Changes in bovine leukemia virus serological status and lymphocyte count between dry-off and early lactation in Michigan dairy cows.

This study addresses how the serological status of bovine leukemia virus (BLV) and lymphocyte count fluctuate from dry-off to early lactation in dairy cattle. Very few studies have investigated how BLV antibody status and lymphocyte count of cows changes longitudinally during the lactation cycle. Blood samples were collected from dairy cattle (n = 149) on 5 commercial dairy herds in Michigan at dry-off, close-up, and 7 to 10 d after calving. Plasma was analyzed for anti-BLV antibodies using a BLV-ELISA and whole blood was analyzed for lymphocyte counts. We found that BLV seroprevalence increased from dry-off (38.9%) to close-up (43.6%), then slightly decreased from close-up to 7 to 10 d after calving (43.0%). However, the change in seroprevalence was only significant from dry-off to close-up. Cows of third or higher parity were more likely to seroconvert than cows of lower parity and had the highest ELISA-negative prevalence of BLV. Lymphocyte counts were significantly higher in ELISA-positive animals, but only among second and third or greater parity animals. These results indicate that the use of lymphocyte counts as a disease severity monitoring tool for BLV should differ by parity group. Future studies should investigate if changes in seroprevalence are due to new infections or natural changes in antibody concentrations as the cow prepares for colostrum production. More accurate lymphocyte guidelines to be used for monitoring the progression of BLV should be created that consider parity and lactation stage.

Genetic and nongenetic profiling of milk pregnancy-associated glycoproteins in Holstein cattle.

Pregnancy-associated glycoproteins (PAG) are secreted by the trophoblast and are detectable in maternal circulation around the time of attachment of the fetal placenta, as well as in blood and milk throughout gestation. The understanding of the genetic mechanisms controlling PAG levels can confer advantages for livestock breeding programs given the precocity and the ease of obtaining this phenotype from routine pregnancy diagnosis. The aim of this study was to characterize PAG levels by estimating genetic parameters and correlations with other dairy traits, and to identify genomic regions and candidate genes associated with PAG levels in milk. The PAG data consisted of pregnancy diagnoses using commercial assays from 2012 to 2017, and genotype data consisted of 54,123 SNP markers for 2,352 individuals (embryos and dams). The model included contemporary group (herd, year, and season) and embryo age as fixed effects, and random embryonic (direct) and maternal (indirect) genetic effects. Using genomic data, the estimated heritability for direct and maternal genetic effects (± standard deviations) were 0.23 ± 0.05 and 0.11 ± 0.05, respectively. The genetic correlation between these effects was almost zero (0.001 ± 0.06). A preliminary analysis revealed low correlations between milk PAG levels and other dairy traits. The genome-wide association analysis was performed using 2 approaches: single-marker and single-step using all markers. Four genomic regions with direct genetic effects were detected on Bos taurus autosome (BTA) 6, BTA7, BTA19, and BTA29 of the embryonic genome. The BTA29 locus was within the bovine PAG gene cluster, suggesting a cis-regulatory quantitative trait locus on the PAG expression. However, other associations, without an obvious link to PAG expression, could be related to the transportation of PAG and their concentration in milk. Only 1 region from the maternal genome, on BTA4, had a significant indirect effect, where WNT2 is a candidate gene related to placenta vascularization and gestation health. Collectively, our results suggest a moderate genetic control of milk PAG levels from both maternal and fetal genomes, but larger studies are needed to fully evaluate the usefulness of milk PAG in the genetic evaluation of fetal growth and cow fertility.

Genome-wide association mapping and pathway analysis of leukosis incidence in a US Holstein cattle population.

Bovine leukosis virus is an oncogenic virus that infects B cells, causing bovine leukosis disease. This disease is known to have a negative impact on dairy cattle production and, because no treatment or vaccine is available, finding a possible genetic solution is important. Our objective was to perform a comprehensive genetic analysis of leukosis incidence in dairy cattle. Data on leukosis occurrence, pedigree and molecular information were combined into multitrait GBLUP models with milk yield (MY) and somatic cell score (SCS) to estimate genetic parameters and to perform whole-genome scans and pathway analysis. Leukosis data were available for 11 554 Holsteins daughters of 3002 sires from 112 herds in 16 US states. Genotypes from a 60K SNP panel were available for 961 of those bulls as well as for 2039 additional bulls. Heritability for leukosis incidence was estimated at about 8%, and the genetic correlations of leukosis disease incidence with MY and SCS were moderate at 0.18 and 0.20 respectively. The genome-wide scan indicated that leukosis is a complex trait, possibly modulated by many genes. The gene set analysis identified many functional terms that showed significant enrichment of genes associated with leukosis. Many of these terms, such as G-Protein Coupled Receptor Signaling Pathway, Regulation of Nucleotide Metabolic Process and different calcium-related processes, are known to be related to retrovirus infection. Overall, our findings contribute to a better understanding of the genetic architecture of this complex disease. The functional categories associated with leukosis may be useful in future studies on fine mapping of genes and development of dairy cattle breeding strategies.

Effect of infection with bovine leukemia virus on milk production in Michigan dairy cows.

The objective of this study was to determine the association between individual cow-level milk production and bovine leukemia virus (BLV) infection as measured by milk BLV-ELISA. Dairy Herd Improvement technicians collected milk samples from 10 cows from each of first, second, third, and 4+ parity cows in 105 Holstein herds with ≥ 120 milking cows. Milk samples were tested for the presence of anti-BLV antibodies by ELISA. Additional data regarding the cows and the herds were collected by farm survey and Dairy Herd Improvement records. A set of mixed-effect models using all cows and only 2+ parity cows were used to investigate the association between BLV ELISA-corrected optical density and 305-d mature equivalents of individual cows. The BLV milk positivity was associated with decreased 305-d mature-equivalent yields, especially among the older cows. Additionally, increasing milk ELISA-corrected optical density was associated with increasing loss of milk production at the cow level. In summary, our results provide evidence that BLV infection is associated with decreased milk production in Michigan dairy cows.

Short communication: Genetic correlation of bovine leukosis incidence with somatic cell score and milk yield in a US Holstein population.

Bovine leukosis (BL) is a retroviral disease caused by the bovine leukosis virus (BLV), which affects only cattle. Dairy cows positive for BL produce less milk and have more days open than cows negative for BL. In addition, the virus also affects the immune system and causes weaker response to vaccines. Heritability estimates of BL incidence have been reported for Jersey and Holstein populations at about 0.08, indicating an important genetic component that can potentially be exploited to reduce the prevalence of the disease. However, before BL is used in selection programs, it is important to study its genetic associations with other economically important traits such that correlated responses to selection can be predicted. Hence, this study aimed to estimate the genetic correlations of BL with milk yield (MY) and with somatic cell score (SCS). Data of a commercial assay (ELISA) used to detect BLV antibodies in milk samples were obtained from Antel BioSystems (Lansing, MI). The data included continuous milk ELISA scores and binary milk ELISA results for 11,554 cows from 112 dairy herds across 16 US states. Continuous and binary milk ELISA were analyzed with linear and threshold models, respectively, together with MY and SCS using multitrait animal models. Genetic correlations (posterior means ± standard deviations) between BL incidence and MY were 0.17 ± 0.077 and 0.14 ± 0.076 using ELISA scores and results, respectively; with SCS, such estimates were 0.20 ± 0.081 and 0.17 ± 0.079, respectively. In summary, the results indicate that selection for higher MY may lead to increased BLV prevalence in dairy herds, but that the inclusion of BL (or SCS as an indicator trait) in selection indexes may help attenuate this problem.

Application of an ELISA Milk Pregnancy Test in Beef Cows.

Pregnancy-associated glycoproteins (PAG) are secreted by the binucleate giant cells of the ruminant placenta and enter maternal circulation at the time of placental attachment. The IDEXX Milk Pregnancy Test (IDEXX, Westbrook, ME) detects a subset of PAG in milk. Although designed as a management tool for dairy cows, there is potential for using the milk PAG test in beef cows. Our objective was to compare the performance of the milk PAG ELISA with a gold standard method for pregnancy diagnosis and determine the agreement between milk and serum PAG analysis in lactating beef cows. Angus and Angus-crossed cows (n = 332) from two Michigan beef herds were enrolled in this study. Cows were subjected either to timed artificial insemination followed by exposure to a bull or exclusively exposed to a bull. The bulls and cows were separated 30 days prior to examination. Serum and milk samples were collected and submitted within 24 h of collection to a commercial laboratory for PAG analysis using the IDEXX Milk Pregnancy Assay (milk) and the IDEXX Bovine Pregnancy Assay (serum). Concurrently with milk and serum collection, each cow was examined transrectally by palpation or ultrasonography. When compared to transrectal examination, the performance (and 95% confidence intervals) of the milk PAG ELISA was sensitivity of 99.7% (99.0-100.0%) and specificity of 80.8% (65.6-95.9%). The lower specificity is likely due to the low prevalence (9.9%) of open cows (n = 30) in the herds examined. Of the 332 cows examined, 1.8% (n = 6) were classified as rechecks using the milk PAG ELISA. Results of the milk and serum PAG ELISA were in high agreement (kappa coefficient = 0.91). The milk PAG ELISA was accurate in predicting pregnancy status using milk collected from beef cattle between days 37 and 125 post-insemination and may be useful for aiding management decisions in beef herds.

Bovine leukemia virus and cow longevity in Michigan dairy herds.

To determine the association between infection with bovine leukemia virus (BLV) and cow longevity, a stratified random sample of 3,849 Holsteins in 112 Michigan dairy herds was followed for an average of 597 d following testing for BLV antibodies with an ELISA milk test. The hazard ratio of 1.23 indicates that BLV-positive cows were 23% more likely than their BLV-negative herd mates to die or be culled during the monitoring period. This result is adjusted for lactation number, which is also positively associated with an increased risk of leaving the herd. Because herd was included in models, the effect of BLV ELISA on cow longevity was a within-herd comparison in which BLV-infected cattle were compared with their uninfected herd mates. The analysis of 4 ELISA optical density (OD) groups demonstrated a dose response such that cows with higher OD values had decreased survival compared with cows with lower OD values. Cows with OD values above 0.5 were at 40% greater risk of dying or being culled than were their uninfected herd mates. These results support the contention that the association of BLV with cow longevity, when added to other economic impacts, may warrant the control of BLV in our US dairy cow population.

Association between bovine leukemia virus, production, and population age in Michigan dairy herds.

The objective of this study was to determine the herd-level effect of bovine leukemia virus (BLV) infection on dairy production, culling, and cow longevity. During routine herd testing, Dairy Herd Improvement Association technicians collected milk samples from about 40 cows from each of 104 randomly selected Michigan dairy herds averaging ≥120 milking cows and 11,686 kg of milk/yr. Milk samples were analyzed for the presence of anti-BLV antibodies by ELISA, and herd- and lactation-specific estimates of BLV prevalence were computed to determine which were the most predictive of herd milk production, culling rate, and cow longevity (proportion of cows in their third or greater lactation). On this basis, the herd BLV index (an unweighted mean BLV prevalence rate for lactation number 1, 2, 3, and ≥4) was selected as the measure of BLV prevalence that was the most highly associated with BLV economic impact. Step-down multivariate analysis was used to determine the extent to which any of 19 herd-level management variables may have confounded the association of BLV index and measures of herd economic impact (milk production and cow longevity). The BLV index was not associated with the 12-mo culling rate, but was negatively associated in the final multivariable model with the proportion of cows that were ≥third lactation, and was negatively associated with herd milk production. In summary, increased prevalence of BLV within Michigan dairy herds was found to be associated with decreased herd milk production and decreased cow longevity. Our results provide evidence that BLV infection is associated with herd-level economic impacts in high-performing dairy herds.

Herd-level prevalence of Johne's disease in Utah and adjacent areas of the intermountain west as detected by a bulk-tank milk surveillance project.

The objectives of this study were to estimate the dairy herd-level prevalence of Johne's disease (JD) in Utah and nearby areas of the intermountain west and to estimate the sensitivity of a single bulk-tank milk test for JD detection. Two milk samples from all bulk tanks on the study farms were collected 1 mo apart. Samples were frozen and shipped to a laboratory for JD testing. An ELISA to measure total IgG antibody specific against Mycobacterium avium ssp. paratuberculosis, the etiological agent that causes JD, and a quantitative real-time PCR to detect M. avium ssp. paratuberculosis DNA were used; both tests were designed for bulk milk. Of the dairy farms in the study area, 170/246 (69%) participated. Positive JD results were found in bulk milk from 67/170 (39%) of dairy farms in Utah and adjacent areas. There were 138 JD-positive bulk-tank results from 241 bulk-tank samples from the 67 positive herds. The sensitivity of the bulk milk testing for detection of JD was 138/241(57%). From the 103 JD-negative farms, 235 bulk-tank samples tested negative for JD. The probability of false-negative results on a single bulk-milk sample was (1 - 0.57) = 0.43. For farms with 1 bulk tank, 2 samples collected 1 mo apart, with both samples testing negative (by both ELISA and quantitative real-time PCR) for JD, the true-negative probability was [1 - (0.43)(2)] = (1 - 0.18) = 82%. For farms with at least 2 bulk tanks, at least 4 samples tested, with all results negative for JD, the true-negative probability was at least 97%. Results support other estimates that prevalence of JD has increased over the last 15 to 20 yr. However, the prevalence detected was 3 times that from a recent report where 13% of dairy herds in the western US were positive. The increase in JD suggests that current control programs, at least as applied, are not effective. Bulk milk testing is a practical way to screen dairy herds for presence of JD. Studies are needed regarding the use of individual cow milk tests for accuracy, practicality, and effectiveness in reducing the prevalence of JD in dairy herds.

Characterization of dose-dependent metabolic responses to close arterial infusion of cimaterol in the hindlimb of steers.

Dose-dependent effects of cimaterol (CIM) on hindlimb metabolism were determined in six steers (247 +/- 22 kg BW) using a close arterial infusion. The external iliac vessels of both hindlimbs were catheterized to accommodate measurement of blood flow, circulating concentrations, and net flux of NEFA, lactate, and alpha-amino nitrogen (AAN) during infusion of CIM at 0, .05, .1, .3, .7, 1 and 3 micrograms/ min. Close arterial infusion of CIM in the hindlimb of steers can be used to achieve a local concentration elevation that is required to differentiate local and systemic effects in vivo. Calculated plasma threshold CIM concentration required to initiate cardiovascular responses was 21 pg/mL, which resulted from an infusion rate of .3 microgram/min. Threshold concentrations of CIM for stimulation of NEFA and lactate net flux in the hindlimb were 38 and 34 pg/mL, respectively, and would be achieved with an infusion rate of .7 microgram/min. All measured responses except AAN net flux exhibited significant linear and quadratic dose effects, and responses in the treated hindlimb were always severalfold greater than in the contralateral control hindlimb. Maximal differences between treated and control hindlimb blood flow occurred with a CIM infusion rate of .7 microgram/min, but the highest infusion rate (3 micrograms/min) was required to maximize differences in NEFA and lactate flux. Therefore, to minimize cardiovascular and other systemic responses and optimize direct hindlimb responses, an infusion rate of .5 microgram of CIM/ min should cause significant stimulation of beta-adrenergic receptors only in the CIM-infused hindlimb of young, growing steers.

The beta-agonist cimaterol directly enhances chronic protein accretion in skeletal muscle.

Our objective was to determine whether the chronic anabolic effects of beta-adrenergic agonists on skeletal muscle are direct and how long they are maintained. We studied acute (6 h) and chronic (1 to 20 d) effects of cimaterol (CIM) on skeletal muscle metabolism and protein accretion by use of close arterial infusion in the hindlimbs of six young steers. Surgical catheterizations were conducted to allow continuous infusion of CIM (.5 microg/min) or saline into the external iliac artery of contralateral hindlimbs and simultaneous sampling for arteriovenous difference measurements. Hindlimb blood flow and net flux of amino acids, glucose, lactate, and NEFA were determined during a basal period before infusion, at 6 h, and at 1, 3, 7, 14, and 20 d of infusion. Cimaterol infusion acutely stimulated blood flow and caused acute mobilization of nitrogen (alanine), NEFA, and lactate from the treated hindlimb. Cimaterol infusion increased net uptake of amino acids (P < .05) in treated and control hindlimbs after 1 d of CIM infusion, but a progressive increase between 1 and 14 d of infusion was observed only in the treated hindlimbs. Net uptake of total amino acids in the treated hindlimb was increased 50 and 80% (P < .05) at 7 and 14 d, respectively, when compared to the control hindlimb and was increased 260% at d 14 when compared with the basal period. Net amino acid uptake was not different between treated and control hindlimbs by d 20 of CIM infusion. Integration of net tyrosine and phenylalanine uptake over the entire infusion period predicted a 10% difference in skeletal muscle protein mass between treated and control hindlimbs. Semitendinosus and semimembranosus muscles in the treated hindlimb contained 9 and 11% greater protein content, respectively (P < .05), at the end of the infusion period. Results provide a quantitative description of the temporal pattern of transient effects of CIM on skeletal muscle metabolism and protein accretion and provide evidence that these are direct effects.

Ensiling characteristics of whole-crop small grains harvested at milk and dough stages.

Dynamics of the ensiling process and final quality of whole-crop wheat, oats, and barley harvested at the milk and the dough stage of maturity were studied using 2.2-liter laboratory silos. An ensiling period of 64 d was used. Silage DM was higher (P less than .05) at the dough than at the milk stage on d 0 and 64. Silage total N content was not different (P greater than .05) among small grain species or stages of maturity either at harvest or 64 d after ensiling. Overall, buffering capacity (BC) on d 0 was greater (P less than .05) in oats and barley than in wheat silages. Small grains had a greater (P less than .05) BC when harvested in the milk stage than when harvested in the dough stage. In all silages, pH had declined (P less than .01) by d 64, when it ranged from 3.8 to 4.1. During the ensiling period, lactic acid concentrations increased (P less than .01) and ranged from 6 to 11% of DM by 64 d of ensiling. Water-soluble carbohydrate (WSC) content was greater (P less than .01) in milk- than in dough-stage silages for each small grain species and declined (P less than .05) during the ensiling period. The WSC decline during ensiling was greater (P less than .01) for milk-stage than for dough-stage harvested forage. Water-soluble N increased during ensiling (P less than .01) and increased 100 to 200% by d 64 for all silages. Ammonia N increased during the 64-d ensiling period but never exceeded .2% of DM.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of cyclopiazonic acid on the development of pale, soft, and exudative pork from pigs of defined malignant hyperthermia genotype.

Malignant hyperthermia (MH) and the mycotoxin cyclopiazonic acid (CPA) are each associated with abnormal calcium homeostasis in skeletal muscle, a key underlying factor in the development of pale, soft, and exudative (PSE) pork. To determine whether the natural presence of CPA in livestock feed ingredients contributes to the varying incidence of PSE in the pork industry, various levels of CPA (.1 to 50 mg/kg of diet) were included in the diets of market weight hogs (n = 52) of defined malignant hyperthermia genotype (NN = normal, Nn = a MH carrier, and nn = MH-positive). Animals with two copies of the MH mutation (nn) displayed improved live animal performance compared with NN and Nn animals (increased feed intake, average daily gain, and feed efficiency) but yielded lower quality loin chops as indicated by lower 45-min pH (P<.01), higher Commission Internationale de l'Eclairage (CIE) L* color coordinate values (P<.05), and higher drip losses (P<.01). The effects of CPA varied. In the first feeding trial, conducted under normal outside temperatures (2 degrees C), CPA had no effect (P> .2) on either live animal performance or meat quality. During the second trial, conducted under extreme outside temperatures (-18 degrees C), CPA-dependent reductions (P<.05) in feed intake, average daily gain, and 45-min pH in nn hogs support the possibility of interactions between malignant hyperthermia and dietary CPA on skeletal muscle calcium homeostasis and the development of PSE pork. These results suggest that this interaction may require stressful environmental conditions or the ingestion of CPA doses much higher than occur under natural conditions.

Analysis and pharmacokinetics of cyclopiazonic acid in market weight pigs.

The pharmacokinetic behavior of cyclopiazonic acid (CPA) was determined in market weight pigs using a competitive indirect ELISA developed for the determination of the mycotoxin in various biological matrices. Sample preparation for corn and skeletal muscle was achieved with a single extraction and recoveries of 53+/-6% over the effective range of the standard curve. The detection limit of CPA was 1 ppb in plasma, which required no extraction, and 20 ppb in corn and skeletal muscle with average intra- and interassay CV of 11 and 23%, respectively. Levels of CPA contamination in corn grown and stored in Michigan were unremarkable compared with published toxicity thresholds; the highest level of CPA found in any sample was 47 ppb. In pigs given a 20-mg i.v. bolus, CPA distributed rapidly among three compartments, with an overall volume of distribution (49 L) nearly equivalent to total body water. Cyclopiazonic acid was eliminated with a half-life of 24 h. Estimates of these pharmacokinetic parameters were supported by the achievement of steady-state plasma CPA levels within 6 d in pigs consuming a diet containing 10 ppm CPA, and by measured concentrations of CPA in plasma (410+/-44 ng/mL) and skeletal muscle (469+/-86 ng/ g). From these and other data, we concluded that the threat of CPA toxicity to livestock from consumption of cereal grains or to humans from consumption of animal products is minimal.

Analysis and pharmacokinetics of cimaterol in growing Holstein steers.

Pharmacokinetic parameters for the beta 2-adrenergic agonist, cimaterol (CIM), were determined in growing Holstein steers. Compartmental analysis was used after measurement of CIM in body fluids by affinity chromatography and HPLC using UV detection. Recoveries from spiked plasma and urine standards were 70 +/- 1.2% and 68 +/- 1.1%, respectively. The minimum detection level in plasma was 1 ng/mL and the average CV was 5.1% for concentrations that ranged from 1 to 30 ng/mL. Four steers (276 +/- 24 kg) received 15 mg of CIM by bolus intravenous injection. Plasma CIM levels declined in a biphasic manner with half-lives of 2.5 min for the distribution phase and 54 min for the elimination phase. A two-compartment open model was used to describe the disappearance of CIM and the following pharmacokinetic parameters were obtained: central compartment volume (Vc) = .76 L/kg, apparent volume of distribution (Vd) = 4.1 L/kg, and transfer rate constants from the central to peripheral compartment (k12) = .177/min, from the peripheral to central compartment (k21) = .054/min and elimination from the central compartment (kel) = .074/min. After 8 h, total urinary CIM accounted for only 18.3% of the administered dose. Results suggest that circulating concentrations of CIM in growing steers are influenced by its accumulation in an unidentified peripheral pool and its conversion into unknown metabolite(s) before elimination.

Skeletal muscle calcium channel ryanodine binding activity in genetically unimproved and commercial turkey populations.

The biochemical basis for the incidence of pale, soft, exudative (PSE) turkey meat was investigated by conducting ryanodine binding experiments on sarcoplasmic reticulum (SR) vesicles prepared from genetically unimproved and commercial turkeys. Ryanodine binding to the Ca2+ channel protein in SR vesicles from both populations of turkeys was activated at a threshold concentration of approximately 0.2 microM Ca2+, reached a plateau over the range of 3 to 30 microM free Ca2+, and was only slightly inhibited at 1 mM Ca2+. The SR fractions, enriched in the Ca(2+)-channel protein, from commercial turkeys exhibited a higher (P < 0.05) mean affinity for ryanodine when compared to that from unimproved turkeys (Kd = 12.2 vs 20.5 nM, respectively). A fourfold difference (P < 0.05) in mean Ca(2+)-channel protein content or Bmax (1.10 pmol/mg vs 4.01 pmol/mg) was observed between commercial and unimproved turkey SR fractions. The apparent difference in channel protein content between the two populations may be partially accounted for by the high abundance of a 75-kDa protein, as yet unidentified, observed in most commercial turkey samples on SDS polyacrylamide gels. The differences in ryanodine binding activity between these two populations of turkeys suggest that altered SR calcium channel protein activity, or altered channel regulation, may be associated with the increased incidence of PSE meat from turkeys selected for growth characteristics.

Performance of a commercial serum ELISA for the detection of antibodies to Neospora caninum in whole and skim milk samples.

Control of Neospora caninum infection in cattle depends on specific, ante-mortem detection of infected animals and limiting their use as breeding stock or by culling. The objective of the present study was to determine appropriate cut-off values and diagnostic performance of a milk ELISA test using whole and skim milk in a commercial serum ELISA test (IDEXX Neospora Ab). Serum and milk samples were obtained from a total of 475 lactating cows from two herds with and two herds without a previous history of N. caninum-associated abortion. Overall seroprevalence determined by the ELISA was 18.3%. Compared to serum ELISA values, correlation and overall performance assessed by receiver operating characteristic analysis was higher when either whole or skim milk samples were diluted 1:2 compared to undiluted or 1:5 diluted samples. Diagnostic performance for analysis of whole and skim milk was compared at cut-off values that achieved a desired operating characteristic of at least 95% specificity. For whole milk diluted 1:2 and a cut-off of 0.14 (S/P ratio), sensitivity and kappa values were 74.7% (95% CI 64.3-83.4) and 0.70 (95% CI 0.61-0.78), respectively. For skim milk diluted 1:2 and a cut-off of 0.30, sensitivity and kappa values were 77.0% (95% CI 66.8-85.4) and 0.72 (95% CI 0.64-0.80), respectively. Using the selected cut-offs, the IDEXX Neospora Ab Test is equally suited for the analysis of whole and skim milk as a screening tool in neosporosis control programs.

Modulation of liposomal membrane fluidity by flavonoids and isoflavonoids.

The polyphenolic structures of flavonoids and isoflavonoids confer them with the ability to scavenge free radicals and to chelate transition metals, a basis for their potent antioxidant abilities. Another possible contributory mechanism toward their antioxidant activities is their ability to stabilize membranes by decreasing membrane fluidity. In this study, the effects of representative flavonoids, isoflavonoids, and their metabolites on membrane fluidity and their preferential localization in the membrane were investigated using large unilamellar vesicles (LUVs) as the membrane models. These results were compared with those of cholesterol and alpha-tocopherol. Changes in fluorescence anisotropy values for a series of n-(9-anthroyloxy) fatty acid probes (n = 6, 12, 16) upon addition of the test compounds were used to monitor alterations in membrane fluidity at graded depths in lipid bilayer. The results of the study suggest that the flavonoids and isoflavonoids, similar to cholesterol and alpha-tocopherol, partition into the hydrophobic core of the membrane and cause a dramatic decrease in lipid fluidity in this region of the membrane. Localization of flavonoids and isoflavonoids into the membrane interiors and their resulting restrictions on fluidity of membrane components could sterically hinder diffusion of free radicals and thereby decrease the kinetics of free radical reactions.

Differential Ca(2+) sensitivity of skeletal and cardiac muscle ryanodine receptors in the presence of calmodulin.

Calmodulin (CaM) activates the skeletal muscle ryanodine receptor Ca(2+) release channel (RyR1) in the presence of nanomolar Ca(2+) concentrations. However, the role of CaM activation in the mechanisms that control Ca(2+) release from the sarcoplasmic reticulum (SR) in skeletal muscle and in the heart remains unclear. In media that contained 100 nM Ca(2+), the rate of (45)Ca(2+) release from porcine skeletal muscle SR vesicles was increased approximately threefold in the presence of CaM (1 microM). In contrast, cardiac SR vesicle (45)Ca(2+) release was unaffected by CaM, suggesting that CaM activated the skeletal RyR1 but not the cardiac RyR2 channel isoform. The activation of RyR1 by CaM was associated with an approximately sixfold increase in the Ca(2+) sensitivity of [(3)H]ryanodine binding to skeletal muscle SR, whereas the Ca(2+) sensitivity of cardiac SR [(3)H]ryanodine binding was similar in the absence and presence of CaM. Cross-linking experiments identified both RyR1 and RyR2 as predominant CaM binding proteins in skeletal and cardiac SR, respectively, and [(35)S]CaM binding determinations further indicated comparable CaM binding to the two isoforms in the presence of micromolar Ca(2+). In nanomolar Ca(2+), however, the affinity and stoichiometry of RyR2 [(35)S]CaM binding was reduced compared with that of RyR1. Together, our results indicate that CaM activates RyR1 by increasing the Ca(2+) sensitivity of the channel, and further suggest differences in CaM's functional interactions with the RyR1 and RyR2 isoforms that may potentially contribute to differences in the Ca(2+) dependence of channel activation in skeletal and cardiac muscle.

Abomasal casein infusion and exogenous somatotropin enhance nitrogen utilization by growing lambs.

Growing Dorset wether lambs (23 kg initial body weight) were used to determine whether the magnitude of nitrogen retention response to daily administration of exogenous somatotropin is limited by post-ruminal amino acid availability in growing ruminants. Eight lambs surgically fitted with abomasal cannulae were fed a total mixed ration of 85% of ad libitum intake. All lambs received a continuous abomasal infusion of 2 L of water or casein and twice daily subcutaneous injections of 0 or 100 micrograms recombinant bovine somatotropin (rbST)/kg body wt for 15 d per treatment in a 2 x 2 single reversal design. The casein solution was infused at a rate (4 to 5 g nitrogen/d) to achieve 25% of nitrogen intake observed with ad libitum feeding prior to initiation of treatments. Each lamb received all four treatments. Nitrogen balance was determined on d 8 to 14 of each treatment. Casein infusion increased nitrogen balance 43.4% (P less than 0.001), and rbST increased nitrogen balance 33.5% (P less than 0.001), without significant interaction (P less than 0.88). Combined effects of casein and rbST were additive, resulting in an 89% increase in nitrogen balance when compared with water plus excipient treatment. Results suggest that the quantity or composition of absorbed amino acids, or both, limit nitrogen retention by growing lambs, and that rbST increases the efficiency of utilization of absorbed amino acids for protein deposition.