Genetic Analysis of Bacterial Catfish Pathogens

 

Justification

   Currently, disease control is the main problem catfish growers must face, accounting for more than 45 percent of total annual fish losses. Because of production costs, nearly all commercially produced catfish are grown in earthen ponds. These closed systems usually support high fish densities and constitute a favorable environment for rapid spread of infectious bacteria and acute outbreaks of disease.

   The most important infectious diseases for catfish are Enteric Septicemia (ESC) and Columnaris disease. The causal agents for these diseases are Edwardsiella ictaluri and Flavobacterium columnare, respectively. These two bacteria are responsible for the US catfish industry losing more than 100 million dollars annually. Despite their significance, little is known about their virulence mechanisms, epidemiology, and ecology. In general, and due to the few cost-effective drugs available on the market, prevention is the best option for disease control in aquaculture. Besides improving husbandry practices, like reducing stress factors, a better understanding of these pathogens is needed in order to reduce economic loses due to disease impact.

Objectives

1. Characterize E. ictaluri and F. columnare isolates using phenotypic and genotypic markers
2. Develop new and improved methods for rapid detection of ESC and columnaris diseases
3. Identify and characterize virulence genes

                                                       Catfish with typical signs of Enteric Septicemia

Research

Intraspecies characterization

   Currently, several methods have been implemented and applied to study the intraspecies variation in E. ictaluri and F. columnare. These methods include phenotypic markers such as fatty acid methyl ester analysis, lipopolysaccharide profiling, biochemically-based commercial techniques and genetic markers including amplified fragment length polymorphisms, ribosomal gene sequencing and single-strand conformation polymorphisms. All data is analyzed and compiled into one single computerized database. This allows the calculation of similarity indexes between isolates using multiple typing methods simultaneously.

PCR-based detection methods

   A new specific and sensitive polymerase chain reaction method has been developed in recent years. This method can detect as few as 100 F. columnare cells per gram of sample in less than 8 hours. We are currently working on developing a combined assay that will allow simultaneous detection of ESC and columnaris infected fish.

Virulence genes

   Several strategies are being used in order to identify and isolate genes that are involved in ESC and columnaris pathogenesis. To date, more than 3,000 clones from a genomic library of a highly virulent F. columnare strain have been sequenced and analyzed. A few putative virulence genes have been identified and their expression under different growing conditions in vitro and in vivo is under investigation.

                                                        Catfish fingerlings with Columnaris disease

Anticipated Impacts

   The first benefit of this project will be an increment in the current knowledge we have about these two fish pathogens. Once different strains/clones have been defined, we will test them for virulence. Avirulent or low-virulent strains will be genetically compared versus the high virulent isolates in order to find virulence markers that will lead to virulence factors identification. Molecular-based detection methods will speed up the identification process of columnaris and ESC diagnosis.

   We also expect to isolate and characterize genes that are involved in pathogenicity. This will help us to better understand the molecular mechanisms of disease and to understand which gene(s) might be critical for infection. This information could be used for developing new vaccines.

Principal Investigators

Cova Arias

Assistant Professor

Auburn University, Department of Fisheries and Allied Aquacultures

ariascr@auburn.edu

Oscar Olivares-Fuster

Postdoctoral Fellow

Auburn University, Department of Fisheries and Allied Aquacultures

olivaoj@auburn.edu

Craig Shoemaker

Affiliate- Assistant Professor

Aquatic Animal Health Laboratory, USDA

cshoemaker@msa-stoneville.ars.usda.gov

Affiliated Institutions

Aquatic Animal Health Research Laboratory, ARS-USDA

Auburn, AL