Submitted to: Applied and Environmental Microbiology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 2/28/2007
Publication Date: 10/26/2007
Citation: Thune, R.L., Fernandez, D.H., Benoit, J.L., Kelly-Smith, M., Rogge, M.L., Booth, N.J., Landry, C.A., Balogna, R.A. 2007. Signature-Tagged Mutagenesis of Edwardsiella ictaluri Identifies Virulence Related Genes, 3 Including a Salmonella Pathogenicity Island-2 Class of Type III Secretion System. Applied and Environmental Microbiology. Interpretive Summary: Using a new technique called Signature Tagged Mutagenesis (STM), a technique used to identify randomly generated bacterial mutants that are no longer able to cause disease, we identified 50 mutants of the bacterium known to cause enteric septicemia that were unable to infect/survive in catfish. Nineteen mutants had disruptions in miscellaneous genes in the chromosome, ten had disruptions in genes that matched to hypothetical proteins, and thirteen had insertions in genes that were not identifiable. Eight disruptions were in genes encoding proteins associated with disease in other pathogens. The results presented here will be of great value in understanding the mechanisms by which this bacterium causes disease in channel catfish.
Technical Abstract: Edwardsiella ictaluri is the leading cause of mortality in channel catfish culture, but little is known about its pathogenesis. The use of signature-tagged mutagenesis in a waterborne infection model resulted in the identification of 50 mutants that were unable to infect/survive in catfish. Nineteen had mini-transposon insertions in miscellaneous genes in the chromosome, ten were in genes that matched to hypothetical proteins, and thirteen were in genes that had no significant matches in the NCBI databases. Eight insertions were in genes encoding proteins associated with virulence in other pathogens, including three in genes involved in LPS biosynthesis, three in genes involved in type III secretion systems (TTSS), and two in genes involved in urease activity. Using sequence from a lambda clone carrying several TTSS genes, Blastn analysis of the partially completed E. ictaluri genome identified a 26,135 bp pathogenicity island encoding 33 genes of a TTSS with similarity to the Salmonella pathogenicity island-2 class of TTSS. Characterization of a TTSS mutant indicated that it retained its ability to invade catfish cells, but was defective in intracellular replication. The mutant also invaded catfish tissues in equal numbers to the wild-type E. ictaluri, but replicated poorly and was slowly cleared from the tissues.