Methane (CH4) production is a ubiquitous, apparently unavoidable side effect of fermentative fibre digestion by symbiotic microbiota in mammalian herbivores. Here, a data compilation is presented of in vivo CH4 measurements in individuals of 37 mammalian herbivore species fed forage-only diets, from the literature and from hitherto unpublished measurements. In contrast to previous claims, absolute CH4 emissions scaled linearly to DM intake, and CH4 yields (per DM or gross energy intake) did not vary significantly with body mass. CH4 physiology hence cannot be construed to represent an intrinsic ruminant or herbivore body size limitation. The dataset does not support traditional dichotomies of CH4 emission intensity between ruminants and nonruminants, or between foregut and hindgut fermenters. Several rodent hindgut fermenters and nonruminant foregut fermenters emit CH4 of a magnitude as high as ruminants of similar size, intake level, digesta retention or gut capacity. By contrast, equids, macropods (kangaroos) and rabbits produce few CH4 and have low CH4 : CO2 ratios for their size, intake level, digesta retention or gut capacity, ruling out these factors as explanation for interspecific variation. These findings lead to the conclusion that still unidentified host-specific factors other than digesta retention characteristics, or the presence of rumination or a foregut, influence CH4 production. Measurements of CH4 yield per digested fibre indicate that the amount of CH4 produced during fibre digestion varies not only across but also within species, possibly pointing towards variation in microbiota functionality. Recent findings on the genetic control of microbiome composition, including methanogens, raise the question about the benefits methanogens provide for many (but apparently not to the same extent for all) species, which possibly prevented the evolution of the hosting of low-methanogenic microbiota across mammals.

Understanding the rumen microbial ecosystem requires the identification of factors that influence the community structure, such as nutrition, physiological condition of the host and host–microbiome interactions. The objective of the current study was to describe the rumen microbial communities before, during and after a complete rumen content transfer. The rumen contents of one donor cow were removed completely and used as inoculum for the emptied rumen of the donor itself and three acceptor cows under identical physiological and nutritional conditions. Temporal changes in microbiome composition and rumen function were analysed for each of four cows over a period of 6 weeks. Shortly after transfer, the cows showed different responses to perturbation of their rumen content. Feed intake depression in the first 2 weeks after transfer resulted in short-term changes in milk production, methane emission, fatty acid composition and rumen bacterial community composition. These effects were more pronounced in two cows, whose microbiome composition showed reduced diversity. The fermentation metrics and microbiome diversity of the other two cows were not affected. Their rumen bacterial community initially resembled the composition of the donor but evolved to a new community profile that resembled neither the donor nor their original composition. Descriptive data presented in the current paper show that the rumen bacterial community composition can quickly recover from a reduction in microbiome diversity after a severe perturbation. In contrast to the bacteria, methanogenic communities were more stable over time and unaffected by stress or host effects.

The rumen microbiome has the important task of supplying ruminants with most of their dietary requirements and is responsible for up to 90% of their metabolic needs. This tremendous feat is possible due to the large diversity of microorganisms in the rumen. The rumen is considered one of the most diverse ecosystems on the planet in terms of species diversity and functional richness. From the moment the feed is ingested, it enters a vast cascade in which specialized microorganisms degrade specific components of the feed turning them into molecules, which in turn are utilized as anabolic precursors and energy sources for the animal. The output of this degradation process not only affects the animal, but also has an extensive impact on the environment. Some of the byproducts that are emitted as waste from this process, such as methane, act as greenhouse gases which greatly contribute to global warming. Recent technological advances developed to study this community enabled a larger overview of its vast taxonomic and functional diversity, thus leading to a better understanding of its ecology and function. This deeper understanding of the forces affecting the microbiome includes the forces that shape composition, the variation among animals, the stability of its key components, the processes of succession on a short- and long-time scales such as primary colonization and diurnal oscillations. These collective understandings have helped to provide insights into the potential effects that these forces have on the outputs observed from the animal itself. Over the recent years, there has been a growing body of evidence demonstrating the link between the microbiome and its effect on productivity of the host animals and the environment, which has placed rumen microbiome studies in the forefront of animal agricultural research. In this review, we focus on the natural variations in community composition, which are not the results of different management or feed but rather intrinsic features of animals. We characterize the rumen microbiome, its potential impact on its host as well as the barriers in implementing the current knowledge to modulate the microbiome and point toward potential avenues to overcome these hurdles.

In order to examine the potential survivability of life in the Martian deep subsurface, we have investigated the effects of temperature (45°C, 55°C and 65°C) and pressure (1, 400, 800 and 1200 atm) on the growth, carbon isotopic data and morphology of chemolithoautotrophic anaerobic methanogenic archaea, Methanothermobacter wolfeii. The growth and survivability of this methanogen were determined by measuring the methane concentration in headspace gas samples after the cells were returned to their conventional growth conditions. Interestingly, this methanogen survived at all the temperatures and pressures tested. M. wolfeii demonstrated the highest methane concentration following exposure to pressure of 800 atm and a temperature of 65°C. We found that the stable carbon isotopic fractionation of methane, δ13C(CH4), was slightly more enriched in 12C at 1 atm and 55°C than the carbon isotopic data obtained in other temperature and pressure conditions. A comparison of the images of the cells before and after the exposure to different temperatures and pressures did not show any obvious alteration in the morphology of M. wolfeii. The research reported here suggests that at least one methanogen, M. wolfeii, may be able to survive under hypothetical Martian subsurface conditions with respect to temperature and pressure.

Methane produced from formate is one of the important methanogensis pathways in the rumen. However, quantitative information of CH4 production from formate has been rarely reported. The aim of this study was to characterize the conversion rate (CR) of formic acid into CH4 and CO2 by rumen microorganisms. Ground lucerne hay was incubated with buffered ruminal fluid for 6, 12, 24 and 48 h. Before the incubation, 13C-labeled H13COOH was also supplied into the incubation bottle at a dose of 0, 1.5, 2.2 or 2.9 mg/g of DM substrate. There were no interactions (P>0.05) between dose and incubation time for all variables evaluated. When expressed as an absolute amount (ml in gas sample) or a relative CR (%), both 13CH4 and 13CO2 production quadratically increased (P<0.01) with the addition of H13COOH. The total 13C (13CH4 and 13CO2) CR was also quadratically increased (P<0.01) when H13COOH was added. Moreover, formate addition linearly decreased (P<0.031) the concentrations of NH3-N, total and individual volatile fatty acids (acetate, propionate and butyrate), and quadratically decreased (P<0.014) the populations of protozoa, total methanogens, Methanosphaera stadtmanae, Methanobrevibacter ruminantium M1, Methanobrevibacter smithii and Methanosarcina barkeri. In summary, formate affects ruminal fermentation and methanogenesis, as well as the rumen microbiome, in particular microorganisms which are directly or indirectly involved in ruminal methanogenesis. This study provides quantitative verification for the rapid dissimilation of formate into CH4 and CO2 by rumen microorganisms.

Ruminant-derived methane (CH4), a potent greenhouse gas, is a consequence of microbial fermentation in the digestive tract of livestock. Development of mitigation strategies to reduce CH4 emissions from farmed animals is currently the subject of both scientific and environmental interest. Methanogens are the sole producers of ruminant CH4, and therefore CH4 abatement strategies can either target the methanogens themselves or target the other members of the rumen microbial community that produce substrates necessary for methanogenesis. Understanding the relationship that methanogens have with other rumen microbes is crucial when considering CH4 mitigation strategies for ruminant livestock. Genome sequencing of rumen microbes is an important tool to improve our knowledge of the processes that underpin those relationships. Currently, several rumen bacterial and archaeal genome projects are either complete or underway. Genome sequencing is providing information directly applicable to CH4 mitigation strategies based on vaccine and small molecule inhibitor approaches. In addition, genome sequencing is contributing information relevant to other CH4 mitigation strategies. These include the selection and breeding of low CH4-emitting animals through the interpretation of large-scale DNA and RNA sequencing studies and the modification of other microbial groups within the rumen, thereby changing the dynamics of microbial fermentation.

Vaccination against rumen methanogens offers a practical approach to reduce methane emissions in livestock, particularly ruminants grazing on pasture. Although successful vaccination strategies have been reported for reducing the activity of the rumen-dwelling organism Streptococcus bovis in sheep and S. bovis and Lactobacillus spp. in cattle, earlier approaches using vaccines based on whole methanogen cells to reduce methane production in sheep have produced less promising results. An anti-methanogen vaccine will need to have broad specificity against methanogens commonly found in the rumen and induce antibody in saliva resulting in delivery of sufficiently high levels of antibodies to the rumen to reduce methanogen activity. Our approach has focussed on identifying surface and membrane-associated proteins that are conserved across a range of rumen methanogens. The identification of potential vaccine antigens has been assisted by recent advances in the knowledge of rumen methanogen genomes. Methanogen surface proteins have been shown to be immunogenic in ruminants and vaccination of sheep with these proteins induced specific antibody responses in saliva and rumen contents. Current studies are directed towards identifying key candidate antigens and investigating the level and types of salivary antibodies produced in sheep and cattle vaccinated with methanogen proteins, stability of antibodies in the rumen and their impact on rumen microbial populations. In addition, there is a need to identify adjuvants that stimulate high levels of salivary antibody and are suitable for formulating with protein antigens to produce a low-cost and effective vaccine.

Understanding the interactions between hydrogen producers and consumers in the rumen ecosystem is important for ruminant production and methane mitigation. The present study explored the relationships between rumen protozoa, methanogens and fermentation characteristics. A total of six donor sheep harbouring (F, faunated) or not (D, defaunated) protozoa in their rumens (D animals were kept without protozoa for a period of a few months (D − ) or for more than 2 years (D+)) were used in in vitro and in vivo experiments. In vitro the absence of protozoa decreased NH3 and butyrate production and had no effect on methane. In contrast, the liquid-associated bacterial and methanogens fraction of D+ inocula produced more methane than D −  and F inoculum (P < 0·05). In vivo fermentation parameters of donor animals showed the same trend on NH3 and butyrate and showed that D+ animals were high methane emitters, while D −  were the lowest ( − 35 %). The concentration of dissolved dihydrogen measured after feeding followed the opposite trend. Methane emissions did not correlate with the relative abundance of methanogens in the rumen measured by quantitative PCR, but there was a trend for higher methanogens concentration in the solid-associated population of D+ animals compared with D −  animals. In contrast, PCR-denaturing gradient gel electrophoresis profiles of methanogens' methyl coenzyme-M reductase A gene showed a clear clustering in liquid-associated fractions for all three groups of donors but fewer differences in solid-associated fractions. These results show that the absence of protozoa may affect differently the methanogen community and methane emissions in wethers.

Ruminant production is under increased public scrutiny in terms of the importance of cattle and other ruminants as major producers of the greenhouse gas methane. Methanogenesis is performed by methanogenic archaea, a specialised group of microbes present in several anaerobic environments including the rumen. In the rumen, methanogens utilise predominantly H2 and CO2 as substrates to produce methane, filling an important functional niche in the ecosystem. However, in addition to methanogens, other microbes also have an influence on methane production either because they are involved in hydrogen (H2) metabolism or because they affect the numbers of methanogens or other members of the microbiota. This study explores the relationship between some of these microbes and methanogenesis and highlights some functional groups that could play a role in decreasing methane emissions. Dihydrogen (‘H2’ from this point on) is the key element that drives methane production in the rumen. Among H2 producers, protozoa have a prominent position, which is strengthened by their close physical association with methanogens, which favours H2 transfer from one to the other. A strong positive interaction was found between protozoal numbers and methane emissions, and because this group is possibly not essential for rumen function, protozoa might be a target for methane mitigation. An important function that is associated with production of H2 is the degradation of fibrous plant material. However, not all members of the rumen fibrolytic community produce H2. Increasing the proportion of non-H2 producing fibrolytic microorganisms might decrease methane production without affecting forage degradability. Alternative pathways that use electron acceptors other than CO2 to oxidise H2 also exist in the rumen. Bacteria with this type of metabolism normally occupy a distinct ecological niche and are not dominant members of the microbiota; however, their numbers can increase if the right potential electron acceptor is present in the diet. Nitrate is an alternative electron sinks that can promote the growth of particular bacteria able to compete with methanogens. Because of the toxicity of the intermediate product, nitrite, the use of nitrate has not been fully explored, but in adapted animals, nitrite does not accumulate and nitrate supplementation may be an alternative under some dietary conditions that deserves to be further studied. In conclusion, methanogens in the rumen co-exist with other microbes, which have contrasting activities. A better understanding of these populations and the pathways that compete with methanogenesis may provide novel targets for emissions abatement in ruminant production.