Changing the Codon Usage of hfq Gene has Profound Effect on Phenotype and Pathogenicity of Salmonella Typhimurium.

Genome recoding with bias codons (synonymous rare codons) or codon pair bias is being used as a method to attenuate virulence mostly in viruses. The target gene chosen for attenuation in general in bacteria is mostly toxin or virulence gene. We have used RNA chaperone hfq, a global post-transcriptional regulator of bacterial gene expression that regulates about 20 % genes in Salmonella, as the target of recoding. The hfq gene was recoded by replacing the codons of hfq gene with synonymous rare codons. Recoding decreased the expression of Hfq protein about two-fold in the mutant as compared to the parent strain. Recoding did not affect growth kinetics, but in growth competition the mutant strain was outcompeted by the parent strain. There was significant decrease in survivability of mutant strain in macrophage as compared to the parent strain. The biofilm formation was significantly impaired in case of recoded mutant. The mutants were also less motile as compared to the parent strain. Intraperitoneal infection of mice with the mutant strain had shown better survival as compared to parent strain. The results show that recoding is an effective method of reducing virulence.