Applying EEM- PARAFAC Analysis With Quantitative Real-Time PCR to Monitor Methanogenic Activity of High-Solid Anaerobic Digestion of Rice Straw

The methanogenic activity is an important indicator to assess the efficiency of high-solid anaerobic digestion. However, it is not yet elucidated clearly how to detect the parameter rapidly and reliably in the rice straw feeding reactor. Co-inoculated with ruminal digesta and anaerobic sludge, the digestion performance was studied at three different organic loading rates (OLRs). The excitation emission matrix–parallel factor analysis (EEM–PARAFAC) was used to detect dynamic changes in the characteristic of fluorescence components. Our results revealed that CH4 productivity reached 280.90 mL/g volatile solid (VS) with a 54.39% CH4 content under the OLR of 2.26 g/(L⋅d), which amount to 80.29% of its theoretical value. At the OLR of 2.47 g/(L⋅d), the average accumulated NH4+ concentration was 1082.63 mg/L, which resulted in the hydrogenotrophic Methanobacteriales decreasing from 1.70 × 109 to 1.04 × 106 copies/g in the solid residues, whereas the acetotrophic Methanosarcinales increased from 7.89 × 106 to 9.44 × 106 copies/g. The dynamics of the methanogenic community consequently influenced the bioconversion efficiency of rice straw, and CH4 productivity was reduced to 256.54 mL/g VS. The three fluorescent components, at the excitation/emission wavelength of 420 nm/470 nm, 340 nm/430 nm, and 280 nm/340 nm, were decomposed by PARAFAC model in the digestate. Fluorescence intensities of coenzyme F420 and NADH reflected the dynamic changes of CH4-producing activity and anaerobic digestion efficiency, respectively. The coenzyme F420, unique to hydrogenotrophic methanogens, was correlated with methane yield, suggesting they played a dominant role in the anaerobic reactor. This study demonstrates that the EEM–PARAFAC combined with Q-PCR can be used to characterize methanogenic activity variation during the high-solid anaerobic digestion of rice straw with 15% total solid (TS).

Marine ultra-small prokaryotes likely affect the cycling of Carbon, Methane, Nitrogen and Sulfur

Recently, we uncovered the genetic components from six carbon fixation autotrophic pathways in cleaned ultra-small size fractions from marine samples (<0.22 micrometres) gathered worldwide by the Tara Oceans Expedition. This first finding suggested that prokaryotic nanoorganisms, phylogenetically distantly related to the known CPR and DPANN groups, could collectively impact carbon cycling and carbon fixation across the world’s ocean. To extend our mining of the functional and taxonomic microbial dark matter from the ultra-small size fraction from the Tara Oceans Expedition, we investigated the distribution of 28 metabolic pathways associated with the cycling of carbon, methane, nitrogen and sulfur. For all of these pathways, we report the existence of novel metabolic homologs in the ultra-small size fraction of the oceanic microbiome, associated with nanoorganisms belonging to the CPR and DPANN lineages, but also of metabolic homologs exclusively found in marine host taxa belonging to other (still unassigned) microbial lineages. Therefore, we conclude that marine nanoorganisms contribute to a greater diversity of key biogeochemical cycles than currently appreciated. In particular, we suggest that oceanic nanoorganisms may be involved in a metabolic loop around Acetyl-CoA, have an underappreciated genetic potential to degrade methane, contribute to sustaining redox-reactions by producing Coenzyme F420, and affect sulfur cycling, notably as they harbour a complete suite of homologs of enzymes of the SOX system.

Effect of liquid digestate recirculation on biogas production and enzyme activities for anaerobic digestion of corn straw

To accelerate the degradation of substrate, 50% liquid digestate recirculation (LDR) was used in the anaerobic digestion (AD) of corn straw. The effects of recirculation on the enzyme activities and biogas production were investigated by comparing with control reactor (ReactorCK). During the AD process, the fermentation system with 50% LDR was more stable. The average biogas and methane production in ReactorLDR were 7,891 mL·d−1 and 347 mL CH4·g−1 VSadded·d−1 respectively. The total volatile fatty acids (TVFAs) concentration in the two reactors both increased at first and then decreased with time. The LDR made the VFAs accumulation significant, especially propionic acid accumulation in 4 ∼ 16 days. The maximum peak value of cellulase, xylanase, dehydrogenase and coenzyme F420 activities in ReactorLDR were 0.51 mg·g−1·h−1, 0.29 mg·g−1·h−1, 4.88 mL·g−1·h−1 and 6.69 μmol·L−1, respectively, which were higher than that in ReactorCK. With or without recirculation, the concentration of TVFAs was positively correlated with cellulase, xylanase and dehydrogenase activities, while was negatively correlated with coenzyme F420 activity. Besides, a very significant correlation existed between hydrolase and dehydrogenase activities and daily biogas production in ReactorCK. And the peaks of cellulase, xylanase and dehydrogenase activities appeared ahead of the peak of daily biogas production with the LDR.

Analysis of Fecal Samples Investigating Methanogenic Potential from Five Orders of Mammals

Animal waste has been studied as potential fuel sources via bio-digestion. Feces have also been used to inoculate biodigesters. Biodigesters are a waste solution which converts organic matter into a fuel source such as methane. They utilize microbes to breakdown organic matter into substrates, which are then converted into methane as fuel. The final step’s productivity depends on the methanogen content of the biodigester. This study serves to look at the feces of captive animals for use as a digester inoculum. The aim was to assess the potential of different feces for methanogen contribution through literature and sample analysis via fluorescent microscopy looking for F420 autofluorescence. Coenzyme F420 is a fluorescent coenzyme involved in redox reactions in methanogens and has been used in their identification and observation. The samples were from herbivores in the orders Rodentia, Lagomorpha (rabbits), Perissodactyla (odd toed ungulates), Artiodactyla (even toed ungulates) and Diprotodontia (some marsupials). The project thus far has been impeded by two dilemmas: the scarcity of methanogens and obscuring from foliage. Due to these contrasting problems the aim has been minimizing foliage obscurity while retaining enough methanogen presence. It was seen with a series of dilutions that a 1:10 dilution reduced foliage impact. Keywords: Fecal, Methanogens, Coenzyme F420

Cadmium Addition Effects on Anaerobic Digestion with Elevated Temperatures

Anaerobic fermentation with biogas as an energy source is influenced by the presence of heavy metals. However, the availability of the heavy metals is dependent on the digestion temperature. In this study, the impacts of Cd on the characteristics of biogas, substrate biodegradation, and enzyme activity during anaerobic co-digestion were investigated under varying digestion temperatures. The results showed that 1 mg/L initial Cd concentration improved cumulative biogas yields by 404.96%, 16.93%, and 5.56% at 55 °C, 45 °C, and 35 °C, respectively. In contrast, at low temperatures (25 °C), the yield decreased by 0.77%. In the 55 °C group, Cd addition improved the activity of cellulase (p < 0.05) and coenzyme F420 (p < 0.01). The total chemical oxygen demand (COD) during the peak period and the transformation of hydrolytic organic components into volatile fatty acids (VFAs) influenced the CH4 and biogas yields. There were no significant differences in cellulase, dehydrogenase, and coenzyme F420 activities with or without Cd addition when the digestion temperature was 45 °C, 35 °C, and 25 °C. Therefore, thermophilic digestion is recommended for the efficient degradation of Cd-contaminated biowaste. Moreover, the impact of metals on the performance of anaerobic digestion should be considered together with temperature conditions in future research and practice.

Metabolic pathway rerouting inParaburkholderia rhizoxinicaevolved long-overlooked derivatives of coenzyme F420

Coenzyme F420is a specialized redox cofactor with a highly negative redox potential. It supports biochemical processes like methanogenesis, degradation of xenobiotics or the biosynthesis of antibiotics. Although well-studied in methanogenic archaea and actinobacteria, not much is known about F420in Gram-negative bacteria. Genome sequencing revealed F420biosynthetic genes in the Gram-negative, endofungal bacteriumParaburkholderia rhizoxinica, a symbiont of phytopathogenic fungi. Fluorescence microscopy, high-resolution LC-MS, and structure elucidation by NMR demonstrated that the encoded pathway is active and yields unexpected derivatives of F420(3PG-F420). Further analyses of a biogas-producing microbial community showed that these derivatives are more widespread in nature. Genetic and biochemical studies of their biosynthesis established that a specificity switch in the guanylyltransferase CofC re-programmed the pathway to start from 3-phospho-D-glycerate, suggesting a rerouting event during the evolution of F420biosynthesis. Furthermore, the cofactor activity of 3PG-F420was validated, thus opening up perspectives for its use in biocatalysis. The 3PG-F420biosynthetic gene cluster is fully functional inEscherichia coli, enabling convenient production of the cofactor by fermentation.

Soybean oil suppresses ruminal methane production and reduces content of coenzyme F420 in vitro fermentation

This experiment examined which type of oils was a superior suppressor to methane mitigation in ruminants. Four oils, peanut, rapeseed, corn and soybean oils, varying in the contents of unsaturated fatty acids as indicated by their iodine values, were used to investigate their effects on methane production and on the content of the F420 enzyme of ruminal methanogens in an in vitro fermentation. The control group was added with calcium palmitate (100% saturated 16C fatty acid). The results showed that the total gas production over a period of 36 h varied from 20.61 mL to 39.67 mL, and were lower in rapeseed, corn and soybean oil treatments than the control (P < 0.05), but not in the peanut oil treatment. The methane concentration in the total gas differed significantly among groups (P < 0.05), and decreased with the increases of unsaturation degree of the oils. The coenzyme F420 content, as indicated by F420 fluorescence intensity in supernatant of the medium, was significantly lower in the oil treatments than in the control (P < 0.05), and the intensity values decreased with the increases of unsaturation degree of the oils, except for the rapeseed oil treatment. Furthermore, there was a close correlation between F420 content and methane production (r = 0.916). By comparison, soybean oil treatment had higher dehydrogenase activity and bacteria density than the other groups (P < 0.05); but was lower in methanogens and genus entodinium (P < 0.05), except for the rapeseed oil treatment. Overall, soybean oil contained a high level of unsaturated fatty acids, and could be used as an ingredient of ruminant diets for methane suppression.