Stress-adapted codon usage enables efficient virulence-gene expression in Salmonella

Abstract Virulence genes in bacterial pathogens are often A/T-rich and horizontally acquired, yet they must be translated efficiently under stress and nutrient limitation encountered during infection. Here we show that synonymous codon usage contributes directly to this problem. Using CodonPipe, a genome-scale framework for synonymous codon-usage analysis, we find that bacterial genomes contain functionally organized codon-usage landscapes that extend well beyond previous knowledge. In Salmonella enterica , virulence genes form a codon signature that is distinct from ribosomal genes and other mobile elements. This signature favors wobble decoding, increased use of rare tRNA isoacceptors, and enrichment of codons previously shown to better preserve translation during amino-acid limitation. Using codon-recoded fluorescent reporters, we show that virulence codon usage outperforms ribosomal codon usage selectively during nutrient starvation and, more strongly, in host-mimicking conditions. In mouse infections, overexpression of virulence-recoded reporters imposes a strong selective cost. Comparative analyses further indicate that related decoding signatures are conserved across diverse Enterobacteriaceae pathogens. These findings support stress-adapted codon usage as a mechanism that promotes virulence-gene expression during infection. More broadly, our work contributes to explain the long-observed A/T-rich codon bias of virulence genes and identifies tRNA charging and modifications as promising broad anti-virulence targets.