Methane production and diurnal variation measured in dairy cows and predicted from fermentation pattern and nutrient or carbon flow

Many feeding trials have been conducted to quantify enteric methane (CH₄) production in ruminants. Although a relationship between diet composition, rumen fermentation and CH₄ production is generally accepted, the efforts to quantify this relationship within the same experiment remain scarce. In the present study, a data set was compiled from the results of three intensive respiration chamber trials with lactating rumen and intestinal fistulated Holstein cows, including measurements of rumen and intestinal digestion, rumen fermentation parameters and CH₄ production. Two approaches were used to calculate CH₄ from observations: (1) a rumen organic matter (OM) balance was derived from OM intake and duodenal organic matter flow (DOM) distinguishing various nutrients and (2) a rumen carbon balance was derived from carbon intake and duodenal carbon flow (DCARB). Duodenal flow was corrected for endogenous matter, and contribution of fermentation in the large intestine was accounted for. Hydrogen (H₂) arising from fermentation was calculated using the fermentation pattern measured in rumen fluid. CH₄ was calculated from H₂ production corrected for H₂ use with biohydrogenation of fatty acids. The DOM model overestimated CH₄/kg dry matter intake (DMI) by 6.1% (R²=0.36) and the DCARB model underestimated CH₄/kg DMI by 0.4% (R²=0.43). A stepwise regression of the difference between measured and calculated daily CH₄ production was conducted to examine explanations for the deviance. Dietary carbohydrate composition and rumen carbohydrate digestion were the main sources of inaccuracies for both models. Furthermore, differences were related to rumen ammonia concentration with the DOM model and to rumen pH and dietary fat with the DCARB model. Adding these parameters to the models and performing a multiple regression against observed daily CH₄ production resulted in R² of 0.66 and 0.72 for DOM and DCARB models, respectively. The diurnal pattern of CH₄ production followed that of rumen volatile fatty acid (VFA) concentration and the CH₄ to CO₂ production ratio, but was inverse to rumen pH and the rumen hydrogen balance calculated from 4×(acetate+butyrate)/2×(propionate+valerate). In conclusion, the amount of feed fermented was the most important factor determining variations in CH₄ production between animals, diets and during the day. Interactions between feed components, VFA absorption rates and variation between animals seemed to be factors that were complicating the accurate prediction of CH₄. Using a ruminal carbon balance appeared to predict CH₄ production just as well as calculations based on rumen digestion of individual nutrients.     

Measurement and prediction of enteric methane emission

The greenhouse gas (GHG) emissions from the agricultural sector account for about 25.5% of total global anthropogenic emission. While CO₂ receives the most attention as a factor relative to global warming, CH₄, N₂O and chlorofluorocarbons (CFCs) also cause significant radiative forcing. With the relative global warming potential of 25 compared with CO₂, CH₄ is one of the most important GHGs. This article reviews the prediction models, estimation methodology and strategies for reducing enteric CH₄ emissions. Emission of CH₄ in ruminants differs among developed and developing countries, depending on factors like animal species, breed, pH of rumen fluid, ratio of acetate:propionate, methanogen population, composition of diet and amount of concentrate fed. Among the ruminant animals, cattle contribute the most towards the greenhouse effect through methane emission followed by sheep, goats and buffalos, respectively. The estimated CH₄ emission rate per cattle, buffaloe, sheep and goat in developed countries are 150.7, 137, 21.9 and 13.7 (g/animal/day) respectively. However, the estimated rates in developing countries are significantly lower at 95.9 and 13.7 (g/animal/day) per cattle and sheep, respectively. There exists a strong interest in developing new and improving the existing CH₄ prediction models to identify mitigation strategies for reducing the overall CH₄ emissions. A synthesis of the available literature suggests that the mechanistic models are superior to empirical models in accurately predicting the CH₄ emission from dairy farms. The latest development in prediction model is the integrated farm system model which is a process-based whole-farm simulation technique. Several techniques are used to quantify enteric CH₄ emissions starting from whole animal chambers to sulfur hexafluoride (SF6) tracer techniques. The latest technology developed to estimate CH₄ more accurately is the micrometeorological mass difference technique. Because the conditions under which animals are managed vary greatly by country, CH₄ emissions reduction strategies must be tailored to country-specific circumstances. Strategies that are cost effective, improve productivity, and have limited potential negative effects on livestock production hold a greater chance of being adopted by producers. It is also important to evaluate CH₄ mitigation strategies in terms of the total GHG budget and to consider the economics of various strategies. Although reductions in GHG emissions from livestock industries are seen as high priorities, strategies for reducing emissions should not reduce the economic viability of enterprises.     

Relationship between rumen methanogens and methane production in dairy cows fed diets supplemented with a feed enzyme additive

Aims: To investigate the relationship between ruminal methanogen community and host enteric methane (CH₄) production in lactating dairy cows fed diets supplemented with an exogenous fibrolytic enzyme additive. Methods and Results: Ecology of ruminal methanogens from dairy cows fed with or without exogenous fibrolytic enzymes was examined using PCR–denaturing gradient gel electrophoresis (PCR–DGGE) analyses and quantitative real‐time PCR (qRT‐PCR). The density of methanogens was not affected by the enzyme additive or sampling times, and no relationship was observed between the total methanogen population and CH₄ yield (as g per head per day or g kg^?1 DMI). The PCR–DGGE profiles consisted of 26 distinctive bands, with two bands similar to Methanogenic archaeon CH1270 negatively correlated, and one band similar to Methanobrevibacter gottschalkii strain HO positively correlated, with CH₄ yield. Three bands similar to Methanogenic archaeon CH1270 or Methanobrevibacter smithii ATCC 35061 appeared after enzyme was added. Conclusions: Supplementing a dairy cow diet with an exogenous fibrolytic enzyme additive increased CH₄ yield and altered the composition of the rumen methanogen community, but not the overall density of methanogens. Significance and Impact of the Study: This is the first study to identify the correlation between methanogen ecology and host CH₄ yield from lactating dairy cows.     

The effect of regular or reduced‐fat distillers grains with solubles on rumen methanogenesis and the rumen bacterial community

Aims

The effect of feeding dried distillers grains with solubles (DDGS) or reduced‐fat DDGS (RFDG) on ruminal methanogenesis and the rumen bacterial community of dairy cattle was evaluated.

Methods and Results

Treatments were CONT, a diet with no distillers grains; DG, inclusion of 20% DDGS; rfDG, inclusion of 20% RFDG; and MIX, inclusion of 10% DDGS and 10% RFDG. Methane emission was measured; rumen bacterial community was evaluated by sequencing the V4 region of the 16S rRNA gene. Total methane production remained unaffected. However, feeding distillers grains tended to reduce methanogenesis per unit of feed intake, decreased the abundance of the phylum Bacteroidetes and tended to increase Firmicutes. The abundance of Prevotellaceae positively correlated with feed intake; methane emission was positively correlated with the abundance of Prevotellaceae and was negatively correlated with the abundance of Succinivibrionaceae.

Conclusions

DDGS or RFDG may reduce methanogenesis per unit of feed intake; shifts in the abundance of predominant ruminal bacterial families may influence methane formation, likely because of their role on hydrogen liberation and utilization pathways.

Significance and Impact of the Study

Replacing corn and soybean meal with DDGS or RFDG in dairy rations may reduce the proportion of dietary energy wasted as methane, without detrimental effects on the overall bacterial population.     

Impact of High-Concentrate Feeding and Low Ruminal pH on Methanogens and Protozoa in the Rumen of Dairy Cows

Non-lactating dairy cattle were transitioned to a high-concentrate diet to investigate the effect of ruminal pH suppression, commonly found in dairy cattle, on the density, diversity, and community structure of rumen methanogens, as well as the density of rumen protozoa. Four ruminally cannulated cows were fed a hay diet and transitioned to a 65% grain and 35% hay diet. The cattle were maintained on an high-concentrate diet for 3 weeks before the transition back to an hay diet, which was fed for an additional 3 weeks. Rumen fluid and solids and fecal samples were obtained prior to feeding during weeks 0 (hay), 1, and 3 (high-concentrate), and 4 and 6 (hay). Subacute ruminal acidosis was induced during week 1. During week 3 of the experiment, there was a significant increase in the number of protozoa present in the rumen fluid (P = 0.049) and rumen solids (P = 0.004), and a significant reduction in protozoa in the rumen fluid in week 6 (P = 0.003). No significant effect of diet on density of rumen methanogens was found in any samples, as determined by real-time PCR. Clone libraries were constructed for weeks 0, 3, and 6, and the methanogen diversity of week 3 was found to differ from week 6. Week 3 was also found to have a significantly altered methanogen community structure, compared to the other weeks. Twenty-two unique 16S rRNA phylotypes were identified, three of which were found only during high-concentrate feeding, three were found during both phases of hay feeding, and seven were found in all three clone libraries. The genus Methanobrevibacter comprised 99% of the clones present. The rumen fluid at weeks 0, 3, and 6 of all the animals was found to contain a type A protozoal population. Ultimately, high-concentrate feeding did not significantly affect the density of rumen methanogens, but did alter methanogen diversity and community structure, as well as protozoal density within the rumen of nonlactating dairy cattle. Therefore, it may be necessary to monitor the rumen methanogen and protozoal communities of dairy cattle susceptible to depressed pH when methane abatement strategies are being investigated.     

Long-term effect of linseed plus nitrate fed to dairy cows on enteric methane emission and nitrate and nitrite residuals in milk

A previous study showed the additive methane (CH₄)-mitigating effect of nitrate and linseed fed to non-lactating cows. Before practical application, the use of this new strategy in dairy cows requires further investigation in terms of persistency of methanogenesis reduction and absence of residuals in milk products. The objective of this experiment was to study the long-term effect of linseed plus nitrate on enteric CH₄ emission and performance in dairy cows. We also assessed the effect of this feeding strategy on the presence of nitrate residuals in milk products, total tract digestibility, nitrogen (N) balance and rumen fermentation. A total of 16 lactating Holstein cows were allocated to two groups in a randomised design conducted in parallel for 17 weeks. Diets were on a dry matter (DM) basis: (1) control (54% maize silage, 6% hay and 40% concentrate; CON) or (2) control plus 3.5% added fat from linseed and 1.8% nitrate (LIN+NIT). Diets were equivalent in terms of CP (16%), starch (28%) and NDF (33%), and were offered twice daily. Cows were fed ad libitum, except during weeks 5, 16 and 17 in which feed was restricted to 95% of dry matter intake (DMI) to ensure complete consumption of meals during measurement periods. Milk production and DMI were measured weekly. Nitrate and nitrite concentrations in milk and milk products were determined monthly. Daily CH₄ emission was quantified in open circuit respiration chambers (weeks 5 and 16). Total tract apparent digestibility, N balance and rumen fermentation parameters were determined in week 17. Daily DMI tended to be lower with LIN+NIT from week 4 to 16 (−5.1 kg/day on average). The LIN+NIT diet decreased milk production during 6 non-consecutive weeks (−2.5 kg/day on average). Nitrate or nitrite residuals were not detected in milk and associated products. The LIN+NIT diet reduced CH₄ emission to a similar extent at the beginning and end of the trial (−47%, g/day; −30%, g/kg DMI; −33%, g/kg fat- and protein-corrected milk, on average). Diets did not affect N efficiency and nutrients digestibility. In the rumen, LIN+NIT did not affect protozoa number but reduced total volatile fatty acid (−12%) and propionate (−31%) concentrations. We concluded that linseed plus nitrate may have a long-term CH₄-mitigating effect in dairy cows and that consuming milk products from cows fed nitrate may be safe in terms of nitrate and nitrite residuals. Further work is required to optimise the doses of linseed plus nitrate to avoid reduced cows performance.     

Direct conversion of cellulose to methane by anaerobic fungus Neocallimastix frontalis and defined methanogens

Co-cultures of N. frontalis with a formate-utilizing methanogen, Methanobacterium formicicum and/or an aceticlastic methanogen, Methanosaeta concilii, were performed for methane production from cellulose. In the co-culture with M. formicicum, ca. 16 mM CH₄ was produced after 7 days without accumulation of H₂ and formate. In the co-culture with M. concilii, 12 mM CH₄ was produced after 17 days with decreasing acetate production. In the tri-culture of N. frontalis with M. formicicum and M. concilii, 24 mM CH₄ was produced after 17 days where acetate still remained at 23 mM, but production of lactate and ethanol decreased. When a 4-times concentrated culture broth of M. concilii was inoculated in this tri-culture system in a bioreactor, 150 mM CH₄ was produced after 24 days by feeding of cellulose, although 57 mM acetate still accumulated.     

Microbial methane cycling in sediments of Arctic thermokarst lagoons

Thermokarst lagoons represent the transition state from a freshwater lacustrine to a marine environment, and receive little attention regarding their role for greenhouse gas production and release in Arctic permafrost landscapes. We studied the fate of methane (CH₄) in sediments of a thermokarst lagoon in comparison to two thermokarst lakes on the Bykovsky Peninsula in northeastern Siberia through the analysis of sediment CH₄ concentrations and isotopic signature, methane-cycling microbial taxa, sediment geochemistry, lipid biomarkers, and network analysis. We assessed how differences in geochemistry between thermokarst lakes and thermokarst lagoons, caused by the infiltration of sulfate-rich marine water, altered the microbial methane-cycling community. Anaerobic sulfate-reducing ANME-2a/2b methanotrophs dominated the sulfate-rich sediments of the lagoon despite its known seasonal alternation between brackish and freshwater inflow and low sulfate concentrations compared to the usual marine ANME habitat. Non-competitive methylotrophic methanogens dominated the methanogenic community of the lakes and the lagoon, independent of differences in porewater chemistry and depth. This potentially contributed to the high CH₄ concentrations observed in all sulfate-poor sediments. CH₄ concentrations in the freshwater-influenced sediments averaged 1.34 ± 0.98 μmol g⁻¹, with highly depleted δ¹³C-CH₄ values ranging from −89‰ to −70‰. In contrast, the sulfate-affected upper 300 cm of the lagoon exhibited low average CH₄ concentrations of 0.011 ± 0.005 μmol g⁻¹ with comparatively enriched δ¹³C-CH₄ values of −54‰ to −37‰ pointing to substantial methane oxidation. Our study shows that lagoon formation specifically supports methane oxidizers and methane oxidation through changes in pore water chemistry, especially sulfate, while methanogens are similar to lake conditions.     

Changes in in vitro gas and methane production from rumen fluid from dairy cows during adaptation to feed additives in vivo

The adaptation of dairy cows to methane (CH₄)-mitigating feed additives was evaluated using the in vitro gas production (GP) technique. Nine rumen-fistulated lactating Holstein cows were grouped into three blocks and within blocks randomly assigned to one of three experimental diets: Control (CON; no feed additive), Agolin Ruminant^R (AR; 0.05 g/kg dry matter (DM)) or lauric acid (LA; 30 g/kg DM). Total mixed rations composed of maize silage, grass silage and concentrate were fed in a 40 : 30 : 30 ratio on DM basis. Rumen fluid was collected from each cow at days −4, 1, 4, 8, 15 and 22 relative to the introduction of the additives in the diets. On each of these days, a 48-h GP experiment was performed in which rumen fluid from each individual donor cow was incubated with each of the three substrates that reflected the treatment diets offered to the cows. DM intake was on average 19.8, 20.1 and 16.2 kg/day with an average fat- and protein-corrected milk production of 30.7, 31.7 and 26.2 kg/day with diet CON, AR and LA, respectively. In general, feed additives in the donor cow diet had a larger effect on gas and CH₄ production than the same additives in the incubation substrate. Incubation substrate affected asymptotic GP, half-time of asymptotic CH₄ production, total volatile fatty acid (VFA) concentration, molar proportions of propionate and butyrate and degradation of organic matter (OMD), but did not affect CH₄ production. No substrate×day interactions were observed. A significant diet×day interaction was observed for in vitro gas and CH₄ production, total VFA concentration, molar proportions of VFA and OMD. From day 4 onwards, the LA diet persistently reduced gas and CH₄ production, total VFA concentration, acetate molar proportion and OMD, and increased propionate molar proportion. In vitro CH₄ production was reduced by the AR diet on day 8, but not on days 15 and 22. In line with these findings, the molar proportion of propionate in fermentation fluid was greater, and that of acetate smaller, for the AR diet than for the CON diet on day 8, but not on days 15 and 22. Overall, the data indicate a short-term effect of AR on CH₄ production, whereas the CH₄-mitigating effect of LA persisted.     

Effects of Propionibacterium strains on ruminal fermentation,nutrient digestibility and methane emissions in beef cattle fed a corn grain finishing diet

Twenty ruminally cannulated beef heifers were fed a high corn grain diet in a randomized block design to determine the effect of three direct fed microbial (DFM) strains of Propionibacterium on ruminal fermentation, nutrient digestibility and methane (CH₄) emissions. The heifers were blocked in five groups on the basis of BW and used in five 28-day periods. Dietary treatments included (1) Control and three strains of Propionibacterium (2) P169, (3) P5, and (4) P54. Strains were administered directly into the rumen at 5×10⁹ CFU with 10 g of a maltodextrin carrier in a gel capsule; Control heifers received carrier only. All heifers were fed the basal diet (10 : 90 forage to concentrate, dry matter basis). Rumen contents were collected on days 15 and 18, ruminal pH was measured continuously between days 15 and 22, enteric CH₄ emissions were measured between days 19 and 22 and diet digestibility was measured from days 25 to 28. Mean ruminal pH was 5.91 and was not affected by treatments. Similarly, duration of time that pH<5.8 and 5.6 was not affected by treatment. Likewise, total and major volatile fatty acid profiles were similar among all treatments. No effects were observed on dry matter intake and total tract digestibility of nutrients. Total enteric CH₄ production (g/day) was not affected by Propionibacterium strains and averaged 139 g/day. Similarly, mean CH₄ yield (g CH₄/kg of dry matter intake) was similar for all the treatments. The relative abundance of total Propionibacteria in the rumen increased with administration of DFM and were greater 3 h post-dosing relative to Control, but returned to baseline levels before feeding. Populations of Propionibacterium P169 were higher at 3 and 9 h as compared with the levels at 0 h. In conclusion, moderate persistency of the inoculated strains within the ruminal microbiome and pre-existing high propionate production due to elevated levels of starch fermentation might have reduced the efficacy of Propionibacterium strains to increase molar proportion of propionate and subsequently reduce CH₄ emissions.     

The Genome Sequence of the Rumen Methanogen Methanobrevibacter ruminantium Reveals New Possibilities for Controlling Ruminant Methane Emissions

Background

Methane (CH₄) is a potent greenhouse gas (GHG), having a global warming potential 21 times that of carbon dioxide (CO₂). Methane emissions from agriculture represent around 40% of the emissions produced by human-related activities, the single largest source being enteric fermentation, mainly in ruminant livestock. Technologies to reduce these emissions are lacking. Ruminant methane is formed by the action of methanogenic archaea typified by Methanobrevibacter ruminantium, which is present in ruminants fed a wide variety of diets worldwide. To gain more insight into the lifestyle of a rumen methanogen, and to identify genes and proteins that can be targeted to reduce methane production, we have sequenced the 2.93 Mb genome of M. ruminantium M1, the first rumen methanogen genome to be completed.

Methodology/Principal Findings

The M1 genome was sequenced, annotated and subjected to comparative genomic and metabolic pathway analyses. Conserved and methanogen-specific gene sets suitable as targets for vaccine development or chemogenomic-based inhibition of rumen methanogens were identified. The feasibility of using a synthetic peptide-directed vaccinology approach to target epitopes of methanogen surface proteins was demonstrated. A prophage genome was described and its lytic enzyme, endoisopeptidase PeiR, was shown to lyse M1 cells in pure culture. A predicted stimulation of M1 growth by alcohols was demonstrated and microarray analyses indicated up-regulation of methanogenesis genes during co-culture with a hydrogen (H₂) producing rumen bacterium. We also report the discovery of non-ribosomal peptide synthetases in M. ruminantium M1, the first reported in archaeal species.

Conclusions/Significance

The M1 genome sequence provides new insights into the lifestyle and cellular processes of this important rumen methanogen. It also defines vaccine and chemogenomic targets for broad inhibition of rumen methanogens and represents a significant contribution to worldwide efforts to mitigate ruminant methane emissions and reduce production of anthropogenic greenhouse gases.     

Does Dietary Mitigation of Enteric Methane Production Affect Rumen Function and Animal Productivity in Dairy Cows?

It has been suggested that the rumen microbiome and rumen function might be disrupted if methane production in the rumen is decreased. Furthermore concerns have been voiced that geography and management might influence the underlying microbial population and hence the response of the rumen to mitigation strategies. Here we report the effect of the dietary additives: linseed oil and nitrate on methane emissions, rumen fermentation, and the rumen microbiome in two experiments from New Zealand (Dairy 1) and the UK (Dairy 2). Dairy 1 was a randomized block design with 18 multiparous lactating cows. Dairy 2 was a complete replicated 3 x 3 Latin Square using 6 rumen cannulated, lactating dairy cows. Treatments consisted of a control total mixed ration (TMR), supplementation with linseed oil (4% of feed DM) and supplementation with nitrate (2% of feed DM) in both experiments. Methane emissions were measured in open circuit respiration chambers and rumen samples were analyzed for rumen fermentation parameters and microbial population structure using qPCR and next generation sequencing (NGS). Supplementation with nitrate, but not linseed oil, decreased methane yield (g/kg DMI; P<0.02) and increased hydrogen (P<0.03) emissions in both experiments. Furthermore, the effect of nitrate on gaseous emissions was accompanied by an increased rumen acetate to propionate ratio and consistent changes in the rumen microbial populations including a decreased abundance of the main genus Prevotella and a decrease in archaeal mcrA (log₁₀ copies/ g rumen DM content). These results demonstrate that methane emissions can be significantly decreased with nitrate supplementation with only minor, but consistent, effects on the rumen microbial population and its function, with no evidence that the response to dietary additives differed due to geography and different underlying microbial populations.     

Methanogen Communities in a Drained Bog: Effect of Ash Fertilization

Forestry practises such has drainage have been shown to decrease emissions of the greenhouse gas methane (CH₄) from peatlands. The aim of the study was to examine the methanogen populations in a drained bog in northern Finland, and to assess the possible effect of ash fertilization on potential methane production and methanogen communities. Peat samples were collected from control and ash fertilized (15,000 kg/ha) plots 5 years after ash application, and potential CH₄ production was measured. The methanogen community structure was studied by DNA isolation, PCR amplification of the methyl coenzyme-M reductase (mcr) gene, denaturing gradient gel electrophoresis (DGGE), and restriction fragment length polymorphism (RFLP) analysis. The drained peatland showed low potential methane production and methanogen diversity in both control and ash-fertilized plots. Samples from both upper and deeper layers of peat were dominated by three groups of sequences related to Rice cluster-I hydrogenotroph methanogens. Even though pH was marginally greater in the ash-treated site, the occurrence of those sequences was not affected by ash fertilization. Interestingly, a less common group of sequences, related to the Fen cluster, were found only in the fertilized plots. The study confirmed the depth related change of methanogen populations in peatland.     

Biological production of H2, CH4 and CO2 in the deep subsurface of the Iberian Pyrite Belt

Most of the terrestrial deep subsurfaces are oligotrophic environments in which some gases, mainly H₂, CH₄ and CO₂, play an important role as energy and/or carbon sources. In this work, we assessed their biotic and abiotic origin in samples from subsurface hard-rock cores of the Iberian Pyrite Belt (IPB) at three different depths (414, 497 and 520 m). One set of samples was sterilized (abiotic control) and all samples were incubated under anaerobic conditions. Our results showed that H₂, CH₄ and CO₂ remained low and constant in the sterilized controls while their levels were 4, 4.1 and 2.5 times higher respectively, in the unsterilized samples compared to the abiotic controls. The δ¹³C_CH4-values measured in the samples (range −31.2 to −43.0 ‰) reveals carbon isotopic signatures that are within the range for biological methane production. Possible microorganisms responsible for the biotic production of the gases were assessed by CARD-FISH. The analysis of sequenced genomes of detected microorganisms within the subsurface of the IPB allowed to identify possible metabolic activities involved in H₂ (Rhodoplanes, Shewanella and Desulfosporosinus), CH₄ (Methanobacteriales) and CO₂ production. The obtained results suggest that part of the H₂, CH₄ and CO₂ detected in the deep subsurface has a biological origin.     

Effect of fumarate reducing bacteria on in vitro rumen fermentation,methane mitigation and microbial diversity

The metabolic pathways involved in hydrogen (H₂) production, utilization and the activity of methanogens are the important factors that should be considered in controlling methane (CH₄) emissions by ruminants. H₂ as one of the major substrate for CH₄ production is therefore should be controlled. One of the strategies on reducing CH₄ is through the use of hydrogenotrophic microorganisms such as fumarate reducing bacteria. This study determined the effect of fumarate reducing bacteria, Mitsuokella jalaludinii, supplementation on in vitro rumen fermentation, CH4 production, diversity and quantity. M. jalaludinii significantly reduced CH₄ at 48 and 72 h of incubation and significantly increased succinate at 24 h. Although not significantly different, propionate was found to be highest in treatment containing M. jalaludinii at 12 and 48 h of incubation. These results suggest that supplementation of fumarate reducing bacteria to ruminal fermentation reduces CH₄ production and quantity, increases succinate and changes the rumen microbial diversity.     

Effects of feed intake on enteric methane emissions from sheep fed fresh white clover (Trifolium repens) and perennial ryegrass (Lolium perenne) forages

Published analyses of enteric methane (CH₄) emissions from sheep and cattle show an inverse relationship between feed intake and CH₄ yield (g CH₄/kg dry matter (DM) intake), which suggests opportunities for reducing CH₄ emissions from feed eaten and per unit of animal production. Most relationships between feed intake and CH₄ yield have been based on animals fed conserved feeds, especially silages and grains. Our research is a series of experiments with fresh white clover (Trifolium repens) and perennial ryegrass (Lolium perenne; ryegrass) forages fed to sheep at a range of feed intake levels. This study was comprised of four experiments where good quality freshly harvested white clover or ryegrass were fed to sheep over a three-fold range in DM intake, and CH₄ emissions were measured in respiration chambers for two consecutive days in each experiment. Measurements were made from 16 sheep in Experiment 1 (fed at 1.6 × metabolizable energy requirements for maintenance; ME_m), 28 sheep in Experiment 2 (at 0.8 and 2.0 × ME_m), eight sheep and two measurement periods in Experiment 3 (at 1.6 × ME_m), and 30 sheep in Experiment 4 (fed at 0.8, 1.2, 1.6, 2.0 and 2.5 × ME_m). Prior to each experiment, sheep had a 10 d acclimatization period to diets. Apparent digestibility was measured over 7 d from sheep in Experiments 1, 3 and 4, along with collection of rumen digesta for volatile fatty acid (VFA) determination. Although CH₄ yields differed when sheep were fed white clover or ryegrass at similar intakes, the differences were inconsistent and mean values similar across all experiments. This, and a similar structure of all experiments, enabled combined analysis of data from all four experiments using the restricted maximum likelihood (REML) procedure to estimate effects of feed intake level on digestibility, digestible nutrient intake, gas emissions, and VFA concentrations in the rumen. The REML analysis showed that when DM intake increased from 0.40 to 1.60 kg/d, the predicted responses were an increase in CH₄ production (g/d) of 187% (12.4–35.6 g/d; P<0.001), and a decline in CH₄ yield of 21% (25.6–20.2 g/kg DM intake; P<0.001). High feed intake levels were associated with increased molar proportions (mM of total VFA) of propionate from 0.17 to 0.21 (P=0.038). Single and multiple regressions were completed on the data from all experiments, with organic matter (OM) intake predicting 0.87 of the variation in CH₄ production, and molar proportion of propionate predicting 0.60 of the variation in CH₄ yield. Increasing feed intakes by 1 kg/d of DM reduced CH₄ yield by 4.5 g/kg DM intake. Plant chemical composition was weakly related to CH₄ yield. High intakes of fresh forages will lower CH₄ yield from fermentation, but effects of feed composition on CH₄ emissions were minor. The interaction between effects of feed intake and rumen function requires further investigation to understand relationships with CH₄ emissions.     

The effect of dietary concentrate and soya oil inclusion on microbial diversity in the rumen of cattle

Aims: Methane emissions from ruminants are a significant contributor to global greenhouse gas production. The aim of this study was to examine the effect of diet on microbial communities in the rumen of steers. Methods and Results: The effects of dietary alteration (50 : 50 vs 90 : 10 concentrate–forage ratio, and inclusion of soya oil) on methanogenic and bacterial communities in the rumen of steers were examined using molecular fingerprinting techniques (T‐RFLP and automated ribosomal intergenic spacer analysis) and real‐time PCR. Bacterial diversity was greatly affected by diet, whereas methanogen diversity was not. However, methanogen abundance was significantly reduced (P = 0·009) in high concentrate–forage diets and in the presence of soya oil (6%). In a parallel study, reduced methane emissions were observed with these diets. Conclusions: The greater effect of dietary alteration on bacterial community in the rumen compared with the methanogen community may reflect the impact of substrate availability on the rumen bacterial community. This resulted in altered rumen volatile fatty acid profiles and had a downstream effect on methanogen abundance, but not diversity. Significance and Impact of the Study: Understanding how rumen microbial communities contribute to methane production and how these microbes are influenced by diet is essential for the rational design of methane mitigation strategies from livestock.     

Comparison of the effects of lanthanum,cerium and praseodymium on rumen fermentation,nutrient digestion and plasma biochemical parameters in beef cattle

The objectives of the trial were to compare the effects of supplementing rare earth elements (REE) lanthanum (La), cerium (Ce) and praseodymium (Pr) on rumen fermentation, nutrient digestion, methane (CH₄) production, nitrogen (N) balance and plasma biochemical parameters in beef cattle. Four Simmental male cattle, aged 12 months, with initial average liveweight of 333 ± 9 kg and fitted with rumen cannulas, were fed with a basal ration composed of concentrate mixture and maize silage. Animals received a basal ration without adding REE (Control) or three treatments, i.e. supplementing LaCl₃, CeCl₃ or PrCl₃ at 204 mg/kg DM to the basal ration, respectively, which were allocated in a 4 × 4 Latin square design. Each experimental period lasted 15 d, consisting of 12 d for pre-treatment and three subsequent days for sampling. Results showed that all tested levels of REE tended to increase neutral detergent fibre digestibility (p = 0.064) and tended to decrease rumen CH₄ production (p = 0.056). Supplementing LaCl₃ and CeCl₃ decreased total N excretion and urinary N excretion, increased N retention (p < 0.05), tended to increase total urinary purine derivatives (PD) (p = 0.053) and microbial N flow (p = 0.095), whereas supplementing PrCl₃ did not affect N retention, urinary PD and microbial N flow. No differences were found in the effects of nutrient digestibility, CH₄ production and plasma biochemical parameters among LaCl₃, CeCl₃ and PrCl₃. Further trials using graded levels of LaCl₃, CeCl₃ and PrCl₃ in a wide range are needed to obtain more pronounced results for comparing effects of La, Ce and Pr on rumen fermentation and nutrient digestion in beef cattle.     

Effects of nitrate adaptation by rumen inocula donors and substrate fiber proportion on in vitro nitrate disappearance,methanogenesis, and rumen fermentation acid

A study was conducted to evaluate the main effects of dietary nitrate adaptation by cattle and alfalfa cell wall to starch ratio in in vitro substrates on nitrate disappearance and nitrite and volatile fatty acid (VFA) concentrations, as well as hydrogen (H₂) and methane (CH₄) accumulations. Rumen fluid from steers fed diets containing urea or nitrate was added into in vitro incubations containing sodium nitrate as the sole nitrogen source and 20 cell wall : 80 starch or 80 cell wall : 20 starch as the carbohydrate source. The results showed that during 24 h incubation, rumen fluid inoculums from steers adapted to dietary nitrate resulted in more rapid nitrate disappearance by 6 h of incubation (P < 0.01), no significant effect on nitrite concentration and diminished CH₄ accumulation (P < 0.05). Cell wall to starch ratio did not affect nitrate disappearance, CH₄ accumulation and total VFA concentration. The higher cell wall ratio had the lower total gas production and H₂ concentration (P < 0.05). Ammonia-N (NH₃-N) concentration increased because of adaptation of donors to nitrate feeding (P < 0.05). Nitrate adaptation did not alter total VFA concentration, but increased acetate, and decreased propionate and butyrate molar proportions (P < 0.01).     

瘤胃甲烷菌第七目的研究进展

随着测序技术和体外培养技术的进步,越来越多的未知甲烷菌被发现。近年来,第七个甲烷菌目"Methanomassiliicoccales"(简称Mmc)被发现并建立。这一目甲烷菌在进化关系上与热原体古菌相近,但又存在较远距离,独立成簇。Mmc分布广泛,遍布哺乳动物和昆虫的消化道以及稻田、湿地等环境,但不同来源菌株表现出生境偏好性。Mmc缺少将CO2还原为甲基辅酶M的完整代谢途径,导致它们严格利用氢气还原甲基底物生成甲烷。全面深入地了解Mmc在反刍动物瘤胃中发挥的功能,将有助于新型高效反刍动物甲烷减排策略的提出。因此,本文主要综述了Mmc菌株的分离培养、生理生化和基因组特性及其在瘤胃甲烷生成中的作用。