Cattle and other ruminants produce large quantities of methane (~110 million metric tonnes per annum), which is a potent greenhouse gas affecting global climate change. Methane (CH₄) is a natural by-product of gastro-enteric microbial fermentation of feedstuffs in the rumen and contributes to 6% of total CH₄ emissions from anthropogenic-related sources. The extent to which the host genome and rumen microbiome influence CH₄ emission is not yet well known. This study confirms individual variation in CH₄ production was influenced by individual host (cow) genotype, as well as the host’s rumen microbiome composition. Abundance of a small proportion of bacteria and archaea taxa were influenced to a limited extent by the host’s genotype and certain taxa were associated with CH₄ emissions. However, the cumulative effect of all bacteria and archaea on CH₄ production was 13%, the host genetics (heritability) was 21% and the two are largely independent. This study demonstrates variation in CH₄ emission is likely not modulated through cow genetic effects on the rumen microbiome. Therefore, the rumen microbiome and cow genome could be targeted independently, by breeding low methane-emitting cows and in parallel, by investigating possible strategies that target changes in the rumen microbiome to reduce CH₄ emissions in the cattle industry. Methane is a potent greenhouse gas and ruminant livestock contribute a substantial amount of total methane from human activities. Variation between cows’ methane production has been found partly due to their genetics (heritable), making genetic selection a promising strategy for breeding low methane emitting cows. We hypothesized that the total methane production by a cow is affected by rumen microbes which are directly responsible for production of methane, as well as the cows’ own genetics and their interaction. We sampled the rumen contents of 750 dairy cows and found the relative abundance of some bacteria and archaea to be heritable and associated with methane production, but the majority of variation in relative abundance of rumen bacteria and archaea is due to non-genetic factors. We compared the amount of variation in methane production associated with host genetics as well as rumen bacteria and archaea and found the host genetics to explain 21% and rumen microbes 13%. Importantly, the two were largely independent of each other, so breeding for low methane emitting cows is unlikely to result in unfavorable changes in the rumen microbiome. However, further functional annotation of rumen microbiota is needed to confirm this. Strategies that target each source of variation can be conducted in parallel to optimize reduction in methane production from dairy cows.

Ruminant livestock is a major source of the potent greenhouse gas methane. The complex rumen microbiome, consisting of bacteria, archaea, and microbial eukaryotes, facilitates anaerobic plant biomass degradation in the cow rumen, leading to methane emissions. Using an integrated approach combining multidomain quantitative metatranscriptomics with gas and volatile fatty acid (VFA) profiling, we aimed at obtaining the most comprehensive picture of the active rumen microbiome during feed degradation to date. Bacterial, archaeal, and eukaryotic biomass, but also methane emissions and VFA concentrations, increased drastically within an hour after feed intake. mRNA profiling revealed a dynamic response of carbohydrate-active enzyme transcripts, transcripts involved in VFA production and methanogenesis. While the relative abundances of functional transcripts did not mirror observed processes, such as methane emissions, transformation to mRNA abundance per gram of rumen fluid echoed ruminant processes. The microbiome composition was highly individual, with, e.g., ciliate, Neocallimastigaceae, Prevotellaceae, Succinivibrionaceae, and Fibrobacteraceae abundances differing between cows. Microbiome individuality was accompanied by inter- and intradomain multifunctional redundancy among microbiome members during feed degradation. This likely enabled the robust performance of the anaerobic degradation process in each rumen. Neocallimastigaceae and ciliates contributed an unexpectedly large share of transcripts for cellulose- and hemicellulose-degrading enzymes, respectively. Methyl-reducing but not CO₂-reducing methanogens were positively correlated with methane emissions. While Methanomassiliicoccales switched from methanol to methylamines as electron acceptors, Methanosphaera became the dominating methanol-reducing methanogen. This study for the first time linked rumen meta-omics with processes and enabled holistic insights into the contribution of all microbiome members to feed degradation. IMPORTANCE Ruminant animals, such as cows, live in a tight symbiotic association with microorganisms, allowing them to feed on otherwise indigestible plant biomass as food sources. Methane is produced as an end product of the anaerobic feed degradation in ruminants and is emitted to the atmosphere, making ruminant animals among the major anthropogenic sources of the potent greenhouse gas methane. Using newly developed quantitative metatranscriptomics for holistic microbiome analysis, we here identified bacterial, archaeal, and eukaryotic key players and the short-term dynamics of the rumen microbiome during anaerobic plant biomass degradation and subsequent methane emissions. These novel insights might pave the way for novel ecologically and economically sustainable methane mitigation strategies, much needed in times of global climate change.

Improving feed efficiency of dairy cows through breeding is expected to reduce enteric methane production per unit of milk produced. This study examined the effect of 2 forage-to-concentrate ratios on methane production, rumen fermentation, and nutrient digestibility in Holstein and Jersey dairy cows divergent in residual feed intake (RFI). Before experimental onset, RFI was estimated using a random regression model on phenotypic herd data. Ten lactating Holstein and 10 lactating Jersey cows were extracted from the herd and allocated to a high or low pre-experimental RFI group of 5 animals each within breed. Cows were fed ad libitum with total mixed rations either low (LC) or high (HC) in concentrates during 3 periods in a crossover design with a back-cross and staggered approach. Forage-to-concentrate ratio was 68:32 for LC and 39:61 for HC. Cows adapted to the diets in 12 to 24 d and feces were subsequently collected on 2 d. Afterward, gas exchange was measured in respiration chambers and rumen liquid was collected once after cows exited the chambers. Pre-experimental RFI was included in the statistical analysis as a class (low and high RFI) or continuous variable. Methane per kilogram of dry matter intake (DMI) was lower for Holsteins than Jerseys and the response to increased concentrate level was more pronounced for Holsteins than Jerseys (27.2 vs.13.8%); a similar pattern was found for the acetate:propionate ratio. However, methane production per kilogram of energy-corrected milk (ECM) was unaffected by breed. Further, total-tract digestibility of neutral detergent fiber was higher for Jerseys than Holsteins. For RFI as a class variable, DMI, methane production regardless of the expression, and digestibility were unaffected by RFI. For RFI as a continuous variable, DMI was lower and methane per kilogram of DMI was higher for cows with negative (efficient) than positive (inefficient) RFI values, and neutral detergent fiber digestibility was higher for Holsteins with negative than positive RFI values, but not for Jerseys. Daily methane production and methane per kilogram of ECM were unaffected by RFI. In conclusion, methane per kilogram of DMI of Jerseys was lowered to a smaller extent in response to the HC diet than of Holsteins. When pre-experimental RFI was used as a continuous variable, higher methane per kilogram of DMI was found for cows with negative RFI than positive RFI values, but not for methane per kilogram of ECM. These findings call for validation in larger studies.

Identifying factors that influence the composition of the microbial population in the digestive system of dairy cattle will be key in regulating these populations to reduce greenhouse gas emissions. In this study, we analyzed rumen and fecal samples from five high residual feed intake (RFI) Holstein cows, five low RFI Holstein cows, five high RFI Jersey cows and five low RFI Jersey cows, fed either a high-concentrate diet (expected to reduce methane emission) or a high-forage diet. Bacterial communities from both the rumen and feces were profiled using Illumina sequencing on the 16S rRNA gene. Rumen archaeal communities were profiled using Terminal-Restriction Fragment Length Polymorphism (T-RFLP) targeting the mcrA gene. The rumen methanogen community was influenced by breed but not by diet or RFI. The rumen bacterial community was influenced by breed and diet but not by RFI. The fecal bacterial community was influenced by individual animal variation and, to a lesser extent, by breed and diet but not by RFI. Only the bacterial community correlated with methane production. Community differences seen in the rumen were reduced or absent in feces, except in the case of animal-to-animal variation, where differences were more pronounced. The two cattle breeds had different levels of response to the dietary intervention; therefore, it may be appropriate to individually tailor methane reduction strategies to each cattle breed.

The complex microbiota that resides within the rumen is responsible for the break-down of plant fibre. The bacteria that attach to ingested plant matter within the rumen are thought to be responsible for initial fibre degradation. Most studies examining the ecology of this important microbiome only offer a ‘snapshot’ in time. We monitored the diversity of rumen bacteria in four New Zealand dairy cows, grazing a rye-grass and clover pasture over five consecutive seasons, using high throughput pyrosequencing of bacterial 16S rRNA genes. We chose to focus on the digesta-adherent bacterial community to learn more about the stability of this community over time. 16S rRNA gene sequencing showed a high level of bacterial diversity, totalling 1539 operational taxonomic units (OTUs, grouped at 96% sequence similarity) across all samples, and ranging from 653 to 926 OTUs per individual sample. The nutritive composition of the pasture changed with the seasons as did the production phase of the animals. Sequence analysis showed that, overall, the bacterial communities were broadly similar between the individual animals. The adherent bacterial community was strongly dominated by members of Firmicutes (82.1%), followed by Bacteroidetes (11.8%). This community differed between the seasons, returning to close to that observed in the same season one year later. These seasonal differences were only small, but were statistically significant (p < 0.001), and were probably due to the seasonal differences in the diet. These results demonstrate a general invariability of the ruminal bacterial community structure in these grazing dairy cattle.

Fatty acids (FA) in bovine milk derive through body mobilization, de novo synthesis or from the feed via the blood stream. To be able to digest feedstuff, the cow depends on its rumen microbiome. The relative abundance of the microbes has been shown to differ between cows. To date, there is little information on the impact of the microbiome on the formation of specific milk FA. Therefore, in this study, our aim was to investigate the impact of the rumen bacterial microbiome on milk FA composition. Furthermore, we evaluated the predictive value of the rumen microbiome and the host genetics on the composition of individual FA in milk. Our results show that the proportion of variance explained by the rumen bacteria composition (termed microbiability or $$h_{B}^{2}$$ ) was generally smaller than that of the genetic component (heritability), and that rumen bacteria influenced most C15:0, C17:0, C18:2 n-6, C18:3 n-3 and CLA cis-9, trans-11 with estimated $$h_{B}^{2}$$ ranging from 0.26 to 0.42. For C6:0, C8:0, C10:0, C12:0, C16:0, C16:1 cis-9 and C18:1 cis-9, the variance explained by the rumen bacteria component was close to 0. In general, both the rumen microbiome and the host genetics had little value for predicting FA phenotype. Compared to genetic information only, adding rumen bacteria information resulted in a significant improvement of the predictive value for C15:0 from 0.22 to 0.38 (P = 9.50e−07) and C18:3 n-3 from 0 to 0.29 (P = 8.81e−18). The rumen microbiome has a pronounced influence on the content of odd chain FA and polyunsaturated C18 FA, and to a lesser extent, on the content of the short- and medium-chain FA in the milk of Holstein cattle. The accuracy of prediction of FA phenotypes in milk based on information from either the animal’s genotypes or rumen bacteria composition was very low.