The recent availability of extensive metagenomic sequences from various environmental microbial communities has extended the postgenomic era to the field of environmental microbiology. Although still restricted to a small number of studies, metaproteomic investigations have revealed interesting aspects of functional gene expression within microbial habitats that contain limited microbial diversity. These studies highlight the potential of proteomics for the study of microbial consortia. However, the application of proteomic investigations to complex microbial assemblages such as seawater and soil still presents considerable challenges. Nonetheless, metaproteomics will enhance the understanding of the microbial world and link microbial community composition to function.

Metaproteomics: Studying functional gene expression in microbial ecosystems

The rumen is a complex ecosystem composed of anaerobic bacteria, protozoa, fungi, methanogenic archaea and phages These microbes interact closely to breakdown plant material that cannot be digested by humans, whilst providing metabolic energy to the host and, in the case of archaea, producing methane Consequently, ruminants produce meat and milk, which are rich in high-quality protein, vitamins and minerals, and therefore contribute to food security As the world population is predicted to reach approximately 97 billion by 2050, an increase in ruminant production to satisfy global protein demand is necessary, despite limited land availability, and whilst ensuring environmental impact is minimized Although challenging, these goals can be met, but depend on our understanding of the rumen microbiome Attempts to manipulate the rumen microbiome to benefit global agricultural challenges have been ongoing for decades with limited success, mostly due to the lack of a detailed understanding of this microbiome and our limited ability to culture most of these microbes outside the rumen The potential to manipulate the rumen microbiome and meet global livestock challenges through animal breeding and introduction of dietary interventions during early life have recently emerged as promising new technologies Our inability to phenotype ruminants in a high-throughput manner has also hampered progress, although the recent increase in "omic" data may allow further development of mathematical models and rumen microbial gene biomarkers as proxies Advances in computational tools, high-throughput sequencing technologies and cultivation-independent "omics" approaches continue to revolutionize our understanding of the rumen microbiome This will ultimately provide the knowledge framework needed to solve current and future ruminant livestock challenges

/pdf/addressing-global-ruminant-agricultural-challenges-through-3336513qsk.pdf

Addressing global ruminant agricultural challenges through understanding the rumen microbiome: past, present and future

Meat and milk from ruminants provide an important source of protein and other nutrients for human consumption. Although ruminants have a unique advantage of being able to consume forages and graze lands not suitable for arable cropping, 2% to 12% of the gross energy consumed is converted to enteric CH4 during ruminal digestion, which contributes approximately 6% of global anthropogenic greenhouse gas emissions. Thus, ruminant producers need to find cost-effective ways to reduce emissions while meeting consumer demand for food. This paper provides a critical review of the substantial amount of ruminant CH4-related research published in past decades, highlighting hydrogen flow in the rumen, the microbiome associated with methanogenesis, current and future prospects for CH4 mitigation and insights into future challenges for science, governments, farmers and associated industries. Methane emission intensity, measured as emissions per unit of meat and milk, has continuously declined over the past decades due to improvements in production efficiency and animal performance, and this trend is expected to continue. However, continued decline in emission intensity will likely be insufficient to offset the rising emissions from increasing demand for animal protein. Thus, decreases in both emission intensity (g CH4/animal product) and absolute emissions (g CH4/day) are needed if the ruminant industries continue to grow. Providing producers with cost-effective options for decreasing CH4 emissions is therefore imperative, yet few cost-effective approaches are currently available. Future abatement may be achieved through animal genetics, vaccine development, early life programming, diet formulation, use of alternative hydrogen sinks, chemical inhibitors and fermentation modifiers. Individually, these strategies are expected to have moderate effects (<20% decrease), with the exception of the experimental inhibitor 3-nitrooxypropanol for which decreases in CH4 have consistently been greater (20% to 40% decrease). Therefore, it will be necessary to combine strategies to attain the sizable reduction in CH4 needed, but further research is required to determine whether combining anti-methanogenic strategies will have consistent additive effects. It is also not clear whether a decrease in CH4 production leads to consistent improved animal performance, information that will be necessary for adoption by producers. Major constraints for decreasing global enteric CH4 emissions from ruminants are continued expansion of the industry, the cost of mitigation, the difficulty of applying mitigation strategies to grazing ruminants, the inconsistent effects on animal performance and the paucity of information on animal health, reproduction, product quality, cost-benefit, safety and consumer acceptance.

https://www.cambridge.org/core/services/aop-cambridge-core/content/view/8F7537B81CBDA633F48663C1ACF33036/S1751731119003100a.pdf/div-class-title-review-fifty-years-of-research-on-rumen-methanogenesis-lessons-learned-and-future-challenges-for-mitigation-div.pdf

Review: Fifty years of research on rumen methanogenesis: lessons learned and future challenges for mitigation

Invited review: Plant polyphenols and rumen microbiota responsible for fatty acid biohydrogenation, fiber digestion, and methane emission: Experimental evidence and methodological approaches.

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.

/pdf/host-genetics-and-the-rumen-microbiome-jointly-associate-4wvfpcmrod.pdf

Host genetics and the rumen microbiome jointly associate with methane emissions in dairy cows.

Methane emissions from ruminal fermentation contribute significantly to total anthropological greenhouse gas (GHG) emissions. New meta-omics technologies are beginning to revolutionise our understanding of the rumen microbial community structure, metabolic potential and metabolic activity. Here we explore these developments in relation to GHG emissions. Microbial rumen community analyses based on small subunit ribosomal RNA sequence analysis are not yet predictive of methane emissions from individual animals or treatments. Few metagenomics studies have been directly related to GHG emissions. In these studies, the main genes that differed in abundance between high and low methane emitters included archaeal genes involved in methanogenesis, with others that were not apparently related to methane metabolism. Unlike the taxonomic analysis up to now, the gene sets from metagenomes may have predictive value. Furthermore, metagenomic analysis predicts metabolic function better than only a taxonomic description, because different taxa share genes with the same function. Metatranscriptomics, the study of mRNA transcript abundance, should help to understand the dynamic of microbial activity rather than the gene abundance; to date, only one study has related the expression levels of methanogenic genes to methane emissions, where gene abundance failed to do so. Metaproteomics describes the proteins present in the ecosystem, and is therefore arguably a better indication of microbial metabolism. Both two-dimensional polyacrylamide gel electrophoresis and shotgun peptide sequencing methods have been used for ruminal analysis. In our unpublished studies, both methods showed an abundance of archaeal methanogenic enzymes, but neither was able to discriminate high and low emitters. Metabolomics can take several forms that appear to have predictive value for methane emissions; ruminal metabolites, milk fatty acid profiles, faecal long-chain alcohols and urinary metabolites have all shown promising results. Rumen microbial amino acid metabolism lies at the root of excessive nitrogen emissions from ruminants, yet only indirect inferences for nitrogen emissions can be drawn from meta-omics studies published so far. Annotation of meta-omics data depends on databases that are generally weak in rumen microbial entries. The Hungate 1000 project and Global Rumen Census initiatives are therefore essential to improve the interpretation of sequence/metabolic information.

/pdf/application-of-meta-omics-techniques-to-understand-4zdhcn1f5l.pdf

Application of meta-omics techniques to understand greenhouse gas emissions originating from ruminal metabolism

Assessment of archaeol as a molecular proxy for methane production in cattle

The targeting of mcrA or 16S rRNA genes by quantitative PCR (qPCR) has become the dominant method for quantifying methanogens in rumen. There are considerable discrepancies between estimates based on different primer sets, and the literature is equivocal about the relationship with methane production. There are a number of problems with qPCR, including low primer specificity, multiple copies of genes and multiple genomes per cell. Accordingly, we have investigated alternative markers for methanogens, on the basis of the distinctive ether lipids of archaeal cell membranes. The membranes of Archaea contain dialkyl glycerol ethers such as 2,3-diphytanayl-O-sn-glycerol (archaeol), and glycerol dialkyl glycerol tetraethers (GDGTs) such as caldarchaeol (GDGT-0) in different proportions. The relationships between estimates of methanogen abundance using qPCR and archaeol measurements varied across primers. Studies in other ecosystems have identified environmental effects on the profile of ether lipids in Archaea. There is a long history of analysing easily accessible samples, such as faeces, urine and milk, to provide information about digestion and metabolism in livestock without the need for intrusive procedures. Purine derivatives in urine and odd-chain fatty acids in milk have been used to study rumen function. The association between volatile fatty acid proportions and methane production is probably the basis for empirical relationships between milk fatty acid profiles and methane production. However, these studies have not yet identified consistent predictors. We have evaluated the relationship between faecal archaeol concentration and methane production across a range of diets in studies on beef and dairy cattle. Faecal archaeol is diagnostic for ruminant faeces being below the limit of detection in faeces from non-ruminant herbivores. The relationship between faecal archaeol and methane production was significant when comparing treatment means across diets, but appears to be subject to considerable between-animal variation. This variation was also evident in the weak relationship between archaeol concentrations in rumen digesta and faeces. We speculate that variation in the distribution and kinetics of methanogens in the rumen may affect the survival and functioning of Archaea in the rumen and therefore contribute to genetic variation in methane production. Indeed, variation in the relationship between the numbers of micro-organisms present in the rumen and those leaving the rumen may explain variation in relationships between methane production and both milk fatty acid profiles and faecal archaeol. As a result, microbial markers in the faeces and milk are unlikely to relate well back to methanogenesis in the rumen. This work has also highlighted the need to describe methanogen abundance in all rumen fractions and this may explain the difficulty interpreting results on the basis of samples taken using stomach tubes or rumenocentesis.

http://www.chm.bris.ac.uk/~chidb/personal/content/paper51.pdf

Chemical markers for rumen methanogens and methanogenesis

http://www.chm.bris.ac.uk/~chidb/personal/content/paper56.pdf

Using archaeol to investigate the location of methanogens in the ruminant digestive tract

Bacteria were isolated from rabbit faeces using equine caecal fluid as a growth medium. Two new isolates of the genus Streptococcus are described in terms of their biochemical properties. One of these has a 16S rRNA gene with 97.7%, and the other 98.5%, identity to Streptococcus thoraltensis. While S. thoraltensis has been described in the intestinal tract of pigs, it is generally considered to inhabit the porcine genital tract. The biochemical properties of these bacteria indicate that both new isolates showed an ability to digest xylose, an adaptation beneficial for survival in a niche where much of the nutrient supply is of plant origin. Moreover, having bacteria able to digest xylose in the digestive tract should be beneficial to the rabbit, allowing more effective utilisation and digestion of food. This work provides one of the few examples of an analysis of the physiological properties of a bacterium found in the hindgut of the rabbit. By building up a number of such studies, the mechanisms of bacterial digestion in the rabbit will become better understood.

Christine A McCartney

Papers

Application of meta-omics techniques to understand greenhouse gas emissions originating from ruminal metabolism

Assessment of archaeol as a molecular proxy for methane production in cattle

Chemical markers for rumen methanogens and methanogenesis

Using archaeol to investigate the location of methanogens in the ruminant digestive tract

Isolation of Streptococcus thoraltensis from Rabbit Faeces