Effects Of Carbon Source On Methanogenic Activities And Pathways Of Anaerobic Microbial Community

Anaerobic digestion is widely applied for removing organic compounds from various wastewaters because of its low cost,resilience to shock loading,and potential to produce methane?CH₄?and hydrogen?H₂?.Anaerobic digestion occurs as a synergistic process between fermentative bacteria and methanogens.Here,direct and indirect interspecies electron transfer has been revealed as a key mechanism during organic matter degradation and methanogenesis.Interspecies electron transfer is affected by intermediate metabolites formed during the syntrophic metabolism of different organic carbon sources.In this study,four 2-L anaerobic sequencing batch reactors?ASBRs?were fed tryptone?ASBR_Try?,volatile fatty acids?ASBR_VFA?,glucose?ASBR_Glu?or ethanol?ASBR_Eth?,representing protein,volatile fatty acids?VFAs?,carbohydrate and alcohol,respectively,were used as the carbon sources.Their effects on the microbial community structure and methanogenic activity were investigated.Explore the effects of four types of organic compounds on composition,content of extracellular polymeric substances?EPS?and soluble microbial products?SMP?.Metagenomic analyses were conducted to better understand the effect of the carbon source on prevalent methanogenic pathways within the enriched communities.Acetic and propionic acids were the dominant VFAs in ASBR-Glu and propionic acid was more prevalent than acetic acid.The VFAs in ASBR-Glu–indicating inhibition of methanogenesis.This confirmed the notion that the accumulation of the intermediate products,particularly propionic acid,led to the poor performance of ASBR-Glu in terms of COD removal.The lowest CH₄ production rate and ultimate CH₄ yield were both obtained in ASBR-Glu,which was consistent with the results of dynamics of COD and VFAs.In contrast to the degradation pathway for acetic acid,propionate acid could not be utilized directly by methanogens.In addition,the imbalance between propionic acid production and degradation rates can cause the accumulation of propionic acid,which may decrease the pH and cause the system failure.This result further indicated that the accumulation of propionic acid can inhibit the methanogenic activity.When protein and ethanol are used as carbon sources,the content of humus in the extracellular polymer is high,and humus may act as an electron mediator and play an important role in the process of electron transfer of cells and increase the methanogenic activity of microorganisms.When sodium acetate/sodium propionate is used as the carbon source,riboflavin is mainly present in the soluble microbial product?SMP?.When ethanol is used as the carbon source,riboflavin is mainly concentrated in the tightly bound extracellular polymer?TB-EPS?.Riboflavin may be involved in electron transport,and riboflavin in TB-EPS,which is more closely linked to the intima of cells,is more conducive to promoting electron transfer activity between methanogens.From this study,it can be concluded that the microbial community structures are strongly affected by the types of carbon substrate available.The archaeal microbial community structure revealed that Methanosarcina was the most abundant methanogen in the four reactors,accounting for 88%,93%,55%and 58%of total archaea in ASBR-Try,ASBR-VFA,ASBR-Glu and ASBR-Eth,respectively.Methanobacterium was another dominant methanogen in ASBR-Glu?44%?and ASBR-Eth?41%?.Based on the results from microbial community and metagenomic analysis,Methanosarcina was pivotal in the hydrogenotrophic and aceticlastic pathway of methanogenesis in ASBR-Try.In contrast,CH₄ was primarily produced by Methanosarcina using the aceticlastic pathway in ASBR-VFA.In ASBR-Glu,genes encoding formate dehydrogenase and formyl-methanofuran dehydrogenase were primarily assigned to Methanobacterium,suggesting that Methanobacterium was responsible for the hydrogenotrophic pathway,while Methanosarcina was responsible for the aceticlastic pathway.In ASBR-Eth,Methanosarcina may be primarily responsible for CH₄ synthesis via the hydrogenotrophic pathway.In ASBR-Eth,genes encoding pili were abundant and 45%of these genes could be assigned to Geobacter.Genes encoding Geobacter pili can be relatively highly expressed at the transcriptional level.Thus,DIET might also be one of the methanogenic pathways in ASBR-Eth.Geobacter can transfer electrons to Methanosarcina via pili,thus enhancing the methanogenic efficiency.In this study,20%of genes encoding pili were assigned to Desulfovibrio.In addition,Desulfovibrio was observed capable of conducting extracellular electron transfer directly via pili.Plausibly,Desulfovibrio may also participate in biological processes utilizing DIET,but this is speculative and will require further investigation.