Homoacetogenesis as an alternative hydrogen sink in the rumen : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Microbiology and Genetics at Massey University, Palmerston North, New Zealand.

Ruminant livestock contribute significantly to global greenhouse gas emissions. This is due to microorganisms, known as methanogens that generate methane from hydrogen and carbon dioxide during feed fermentation in the rumen. Mitigation strategies are being developed to reduce methane emissions from ruminants. However, inhibiting methane production may cause accumulation of unused hydrogen in the rumen, which may slow down rumen fermentation and affect animal productivity. Homoacetogens, microbes known to reside in the rumen, can use hydrogen and carbon dioxide to form acetate. Homoacetogens could take over the role of ruminal hydrogen disposal following inhibition of methanogens. The aims of this study were to quantify the involvement of alternative hydrogen utilisers, such as homoacetogens, in hydrogen or electron utilisation. Chemical compounds were screened to identify specific inhibitors of methanogens (BES, acetylene), and both methanogens and homoacetogens (chloroform). Homoacetogenesis was measured via incorporation of 13CO2 into 13C-acetate using a short-term in vitro assay. This short-term in vitro assay measured and confirmed the occurrence of homoacetogenesis in sheep rumen fluid, and it accounted for 1.67% of electron utilisation in fresh rumen fluid. Homoacetogenesis increased in the assay when BES was added, suggesting homoacetogens could increase their activity in the absence of methanogens. Homoacetogenesis decreased with the addition of chloroform, which is known to partially inhibit homoacetogens. Methane formation was inhibited by acetylene in an in vitro serial batch fermentation inoculated with sheep rumen fluid. Homoacetogenesis did not increase, but the homoacetogens were able to grow and maintain themselves as the rumen material was repeatedly diluted and supplemented with fresh feed. Their activity accounted for 2.32% of electron utilisation. To study their significance in the rumen, methane formation was inhibited in sheep using acetylene. Homoacetogenesis increased and accounted for 6.53% of electron utilisation. However, propionate appeared to be the major electron sink (58-88%) in the absence of methanogenesis both in vitro and in vivo. In the future, knowledge of these hydrogen-utilising microorganisms could be used to divert hydrogen or electrons into more beneficial end-products, leading to the transition from a normal methane-producing rumen to an equally or even more productive low methane one.