Antivirulence activity of the human gut metabolome.
Bottom Line: Importance: Microbes can communicate through the production and sensing of small molecules.Within the complex ecosystem formed by commensal microbes living in and on the human body, it is likely that these molecular messages are used extensively during the interactions between different microbial species as well as with host cells.Deciphering such a molecular dialect will be fundamental to our understanding of host-microbe interactions in health and disease and may prove useful for the design of new therapeutic strategies that target these mechanisms of communication.
Affiliation: Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.Show MeSH
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Mentions: The results described above established that the inhibitory activity is produced by the intestinal microbiota in the absence of any host factors. In order to determine if individual microbial isolates can produce the bioactive molecule or if a community of microbes is required for such activity, we screened individual isolates from one of the chemostat microbial communities used (donor A, Table 2) for specific strains with inhibitory activity against hilA. This was done using a reporter strain containing a fusion between the promoter of hilA and gfp in pFPV25 (28, 29). The bacterial strains were grown in culture medium, as described in Materials and Methods, and extracted with ethyl acetate. Extracts were dried and resuspended in Luria-Bertani (LB) broth. Salmonella was then grown in LB medium supplemented with these extracts or with an ethyl acetate extract of culture medium alone to the late logarithmic growth phase, and green fluorescent protein (GFP) production was tested using flow cytometry. As can be seen in Fig. 7, most microbial isolates showed little to no inhibitory activity against hilA. However, a specific strain of Clostridium citroniae caused strong inhibition of invasion gene expression. Therefore, this determined not only that the microbiota is involved in the production of the active molecule but also that a single microbial species can produce the biological activity in the laboratory. We tested several other C. citroniae strains as well as strains of closely related species for inhibitory activity using this reporter system and found that multiple strains of C. citroniae were active. In addition, multiple isolates of Clostridium aldenense also produced active molecules, suggesting that a closely related clade within the Clostridiales cluster XIVa (otherwise known as the Lachnospiraceae family) is involved in this phenomenon (Fig. 8A). We also determined hilA mRNA levels using RT-PCR for selected C. citroniae and C. aldenense strains and confirmed that extracts from cultures of these microbes showed strong inhibitory effects on the expression of Salmonella invasion genes (Fig. 8B).
Affiliation: Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.