Glucose, Cholesterol, Total Protein and Growth Factor Insulin-like Type I in Follicular Fluid of River Buffaloes (Bubalus bubalis)

In order to establish the concentrations of glucose, cholesterol, total protein and growth factor insulin-like type I (IGF-I) in the follicular fluid, 26 Murrah breed river buffaloes, between 45 and 70 days postpartum, empty, multiparous, with average live weight of 675 +/- 56 kg and average body condition of 3.5 points on a scale of 1-5, were used in this study. The fluid was collected from dominant follicles with diameters between 8 and 12 mm by OPU, and was not taken into account the stage of the estrous cycle. Using this technique, the wave of follicular development was synchronized six days prior to collection. Biochemical analysis was performed to glucose and cholesterol through the enzymatic colorimetric method using commercial kit glicose CHOLESTEROL GOD-PAP and CHOD-PAP (Kovalent), respectively. Determination of total protein was carried out by using total protein commercial kit (Kovalent) Biuret method, and the readings were performed using absorption spectrophotometry with visible light. Concentration of IGF-I was measured by Radioimmunoassay (RIA) technique using commercial IRMA Kit IGF-I (INMUNOTECH). Descriptive statistics were developed using the PROC MEANS procedure of SAS (2009). Concentration of glucose (4.0 +/- 0.75 mmol / L-1) and IGF-I (340 +/- 129.83 ng / mL (-1)) were higher than those reported by other authors in river buffaloes and cows, respectively. However, cholesterol levels (0.51 +/- 0.12 mmol / L (-1)) and total protein (58.4 +/- 4.43 g / L (-1)) behaved inferior to other studies in same species. The results indicated that there is relationship among the nutritional aspects, diameter of follicles aspirated and productive period in the concentration of biochemical indicators.

Genetic parameters for body weight in buffaloes (Bubalus bubalis) in Colombia using random regression models

The objective of this research was to estimate (co) variance functions and genetic parameters for body weight in Colombian buffalo populations using random regression models with Legendre polynomials. Data consisted of 34,738 weight records from birth to 900 days of age from 7815 buffaloes. Fixed effects in the model were contemporary group and parity order of the mother. Random effects were direct and maternal additive genetic, as well as animal and maternal permanent environmental effects. A cubic orthogonal Legendre polynomial was used to model the mean curve of the population. Eleven models with first to sixth order polynomials were used to describe additive genetic direct and maternal effects, and animal and maternal permanent environmental effects. The residual was modeled considering five variance classes. The best model included fourth and sixth order polynomials for direct additive genetic and animal permanent environmental effects, respectively, and third-order polynomials for maternal genetic and maternal permanent environmental effects. The direct heritability increased from birth until 120 days of age (0.32 +/- 0.05), decreasing thereafter until one year of age (0.18 +/- 0.04) and increased again, reaching 0.39 +/- 0.09, at the end of the evaluated period. The highest maternal heritability estimates (0.11 +/- 0.05), were obtained for weights around weaning age (weaning age range is between 8 and 9.5 months). Maternal genetic and maternal permanent environmental variances increased from birth until about one year of age, decreasing at later ages. Direct genetic correlations ranged from moderate (0.60 +/- 0.060) to high (0.99 +/- 0.001), maternal genetic correlations showed a similar range (0.41 +/- 0.401 and 0.99 +/- 0.003), and all of them decreased as time between weighings increased. Direct genetic correlations suggested that selecting buffalos for heavier weights at any age would increase weights from birth through 900 days of age. However, higher heritabilities for direct genetic weights effects after 600 days of age suggested that selection for these effects would be more effective if done during this age period. A greater response to selection for maternal ability would be expected if selection used maternal genetic predictions for weights near weaning. (C) 2013 Elsevier B.V. All rights reserved.

Genetic Parameters for Reproductive Traits of Crossbred Buffaloes from Brazil, Estimated by Bayesian Inference

The objective of the study was to estimate heritability for calving interval (CI) and age at first calving (AFC) and also calculate repeatability for CI in buffaloes using Bayesian inference. The Brazilian Buffaloes Genetic Improvement Program provided the database. Data consists on information from 628 females and four different herds, born between 1980 and 2003. In order to estimate the variance, univariate analyses were performed employing Gibbs sampler procedure included in the MTGSAM software. The model for CI included the random effects direct additive and permanent environment factors, and the fixed effects of contemporary groups and calving orders. The model for AFC included the direct additive random effect and contemporary groups as a fixed effect. The convergence diagnosis was obtained using Geweke that was implemented through the Bayesian Output Analysis package in R software. The estimated averages were 433.2 days and 36.7months for CI and AFC, respectively. The means, medians and modes for the calculated heritability coefficients were similar. The heritability coefficients were 0.10 and 0.42 for CI and AFC respectively, with a posteriori marginal density that follows a normal distribution for both traits. The repeatability for CI was 0.13. The low heritability estimated for CI indicates that the variation in this trait is, to a large extent, influenced by environmental factors such as herd management policies. The age at first calving has clear potential for yield improvement through direct selection in these animals.

Population Parameters Based on Known Pedigree Records from Brazilian Jaffarabadi Buffaloes

The objective of this study was to evaluate the effective number of founders and ancestors, generation intervals and completeness of pedigree in Jaffarabadi breed buffaloes raised in Brazil. Pedigree records of 1,272 animals born from 1966 were used. The parameters were estimated using ENDOG, computational population genetic software. The obtained value for completeness of pedigree was 99.5, 50.9, and 20.5 for, the first, second and third generations, respectively. Generation interval estimates expressed in years and considering different pathways were 12.28 +/- 6.90 (sire-son), 11.55 +/- 6.07 (sire-daughter), 8.20 +/- 2.63 (dam-son) and 8.794 +/-.33 (dam-daughter). The overall average generation interval was 10.17 +/- 5.43 years. The number of founders, equivalent founders and ancestor animals that contributed for the genetic diversity in the reference population (1059) were 136, 130 and 134, respectively. Effective number of founder (f(e)=8) and ancestors (f(a)=7) were small, and the calculated expected inbreeding increase per generation was 4.99%. Four ancestors explained 50% of the genetic variability in the population and the major ancestor contributed with approximately 33% of the total population genetic variation. The genetic diversity within the current population is low as a consequence of a reduced number of ancestors.

Genetic Parameters for Milk Yield and Lactation Length of Crossbred Buffaloes from Brazil by Bayesian Inference

The objective of the study was to estimate heritability and repeatability for milk yield (MY) and lactation length (LL) in buffaloes using Bayesian inference. The Brazilian genetic improvement program of buffalo provided the data that included 628 females, from four herds, born between 1980 and 2003. In order to obtain the estimates of variance, univariate analyses were performed with the Gibbs sampler, using the MTGSAM software. The model for MY and LL included direct genetic additive and permanent environment as random effects, and contemporary groups, milking frequency and calving number as fixed effects. The convergence diagnosis was performed with the Geweke method using an algorithm implemented in R software through the package Bayesian Output Analysis. Average for milk yield and lactation length was 1,546.1 +/- 483.8 kg and 252.3 +/- 42.5 days, respectively. The heritability coefficients were 0.31 (mode), 0.35 (mean) and 0.34 (median) for MY and 0.11 (mode), 0.10 (mean) and 0.10 (median) for LL. The repeatability coefficient (mode) were 0.50 and 0.15 for MY and LL, respectively. Milk yield is the only trait with clear potential for genetic improvement by direct genetic selection. The repeatability for MY indicates that selection based on the first lactation could contribute for an improvement in this trait.

Polymorphisms in the MTRN1A gene and their effects on the productive and reproductive traits in buffaloes

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Application of Reactions Norms in Study of Genotype Environmental Interaction for Milk Yield of Buffaloes

The aim of this study was to estimate genetic parameters for milk yield (MY) in buffaloes using reaction norms. Model included the additive direct effect as random and contemporary group (herd and year of birth) were included as fixed effects and cow age classes (linear) as covariables. The animal additive direct random effect was modeled through linear Legendre polynomials on environment gradient (EG) standardized means. Mean trends were taken into account by a linear regression on Legendre polynomials of environmental group means. Residual variance was modeled trough 6 heterogeneity classes (EG). These classes of residual variance was formed : EG1: mean = 866,93 kg (621,68 kg-1011,76 kg); EG2: mean = 1193,00 kg (1011,76 kg-1251,49 kg); EG3: mean = 1309,37 kg (1251,49 kg -1393,20 kg); EG4: mean = 1497,59 kg (1393,20 kg-1593,53 kg); EG5: mean = 1664,78 kg (1593,53 kg -1727,32kg) e EG6: mean = 1973,85 kg (1727,32 kg -2422,19 kg).(Co) variance functions were estimated by restricted maximum likelihood (REML) using the GIBBS3F90 package. The heritability estimates for MY raised as the environmental gradient increased, varying from 0.20 to 0.40. However, in intermediate to favorable environments, the heritability estimates obtained with Considerable genotype-environment interaction was found for MY using reaction norms. For genetic evaluation of MY is necessary to consider heterogeneity of variances to model the residual variance.

Polymorphisms in Osteopontin Gene in Amazon Buffaloes

The aim of this work was to identify polymorphisms in the osteopontin gene. It was used in this experiment 306 male buffaloes, older than 18 months, bred in two farms, one in the State of Amapa and the other farm in the State of Para. There was identified three SNP polymorphisms for the region amplified by the primer OS4 (5'upstream) and four SNP polymorphisms for the region amplified by the primer OS9 (exon 5 to exon 6). The polymorphisms were in positions 1478, 1513 and 1611 in the region amplified by OS4 and positions 6690, 6737, 6925 and 6952 in the region amplified by OS9. These data indicate that the osteopontin gene is important because it can have a substantial influence on the reproductive traits of male buffaloes.

Bone Marrow'S Harvest in The Coxal Tuberosity for Isolation and Culture of Mesenchymal Stem Cells of Buffaloes (Bubalus Bubalis)

Several studies with mesenchymal stem cells (MSCs) have been developed in many species because of its ability to differentiate into other mesoderm lineages, capacity of self-regeneration, low immunogenicity, paracrine, anti-inflamatory, immunomodulatory and antiapoptotic effects which make then a promissory source to be used in therapeutic strategies. The aim of this study is to report the technique of harvest of bone marrow (BM) in the coxal tuberosity (CT) of buffaloes. For this, the animals were selected, identified and contained in a stock. Then trichotomy was performed in the region corresponding to the CT. After identifying the anatomic site it was performed antisepsis, local anesthetic block and introduction of a myelogram's needle (Lang(R)) for BM aspiration. Once the needle was firmly fixed in the CT, the mandril was removed and proceeded to BM aspiration with a syringe (20 mL) containing 1 ml of heparin at 1000 IU / mL and 1 mL of PBS. After the collection, each sample collected was manually homogenized, identified and referred to the LRACT - FMVZ / UNESP-BRAZIL for the correct processing. The anatomical site tested showed to be an alternative site of harvest of BM once provided the appropriate isolation and culture of the mononuclear fraction. Moreover, the procedure was performed without difficulty and with great security. Based on this, it can be conclude that CT is an excellent anatomical site for isolation and culture of MSCs and the proposed technique is viable and feasible to be held in buffaloes.

Isolation, Culture and Differentiation of Buffaloes Bone Marrow Mesenchymal Stem Cells Obtained from the Coxal Tuberosity

Currently, much attention has been devoted to the renewal of knowledge about Stem Cells and Cell Therapy in domestic species. In this sense, the present work aimed to develop a methodology for collecting, processing and cultivation of mesenchymal stem cells obtained from bone marrow of coxal tuberosity in buffaloes. The collection was performed using a Komiyashiki needle, which was introduced in the coxal tuberosity and the bone marrow aspirated into a heparinized syringe with the aid of negative pressure. Directly after collection samples were processed at the laboratory at FMVZ - UNESP. The samples took approximately 32 days to reach 80% confluence, when the first passage and differentiation was performed. To confirm the mesenchymal origin, cells were induced to differentiate into adipogenic and osteogenic lineages. Samples showed morphological changes during differentiation protocol, but not all presented production of extracellular deposits of calcium or intracellular fat droplets, observed after staining with Alizarin Red and Oil Red respectively. Compared with the material obtained from other species and processed in the same laboratory, the primary culture was longer. Therefore, more studies are needed to standardize the age of animals used and to test other inducers of cell differentiation.

Scrotal Thermography and Doppler Ultrasonography of the Testicular Artery of Buffaloes Subjected to Environmental Heat Stress

The process of spermatic division and differentiation (spermatogenesis) occurs with intratesticular temperature lower that the corporal temperature and for that is essential that the testicular thermoregulation mechanism occurs properly. For evaluation of the scrotal surface temperature can be used the infrared thermography or testicular sensors, besides that, can be evaluated the blood flux in the spermatic cord through the Doppler ultrasonography. Therefore the objective of this study was the evaluation of the scrotal thermography and Doppler flowmetry of the testicular artery of buffaloes subjected to environmental heat stress. For that were used seven healthy buffaloes, with age of 3 and 4 years, of the Murrah breed. For the surface scrotal temperature measurement (SST, degrees C) and superficial neck temperature (SNT, degrees C) was used the infrared termography (Infra Cam (TM) of the brand FLIR Systems Inc.), then Doppler flowmetry of the testicular artery in the region of the spermatic cord through the ultrasonography (Mylab 5, Esaote (R)) and measurement of the rectal temperature (RT, degrees C). The evaluations were done in two moments: moment 1 (M1) with all the animals in the shade (Temperature=32,2 degrees C) and moment 2 (M2) after 3 hours of exposure of animals to the sun (Temperature=38,7 degrees C To calculate the resistivity index (RI) and pulsatility index (PI), spectra were obtained from pulsed Doppler in three random regions of the testicular artery in the spermatic cord. Data were subjected to analysis of variance (ANOVA) followed by T test, using a significance level of 5%. There was an increase (p<0,05) of RT (37,4 +/- 0,4(a) vs 39,0 +/- 0,3(b); M1 and M2 respectively), SST (30,6 +/- 1,4(a) vs 35,2,0 +/- 1,0(b); M1 and M2 respectively) and SNT (33,1 +/- 2,5(a) vs 38,5,0 +/- 0,3(b); M1 e M2 respectively) e RI (0,67 +/- 0,1(a) vs 0,74 +/- 0,1(b); M1 e M2 respectively) in M2. Increasing trend was observed (0,05>p>0,01) in PI (1,10 +/- 0,4(a) vs 1,23 +/- 0,2(b); M1 and M2 respectively) in M2. The results of the present study allow us to conclude the healthy buffaloes have the scrotal average surface temperature 3 degrees C lower that the body temperature and that the exposure of 3 hours to sun in healthy buffaloes causes thermal stress to the animals and changes in its surface scrotal temperature, and the Doppler flowmetry of the testicular artery demonstrating the importance of thermal management for breeding buffaloes. Besides that, the thermography and the Doppler ultrasonography presented great potential to detect changes of testicular perfusion, being a promising additional test in the buffalo andrological evaluation.

Morphometric and Functional Characteristics of Buffaloes and Cows Corpus Luteum at Different Stages of the Estrous Cycle

The aims of the present study were to evaluate the morphometry of corpus luteum (CL) and progesterone (P-4) plasma concentration of 86 buffaloes (33 pregnant and 53 non-pregnant) and 95 cows (36 pregnant and 59 non-pregnant) at the moment of slaughter. Seventy CLs of buffaloes and 110 CL of cattle were analyzed. The CL classified as II and III were more common in both species (35.7 and 41.4% for buffaloes and 43.6 and 35.5% for cows). The 29 non-pregnant buffaloes had a total of 36 CL, being 19.4% CLI; 33.3% CL II; 27.8% CL III and 19.4% CL IV. The 51 nonpregnant cows had a total of 71 CL, being 26.8% CL I; 47.9% CL II; 21.1% CL III and 4.2% CL IV. The average diameters of bubaline and bovine CL were 5.2 +/- 0.9 and 6.4 +/- 1.8 mm (CL I); 17.6 +/- 2.6 and 19.8 +/- 3.2 mm (CL II); 17.2 +/- 2.1 and 20.0 +/- 3.2 mm (CL III); 7.8 +/- 1.8 and 8.7 +/- 2.7 mm (CL IV), respectively. The mean plasma concentrations of P4 were 5.6 (CL I); 5.4 (CL II); 4.7 (CL III) and 0.5 (CL IV) ng/mL for buffaloes and 0.02 (CL I); 6.3 (CL II) and 6.4 (CL III) ng/mL for cows. In both species, P4 concentration was similar between stages II and III. The results indicated that the characterization of the CL provides important information about the status of estrous cycle.

Inbreeding, Average Relatedness Coefficient and Effective Population Size In Jaffarabadi Buffaloes Raised In Brazil

The major aim of this study was to evaluate the inbreeding (F), average relatedness coefficient (AR) and effective population size (Ne) in the Jaffarabadi buffalo breed from Brazil. Pedigree information of 1,272 animals born from 1966 was used. The effective population size was calculated in two ways: first, computed via individual increase in inbreeding and second estimated by individual increase in coancestry. The known generation numbers were 1.24, 1.76 and 2.64 for complete, equivalent and maximum generation, respectively. The effective size computed via individual increase in coancestry was small with a value of 10.82 +/- 1.29. The effective size computed by individual increase in inbreeding (10.40 +/- 3.69) was very similar but a little smaller than the previous reported value. The average values of F and AR for the population reference (1,059) were 4.22 and 12.5 percent. The mean of F for inbred animals (319) was 14.0%. The F and AR means were 5.7 and 13.3% for animals with at least 1.5 known equivalent generation and 9.3 and 15.97% for individuals having at least 2.5 equivalent generations known. It was found 78 matings between half sibs (6.14%) and 67 matings (5.27%) between parent-offspring. The estimated inbreeding increase per generation by considering maximum generation, complete generation and equivalent generation were 1.21%, 5.18% and 3.57%, respectively. Considering the uncompleted pedigree, the estimated inbreeding for the reference population could be underestimated.