by industry and public health agency use. Detection method testing on samples from a network of industry partners will be used to develop an optimal regional monitoring approach and to inform risk assessment thresholds, verification of management strategies, and serve as a predictive tool for public health risks. This study is unique in integrating Vv,Vp and Vc data with water quality and climatic data to predict risks. Post-harvest treatments currently under evaluation will be modified according to factors identified that are associated with reduced Vibrio levels. Relayed shellfish will be analyzed at day 0, 7, 10 and 14 for Vv,Vp and Vc from July-November. Metagenetic analysis of shellfish microbial communities will inform future microbial manipulations in concert with environment factors for improving post-harvest treatments. The results will be used to help identify bacteria that may correlate with disappearance (relay) of pathogenic Vibrios to optimize consistent relay tank systems for industry use. Detection methods will be analyzed to determine the relationship between total Vibrios, pathogenic species and virulent strains, and the environmental and climatic conditions conducive to presence/absence of the different Vibrio measures. Results will be analyzed for seasonal and geographical differences in Vibrio levels. Post-harvest treatment reductions will be determined and the most significant factors affecting Vibrio levels will be determined using stepwise multiple regression analysis. Study findings will be communicated to NH Sea Grant and UNH Cooperative Extension, Extension specialists involved in fisheries and food safety issues. Results will be presented as educational information in local and regional forums and shared with shellfish industry representatives, aquaculturists, public health agency personnel, scientists and Northeast extension specialists through a network established as part of a closely related, project. Findings will be taught to students at UNH and in local high school classrooms. Success will be evaluated by quantifying how many people adopt relaying after summertime harvest, the number of people informed about the detection methods, and the number that adopt their use.