Background: Stability and diversity of the infant gut microbiome are essential for healthy digestion, immune development, metabolism, and protection from infection. Dysbiosis of the intestinal microbiota at young ages has been correlated with poor health outcomes, including metabolic disorders, allergy, and inflammatory diseases. In the context of cystic fibrosis (CF), infants as young as six weeks old exhibit gut microbial dysbiosis as judged by the analysis of stool. One signature of this observed dysbiosis is a significant depletion of a key gut microbe, Bacteroides, as well as an enrichment of E. coli. The goal of this proposal is to understand the potential mechanism(s) of Bacteroides depletion in the infant CF gut. Our central hypothesis is that the observed paucity of Bacteroides in the CF gut is driven, at least in part, by (i) the differential environment of the CF intestine and (ii) increased microbial competition with E. coli, both of which contribute to significant shifts in metabolic function of the microbiome and, ultimately, overall negative health outcomes.
Methods: To test the hypothesis that the CF intestinal environment contributes to microbial dysbiosis, we are developing a novel in vitro CF gut medium. This medium utilizes an established gut microbiome medium, MiPro, supplemented with a variety of factors reported to be differentially abundant in the CF gut: fat, oxidative stress, inflammatory markers, nitrate, sulfate, pH, bile salts, antibiotics and mucin. We culture clinical isolates of both Bacteroides and E. coli in each component of this medium to examine which physiological feature(s) may be modulating microbial viability. To test the hypothesis that the Bacteroides-E. coli interaction contributes to Bacteroides depletion, we utilize a co-culture model system consisting of CF clinical isolates of both microbes inoculated at equal densities. After competition (48-72h) in both media, we quantify the growth and survival of each microbe in competition, compared to its monoculture control.
Results: Preliminary results reveal that E. coli isolates can kill Bacteroides in co-culture, and specific features of the CF gut environment modulate growth and survival of Bacteroides in a dose-dependent manner. Under CF and healthy conditions, E. coli growth is unaffected. When in co-culture with E. coli, Bacteroides depletion is exacerbated in the presence of glycerol, likely due to shifts in metabolism by both microbes. Additionally, due to E. coli’s catalase activity, the growth of Bacteroides is rescued when hydrogen peroxide is present.
Conclusions: We have been able to identify physiological factors of the CF gut that may be driving microbial dysbiosis, with a focus on the immune-training microbe, Bacteroides. Concomitantly, we are developing a new in vitro CF gut medium, which will allow us, for the first time, to accurately investigate microbial dynamics of this system. Elucidation of putative causes of microbial dysbiosis will provide insight into effective interventions, which may be used to ameliorate or prevent adverse health outcomes in children with CF.
