Degree Type

Dissertation

Date of Award

2011

Degree Name

Doctor of Philosophy

Department

Veterinary Microbiology and Preventive Medicine

First Advisor

Bradley J. Blitvich

Abstract

Infectious diseases are significant impediments to the growth and sustainability of commercial shrimp aquaculture. Endemic diseases such as that caused by the obligate intracellular parasite the Necrotizing Hepatopancreatitis Bacterium (NHPB), cause significant losses to shrimp producers thoughout the Americas. New and emerging viral diseases, such as that caused by Infectious myonecrosis virus (IMNV), have caused tremendous economic losses in farmed Pacific white shrimp (Litopenaeus vannamei), since it emerged in Brazil and subsequently spread across the globe to Indonesia. No methods exist to propagate and model these infections in vitro), as no cell culture lines are available for shrimp. Therefore, we describe a process of maintaining and propagating NHPB in vivo), for the use in conjunction with challenge models to simulate infections in ponds, as well as a challenge model to repeatably induce IMNV infections. Additionally, no existing antiviral treatments or interventions, outside of pathogen exclusion, exist to mitigate disease in commercial shrimp operations. Novel RNA interference (RNAi) strategies that employ exogenous double-stranded RNA (dsRNA) are a promising means to approach disease control. Here we describe an iterative process of panning the viral genome of IMNV to discover RNAi trigger sequences that initiate a robust and long-lasting protective response against viral disease inL. vannamei. Using this process, a single, low dose of a short dsRNA fragment targeting the 5' end of the genome was discovered, which protected 100% of animals from disease and mortality caused by IMNV. Furthermore, animals that were treated with this dsRNA survived an initial infection and were resistant to subsequent infections 50 days later with a hundred-fold greater dose of IMNV. dsRNA also protected 50% of animals treated 2 days after challenge with a lethal dose of IMNV. Interestingly, non-sequence specific or heterologous dsRNA did not provide any degree of protection to animals as had been described for other shrimp viruses. Our data indicate that the targeted region for dsRNA is a crucial factor in maximizing the degree of protection and lowering the dose required to induce a protective effect against IMNV infection in shrimp. This data enables further development of RNAi based antivirals in farm-raised shrimp.

DOI

https://doi.org/10.31274/etd-180810-3130

Copyright Owner

John Dustin Loy

Language

en

Date Available

2012-04-28

File Format

application/pdf

File Size

121 pages

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