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 McIlvaines 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.
| Identifer | oai:union.ndltd.org:LSU/oai:etd.lsu.edu:etd-04062016-083339 |
| Date | 29 April 2016 |
| Creators | Maite, Morgan |
| Contributors | Janes, Marlene, Losso, Jack, Johnson, Crystal, LeBlanc, Brian |
| Publisher | LSU |
| Source Sets | Louisiana State University |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.lsu.edu/docs/available/etd-04062016-083339/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached herein a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to LSU or its agents the non-exclusive license to archive and make accessible, under the conditions specified below and in appropriate University policies, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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