Title page for ETD etd-12152003-111636


Type of Document Dissertation
Author Harbottle, Heather C.
URN etd-12152003-111636
Title Investigations into DNA Vaccination against Channel Catfish Virus
Degree Doctor of Philosophy (Ph.D.)
Department Veterinary Microbiology & Parasitology (Veterinary Medical Sciences)
Advisory Committee
Advisor Name Title
Ronald L. Thune Committee Chair
D. A. Rutherford Committee Member
J. F. Siebenaller Committee Member
John Hawke Committee Member
K. G. Kousoulas Committee Member
Keywords
  • fish disease
  • ictalurid herpesvirus 1
  • dna vaccines
Date of Defense 2003-11-24
Availability unrestricted
Abstract
The leading viral killer of commercially produced channel catfish is Channel Catfish Virus (CCV). Studies conducted to evaluate DNA vaccination against CCV compared encoded gene, dose, multiple DNA vaccines, and immune response to vaccination. Genes were selected (ORFs 1 and 3 [immediate early genes], ORFs 6, 19, and 46 [membrane genes], and ORF59 [putative major envelope glycoprotein gene], cloned into a plasmid, and expressed in mammalian and fish cell culture to detect predicted molecular weight proteins. Plasmid vaccines were injected into fish muscle in doses of 50 μg, 25 μg, 5 μg, or 1 μg and efficacy was evaluated upon challenge. Immune responses were measured by Mx gene expression (α/β interferon indicator), serum neutralization, specific antibody stimulation (ELISA), and DNA vaccine specific expression. Comparisons were made between published live attenuated CCVTK-, DNA vaccines (KN59 and KN6), and DNA vaccines tested in this study. In all experiments, no protection was observed. Multiple groups of vaccines delivered per fish were tested for efficacy and protection was not observed. The live attenuated CCVTK- and published CCV DNA vaccines (KN59 and KN6) were not protective. Mx gene expression in response to DNA vaccination showed positive expression profiles at all time points, but most often at day 3 in all experiments. In the multiple group vaccination, Mx gene expression was detected at a higher level overall and at day 35, indicating that multiple antigens induce a stronger innate response with longer duration. In all experiments, serum neutralizing titers were low in most treatments (< 10), but weakly reactive in the multiple group vaccination (3 groups with titers > 10) and the comparison vaccination (4 groups with titers > 10). ELISA of vaccinated fish sera detected low reactivity, but significant levels of reactivity were observed between sera from pORF46 and pORF3 and the negative control. In the comparison vaccination, DNA vaccine specific gene expression was detected in all groups and at most time points, indicating the DNA vaccines were transcriptionally functional. A protective vaccine against CCV is a goal to be striven for, because currently available vaccines may not be suitable for commercial use.
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