The spore‐associated protein BclA1 affects the susceptibility of animals to colonization and infection by Clostridium difficile

The BclA protein is a major component of the outermost layer of spores of a number of bacterial species and Clostridium difficile carries three bclA genes. Using insertional mutagenesis each gene was characterized and spores devoid of these proteins had surface aberrations, reduced hydrophobicity and germinated faster than wild‐type spores. Therefore the BclA proteins were likely major components of the spore surface and when absent impaired the protective shield effect of this outermost layer. Analysis of infection and colonization in mice and hamsters revealed that the 50% infectious dose (ID₅₀) of spores was significantly higher (2‐logs) in the bclA1^? mutant compared to the isogenic wild‐type control, but that levels of toxins (A and B) were indistinguishable from animals dosed with wild‐type spores. bclA1^? spores germinated faster than wild‐type spores yet mice were less susceptible to infection suggesting that BclA1 must play a key role in the initial (i.e. pre‐spore germination) stages of infection. We also show that the ID₅₀ was higher in mice infected with R20291, a ‘hypervirulent’ 027 strain, that carries a truncated BclA1 protein.