FIG 6.

Allele-specific effects of combining SipL and SpoIVA mutations on functional spore formation. (A) The indicated C. difficile strains were induced to sporulate for 20 to 24 h. The SpoIVA ATPase motif mutations are carried in the native spoIVA locus, while the SipL LysM domain mutations are carried in a sipL gene integrated into the pyrE locus of a ΔsipL strain. The ratio of heat-resistant spores to total CFU was determined from a minimum of three biological replicates. The ratio shown is the mean and standard deviation for a given strain relative to wild type. The limit of detection of the assay is 10⁻⁶ (black line). The red and blue lines indicate the spore ratio for the sipL I463R and W475E complementation strains, respectively, which had the most severe phenotype of the sipL and spoIVA single point mutations. Statistical significance for all assays was determined relative to the wild type using a one-way ANOVA and Tukey’s test. (B) Western blot analyses of SpoIVA and SipL.