Ompx Contribute to Biofilm Formation, Osmotic Response and Swimming Motility in Citrobacter Werkmanii

Citrobacter werkmanii, an aerobe and mesophilic Proteobacterium, is universal in industrial putrefaction, coastal water, and human blood. Our previous studies have discovered that outer membrane protein X (OmpX) of C. werkmanii is involved in calcium response, but the underlying mechanisms and its molecular characteristics remain elusive. To that end, the ompX gene was deleted from the genome of C. werkmanii and its phenotypic variations were thoroughly investigated in conjunction with the wild type (WT) and complementary strains using biochemical and molecular techniques such as RNA-Seq, respectively. The results demonstrated that deleting ompX reduces biofilm formation on polystyrene and glass surfaces. Meanwhile, Delta ompX's swimming ability but not for its twitching or swarming abilities, was also reduced on semi-solid plates compared with WT, which was caused by inhibition of flagellar assembly genes, such as flgC, flhB, and fliE, etc. Furthermore, ompX inactivation altered susceptibility to various bactericide classes, as well as responses to Ca2+ and Mg2+ stress. In addition, when compared to WT, Delta ompX captures a total of 1,357 deferentially expressed genes (DEGs), of which 465 were up-regulated and 892 were down-regulated, which can be enriched into various GO ontology and KEGG pathway terms. Furthermore, ompX, as well as ompD and ompW, can be modulated at the transcriptional levels by rbsR and tdcA. Overall, the ompX gene contributed to a variety of biological functions in C. werkmanii and could be served as a targeted site for controlling biofilm formation and developing new bactericides.