Characterization of antisperm antibody binding patterns in relation to sperm phenotypic attributes and field fertility in dairy bulls.

To test our hypothesis that antisperm antibodies (ASA) might alter sperm phenotypic attributes thus leading to sub-fertility/infertility in bulls, ASA were generated in crossbred male calves by immunizing with sperm two times. Cryopreserved spermatozoa from crossbred bulls (n = 24) with different field fertility ratings were incubated with ASA and different patterns of ASA immunolocalization were studied. In addition, sperm membrane integrity, acrosomal integrity and cryo-capacitation status were also assessed. Immunolocalization of sperm antigens using antisperm antibody revealed three major patterns (Acrosomal-AR, apical-AP and, acrosome and tail-AT). The proportion of ASA reactive spermatozoa was significantly (P < 0.05) higher in low- and medium-fertile bulls compared to high-fertile bulls. Among the three patterns, the proportion of spermatozoa with AR pattern was significantly (P < 0.05) higher in low- and medium-fertile bulls compared to high-fertile bulls. The proportion of membrane and acrosome intact spermatozoa was significantly (P < 0.05) higher in high-fertile bulls compared to medium- and low-fertile bulls. There were no significant differences in the proportion of cryo-capacitated spermatozoa among high-, medium- and low-fertile bulls. The relationship between ASA reactive spermatozoa and conception rates (CR) of bulls was highly (P < 0.01) significant and negative. Similarly, AR and AT pattern were also significantly (P < 0.01) and negatively related to CR of bulls. The reactivity of spermatozoa with ASA was also significantly (P < 0.01) and negatively related to the membrane and acrosome integrity of spermatozoa. It was concluded that the proportion of spermatozoa responding to ASA was higher in low-compared to high-fertile bulls and ASA localization in sperm acrosomal area was negatively related to sperm membrane and acrosomal integrity and bull fertility.

Improvement in sperm functional competence through modified low-dose packaging in French mini straws of bull semen.

To achieve the targeted artificial insemination coverage with the current rate of semen production, without affecting the conception rate, it needs to reduce the number of spermatozoa per insemination dose in India as per international practice. Therefore, this study was planned to perform different levels of semen dilution, compare in vitro post-thaw semen quality and develop a modified low-dose semen packaging method in French mini straw to minimise semen dilution effect. Sixteen ejaculates were collected from Karan Fries bulls (n = 4). The mean percentage post-thaw motility, viability, membrane integrity, acrosome integrity, lipid peroxidation and capacitation status were estimated as post-thaw sperm function assays in semen sample diluted to 20, 15, 10 and 5 million spermatozoa per 0.25 ml and filled in the French mini straw by conventional packaging. No significant (p > .05) difference in post-thaw sperm quality was observed between 15 and 20 million doses; however, below 15 million sperm quality get reduced. There was no significant difference in post-thaw semen quality traits between 20 million conventional packaging and 5 million spermatozoa/dose in modified packaging. In conclusions, the modified packaging is a very effective method for low-dose cryopreservation with acceptable post-thaw semen quality.

Biochemical analysis of uterine fluid for identification of indicators for subclinical endometritis in the water buffalo (Bubalus bubalis).

Alterations in biochemical constituents of uterine fluid have been suggested for diagnosis of subclinical uterine infection in the bovine. This study was undertaken to investigate whether uterine fluid biomolecules could act as tool for diagnosis of subclinical endometritis in the buffalo. Uterine fluid samples from normal (n = 22) and subclinical endometritis (n = 18; diagnosed based on uterine cytology)-affected buffaloes were subjected to biochemical analysis. Among the different biochemical constituents estimated, urea, urea N, cholesterol, total bilirubin and alkaline phosphatase (ALP) concentrations were significantly (p < 0.05) higher in uterine fluid obtained from subclinical endometritis-affected buffaloes. The extent of difference between normal and subclinical endometritis-affected buffaloes was highest in ALP (69%) followed by cholesterol (55%), bilirubin (48%), urea (30%) and urea N (30%) concentrations. Receiver operating characteristic (ROC) analysis indicated that the likelihood ratio (LR) was 3.63 for urea, indicating that buffaloes having less than the threshold concentration (47.5 mg/dl) of urea in their uterine fluid were at 3.6 times more risk to be affected with SE. The LRs for urea N, cholesterol, ALP and bilirubin were 2.33, 2.54, 2.12 and 1.65, respectively. It was concluded that ALP, urea, urea N and cholesterol concentrations in uterine fluid may serve as an aid for diagnosing subclinical endometritis in the buffalo.

Nonsurgical endodontic treatment of teeth associated with large periapical lesion using triple antibiotic paste and mineral trioxide aggregate apical plug: A case series.

Periapical diseases are induced as a result of the direct or indirect involvement of oral bacteria. The etiologic factor being the degenerating pulp tissue. A periapical lesion is formed within an area of apical periodontitis which cannot form by itself and is inflammatory in origin. If the microbial etiology of periapical lesions and in the root canal is removed by nonsurgical root canal therapy the lesions regress. Mere surgical removal of the periapical lesions without proper root canal disinfection and obturation will not result in the healing of periapical tissues. Nonsurgical treatment with triple antibiotic paste offers a high success rate in the healing of large periapical lesions. The present clinical cases show the nonsurgical endodontic management of large periapical pathosis using triple antibiotic paste and mineral trioxide aggregate.

Development of an in vitro oviduct epithelial explants model for studying sperm-oviduct binding in the buffalo.

In this study, we developed an in vitro model for studying sperm-oviduct binding in the buffalo. Oviduct explants were prepared by overnight culture of epithelial cells in TCM-199 medium under 5% CO2 at 38.5 °C. Cryopreserved spermatozoa from buffalo bulls (n = 4) were incubated with the oviduct explants, and the sperm-oviduct explants complex was stained with JC-1. The effect of sperm concentration (2, 3 and 4 million), size of the oviduct explants (<0.2, 0.2-0.3, 0.3-0.4 and >0.4 mm2 ) and time of incubation (1 hr and 4 hr) on binding index (BI-number of sperm bound to unit area of explants) was studied. No significant difference was observed in the BI among <0.2, 0.2-0.3 and 0.3-0.4 mm2 size of explants; however, the BI decreased significantly (p < .05) when the size of explants exceeded 0.4 mm2 . The BI decreased significantly (p < .05) when the sperm concentration was increased to 4 million, while the duration of incubation did not have any significant effect on the BI. The interaction of bulls with explants size, sperm concentration and incubation time was not significant. The developed assay has the potential to be used as an in vitro model for studying sperm-oviduct binding in the buffalo.