Pathoadaptive Alteration of Biofilm Formation in Response to the Gallbladder Environment.

Typhoid fever, a human-specific disease, is primarily caused by the pathogen serovar Typhi. It is estimated that 3 to 5% of people infected with typhoid fever become chronic carriers. Studies have demonstrated that a mechanism of chronic carriage involves biofilm formation on gallstone surfaces. In the course of a previous study using a chronic carriage mouse model, a serovar Typhimurium isolate was recovered from a mouse gallstone that exhibited a 2-fold increase in biofilm formation over the wild type. In order to identify the gene(s) responsible for the phenotype, the genomic sequences of this isolate and others were determined and compared. These sequences identified single nucleotide polymorphisms (SNPs) in 14 genes. Mutations in the most promising candidates, and were created, but neither showed increased biofilm-forming ability separately or in combination. The hyperbiofilm isolate did, however, present variations in cellular appendages observable using different techniques and a preferential binding to cholesterol. The isolate was also examined for systemic virulence and the ability to colonize the gallbladder/gallstones in a mouse model of chronic infection, demonstrating a systemic virulence defect and decreased gallbladder/gallstone colonization. Finally, to determine if the appearance of hyperbiofilm isolates could be replicated and if this was a common event, wild-type spp. were grown long term under gallbladder-mimicking conditions, resulting in a high proportion of isolates that replicated the hyperbiofilm phenotype of the original isolate. Thus, spp. acquire random mutations under the gallbladder/gallbladder-simulating conditions that may aid persistence but negatively affect systemic virulence. Chronic carriers are the main reservoirs for the spread of typhoid fever in regions of endemicity. Typhi forms biofilms on gallstones in order to persist. A strain with enhanced biofilm-forming ability was recovered after a nine-month chronic-carriage mouse study. After sequencing this strain and recreating some of the mutations, we could not duplicate the phenotype. The isolate did show a difference in flagella, a preference to bind to cholesterol, and a systemic virulence defect. Finally, gallbladder conditions were simulated After 60 days, there was a 4.5-fold increase in hyperbiofilm isolates when a gallstone was present. These results indicate that spp. can undergo genetic changes that improve persistence in gallbladder albeit at the cost of decreased virulence.