Potential to reduce methane production of using cultivated seaweeds supplementation to reshape the community structure of rumen microorganisms.

Methane is a short-lived greenhouse gas but has a far greater warming effect than carbon dioxide. At the same time, the livestock sector serves as a large contributor to global emissions of anthropogenic methane. Herein, this work aimed to use cultivated seaweed supplementation to reduce methane emissions and investigate the potential influencing mechanism. To evaluate the feasibility, two cultivated seaweeds, Laminaria japonica Aresch, and Porphyra tenera, along with the enzymatic hydrolysates derived from L. japonica, underwent in vitro trials, and they were both added into corn silage feed (CSF) with different concentrations (1%, 5%, and 10% of CSF) for methane reduction evaluation. The results indicated that >75% and 50% reductions in methane production were observed for the seaweeds and seaweed enzymatic hydrolysates in 9- and 30-day, respectively. Combined high-throughput sequencing and multivariate analysis revealed that supplementation with seaweed and seaweed enzymatic hydrolysates had a notable impact on the prokaryotic community structure. Mantel tests further revealed that significant correlations between the prokaryotic community and methane accumulation (P < 0.05), implying the prokaryotic community plays a role in reducing methane emissions within the rumen. Correspondingly, the networks within the prokaryotic community unveiled the crucial role of propionate/butyrate-producing bacteria in regulating methane emissions through microbial interactions. The predicted function of the prokaryotic community exhibited a significant reduction in the presence of the narB gene in seaweed-supplemented treatments. This reduction may facilitate an increased rate of electron flow toward the nitrate reduction pathway while decreasing the conversion of H2 to methane. These results indicated the supplementation of cultivated seaweeds and the enzymatic hydrolysates has the potential to reshape the community structure of rumen microbial communities, and this alteration appears to be a key factor contributing to their methane production-reduction capability.

Associations between the rumen microbiota and carcass merit and meat quality in beef cattle.

The rumen microbiota is important for energy and nutrient acquisition in cattle, and therefore its composition may also affect carcass merit and meat quality attributes. In this study, we examined the associations between archaeal and bacterial taxa in the rumen microbiota of beef cattle and 12 different attributes, including hot carcass weight (HCW), dressing percentage, ribeye area (REA), intramuscular fat content, marbling score, fat thickness, yield grade, moisture content, purge loss, and shear force. There were significant correlations between the relative abundance of certain archaeal and bacterial genera and these attributes. Notably, Selenomonas spp. were positively correlated with live weight and HCW, while also being negatively correlated with purge loss. Members of the Christensenellaceae R-7, Moryella, and Prevotella genera exhibited positive and significant correlations with various attributes, such as dressing percentage and intramuscular fat content. Ruminococcaceae UCG-001 was negatively correlated with live weight, HCW, and dressing percentage, while Acidaminococcus and Succinivibrionaceae UCG-001 were negatively correlated with intramuscular fat content, moisture content, and marbling score. Overall, our findings suggest that specific changes in the rumen microbiota could be a valuable tool to improve beef carcass merit and meat quality attributes. Additional research is required to better understand the relationship between the rumen microbiota and these attributes, with the potential to develop microbiome-targeted strategies for enhancing beef production. KEY POINTS: • Certain rumen bacteria were associated with carcass merit and meat quality • Moryella was positively correlated with intramuscular fat in beef carcasses • Acidaminococcus spp. was negatively correlated with marbling and intramuscular fat.

Host and rumen microbiome contributions to feed efficiency traits in Holstein cows.

It is now widely accepted that dairy cow performance is influenced by both the host genome and rumen microbiome composition. The contributions of the genome and the microbiome to the phenotypes of interest are quantified by heritability (h2) and microbiability (m2), respectively. However, if the genome and microbiome are included in the model, then the h2 reflects only the contribution of the direct genetic effects quantified as direct heritability (hd2), and the holobiont effect reflects the joint action of the genome and the microbiome, quantified as the holobiability (ho2). The objectives of this study were to estimate h2, hd2,m2, and ho2 for dry matter intake, milk energy, and residual feed intake; and to evaluate the predictive ability of different models, including genome, microbiome, and their interaction. Data consisted of feed efficiency records, SNP genotype data, and 16S rRNA rumen microbial abundances from 448 mid-lactation Holstein cows from 2 research farms. Three kernel models were fit to each trait: one with only the genomic effect (model G), one with the genomic and microbiome effects (model GM), and one with the genomic, microbiome, and interaction effects (model GMO). The model GMO, or holobiont model, showed the best goodness-of-fit. The hd2 estimates were always 10% to 15% lower than h2 estimates for all traits, suggesting a mediated genetic effect through the rumen microbiome, and m2 estimates were moderate for all traits, and up to 26% for milk energy. The ho2 was greater than the sum of hd2 and m2, suggesting that the genome-by-microbiome interaction had a sizable effect on feed efficiency. Kernel models fitting the rumen microbiome (i.e., models GM and GMO) showed larger predictive correlations and smaller prediction bias than the model G. These findings reveal a moderate contribution of the rumen microbiome to feed efficiency traits in lactating Holstein cows and strongly suggest that the rumen microbiome mediates part of the host genetic effect.

Short- and long-term effects of different forage types supplemented in preweaning dairy calves on performance and milk production into first lactation.

We investigated the short- and long-term effects of different forage types supplemented in preweaning dairy calves on growth performance, blood metabolites, rumen fermentation, bacterial community, and milk production during first lactation. A total of 60 healthy 30-d-old female Holstein calves were blocked by birth date and body weight and randomly assigned to 1 of 3 groups (n = 20): normal milk and pelleted starter feeding (CON), supplemented with chopped oat hay (OAH; 75.0 g/d per calf [DM basis]), or alfalfa hay (ALF; 75.0 g/d per calf [DM basis]). The forage supplementation started when calves were 30 d old (d 1 of the experimental period) and ended when they were 73 d old (d 44 of the experimental period, when calves were weaned). Milk and feed intakes and fecal consistency scores were recorded daily. Growth performance, rumen fluid, and blood samples were collected biweekly. After weaning, all the calves were integrated with the same barn and diets. After calving, the milk production was recorded daily. During the experimental period, the OAH group had greater solid feed and total DM intakes and greater rumen pH than the CON group (P ≤ 0.04), but had lower forage intake and CP digestibility than the ALF group (P ≤ 0.04). The ALF group had higher rumen pH and blood BHB concentration (P ≤ 0.04), lower fecal score (P = 0.02), and greater ether extract digestibility (P = 0.02) than the CON group. The ALF and OAH groups had lower concentrations of ruminal total VFA (P = 0.01). Still, the ALF group had a greater proportion of acetate and a relative abundance of cellulose degradation-related bacteria (Lachnoclostridium_1 and Oribacterium) and a lower relative abundance of inflammation-related bacteria (Erysipelotrichaceae_UCG-009) in the rumen compared with CON. Interestingly, the average milk production from 6 to 200 DIM was greater in the ALF group (P < 0.01), even though no significant effects were found on the rumen fermentation parameters and blood metabolites at 200 DIM. Generally, alfalfa hay supplementation in preweaning dairy calves had positive effects in the short- and long-term for rumen development, health status, and future milk production.

Effects of Chitosan-Based Additive on Rumen Fermentation and Microbial Community, Nutrients Digestibility and Lactation Performance in Goats.

Recently, the potential of using chitosan (CHI) as a feed additive to enhance ruminal fermentation and improve animal performance has gained increasing attention in ruminant nutrition. This study was undertaken to investigate the effect of dietary supplementation with increasing doses of CHI on rumen fermentation attributes and microbial composition, digestibility and milk performance in Dhofari goats. Twenty-four lactating goats (27 ± 1.8 kg of initial live body weight) were fed a control diet comprising of Rhodes grass hay plus a concentrate feed mixture. Goats were assigned to one of three experimental treatments (n = 8 per treatment) as: (1) control diet with no supplement (CTRL), (2) control diet with 0.300 g/day CHI (CHI0.3) and (3) control diet supplemented with 0.600 g/day CHI (CHI0.6) for a 45-day experimental period. Dietary supplementation with increasing doses of CHI decreased (p < 0.05) linearly ruminal pH (p = 0.023), total short chain fatty acids concentrations (p = 0.011), acetate (p = 0.013) and butyrate (p = 0.042) proportions, acetate to propionate ratio (p < 0.001), estimated methane (CH4) production (p < 0.001), ammonia nitrogen concentrations (p = 0.003) and protozoa abundance (p = 0.003). However, the ruminal propionate proportion augmented (p = 0.002) linearly with increasing doses of CHI in the diet. Increasing doses of CHI linearly increased the abundance of the ruminal propionate-producing bacteria, while diminished acetate and CH4-producing bacteria (p < 0.05). Serum total protein (p = 0.037) and glucose (p = 0.042) levels linearly increased as CHI doses increased in the diet. However, serum UREA levels decreased linearly (p = 0.002) by 21% with increasing CHI amounts in the diet. The digestibility of organic matter, crude protein and neutral detergent fibre increased linearly with the increasing CHI doses (p < 0.05). Neither linear nor quadratic responses (p > 0.05) were observed in daily milk yield and feed efficiency by supplementing the diet with CHI. In conclusion, supplementing the diet with CHI at a dose of 0.600 g/day as a feed additive for dairy goats reduced estimated CH4 generation and improved fibre and protein digestion, with no influence on feed intake, milk yield or composition.

Large-scale analysis of sheep rumen metagenome profiles captured by reduced representation sequencing reveals individual profiles are influenced by the environment and genetics of the host.

BACKGROUND: Producing animal protein while reducing the animal's impact on the environment, e.g., through improved feed efficiency and lowered methane emissions, has gained interest in recent years. Genetic selection is one possible path to reduce the environmental impact of livestock production, but these traits are difficult and expensive to measure on many animals. The rumen microbiome may serve as a proxy for these traits due to its role in feed digestion. Restriction enzyme-reduced representation sequencing (RE-RRS) is a high-throughput and cost-effective approach to rumen metagenome profiling, but the systematic (e.g., sequencing) and biological factors influencing the resulting reference based (RB) and reference free (RF) profiles need to be explored before widespread industry adoption is possible. RESULTS: Metagenome profiles were generated by RE-RRS of 4,479 rumen samples collected from 1,708 sheep, and assigned to eight groups based on diet, age, time off feed, and country (New Zealand or Australia) at the time of sample collection. Systematic effects were found to have minimal influence on metagenome profiles. Diet was a major driver of differences between samples, followed by time off feed, then age of the sheep. The RF approach resulted in more reads being assigned per sample and afforded greater resolution when distinguishing between groups than the RB approach. Normalizing relative abundances within the sampling Cohort abolished structures related to age, diet, and time off feed, allowing a clear signal based on methane emissions to be elucidated. Genus-level abundances of rumen microbes showed low-to-moderate heritability and repeatability and were consistent between diets. CONCLUSIONS: Variation in rumen metagenomic profiles was influenced by diet, age, time off feed and genetics. Not accounting for environmental factors may limit the ability to associate the profile with traits of interest. However, these differences can be accounted for by adjusting for Cohort effects, revealing robust biological signals. The abundances of some genera were consistently heritable and repeatable across different environments, suggesting that metagenomic profiles could be used to predict an individual's future performance, or performance of its offspring, in a range of environments. These results highlight the potential of using rumen metagenomic profiles for selection purposes in a practical, agricultural setting.

Enhancing the Conventional Culture: the Evaluation of Several Culture Media and Growth Conditions Improves the Isolation of Ruminal Bacteria.

The rumen microbiota is critical in cattle digestion. Still, its low cultivability makes it difficult to study its ecological function and biotechnological potential. To improve the recovery of ruminal microorganisms, this study combined the evaluation of several cultivation parameters with metabarcoding analysis. The parameters tested comprised eight media cultures, three sample dilutions (10-2, 10-6, 10-12), and two incubation times (3 and 7 days). Bacterial populations were determined through Illumina sequencing of 16S rRNA from three biological replicates. The results indicate that none of the culture media recovered all rumen populations and that there was an altered relative abundance of the dominant phyla. In the rumen, Bacteroidetes and Firmicutes comprised 75% and 15% of the relative abundance, respectively, while in the culture media, these were 15% and 60%, respectively. Principal coordinate analysis (PCoA) of the bacterial community revealed significant shifts in population composition due to dilution, with 10-2 and 10-6 dilutions clustered closely while the 10-12 dilution differed markedly. In contrast, incubation duration did not influence population diversity. According to the results, two media, CAN and KNT, were selected based on their ability to recover more similar populations compared to the rumen sample. The metataxonomic study showed that CAN media had consistent reproducibility over time, while KNT showed enrichment of different taxa due to the use of rumen fluid as a substrate. From these, 64 pure cultures were obtained and 54 were identified through 16S rRNA gene sequencing. Being Streptococcus the most frequently isolated genus, this prevalence contrasts with the liquid media composition, underscoring the importance of refining single colony isolation strategies. Although no culture medium could replicate the native rumen bacterial population perfectly, our findings highlight the potential of CAN and KNT media in recovering populations that are more closely aligned to natural rumen conditions. In conclusion, our study emphasizes the importance of integrating molecular approaches in selecting suitable cultivation media and parameters to depict rumen bacteria accurately.

Rumen multi-omics addressing diet-host-microbiome interplay in farm animals: a review.

Continuous improvement in the living standards of developing countries, calls for an urgent need of high quality meat and dairy products. The farm animals have a micro-ecosystem in gastro-intestinal tract, comprising of a wide variety of flora and fauna which converts roughages and agricultural byproducts as well as nutrient rich concentrate sources into the useful products such as volatile fatty acids and microbial crude proteins. The microbial diversity changes according to composition of the feed, host species/breed and host's individual genetic makeup. From culture methods to next-generation sequencing technologies, the knowledge has emerged a lot to know-how of microbial world viz. their identification, enzymatic activities and metabolites which are the keys of ruminant's successful existence. The structural composition of ruminal community revealed through metagenomics can be elaborated by metatranscriptomics and metabolomics through deciphering their functional role in metabolism and their responses to the external and internal stimuli. These highly sophisticated analytical tools have made possible to correlate the differences in the feed efficiency, nutrients utilization and methane emissions to their rumen microbiome. The comprehensively understood rumen microbiome will enhance the knowledge in the fields of animal nutrition, biotechnology and climatology through deciphering the significance of each and every domain of residing microbial entity. The present review undertakes the recent investigations regarding rumen multi-omics viz. taxonomic and functional potential of microbial populations, host-diet-microbiome interactions and correlation with metabolic dynamics.

Effect of a garlic and citrus extract supplement on performance, rumen fermentation, methane production, and rumen microbiome of dairy cows.

The aim of this trial was to determine the effect of a garlic and citrus extract supplement (GCE) on the performance, rumen fermentation, methane emissions, and rumen microbiome of dairy cows. Fourteen multiparous Nordic Red cows in mid-lactation from the research herd of Luke (Jokioinen, Finland) were allocated to 7 blocks in a complete randomized block design based on body weight, days in milk, dry matter intake (DMI), and milk yield. Animals within each block were randomly allocated to a diet with or without GCE. The experimental period for each block of cows (one for each of the control and GCE groups) consisted of 14 d of adaptation followed by 4 d of methane measurements inside the open circuit respiration chambers, with the first day being considered as acclimatization. Data were analyzed using the GLM procedure of SAS (SAS Institute Inc.). Methane production (g/d) and methane intensity (g/kg of energy-corrected milk) were lower by 10.3 and 11.7%, respectively, and methane yield (g/kg of DMI) tended to be lower by 9.7% in cows fed GCE compared with the control. Dry matter intake, milk production, and milk composition were similar between treatments. Rumen pH and total volatile fatty acid concentrations in rumen fluid were similar, whereas GCE tended to increase molar propionate concentration and decrease the molar ratio of acetate to propionate. Supplementation with GCE resulted in greater abundance of Succinivibrionaceae, which was associated with reduced methane. The relative abundance of the strict anaerobic Methanobrevibacter genus was reduced by GCE. The change in microbial community and rumen propionate proportion may explain the decrease in enteric methane emissions. In conclusion, feeding GCE to dairy cows for 18 d modified rumen fermentation and microbiota, leading to reduced methane production and intensity without compromising DMI or milk production in dairy cows. This could be an effective strategy for enteric methane mitigation of dairy cows.

Rumen metaproteomics: Closer to linking rumen microbial function to animal productivity traits.

The rumen microbiome constitutes a dense and complex mixture of anaerobic bacteria, archaea, protozoa, virus and fungi. Collectively, rumen microbial populations interact closely in order to degrade and ferment complex plant material into nutrients for host metabolism, a process which also produces other by-products, such as methane gas. Our understanding of the rumen microbiome and its functions are of both scientific and industrial interest, as the metabolic functions are connected to animal health and nutrition, but at the same time contribute significantly to global greenhouse gas emissions. While many of the major microbial members of the rumen microbiome are acknowledged, advances in modern culture-independent meta-omic techniques, such as metaproteomics, enable deep exploration into active microbial populations involved in essential rumen metabolic functions. Meaningful and accurate metaproteomic analyses are highly dependent on representative samples, precise protein extraction and fractionation, as well as a comprehensive and high-quality protein sequence database that enables precise protein identification and quantification. This review focuses on the application of rumen metaproteomics, and its potential towards understanding the complex rumen microbiome and its metabolic functions. We present and discuss current methods in sample handling, protein extraction and data analysis for rumen metaproteomics, and finally emphasize the potential of (meta)genome-integrated metaproteomics for accurate reconstruction of active microbial populations in the rumen.

Fermented soybean meal modified the rumen microbiome to enhance the yield of milk components in Holstein cows.

The study was conducted to evaluate the rumen microbiota as well as the milk composition and milk component yields of Holstein cows supplemented with fermented soybean meal (FSBM). Eighteen Holstein cows in their 2nd parity with 54.38 ± 11.12 SD days in milking (DIM) were divided into two dietary groups (CON and TRT) of nine cows per group. The cows in the TRT group received 300 g of FSBM per cow per day in addition to the conventional diet, while each cow in the CON group was supplemented with 350 g of soybean meal (SBM) in their diet daily throughout the 28-day feeding trial. Rumen bacterial composition was detected via 16S rRNA sequencing, and the functional profiles of bacterial communities were predicted. Milk composition, milk yield, as well as rumen fermentation parameters, and serum biochemistry were also recorded. The inclusion of FSBM into the diets of Holstein cows increased the milk urea nitrogen (MUN), milk protein yield, fat corrected milk (FCM), and milk fat yield while the milk somatic cell count (SCC) was decreased. In the rumen, the relative abundances of Fibrobacterota, and Spirochaetota phyla were increased in the TRT group, while the percentage of Proteobacteria was lower. In addition, the supplementation of FSBM to Holstein cows increased the acetate percentage, rumen pH, and acetate to propionate ratio, while the proportion of propionate and propionate % was observed to decrease in the TRT group. The KEGG pathway and functional prediction revealed an upregulation in the functional genes associated with the biosynthesis of amino acids in the TRT group. This enrichment in functional genes resulted in an improved synthesis of several essential amino acids including lysine, methionine, and branch chain amino acids (BCAA) which might be responsible for the increased milk protein yield. Future studies should employ shotgun metagenomics, transcriptomics, and metabolomics technology to investigate the effects of FSBM on other rumen microbiomes and milk protein synthesis in the mammary gland in Holstein cows. KEY POINTS: • The supplementation of fermented soybean meal (FSBM) to Holstein cows modified the proportion of rumen bacteria. • Predicted metabolic pathways and functional genes of rumen bacteria revealed an enrichment in pathway and genes associated with biosynthesis of amino acids in the group fed FSBM. • The cows supplemented with FSBM record an improved rumen fermentation. • Cows supplemented with FSBM recorded an increased yield of milk protein and milk fat.

Rumen bacterial diversity in relation to nitrogen retention in beef cattle.

This study aimed to characterize the rumen bacterial diversity of beef steers differing in the efficiency of nitrogen retention (ENR). Eight castrated steers and fitted with ruminal silicone - and duodenal T-type cannulas were used in a cross-over design with three consecutive periods and three diets. During each experimental period, nitrogen balance was measured, and based on the efficiency of N utilization data, steers were split into three ENR groups: high (HNR, 56.6% ± 3.3%, n = 10), medium (MNR, 45.8% ± 2.2%, n = 6), and low (LNR, 37.7% ± 1.9%, n = 8) using the NbClust package version 2.0.4 in R. Prevotellaceae, Lactobacillaceae, Leuconostocaceae, Clostridiales_Incertae_Sedis_XIII, Lachnospiraceae, and Peptostreptococcaceae were more abundant in LNR (P < 0.05) compared to HNR or MNR. Negative correlations were found between N retention and Mogibacterium, Anaerofustis, Butyrivibrio, Coprococcus, Hespellia, Lactonifactor and Lachnospiraceae (r ≤ -0.61; P ≤ 0.05). Prevotella, Hespellia, Lactonifactor, Lachnospiraceae_other, and Anaerobiospirillum were positively correlated between urinary N excretion (r > 0.55; P < 0.01), and negative correlations were found with Elusimicrobia, Victivallis and Treponema (r < -0.41; P < 0.05). The adjustment of the rumen bacterial community differed significantly between the N use retention groups. The high N retention in beef cattle was associated with less abundant bacteria in the rumen; however, N fixation capacity and uncharacterized rumen microorganisms need to be elucidated in future studies. In contrast, lower N utilization was associated with high abundance of bacteria that promote greater urinary N excretion through ruminal protein degradation.

The effects of 2-hydroxy-4-methylthio-butanoic acid supplementation on the rumen microbial population and duodenal flow of microbial nitrogen.

Four multiparous, lactating Holstein cows (average DIM 169.5 ± 20.5 d), fitted with ruminal and duodenal cannulas, were used in a 4 × 4 Latin square with a 2 × 2 factorial arrangement of treatments to investigate the effects of 2-hydroxy-4-methylthio-butanoic acid (HMTBA) when fed with diets differing in metabolizable protein (MP) supply and equal levels of crude protein on milk production and composition, rumen microbial activity, duodenal protein flow, and rumen bacterial community composition in vivo and in vitro. Experimental periods were 28 d in length. Cows were housed in individual tie stalls and were randomly assigned to 4 dietary treatments: low MP or high MP, supplemented with or without 25 g of HMTBA, which was top-dressed once daily at 0930 h. No interactions were observed between HMTBA and level of dietary MP, with the exception of ruminal acetate-to-propionate ratio. Milk yield was not affected by treatment and averaged 23.8 ± 2.06 kg/d. There was a tendency for increased milk protein percent in cows receiving low MP diets, averaging 3.30 ± 0.09% and 3.21 ± 0.09% for low MP and high MP, respectively. The total-tract apparent digestibility of organic matter, neutral detergent fiber, and nitrogen were greater in cows consuming the low MP diet. Rumen pH was lower in cows consuming high MP diets as well as in those consuming HMTBA. Rumen ammonia concentrations tended to be greater in cows consuming HMTBA, and volatile fatty acid concentrations were greater in cows consuming HMTBA. Duodenal dry matter flow, nitrogen flow, and microbial nitrogen flow did not differ between treatments. The bacterial community structure of cows receiving HMTBA was not affected at the phylum level. The relative abundance of bacterial phyla in vivo differed when compared with in vitro conditions for Firmicutes, Bacteroidetes, Proteobacteria, TM7, Tenericutes, Spirochaetes, SR1, and Verrucomicrobia.

3-Nitrooxypropanol supplementation had little effect on fiber degradation and microbial colonization of forage particles when evaluated using the in situ ruminal incubation technique.

3-Nitrooxypropanol (3-NOP) is an investigational compound that acts as an enzyme inhibitor to decrease ruminal methanogenesis. We hypothesized that when feeding 3-NOP to cattle fed a high-forage diet, H2 would accumulate in the rumen, which could suppress microbial colonization of feed particles and fiber degradation. Therefore, the study investigated the effects of supplementing a high-forage diet with 3-NOP on ruminal fiber degradability and microbial colonization of feed particles using the in situ technique. Eight ruminally cannulated beef cattle were allocated to 2 groups (4 cattle/group) in a crossover design with 2 periods and 2 dietary treatments. The treatments were control (basal diet) and 3-NOP (basal diet supplemented with 3-NOP, 150 mg/kg of dry matter). The basal diet consisted of 45% barley silage, 45% chopped grass hay, and 10% concentrate (dry matter basis). Samples of dried, ground barley silage and grass hay were incubated in the rumen of each animal for 0, 4, 12, 24, 36, 48, 96, 120, 216, and 288 h to determine neutral detergent fiber (NDF) degradation kinetics. An additional 2 bags were incubated for 4 and 48 h to evaluate the bacterial community attached to the incubated forages. Dietary supplementation of 3-NOP decreased (-53%) the dissolved methane concentration and increased (+780%) the dissolved H2 concentration in ruminal fluid, but did not substantially alter in situ NDF degradation. The addition of 3-NOP resulted in a decrease in the α-diversity of the microbial community with colonizing communities showing reduced numbers of amplicon sequence variants and phylogenetic diversity compared with control diets. Principal coordinate analysis plots indicated that forages incubated in animals fed 3-NOP resulted in highly specific changes to targeted microbes compared with control diets based on unweighted analysis (considering only absence and presence of taxa), but did not alter the overall composition of the colonizing community based on weighted UniFrac distances; unchanged relative abundances of major taxa included phyla Bacteroidetes, Firmicutes, and Fibrobacteres. The effect of 3-NOP on colonizing methanogenic microbes differed depending upon the forage incubated, as abundance of genus Methanobrevibacter was decreased for barley silage but not for grass hay. In conclusion, 3-NOP supplementation of a high-forage diet decreased ruminal methanogenesis and increased dissolved H2 concentration, but had no negative effects on ruminal fiber degradation and only minor effects on relative abundances of the major taxa of bacteria adhered to forage substrates incubated in the rumen.

Variations in methane yield and microbial community profiles in the rumen of dairy cows as they pass through stages of first lactation.

Considerable interest exists both from an environmental and economic perspective in reducing methane emissions from agriculture. In ruminants, CH4 is produced by a complex community of microorganisms that is established in early life but can be influenced by external factors such as feed. Although CH4 emissions were thought to be constant once an animal reached maturity, recent studies have shown that CH4 yield significantly increases from early to late lactation in dairy cows. The aim of this study was to test the hypothesis that increases in CH4 yield over the lactation cycle are related to changes in rumen microbial community structure. Nine cows were monitored throughout their first lactation cycle. Methane and dry matter intake were measured to calculate CH4 per dry matter intake (CH4 yield) and ruminal fluid was collected during early, mid, and late lactation. A significant difference in bacterial and archaeal community structure during early and late lactation was observed. Furthermore, when ruminal short-chain fatty acid concentrations were measured, the ratio of acetate and butyrate to propionate was significantly higher in late lactation compared with early lactation. Propionate concentrations were higher in cows with low CH4 yield during late lactation, but no differences were observed in bacterial or archaeal community structures. Prevotella dominated the rumen of cows followed by Succinclasticum; Treponema, Fibrobacter, Ruminococcus, and Bifidobacterium were also in high abundance relative to other bacterial genera. In general, positive correlations were stronger between the most relatively abundant bacterial genera and acetate and butyrate concentrations in the cows with high CH4 and weaker between these genera and propionate concentration. This study indicates that increased CH4 yield in late lactation is reflected in significant changes in microbial community structure.

Metatranscriptomic Profiling Reveals Linkages between the Active Rumen Microbiome and Feed Efficiency in Beef Cattle.

Exploring compositional and functional characteristics of the rumen microbiome can improve the understanding of its role in rumen function and cattle feed efficiency. In this study, we applied metatranscriptomics to characterize the active rumen microbiomes of beef cattle with different feed efficiencies (efficient, n = 10; inefficient, n = 10) using total RNA sequencing. Active bacterial and archaeal compositions were estimated based on 16S rRNAs, and active microbial metabolic functions including carbohydrate-active enzymes (CAZymes) were assessed based on mRNAs from the same metatranscriptomic data sets. In total, six bacterial phyla (Proteobacteria, Firmicutes, Bacteroidetes, Spirochaetes, Cyanobacteria, and Synergistetes), eight bacterial families (Succinivibrionaceae, Prevotellaceae, Ruminococcaceae, Lachnospiraceae, Veillonellaceae, Spirochaetaceae, Dethiosulfovibrionaceae, and Mogibacteriaceae), four archaeal clades (Methanomassiliicoccales, Methanobrevibacter ruminantium, Methanobrevibacter gottschalkii, and Methanosphaera), 112 metabolic pathways, and 126 CAZymes were identified as core components of the active rumen microbiome. As determined by comparative analysis, three bacterial families (Lachnospiraceae, Lactobacillaceae, and Veillonellaceae) tended to be more abundant in low-feed-efficiency (inefficient) animals (P < 0.10), and one archaeal taxon (Methanomassiliicoccales) tended to be more abundant in high-feed-efficiency (efficient) cattle (P < 0.10). Meanwhile, 32 microbial metabolic pathways and 12 CAZymes were differentially abundant (linear discriminant analysis score of >2 with a P value of <0.05) between two groups. Among them, 30 metabolic pathways and 11 CAZymes were more abundant in the rumen of inefficient cattle, while 2 metabolic pathways and 1 CAZyme were more abundant in efficient animals. These findings suggest that the rumen microbiomes of inefficient cattle have more diverse activities than those of efficient cattle, which may be related to the host feed efficiency variation.IMPORTANCE This study applied total RNA-based metatranscriptomics and showed the linkage between the active rumen microbiome and feed efficiency (residual feed intake) in beef cattle. The data generated from the current study provide fundamental information on active rumen microbiome at both compositional and functional levels, which serve as a foundation to study rumen function and its role in cattle feed efficiency. The findings that the active rumen microbiome may contribute to variations in feed efficiency of beef cattle highlight the possibility of enhancing nutrient utilization and improve cattle feed efficiency through modification of rumen microbial functions.

Factors influencing ruminal bacterial community diversity and composition and microbial fibrolytic enzyme abundance in lactating dairy cows with a focus on the role of active dry yeast.

The objective of the current study was to employ a DNA-based sequencing technology to study the effect of active dry yeast (ADY) supplementation, diet type, and sample location within the rumen on rumen bacterial community diversity and composition, and to use an RNA-based method to study the effect of ADY supplementation on rumen microbial metabolism during high-grain feeding (HG). Our previous report demonstrated that the supplementation of lactating dairy cows with ADY attenuated the effect of subacute ruminal acidosis. Therefore, we used samples from that study, where 16 multiparous, rumen-cannulated lactating Holstein cows were randomly assigned to 1 of 2 dietary treatments: ADY (Saccharomyces cerevisiae strain Y1242, 80 billion cfu/animal per day) or control (carrier only). Cows received a high-forage diet (77:23, forage:concentrate), then were abruptly switched to HG (49:51, forage:concentrate). Rumen bacterial community diversity and structure were highly influenced by diet and sampling location (fluid, solids, epimural). The transition to HG reduced bacterial diversity, but epimural bacteria maintained a greater diversity than fluid and solids. Analysis of molecular variance indicated a significant separation due to diet × sampling location, but not due to treatment. Across all samples, the analysis yielded 6,254 nonsingleton operational taxonomic units (OTU), which were classified into several phyla: mainly Firmicutes, Bacteroidetes, Fibrobacteres, Tenericutes, and Proteobacteria. High forage and solids were dominated by OTU from Fibrobacter, whereas HG and fluid were dominated by OTU from Prevotella. Epimural samples, however, were dominated in part by Campylobacter. Active dry yeast had no effect on bacterial community diversity or structure. The phylum SR1 was more abundant in all ADY samples regardless of diet or sampling location. Furthermore, on HG, OTU2 and OTU3 (both classified into Fibrobacter succinogenes) were more abundant with ADY in fluid and solids than control samples. This increase with ADY was paralleled by a reduction in prominent Prevotella OTU. Metatranscriptomic profiling of rumen microbiome conducted on random samples from the HG phase showed that ADY increased the abundance of the cellulase endo-ß-1,4-glucanase and had a tendency to increase the hemicellulase α-glucuronidase. In conclusion, the shift from high forage to HG and sampling location had a more significant influence on ruminal bacterial community abundance and structure compared with ADY. However, evidence suggested that ADY can increase the abundance of some dominant anaerobic OTU belonging to F. succinogenes and phylum SR1. Further, microbial mRNA-based evidence suggested that ADY can increase the abundance of a specific microbial fibrolytic enzymes.

Changes in rumen microbiota composition and in situ degradation kinetics during the dry period and early lactation as affected by rate of increase of concentrate allowance.

Changes in rumen microbiota and in situ degradation kinetics were studied in 12 rumen-cannulated Holstein Friesian dairy cows during the dry period and early lactation. The effect of a rapid (RAP) or gradual (GRAD) postpartum (pp) rate of increase of concentrate allowance was also investigated. Cows were fed for ad libitum intake and had free access to a mixed ration consisting of chopped wheat straw (dry period only), grass silage, corn silage, and soybean meal. Treatment consisted of either a rapid (1.0 kg of dry matter/d; n = 6) or gradual (0.25 kg of dry matter/d; n = 6) increase of concentrate allowance (up to 10.9 kg of dry matter/d), starting at 4 d pp. In whole rumen contents, bacterial community composition was assessed using samples from 50, 30, and 10 d antepartum (ap), and 3, 9, 16, 30, 44, 60, and 80 d pp, and protozoal and archaeal community composition using samples from 10 d ap, and 16 and 44 d pp. Intake of fermentable organic matter, starch, and sugar was temporarily greater in RAP than GRAD at 16 d pp. Bacterial community richness was higher during the dry period than during the lactation. A rapid increase in concentrate allowance decreased bacterial community richness at 9 and 16 d pp compared with a gradual increase in concentrate allowance, whereas from 30 d pp onward richness of RAP and GRAD was similar. In general, the relative abundances of Bacteroidales and Aeromonadales were greater, and those of Clostridiales, Fibrobacterales, and Spirochaetales were smaller, during the lactation compared with the dry period. An interaction between treatment and sampling day was observed for some bacterial community members, and most of the protozoal and archaeal community members. Transition to lactation increased the relative abundance of Epidinium and Entodinium, but reduced the relative abundance of Ostracodinium. Archaea from genus Methanobrevibacter dominated during both the dry period and lactation. However, during lactation the abundance of the methylotrophic Methanomassiliicoccaceae and Methanosphaera increased. The in situ degradation of organic matter, neutral detergent fiber, starch, and crude protein was neither affected by treatment nor by transition from the dry period to lactation. Results show that the composition of the rumen microbiota can change quickly from the dry period to the lactation period, in particular with a rapid increase in fermentable substrate supply postpartum, but this was not associated with changes in rumen degradation kinetics.

Metagenomic assessment of the functional potential of the rumen microbiome in Holstein dairy cows.

The microbial ecology of the rumen microbiome is influenced by the diet and the physiological status of the dairy cow and can have tremendous influence on the yield and components of milk. There are significant differences in milk yields between first and subsequent lactations of dairy cows, but information on how the rumen microbiome changes as the dairy cow gets older has received little attention. We characterized the rumen microbiome of the dairy cow for phylogeny and functional pathways by lactation group and stage of lactation using a metagenomics approach. Our findings revealed that the rumen microbiome was dominated by Bacteroidetes (70%), Firmicutes (15-20%) and Proteobacteria (7%). The abundance of Firmicutes and Proteobacteria were independently influenced by diet and lactation. Bacteroidetes contributed to a majority of the metabolic functions in first lactation dairy cows while the contribution from Firmicutes and Proteobacteria increased incrementally in second and third lactation dairy cows. We found that nearly 70% of the CAZymes were oligosaccharide breaking enzymes which reflect the higher starch and fermentable sugars in the diet. The results of this study suggest that the rumen microbiome continues to evolve as the dairy cow advances in lactations and these changes may have a significant role in milk production.

Using the second law of thermodynamics for enrichment and isolation of microorganisms to produce fuel alcohols or hydrocarbons.

Fermentation of crops, waste biomass, or gases has been proposed as a means to produce desired chemicals and renewable fuels. The second law of thermodynamics has been shown to determine the net direction of metabolite flow in fermentation processes. In this article, we describe a process to isolate and direct the evolution of microorganisms that convert cellulosic biomass or gaseous CO2 and H2 to biofuels such as ethanol, 1-butanol, butane, or hexane (among others). Mathematical models of fermentation elucidated sets of conditions that thermodynamically favor synthesis of desired products. When these conditions were applied to mixed cultures from the rumen of a cow, bacteria that produced alcohols or alkanes were isolated. The examples demonstrate the first use of thermodynamic analysis to isolate bacteria and control fermentation processes for biofuel production among other uses.