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United States Department of Agriculture

Agricultural Research Service

Research Project: Integrated Approaches for Improving Aquatic Animal Health in Cool and Cold Water Aquaculture

Location: Cool and Cold Water Aquaculture Research

2011 Annual Report


1a.Objectives (from AD-416)
Endemic and emerging diseases limit U.S. aquatic farm-animal production. This project focuses on improving fish health by selectively breeding rainbow trout for increased disease resistance and utilizing divergent fish phenotypes to understand host resistance and pathogen virulence mechanisms under laboratory and farm conditions.

1: Conduct selective breeding and determine, under field and laboratory conditions, whether rainbow trout bred for bacterial cold water disease resistance exhibit superior performance traits. • 1.a. Evaluate and selectively breed NCCCWA rainbow trout for increased disease resistance and quantify improvement due to selection. • 1.b. Compare improved and reference lines of NCCCWA rainbow trout under farm conditions.

2: Characterize virulence determinants involved in the emergence of vaccine-resistant Yersinia ruckeri strains and develop novel therapeutics to treat or prevent disease. • 2.a. Elucidate the genetic basis of the biotype 2 phenotype. • 2.b. Evaluate the role of flagellar secretion in the function of the traditional ERM immersion vaccine and vaccine failure. • 2.c. Develop novel phage therapeutics for control of ERM and BCWD.

3: Identify genes and non-genetic factors in rainbow trout critical to innate and acquired immunity. • 3.a. Determine if heritable differences in spleen structure and function account for the superior BCWD survival and clearance traits of resistant fish. • 3.b. Determine whether orally-delivered immunostimulants modulate the gut immune system and whether variation exists between resistant and susceptible lines of rainbow trout. • 3.c. Initiate development of a rainbow trout immune-pathway database that is integrated with pathogen genomic information.


1b.Approach (from AD-416)
Our approach incorporates a comprehensive and multidisciplinary strategy that combines selective breeding, functional genomics of pathogenic bacteria, and immunological studies. In the first objective, we utilize conventional family-based selective breeding to increase bacterial cold water disease resistance and evaluate select and control fish performance in on-farm trials. In the second objective, we characterize indicators of virulence in emerging strains of vaccine-resistant Y. ruckeri using molecular and genomic approaches and develop novel therapeutics to treat or prevent bacterial disease. In the third objective, we identify genes and non-genetic factors in trout that are critical to expression of innate and acquired immune responses using molecular and immunological techniques. We will identify and measure mucosal immune system function and dietary immunomodulation of local immunity. The overall impact of this research is improved animal well-being, reduced antibiotic use and increased production efficiency.


3.Progress Report
Efforts to selectively breed disease resistance are conducted under NP106 Components 1 and 4. Under Objective 1, we completed the spawning and phenotyping of our 2011 year-class of rainbow trout which has been bred for increased resistance to bacterial cold water disease. The resistant line, designated ARS-Fp-R, now represents 3 generations of selection for increased resistance. We have also developed two control lines: a selection control line, designated ARS-Fp-C; and a reference susceptible line, designated ARS-Fp-S. This year we created over 159 viable crosses and experimentally evaluated the disease resistance of each cross. The results from the disease challenge studies verified our selective breeding efforts and quantified the relative disease resistance of each genetic line.

A major focus of our current work is to determine if the ARS-Fp-R line exhibits significant resistance when exposed to natural challenge under farm conditions. This year, we completed field trials initiated in 2010 and also initiated another set of field trials in 2011. In the 2010 experiments, eggs were shipped to three locations for evaluation by cooperators Clear Springs Foods Inc (agreement #58-1930-0-038) and Utah State Fish and Wildlife Service (agreement # 58-1930-0-042N). Egg hatch rate was above 80% at each location. In the Utah trials, survival of ARS-Fp-R line was compared to age-matched, BCWD susceptible rainbow trout (Gunnison River/Harrison Lake triploids, GHTP). Survival was measured over an 80 day period post-first feeding when bacterial cold water disease is most severe. The GHTP line exhibited an outbreak of bacterial cold water disease at two Utah trial sites. At these sites, survival of the ARS-Fp-R line was significantly greater than the GHTP line and no bacterial cold water disease was observed in the resistant-line fish. In the Idaho field trial, no bacterial cold water disease was diagnosed in either ARS-Fp-R or control line ARS-Fp-S fish. The overall survival and growth of the ARS-Fp-R line was positive and warranted additional field evaluation. Progress towards Objective 3 (NP106 Component.
4)involves characterizing the intestinal immune response to bacterial challenge which is poorly understood in rainbow trout. Two challenge model systems, injection and immersion of bacteria, were used to examine whether expression of immune genes changed in the lower intestine following exposure to pathogenic bacteria. Several immune genes representing both innate and acquired responses showed increases in abundance following both types of challenge paradigms. Our results demonstrate that the lower intestine responds following bacterial challenge with Y. ruckeri or F. psychrophilum. These findings will aid in the development of orally-delivered vaccines.


4.Accomplishments
1. Novel diagnostic assays differentiate virulent strains of the fish pathogen, Yersinia ruckeri. Yersinia ruckeri biotype 2 strains pose an emerging threat to rainbow trout aquaculture and improved diagnostic assays are needed. ARS researchers at the National Center for Cool and Cold Water Aquaculture (Kearneysville, WV) have developed assays for the rapid and precise identification of the three specific lineages of BT2 Yersinia ruckeri that are currently circulating in the US and Europe. The assays are easy to perform and interpret and require equipment common to diagnostic laboratories. They will be used to identify infected fish populations, and will facilitate the application of specific vaccines or other management practices aimed at controlling these newly emerging strains of bacteria in aquaculture.


Review Publications
Welch, T.J. 2011. Rapid genotyping assays for the identification and differentiation of Yersinia ruckeri biotype 2 strains. Letters in Applied Microbiology. DOI: 10.1111/j.1472-765X.2011.03114.x.

Welch, T.J., Verner-Jeffreys, D.W., Dalsgaard, I., Wiklund, T., Evenhuis, J., Garcia Cabrera, J.A., Hinshaw, J.M., Drennan, J.D., Lapatra, S.E. 2011. Independent emergence of biotype 2 Yersinia ruckeri in the United States and Europe. Applied and Environmental Microbiology. 2011 May;77(10):3493-9

Carrias, A., Welch, T.J., Waldbieser, G.C., Mead, D.A., Terhune, J.S., Liles, M.R. 2011. Comparative genomic analysis of bacteriophages specific to the channel catfish pathogen Edwardsiella ictaluri. Virology Journal. 8:6. DOI: 10.1186/1743-422X-8-6.

Wiens, G.D. 2011. Bacterial kidney disease (Renibacterium salmoninarum). Fish Diseases and Disorders. 3:338-374.

Wiens, G.D., Glenney, G.W. 2011. Origin and evolution of TNF and TNF receptor superfamilies. Developmental and Comparative Immunology. DOI: 10.1016/j.dci.2011.03.031.

Last Modified: 12/18/2014
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