1a.Objectives (from AD-416):
Development and validation of sensitive, rapid and practical methods to detect and genotype human noroviruses in relevent sample matrices (food, water,environmental, clinical).
1b.Approach (from AD-416):
Use DNA microarray platform in conjunction with rapid colorimetric procedure (ampliPHOX system) for genotyping human noroviruses.
The objective of this project is to develop a rapid and cost-effective low density DNA microarray for viral detection to identify genetic differences between human noroviruses (HuNV) in support of virus detection objective of a large NIFA-AFRI-funded project at NCSU. A large set of oligonucleotides has been designed for the many HuNV genogroups and subgroups and will be tested with a reference collection of HuNV RNA provided by CDC. The specific colorimetric detection of positive signals on the microarray is determined with a novel commercial system. To confirm the presence of HuNV in the samples, the amplicons hybridized to the DNA microarray will be sequenced, and their sequence similarity to reference HuNV strains will be determined as previously described. Low density arrays will be available for use by other labs who are part of the consortium project. An additional objective for this project is a preliminary study of watersheds for HuNV. HuNV in the environment of food production is of concern to regulatory agencies, but has not been evaluated well due to the difficulties in detecting HuNV. We have developed a receptor-binding capture method to concentrate HuNV from large volumes of water and detection by reverse transcriptase PCR that will be used to analyze water from watersheds in the vicinity of leafy greens production. This work relates to objectives of the NP108 to develop “technologies … for the entire food chain which allows the most effective and rapid detection and characterization… and the highest level of detection/characterization capability”…and with “Coordination among various agencies.”