Characterization of the methanogenic Archaea within two-phase biogas reactor systems operated with plant biomass

The two-phase leach-bed system is a biogas reactor system optimized for the utilization of energy crop silages at maximized loading rates under maintenance of an optimal microbial activity. In this study, a characterization of the methanogenic microbial community within this reactor system was conducted for the first time. Accordingly, effluent samples from the anaerobic filter and the silage digesting leach-bed reactors of both a laboratory-scale two-phase biogas reactor system and a scaled-up commercial on-farm pilot plant were investigated. In total, five Archaea-specific 16S rDNA libraries were constructed and analyzed by amplified rDNA restriction analysis (ARDRA), with subsequent phylogenetic analysis of nucleotide sequences for individual ARDRA patterns. A quantification of major methanogenic Archaea groups was conducted by real-time PCR. A total of 663 clones were analyzed and 45 operational taxonomic units (OTUs) related to methanogenic Archaea were detected. These OTUs were related to the orders Methanosarcinales, Methanomicrobiales and Methanobacteriales, as well as the hitherto uncultured CA-11 and ARC-I groups, and most of them occurred throughout all the compartments of both two-phase biogas reactors. The proportion of acetotrophic to hydrogenotrophic methanogens differed between the laboratory and the pilot scale system. A total of 56% of the clones from the 16S rDNA library derived from the laboratory biogas system were assigned to presumably acetotrophic members of Methanosarcinales. In contrast, these OTUs were less abundant in the 16S rDNA library derived from samples of the pilot plant. Therein, the most dominant OTUs were Methanoculleus-related OTUs, which presumably indicated the predominant presence of hydrogenotrophic methanogens. These findings were confirmed by group-specific quantitative real-time PCR assays. The results indicated that the fraction of acetotrophic and hydrogenotrophic methanogens within a biogas reactor caused certain variations, which may reflect varying substrate utilization during methanogenesis.     

Influence of source of organics and soil organic matter content on cyanobacterial nitrogen fixation and distributional pattern under different water regimes

Organic substances viz., sewage and slurry, compost, farmyard manure (FYM) and green manure used in the study under different moisture regimes had a varied effect on cyanobacterial nitrogenase activity and its distributional pattern. Green manure source was the most effective in enhancing cyanobacterial nitrogenase activity, followed by FYM, compost and sewage and slurry. A better response was observed in soils under waterlogged than under moist conditions. A gradual increase in nitrogenase activity was recorded up to one per cent organic carbon both in vitro and in vivo. The cyanobacterial diversity was maximum in sewage and slurry sites.     

Archaea diversity within a commercial biogas plant utilizing herbal biomass determined by 16S rDNA and mcrA analysis

Aims: The Archaea diversity was evaluated in an agricultural biogas plant supplied with cattle liquid manure and maize silage under mesophilic conditions. Methods and Results: Two different genes (16S rRNA; methyl‐coenzyme‐M‐reductase, MCR) targeted by three different PCR primer sets were selected and used for the construction of three clone libraries comprising between 104 and 118 clones. The clone libraries were analysed by restriction fragment polymorphism (RFLP). Between 11 and 31 operational taxonomic units (OTUs) were detected and assigned to orders Methanomicrobiales, Methanosarcinales and Methanobacteriales. Over 70% of all Archaea OTUs belong to the order Methanomicrobiales which mostly include hydrogenotrophic methanogens. Acetotrophic methanogens were detected in minor rates. Similar relative values were obtained by a quantitative real‐time PCR analysis. Conclusions: The results implied that in this biogas plant the most of the methane formation resulted from the conversion of H₂ and CO₂. Significance and Impact of the Study: This study reports, for the first time, a molecular analysis of the archaeal community in this type of agricultural biogas plants. Therein the hydrogenotrophic methanogenesis seems to be the major pathway of methane formation. These results are in contrast with the common thesis that in biogas fermentations the primary substrate for methanogenesis is acetate.     

Fate of bacterial pathogens in cattle dung slurry subjected to anaerobic digestion

The survival of certain pathogenic bacteria was studied in anaerobic batch digesters at room temperature (18–25 °C) as well as at 35 °C under laboratory conditions. The survival of Escherichia coli and Salmonella typhi at room temperature was upto 20 days whereas at 35 °C it was only upto 10 days. Shigella dysenteriae was found to be the most sensitive organism which could survive upto 10 days at room temperature and upto 5 days at 35 °C. The longest survival was observed in case of Streptococcus faecalis which could survive upto 35 days at room temperature and 15 days at 35 °C. The survival time of Salmonella typhi increased when the solid contents of the digester were elevated from 9% to 15%.     

Mathematical model of anaerobic digestion in a chemostat: effects of syntrophy and inhibition

Three of the four main stages of anaerobic digestion: acidogenesis, acetogenesis, and methanogenesis are described by a system of differential equations modelling the interaction of microbial populations in a chemostat. The microbes consume and/or produce simple substrates, alcohols and fatty acids, acetic acid, and hydrogen. Acetogenic bacteria and hydrogenotrophic methanogens interact through syntrophy. The model also includes the inhibition of acetoclastic and hydrogenotrophic methanogens due to sensitivity to varying pH-levels. To examine the effects of these interactions and inhibitions, we first study an inhibition-free model and obtain results for global stability using differential inequalities together with conservation laws. For the model with inhibition, we derive conditions for existence, local stability, and bistability of equilibria and present a global stability result. A case study illustrates the effects of inhibition on the regions of stability. Inhibition introduces regions of bistability and stabilizes some equilibria.     

Elevated methane emissions from a paddy field in southeast China occur after applying anaerobic digestion slurry

One hundred million tons of farm stalk waste and livestock and poultry excrement are used every year in China for the production of clean energy (biogas) by anaerobic digestion. Consequently, a large amount of fermented liquid is produced, and if disposed of improperly, it will result in secondary pollution. Agricultural application of this anaerobic slurry as a liquid fertilizer would reduce possible eutrophication of water sources from random slurry discharge and supply a superior organic fertilizer for farming. This study investigated the effect of applying anaerobic digestion slurry as a liquid fertilizer on the methane emitted from a paddy field. A two‐year (2008–2009) field experiment replacing chemical fertilizer with liquid fertilizer from anaerobically digested pig manure slurry was conducted in a paddy field in Yixing, Jiangsu, China. A static closed chamber method was used to measure methane fluxes over the period from June 2008 to October 2009. All fertilizer treatments increased methane emissions relative to untreated controls, with increases in methane ranging from 40–70% in 2008 to 48–84% in 2009. Paddy fields treated with anaerobically digested pig manure slurry had greater methane emissions (8–84% in 2008 and 3–26% in 2009) than those treated with chemical fertilizer. This suggests that the anaerobic digestion slurry would increase methane emission and so is unsuitable as a liquid fertilizer in paddy fields without development of cultivation practices to limit these emissions.     

pH值对沼液培养的普通小球藻生长及油含量积累的影响

以50％的沼液为普通小球藻的全营养培养基,考察培养基的起始pH值对小球藻生长及油脂含量的影响,普通小球藻对不同初始pH的沼液中氮、磷的去除情况。设定了2组实验,一组只调节初始接种培养液的pH,分别为6.0、6.5、7.0、7.5、8.0、8.5;另一组将培养液pH分别固定在6.0、6.5、7.0、7.5、8.0、8.5,pH用稀HCl和NaOH进行调节。研究发现在pH 6.5和pH 7.0的偏酸环境有利于小球藻生长,而pH在7.0~8.5的偏碱性条件下有利于小球藻油脂的积累,因此综合小球藻生长和油脂积累2个因素,得到最适合小球藻生长和油脂积累的pH为7.0。培养结束后沼液中氮磷的去除率分别达到了95％和97％,沼液中的总氮由原来的134.91 mg/L降至4.86 mg/L,总磷由10.19 mg/L降到0.32 mg/L。