T. A. Wickersham

High-throughput Methods Redefine the Rumen Microbiome and Its Relationship with Nutrition and Metabolism

Diversity in the forestomach microbiome is one of the key features of ruminant animals. The diverse microbial community adapts to a wide array of dietary feedstuffs and management strategies. Understanding rumen microbiome composition, adaptation, and function has global implications ranging from climatology to applied animal production. Classical knowledge of rumen microbiology was based on anaerobic, culture-dependent methods. Next-generation sequencing and other molecular techniques have uncovered novel features of the rumen microbiome. For instance, pyrosequencing of the 16S ribosomal RNA gene has revealed the taxonomic identity of bacteria and archaea to the genus level, and when complemented with barcoding adds multiple samples to a single run. Whole genome shotgun sequencing generates true metagenomic sequences to predict the functional capability of a microbiome, and can also be used to construct genomes of isolated organisms. Integration of high-throughput data describing the rumen microbiome with classic fermentation and animal performance parameters has produced meaningful advances and opened additional areas for study. In this review, we highlight recent studies of the rumen microbiome in the context of cattle production focusing on nutrition, rumen development, animal efficiency, and microbial function.

Effect of increasing amounts of postextraction algal residue on straw utilization in steers

Algal biomass has been identified as a third-generation biofuel. Significant quantities of the coproduct postextraction algal residue (PEAR) remain after lipid extraction. After extraction, PEAR is concentrated in protein (17.9% CP on a DM basis and 32.5% CP on an ash-free basis), suggesting it may be an alternative to cottonseed meal (CSM) as a protein supplement. Our objectives were to determine the optimal level of PEAR supplementation to steers consuming straw and to compare the effects of PEAR supplementation on straw utilization and N metabolism with an isonitrogenous level of CSM. Five steers (198.2 ± 6.1 kg of BW), in a 5 × 5 Latin square, had ad libitum access to oat straw (80% NDF and 4.5% CP on a DM basis). Treatments were infused ruminally once daily and included no supplemental protein (CON); PEAR at 50, 100, and 150 mg N/kg BW; and CSM at 100 mg N/kg BW. Provision of PEAR increased total digestible OM intake (TDOMI) quadratically (P = 0.01) from 0.9 (CON) to 1.6 kg/d (100 mg N/kg BW of PEAR). Organic matter digestibility (OMD) increased quadratically (P < 0.01) with supplementation and was maximized (55% OMD) at 50 mg N/kg BW of PEAR. At isonitrogenous levels of PEAR and CSM, TDOMI was similar (P = 0.13) as was OMD (P = 0.50). Negative N balance was observed for all treatments except PEAR provided at 100 or 150 mg of N/kg BW. Nitrogen balance was quadratic (P < 0.01) with the greatest retention (1.84 g N/d) occurring at 100 mg N/kg BW of PEAR. There were no differences (P ≥ 0.22) between isonitrogenous PEAR and CSM supplementation in measurements of ruminal ammonia or VFA concentrations. Straw utilization was maximized when PEAR was provided at 100 mg N/kg BW. Our observations suggest cattle provided PEAR utilize straw in a manner similar to those supplemented CSM, indicating PEAR has potential to substitute for CSM as a protein supplement in forage-based operations.

Effect of postextraction algal residue supplementation on the ruminal microbiome of steers consuming low-quality forage

Cattle consuming low-quality forages (LQF) require protein supplementation to increase forage utilization via ruminal fermentation. Biofuel production from algal biomass results in large quantities of postextraction algal residue (PEAR), which has the potential to elicit LQF utilization responses similar to cottonseed meal (CSM); however, its effect on ruminal bacterial communities is unknown. Five ruminally and duodenally cannulated Angus steers in a 5 × 5 Latin square had ad libitum access to oat straw diets. Treatments were infused ruminally and consisted of an unsupplemented control; PEAR at 50, 100, and 150 mg N/kg BW; and CSM at 100 mg N/kg BW. Ruminal samples were collected 4 h after supplementation on d 14 of each period and separated into solid and liquid fractions. Each sample was extracted for genomic DNA, PCR amplified for the V4 to V6 region of the 16S rRNA, sequenced on the 454 Roche pyrosequencing platform, and analyzed using the QIIME pipeline. Weighted UniFrac analysis and Morisita-Horn index demonstrated different community composition between liquid and solid fractions. Measures of richness including observed operational taxonomic units (OTU) and abundance coverage estimator metric decreased with greater PEAR provision (P ≤ 0.09). There were 42 core microbiome OTU observed in all solid fraction samples while the liquid fraction samples contained 30 core OTU. Bacteroidetes was the predominant phylum followed by Firmicutes in both fractions, which together characterized more than 90% of sequences. Relative abundance of Firmicutes increased with PEAR supplementation in the liquid fraction (linear, P = 0.02). Among Firmicutes, Lachnospiraceae, Ruminococcaceae, and Clostridiaceae families increased in the liquid fraction with greater PEAR supplementation (linear, P ≤ 0.03). Prevotella represented over 25% of sequences in all treatments, and relative abundance decreased in the solid fraction with increasing PEAR provision (linear, P = 0.01). Fibrobacter and Treponema decreased in the liquid fraction with increasing PEAR (linear, P < 0.10). Results suggest PEAR supplementation increased forage utilization by increasing members of Firmicutes within the liquid fraction of the rumen microbiome.

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.

Effects of a slow-release urea product on performance, carcass characteristics, and nitrogen balance of steers fed steam-flaked corn

Two experiments were conducted to examine the impact of source, urea (U) or Optigen II (OP), and level of dietary NPN on performance (Exp.1) and N balance (Exp. 2) of growing cattle. Sixty Angus crossbred steers (initial BW = 353 ± 13.9 kg) were used to evaluate performance, and fed 1 of 3 steam-flaked corn based diets: U (U(1.2), 1.2% NPN), OP (OP(1.3), 1.3% NPN), or OP without cottonseed meal (OP(3.1), 3.1% NPN). U(1.2)and OP(1.3) contained cottonseed meal and NPN as CP sources, whereas OP(3.1) contained only NPN. For Exp. 1, steers were blocked by postweaning BW and assigned to treatment (TRT) and pen within block. Body weight was collected every 14 d during the 105-d trial. Six steers from each TRT were selected based on residual feed intake (RFI) and harvested. Carcass and organ measurements were obtained. Cumulative animal performance was evaluated in 3 periods (0 to 35, 0 to 70, and 0 to 105 d) using a mixed coefficient model with initial BW as a covariate. For Exp. 2, 5 ruminally cannulated Holstein steers in a 5 × 5 Latin square design were used to evaluate N balance. Steers were fed a steam-flaked corn based diet with either no NPN, 0.75% U or N equivalent OP, or 1.5% U or N equivalent OP. Intake was measured, and feed, orts, urine, and fecal samples were obtained and composited for each steer by period. Data were analyzed using a mixed coefficient model. Orthogonal contrasts were used to evaluate differences between OP and U, and high and low level of NPN. For Exp. 1, there were no differences (P > 0.10) in initial BW, final BW, ADG, or DMI among TRT for any of the periods. However, for period 1 steers on OP(3.1) had reduced F:G than U(1.2) (5.71 kg/kg vs. 7.39 kg/kg; P = 0.03), and steers fed OP(1.3) tended to have less F:G than those fed U(1.2) (6.07 kg/kg vs. 7.39 kg/kg; P = 0.07). In period 2, OP(3.1) had reduced F:G than U(1.2) (5.58 kg/kg vs. 6.56 kg/kg; P = 0.03), but did not differ from OP(1.3) (5.97). For Exp. 2, steers fed OP tended (P = 0.09) to have less apparent N absorption than those fed U. Apparent N absorption differed (P < 0.05) for level of NPN. There were no differences (P > 0.10) in intake or digestibility among source or level of NPN. No major differences (P > 0.10) on performance and carcass composition were observed between U and OP diets. Steers had better initial F:G (Period 1 and 2) when OP was used as the only source of feed N (OP(3.1)), suggesting that OP may replace true protein feeds in finishing cattle diets, minimizing feed use with no impact on carcass quality.

Influence of dietary methionine concentration on growth and nitrogen balance in weanling Quarter Horses

Twenty-four Quarter Horse weanlings (120 ± 10 d) were blocked by age into 4 groups (n=6) for a 56-d trial to evaluate the influence of dietary Met concentration on growth and N balance. Weanling horses were housed by block and individually fed concentrates twice daily at 1.75% BW (as-fed basis). Weanling horses were randomly assigned to 1 of 4 concentrate treatments: basal (0.20 Met), basal + 0.03% Met (0.23 Met), basal + 0.07% Met (0.27 Met), and basal + 0.11% Met (0.31 Met). Diets were formulated to be isonitrogenous and isocaloric and contained equal amounts of Lys and Thr. Coastal bermudagrass hay (Cynodon dactylon) was individually fed at 0.75% BW (as-fed basis). Growth measurements, BW, rump fat, and plasma were obtained every 7 d. The final 4 d consisted of total collection of urine and feces. Feed, fecal, and urine samples were analyzed for N content, and N balance was calculated. Urine was analyzed for urea and ammonia concentrations. Plasma was analyzed for urea concentration. Grain, hay, and fecal samples were analyzed for nutrient composition. Data were analyzed using the PROC MIXED procedure of SAS. Linear, quadratic, and cubic effects were tested in the form of contrasts. There was no influence (P>0.25) of treatment on growth measurements, N balance, or urinary urea or ammonia. Intake of Lys and Thr did not differ (P>0.08) among treatments. Methionine intake increased as expected with increasing Met inclusion in the diet (P<0.01). Plasma urea N responded quadratically (P=0.04) to treatment. An unexpected peak in PUN was observed with 0.27 Met. The results indicate future studies that can ensure Met as the only dietary variable are necessary to better explain the Met requirements of weanling horses.

Estimation of Rhizome Composition and Overwintering Ability in Perennial Sorghum spp. Using Near-Infrared Spectroscopy (NIRS)

Temperately adapted perennial sorghum feedstocks have recently begun to receive increasing interest as candidate energy crops, producing significant biomass and contributing agroecological benefits including increased soil organic carbon, reduced soil erosion, reduced input requirements, and higher net energy return. Rhizomes are the primary morphological feature facilitating overwintering in Sorghum species; however, underlying physiological mechanisms governing rhizome overwintering remain poorly characterized. In this study, we investigated the composition of sorghum rhizomes from diverse germplasm before and after overwintering at two locations and three experimental environments. Significant positive correlations were found between rhizome overwintering and water-soluble carbohydrates, ethanol soluble carbohydrates, and fructan concentrations, while significant negative correlations were found between rhizome overwintering and both crude fat and starch. Near-infrared spectroscopy (NIRS) calibration equations were developed to quickly and efficiently predict the concentrations of each of these assimilates in rhizomes.

The influence of taste in willingness-to-pay valuations of sirloin steaks from postextraction algal residue–fed cattle

Consumer preferences and willingness-to-pay (WTP) for beef sirloin steaks with differing production, physical, and credence attributes related to the use of postextraction algal residue (PEAR), a novel feed ingredient, were estimated. Ninety-six consumers participated in a sensory tasting panel before completing a choice set survey; 127 consumers completed only the choice set survey without sampling products. Steaks from grain- and PEAR-fed steers had similar Warner-Bratzler shear force (WBSF) scores (1.89 kg and 2.01 kg, respectively; = 0.77) and had lower WBSF scores than steaks from grass-fed steers (3.37 kg; < 0.05). Eicosapentaenoic acid (20:5) was not different among steaks from grain- and PEAR-fed steers ( = 0.39) but was greater compared with steaks from grass-fed cattle ( ≤ 0.03). Panelists in the sensory portion of the study evaluated beef samples for like/dislike of overall sample, overall flavor, beefy flavor, and juiciness. Panelist rating of overall like, overall flavor like, and beefy flavor like were not different between the PEAR- and grain-fed treatments ( > 0.26). Panelists rated the juiciness like/dislike of steaks from PEAR-fed cattle the highest ( < 0.01) among the 3 samples. Sensory tasting of the products was observed to alter the preferences of consumers. Consumers who completed only the survey negatively perceived beef from PEAR-fed cattle compared with beef from grain-fed cattle, with a WTP discount of -US

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1.17/kg. However, with sensory tasting, the WTP for beef from PEAR-fed cattle was not discounted relative to beef from grain-fed cattle ( = 0.21). The nontasting consumers had much higher stated WTP values for credence attributes. Factors that influence the eating experience (tenderness and quality grade) dominated as the most important attributes on WTP among the tasting group. The use of no hormones and no antibiotics in production had a premium of

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2.34/kg among the nontasting group, but with tasting, the premium was