Differential expression of key systems underpins colony biofilm formation in Clostridioides difficile

Clostridioides difficile is the most common bacterial healthcare-associated infection in the USA (CDC
2019) and disease recurrence has been linked to biofilm formation in the gut. In previous studies,
disruption of the key chaperone gene, dnaK, by ClosTron mutagenesis yielded a DnaK deficient strain
(630Δerm:dnaK) which exhibited 50 % cell elongation, increased cell hydrophobicity, and increased
biofilm production (Jain et al. 2011).
Here, we used a colony biofilm model to investigate gene expression in 24 h old C. difficile biofilms and
in planktonically cultured cells using mutant strain (630Δerm:dnaK), parent strain (630Δerm) and wildtype (630). RNA sequencing revealed that the dnaK operon expression was significantly decreased (up to
14.5-fold) in 630 and 630Δerm biofilms (p < 0.001). However, in the dnaK mutant (630Δerm:dnaK)
biofilm only very modest changes in gene expression were observed for the dnaK operon. Thus, unlike
630 and 630Δerm biofilms, the chaperone genes in the dnaK mutant exhibit much less extreme
expressional changes when the organism grows as a biofilm.
More generally, biofilm transcriptomes showed a significant decrease in motility gene expression while
the majority of sporulation associated genes were increased in expression, some by >1000 fold. A large
number of cell wall function related genes exhibited both increased and decreased expression, revealing
a shift in cell wall architecture related to the biofilm mode of life.
Further studies will focus on elucidating the core genes involved in biofilm formation in C. difficile, and
should facilitate for the identification of key biochemical pathways underpinning biofilm development.