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Expression of duplicate msa genes in the salmonid pathogen Renibacterium salmoninarum
Linda D Rhodes1, Alison M Coady, Mark S Strom
1Northwest Fisheries Science Center, National Marine Fisheries Service, National Oceanic and Atmospheric Administration, United States Department of Commerce, Seattle, Washington 98112, USA. Linda.Rhodes@noaa.gov
Abstract:
Renibacterium salmoninarum is a gram-positive bacterium responsible for bacterial kidney disease of salmon and trout. R. salmoninarum has two identical copies of the gene encoding major soluble antigen (MSA), an immunodominant, extracellular protein. To determine whether one or both copies of msa are expressed, reporter plasmids encoding a fusion of MSA and green fluorescent protein controlled by 0.6 kb of promoter region from msa1 or msa2 were constructed and introduced into R. salmoninarum. Single copies of the reporter plasmids integrated into the chromosome by homologous recombination. Expression of mRNA and protein from the integrated plasmids was detected, and transformed cells were fluorescent, demonstrating that both msa1 and msa2 are expressed under in vitro conditions. This is the first report of successful transformation and homologous recombination in R. salmoninarum.
Insights
This study shows that both copies of the major soluble antigen (MSA) gene in Renibacterium salmoninarum are expressed. This research is the first to achieve successful transformation and homologous recombination in this fish pathogen.
Area of Science:
- Microbiology
- Fish Pathology
- Molecular Biology
Background:
- Renibacterium salmoninarum causes bacterial kidney disease in salmon and trout.
- The bacterium possesses two identical copies of the major soluble antigen (MSA) gene.
- MSA is an immunodominant, extracellular protein crucial for pathogenesis.
Purpose of the Study:
- To investigate the expression of both msa gene copies in R. salmoninarum.
- To determine if one or both msa genes are transcribed and translated.
- To establish a method for genetic manipulation of R. salmoninarum.
Main Methods:
- Constructed reporter plasmids fusing MSA with green fluorescent protein (GFP) under the control of msa1 or msa2 promoter regions.
- Introduced reporter plasmids into R. salmoninarum.
- Achieved homologous recombination for single-copy integration of plasmids into the bacterial chromosome.
Main Results:
- Detected expression of mRNA and protein from the integrated reporter plasmids.
- Transformed R. salmoninarum cells exhibited fluorescence, confirming protein expression.
- Demonstrated that both msa1 and msa2 genes are expressed under laboratory conditions.
Conclusions:
- Both copies of the major soluble antigen (MSA) gene (msa1 and msa2) are actively expressed by Renibacterium salmoninarum in vitro.
- This study reports the first successful transformation and homologous recombination in R. salmoninarum, opening avenues for further genetic research.
- Understanding msa gene expression is vital for developing strategies against bacterial kidney disease in salmonid aquaculture.