Biochemical and microbiological characteristics of in situ biofilm formed on materials containing fluoride or amorphous calcium phosphate

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Molecular Evaluation of Developmental Competence of Oocytes Collected In Vivo from Buffalo and Bovine Heifers during Winter and Summer

Buffaloes and bovines are polyestrous and seasonal or annual livestock, respectively, that show reduced fertility during heat stress. To investigate whether reduced fertility is related to oocyte competence in both species, immature oocytes from buffalo and bovine heifers were collected during winter and summer and subjected to molecular analyses. In each season, heifers of both species had their follicular wave emergence synchronized with a standard protocol (Ferreira et al., 2011). Before being subjected to ovum pick up (OPU), cutaneous (CT; degrees C) and rectal (RT; degrees C) temperatures and respiratory rate (RR; breaths/min) were measured. Oocytes' RNA was extracted to evaluate the expression of target genes related to mtDNA replication/transcription (PPARGC1A, TFAM and MT-CO1), apoptosis (BAX and BCL2) and HS (HSP90AA1 and HSPA1AB). ACTB, HIST1H2AG and GAPDH were initially chosen as housekeeping genes. In buffaloes, CT (35.0 +/- 0.4 vs 23.8 +/- 0.5), RT (38.7 +/- 0.1 vs 38.0 +/- 0) and RR (21.3 +/- 1.2 vs 15.4 +/- 1.1) were higher during summer than winter. However, in bovine heifers, RT (38.7 +/- 0.1 vs 38.6 +/- 0.1) and RR (44.8 +/- 1.5 vs 40.6 +/- 1.5) were similar in both seasons, while CT (31.6 +/- 0.3 vs 30.2 +/- 0.3) was increased during summer. Reduced expression of ACTB, HIST1H2AG and GAPDH was evidenced during summer, disqualifying them as housekeeping genes. Similarly, the expression of all target genes was reduced during summer in oocytes of both species. In summary, physiological responses to heat stress seem to be more intense in buffalo than bovine heifers. However, in both species, negative effects of heat stress upon oocyte quality occur at the molecular level and affects genes related to several biological functions.

Productive and reproductive traits in Murrah breed from Brazil

Major objective of this study was to estimate heritability and genetic correlations between milk yield (MY) and calving interval (CI) and lactation length (LL) in Murrah buffaloes using Bayesian inference. The database used belongs to the genetic improvement program of four buffalo herds from Brazil. To obtain the estimates of variance and covariance, bivariate analyses were performed with the Gibbs sampler, using the program MTGSAM. The heritability coefficient estimates were 0.28, 0.03 and 0.15 for MY, CI and LL, respectively. The genetic correlations between MY and LL was moderate (0.48). However, the genetic correlation between MY and CI showed large HPD regions (highest posterior density interval). Milk yield was the only trait with clear potential for genetic improvement by direct mass selection. The genetic correlation between MY-LL indicates that indirect selection using milk yield is a potentially beneficial strategy. The interpretation of the estimated genetic correlation between MY-CI is difficult and could be spurious.

Genetic Parameter Estimates for Milk Yield and Lactation Length in Buffalo

The objective of this study was to estimate genetic parameters for milk yield at 244 days and lactation length in graded buffalo cows at the El Cangre Cattle Genetic Enterprise. Data were gathered from 2575 lactations, 1377 buffalo cows, 37 milking units and between 2002-2009 calving years. It was employed the Restricted Maximum Likelihood method (REML) for estimating (co) variance components with multi trait model. Average of milk yield at 244 days and lactation length were 864 kg and 240 days, respectively. Heritability was 0.15 for milk yield and 0.13 for lactation length. Genetic correlation between these traits was 0.63. It was concluded that it is necessary to intensify selection and to increase control of the information of the genetic herds to obtain high precision in the estimates and therefore, obtain bigger genetic progress in of this species in our country.

Genetic Parameters for Productive and Reproductive Traits for Milk Buffalo in Brazil

Knowing the genetic parameters of productive and reproductive traits in milking buffaloes is essential for planning and implementing of a program genetic selection. In Brazil, this information is still scarce. The objective of this study was to verify the existence of genetic variability in milk yield of buffaloes and their constituents, and reproductive traits for the possibility of application of the selection. A total of 9,318 lactations records from 3,061 cows were used to estimate heritabilities for milk yield (MY), fat percentage (%F), protein percentage (%P), length of lactation (LL), age of first calving (AFC) and calving interval (CI) and the genetic correlations among traits MY, %F and %P. The (co) variance components were estimated using multiple-trait analysis by Bayesian inference method, applying an animal model, through Gibbs sampling. The model included the fixed effects of contemporary groups (herd-year and calving season), number of milking (2 levels), and age of cow at calving as (co) variable (quadratic and linear effect). The additive genetic, permanent environmental, and residual effects were included as random effects in the model. Estimated heritability values for MY, % F, % P, LL, AFC and CI were 0.24, 0.34, 0.40, 0.09, 0.16 and 0.05, respectively. The genetic correlation estimates among MY and % F, MY and % P and % F and % P were -0.29, -0.18 and 0.25, respectively. The production of milk and its constituents showed enough genetic variation to respond to a selection program. Negative estimates of genetic correlations between milk production and its components suggest that selection entails a reduction in the other.

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.

Interference of Vaccinal Antibodies on Serological Diagnostic of Leptospirosis in Vaccinated Buffalo using Two Types of Commercial Vaccines

The vaccinal antibodies interference represents one of the problems in the leptospirosis diagnostic on serum. The present study aimed to determine the pattern of serum agglutinins anti-Leptospirae spp in vaccinated female buffaloes against leptospirosis using two types of commercial vaccines: bacterin and extern membrane. The temporal interference of vaccinal titers on serum diagnostic was evaluated. Three groups of 11 adult female buffaloes were established as follows: G1 control, non-vaccinated; G2: vaccinated with bacterin containing six serovars and G3 with extern membrane vaccine containing five serovars. A booster was administered at 30 days from the first vaccination (dfv) and two re- vaccinations were performed in each semester (days 210 and 390). Serum samples were collected on days 0, 15, 30, 45 and 60 and every 30 days until 540 dfv, being submitted to Serum Agglutination Microscopy (SAM) against the serovars present in the vaccine. G1 remained always negative. Both vaccines induced serologic responses when assessed by SAM at 150 days post first vaccination against all serovars and they revealed maximum titers around days 45 and 60 after first vaccination. At the re-vaccination there was an increase on agglutinin levels, but of less intensity than the levels previously observed. After six months from the second revaccination (540 dfv), they were almost zero, which demonstrates the short duration of diagnostic interference. The serologic monitoring of the vaccinated herds can be an efficient method to evaluate the status of protection provided by the vaccine.

Allometry of Organs and the Gastrintestinal Tract from Water Buffaloes (Bubalus bubalis) Finished in Feedlot

The objective of present study was to evaluate the relative growth of organs and viscera from water buffaloes. Fifteen Mediterranean intact males, averaging, 356.7 kg initial live weight and twenty four months of age, were used. The animals were ramdomly assigned into three groups (categories). One group was ramdomly assigned to immediate slaughter (AR), the rest two groups were full-fed a ration containing 50% concentrate, dry matter basis until reaching the slaughter weights of 450 and 500kg, respectively. At slaughter the empty body weight was determined and the weights of head, feet, leather, lungs, heart, liver, spleen, rumen-reticulum, omasum, abomasum, small intestine, large intestine were recorded. Regression equations of log weight of organs and viscera as a function of log empty-body-weight (EBW), were fitted. All body components studied, with exception of liver and spleen, developed slower than in relation to EBW.

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.

Single Nucleotide Variations in the Buffalo Kappa-Casein Gene (CSN3)

Caseins are the major milk proteins associated with lactation performance, milk composition and cheese yield efficiency, representing around 80% of the total amount of proteins found in milk. Among the caseins, kappa-casein is the protein that stabilizes micelle structure during milk coagulation process and being used during the cheese production. The kappa-casein gene (CSN3) has been previously mapped to buffalo chromosome 7 using a radiation hybrid panel and a comparative map was established using the sequence from bovine chromosome 6. The molecular structure of this gene has also been established in river buffalo, with a total length of 13,191 bp (GenBank: AM900443.1) and containing five exons. In this study we searched for single nucleotide variations in specific regions of the CSN3 gene in three animals representing the Murrah breed. Sequencing reactions were performed using ABI3730xl sequencer. The primer walking method was used to span the 5'-UTR and intron 2 regions of the gene, for which ten primer pairs were designed using Oligo 6 software. BLAST tool was used to verify the primers specificity. DNA sequences assemblies from all three animals were performed with Sequencher (R) software 4.1, while multiple alignments were performed using Clustal W software to identify single nucleotide variations. The sequencing revealed a total of 19 single nucleotide variations with 13 located in the upstream regulatory region of the gene (5'-UTR) and six on intron 2. These variations can be validated using commercial populations segregating specific economic traits.

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.

Control of Buffalo Follicular Dynamics for Artificial Insemination, Superovulation and In Vitro Embryo Production

Currently, timed ovulation induction and timed artificial insemination (TAI) can be performed in buffalo using GnRH or estradiol plus progesterone/progestin (P4)-releasing devices and prostaglandin F-2 alpha (PGF(2 alpha)). The control of the emergence of follicular waves and of ovulation at predetermined times, without the need for estrus detection, has facilitated the management and improved the efficiency of AI programs in buffalo during the breeding and nonbreeding season. Multiple ovulations, embryo transfer, ovum collection and in vitro embryo production have been shown to be feasible in buffalo, although low efficiency and limited commercial application of these techniques have been documented as well. These results could be associated with low ovarian follicular pools, high levels of follicular atresia and failures of the oocyte to enter the oviduct after superstimulation of follicular growth. This review discusses a number of key points related to the manipulation of ovarian follicular growth to improve pregnancy rates following TAI and embryo transfer of in vivo- and in vitro-derived embryos in buffalo.