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.

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.

Survival in crossbred lambs: Breed and heterosis effects.

The objective of this study was to estimate genetic and nongenetic effects affecting survival in crossbred lambs during 3 time periods: through 1 d of age ( = 7,933), 2 to 30 d of age ( = 5,370), and 2 to 60 d of age ( = 5,216) in a population being upgraded to the dairy breeds of East Friesian and Lacaune in which lambs were artificially reared on milk replacer. Survival was analyzed for lambs born in 14 yr from 1998 to 2011 using pedigree information from 14,339 animals born in 23 yr from 1989 to 2011. Date of death, sex, age of the dam, birth type, month and year of birth, and breed composition were recorded, and the proportion of retained heterosis was calculated for each animal. Lambs were crossbreds of 2 or more breeds with 14 breeds represented in the population. Due to low mean genetic contribution of the 12 nondairy breeds, they were placed into 2 groups: meat breeds (Hampshire, Suffolk, and Texel) and maternal breeds (Romanov, Finnsheep, Dorset, Targhee, Rambouillet, Polypay, Katahdin, Arcott Rideau, and Commercial). The proportion of individual retained heterosis was positively associated ( < 0.05) with lamb survival from 2 to 30 and from 2 to 60 d of age. The predicted increase in survival of F lambs compared to purebred lambs was +8.8 and +14.6%, respectively. Predicted survival of meat breed lambs and maternal breed lambs was greater ( ≤ 0.01) than Lacaune lambs during all 3 periods. Predicted survival of East Friesian lambs was consistently lower ( ≤ 0.01) than meat breed and maternal breed lambs during all periods. The predicted survival of East Friesian lambs was numerically greater but not significantly different from Lacaune lambs. There was a lower ( < 0.01) survival of females compared to males through 1 d of age (-5.6%), but females had higher ( < 0.01) survival than males in the other 2 periods (2 to 30 d = +3.3% and 2 to 60 d = +6.0%). Through 1 d of age, lambs of triplet and greater birth types had lower ( < 0.01) survival than single lambs (-6.2%), and lambs from 1-yr-old dams had lower ( < 0.01) survival than lambs from 2-yr-old dams (+4.5%). Estimates of heritability of lamb survival were 0.14 (SE = 0.03), 0.03 (SE = 0.04), and 0.06 (SE = 0.03) for the 3 time periods, respectively. An increase in the proportion of individual retained heterosis was the most important genetic factor associated with increased lamb survival in this study.

Assessment of bagging GBLUP for whole-genome prediction of broiler chicken traits.

Bootstrap aggregation (bagging) is a resampling method known to produce more accurate predictions when predictors are unstable or when the number of markers is much larger than sample size, because of variance reduction capabilities. The purpose of this study was to compare genomic best linear unbiased prediction (GBLUP) with bootstrap aggregated sampling GBLUP (Bagged GBLUP, or BGBLUP) in terms of prediction accuracy. We used a 600 K Affymetrix platform with 1351 birds genotyped and phenotyped for three traits in broiler chickens; body weight, ultrasound measurement of breast muscle and hen house egg production. The predictive performance of GBLUP versus BGBLUP was evaluated in different scenarios consisting of including or excluding the TOP 20 markers from a standard genome-wide association study (GWAS) as fixed effects in the GBLUP model, and varying training sample sizes and allelic frequency bins. Predictive performance was assessed via five replications of a threefold cross-validation using the correlation between observed and predicted values, and prediction mean-squared error. GBLUP overfitted the training set data, and BGBLUP delivered a better predictive ability in testing sets. Treating the TOP 20 markers from the GWAS into the model as fixed effects improved prediction accuracy and added advantages to BGBLUP over GBLUP. The performance of GBLUP and BGBLUP at different allele frequency bins and training sample sizes was similar. In general, results of this study confirm that BGBLUP can be valuable for enhancing genome-enabled prediction of complex traits.

Genetic analysis of leukosis incidence in United States Holstein and Jersey populations.

Bovine leukosis (BL) is a retroviral disease caused by the bovine leukosis virus that affects only cattle. It is associated with decreased milk production and increased cull rates due to development of lymphosarcoma. The virus also affects the immune system. Infected cows display a weak response to some vaccinations. It is important to determine if the heritability of BL susceptibility is greater than zero, or if the environment is the only factor that can be used to reduce the transmission and incidence of the disease. Accordingly, the aim of this study was to estimate the heritability for BL incidence and the genetic merit of sires for leukosis resistance in Holstein and Jersey cattle. Continuous scores and binary milk ELISA results for 13,217 Holstein cows from 114 dairy herds across 16 states and 642 Jersey cows from 8 dairy herds were considered. Data were obtained from commercial testing records at Antel BioSystems (Lansing, MI). Out of the 13,859 animals tested, 38% were found to be infected with the disease. Linear and threshold animal models were used to analyze the continuous and binary data, respectively. Results from both models were similar in terms of estimated breeding values and variance components in their respective scales. Estimates of heritability obtained with the 2 approaches were approximately 8% for both breeds, indicating a considerable genetic component underlying BL disease incidence. The correlation between the estimated breeding values from the 2 models was larger than 0.90, and the lists of top 10% bulls selected from each model had about 80% overlap for both breeds. In summary, results indicate that a simple linear model using the continuous ELISA scores as the response variable was a reasonable approach for the genetic analysis of BL incidence in cattle. In addition, the levels of heritability found indicate that genetic selection could also be used to decrease susceptibility to bovine leukosis virus infection in Holstein and Jersey cattle. Further research is necessary to investigate the genetic correlations of BL with other production and reproduction traits, and to search for potential genomic regions harboring major genes affecting BL susceptibility.

Características reprodutivas e suas associações com outras características de importância econômica na raça Nelore / Reproductive traits and their associations with other economically important traits in Nelore

Objetivou-se com este trabalho estimar as herdabilidades (h²) e as correlações genéticas (r g) entre idade ao primeiro parto (IPP) e primeiro intervalo de partos (PIEP) e outras características como peso (PS) ao ano (A) e ao sobreano (S), altura do posterior (ALT) e perímetro escrotal (PE450) em animais da raça Nelore. Os parâmetros genéticos foram estimados em uma análise multicaracterística por modelo animal, utilizando-se a inferência bayesiana via algoritmo de "Gibbs Sampling". Os parâmetros genéticos estimados sugerem a existência de variabilidade genética para IPP (h² = 0,26), sendo que a seleção para a diminuição da IPP de fêmeas Nelore deve responder à seleção individual, sem causar antagonismo do valor genético dos animais para PS (r g = -0,22 (A) e -0,44 (S)) e PE450 (r g = 0,02). A seleção para a diminuição da IPP, no longo prazo, pode levar a um aumento da ALT dos animais, embora essa associação seja relativamente baixa (-0,35). A estimativa de herdabilidade a posteriori para a característica PIEP foi baixa, 0,11±0,03. As r g entre PIEP e as demais características estudadas indicam que a seleção para essas características de crescimento não afetará o PIEP.

Heritability (h²) and genetic correlations (r g) were estimated between reproductive traits such as age at first calving (AFC), first calving interval (FCI) and other economically relevant traits, i.e., weight (W) at year (Y) and at 18 months of age (S), scrotal circumference (SC), and hip height (HH) in Nelore cattle. The genetic parameters were estimated in a multiple-trait analysis, with animal models using the Bayesian inference by Gibbs Sampling algorithm. The genetic parameters estimated in this work suggest the existence of genetic variability for AFC (h² = 0.26), where the selection for the reduction of Nelore females AFC should respond to mass selection, without causing genetic antagonism in the selection of W (r g = -0,22 (Y) and -0,44 (S)), and SC (r g = 0,02). The selection for the AFC in the long term could lead to an increase in the animal's frame, although this association is relatively low (-0.35). The posteriori heritability estimate for FCI was low, 0.11±0.03. The r g between FCI and the other traits studied indicate that selection for these growth traits will not affect the FCI.

Analysis of the bovine neutrophil transcriptome during glucocorticoid treatment.

The objective of this study was to characterize a large portion of the bovine neutrophil transcriptome following treatment with the anti-inflammatory glucocorticoid dexamethasone (Dex). Total RNA was isolated from blood neutrophils of healthy cattle (5 castrated male Holsteins) immediately following cell purification (0 h) or after ex vivo aging for 4 h with or without added Dex. Additional neutrophils were cotreated with a glucocorticoid receptor (GR) antagonist (RU486) and Dex for 4 h. RNA was amplified, dye labeled (Cy3 or Cy5), and hybridized to a series of National Bovine Functional Genomics Consortium (NBFGC) microarrays. LOWESS data normalization followed by mixture model analyses showed that 11.15% of the spotted NBFGC cDNAs (2,036/18,263) were expressed in 4-h (untreated) neutrophils. Subsequent two-step mixed-model analysis detected (P < or = 0.05) 1,109 differentially expressed genes, of which contrast analysis indicated those that were independently responsive to aging (1,064), Dex (502), RU486 + Dex (141), or RU486 (357). In silico analysis revealed that 416 of the differentially expressed genes are unknown, 59 did not cluster well based on known function, and 634 clustered into 20 ontological categories. Independent validation of differential expression was done for 14 of the putatively Dex-responsive genes across these categories. Results showed that Dex induced rapid translocation of GR into the neutrophil nucleus and signaled dramatic alterations in expression of genes that delay apoptosis, enhance bactericidal activity, and promote tissue remodeling without inflammation or fibrosis. Thus these findings revealed hitherto unappreciated plasticity of blood neutrophils and potentially novel anti-inflammatory/wound-healing actions of glucocorticoids.

Comparison of ovarian function and circulating steroids in estrous cycles of Holstein heifers and lactating cows.

Ovarian function was compared between nulliparous heifers (n = 29; 10 to 16 mo old) and lactating Holstein cows (n = 31; 55.9 +/- 3.5 d postpartum). Follicular dynamics, corpus luteum growth, and regression, and serum steroid concentrations were evaluated through ultrasonography and daily blood sampling. Most heifers (27 of 29) but only 14 of 31 cows had typical spontaneous estrous cycles after cycles were initiated. Twelve cows had atypical cycles, and 5 became anovulatory during the study. The 12 cows with atypical estrous cycles had low serum estradiol after luteolysis and failed to ovulate the dominant follicle present at luteolysis. Heifers and cows with typical cycles were compared directly. Interovulatory intervals were similar between heifers (22.0 +/- 0.4 d) and cows (22.9 +/- 0.7 d). Those animals had estrous cycles with either 2 (15 heifers; 11 cows), 3 (9 heifers; 2 cows), or 4 follicular waves (3 heifers; 1 cow). Cows ovulated later after luteolysis than heifers (5.2 +/- 0.2 vs. 4.6 +/- 0.1 d, respectively), and had more multiple ovulations (17.9 vs. 1.9%). Maximal serum estradiol concentration preceding ovulation was lower in cows than in heifers (7.9 +/- 0.8 vs. 11.3 +/- 0.6 pg/mL) even though ovulatory follicles were larger in cows (16.8 +/- 0.5 vs. 14.9 +/- 0.2 mm). Similarly, maximal serum progesterone concentration was lower for cows (5.6 +/- 0.5 vs. 7.3 +/- 0.4 ng/mL), whereas maximal volume of luteal tissue was larger for cows than heifers (11,120 +/- 678 vs. 7303 +/- 308 mm3). Thus, higher incidence of reproductive anomalies in lactating cows, such as low conception rate, ovulation failure, delayed ovulation, and multiple ovulations, may be due to lower circulating steroid concentrations in spite of larger ovulatory follicles and luteal structures.

Improvement in recovery of embryos/ova using a shallow uterine horn flushing technique in superovulated Holstein heifers.

The aim of this study was two-fold: (1). to compare recovery of embryos/ova from superovulated Holstein heifers by flushing the uterine horns through insertion of the catheter very close to the tip of the horn (deep) or just after the uterine bifurcation (shallow) and (2). to evaluate the hormonal and superovulatory response to estradiol benzoate (EB) treatment prior to superovulation. Ten Holstein heifers (12-16 months) underwent two superovulatory treatments in a cross-over design. Heifers were treated with decreasing doses of FSH from Days 8 to 12.5 of a synchronized estrous cycle. At 4 days prior to superovulation, half of the heifers received EB (5mg, i.m.) or served as Controls, followed by the alternative treatment in the subsequent superovulation. At embryo recovery, one uterine horn was flushed with deep ( approximately 7 cm caudal to the tip of the horn) and the other with shallow ( approximately 5 cm cranial to the beginning of the uterine bifurcation) flushing techniques. Embryos/ova were recovered, counted, and scored. Number of ovulations was estimated by ultrasound. Pretreatment with EB reduced circulating FSH and regressed the first wave dominant follicle with no change in number of large follicles, number of ovulations, number of embryos/ova recovered, or number of transferable embryos. The shallow flushing technique was superior to the deep technique for number of embryos/ova recovered per horn (5.4+/-1.1 versus 3.9+/-0.8) or percentage of embryos/ova recovered per CL (63.9+/-8.6% versus 37.4+/-6.5%). Thus, flushing the entire uterine horn increased recovery of embryos/ova.

Ovarian structures and circulating steroids in heifers and lactating cows in summer and lactating and dry cows in winter.

Two experiments compared follicular and luteal development and circulating steroid concentrations from induced luteolysis to ovulation in lactating Holstein cows (n = 27; 40.0 +/- 1.5 kg milk/day) vs. nulliparous heifers (n = 28; 11 to 17 mo-old) during summer (Experiment 1), and in lactating (n = 27; 45.9 +/- 1.4 kg milk/d) vs. dry cows (n = 26) during winter (experiment 2). All females received PGF2,, 6 d after ovulation and were monitored until next ovulation by daily ultrasound and assay of serum progesterone (P4) and estradiol (E2). Every female was used two or three times. In Experiment 1, lactating cows had high incidence of multiple ovulation (63.5%) compared with heifers (1.3%). Among single ovulators, there was no difference in maximal size of ovulatory follicles between lactating cows and heifers (15.8 vs. 16.5 mm, respectively). However, lactating cows had lower peak serum E2 (8.6 vs. 12.1 pg/ml), took longer to ovulate after luteolysis (4.6 vs. 3.8 d), developed more luteal tissue volume (7,293.6 vs. 5,515.2 mm3), and had lower serum P4 on d 6 after ovulation (2.0 vs. 3.0 ng/ml) than heifers (data included multiple ovulators). In experiment 2, multiple ovulations were similar between lactating and dry cows (17.9 vs. 17.2%, respectively). Peak serum E2 was also similar between lactating and dry cows (7.6 vs. 8.5 pg/ml) although lactating cows had larger ovulatory follicles (18.6 vs. 16.2 +/- 0.4 mm). Lactating cows took longer to ovulate (4.8 vs. 4.2 d), developed more luteal tissue (7,599 vs. 5,139 +/- 468 mm3), but had similar serum P4 (2.2 vs. 1.9 ng/ ml) compared with dry cows. Therefore, lactating cows had similar or lower circulating steroid concentrations than dry cows or heifers, respectively, despite having larger ovarian structures.

Avaliaçöes ultra-sonográfica, macroscópica e histológica da biopsia testicular em ovinos / Assessment of ultrasonographic images and gross and microscopic lesions of the testicular biopsy in sheep

Devido ao fato da biopsia testicular poder acarretar hemorragia, inflamaçäo, degeneraçäo, aderência e fibrose, especialmente com as técnicas incisionais ou abertas, este trabalho avaliou a aplicabilidade de uma técnica menos invasiva (biopsia com agulha Tru-Cut) em ovinos na obtençäo de material para histologia e acompanhou as lesöes testiculares posteriores. Trinta carneiros foram igualmente divididos em três grupos: 1) controle, animais näo submetidos à biopsia; 2) submetidos à biopsia + cola de fibrina nos locais da biopsia testicular e incisöes da pele; e 3) submetidos à biopsia + sutura da pele escrotal após a biopsia. Exames ultra-sonográficos foram realizados pré e pós biopsia. No centésimodia, os testículos foram avaliados macro e microscopicamente após orquiectomia. A ultra-sonografia permitiu mapear as alteraçöes ocorridas e acompanhar a evoluçäo das seqüelas. Ao exame macroscópico, pequenas áreas de calcificaçäo foram observadas em 55 e 70 por cento dos testículos nos grupos 2 e 3, respectivamente. A biopsia com agulha Tru-Cut forneceu material suficiente para histologia mas ocasionou lesöes focais restritas à área biopsiada. Apesar da possível ocorrência de calcificaçäo e outras lesöes mínimas, foi demonstrado que a biopsia com agulha Tru-Cut em ovinos é um procedimento seguro por näo ter comprometido significativamente as características estruturais e funcionais dos testículos

Effect of gamma irradiation on the behavioral properties of crotoxin

Crotoxin has been detoxified with gamma radiation in order to improve crotalic antiserum production. Nevertheless, present knowledge of the biological characteristics of irradiated crotoxin is insufficient to propose it as an immunizing agent. Crotoxin is known to increase the emotional state of rats and to decrease their exploratory behavior (Moreira EG, Nascimento N, Rosa GJM, Rogero JR and Vassilieff VS (1996) Brazilian Journal of Medical and Biological Research, 29: 629-632). Therefore, we decided 1) to evaluate the effects of crotoxin in the social interaction test, which has been widely used for the evaluation of anxiogenic drugs, and 2) to determine if irradiated crotoxin induces behavioral alterations similar to those of crotoxin in the social interaction, open-field and hole-board tests. Male Wistar rats (l8O-220 g) were used. Crotoxin (100, 250, and 500 mug/kg) was injected intraperitoneally 2 h before the social interaction test. Similarly, irradiated crotoxin (2000 Gy gamma radiation from a 60Co source) was administered at the doses of 100, 250, and 500 mug/kg for the hole-board test, and at the doses of 1000 and 2500 mug/kg for the open-field and social interaction tests. ANOVA complemented with the Dunnett test was used for statistical analysis (P<0.05). Crotoxin decreased the social interaction time(s) at the doses of 1OO, 250 and 500 mug/kg (means + SEM) from 51.6 ñ 4.4 to 32.6 ñ 3.7,28.0 ñ 3.6 and 31.6 ñ 4.4, respectively. Irradiated crotoxin did not induce behavioral alterations. These results indicate that 1) crotoxin may be an anxiogenic compound, and 2) in contrast to crotoxin, irradiated crotoxin was unable to induce behavioral alterations, which makes it a promising compound for the production of crotalic antiserum.

Crotoxin-induced behavioral effects in rats

Crotoxin is the major component of Crotalus durissus terrificus venom. In view of the presence of high-affinity specific binding sites for crotoxin in the brain, the objective of this work was to investigate whether crotoxin induces behavioral effects in the open-fleld and hole-board tests. Adult male Wistar rats (l80-220 g) treated with crotoxin, 1OO, 250 and 500 mug/kg, ip, administered 2 h before the test, presented statistically significant behavioral alterations (ANOVA for one-way classification complemented with Dunnet test, P<0.05). In the open-field test, 250 and 500 mug/kg of crotoxin increased freezing (from 3.22 sec to 10.75 sec) and grooming (from 13.44 sec to 22.75 sec and 21.22 sec) and decreased ambulation (from 64.8 to 39.38 and 45.8). The dose of 500 mug/kg also decreased rearing (from 24.9 to 17.5). In the hole-board test, 500 mug/kg of crotoxin decreased head-dip count (from 6.33 to 4.00). All the crotoxin- induced behavioral effects were antagonized by an anxiolytic dose of diazepam (1.5 mg/kg, ip, 30 min before the tests). These results show that crotoxin reduced open-field activity and exploratory behavior as well. We suggest that these effects express an increased emotional state induced by this toxin.