A mix of six different bacteria found naturally in the healthy gut system of mice and isolated from feces can succeed where antibiotics fail, and eradicate murine infection by highly contagious strains of the Gram-positive anaerobe Clostridium difficile, including the human epidemic 027/BI strain, report scientists at the Wellcome Trust Sanger Institute. The findings point to a promising approach for treating antibiotic-resistant C. difficile infection and controlling its spread by harnessing naturally occurring microbial communities.
Studies by Gordon Dougan, Ph.D., and colleagues demonstrated that the 027/BI strain of C. difficile establishes a persistent and highly contagious infection in mice that is associated with high levels of spore shedding (supershedding), chronic intestinal pathology, and persistent dysbiosis; essentially an imbalance in intestinal microbiota. While treating these infected animals using the antibiotic vancomycin rapidly suppressed C. difficile excretion to below the culture detection limit, as soon as the drug was withdrawn, the bacterium invariably took hold again and high-level shedding was evident within a week.
Human studies have suggested that as an alternative to antibiotic therapy, administration of homogenized feces from a healthy donor can treat recurrent C. difficile infection by reintroducing the right mix of healthy gut bacteria that can overpower the pathogen. When the Sanger team tested this form of treatment in their supershedding 027/BI C. difficile mouse model, they found that just a single oral treatment of homogenized feces from a healthy mouse donor suppressed shedding levels in the infected animals to below detection limits within just days. The treatment worked every time, and suppression of shedding was associated with a marked loss in contagiousness. Encouragingly, and in contrast with vancomycin therapy, bacterial suppression lasted for months after just a single treatment. The recipient animals in addition demonstrated restored intestinal pathology and a reduction in the expression of proinflammatory genes associated with supershedding C. difficile.
Notably, suppression of C. difficile-shedding levels after the fecal treatment coincided with a rebalancing of gut microbiota in the treated mice. The C. difficile supershedding state is associated with a markedly reduced diversity of gut microbial species, but following fecal therapy this species diversity increased, and the profile of microbiota in the recipients was restored to that found in healthy animals.
Taking their findings one step further the investigators reasoned that there are probably a number of key species of bacteria within healthy fecal samples that are responsible for suppressing C. difficile. To investigate this further the team first incubated the feces used for therapy in nutrient broth to enrich for readily culturable bacteria. They then confirmed that this mix of broth-grown bacteria was still capable of suppressing the C. difficile supershedding state in mice, and shifting gut microbiota populations back towards a "healthy composition" profile.
Next, the team cultured 18 different species from the broth-grown bacteria, including representatives of the four phyla that make up the bulk of mammalian intestinal microbiota: Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria. It was then a case of treating supershedding C. difficile-infected mice with different combinations of these 18 bacteria, to see which combinations were capable of overthrowing the C. difficile infection.
The results showed that while many of the combinations failed to impact on the infection, one mixture of six bacteria effectively and reproducibly repressed the C. difficile supershedder state.
Treatment with this MixB collection of bacteria led to a resolution of intestinal disease and contagiousness, allowed bacterial diversity in the gut to increase, and shifted the recipient animals’ intestinal microbiota profile back to that found in healthy animals. Importantly, the researchers state, much of the increased diversity after treatment was derived from commensal bacteria that were present at low levels pretreatment. This suggested that the MixB bacteria had disrupted colonization by C. difficile 027/BI by triggering an expansion of the suppressed health-associated bacteria and a redistribution of the microbiota to a healthy composition.
Sequencing and phylogenetic analyses indicated that the six mouse feces-derived MixB bacteria were phylogenetically diverse. Three were already known—Staphylococcus warneri, Enterococcus hirae, and Lactobacillus reuteri—but three were novel species of the genera Anaerostipes, Bacteroidetes, and Enterorhabdus. “This mix of bacteria is therefore phylogenetically diverse, including both obligate and facultative anaerobic species, and represents three of the four predominant intestinal microbiota phyla,” the researchers note.
“Our results open the way to reduce the over-use of antibiotic treatment and harness the potential of naturally occurring microbial communities to treat C. difficile infection and transmission, and potentially other diseases associated with microbial imbalances,” Dr. Gordon concludes. “Fecal transplantation is viewed as an alternative treatment but it is not widely used because of the risk of introducing harmful pathogens as well as general patient aversion. This model encapsulates some of the features of fecal therapy and acts as a basis to develop standardized treatment mixture.”
The researchers report their findings in a paper in PLoS Pathogens, titled “Targeted Restoration of the Intestinal Microbiota with a Simple, Defined Bacteriotherapy Resolves Relapsing Clostridium difficile Disease in Mice.”