Longitudinal shifts in bacterial diversity and fermentation pattern in the rumen of steers grazing wheat pasture.

Grazing steers on winter wheat forage is routinely practiced in the Southern Great Plains of the US. Here, we investigated the dynamics in bacterial populations of both solid and liquid ruminal fractions of steers grazing on maturing wheat forage of changing nutritive quality. The relationship between bacterial diversity and fermentation parameters in the liquid fraction was also investigated. During the first 28 days, the wheat was in a vegetative phase with a relatively high crude protein content (CP; 21%), which led to the incidence of mild cases of frothy bloat among steers. Rumen samples were collected on days 14, 28, 56 and 76, separated into solid and liquid fractions and analyzed for bacterial diversity using 16S pyrotag technology. The predominant phyla identified were Bacteroidetes (59-77%) and Firmicutes (20-33%) across both ruminal fractions. Very few differences were observed in the rumen bacterial communities within solid and liquid fractions on day 14. However, by day 28, the relatively high CP content complemented a distinct bacterial and chemical composition of the rumen fluid that was characterized by a higher ratio (4:1) of Bacteroidetes:Firmicutes and a corresponding lower acetate:propionate (3:1) ratio. Further, a greater accumulation of biofilm (mucopolysaccharide complex) on day 28 was strongly associated with the abundance of Firmicutes lineages such as Clostridium, Ruminococcus, Oscillospira and Moryella (P<0.05) in the fiber fraction. Such changes were diminished as the CP concentration declined over the course of the study. The abundance of Firmicutes was noticeable by 76 d in both fractions which signifies the development of a core microbiome associated with digestion of a more recalcitrant fiber in the mature wheat. This study demonstrates dynamics in the rumen microbiome and their association with fermentation activity in the rumen of steers during the vegetative (bloat-prone) and reproductive stages of wheat forage.

Rumen bacterial diversity dynamics associated with changing from bermudagrass hay to grazed winter wheat diets.

Rumen bacterial communities in forage-fed and grazing cattle continually adapt to a wide range of changing dietary composition, nutrient density, and environmental conditions. We hypothesized that very distinct community assemblages would develop between the fiber and liquid fractions of rumen contents in animals transitioned from bermudagrass hay diet to a grazed wheat diet. To address this hypothesis, we designed an experiment utilizing a 16S-based bTEFAP pyrosequencing technique to characterize and elucidate changes in bacterial diversity among the fiber and liquid rumen fractions and whole rumen contents of 14 (Angus x Hereford) ruminally cannulated steers sequentially fed bermudagrass hay (Cynodon dactylon; 34 days) and grazing wheat forage (28 days). Bermudagrass hay was a conserved C4 perennial grass lower in protein and higher in fiber (11% and 67%, respectively) content than grazed winter wheat (Triticum aestivum), a C3 annual grass with higher protein (20%) and a large (66%) soluble fraction.Significant differences in the OTU estimates (Chao1, Ace,and Rarefaction) were detected between fractions of both diets, with bermudagrass hay supporting greater diversity than wheat forage. Sequences were compared with a 16S database using BLASTn and assigned sequences to respective genera and genera-like units based on the similarity value to known sequences in the database. Predominant genera were Prevotella (up to 33%) and Rikenella-like (upto 28%) genera on the bermudagrass diet and Prevotella (upto 56%) genus on the wheat diet irrespective of the fractions. Principle component analyses accounted for over 95% of variation in 16S estimated bacterial community composition in all three fractions and clearly differentiated communities associated with each diet. Overall, bermudagrass hay diets clustered more clearly than wheat diets.These data are the first to explore bacterial diversity dynamics in a common population of animals in response to contrasting grass forage diets.

Metagenomic Analysis of the Rumen Microbiome of Steers with Wheat-Induced Frothy Bloat.

Frothy bloat is a serious metabolic disorder that affects stocker cattle grazing hard red winter wheat forage in the Southern Great Plains causing reduced performance, morbidity, and mortality. We hypothesize that a microbial dysbiosis develops in the rumen microbiome of stocker cattle when grazing on high quality winter wheat pasture that predisposes them to frothy bloat risk. In this study, rumen contents were harvested from six cannulated steers grazing hard red winter wheat (three with bloat score "2" and three with bloat score "0"), extracted for genomic DNA and subjected to 16S rDNA and shotgun sequencing on 454/Roche platform. Approximately 1.5 million reads were sequenced, assembled and assigned for phylogenetic and functional annotations. Bacteria predominated up to 84% of the sequences while archaea contributed to nearly 5% of the sequences. The abundance of archaea was higher in bloated animals (P < 0.05) and dominated by Methanobrevibacter. Predominant bacterial phyla were Firmicutes (65%), Actinobacteria (13%), Bacteroidetes (10%), and Proteobacteria (6%) across all samples. Genera from Firmicutes such as Clostridium, Eubacterium, and Butyrivibrio increased (P < 0.05) while Prevotella from Bacteroidetes decreased in bloated samples. Co-occurrence analysis revealed syntrophic associations between bacteria and archaea in non-bloated samples, however; such interactions faded in bloated samples. Functional annotations of assembled reads to Subsystems database revealed the abundance of several metabolic pathways, with carbohydrate and protein metabolism well represented. Assignment of contigs to CaZy database revealed a greater diversity of Glycosyl Hydrolases dominated by oligosaccharide breaking enzymes (>70%) in non-bloated samples. However, the abundance and diversity of CaZymes were greatly reduced in bloated samples indicating the disruption of carbohydrate metabolism. We conclude that mild to moderate frothy bloat results from tradeoffs both within and between microbial domains due to greater competition for substrates that are of limited availability as a result of biofilm formation.

Effects of condensed tannins supplementation level on weight gain and in vitro and in vivo bloat precursors in steers grazing winter wheat.

Research was conducted to determine the effects of level of supplementation with quebracho condensed tannins (CT) on in vitro ruminal fluid gas production, in vivo ruminal fluid protein fractions, bloat dynamics, and ADG of steers grazing winter wheat. Two experiments were conducted to 1) enumerate the effect of ruminal fluid from steers fed quebracho CT (0, 1, and 2% CT/kg of DMI) on in vitro gas and methane production from minced fresh wheat forage; and 2) quantify the influence of CT supplementation on ruminal protein characteristics, biofilm complexes, bloat potential, and ADG of steers grazing wheat pasture. Eighteen ruminally cannulated steers (386 +/- 36 kg of BW) were randomly allocated to 1 of 3 treatments that included a control (water infusion) and 2 CT treatment levels (1 or 2% CT/kg of DMI). Treatments were administered daily (63 d) through the rumen cannula as pre-mixes with warm water (approximately 30 degrees C). Rumen contents were collected 2 h postinfusion (at 1030 to 1130) on d 0, 20, 40, 50, and 60. Bloat was visually scored daily for 5 d each wk. In Exp. 1, supplementation of CT decreased the rate of in vitro gas production in a dose-dependent response. In Exp. 2, ADG increased (P < 0.04) at both levels of CT supplementation. Mean bloat score across stage of growth and replicates decreased linearly with increasing CT supplementation; bloat scores were greater (P < 0.001) for the vegetative than for the reproductive stage of plant growth. Biofilm production and rumen fluid protein fractions varied among CT treatments and stage of growth. Addition of CT reduced the severity of bloat, principally through reducing microbial activities, biofilm production, and ruminal gas production. Quebracho CT is potentially a value-added supplement that can decrease the impacts of frothy bloat and increase BW gains in stocker cattle-wheat systems.

Wheat pasture bloat dynamics, in vitro ruminal gas production, and potential bloat mitigation with condensed tannins.

The aim of this study was to determine the effect of winter wheat (Triticum aestivum L.) forage growth stage, forage allowance, time of day, and commercial condensed tannins (CT) on steer bloat dynamics and in vitro ruminal gas production. Twenty-six crossbreed steers (Angus x Hereford x Salers; average initial BW = 194 +/- 26 kg) were used. Wheat forage allowances were either 18 kg (high forage allowance) and 6 kg (low forage allowance) of DM/(100 kg BW.d). In each bloat observation period, fresh wheat forage samples were hand-clipped to ground level in all study pastures for nutrient and in vitro ruminal gas production analyses. In vitro ruminal gas accumulation was measured at 0, 1, 2, 3, 4, 5, 6, and 12 h. Commercial CT was added at 0, 10, 15, and 20 mg of CT/g of DM. Bloat was scored once per week on two consecutive days at 0800 and 1500 during the vegetative stage and once every 2 wk during the reproductive stage of wheat development. Mean bloat score was calculated for each steer by time of day, stage of plant growth, and forage allowance. Bloat was detected in 65.8% of the observation periods. Average bloat scores were four and 2.5 times greater (P < 0.05) in cattle grazing at a high forage allowance than at a low forage allowance in the vegetative and reproductive growth phases of wheat, respectively. Rate of gas production was greater (P < 0.001) in the vegetative stage than in the reproductive stage. Steer bloat score was positively correlated with forage CP (r = 0.22; P < 0.05) and IVDMD (r = 0.32; P < 0.05). Rate of ruminal gas production was positively correlated (P < 0.01) to forage CP (r = 0.48), NPN (r = 0.40), soluble protein (r = 0.32), and IVDMD (r = 0.47). Conversely, negative correlations were found for forage DM (r = -0.20; P < 0.05), insoluble protein (r = -0.40), NDF (r = -0.69), and forage height (r = -0.49; P < 0.01) on the rate of ruminal gas production. Addition of CT at levels greater than 10 mg of CT/g of DM decreased (P < 0.05) the rate of in vitro ruminal gas and methane gas production after 5 h of incubation. Wheat pasture bloat is a complex disorder that varies across an array of forage and environmental conditions. Condensed tannins have the potential to decrease bloat by altering ruminal gas production and soluble protein digestibility from wheat forage.