ABSTRACT Transcriptomic analysis of the genome sequenced Ligilactobacillus salivarius strain IBB3154 grown at two different temperatures (37°C vs 42°C) identified differentially expressed genes involved in metabolic pathways, osmoregulation, and surface protein expression. Two highly expressed genes, sasA1 and sasA2 , which encode cell wall-anchored proteins belonging to the serine-rich repeat protein group, were found to be temperature-inducible. Moonlighting proteins with various functions, such as glyceraldehyde 3-phosphate dehydrogenase, fructose-bisphosphate aldolase, elongation factor Tu, and enolase, were highly expressed at both temperatures. The efficiency of promoters has been confirmed by the β-glucuronidase activity test; however, temperature dependence was not detected. We also found that the P _sasA1 promoter retained its activity in the presence of bile salts. Knowledge of promoters that are highly active in L. salivarius cells can be used to produce strains that are carriers of immunogenic proteins. IMPORTANCE The genome of the strain Ligilactobacillus salivarius IBB3154 was sequenced, and transcriptome analysis was carried out at two different temperatures, allowing the determination of gene expression levels in response to environmental changes (temperature). Genes with higher expression at 42°C were identified. The use of a reporter gene (β- glucuronidase) did not confirm the transcriptomic results; it was found that the promoters of the genes sasA1 and sasA2 were active in the presence of bile salts. This opens up new opportunities for the overexpression of genes of other bacterial species in Ligilactobacillus cells in the intestinal environment.