Association analysis of novel polymorphisms in 2′, 5′‐oligoadenylate synthetase gene with reproductive traits in indigenous and cross‐bred cattle of Indian Origin

2′, 5′‐Oligoadenylate synthetases (OAS) are important components of an interferon‐mediated antiviral pathway. No polymorphisms in exonic regions of bovine OAS1 gene have been identified and associated with reproduction traits. The objective of the study was to detect and evaluate the effects of mutations in exonic region of bovine OAS1 gene with reproduction traits in cattle. DNA samples collected from 250 individual cows of two Indian dairy breeds (Sahiwal and Frieswal) of cattle were used in the study. The genetic variants of the OAS1 gene were identified with polymerase chain reaction–single‐strand conformation polymorphism (PCR‐SSCP) and sequence analysis using seven set of primer pairs. The PCR‐SSCP analysis revealed polymorphism in the fragments comprising of exon 2, exon 5 and first fragment of exon 6 while the fragments of exons 1, 3, 4 and second fragment of exon 6 were monomorphic in Sahiwal and Frieswal cattle. The mutations in the amplified region comprising of exon 2 were found to have significant association with age at first breeding and calving, service period, dry period and pregnancy rate. Significant associations were found between SNPs in the exon 5 and service and dry periods of the animal, whereas the genetic variants in the first fragment of the exon 6 showed significant association with age at first breeding and calving. To our knowledge, this study demonstrated for the first time that the polymorphisms in OAS1 gene were associated with reproductive traits and it can be chosen as a candidate gene for improvement of reproductive performance of cattle.     

Effect of feeding tannin degrading bacterial culture (Streptococcus gallolyticus strain TDGB 406) on nutrient utilization,urinary purine derivatives and growth performance of goats fed on Quercus semicarpifolia leaves

To study the effect of supplementation of tannin degrading bacterial culture (Streptococcus gallolyticus strain TDGB 406) on growth performance, nutrient utilization and urinary purine derivatives of goats fed on oak (Quercus semicarpifolia) leaves. For growth study, eighteen billy goats (4 month old, average body weight 9.50 ± 1.50 kg) were distributed into three groups of six animals each. The animals of group 1 served as control while animals of groups 2 (T1) and 3 (T2) were given (@ 5 ml/kg live weight) autoclaved and live culture of isolate TDGB 406 (10⁶ cells/ml) respectively. The animals were fed measured quantity of dry oak leaves as the main roughage source and ad libitum maize hay along with fixed quantity of concentrate mixture. The feeding of live culture of isolate TDGB 406 (probiotic) did not affect dry matter intake and digestibility of nutrients except that of dry matter and crude protein, which was higher in T2 group as compared to control. All the animals were in positive nitrogen balance. There was no significant effect of feeding isolate TDGB 406 on urinary purine derivatives (microbial protein production) in goats. The body weight gain and average live weight gain was significantly higher (p = 0.071) in T2 group as compared to control. Feed conversion efficiency was also better in the goats fed on live culture of TDGB 406 (T2). The feeding of tannin degrading bacterial isolate TDGB 406 as probiotic resulted in improved growth performance and feed conversion ratio in goats fed on oak leaves as one of the main roughage source.     

Molecular Characterization of Oxytocin Receptor Gene in Water Buffalo (Bubalus bubalis)

Buffaloes are known for their productivity as compared to average yielding cows due to higher fat percentage, better feed conversion ability and disease resistance. On the other hand, the reproductive performances of buffaloes are often considered as poor owing to late sexual maturity, weak/silent oestrus, repeat breeder and prolonged intercalving interval. The study of cascade of events during oestrus and oestrous cycle can be useful for the improvement of reproductive efficiency of buffaloes. More precisely, the hormonal changes initiated at the molecular level within the animal determine the reproductive nature of the species. Nucleotide/protein sequence analysis serves as a vital tool in analysing the binding of the hormones for their effect or functions. In this study, we have reported cloning and characterization of the complete coding (cDNA) sequence of oxytocin receptor gene (OXTR) in buffaloes. Buffalo OXTR gene contains an uninterrupted ORF of 1176 nucleotides corresponding to an inferred polypeptide length of 391 amino acids (aa). The molecular weight of the deduced aa sequence was found to be 43 kDa with an isoelectric point of 9.253 and 16.328 charge at pH 7.0. The deduced protein sequence consists of 38 strongly basic (+) (K,R), 22 strongly acidic (?) (D,E), 186 hydrophobic (A, I, L, F, W, V) and 95 Polar (N, C, Q, S, T, Y) aa. Results indicated that aspartate (D) at aa position 85 and D, R and C at aa positions 136, 137 and 138, respectively, are conserved in buffaloes. The buffalo OXTR gene shared a per cent similarity ranging from 84.7 to 98.1 and 88.5 to 97.7 at nucleotide and deduced aa sequence levels, respectively, with that of other species. Phylogram constructed on the basis of either nucleotide or deduced aa sequences of buffalo OXTR gene showed that buffalo, cattle and sheep have diverged from human and swine and formed a separate clad. The buffalo sequence has shown maximum similarity and closeness with cattle followed by sheep both at nucleotide and at aa level.     

IFN‐τ Acts in a Dose‐Dependent Manner on Prostaglandin Production by Buffalo Endometrial Stromal Cells Cultured in vitro

Interferon‐τ (IFN‐τ) has been recognized as the primary embryonic signal responsible for maternal recognition of pregnancy. Uterine endometrium produces both prostaglandin F_2α (PGF_2α) and prostaglandin E₂ (PGE₂). PGF_2α is responsible for the luteolysis; however, PGE₂ favours establishment of pregnancy by its luteoprotective action. In this study, the dose‐response effect of recombinant bovine IFN‐τ (rbIFN‐τ) on prostaglandin (PG) production by buffalo endometrial stromal cells cultured in vitro was studied. Buffalo endometrial stromal cells were isolated by double enzymatic digestion, initially with trypsin III followed by a cocktail of trypsin III, collagenase type II and DNase I and subsequently cultured till confluence. Further, cells were treated with different doses of rbIFN‐τ (0.001, 0.01, 0.1, 1.0 and 10 μg/ml) and keeping a separate set of control. Culture supernatant was collected after 6, 12 and 24 h of treatment. PG levels in the culture supernatant were measured by enzyme immune assay (EIA) and total cellular protein estimated by Bradford method. Results indicated that buffalo endometrial stromal cells following rbIFN‐τ treatment enhanced the secretion of both PGE₂ and PGF_2α, and also its ratio in a strict dose‐dependent manner with a significant increase (p < 0.01) in PGE₂ production at 1 μg/ml dose of rbIFN‐τ and maximal stimulation for both PG was observed at 10 μg/ml. Further, both PG production and its ratio were increased significantly (p < 0.01) in a time‐dependent fashion in all the groups at 6, 12 and 24 h post‐treatment with highest level achieved at 24 h as compared with control. Absolute levels of PGE₂ remained higher than PGF_2α indicating PGE₂ as the major PG produced by endometrial stromal cells. The dose‐dependent response of rbIFN‐τ signifies the importance of optimum concentration of IFN‐τ for the embryonic development especially during the critical period to establish successful pregnancy.     

Ghrelin Accelerates In Vitro Maturation of Bovine Oocytes

Ghrelin, apart from its metabolic role, is nowadays considered as a basic regulator of reproductive functions of mammals, acting at central and gonadal levels. Here, we investigated for possible direct actions of ghrelin on in vitro maturation of bovine oocytes and for its effects on blastocyst yield and quality. In experiment 1, cumulus oocyte complexes (COCs) were matured in the presence of four different concentrations of ghrelin (0, 200, 800 and 2000 pg/ml). In vitro fertilization and embryo culture were carried out in the absence of ghrelin, and blastocyst formation rates were examined on days 7, 8 and 9. In experiment 2, only the 800 pg/ml dose of ghrelin was used. Four groups of COCs were matured for 18 or 24 h (C18, Ghr18, C24 and Ghr24), and subsequently, they were examined for oocyte nuclear maturation and cumulus layer expansion; blastocysts were produced as in experiment 1. The relative mRNA abundance of various genes related to metabolism, oxidation, developmental competence and apoptosis was examined in snap‐frozen cumulus cells, oocytes and day‐7 blastocysts. In experiment 1, ghrelin significantly suppressed blastocyst formation rates. In experiment 2, more ghrelin‐treated oocytes matured for 18 h reached MII compared with controls, while no difference was observed when maturation lasted for 24 h. At 18 and 24 h, the cumulus layer was more expanded in ghrelin‐treated COCs than in the controls. The blastocyst formation rate was higher in Ghr18 (27.7 ± 2.4%) compared with Ghr24 (17.5 ± 2.4%). Differences were detected in various genes’ expression, indicating that in the presence of ghrelin, incubation of COCs for 24 h caused over‐maturation (induced ageing) of oocytes, but formed blastocysts had a higher hatching rate compared with the controls. We infer that ghrelin exerts a specific and direct role on the oocyte, accelerating its maturational process.     

Hepatoprotective activity of Beta vulgaris against CCl4-induced hepatic injury in rats

Ethanolic extract of Beta vulgaris roots given orally at doses of 1000, 2000 and 4000 mg/kg exhibited significant dose-dependent hepatoprotective activity against carbontetrachloride (CCl(4))-induced hepatotoxicity in rats. Hepatotoxicity and its prevention were assessed by serum markers viz. cholesterol, triglyceride, alanine amino transferase and alkaline phosphatase.     

Effect of Terminalia chebula and Allium sativum on in vivo methane emission by sheep

A study was conducted to evaluate some plant parts (already tested for their antimethanogenic activity in in vitro gas production test in the authors’ laboratory) as feed additive to combat methane emission from sheep. Sixteen male sheep with average body weight of 29.96 ± 1.69 kg (22 months of age) were divided into four groups in a randomized block design. The animals were fed on a diet containing forage to concentrate ratio of 1:1. The concentrate fraction composed (in parts) of maize grain, 32; wheat bran, 45; deoiled soybean meal, 20; mineral mixture, 2 and common salt, 1. The four treatments were control (without additive), seed pulp of Terminalia chebula (Harad), bulb of Allium sativum (Garlic) and a mixture (Mix) of the latter two in equal proportions at the rate of 1% of dry matter (DM) intake. There was no effect on DM intake due to the inclusion of these feed additives. The digestibilities of DM and organic matter tended to be higher (p < 0.1) in the groups with T. chebula and A. sativum, whereas, neutral detergent fibre (NDF), acid detergent fibre (ADF) and cellulose digestibilities were higher (p < 0.05) in all the three experimental groups compared with control. The nitrogen balance and plane of nutrition were not affected by inclusion of any of the feed additives. Methane emission (L/kg digested DM intake) as estimated by open circuit respiration chamber and methane energy loss as per cent of digestible energy intake tended to be lower in T. chebula (p = 0.09) and Mix (p = 0.08) groups compared with control. The data indicated that T. chebula showed antimethanogenic activity, whereas both T. chebula and A. sativum improved nutrient digestibility. Therefore, these two plants appear to be suitable candidates for use as feed additive to mitigate methane emission and to improve nutrient utilization by sheep.     

Complement factor H variant increases the risk of age-related macular degeneration

Age-related macular degeneration (AMD) is a leading cause of visual impairment and blindness in the elderly whose etiology remains largely unknown. Previous studies identified chromosome 1q32 as harboring a susceptibility locus for AMD. We used single-nucleotide polymorphisms to interrogate this region and identified a strongly associated haplotype in two independent data sets. DNA resequencing of the complement factor H gene within this haplotype revealed a common coding variant, Y402H, that significantly increases the risk for AMD with odds ratios between 2.45 and 5.57. This common variant likely explains approximately 43% of AMD in older adults.     

The Pattern of Embryonic Fixation and its Relationship to Pregnancy Loss in Thoroughbred Mares

Ultrasonographic pregnancy records of 195 mares from six Thoroughbred stud farms, over a period of 7 years were retrospectively analysed to assess the effect of various factors on embryonic vesicle (EV) fixation pattern and pregnancy outcome. Of the total of 746 pregnancies analysed, significantly (p < 0.01) more EV fixations were evident in the right uterine horn than in the left (53.35% vs 46.65% respectively). There was no significant effect of either, the side of ovulation, or age of the mare, on the side of EV fixation. However, EV fixation, was significantly (p < 0.001) more likely to occur in the right uterine horn in maiden and barren mares (65.75% vs 57.45% respectively). The age and reproductive status of the mare as well as foal heat breeding failed to demonstrate a consistent effect on pregnancy loss relative to the side of EV fixation. In lactating and foal heat bred mares, EVs were significantly (p < 0.0001) more frequently established in the contralateral horn to the one from which the mare delivered her most recent foal. In lactating mares, significantly (p < 0.05) higher embryonic and pregnancy losses were observed in the ipsilateral horn. In conclusion, (a) side of EV fixation was (i) independent of the side of ovulation and mare age (ii) significantly (p < 0.001) affected by reproductive status, (b) neither age of mare nor reproductive status had any effect on pregnancy loss rates relative to the side of EV fixation and (c) in lactating mares the EV had a greater chance of fixation and survival in the horn contralateral to the one from which the mare delivered her most recent foal.     

Induction of Ovulation of Ovulatory Size Non‐Ovulatory Follicles and Initiation of Ovarian Cyclicity in Summer Anoestrous Buffalo Heifers (Bubalus bubalis) Using Melatonin Implants

This study was conducted on summer anoestrous buffalo heifers to monitor the efficacy of melatonin for induction of ovulation and ovarian cyclicity. During pre‐treatment period of 24 days, the ovarian dynamics of five cycling and 10 summer anoestrous heifers was monitored on each alternate day using a transrectal ultrasound scanner. Thereafter, during treatment period, these 10 anoestrous heifers along with additional seven anoestrous heifers were randomly allocated into non‐implanted (n = 5) and implanted (n = 12, one melatonin implant/50 kg, 18 mg melatonin/implant) group. Non‐implanted heifers were monitored on each alternate day till the confirmation of second‐ovulation in implanted heifers. Pre‐treatment period revealed the presence of dominant follicles in anoestrous heifers which attained the diameter comparable with ovulatory follicles of cycling heifers but failed to ovulate and regressed. Between 6 and 36 days (15.3 ± 2.9 days) post‐treatment, all the implanted heifers (p < 0.05) exhibited ovulation of dominant follicles; however none of the non‐implanted heifers ovulated during the corresponding period. The first‐interovulatory period in implanted heifers ranged between 8 and 28 days (18.0 ± 1.8 days). The implanted heifers with short (≤16 days) interovulatory period had short‐lived corpus luteum (CL) that had smaller diameter and secreted less progesterone (p < 0.05). The diameter of CL was large (p < 0.05) and plasma progesterone was high (p < 0.05) following second‐ovulation compared with first‐ovulation in implanted heifers. In conclusion, using melatonin implants, ovulatory size nonovulatory follicles observed in summer anoestrous buffalo heifers can be successfully ovulated to initiate ovarian cyclicity.