Metagenomic analysis of bacterial communities during estrous cycle in Bos indicus.

The present study, using 16 s rRNA sequencing of the V3-V4 hypervariable region, was aimed to check diversity of vaginal microbiota throughout different stages of the estrous cycle in Bos indicus, with attention to changes in progesterone hormone and microorganism diversity. Metagenomic research was conducted on vaginal swabs obtained from nine healthy Indian Gir cows at estrus (day 0), metestrus (day 04), diestrus (day 12), and proestrus (day 16) phases of the estrous cycle. The findings revealed that the diestrus phase has a different bacterial diversity than the other three estrous cycle phases, implying that progesterone affects bacterial diversity. Proteobacteria and Firmicutes were the most abundant phyla at the phylum level, accounting for 94% of bacterial diversity. Actinobacteriota, Patescibacteria, Cyanobacteria, and Bacteroidota were among the less prevalent phyla observed in all samples. After statistical analysis, Bacillaceae, Alcaligenes, Enterobacteriaceae, and Morganellaceae families were more significant. The Enterobacteriaceae family was found to be lower in the diestrus phase than in the other three phases; in contrast, all statistically significant genera were high at the diestrus phase. The luteal stage had higher levels of Micrococcus, Stenotrophomonas, UGC-010, Massilia, and Methylobacillus than the follicular stage, and statistical analysis revealed substantial difference between the luteal and follicular stages. Lactobacillus genus was present in both the estrus and diestrus phases. This study represents an important step toward the understanding of microbial diversity within different stages of the estrous cycle of Indian cows.

Effect of different antioxidant additives in semen diluent on cryopreservability (-196°C) of buffalo semen.

AIM: The aim of this study was to evaluate the effect of different antioxidant additives in standard tris-fructose-egg yolk-glycerol (TFYG) extender on the cryopreservability of buffalo semen. MATERIALS AND METHODS: Semen collection using artificial vagina, twice weekly for 5 weeks from three pedigreed health breeding bulls of Mehsani breed, aged between 6 and 8 years. Immediately after initial evaluation all 30 qualifying ejaculates (10/bull) were split into three aliquots and diluted at 34°C keeping the concentration of 100 million spermatozoa/ml with standard TFYG extender as control and TFYG having two antioxidant additives - Cysteine HCl at 1 mg/ml and ascorbic acid at 0.2 mg/ml to study their comparative performance. Semen filled in French Mini straws using IS-4 system and gradually cooled to 4°C and equilibrated for 4 h in cold handing cabinet. After completion of equilibration, straws were cryopreserved in LN2 by Programmable Bio-freezer. Semen was examined at post-dilution, post-equilibration, and post-thaw stages for sperm quality parameters, and at each stage plasma was separated for enzymatic analysis of aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and alkaline phosphatase (AKP). RESULTS: The mean percentage of sperms in TFYG, TFYG + cysteine HCl and TFYG + ascorbic acid diluents at post-thaw stage in terms of progressive motility (52.83±0.52, 57.83±0.52, 57.83±0.52), livability (78.70±0.21, 82.33±0.23, 81.73±0.22), and abnormality (5.43±0.21, 5.03±0.17, 5.23±0.18) varied significantly (p<0.05) between control TFYG and TFYG having antioxidant additives. The mean U/L activities of AST (78.70±0.47, 72.80±0.48, 73.30±0.54), LDH (172.70±0.41, 155.78±0.42, 156.33±0.41), and AKP (103.61±0.34, 90.20±0.34, 91.03±0.34) in semen diluted with TFYG, TFYG + cysteine HCl and TFYG + ascorbic acid diluents at post-thaw stage, respectively, which showed significantly (p<0.05) higher leakage of enzymes in control TFYG than TFYG incorporated with additives. CONCLUSION: Incorporation of antioxidant additives such as cysteine HCl and ascorbic acid in standard TFYG diluents improves sperm quality parameters, reduces enzyme leakage, and ultimately advances cryopreservability of buffalo semen.