Paul J. Kononoff

Evaluation of bacterial diversity in the rumen and feces of cattle fed different levels of dried distillers grains plus solubles using bacterial tag-encoded FLX amplicon pyrosequencing.

Dietary components and changes cause shifts in the gastrointestinal microbial ecology that can play a role in animal health and productivity. However, most information about the microbial populations in the gut of livestock species has not been quantitative. In the present study, we utilized a new molecular method, bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP) that can perform diversity analyses of gastrointestinal bacterial populations. In the present study, cattle (n = 6) were fed a basal feedlot diet and were subsequently randomly assigned to 1 of 3 diets (n = 2 cows per diet). In each diet, 0, 25, or 50% of the concentrate portion of the ration was replaced with dried distillers grain (DDGS). Ruminal and fecal bacterial populations were different when animals were fed DDGS compared with controls; ruminal and fecal Firmicute:Bacteroidetes ratios were smaller (P = 0.07) in the 25 and 50% DDG diets compared with controls. Ruminal pH was decreased (P < 0.05) in ruminal fluid from cattle fed diets containing 50% compared with 0% DDGS. Using bTEFAP, the normal microbiota of cattle were examined using modern molecular methods to understand how diets affect gastrointestinal ecology and the gastrointestinal contribution of the microbiome to animal health and production.

The Effect of Feeding Dried Distillers Grains Plus Solubles on Milk Production and Excretion of Urinary Purine Derivatives

Two studies were performed to evaluate the effects of dried distillers grains with solubles (DDGS) on the lactational performance of dairy cows. The intent of experiment 1 was to evaluate the effects of feeding increasing concentrations of DDGS on the feed intake and production of Holstein dairy cows. Twenty multiparous Holstein cows averaging 76 +/- 24 d in milk and 638 +/- 68 kg of body weight were randomly assigned to one of five 4 x 4 Latin squares. During each of the 28-d periods, cows were offered 1 of 4 diets: 1) control, 0% DDGS, 2) 10% DDGS, 3) 20% DDGS, or 4) 30% DDGS. For the treatment diets, DDGS replaced a portion of both forages and concentrates. Dry matter intake increased linearly with increasing concentrations of DDGS (21.4, 22.4, 23.0, and 24.0 +/- 0.98 kg/d). Similarly, milk production increased linearly (27.4, 28.5, 29.3, and 30.6 +/- 1.44 kg/d). The intent of experiment 2 was to evaluate the effect of feeding DDGS on feed intake, milk production, and excretion of urinary purine derivatives (PD). Excretion of PD was used to estimate the effects on rumen microbial crude protein production. Twenty-one multiparous and 13 primiparous Holstein cows, averaging 178 +/- 36 d in milk and 651 +/- 65 kg of body weight were randomly assigned to 1 of 2 diets in a 3-period crossover design. Cows were offered 1 of 2 rations during each 21-d period. Dietary treatments were either a control (0% DDGS) or 30% dietary dry matter of DDGS. Dry matter intake increased when feeding DDGS (22.8 vs. 24.1 +/- 0.74 kg/d for 0 and 30% DDGS, respectively) but milk production, percentages of milk fat and protein, and the ratio of PD to creatinine were not significantly different between the control and DDGS diets. Results of this study suggest a dairy ration may be formulated to contain as much as 30% of dietary dry matter as DDGS.

The effect of a leptin single nucleotide polymorphism on quality grade, yield grade, and carcass weight of beef cattle

Feedlot producers could optimize the value of cattle in a given market grid if they were able to improve the uniformity of the body composition between cattle among loads. Allelic variation due to a single nucleotide transition (cytosine [C] to thymine [T] transition that results in a Arg25Cys) has been demonstrated to be associated with higher leptin mRNA levels in adipose tissue and increased fat deposition in mature beef, but the effect on economically important carcass traits has not been investigated in either market-ready steers or heifers. Therefore, the objective of this study was to determine the effects of a leptin SNP on the quality grade (QG), yield grade (YG), and weight of beef carcasses. A slaughter trial was conducted using 1,435 crossbred finished heifers and 142 crossbred finished steers as they entered the slaughter facility. Canada QG tended (main effect of genotype P = 0.16, but P < 0.01 for both CC vs. TT and CT vs. TT) to be affected by leptin genotype. Specifically, 7.6 and 7.1% more TT carcasses graded Canada AAA or higher than the CT and CC carcasses, respectively, which supports the suggestion that the leptin SNP is associated with carcass fat. The proportion of carcasses grading Canada YG 1, 2, or 3 was affected (P < 0.01, P = 0.05, and P = 0.02 for YG 1, 2, and 3) by leptin genotype. The proportion of TT carcasses of Canada YG 1 was 12.5 and 15.1% lower than that of CT and CC carcasses, respectively, indicating that rearing animals under the same management and feeding system may result in greater carcass fat and a lower probability of the proportion of carcasses grading YG 1 within certain genotypes. The carcass weights of animals with the CC genotype tended (P = 0.07) to be higher than those of the TT genotype (365.5 vs. 362.3 kg). No significant difference was observed between the TT and CT genotypes in carcass weight. The observed associations between leptin genotype and carcass characteristics may represent an opportunity to genetically identify animals that are most likely to reach specific marketing groups.

Effects of chemical composition variation on the dynamics of ruminal fermentation and biological value of corn milling (co)products.

The objectives of this study were to evaluate the dynamics of gas production of several corn (co)products, to develop equations to predict the rate of ruminal fiber digestion, to estimate total digestible nutrients (TDN) and net energy for lactation (NE(L)), and to assess the stochasticity of chemical composition and nutritive value variability. Four corn milling (co)products were evaluated in this study: high protein dried distillers grains (HP-DDG), corn bran (BRAN) and dehydrated germ (GERM), and a dried distillers grains plus soluble produced with a low-heat drying process (BPX). Alfalfa hay was used as an internal standard feed in the in vitro fermentation dynamics analysis. Standard chemical analyses, in vitro digestibility, and in vitro gas production techniques were used to obtain the necessary physicochemical characterization of feeds. The in vitro dry matter digestibility at 24 and 48 h of incubation decreased exponentially as acid detergent insoluble nitrogen increased. However, the degree of in vitro dry matter digestibility reduction was more accentuated at 24 than at 48 h of incubation. The difference among these feeds regarding the dynamics of the anaerobic fermentation within different substrates (intact feed, and fiber and defatted residues) was evaluated. Results suggested that the proportion of fiber digested in the rumen was affected by the degree of sample processing and fat removal. Fractional fermentation rate (kf) of neutral detergent residue (without sodium sulfite) and defatted fiber residue for BRAN, GERM, HP-DDG, and BPX was estimated to be 0.0635 and 0.0852 h(-1), 0.0803 and 0.0914 h(-1), 0.118 and 0.117 h(-1), and 0.0695 and 0.0844 h(-1), respectively. The most influential variables affecting kf(NDR) of HP-DDG and BPX also affected the predicted TDN, suggesting that fiber quality is essential to ensure higher TDN values for these feeds. Our study indicated that it is possible to routinely quantify the rate of fiber digestion and this approach may be based on common analytical procedures namely estimates of neutral detergent fiber, acid detergent fiber, acid detergent insoluble nitrogen, ether extract, and acid detergent lignin. Our simulations of TDN values demonstrated that differences in fermentability and chemical composition of these corn (co)products might considerably affect the supply of energy to lactating dairy cow. The analytical methods developed in this study may serve as a valuable tool to assess nutrient quality and uniformity when samples differ in chemical composition.

Evaluation of protein fractionation and ruminal and intestinal digestibility of corn milling co-products

Novel corn milling co-products developed from technological advancements in ethanol production vary widely in chemical composition and nutrient availability. The objectives of this study were to characterize feed protein fractions and evaluate differences in rumen-undegradable protein (RUP) and its digestible fraction (dRUP), amino acid concentration, and in vitro gas production of 7 corn milling co-products. The crude protein (CP; % of dry matter) of co-products was 12.7 for germ, 26.9 for dried distillers grains plus solubles that had no heat exposure before fermentation (DDGS1), 45.4 for high-protein dried distillers grains (HPDDG), 12.7 for bran, 30.2 for wet distillers grains plus solubles (WDGS), 23.1 for wet corn gluten feed (WCGF), and 26.0 for dried distillers grains plus solubles that had heat exposure before fermentation (DDGS2). Two ruminally and duodenally fistulated Holstein steers weighing 663+/-24 kg were used to determine RUP and dRUP with the in situ and mobile bag techniques. Samples of each feed were ruminally incubated for 16 h, and mobile bags were exposed to simulated abomasal digestion before insertion into the duodenum and subsequent collection in the feces. Protein fractions A, B(1), B(2), B(3), and C were characterized as follows (% CP): germ=30.0, 15.0, 38.1, 13.5, 3.4; DDGS1=17.0, 7.0, 67.0, 4.8, 4.2; HPDDG=7.4, 0.6, 82.4, 8.8, 0.8; bran=33.5, 4.0, 54.3, 6.0, 2.2; WDGS=18.6, 2.4, 53.1, 11.0, 14.9; WCGF=36.6, 15.9, 33.2, 10.1, 4.1; and DDGS2=17.9, 2.1, 41.1, 11.1, 27.9. The proportions of RUP and dRUP were different and are reported as follows (% CP): DDGS2=56.3, 91.9; HPDDG=55.2, 97.7; WDGS=44.7, 93.1; DDGS1=33.2, 92.1; bran=20.7, 65.8; germ=16.5, 66.8; and WCGF=11.5, 51.1. The concentrations of Lys and Met in the RUP were different and are listed as follows (% CP): germ=2.9, 2.0; DDGS1=1.9, 2.0; HPDDG=2.0, 3.2; bran=3.2, 1.5; WDGS=1.9, 2.3; WCGF=3.5, 1.6; and DDGS2=1.9, 2.4. In vitro gas production (mL/48h) was highest for germ (52.1) followed by bran (50.1), WDGS (40.7), DDGS2 (40.1), WCGF (39.0), DDGS1 (38.6), and HPDDG (37.5). Comparison of co-products defined differences in chemical composition, protein fractionation, ruminal availability, and microbial fermentation.

Effects of feeding three types of corn-milling coproducts on milk production and ruminal fermentation of lactating Holstein cattle

Two experiments were conducted to determine the effects of feeding 3 corn-milling coproducts on intake, milk production, ruminal fermentation, and digestibility of lactating Holstein cows. In experiment 1, three corn-milling coproducts were fed at 15% of the diet dry matter (DM) to 28 Holstein cows averaging (+/-SD) 625 +/- 81 kg of body weight and 116 +/- 33 d in milk to determine effects on DM intake and milk production. In experiment 2, the same rations were fed to 4 ruminally fistulated, multiparous Holstein cows averaging 677 +/- 41 kg of body weight and 144 +/- 5 d in milk to determine the effects on ruminal fermentation and digestibility. In both experiments, cows and treatments were assigned randomly in 4 x 4 Latin squares over four 21-d periods. Treatments were formulated by replacing portions of forage and concentrate feeds with 15% coproduct and included 1) 0% coproduct (control), 2) dried distillers grains plus solubles (DDGS), 3) dehydrated corn germ meal (germ), and 4) high-protein dried distillers grains (HPDDG). Feed intake was recorded daily, and milk samples were collected on d 19 to 21 of each period for analysis of major components. Rumen fluid was collected at 10 time points over 24 h post feeding on d 21 of experiment 2. In experiment 1, DM intake was greater for the germ (24.3 kg/d) and DDGS treatments (23.8 kg/d), but DDGS was not different from the control (22.9 kg/d) and HPDDG treatments (22.4 kg/d). Milk production paralleled DM intake and tended to be greater for the germ (32.1 kg/d) and DDGS treatments (30.9 kg/d), but the DDGS treatment was not different from the control (30.6 kg/d) and HPDDG treatments (30.3 kg/d). However, yields of milk fat, milk protein, and 3.5% FCM were similar and averaged (+/-SEM) 1.1 +/- 0.1, 0.9 +/- 0.03, and 31.7 +/- 1.3 kg/d. Milk urea nitrogen was greater for the HPDDG (15.9 mg/dL) and germ treatments (15.5 mg/dL) than for the control (15.0 mg/dL) and DDGS treatments (14.9 mg/dL). In experiment 2, DM intake and milk production were not different across treatments and averaged 26.1 +/- 2.3 and 28.3 +/- 3.9 kg/d. Ruminal pH (6.26 +/- 0.08) and total concentration of volatile fatty acids (125.3 +/- 4.2 mM) were similar. Acetate concentration was higher for the control treatment than the DDGS, germ, and HPDDG treatments (81.7 vs. 75.8, 75.0, and 78.4 mM). Concentrations of propionate and butyrate were not different and averaged 27.8 +/- 1.2 and 14.3 +/- 0.9 mM across treatments. The acetate:propionate ratios for the control, germ, and HPDDG treatments were greater than for the DDGS treatment (3.02, 2.88, and 2.91 vs. 2.62). Dry matter, organic matter, and neutral detergent fiber digestibilities were similar across treatments and averaged 63.5 +/- 2.7, 67.3 +/- 2.2, and 43.5 +/- 4.2%. Milk production followed DM intake in experiment 1, and yield of major milk components was not affected. Results of these experiments indicate that dairy rations can be successfully formulated to include 15% of diet DM as corn-milling coproducts while maintaining or increasing DM intakes and yields of milk and milk components.

Nutrient variability for distillers grains plus solubles and dry matter determination of ethanol by-products1

ABSTRACT Three experiments were conducted to evaluate nutrient content and DM determination methods of dry milling byproducts. In Exp. 1, nutrient composition was determined for wet distillers grains plus solubles (WDGS) and modified distillers grains plus solubles (MDGS) from 6 ethanol plants with 10 samples collected per day, across 5 d, and sampling was repeated over 4 separate months. Mean composition was 31.0% CP, 11.9% fat, 0.84% P, and 0.77% S (DM basis). Coefficients of variation for DM content were greater for some plants than others, and variation occurred within and across days. Variability was small for CP and P, whereas fat differed among ethanol plants. Large variation in means and CV were observed for S in period 1, but variation subsequently decreased. Coefficients of variation for S were similar for samples collected within the same day and across days. In Exp. 2, samples of WDGS, MDGS, Dakota Bran Cake, and distillers solubles were used to determine DM content by drying samples at 105°C for 3, 8, and 24 h and 60°C for 24 and 48 h, vacuum oven drying, toluene distillation, and Karl Fischer titration. Compared with toluene distillation, drying at 105°C resulted in less DM (P ≤ 0.10) and vacuum drying and Karl Fischer titration resulted in greater DM (P

Evaluation of in vitro gas production and rumen bacterial populations fermenting corn milling (co)products

The objective of this study was to evaluate the fermentation dynamics of 2 commonly fed corn (co)products in their intact and defatted forms, using the in vitro gas production (IVGP) technique, and to investigate the shifts of the predominant rumen bacterial populations using the 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP) technique. The bTEFAP technique was used to determine the bacterial profile of each fermentation time at 24 and 48 h. Bacterial populations were identified at the species level. Species were grouped by substrate affinities (guilds) for cellulose, hemicellulose, pectin, starch, sugars, protein, lipids, and lactate. The 2 (co)products were a dried distillers grain (DDG) plus solubles produced from a low-heat drying process (BPX) and a high-protein DDG without solubles (HP). Chemical analysis revealed that BPX contained about 11.4% ether extract, whereas HP contained only 3.88%. Previous studies have indicated that processing methods, as well as fat content, of corn (co)products directly affect fermentation rate and substrate availability, but little information is available regarding changes in rumen bacterial populations. Fermentation profiles of intact and defatted BPX and HP were compared with alfalfa hay as a standard profile. Defatting before incubation had no effect on total gas production in BPX or HP, but reduced lag time and the fractional rate of fermentation of BPX by at least half, whereas there was no effect for HP. The HP feed supported a greater percentage of fibrolytic and proteolytic bacteria than did BPX. Defatting both DDG increased the fibrolytic (26.8 to 38.7%) and proteolytic (26.1 to 37.2%) bacterial guild populations and decreased the lactate-utilizing bacterial guild (3.06 to 1.44%). Information regarding the fermentation kinetics and bacterial population shifts when feeding corn (co)products may lead to more innovative processing methods that improve feed quality (e.g., deoiling) and consequently allow greater inclusion rates in dairy cow rations.

Rumen Bacterial Community Composition in Holstein and Jersey Cows Is Different under Same Dietary Condition and Is Not Affected by Sampling Method

The rumen microbial community in dairy cows plays a critical role in efficient milk production. However, there is a lack of data comparing the composition of the rumen bacterial community of the main dairy breeds. This study utilizes 16S rRNA gene sequencing to describe the rumen bacterial community composition in Holstein and Jersey cows fed the same diet by sampling the rumen microbiota via the rumen cannula (Holstein cows) or esophageal tubing (both Holstein and Jersey cows). After collection of the rumen sample via esophageal tubing, particles attached to the strainer were added to the sample to ensure representative sampling of both the liquid and solid fraction of the rumen contents. Alpha diversity metrics, Chao1 and observed OTUs estimates, displayed higher (P = 0.02) bacterial richness in Holstein compared to Jersey cows and no difference (P > 0.70) in bacterial community richness due to sampling method. The principal coordinate analysis displayed distinct clustering of bacterial communities by breed suggesting that Holstein and Jersey cows harbor different rumen bacterial communities. Family level classification of most abundant (>1%) differential OTUs displayed that OTUs from the bacterial families Lachnospiraceae and p-2534-18B5 to be predominant in Holstein cows compared to Jersey cows. Additionally, OTUs belonging to family Prevotellaceae were differentially abundant in the two breeds. Overall, the results from this study suggest that the bacterial community between Holstein and Jersey cows differ and that esophageal tubing with collection of feed particles associated with the strainer provides a representative rumen sample similar to a sample collected via the rumen cannula. Thus, in future studies esophageal tubing with addition of retained particles can be used to collect rumen samples reducing the cost of cannulation and increasing the number of animals used in microbiome investigations, thus increasing the statistical power of rumen microbial community evaluations.

In vivo determination of rumen undegradable protein of dried distillers grains with solubles and evaluation of duodenal microbial crude protein flow

The objectives of this trial were to determine the rumen undegradable protein (RUP) of dried distillers grains with solubles (DDGS), to compare the estimates of duodenal bacterial CP (BCP) flow using diaminopumelic acid (DAPA) or DNA as bacterial markers, and to estimate duodenal protozoal CP (PCP) and yeast CP (YCP) flow when DDGS are fed. Three crossbred steers fitted with ruminal and double L-shaped duodenal cannulae (average BW 780 ± 137 kg) were used in a 3 treatment, 6 period crossover design. Animals were housed in individual free stalls and fed twice daily at 0700 and 1900 h. Diets (DM basis) were 1) CONTROL, which is 0% DDGS but with 19.5% corn bran, 20% sorghum silage, 60% brome hay, 0.5% trace minerals, and 0.25% urea, 2) LOW DDGS, which is inclusion of 9.75% DDGS replacing equal percentage of corn bran, and 3) HIGH DDGS, which is inclusion of 19.5% DDGS completely replacing corn bran. Duodenal BCP flow was estimated using DAPA and DNA as bacterial markers. In addition, duodenal PCP and YCP flow were estimated using DNA markers. The value of DDGS RUP as a percent of CP was determined to be 63.0 ± 0.64%. Estimates of duodenal BCP flow using DAPA were 473, 393, and 357 ± 78 g/d (P = 0.09) for CONTROL, LOW DDGS, and HIGH DDGS, respectively. Estimates of duodenal BCP flow using DNA were 479, 397, and 368 ± 74 g/d (P = 0.14), respectively. Average BCP flow across treatments was unaffected (P = 0.71) by marker type and were 404 and 417 ± 83 g/d for DAPA and DNA markers, respectively. Estimates of duodenal PCP flow were 82, 80, and 78 ± 12 g/d (P = 0.64) for CONTROL, LOW DDGS, and HIGH DDGS, respectively. Estimates of duodenal YCP flow were 0.15, 1.94, and 4.80 ± 0.66 g/d (P < 0.01) for CONTROL, LOW DDGS, and HIGH DDGS, respectively. Duodenal BCP flow tended to decrease with DDGS inclusion, but estimates were not affected by marker type. In addition, DDGS did not affect duodenal PCP supply and provided small amounts of duodenal YCP. Overall, the value of DDGS RUP determined in this study will contribute to a better understanding of the effect of this coproduct in ruminant nutrition.