| dc.description.abstract | Bacillus anthracis is a gram-positive bacterium that causes the deadly anthrax disease. ClpX is a subunit of ClpXP protease that is known to be essential in virulence as well as providing resistance to cell-envelope targeting antibiotics such as penicillin, daptomycin, and the antimicrobial peptide LL-37. While clpX is critical for virulence in B. anthracis, it is unlikely to be directly mediating the effect. Hence, our lab investigated the genes that are differentially expressed in the ¿clpX mutant compared to the wild type B. anthracis through microarray analysis. We found 119 genes that were highly differentially expressed in the ¿clpX mutant. In this study, we focused on three genes sigM and glpF, which are downregulated in the ¿clpX mutant, and msrA, which is upregulated in the ¿clpX mutant because sigM confer resistance to cell-wall targeting antibiotics in the closely related gram-positive bacterial species, Bacillus subtilis while both glpF and msrA confer resistance in Staphylococcus aureus. Our objective was to determine whether loss of siM, glpF and msrA will lead to similar phenotypes as loss of clpX in B. anthracis Sterne. We found that sigM and glpF are more susceptible to penicillin, although in a growth phase dependent manner, and only glpF is critical for daptomycin resistance. Loss of msrA increased the susceptibility to penicillin in both log and stationary phase which was comparable to loss of clpX. Our findings from the heat stress test at 42 °C indicate that the sigM, glpF, or msrA genes might not be required to confer heat stress resistance in B. anthracis since all these mutants grew at the same rate as WT. The loss of glpF reduced the number of surviving cfu/ml in the acidic stress test at pH 4, which was comparable to the ¿clpX mutant, indicating that glpF is crucial for B. anthracis to provide acidic stress resistance. Future studies will examine the susceptibility of these mutants to LL-37 and vancomycin. Complementation of these mutants will serve to further support the importance of these genes for the roles we examined. This research will aid in understanding the mechanism of antibiotic resistance and virulence in the ClpX regulatory network in B. anthracis. | en_US |
