L. L. Guan

Effects of sampling location and time, and host animal on assessment of bacterial diversity and fermentation parameters in the bovine rumen

Aims:  To investigate, using culture‐independent methods, whether the ruminal bacterial structure, population and fermentation parameters differed between sampling locations and time.

Effect of dietary forage to concentrate ratio on volatile fatty acid absorption and the expression of genes related to volatile fatty acid absorption and metabolism in ruminal tissue.

The objective of the study was to investigate the fractional rate of volatile fatty acid (VFA) absorption and the expression of genes encoding for transporters and enzymes involved in the absorption and metabolism of VFA in ruminal tissue when cattle were fed high or low concentrate diets. Twelve ruminally cannulated Holstein cows were used in a randomized complete block design. The low concentrate (LC) and high concentrate (HC) diets contained 8 and 64% dietary concentrate (dry matter basis), respectively. Cows were fed their respective diet for at least 28 d, following which data and samples were collected over 6 d. Ruminal pH was measured continuously for 72 h, and the in vivo VFA absorption and passage rates were measured using Co-EDTA and n-valeric acid as markers. Ruminal tissue was collected postslaughter from the ventral sac of the rumen, and gene expression was evaluated using quantitative real-time PCR. Dry matter intake was not affected by treatment, averaging 14.9 kg/d, but cows fed HC had lower mean ruminal pH (6.03 vs. 6.48), and a greater duration (376 vs. 10 min/d) that ruminal pH was <5.8. Ruminal VFA concentration was 24 mM higher for cows fed HC compared with LC; however, the fractional rate of VFA absorption and passage from the rumen was not affected by dietary treatment, averaging 23.4 and 9.6%/h, respectively. The expression of genes encoding for enzymes involved in VFA activation and ketogenesis were not affected by treatment. Cows fed HC tended to have a relative abundance of pyruvate dehydrogenase lipoamide alpha 1 mRNA transcripts that was 1.4 times lower than that of cows fed LC, but other enzymes involved in pyruvate metabolism or regulation of the citric acid cycle were not affected. Collectively, these results suggest that the dietary forage to concentrate ratio does not affect the fractional rate of VFA absorption in vivo, but potentially alters energy metabolism in ruminal tissue.

Relationship between rumen methanogens and methane production in dairy cows fed diets supplemented with a feed enzyme additive

Aims:  To investigate the relationship between ruminal methanogen community and host enteric methane (CH4) production in lactating dairy cows fed diets supplemented with an exogenous fibrolytic enzyme additive.

A fibrolytic enzyme additive for lactating Holstein cow diets: Ruminal fermentation, rumen microbial populations, and enteric methane emissions

The objective was to determine if supplementing a dairy cow diet with an exogenous fibrolytic enzyme additive (Econase RDE; AB Vista, Marlborough, Wiltshire, UK) altered fermentation, pH, and microbial populations in the rumen or enteric methane (CH(4)) emissions. In a companion study, this enzyme additive improved efficiency of fat-corrected milk production in a dose-dependent manner by up to 11% for early lactation dairy cows. Nine ruminally cannulated, lactating Holstein cows were used in a replicated 3 × 3 Latin square design with 21-d periods. Dietary treatments were 0 (control), 0.5 (low), and 1.0 (high) mL of enzyme/kg of total mixed ration dry matter. Rumen contents were collected on 2 d (d 15 and 19), ruminal pH was measured continuously for 6 d (d 13 to 18) by using an indwelling system, and enteric CH(4) production was measured for 3 d (d 16 to 18) using the sulfur hexafluoride tracer gas technique. The enzyme additive did not alter volatile fatty acids, NH(3), pH, or population densities of total protozoa, bacteria, and methanogens in ruminal fluid. However, population densities of certain bacteria, calculated as copy number of species-specific 16S-rRNA, were affected by enzyme treatment. Population density of Ruminobacter amylophilus was increased and that of Fibrobacter succinogenes tended to be increased by the high enzyme treatment. Selenomonas ruminantium tended to increase linearly with increasing levels of enzyme in the diet, although its population density was only numerically increased by the high enzyme treatment. Streptococcus bovis, however, tended to be decreased by the low enzyme treatment. Increasing the level of enzyme supplement in the diet also linearly increased enteric CH(4) production, even when adjusted for feed intake or milk production (19.3, 20.8, and 21.7 g of CH(4)/kg of dry matter intake or 12.9, 13.6, and 15.1g of CH(4)/kg of milk for the control, low, and high enzyme treatments, respectively). This shift in ruminal bacterial communities and higher CH(4) emissions could imply increased ruminal digestion of feed, which needs to be substantiated in longer term studies.

The relationship between rumen acidosis resistance and expression of genes involved in regulation of intracellular pH and butyrate metabolism of ruminal epithelial cells in steers.

Past research has focused on the prevention and management of subacute rumen acidosis by manipulating the ration; however, the severity of acidosis varies even among animals fed a common high-grain diet. The objectives of this study were to compare the ruminal volatile fatty acid (VFA) profile and expression of genes involved in the metabolism of butyrate, the VFA most extensively metabolized by the ruminal epithelium, and intracellular pH regulation in ruminal epithelial cells between acidosis-resistant (AR) and acidosis-susceptible (AS) steers. Acidosis indexes (area per day under pH 5.8 divided by dry matter intake) were measured for 17 steers fed a common high-grain diet, and the 3 steers with the lowest (1.4 ± 1.2 pH∙min/kg) and the 3 with the highest values (23.9 ± 7.4 pH∙min/kg) were classified as AR and AS, respectively, and used in the subsequent study. The steers were force-fed a diet containing 85% grain at 60% of the expected daily intake (5.8 ± 0.8 and 5.6 ± 0.6 kg for AR and AS, respectively) within 30 min. Mean ruminal pH over the postprandial 6-h period was higher for AR compared with AS (6.02 vs. 5.55), and mean total VFA concentration was 74% for AR compared with AS (122 vs. 164 mM). Molar proportion of butyrate in the ruminal fluid was 139% higher for AR compared with AS (17.5 vs. 7.33 mol/100 mol of VFA). Expression of monocarboxylate cotransporter isoform 1, sodium hydrogen exchanger isoforms 1 and 2, and anion exchangers (downregulated in adenoma and putative anion exchanger, isoform 1) did not differ between AR and AS steers. However, expression of sodium hydrogen exchanger isoform 3, which imports Na(+) to the epithelial cell and exports H(+) to the rumen, was 176% higher in AR steers than in AS steers. Higher ruminal pH for AR might be partly due to a faster rate of VFA absorption, lower VFA production, or both.

The effects of feeding 3-nitrooxypropanol on methane emissions and productivity of Holstein cows in mid lactation.

The objective of the current study was to determine the effects of adding 3-nitrooxypropanol to the diet of lactating Holstein cows on methane emissions, rumen fermentation, ruminal microbial profile, and milk production. Twelve ruminally cannulated Holstein cows in midlactation were used in a crossover design study with 28-d periods. Cows were fed a diet containing 38% forage on a dry matter basis with either 2,500 mg/d of 3-nitrooxypropanol (fed as 25 g of 10% 3-nitrooxypropanol on silicon dioxide) or 25 g/d of silicon dioxide (control). After a 21-d diet adaptation period, dry matter intake (DMI) and milk yield were recorded daily. Rumen fluid and digesta were collected on d 22 and 28 for volatile fatty acid analysis and microbial profiling. Enteric methane emissions were measured on d 23 to 27 using the sulfur hexafluoride tracer gas technique. Feeding 3-nitrooxypropanol did not affect DMI; however, methane production was reduced from 17.8 to 7.18 g/kg of DMI. No change in milk or milk component yields was observed, but cows fed 3-nitrooxypropanol gained more body weight than control cows (1.06 vs. 0.39 kg/d). Concentrations of total volatile fatty acids in ruminal fluid were not affected by treatment, but a reduction in acetate proportion and a tendency for an increase in propionate proportion was noted. As such, a reduction in the acetate-to-propionate ratio was observed (2.02 vs. 2.36). Protozoa counts were not affected by treatment; however, a reduction in methanogen copy count number was observed when 3-nitrooxypropanol was fed (0.95 vs. 2.69 × 10(8)/g of rumen digesta). The data showed that feeding 3-nitrooxypropanol to lactating dairy cows at 2,500 mg/d can reduce methane emissions without compromising DMI or milk production.

Effects of feeding Fermenten on ruminal fermentation in lactating Holstein cows fed two dietary sugar concentrations

This study was conducted to determine the effects of feeding Fermenten (Church and Dwight Co., Princeton, NJ) with or without dietary sucrose on ruminal fermentation, apparent total-tract nutrient digestibility, and nutrient utilization. Eight ruminally cannulated Holstein cows (163 +/- 55 d in milk; mean +/- standard deviation) were used in a replicated 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments. Experimental diets were formulated with and without Fermenten (0 vs. 3.3% of dietary DM) at 2 dietary sugar concentrations (2.8 vs. 5.7%). Dietary treatment did not affect dry matter intake or apparent total-tract nutrient digestibility. Feeding Fermenten did not affect ruminal pH, but high-sugar diets tended to increase the daily minimum pH (5.61 vs. 5.42) and mean pH (6.17 vs. 6.30) compared with low-sugar diets. Ruminal ammonia concentration tended to be greater for cows fed Fermenten compared with control (18.1 vs. 15.9 mg/dL), but was not affected by dietary sugar concentration. Significant interactions between Fermenten and dietary sugar concentration were detected for some milk production responses. Fermenten treatment numerically increased milk fat yield (0.92 vs. 0.82 kg/d), 4% fat-corrected milk yield (24.3 vs. 21.9 kg/d), and milk energy output (18.2 vs. 16.4 Mcal/d) compared with control for cows fed low-sugar diets, but not for cows fed high-sugar diets. Increasing dietary sugar concentration did not enhance the effects of Fermenten, providing no support for the theory that synchronizing the availability of N and fermentable energy in the rumen improves nutrient utilization in lactating dairy cows.

The potential of 3-nitrooxypropanol to lower enteric methane emissions from beef cattle.

This study evaluated if 3-nitrooxypropanol reduces enteric methane (CH4) emissions when added to the diet of beef cattle. The effects of 3-nitrooxypropanol on related variables including diet digestibility, ruminal fermentation, and ruminal microorganisms were also investigated. Eight ruminally cannulated Angus heifers (549 ± 64.3 kg [mean BW ± SD]) were fed a high forage diet (backgrounding diet) supplemented with 4 levels of 3-nitrooxypropanol (0, 0.75, 2.25 and 4.50 mg/kg BW). The experiment was designed as a duplicated 4 × 4 Latin square with 2 groups of heifers and four 28-d periods. Methane emissions were measured during 3 consecutive days using metabolic chambers. Up to a 5.8% reduction in ad libitum DMI was observed when 2.5 mg/kg BW of 3-nitrooxypropanol was fed (P = 0.03). Increasing level of 3-nitrooxypropanol linearly (P < 0.001) reduced CH4, with 33% less CH4 (corrected for DMI) at the highest level of supplementation compared with the control. Feed energy lost as CH4 was also reduced when 3-nitrooxypropanol was supplemented (P < 0.001). Molar proportion of acetate was reduced (P < 0.001) and that for propionate increased (P < 0.001) with increasing dose of 3-nitrooxypropanol, which in turn led to a reduction in the acetate to propionate ratio (P < 0.001). Total copy numbers of 16S ribosomal RNA (rRNA) genes for bacteria, methanogens, and 18S rRNA genes for protozoa in ruminal contents were not affected by 3-nitrooxypropanol supplementation (P ≥ 0.31). There was no effect of 3-nitrooxypropanol on DM (P = 0.1) digestibility in the total tract. The use of 4.5 mg/kg BW of 3-nitrooxypropanol in beef cattle consuming a backgrounding diet was effective in reducing enteric CH4 emissions without negatively affecting diet digestibility.

Broiler egg storage induces cell death and influences embryo quality

It is well known that egg storage reduces embryo performance, but the fundamental reasons for reduced embryo quality remain unclear. The objective of this study was to investigate possible cellular and molecular mechanisms that might reduce embryo quality after egg storage. Broiler hatching eggs were obtained from the Ross 308 broiler strain, divided into 2 groups, and stored (4 and 14 d) under the same temperature and humidity conditions. Samples of the eggs were used to assess embryo quality by determining daily embryo weight (wet and dry) from 4 to 21 d of incubation. To understand possible cellular and molecular mechanisms that might affect embryo quality, blastoderms (unincubated embryos) were isolated from a sample of eggs from each storage group, dissociated into single cells, and subjected to flow cytometry analysis to differentiate between viable, apoptotic, and necrotic cell populations. Quantitative real-time PCR analysis was used to compare the expression of selected apoptotic genes (Bcl-2 homologous antagonist/killer gene, Bcl-2-associated X gene, Bcl-2-related ovarian killer gene, B-cell lymphoma 2 gene, and B-cell lymphoma xL gene) in blastoderms and embryos (6 d old after incubation). Data were analyzed by the MIXED model procedure of SAS (SAS Institute, Cary, NC), with significance set at P ≤ 0.05. After covariance analysis of initial egg weights, the results showed decreased daily embryo weights (wet and dry), an indication of decreased embryo quality that could affect hatch quality. In addition, a decrease in blastodermal cell viability was associated with an increased percentage of apoptotic cell deaths (P < 0.0001). Expression of pro-apoptotic genes (Bcl-2 homologous antagonist/killer gene, Bcl-2-associated X gene, and Bcl-2-related ovarian killer gene) were upregulated at the blastodermal level as the storage duration increased, but all genes were downregulated after 6 d of incubation. This suggests that an increase in egg storage duration could activate mechanisms of apoptotic cell death at the blastodermal level, which may be one of the molecular mechanisms that leads to reduced daily embryonic weight during incubation. Experimental controls capable of reducing the cellular and molecular mechanisms of egg storage should be used to increase embryo quality.

Transcriptome analysis of subcutaneous adipose tissues in beef cattle using 3′ digital gene expression-tag profiling 1

The molecular mechanisms that regulate fat deposition in bovine adipose tissue have not been well studied. To elucidate the genes and gene networks involved in bovine fat development, transcriptional profiles of backfat (BF) tissues from Hereford × Aberdeen Angus (HEAN, n = 6) and Charolais × Red Angus (CHRA, n = 6) steers with high or low BF thickness were characterized by digital gene expression-tag profiling. Approximately 9.8 to 21.9 million tags were obtained for each library, and a total of 18,034 genes were identified. In total, 650 genes were found to be differentially expressed, with a greater than 1.5-fold difference between the 2 crossbreds (Benjamini-Hochberg false discovery rate ≤ 0.05). The majority of differentially expressed genes that were more highly expressed in CHRA vs. HEAN were associated with development, whereas the differentially expressed genes with greater expression in HEAN vs. CHRA were overrepresented in biological processes such as metabolism and immune response. Thirty-six and 152 differentially expressed genes were detected between animals with high (n = 3) and low (n = 3) BF thickness in HEAN and CHRA, respectively (Benjamini-Hochberg false discovery rate ≤0.05). The differentially expressed genes between high and low groups in CHRA were related to cell proliferation and development processes. In addition, lipid metabolism was 1 of the top 5 molecular and cellular functions identified in both crossbreds. Ten and 17 differentially expressed genes were found to be involved in fat metabolism in HEAN and CHRA, respectively. Genes associated with obesity, such as PTX3 (pentraxin 3, long) and SERPINE1 (serpin peptidase inhibitor, clade E, member 1), were more highly expressed (P < 0.05) in the subset of CHRA animals with greater BF thickness. Our study revealed that the expression patterns of genes in BF tissues differed depending on the genetic background of the cattle.