Clostridium difficile is a leading cause of
nosocomial infections worldwide and has become an urgent public health threat requiring immediate attention. Epidemic lineages of the BI/
NAP1/027 strain type have emerged and spread through health care systems across the globe over the past decade. Limiting person-to-person transmission and eradicating C. difficile, especially the BI/
NAP1/027 strain type, from health care facilities are difficult due to the abundant shedding of spores that are impervious to most interventions. Effective prophylaxis for C. difficile
infection (CDI) is lacking. We have genetically modified a contractile R-type
bacteriocin ("diffocin") from C. difficile strain CD4 to kill BI/
NAP1/027-type strains for this purpose. The natural receptor
binding protein (RBP) responsible for diffocin targeting was replaced with a newly discovered RBP identified within a prophage of a BI/
NAP1/027-type target strain by genome mining. The resulting modified diffocins (a.k.a.
Avidocin-CDs), Av-CD291.1 and Av-CD291.2, were stable and killed all 16 tested BI/
NAP1/027-type strains. Av-CD291.2 administered in
drinking water survived passage through the mouse gastrointestinal (GI) tract, did not detectably alter the mouse gut microbiota or disrupt natural colonization resistance to C. difficile or the
vancomycin-resistant Enterococcus faecium (VREF), and prevented
antibiotic-induced colonization of mice inoculated with BI/
NAP1/027-type spores. Given the high incidence and virulence of the pathogen, preventing colonization by BI/
NAP1/027-type strains and limiting their transmission could significantly reduce the occurrence of the most severe CDIs. This modified diffocin represents a prototype of an
Avidocin-CD platform capable of producing targetable, precision anti-C. difficile agents that can prevent and potentially treat CDIs without disrupting protective indigenous microbiota.
IMPORTANCE: Treatment and prevention strategies for
bacterial diseases rely heavily on traditional
antibiotics, which impose strong selection for resistance and disrupt protective microbiota. One consequence has been an upsurge of opportunistic pathogens, such as Clostridium difficile, that exploit
antibiotic-induced disruptions in gut microbiota to proliferate and cause life-threatening diseases. We have developed alternative agents that utilize contractile bactericidal
protein complexes (R-type
bacteriocins) to kill specific C. difficile pathogens. Efficacy in a preclinical animal study indicates these molecules warrant further development as potential prophylactic agents to prevent C. difficile
infections in humans. Since these agents do not detectably alter the indigenous gut microbiota or colonization resistance in mice, we believe they will be safe to administer as a prophylactic to block transmission in high-risk environments without rendering patients susceptible to enteric
infection after
cessation of treatment.