Parameter Optimization And Characteristics Of Microbial Community In Two-stage Process Integrating Hydrogen Fermentation And Anaerobic Digestion From Fresh Livestock Dung

In recent years, with the intensive development of animal industry in China, it can not be ignored that the problem of disposing a number of livestock manures. The Hydrogen and methane production through integrating hydrogen fermentation and anaerobic digestion has become an effective way to deal with cellulose organic matter, it not only obtain clean energy, but also solve the environmental problems caused by organic waste, has double significances. But it is rarely studyed for hydrogen and methane production using fresh cow manure as substrate. So, this paper attempted to construct a two stage fermentation systerm of hydrogen and methane production using fresh cow dung as raw material. Firstly, single factor experimental designs and the response surface methodology（RSM） were used to optimize parameters for fermentation hydrogen production. And than gas production and energy recovery rate were compared between two-step hydrogen and methane fermentation and direct methane production system. The community characteristics of different stages in the fermentation process were also analyzed using molecular biology method. The results were as follows:1. Bacteria which can directly use microcrystalline cellulose for hydrogen production were achieved by domesticating, and hydrogen concent was 30% in hydrogen production process. Results of high-throughput sequencing indicated that Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria were the main community during fermentation process.2. The effects of inoculum, pH, Fe, bioaugmentation dose and C/N ratio on hydrogen production from fresh cow dung（FCD） were investigated separately. The highest hydrogen yield 55.19 m L/g-TVS and hydrogen content 31.4% were achieved under optimal conditions: river sludge as inoculum, pH 5.0, Fe 600 mg/L, bioaugmentation dose 8%, and C/N ratio 24. Then RSM was performed to optimize the media for hydrogen production. The maximum hydrogen yield of 59.94 mL/g-TVS was obtained under the optimal conditions with pH 5.95, bioaugmentation dose 8.32% and C/N 25.2. The microbial community analysised by high-throughput metagenomics sequencing and Polymerase chain reaction-denaturing gradient gel electrophoresis（ PCR-DGGE） method present diversity with adding the bioaugmentation bacteria. And the number of Clostridiales increased significantly.3. The yields of hydrogen and methane in two-stage digestion were 59.85 m L H2/g-TVS and 447.3m L CH₄/g-TVS under optimal conditions,respectively. When the C/N ratio was 25.2 under the single-stage anaerobic digestion process, the cumulative CH₄ yield of cow dung reached to 223.9 m L CH₄/g-TVS. Compared with the one-stage digestion system,COD removal rate and energy recovery were 78% and 44.06% respectely in two-stage digestion, which increased by 13.42% and 10.7% respectely.4. The microbial community of different fermentation times were determined using high-throughput metagenomics sequencing and PCR-DGGE method. The results showed that microorganism population and the diversity of microbial community increased with increase of culture time and then decreased in the fermentation process, the number of Clostridiales and Coriobacteriales increased obviously in hydrogen fermentation stage. Compared with the hydrogen producing process, Firmicutes and Bacteroidetes were the dominant community and Spirochaetae, Synergistetes and Chloroflexi were increased in methane fermentation stage.