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Applied and Environmental Microbiology, December 2007, p. 7934-7946, Vol. 73, No. 24
0099-2240/07/$08.00+0     doi:10.1128/AEM.01115-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Signature-Tagged Mutagenesis of Edwardsiella ictaluri Identifies Virulence-Related Genes, Including a Salmonella Pathogenicity Island 2 Class of Type III Secretion Systems{triangledown}

Ronald L. Thune,1,2* Denise H. Fernandez,1,2 Jennifer L. Benoit,1 Maria Kelly-Smith,1 Matthew L. Rogge,2 Natha J. Booth,1 Christie A. Landry,1,2 and Rachel A. Bologna1

Department of Veterinary Science, Louisiana State University Agricultural Center, Louisiana State University, Baton Rouge, Louisiana 70803,1 Department of Pathobiological Sciences, School of Veterinary Medicine, Skip Bertman Drive and River Road, Louisiana State University, Baton Rouge, Louisiana 708032

Received 18 May 2007/ Accepted 15 October 2007

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 minitransposon insertions in miscellaneous genes in the chromosome, 10 were in genes that matched to hypothetical proteins, and 13 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 lipopolysaccharide biosynthesis, three in genes involved in type III secretion systems (TTSS), and two in genes involved in urease activity. With the use of a 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 containing 33 genes of a TTSS with similarity to the Salmonella pathogenicity island 2 class of TTSS. The characterization of a TTSS apparatus mutant indicated that it retained its ability to invade catfish cell lines and macrophages but was defective in intracellular replication. The mutant also invaded catfish tissues in numbers equal to those of invading wild-type E. ictaluri bacteria but replicated poorly and was slowly cleared from the tissues, while the wild type increased in number.

* Corresponding author. Mailing address: LSU School of Veterinary Medicine, Skip Bertman Drive and River Rd., Baton Rouge, LA 70803. Phone: (225) 578-9680. Fax: (225) 578-9701. E-mail: thune{at}vetmed.lsu.edu

{triangledown} Published ahead of print on 26 October 2007.

Applied and Environmental Microbiology, December 2007, p. 7934-7946, Vol. 73, No. 24
0099-2240/07/$08.00+0     doi:10.1128/AEM.01115-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Booth, N. J., Beekman, J. B., Thune, R. L. (2009). Edwardsiella ictaluri Encodes an Acid-Activated Urease That Is Required for Intracellular Replication in Channel Catfish (Ictalurus punctatus) Macrophages. Appl. Environ. Microbiol. 75: 6712-6720 [Abstract] [Full Text]  
  • Karsi, A., Gulsoy, N., Corb, E., Dumpala, P. R., Lawrence, M. L. (2009). High-Throughput Bioluminescence-Based Mutant Screening Strategy for Identification of Bacterial Virulence Genes. Appl. Environ. Microbiol. 75: 2166-2175 [Abstract] [Full Text]  


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