Polymorphisms in the melatonin receptor gene promoter and their associations with fertility characteristics in buffalo herd in Eastern Amazon.

Buffalo production is spreading globally because of its economic advantage. Then, it has become necessary to improve the reproductive and productive efficiency of these animals, as well as to look for genetic factors that increase this efficiency. The objectives of this study were to characterize the promoter region of the melatonin 1A receptor gene (MTRN1A), to detect possible SNPs and associate them with fertility characteristics, and identify binding sites of transcription factors involved in the regulation of genetic expression in buffaloes in the Amazon. The conventional PCR method was carried out using the two primers designed from the reference sequence deposited in the GenBank AY52466.1. The products of the PCRs were purified, sequenced, and subsequently edited and aligned. Twenty-six SNPs were found, where 73% presented allele frequencies of wild nucleotides above 0.5, and 73% presented deviations from the Hardy-Weinberg equilibrium (P < 0.05) and FIS varying between 0.06 and 1.00, characterizing high degrees of inbreeding within the population. A block of ACAA deletion (position -1483) was observed in 25% of samples. The associations between these SNPs and reproductive characteristics were observed for calving interval and 5 SNPs: -1289, -1139, -911, -724, and -656 (P < 0.05), and three other SNPs: -1395, -724, and -94 (P < 0.05) were associated significantly with age at first calving, and were not associated with calving concentration. The promoter region was characterized by the different types of binding factors, where only 11 sites are significantly strong enough for transcription factor bindings. The ACAA deletion also exhibited a strong association with transcription factors. As a result, it would be necessary to test the SNPs above with other reproductive characteristics of economic relevance to approve the gene as a strong candidate for the selection of buffaloes in the Amazon.

Novel polymorphism in exon 1 of the melatonin receptor gene unassociated with reproductive characteristics of buffaloes in the Amazon Region.

The objective of this study was to sequence part of the exon 1 in the melatonin receptor 1A gene (MTRN1A) in buffaloes to detect a novel polymorphism with which to associate reproductive characteristics, such as age at first birth and the interval between births, in buffaloes from the northeastern region of the State of Pará (Brazil). Buffalo hair samples (77) were collected from the Terra Firme region of Pará. DNA was extracted and polymerase chain reactions (PCRs) were carried out with a primer that was designed using the GenBank accession No. AY524665 reference sequence. PCR products were purified and sequenced. After editing and analysis of the sequences, a mutation was observed at the 62nd position in exon 1 of MTRN1A (T↔C), which corresponded with a change in the 21st amino acid from leucine to proline. All possible genotypes were observed, with the most common being genotype CC (0.481). The allele frequencies were T = 0.377 and C = 0.623. Statistical analysis of FIS showed inbreeding within the sample group (FIS = 0.397) and deviations from the Hardy- Weinberg equilibrium were observed (P < 0.05). Associations between genotypes and reproductive characteristics were not significant (P > 0.05). Although the related SNP was not synonymous, there were no observable effects on the reproductive characteristics under investigation. As such, it would be ideal to detect other SNPs in exon 1 of the MTRN1A gene that can be associated with reproductive characteristics in Amazonian buffaloes.

Corrigendum Polymorphism in the melatonin receptor gene in buffalo populations of the Brazilian Amazon - Genet. Mol. Res. 15 (2): gmr.15027960.

The correction is only in the name of the first author and should be: E.M. Barbosa(1), B.B. Souza(2), R.C. Guimarães(2), J.S.N. Azevedo(3), E.C. Gonçalves(4), H.F.L. Ribeiro(2), S.T. Rolim Filho(2), E. Silva Filho(2).

Polymorphism in the melatonin receptor gene in buffalo populations of the Brazilian Amazon.

Buffalo farming in Brazil is increasing, as is the challenge of identifying molecular markers that will improve productivity. Therefore, the aim of this study was to analyze single nucleotide polymorphisms of the receptor gene for the hormone melatonin in buffaloes from northern Brazil by polymerase chain reactions (PCRs) and restriction fragment length polymorphism assays. The PCR products exhibited a cutting point for HpaI at the 318th position of the gene, indicating a transition substitution (T↔C). This substitution was synonymic, and did not alter the stability of the mRNA structure. Allelic and genotypic frequencies differed between the populations studied, and all of the populations demonstrated endogamy and were in Hardy-Weinberg equilibrium. Therefore, the HpaI restriction marker in the melatonin receptor gene cannot be used for genetic improvement, but is an excellent marker for population genetic studies.

Efeitos do trimetilaminoetano (TES) e ringer lactato em sêmen de macacos-aranha mantidos em cativeiro ( Ateles paniscus e A. marginatus) / Effects of TES and a ringer-lactato diluent in semen from captive spider monkeys (Ateles paniscus e A. marginatus)

The performances of the diluents TES and CEBRAN II were compared as cryopreservatives of semen from non human primates of the genus Ateles. The experiment was carried out using one Ateles marginatus and two Ateles paniscus specimens, males and adults, maintained in the same captivity conditions at the National Center of Primates (CENP-SVS/MS). The animals were subjected to clinical and andrological examinations - testicular biometry - before the semen collection by eletroejaculation. Evaluations of motility and forward movement in the fresh semen were made. Semen were made dilution was made with the diluents TES and CEBRAN II. The ejaculates were diluted with the diluents (2:1proportion), packed in 0.25mL plastic straws and cryopreserved in liquid nitrogen. After thawing, the packed ejaculates were appraised in thermo resistance test (TTR). The averages of volume and concentration were, respectively, 1.94mL (0.83) and 3,020,000 sptz/mL (275.97). The pH 8 and seminal coagulation were observed in all samples. The results suggest that the TES diluent presents better efficiency in the preservation of Ateles semen than CEBRAN II.

Identification of polymorphisms in the osteopontin gene and their associations with certain semen production traits of water buffaloes in the Brazilian Amazon.

The osteopontin gene may influence the fertility of water buffaloes because it is a protein present in sperm. The aim of this work was to identify polymorphisms in this gene and associate them with fertility parameters of animals kept under extensive grazing. A total of 306 male buffaloes older than 18 months, from two farms, one in the state of Amapá and the other in the state of Pará, Brazil were used in the study. Seven SNPs were identified in the regions studied. The polymorphisms were in gene positions 1478, 1513 and 1611 in the region 5'upstrem and positions 6690, 6737, 6925 and 6952 in the region amplified in intron 5. The SNPs were associated with the traits, namely scrotal circumference, scrotal volume, sperm motility, sperm concentration and sperm pathology. There were significant SNPs (p < 0.05) for all the traits. The SNP 6690 was significant for scrotal circumference, sperm concentration, sperm motility and sperm pathology and the SNP 6737 for scrotal volume. The genotype AA of SNP 6690 presented the highest averages for scrotal circumference, sperm concentration and motility and the lowest total number of sperm pathologies. For the scrotal volume trait, the animals with the largest volume were correlated with the presence of the genotype GG of SNP 6737. These results indicate a significance of the osteopontin gene as it seems to exert a substantial influence on the semen production traits of male buffaloes.

Characterization of folliculogenesis and the occurrence of apoptosis in the development of the bovine fetal ovary.

The aim of this research was to perform in situ quantification, morphometry evaluation, and apoptosis analysis of ovarian follicular wall cells in mechanically isolated follicles obtained from ovaries of bovine fetuses (Bos taurus indicus) between 3 and 9 months of age. Apoptosis was evaluated using the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. The number of isolated follicles increased from 3 months onward (102.5 ± 141.1, mean ± SEM), peaked at 6 months (12855.0 ± 9030.1), and then decreased by 7 months (3208.7 ± 3249.5), consistent with atresia occurring at these stages. Follicular density was greatest at 4 months, consistent with a sudden boost in follicular activity independent of a corresponding increase in ovarian size. Antral follicles were first observed at 5 months. As fetal age increased, there was a tendency for the percentage of primordial and primary follicles to decrease, and the percentage of secondary follicles to increase. However, the high variability (P < 0.05) for all follicle populations up to 5 months of age precluded further interpretation of these results. Oocyte diameter increased from the primordial (23.6 ± 4.4 µm) to the secondary follicular stages (38.0 ± 14.9 µm). Apoptosis was observed in ovaries from all fetal ages analyzed. We concluded that preantral follicles could be isolated from bovine fetuses by 3 months of age, with apoptosis affecting ovarian follicular dynamics throughout fetal life.

Níveis de progesterona no leite durante o ciclo estral da búfala / Milk progesterone profiles during the oestrus cycle of the water bufallo cow

Os níveis de progesterona (P4) no leite sem gordura, foram determinados através do método do Radioimunoteste (RIA) em fase sólida, durante 28 ciclos estrais ajustados com duraçäo entre 20 e 24 dias (x = 22,4 ñ 1,58 dias) em 11 vacas bubalinas. Os níveis de P4 no estro variaram ente 0,33 e 0,48 ng/ml, acusando, entre o 3§ e 5§ dia do ciclo, aumento que varia de 1,3 a 1,8 ng/ml. Os níveis de hormônio aumentaram gradativamente até 2,2 e 3,2 ng/ml entre o 7§ e 9§ dia e atingiram pique máximo de 3,5 a 5,8 ng/ml entre o 11§ e 13§ dia do ciclo. A partir do 15§ e 17§ um nível de 3,1 e 4,8 ng/ml foi observado o qual decresceu para 2,3 e 2,8 ng/ml entre o 19§ e 21§ dias e voltou ao nível basal de 0,4 ng/ml no estro subseqüente