Factors that affect heat production in lactating Jersey cows.

Heat production (HP) represents a major energy cost in lactating dairy cows. Better understanding of factors that affect HP will improve our understanding of energy metabolism. Our objective was to derive models to explain variation in HP of lactating Jersey cows. Individual animal-period data from 9 studies (n = 293) were used. The data set included cows with a wide range (min to max) in days in milk (44-410) and milk yield (7.8-43.0 kg/d). Diets included corn silage as the predominate forage source, but diets varied (min to max on DM basis) in crude protein (CP; 15.2-19.5%), neutral detergent fiber (NDF; 35.5-43.0%), starch (16.2-31.1%), and crude fat (2.2 to 6.4%) contents. Average HP was (mean ± standard deviation) 22.1 ± 2.86 Mcal/d, or 28.1 ± 3.70% of gross energy intake. Eight models were fit to explain variation in HP: (1) dry matter intake (DMI; INT); (2) milk fat, protein, and lactose yield (MILKCOMP); (3) INT and milk yield (INT+MY); (4) INT and MILKCOMP/DMI (INT+MILKCOMP); (5) mass of digested NDF, CP, and starch (DIG); (6) INT and digested energy (INT+DE); (7) INT and NDF, CP, and starch digestibility (INT+DIG); or (8) INT+MILKCOMP model plus urinary N excretion (INT+MILKCOMP+UN). For all HP models, metabolic body weight was included. All models were derived via a backward elimination approach and included the random effects of study, cow, and period within block within study. The INT models adequately explained variation in HP with a nonrandom effect-adjusted concordance correlation coefficient of 0.84. Similar adjusted concordance correlation coefficients (0.79-0.85) were observed for other HP models. The HP associated with milk protein yield and supply of digestible protein was greater than other milk production and nutrient digestibility variables. The HP associated with urinary N excretion was 5.32. Overall, HP can be adequately predicted from metabolic body weight and DMI. Milk component yield, nutrient digestibility, or urinary N excretion explained similar variation as DMI. Coefficients for milk protein and protein digestion suggest that digestion and metabolism of protein and synthesis of milk protein contribute substantially to HP of a dairy cow.

Development of feed composition tables using a statistical screening procedure.

Millions of feed composition records generated annually by testing laboratories are valuable assets that can be used to benefit the animal nutrition community. However, it is challenging to manage, handle, and process feed composition data that originate from multiple sources, lack standardized feed names, and contain outliers. Efficient methods that consolidate and screen such data are needed to develop feed composition databases with accurate means and standard deviations (SD). Considering the interest of the animal science community in data management and the importance of feed composition tables for the animal industry, the objective was to develop a set of procedures to construct accurate feed composition tables from large data sets. A published statistical procedure, designed to screen feed composition data, was employed, modified, and programmed to operate using Python and SAS. The 2.76 million data received from 4 commercial feed testing laboratories were used to develop procedures and to construct tables summarizing feed composition. Briefly, feed names and nutrients across laboratories were standardized, and erroneous and duplicated records were removed. Histogram, univariate, and principal component analyses were used to identify and remove outliers having key nutrients outside of the mean ± 3.5 SD. Clustering procedures identified subgroups of feeds within a large data set. Aside from the clustering step that was programmed in Python to automatically execute in SAS, all steps were programmed and automatically conducted using Python followed by a manual evaluation of the resulting mean Pearson correlation matrices of clusters. The input data set contained 42, 94, 162, and 270 feeds from 4 laboratories and comprised 25 to 30 nutrients. The final database included 174 feeds and 1.48 million records. The developed procedures effectively classified by-products (e.g., distillers grains and solubles as low or high fat), forages (e.g., legume or grass-legume mixture by maturity), and oilseeds versus meal (e.g., soybeans as whole raw seeds vs. soybean meal expellers or solvent extracted) into distinct sub-populations. Results from these analyses suggest that the procedure can provide a robust tool to construct and update large feed data sets. This approach can also be used by commercial laboratories, feed manufacturers, animal producers, and other professionals to process feed composition data sets and update feed libraries.

Increasing the concentration of linolenic acid in diets fed to Jersey cows in late lactation does not affect methane production.

Although the inclusion of fat has reduced methane production in ruminants, relatively little research has been conducted comparing the effects of source and profile of fatty acids on methane production in lactating dairy cows. A study using 8 multiparous (325 ± 17 DIM; mean ± SD) lactating Jersey cows was conducted to determine effects of feeding canola meal and lard versus extruded byproduct containing flaxseed as a high-C18:3 fat source on methane production and diet digestibility in late-lactation dairy cows. A crossover design with 32-d periods (28-d adaptation and 4-d collections) was used to compare 2 different fat sources. Diets contained approximately 50% forage mixture of corn silage, alfalfa hay, and brome hay; the concentrate mixture changed between diets to include either (1) a conventional diet of corn, soybean meal, and canola meal with lard (control) or (2) a conventional diet of corn and soybean meal with an extruded byproduct containing flaxseed (EXF) as the fat source. Diets were balanced to decrease corn, lard, and canola meal and replace them with soybean mean and EXF to increase the concentration of C18:3 (0.14 vs. 1.20% of DM). Methane production was measured using headbox-style indirect calorimeters. Cattle were restricted to 95% ad libitum feed intake during collections. Milk production (17.4 ± 1.04 kg/d) and dry matter intake (15.4 ± 0.71 kg/d) were similar among treatments. Milk fat (5.88 ± 0.25%) and protein (4.08 ± 0.14%) were not affected by treatment. For methane production, no difference was observed for total production (352.0 vs. 349.8 ± 16.43 L/d for control vs. EXF, respectively). Methane production per unit of dry matter intake was not affected and averaged 23.1 ± 0.57 L/kg. Similarly, methane production per unit of energy-corrected milk was not affected by fat source and averaged 15.5 ± 0.68 L/kg. Heat production was similar, averaging 21.1 ± 1.02 Mcal/d. Digestibility of organic matter, neutral detergent fiber, and crude protein was not affected by diet and averaged 69.9, 53.6, and 73.3%, respectively. Results indicated that increasing C18:3 may not affect methane production or digestibility of the diet in lactating dairy cows.

Reducing methane production with corn oil and calcium sulfate: Responses on whole-animal energy and nitrogen balance in dairy cattle.

The addition of fat and calcium sulfate to diets fed to ruminants has resulted in a reduction in methane production, but the effects on energy balance have not been studied. A study using indirect calorimetry and 16 multiparous (8 Holstein and 8 Jersey; 78 ± 15 d in milk; mean ± standard deviation) lactating dairy cows was conducted to determine how mitigating methane production by adding corn oil or calcium sulfate to diets containing reduced-fat distillers grains affects energy and nitrogen balance. A replicated 4 × 4 Latin square design with 35-d periods (28 d of adaption and 4 d of collections) was used to compare 4 different dietary treatments. Treatments were composed of a control (CON) diet, which did not contain reduced-fat distillers grain and solubles (DDGS), and treatment diets containing 20% (dry matter basis) DDGS (DG), 20% DDGS with 1.38% (dry matter basis) added corn oil (CO), and 20% DDGS with 0.93% (dry matter basis) added calcium sulfate (CaS). Compared with CON, dry matter intake was not affected by treatment, averaging 29.6 ± 0.67 kg/d. Milk production was increased for diets containing DDGS compared with CON (26.3 vs. 27.8 ± 0.47 kg/d for CON vs. DDGS, respectively), likely supported by increased energy intake. Compared with CON, energy-corrected milk was greater in DG and CO (30.1 vs. 31.4, 31.7, and 31.0 ± 0.67 kg/d for CON, DG, CO, and CaS, respectively). Compared with CON, the addition of calcium sulfate and corn oil to diets containing DDGS reduced methane production per kg of dry matter intake (22.3, 19.9, and 19.6 ± 0.75 L/kg per d for CON, CO, and CaS, respectively). Similarly, methane production per kilogram of energy-corrected milk was reduced with the addition of calcium sulfate and corn oil to diets containing DDGS (14.2, 12.5, and 12.4 ± 0.50 L/kg per d for CON, CO, and CaS, respectively). Compared with CON and CaS, the intake of digestible energy was greater for DG and CO treatments (57.7, 62.1, 62.0, and 59.0 ± 1.38 Mcal/d for CON, DG, CO, and CaS, respectively). Intake of metabolizable energy was greater in all treatments containing DDGS compared with CON (50.5 vs. 54.0 ± 1.08 Mcal/d for CON vs. DDGS, respectively). Net balance (milk plus tissue energy) per unit of dry matter was greater in CO (containing DDGS and oil) than CON (1.55 vs. 1.35 ± 0.06 Mcal/kg for CO vs. CON, respectively). Tissue energy was greater in DG and CO compared with CON (6.08, 7.04, and 3.16 ± 0.99 Mcal/d for DG, CO, and CON, respectively. Results of this study suggest that the addition of oil and calcium sulfate to diets containing DDGS may be a viable option to reduce methane production and in the case of oil also improve net energy balance in lactating dairy cows.

Energy balance and diurnal variation in methane production as affected by feeding frequency in Jersey cows in late lactation.

Methane (CH4) production of ruminants typically increases with increased dry matter intake (DMI). However, few studies have observed the effects of feeding multiple times a day and its effects on diurnal variation in CH4 production and energy balance in late-lactation dairy cattle. A study using headbox-style indirect calorimetry and 12 multiparous (225 ± 16.2 d in milk; mean ± SD) lactating Jersey cows was conducted to determine the effects of feeding twice daily on diurnal variation in CH4 production and total energy balance. A crossover design with 14-d periods (10 d of adaption and 4 d of collection) was used to compare 2 treatments. Treatments consisted of either once a day feeding (1×; 100% of feed given at 1000 h) or twice a day feeding (2×; 50% of feed given at 1000 h and the final 50% at 2000 h) with a common diet fed in both treatments. Dry matter intake was not different between treatments, with a mean of 16.9 ± 0.88 kg/d. Once a day feeding tended to have greater milk yield compared with twice a day feeding (21.2 vs. 20.4 ± 1.59 kg/d, respectively). Milk fat and milk protein percentage were not different, with means of 6.18 ± 0.20% and 3.98 ± 0.08%, respectively. Total CH4 production did not differ between treatments, with a mean of 402.1 ± 20.8 L/d. Similarly, CH4 per unit of milk yield and DMI was not different between treatments, with means of 20.5 ± 1.81 and 23.8 ± 1.21 L/kg, respectively. Feeding frequency did not affect diurnal variation of hourly CH4 production, with a mean of 17.1 ± 0.74 L/h. A trend was observed for a treatment × hour interaction. Methane production per hour increased after the second feeding for cattle fed twice versus once daily. Gross energy, digestible energy, metabolizable energy, and balance (milk plus tissue) per kilogram of DMI did not differ by feeding frequency, with means of 4.41 ± 0.01, 3.05 ± 0.03, 2.63 ± 0.03, and 1.32 ± 0.08 Mcal/kg of DM, respectively. Metabolizable energy for maintenance was 146 kcal/kg of metabolic body weight, with an efficiency of converting metabolizable energy to net energy balance (milk plus tissue) of 76%. Nitrogen balance did not differ among treatments, with a mean balance of 17.3 ± 13.0 g/d. Therefore, total CH4 production and energy maintenance were not affected by feeding frequency. However, CH4 was variable throughout the day, and caution should be exercised when collecting CH4 samples at a limited number of time points because this may under- or overestimate total production.

The influence of fat and hemicellulose on methane production and energy utilization in lactating Jersey cattle.

Feeding fat to lactating dairy cows may reduce methane production. Relative to cellulose, fermentation of hemicellulose is believed to result in less methane; however, these factors have not been studied simultaneously. Eight multiparous, lactating Jersey cows averaging (±SD) 98 ± 30.8 d in milk and body weight of 439.3 ± 56.7 kg were used in a twice-replicated 4 × 4 Latin square to determine the effects of fat and hemicellulose on energy utilization and methane production using a headbox-type indirect calorimetry method. To manipulate the concentration of fat, porcine tallow was included at either 0 or 2% of the diet dry matter. The concentration of hemicellulose was adjusted by manipulating the inclusion rate of corn silage, alfalfa hay, and soybean hulls resulting in either 11.3 or 12.7% hemicellulose (dry matter basis). The resulting factorial arrangement of treatments were low fat low hemicellulose (LFLH), low fat high hemicellulose (LFHH), high fat low hemicellulose (HFLH), and high fat high hemicellulose (HFHH). Neither fat nor hemicellulose affected dry matter intake, averaging 16.2 ± 1.18 kg/d across treatments. Likewise, treatments did not affect milk production, averaging 23.0 ± 1.72 kg/d, or energy-corrected milk, averaging 30.1 ± 2.41 kg/d. The inclusion of fat tended to reduce methane produced per kilogram of dry matter intake from 24.9 to 23.1 ± 1.59 L/kg, whereas hemicellulose had no effect. Increasing hemicellulose increased neutral detergent fiber (NDF) digestibility from 43.0 to 51.1 ± 2.35%. Similarly, increasing hemicellulose concentration increased total intake of digestible NDF from 6.62 to 8.42 ± 0.89 kg/d, whereas fat had no effect. Methane per unit of digested NDF tended to decrease from 64.8 to 49.2 ± 9.60 L/kg with increasing hemicellulose, whereas fat had no effect. An interaction between hemicellulose and fat content on net energy balance (milk plus tissue energy) was observed. Specifically, increasing hemicellulose in low-fat diets tended to increase net energy balance, but this was not observed in high-fat diets. These results confirm that methane production may be reduced with the inclusion of fat, whereas energy utilization of lactating dairy cows is improved by increasing hemicellulose in low-fat diets.

Energy content of reduced-fat dried distillers grains with solubles for lactating dairy cows.

Eight Holstein and 8 Jersey multiparous, lactating cows were used to complete 56 energy balances to determine the energy content of reduced-fat dried distillers grains with solubles (RFDDGS). A repeated switchback design was used to compare treatments with and without RFDDGS. Diets consisted of 24.2% corn silage, 18.4% alfalfa hay, 6.94% brome hay with either 22.9% rolled corn or 14.8% soybean meal (control), or 8.95% rolled corn, 28.8% RFDDGS, and 0% soybean meal [Co-P; dry-matter (DM) basis]. The inclusion of RFDDGS did not affect DM intake, averaging 21.4 ± 0.53 kg of DM for all cows, but milk production tended to increase from 29.8 to 30.9 ± 1.46 kg/d for control and Co-P treatments, respectively. Milk fat percentage and energy-corrected milk did not differ between treatments, averaging 4.33 ± 0.14% and 34.1 kg/d, respectively. Milk protein was significantly decreased by the Co-P treatment (3.56 and 3.41 ± 0.08% for control and Co-P treatments), but protein yield was not affected. Milk energies were 1.40 Mcal/d greater with Co-P. Energy lost as methane was reduced by 0.31 Mcal/d with the addition of RFDDGS to the diet. Heat loss averaged 29.9 ± 0.55 Mcal/d and was not different between diets. Average energy retained as tissue energy was -2.99 ± 0.93 Mcal/d and did not differ between treatments. Intake of digestible and metabolizable energy were not different between the control and Co-P treatments, averaging 2.68 and 2.31 Mcal/kg of DM, respectively. The net energy of lactation values of control and Co-P diets were calculated to be 1.43 and 1.47 Mcal/kg of DM, respectively. These energy estimates suggest greater energy content of diets containing RFDDGS than diets containing a mixture of corn and soybean meal in lactating dairy cows.

Genome-wide prediction of age at puberty and reproductive longevity in sows.

Traditional selection for sow reproductive longevity is ineffective due to low heritability and late expression of the trait. Incorporation of DNA markers into selection programs is potentially a more practical approach for improving sow lifetime productivity. Using a resource population of crossbred gilts, we explored pleiotropic sources of variation that influence age at puberty and reproductive longevity. Of the traits recorded before breeding, only age at puberty significantly affected the probability that females would produce a first parity litter. The genetic variance explained by 1-Mb windows of the sow genome, compared across traits, uncovered regions that influence both age at puberty and lifetime number of parities. Allelic variants of SNPs located on SSC5 (27-28 Mb), SSC8 (36-37 Mb) and SSC12 (1.2-2 Mb) exhibited additive effects and were associated with both early expression of puberty and a greater than average number of lifetime parities. Combined analysis of these SNPs showed that an increase in the number of favorable alleles had positive impact on reproductive longevity, increasing number of parities by up to 1.36. The region located on SSC5 harbors non-synonymous alleles in the arginine vasopressin receptor 1A (AVPR1A) gene, a G-protein-coupled receptor associated with social and reproductive behaviors in voles and humans and a candidate for the observed effects. This region is characterized by high levels of linkage disequilibrium in different lines and could be exploited in marker-assisted selection programs across populations to increase sow reproductive longevity.

Targeting of an interrupted polypurine:polypyrimidine sequence in mammalian cells by a triplex-forming oligonucleotide containing a novel base analogue.

The DNA triple helix consists of a third strand of nucleic acid lying in the major groove of an intact DNA duplex. The most stable triplexes form on polypurine:polypyrimidine sequences, and pyrimidine interruptions in the purine strand are destabilizing. Sequence stringency is imparted by specific Hoogsteen hydrogen bonds between third strand bases and the purine bases in the duplex. Appropriate base and sugar modifications of triple helix-forming oligonucleotides (TFOs) confer chromosome targeting activity in living cells. However, broad utilization of TFOs as gene targeting reagents in mammalian cells has been limited by the requirement for homopurine target sequences. Although there have been a number of base analogues described that appear to be promising as candidates for triplex target expansion, none has been examined in a biological system. We have employed a postsynthetic strategy to prepare a collection of TFOs with base analogues at a defined position. Following assessment of affinity for a triplex target with a single C:G inversion, TFOs with a second generation of analogues were synthesized. One of these, TFO-5a, with 2'-OMe-guanidinylethyl-5-methylcytosine at the position corresponding to the C:G interruption in the target sequence, was further modified to confer bioactivity. The activity of this TFO, linked to psoralen, was measured in a mammalian cell line that was engineered by directed sequence conversion to carry a triplex target with a single C:G interruption. TFO-5a was active against this target and inactive against the corresponding target with an uninterrupted polypurine:polypyrimidine sequence.

Extremophile-inspired strategies for enzymatic biomass saccharification.

Domestic ethanol production in the USA relies on starch feedstocks using a first generation bioprocess. Enzymes that contribute to this industry remain of critical value in new and established markets as commodity additives and for in planta production. A transition to non-food feedstocks is both desirable and essential to enable larger scale production. This objective would relieve dependence on foreign oil and strengthen the national economy. Feedstocks derived from corn stover, wheat straw, perennial grasses and timber require pretreatment to increase the accessibility of the cellulosic and hemicellulosic substrates to commodity enzymes for saccharification, which is followed by fermentation-based conversion of monosaccharides to ethanol. Hot acid pretreatment is the industrial standard method used to achieve deconstruction of lignocellulosic biomass. Therefore, enzymes that tolerate both acid and heat may contribute toward the improvement of lignocellulosic biomass processing. These enzymes are produced naturally by extremely thermophilic microbes, sometimes called extremophiles. This review summarizes information on enzymes from selected (acido)thermophiles that mediate saccharification of alpha- and beta-linked carbohydrates of relevance to biomass processing.

GENOMICS SYMPOSIUM: Using genomic approaches to uncover sources of variation in age at puberty and reproductive longevity in sows.

Genetic variants associated with traits such as age at puberty and litter size could provide insight into the underlying genetic sources of variation impacting sow reproductive longevity and productivity. Genomewide characterization and gene expression profiling were used using gilts from the University of Nebraska-Lincoln swine resource population ( = 1,644) to identify genetic variants associated with age at puberty and litter size traits. From all reproductive traits studied, the largest fraction of phenotypic variation explained by the Porcine SNP60 BeadArray was for age at puberty (27.3%). In an evaluation data set, the predictive ability of all SNP from high-ranked 1-Mb windows (1 to 50%), based on genetic variance explained in training, was greater (12.3 to 36.8%) compared with the most informative SNP from these windows (6.5 to 23.7%). In the integrated data set ( = 1,644), the top 1% of the 1-Mb windows explained 6.7% of the genetic variation of age at puberty. One of the high-ranked windows detected (SSC2, 12-12.9 Mb) showed pleiotropic features, affecting both age at puberty and litter size traits. The RNA sequencing of the hypothalami arcuate nucleus uncovered 17 differentially expressed genes (adjusted < 0.05) between gilts that became pubertal early (<155 d of age) and late (>180 d of age). Twelve of the differentially expressed genes are upregulated in the late pubertal gilts. One of these genes is involved in energy homeostasis (), a function in which the arcuate nucleus plays an important contribution, linking nutrition with reproductive development. Energy restriction during the gilt development period delayed age at puberty by 7 d but increased the probability of a sow to produce up to 3 parities ( < 0.05). Identification of pleotropic functional polymorphisms may improve accuracy of genomic prediction while facilitating a reduction in sow replacement rates and addressing welfare concerns.

Body composition, tissue deposition, and lysine utilization for protein deposition of barrows and gilts fed crystalline or protein-bound lysine.

An experiment with 2 trials (28 d/trial) was conducted to determine body composition, tissue deposition, and utilization of Lys for protein deposition (PD) of barrows and gilts fed -Lys·HCl (CLys) or protein-bound Lys in soybean meal (SBM). Thirty-two growing pigs (16 barrows and 16 gilts; average initial BW of 18.6 kg) were used in each of 2 trials. Four pigs (2 barrows and 2 gilts) were euthanized at the start of each trial to determine initial body composition. The remaining pigs were euthanized at the end of the trials to determine empty-body composition and deposition rates of water, protein, fat, ash, and AA. Pigs were randomly allotted to 1 of 7 dietary treatments. There were 2 replications per treatment in each trial for a total of 4 replications. Dietary treatments consisted of a corn-SBM basal diet (0.48% Lys) and diets containing 0.56%, 0.65%, and 0.74% standardized ileal digestible (SID) Lys that were achieved by adding Lys to the basal diet from either SBM or CLys. Pigs fed the CLys-supplemented diets at 0.65% SID Lys had more ( < 0.05) body water (663 vs. 624 g/kg) and less ( < 0.01) body protein (153 vs. 160 g/kg) than pigs fed the SBM-supplemented diets. Body fat content decreased ( < 0.01) as the dietary Lys increased similarly for pigs fed Lys from SBM and pigs fed CLys. Gilts had greater ( = 0.05) body Lys content in body protein than barrows (7.68 vs. 7.52 g/100 g). Empty-body ash contents were not different between pigs fed CLys or SBM-supplemented diets. Water deposition and PD increased linearly ( < 0.01) with dietary Lys and were least ( < 0.01) in pigs fed the basal diet but were similar when comparing pigs fed CLys and SBM-supplemented diets at the same dietary Lys concentration. Lysine deposition showed a linear increase ( < 0.01) with dietary Lys but was not different between pigs fed the 2 Lys sources at the same concentration. Barrows and gilts did not differ in tissue deposition rates. Overall, empty-body contents and deposition rates of essential and nonessential AA were not different between pigs fed CLys and pigs fed SBM-bound Lys. The amount of SID Lys required for PD ranged between 0.09 and 0.13 g/g for both sources of Lys. The Lys deposition:SID Lys intake ratio was greater ( < 0.01) in gilts than barrows (0.62 vs. 0.56). Body composition, tissue deposition, and utilization of Lys for PD and Lys deposition were not different in pigs fed diets supplemented with -Lys·HCl with respect to protein-bound Lys in SBM.

Evaluation of collection method and diet effects on apparent digestibility and energy values of swine diets.

Two experiments were conducted to investigate the effects of collection method and diet type on digestibility coefficients. In Exp. 1, 24 barrows were fed either a corn-soybean meal (CSBM) diet or CSBM with 20% dried distillers' grains with solubles (CSBM-DDGS). In Exp. 2, the effects of basal diet and collection method on determination of dried distillers' grains with solubles (DDGS) digestibility were studied using 24 barrows. The 4 diets used in Exp. 2 were: a CSBM (basal 1) , a barley-canola meal (BCM; basal 2), 80% basal 1 with 20% DDGS (CSBM-DDGS), and 80% basal 2 with 20% DDGS (BCM-DDGS). In both experiments, feces were collected using a time-based collection method (DY) or a "marker-to-marker" collection method (MM). Diets contained 0.5% of titanium dioxide (TiO) for estimating digestibility using the index marker approach (IM). The apparent total tract digestibility (ATTD) of DM and GE were lower ( < 0.05) in the CSBM-DDGS diet than in the CSBM diet in Exp. 1 but were not different in Exp. 2. All the estimates of BCM-based diets were consistently lower ( < 0.05) than those of CSBM-based diets. In Exp. 1, digestibility coefficients determined by the DY and MM were not different from each other, whereas those estimates were lower ( < 0.05) using the IM than those using the total collection approach (TC; DY and MM). In Exp. 2, interactions ( < 0.05) were observed between diet type and method for dietary digestibility coefficients. Digestibility and energy values estimated by the DY and MM were not different in pigs fed CSBM-based diets and the BCM-DDGS diet, whereas those estimates were greater ( < 0.05) using the DY than those using the MM in pigs fed the BCM. There were no interactions between basal diet and method for estimating DDGS digestibility. The ATTD of DM and GE of DDGS using the MM were greater ( < 0.05) than those using the IM, and ATTD of N tended to be greater ( < 0.10) using the MM than that using the IM. All estimates using the DY were not different from those using the MM or the IM, except that DE of DDGS was greater ( < 0.05) using the DY than when using the IM. Digestibility estimates of DDGS were not affected by basal diets. The mean DE and ME (as-fed basis) of DDGS were 3,994 and 3,688 kcal/kg, respectively, when estimated using the basal 1 diet and were 3,919 and 3,547 kcal/kg, respectively, when estimated using the basal 2 diet. In conclusion, both collection methods can be used to estimate energy and nutrient digestibility of diets and DDGS when using CSBM-based diets.

Pyrimidine motif triplexes containing polypurine RNA or DNA with oligo 2'-O-methyl or DNA triplex forming oligonucleotides.

Triplex forming oligonucleotides (TFOs) are potentially useful in targeting RNA for antisense therapeutic applications. To determine the feasibility of targeting polypurine RNA with nuclease-resistant oligonucleotides, TFOs containing 2'-deoxy or 2'-O-methyl (2'-OMe) backbones, designed to form pyrimidine motif triplexes with RNA, were synthesized. TFOs were made which can form trimolecular triplexes, or bimolecular, 'clamp' triplexes with polypurine RNA and DNA. It was found that the relative stabilities of the triplexes formed followed the order: M.DM(clamp)>>>D.DD approximately M.DD>M. RM>D.DM>M.RD approximately M.DM, where M is a 2'-OMe, D is a DNA and R is an RNA backbone. The third strand is listed first, separated by a dot from the purine strand of the Watson-Crick duplex, followed by the pyrimidine strand of the duplex. The results described here provide insight into the feasibility of using TFOs containing a 2'-OMe backbone as antisense agents.

Studies on anti-human immunodeficiency virus oligonucleotides that have alternating methylphosphonate/phosphodiester linkages.

Preliminary investigations of the physical properties of oligonucleotide analogs that contain alternating methylphosphonate/phosphodiester linkages are described. An alternating oligo-2'-O-methylribonucleoside methylphosphonate, oligomer 1676, whose sequence is complementary to the upper hairpin region of human immunodeficiency virus TAR RNA, has been synthesized. This 15-mer forms a very stable duplex with its complementary RNA target, whose melting temperature is 71 degrees C. Introduction of two mismatched bases reduces the melting temperature by 16 degrees C. Similar results were obtained with the all-phosphodiester version of oligomer 1676, which demonstrates that introduction of the methylphosphonate linkages does not significantly perturb the ability of the oligo-2'-O-methylribonucleoside methylphosphonate to bind to RNA. Unlike the phosphodiester oligomer, however, oligomer 1676 is completely resistant to hydrolysis by the 3'-exonuclease activity found in mammalian serum. The interactions between nuclease-resistant, 5'-psoralen-derivatized, alternating oligo-2'-deoxypyrimidine methylphosphonates and double-stranded DNA were also studied. A 15-mer that contains thymine, 5-methylcytosine, and 5-propynyl-uracil forms a triplex with a polypurine tract found in the env gene of human immunodeficiency virus proviral DNA with an apparent dissociation constant of 400 nM at 22 degrees C. Maximal triplex formation by these oligomers is observed at approximately 2.5 mM magnesium, whereas maximal triplex formation by the corresponding all-phosphodiester oligomers occurs between 10 and 20 mM magnesium. This reduced magnesium dependence most likely results from reduced charge repulsion between the backbones of the methylphosphonate oligomer and purine strand of the target. The nuclease stability and ability of the methylphosphonate oligomers to form stable complexes with their target nucleic acids suggest that these oligomers are potential candidates for use as antisense or antigene agents in cell culture.

Major histocompatibility complex variation in the endangered Przewalski's horse.

The major histocompatibility complex (MHC) is a fundamental part of the vertebrate immune system, and the high variability in many MHC genes is thought to play an essential role in recognition of parasites. The Przewalski's horse is extinct in the wild and all the living individuals descend from 13 founders, most of whom were captured around the turn of the century. One of the primary genetic concerns in endangered species is whether they have ample adaptive variation to respond to novel selective factors. In examining 14 Przewalski's horses that are broadly representative of the living animals, we found six different class II DRB major histocompatibility sequences. The sequences showed extensive nonsynonymous variation, concentrated in the putative antigen-binding sites, and little synonymous variation. Individuals had from two to four sequences as determined by single-stranded conformation polymorphism (SSCP) analysis. On the basis of the SSCP data, phylogenetic analysis of the nucleotide sequences, and segregation in a family group, we conclude that four of these sequences are from one gene (although one sequence codes for a nonfunctional allele because it contains a stop codon) and two other sequences are from another gene. The position of the stop codon is at the same amino-acid position as in a closely related sequence from the domestic horse. Because other organisms have extensive variation at homologous loci, the Przewalski's horse may have quite low variation in this important adaptive region.

A cooperative study on the standardized total-tract digestible phosphorus requirement of twenty-kilogram pigs.

A cooperative study comprising growth performance, bone mineralization, and nutrient balance experiments was conducted at 11 stations to determine the standardized total-tract digestible (STTD) P requirement of 20-kg pigs using broken-line regression analysis. Monocalcium phosphate and limestone were added to a corn-soybean meal-based diet at the expense of cornstarch to establish 6 concentrations of STTD P from 1.54 to 5.15 g/kg in increments of 0.62 g/kg at a constant Ca:total P of 1.52:1.0. Diets were fed to 936 pigs (average initial BW of 19 kg) in 240 pens for 20 replicate pens of barrows and 20 replicate pens of gilts per diet. As STTD P increased from 1.54 to 5.15 g/kg of the diet for d 0 to 14, 14 to 28, and 0 to 28, the ADG, ADFI, and G:F increased ( < 0.01). Barrows gained and ate more ( < 0.05) than gilts during d 14 to 28 and 0 to 28. There was no interaction between sex and STTD P concentration for any of the growth performance response criteria. There were both linear and quadratic increases ( < 0.05) in mineral density and content of ash, Ca, and P in the femur expressed as a percentage of dry, fat-free metacarpal as dietary STTD P increased. Furthermore, the maximum load of the femur and mineral density and content and maximum load as well as the Ca and P expressed as a percentage of metacarpal ash linearly increased ( < 0.01) with increasing dietary concentrations of STTD P. There were both linear and quadratic increases ( < 0.01) in apparent digestibility and retention of P with increasing concentrations of STTD P in the diets. Digestibility and retention of Ca linearly ( < 0.01) increased with increasing dietary concentrations of STTD P. Breakpoints determined from nonlinear broken-line regression analyses revealed estimates of 4.20 ± 0.102, 3.20 ± 0.036, or 3.87 ± 0.090 g/kg for ADG during d 0 to 14, 14 to 28, or 0 to 28, respectively. Corresponding estimates using G:F as the response criterion were 4.34 ± 0.146, 3.38 ± 0.139, or 4.08 ± 0.195 g/kg. When mineralization of the femur was used as criteria of response, estimates of STTD P requirement were 4.28, 4.28, or 4.34, g/kg for mineral density, mineral content, or maximum load, respectively. Using mineralization of the metacarpal as criteria of response, estimates of STTD P requirement ranged from 3.5 to 5.0 g/kg depending on the metacarpal response criteria. The study provided empirical estimates of STTD P requirements of 20- to 40-kg pigs.

Association of low serum anticholinergic levels and cognitive impairment in elderly presurgical patients.

Low-dose scopolamine, given as presurgery medication, resulted in low levels of serum anticholinergic activity and caused measurable cognitive impairment in 18 psychiatrically healthy elderly patients. The degree of impairment was directly related to serum anticholinergic activity levels and, in the small subgroup of patients scheduled for spinal anesthesia, to CSF anticholinergic activity. Two of the mental status tests used, the Rey Auditory-Verbal Learning Test and the Saskatoon Delirium Checklist, were sensitive enough to detect these mild drug-induced changes, while two other tests, the Mini-Mental State and the Symbol Digit Modalities Test, were not.

Control of ribonucleic acid function by oligonucleoside methylphosphonates.

Oligodeoxyribonucleoside methylphosphonates contain nonionic 3'-5' linked methylphosphonate internucleotide bonds in place of the normal charged phosphodiester linkage of natural nucleic acids. These oligomers are resistant to nuclease hydrolysis, can pass through the membranes of mammalian cells in culture and can form stable hydrogen-bonded complexes with complementary nucleotide sequences of cellular RNAs such as mRNA. The oligomers are readily synthesized on insoluble polymer supports. Their chainlength and nucleotide sequence can be determined by chemical sequencing procedures. Oligonucleoside methylphosphonates which are complementary to the 5'-end, initiation codon region, or coding region of rabbit globin mRNA inhibit translation of the mRNA in rabbit reticulocyte lysates and globin synthesis in rabbit reticulocytes. This inhibition is due to the interaction of the oligomers with mRNA and the extent of inhibition is influenced by the secondary structure of the mRNA and the location of oligomer binding site on the mRNA. Oligomers complementary to the initiation codon regions of N, NS and G protein mRNAs of Vesicular stomatitis virus (VSV) inhibit virus protein synthesis in VSV-infected Mouse L-cells. These oligomers do not affect L-cell protein synthesis or growth. Virus protein synthesis and growth can also be selectively inhibited by oligonucleoside methylphosphonates which are complementary to the donor or acceptor splice junctions of virus pre mRNA. An oligomer complementary to the donor splice junction of SV40 large T antigen mRNA inhibits T-antigen synthesis in SV40-infected African green monkey kidney cells but does not inhibit overall cellular protein synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Rare alleles, MHC and captive breeding.

In recent years, more detailed genetic information has become available for individuals of endangered species in captive breeding programs. There have been suggestions that this information be used to identify rare alleles, particularly those at the MHC, that can be subsequently selected for captive breeding programs. First, we summarize the current information on the MHC relevant to conservation genetics, so that such a possible breeding program is seen in a proper perspective. For example, very few specific alleles at the MHC have been identified as selectively advantageous, even though there has been substantial effort to find such alleles in humans and a few other organisms. Further, many of the balancing selection models suggested for MHC variation are based on heterozygotes in general having a higher fitness than homozygotes and not on specific selectively advantageous alleles. Because there is no detailed data on MHC variability in captive populations, we used transferrin data in Przewalski's horses to evaluate a breeding program to select for rare alleles. In this species, one individual, 1060, has been identified to have the transferrin allele J. We determine the effect on founder contribution of multiply mating 1060 to increase the number of copies of this allele. Since there were 485 individuals in the population at this time, this extra mating had little detrimental effect on the distribution of founder contributions and the number of founder equivalents. We then selected 65, an ancestor of 1060, which had a high likelihood of being the individual that passed on the J allele in the lineage of 1060. We examined the effect of increasing the number of copies of alleles of 65 at a time when the population had only 22 other individuals. In this case, even though the founder contributions were changed more, there was also little effect on the founder contributions and the number of founder equivalents. Overall, it appears that selection that results in a limited change in the number of copies of rare alleles may not always have an overall detrimental effect. However, because other pedigrees may have very different properties, it is essential to perform a detailed pedigree analysis of any such selective breeding program to determine its effect before such a selection program is implemented.