Antibiotic-induced microbiota disruption and its persistence
create
conditions for dysbiosis and colonization by opportunistic pathogens,
such as those causing Clostridioides difficile (C. difficile) infection (CDI), which is the most severe
hospital-acquired intestinal infection. Given the wide differences
in microbiota across hosts and in their recovery after antibiotic
treatments, there is a need for assays to assess the influence of
dysbiosis and its recovery dynamics on the susceptibility of the host
to CDI. Germination of C. difficile spores is a key
virulence trait for the onset of CDI, which is influenced by the level
of primary vs secondary bile acids in the intestinal milieu that is
regulated by the microbiota composition. Herein, the germination of C. difficile spores in fecal supernatant from mice that
are subject to varying degrees of antibiotic treatment is utilized
as an ex vivo assay to predict intestinal dysbiosis
in the host based on their susceptibility to CDI, as determined by in vivo CDI metrics in the same mouse model. Quantification
of spore germination down to lower detection limits than the colony-forming
assay is achieved by using impedance cytometry to count single vegetative
bacteria that are identified based on their characteristic electrical
physiology for distinction vs aggregated spores and cell debris in
the media. As a result, germination can be quantified at earlier time
points and with fewer spores for correlation to CDI outcomes. This
sets the groundwork for a point-of-care tool to gauge the susceptibility
of human microbiota to CDI after antibiotic treatments.