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Salmonella spp. are facultative anaerobic, Gram-negative, rod-shaped bacteria and belongs to the Enterobacteriaceae family. Although much has been known about Salmonella pathogenesis, the functional characterizations of certain genes are yet to be explored. The rspA (STM14_1818) is one such gene with putative dehydratase function, and its role in pathogenesis is unknown. The background information showed that rspA gene is upregulated in Salmonella when it resides inside macrophages, which led us to investigate its role in Salmonella pathogenesis. We generated the rspA knockout strain and complement strain in S. Typhimurium 14028. Ex-vivo and in-vivo infectivity was looked at macrophage and epithelial cell lines and Caenorhabditis elegans (C. elegans). The mutant strain differentially formed the biofilm at different temperatures by altering the expression of genes involved in the synthesis of cellulose and curli. Besides, the mutant strain is hyperproliferative intracellularly and showed increased bacterial burden in C. elegans. The mutant strain became more infectious and lethal, causing faster death of the worms than the wild type, and also modulates the worm's innate immunity. Thus, we found that the rspA deletion mutant was more pathogenic. In this study, we concluded that the rspA gene differentially regulates the biofilm formation in a temperature dependent manner by modulating the genes involved in the synthesis of cellulose and curli and negatively regulates the Salmonella virulence for longer persistence inside the host., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The authors have no conflict of interest to declare related to this article. Vidya Devi Negi reports financial support, administrative support, equipment, drugs, or supplies, and travel were provided by National Institute of Technology Rourkela., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Adaptation is an important phenomenon for the survival of any organism in various in-vitro and in-vivo conditions leading to their better survival. We studied Salmonella, for the evolutionary adaptation under different conditions. When Salmonella was passaged for 12 rounds in Luria Bertani (P12-STM) media, F media (F12-STM) and Caenorhabditis elegans (Ce12-STM), we found certain phenotypic variations with respect to unpassaged strain. The passaged strains showed a significant increase in motility, body size and number of flagella they possessed, increased tolerance to ROS and RNS under in-vitro condition. They were more invasive in the human intestinal epithelial cells (Int-407) and showed a higher proliferation rate in the phagocytic cells (U-937 and RAW-264.7) suggesting their efficient intracellular proliferation. The evolved strains were hyper-infectious and reduced host viability significantly in C. elegans. The upregulation of various virulent and regulatory genes observed could be associated with the evolved adaptive virulent phenotype by Salmonella in a given condition. Whole genome sequencing also indicates the genetic alteration including insertion and deletions, single nucleotide polymorphisms which are common in all the strains as well as exposure specific, giving varied phenotype among the passaged strains. This study indicates that Salmonella is showing adaptation, genetic alteration and have become hypervirulent hence is capable of causing disease severity., (Copyright © 2019 Elsevier Ltd. All rights reserved.)