Poster Presentation Australian Society for Microbiology Annual Scientific Meeting 2023

A previously uncharacterised hydrogenase dominates hydrogen production in the gut (#211)

Cait Welsh 1 , Rachael Lappan 1 , Thomas D Watts 1 , Princess Cabotaje 2 , Ping Huang 2 , Gemma D' Adamo 3 , Nhu Quynh Doan 1 , Natasha Bong 1 , Jodee Gould 3 , Patricia Wolf 4 , Jason M Ridlon 4 , H. Rex Gaskins 4 , Emily Gulliver 3 , Remy Young 3 , Dena Lyras 1 , Edward Giles 5 , Gustav Berggren 2 , Sam Forster 3 , Chris Greening 1
  1. Monash University, Melbourne, VIC, Australia
  2. Department of Chemistry, Uppsala University, Uppsala, Sweden
  3. Centre for Innate Immunity and Infectious Diseases, Hudson Institute of Medical Research, Melbourne, VIC, Australia
  4. University of Illinois, Urbana-Champaign, Illinois, United States of America
  5. Paediatrics, Monash Health, Melbourne, Victoria, Australia

Bacteria within the human gastrointestinal tract (GIT) primarily grow via anaerobic fermentation of carbohydrates. An important metabolite produced during fermentation is molecular hydrogen (H2), which ensures redox balance is maintained by regeneration of the key electron carriers ferrodoxin and NAD+. Most of the H2 produced in the gut is predicted to be consumed by hydrogenotrophic microbes, although the microbial determinants of H2 cycling in the gut remain largely uncharacterised. In addition, H2 imbalance in the gut is associated with GIT disorders including inflammatory bowel disorder (IBD), and pathogenic bacteria such as Salmonella spp. and Campylobacter spp. were shown to use hydrogen during pathogenesis. In this study, we used metagenomics and metatranscriptomics to probe the microbial community in faecal and gut biopsy samples, in addition to culture-based approaches, to illuminate the key determinants of hydrogen production in the gut. Our results reveal that a previously uncharacterised Group B [FeFe] hydrogenase enzyme is abundant and highly expressed in bacteria within the GIT. Phylogenetic analysis illustrated that these enzymes are present primarily in the Bacteroidetes and Firmicutes. To determine if these putative Group B [FeFe] hydrogenases were functional, expressed and important for H2 production, GIT isolates encoding these hydrogenases were isolated and tested for their capacity to produce H2. The results showed that 15/16 isolates encoding this hydrogenase produced H2 during anaerobic culture in vitro. In addition, RNAseq showed that Group B [FeFe] hydrogenases were expressed during H2 production in the majority of isolates. The organism that produced the most H2 (>400,000 ppm), Clostridium perfringens, encodes four [FeFe] hydrogenases, including two putative Group B [FeFe] hydrogenases. To further dissect the function of these hydrogenases, future experiments will seek to delete the genes that encode them, and profile their expression during growth. In summary, our data identify the major microbial determinants of H2 production in the GIT, and begin to provide insight into the process of hydrogen cycling in this unexplored niche.