Dissecting individual pathogen-commensal interactions within a complex gut microbiota community

Abstract

14Interactions of commensal bacteria within the gut microbiota and with invading pathogens 15 are critical in determining the outcome of an infection. While murine studies have been 16 valuable, we lack in vitro tools to monitor community responses to pathogens at a single-17 species level. We have developed a multi-species community of nine representative gut 18 species cultured together as a mixed biofilm and tracked numbers of individual species over 19 time using a qPCR-based approach. Introduction of the major nosocomial gut pathogen, 20Clostridiodes difficile, to this community resulted in increased adhesion of commensals and 21 inhibition of C. difficile multiplication. Interestingly, we observed an increase in individual 22Bacteroides species accompanying the inhibition of C. difficile. Furthermore, Bacteroides 23 dorei reduced C. difficile growth within biofilms, suggesting a role for Bacteroides spp in 24 prevention of C. difficile colonisation. We report here an in vitro tool with excellent 25 applications for investigating bacterial interactions within a complex community. 26 27 28 Disturbed microbiota and the loss of colonisation resistance are associated with several 49 pathogen infections including Clostridioides difficile 8 , Enterohemorrhagic E. coli 15 , and 50 Campylobacter jejuni 16 . 51 In the case of C. difficile, a leading cause of healthcare-associated diarrhoea worldwide, 52 colonisation occurs only when the microbiota is altered, usually due to treatment with 53 antibiotics such as fluoroquinolones 17 . The increased susceptibility of C. difficile infection (CDI) 54 after antibiotic-induced dysbiosis of the gut microbiota is well documented 18,19 . While most 55 4 studies demonstrating the link between CDI and antibiotic therapy are based on changes in 56 microbial populations by microbiota sequencing, recent studies have reported mechanisms by 57 which the microbiota can prevent C. difficile infections 20 . The microbiota in a healthy state 58 consumes or converts primary bile acid into secondary bile acids, reducing the ability of C. 59 difficile to germinate 21 . Secondary bile acids such as deoxycholic acid (DCA) and lithocholic 60 acid (LCA), are toxic to vegetative C. difficile 22,23 . Additionally, gut bacteria like Clostridium 61 scindens which encode secondary bile acid synthesis enzymes have been associated to 62 resistance to C. difficile infection 24 . The microbiota not only competes for resources but 63 actively inhibits C. difficile through production of bacteriocins such as the thuricin CD, 64 produced by Bacteroides thuringiensis 25 . 65While the microbiota is clearly important in preventing infections, current knowledge is mainly 66 based on microbiota profiles from faeces. The gut microbiota is composed of bacteria within 67 the lumen, which are usually detected in faeces and bacteria associated with the gut mucosa. 68Few studies have profiled the adherent microbiota population of healthy human guts as they 69 require invasive biopsies 26 . While not much is known abou...

Species richness determinesC. difficileinvasion outcome in synthetic human gut communities

“…Bacteroides have been found to both inhibit and promote C. difficile growth in different environments 8,10,38,53 , but in our system all Bacteroides species in the community strongly inhibited C. difficile. We did not observe a strong inhibition of C. difficile by C. scindens which has been documented to occur via production of secondary bile acids that inhibit C. difficile germination 9 because our media does not contain bile acids.…”