Identifier

etd-04062016-083339

Degree

Doctor of Philosophy (PhD)

Department

School of Nutrition and Food Sciences

Document Type

Dissertation

Abstract

Norovirus (NoV) is the principal cause of viral gastroenteritis in the United States. It has been linked to filter-feeding molluscan shellfish, that bioaccumulate the virus from contaminated surrounding waters. The consumption of raw or undercooked contaminated oysters may result in acute gastroenteritis. We investigated the occurrence of NoV GI and GII and microbial indicators of fecal contamination in oysters and harvesting water from areas along the Louisiana Gulf Coast. We developed a filtration and concentration method for the detection of NoV from oyster harvesting waters. Lastly, this body of work compares commonly used molecular techniques (RT-PCR) and a commercial enzyme immunoassay for the detection of NoV. One oyster sample was positive for norovirus GII at 3.5 ± 0.2 log10 genomic equivalent copies/g digestive tissues, however the surrounding water tested negative for NoV. Zeolite granules were used for the filtration of norovirus-seeded waters. Beef Extract (10%) in McIlvaine’s buffer was the optimal elution buffer resulting in an average percent recovery of 41.76 + 0.07 (p<0.05). Artificial and environmental waters with 20ppt salt had an observed average percent recovery of 40.79 + 0.19 and 18.95 + 0.24, respectively which was significantly higher than 0, 5, 10, 15, and 25ppt (p<0.05). The observed percent recoveries for artificial and environmental waters were 44.03 + 0.20 and 34.36 + 0.02, respectively. The percent recovery for artificial and environmental water using TaqMan® Fast Virus 1-Step RT-qPCR was 38.85% + 0.27 and 19.77% + 0.07, respectively. In comparison, SuperScript® III Platinum One-Step qRT-PCR exhibited an average percent recovery of 11.12% + 0.183 and 15.55% + 0.225 for artificial and environmental waters. The EIA assay assay was not sensitive enough to detect NoV in the elution samples despite RT-qPCR methods quantifying the virus concentration between 104 and 105 genomic copies/ml. As such, it is not an effective method for the detection of NoV from environmental water matrices without RT-qPCR as a secondary validation method. This body of work provides an effective method to detect norovirus in oyster harvesting waters. Our results emphasize the need for regular monitoring of pathogenic viruses in oyster harvesting areas to reduce viral gastroenteritis incidences.

Date

2016

Document Availability at the Time of Submission

Release the entire work immediately for access worldwide.

Committee Chair

Janes, Marlene

Included in

Life Sciences Commons

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