STTR Phase I: Feasibility of a two-phase test tube assay for rapid detection and enumeration of Vibrio vulnificus and Vibrio parahaemolyticus in oysters

这一小型企业技术转让项目的更广泛的影响/商业潜力将是开发一种检测分析方法，以减少弧菌相关感染的发生率。副溶血性弧菌和创伤弧菌的感染分别是通过海鲜传播的感染和死亡的主要原因，主要来源于生牡蛎。开发和实施快速、准确、廉价和易于使用的检测方法将扩大弧菌数据的收集，并加强国家贝类实验室(监测捕获区)、牡蛎加工者和收获后加工者(设施的质量控制)的预防措施。这些化验结果将以简单、现成的试剂盒提供给用户。这些试剂盒将使非专业人员能够在不中断物流的情况下轻松检测仍处于冷藏状态的牡蛎批次，并帮助美国牡蛎加工商和收获后加工商改进风险管理工具，并在限制弧菌水平的情况下进入出口市场。这些测试还将改善国家贝类实验室、FDA、国家海洋和大气管理局以及国家贝类卫生计划其他成员对捕捞区的环境风险评估。这些检测将在美国和其他大型牡蛎生产国销售。STTR第一阶段项目建议开发两相耐热试管培养基配方，用于检测和计数牡蛎中致病的副溶血性弧菌、总副溶血性弧菌和创伤弧菌。目标是证明：a)用纯培养物和牡蛎样本进行每项检测的灵敏度和特异度为95%；b)24小时检测时间；c)根据时间、步骤、设备、材料和行业反馈，易于使用现成试剂盒；以及d)低成本。主要的技术障碍是：1)增加试管分析的配方容量，因为试管介质配方中琼脂层体积的线性增加可能会影响弧菌的生物运动性，从而影响检测速度和准确性；以及2)将创伤弧菌试验中非热稳定的抗生素成分混合B替换为热稳定的替代品，如氨苄西林或林可霉素，以提高易用性。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The broader impact/commercial potential of this Small Business Technology Transfer (STTR) project will be the development of a detection assay to reduce the incidence of Vibrio-related infections. Infections from Vibrio parahaemolyticus and Vibrio vulnificus are the leading causes of seafood-borne infections and fatalities, respectively, and stem primarily from raw oysters. Development and implementation of rapid, accurate, inexpensive and easy-to-use testing methods would expand Vibrio data collection and enhance preventive measures by state shellfish labs (monitor harvest areas), and oyster processors and post-harvest processors (quality control at facilities). The assays will be available to users in simple, ready-to-prepare kits. These kits will make it easy for non-specialized personnel to test oyster lots still under refrigeration without disrupting logistics, and help U.S. oyster processors and post-harvest processors improve risk management tools and access export markets with restrictions on Vibrio levels. The tests also will improve environmental risk evaluations for harvest areas by state shellfish labs, FDA, National Oceanic and Atmospheric Administration, and other members of the National Shellfish Sanitation Program. The tests will be marketed in the U.S., and other large oyster producing nations.This STTR Phase I project proposes to develop two-phase, thermostable test tube media formulations for the detection and enumeration of pathogenic V. parahaemolyticus, total V. parahaemolyticus, and V. vulnificus in oysters. The goal is to demonstrate a) 95% sensitivity and specificity of each test with pure cultures and oyster samples; b) 24-hr detection time; c) ease-of-use of the ready-to-prepare kits, based on time, steps, equipment, materials and feedback from industry; and d) low cost. The main technical hurdles are: 1) Increasing formulation volumes for test tube assays since linear increases of agar layer volumes in test tube media formulations could impact Vibrio organism motility, and consequently detection speed and accuracy; and 2) replacing a non-heat stable antibiotic component, Polymixin B of the V. vulnificus test, with a heat stable alternative, like ampicillin or lincomycin, to improve ease-of-use.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.