The study revealed noticeable differences in the cumulus gene expression profile at different stages of IVM between ovine oocytes of differential developmental ability. The results indicated that the loss of cumulus gene expression along the maturation period in the poor oocytes was related to their intrinsic poor quality in the ovarian follicle.

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SummaryThe objective of the study was to investigate the effect of l-ergothioneine (l-erg) (5 mM or 10 mM) supplementation in maturation medium on the developmental potential and OCTN1-dependant l-erg-mediated (10 mM) change in mRNA abundance of apoptotic (Bcl2, Bax, Casp3 and PCNA) and antioxidant (GPx, SOD1, SOD2 and CAT) genes in sheep oocytes and developmental stages of embryos produced in vitro. Oocytes matured with l-erg (10 mM) reduced their embryo toxicity by decreasing intracellular ROS and increasing intracellular GSH in matured oocytes that in turn improved developmental potential, resulting in significantly (P < 0.05) higher percentages of cleavage (53.72% vs 38.86, 46.56%), morulae (34.36% vs 20.62, 25.84%) and blastocysts (14.83% vs 6.98, 9.26%) compared with other lower concentrations (0 mM and 5 mM) of l-erg without change in maturation rate. l-Erg (10 mM) treatment did not influence the mRNA abundance of the majority of apoptotic and antioxidant genes studied in the matured oocytes and developmental stages of embryo. A gene expression study found that the SLC22A4 gene that encodes OCTN1, an integral membrane protein and specific transporter of l-erg was not expressed in oocytes and developmental stages of embryos. Therefore it was concluded from the study that although there was improvement in the developmental potential of sheep embryos by l-erg supplementation in maturation medium, there was no change in the expression of the majority of the genes studied due to the absence of the SLC22A4 gene in oocytes and embryos that encode OCTN1, which is responsible for transportation of l-erg across the membrane to alter gene expression.

Assessment of intracellular reactive oxygen species (ROS) is important for evaluating the developmental ability of cumulus-oocyte complexes (COC) and embryos. Although, fluorescence-based 2 ,7-dichlorodihydrofluorescein diacetate (DCFH-DA) staining method is used widely for detecting intracellular ROS in COC and embryos, it is associated with several limitations. This study aimed to develop an alternative method for detecting and quantifying intracellular ROS in oocytes, cumulus cells and embryos based on nitroblue tetrazolium (NBT) staining and bright-field microscopy. Nitroblue tetrazolium reacts with ROS and forms formazan precipitate that can be detected as dark purple/blue spots under bright-field microscope. Ovine COC were matured in vitro without (control) or with the supplementation of Interleukin-7 (IL-7; for stimulating intracellular ROS), Tempol (superoxide scavenger) or combination of IL-7 and Tempol. The matured COC were stained with NBT and the formation of intracellular formazan precipitates was assessed. Additionally, the matured COC were stained with DCFH-DA to compare the level of intracellular ROS. Further, ovine embryos (8-cell, morula, and degenerating) were generated in vitro and stained with NBT for assessing intracellular ROS. The level of intracellular ROS was expressed as the proportion (%) of the NBT stained area of oocytes, compact cumulus cell masses or embryos. The proportions of NBT stained area in the matured oocytes and cumulus cells was found significantly lesser in the control as compared to the IL-7 (1 and 5 ng/ml) treated groups. A similar trend in the intracellular ROS level was also observed in the matured COC, when assessed based on the DCFH-DA staining. Following the treatment with Tempol (100 mM), negligible NBT stained area in oocytes and cumulus cells was observed. The NBT staining patterns of the oocytes and cumulus cells following the combined

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The present study was aimed to see the faecal bacterial fingerprints of pigs applying terminal restriction fragment length polymorphism (T-RFLP) analysis. Sixteen crossbred (Large White Yorkshire X desi) grower pigs (body weight of 15.8±1.1 kg), were divided into four groups (control, antibiotic, herbal residue and prebiotic) with four in each treatment. T-RFLP analysis revealed comparable bacterial abundance in control - antibiotic and herbal residue - prebiotic groups. Abundance of Peptostreptococcus anaerobius (an antibiotic resistant pathogen) was higher in antibiotic supplemented pigs. Number of significantly responded operational taxonomic units (OTU) was higher in herbal residue and prebiotic supplemented pigs than control or antibiotic pigs. Bacterial abundance was significantly higher in pigs supplemented with prebiotic, followed by herbal residue, antibiotic, and control. The abundance of beneficial bacteria such as Sarcina maxima, Bacteroidetes sp., Cetobacterium somerae, Selenomonas sputigera, Fecalibacterium prausnitzi were significantly higher in prebiotic groups. The present findings established that herbal residue and prebiotic are alternative to feed antibiotic in pigs.

Large number of proteases have been identified in different parts of the male reproductive tract which play a crucial role during testicular development, epididymal sperm maturation, and sperm-oocyte fusion. The ADAMTS10 is a member of metalloproteinase family, the expression of which is important in sperm-oocyte interaction at the time of fertilization in mouse. In this species, the expression of ADAMTS10 has been demonstrated in testis, and on surface of spermatozoa from caput, corpus and cauda epididymis. Whether ADAMTS10 plays any role in male fertility of any of the domestic ruminant species including buffaloes is not known so far. Thus, a study was conducted to detect the expression of ADAMTS10 in testis, different parts of the epididymis and ejaculated spermatozoa of water buffalo (Bubalus bubalis) using real-time PCR, Western blot, and indirect immunofluorescence techniques. The highest expression of ADAMTS10 transcript was observed in corpus epididymis, followed by cauda, caput and testis. The Western blot detection of ADAMTS10 using the heterologous antibody has demonstrated 31-, 44-, 50-, 65 kDa immune-reactive protein bands from testis and 82 kDa protein from both testes and corpus epididymis and a major 36 kDa protein from ejaculated spermatozoa. The indirect immunofluorescence study has demonstrated high fluorescence signal in post-acrosome and the middle piece region of the ejaculated spermatozoa. Thus the differential expression pattern of ADAMTS10 in different parts of male reproductive tract of buffalo indicates their role in sperm maturation process. However, specific role of ADAMTS10 in buffalo male reproduction can only be ascertained after further studies.