Renibacterium salmoninarum is the causative agent of bacterial kidney disease and a significant threat to healthy and sustainable production of salmonid fish worldwide. This pathogen is difficult to culture in vitro, genetic manipulation is challenging, and current therapies and preventative strategies are only marginally effective in preventing disease. The complete genome of R. salmoninarum ATCC 33209 was sequenced and shown to be a 3,155,250-bp circular chromosome that is predicted to contain 3,507 open-reading frames (ORFs). A total of 80 copies of three different insertion sequence elements are interspersed throughout the genome. Approximately 21% of the predicted ORFs have been inactivated via frameshifts, point mutations, insertion sequences, and putative deletions. The R. salmoninarum genome has extended regions of synteny to the Arthrobacter sp. strain FB24 and Arthrobacter aurescens TC1 genomes, but it is approximately 1.9 Mb smaller than both Arthrobacter genomes and has a lower G؉C content, suggesting that significant genome reduction has occurred since divergence from the last common ancestor. A limited set of putative virulence factors appear to have been acquired via horizontal transmission after divergence of the species; these factors include capsular polysaccharides, heme sequestration molecules, and the major secreted cell surface antigen p57 (also known as major soluble antigen). Examination of the genome revealed a number of ORFs homologous to antibiotic resistance genes, including genes encoding ␤-lactamases, efflux proteins, macrolide glycosyltransferases, and rRNA methyltransferases. The genome sequence provides new insights into R. salmoninarum evolution and may facilitate identification of chemotherapeutic targets and vaccine candidates that can be used for prevention and treatment of infections in cultured salmonids.

Renibacterium salmoninarum, the causative agent of bacterial kidney disease in salmonid fishes, is a Gram-positive diplococcobacillus belonging to the family Micrococcaceae. Analysis of the genome sequence of the bacterium demonstrated the presence of a sortase homolog (srtD), a gene specifying an enzyme found in Gram-positive bacteria and required for covalent anchoring of cell surface proteins. Interference of sortase activity is being examined as a target for therapeutic prevention of infection by several pathogenic Gram-positive bacterial species. In silico analysis identified 8 open reading frames containing sortase recognition motifs, suggesting these proteins are translocated to the bacterial cell wall. The sortase and potential sortase substrate genes are transcribed in R. salmoninarum, suggesting they encode functional proteins. Treatment of R. salmoninarum with phenyl vinyl sulfone (PVS) significantly reduced bacterial adherence to Chinook salmon fibronectin. In addition, the ability of the PVS-treated bacteria to adhere to Chinook salmon embryo cells in vitro was dramatically reduced compared to that of untreated bacteria. More importantly, PVS-treated bacteria were unable to invade and replicate within CHSE-214 cells (demonstrated by an intracellular growth assay and by light microscopy). When treated with PVS, R. salmoninarum was not cytopathic to CHSE-214 cells, whereas untreated bacteria produced cytopathology within a few days. These findings clearly show that PVS, a small molecule drug and a known sortase inhibitor, can interfere with the ability of R. salmoninarum to adhere and colonize fish cells, with a corresponding decrease in virulence.KEY WORDS: Renibacterium salmoninarum · Bacterial kidney disease · Anti-virulence chemotherapy · Adherence · Invasion · Host cell sortase inhibitor · Phenyl vinyl sulfone Resale or republication not permitted without written consent of the publisherDis Aquat Org 78: [115][116][117][118][119][120][121][122][123][124][125][126][127] 2007 1994, Kehoe 1994). MSCRAMs have specific C-terminal anchoring signals or cell wall sorting (CWS)-motifs that are recognized by a membrane transpeptidase called sortase. Sortase catalyzes covalent anchoring of these proteins to the peptidoglycan surface during cell wall synthesis (reviewed in Navarre & Schneewind 1999, Ton-That et al. 2004, Marraffini et al. 2006). Elimination of sortase activity and the consequent disruption of surface protein anchoring correlates with a dramatic decrease in pathogenicity of Gram-positive bacteria in animal infections (Mazmanian et al. 2000, Bolken et al. 2001, Bierne et al. 2002, Garandeau et al. 2002, Jonsson et al. 2002, Weiss et al. 2004. Sortase is also involved in the assembly of cell surface pili on Gram-positive bacteria, which often aid attachment to host cells (Ton-That & Schneewind 2003). Collectively, these and other findings suggest that sortases are promising targets for new antibacterial drugs (Schneewind et al. 1993, Frankel et al. 2004, Ton-That et al. 2004, Weis...