Adaptation of the WISH-barcoding approach for ATCC14028s. Dynamics of ATCC14028s WISH-barcoded populations in the streptomycin-pretreated mouse model

Salmonella Typhimurium is a foodborne pathogen that causes gastrointestinal disease. In recent years, barcoded approaches have been established to simultaneously test multiple S. Typhimurium mutants within the same mouse model. This method has been specifically adapted to one of the two most prevalent S. Typhimurium strains in research, SL1344. Here, we adapted a barcoding approach to the second widely used strain, ATCC14028s. A key prerequisite for barcoded pool experiments involving multiple strains in the same mouse model is the stable colonization of wild-type strains. To assess this for ATCC14028s, we barcoded S. Typhimurium with 12 unique barcodes and additionally tested invG- and ssaV-deficient ATCC14028s mutants. Interestingly, we observed random strain loss for wild-type ATCC14028s in two streptomycin-pretreated mouse models C57BL/6J and 129S6/SvEvTac, a phenomenon that was less pronounced in the ΔinvG and ΔssaV parental strains. Furthermore, both mutants exhibited similar colonization kinetics and were still capable of inducing inflammation, albeit to a lesser extent than the ATCC14028s wild type. For future barcoding approaches, we recommend using an ssaV-deficient strain, as it demonstrated more stable colonization while maintaining wild-type-like tissue invasion. However, further experiments are needed to evaluate the potential effects of the ssaV mutation on downstream functions, such as carbohydrate utilization.