Evaluation of Sperm Functional Attributes in Relation to in Vitro Sperm-zona Pellucida Binding Ability and Cleavage Rate in Assessing Frozen Thawed Buffalo (Bubalus Bubalis) Semen Quality

The objective of this study was to evaluate sperm functional attributes in relation to in vitro sperm-zona binding ability and cleavage rate in assessing frozen thawed buffalo (Bubalus bubalis) semen quality. Frozen-thawed forty-eight ejaculates from eight Surti buffalo bulls (six ejaculates/bull) obtained by artificial vagina were used. Frozen semen from each bull was thawed, pooled, and subjected for sperm functional (six replicates) and in vitro fertilization (four replicates) tests. The progressive forward motility, plasmalemma functional integrity assessed by fluorogenic [6-carboxyfluorescein diacetate (CFDA), and propidium iodide (PI)], hypoosmotic swelling (HOS), and hypoosmotic swelling-Giemsa (HOS-G) test, mitochondrial membrane potential, sperm nuclear morphology, the number of sperm bound to zona and cleavage rate differed significantly (P<0.05) between bulls. When the animals were grouped based on cleavage rate (group I, >40% cleavage rate, n=5, and group II, <40% cleavage rate, n=3), in vitro fertility parameters and all the sperm functional attributes except sperm nuclear morphology differed significantly (P<0.05). The proportions of sperm with functional plasmalemma in the tail and intact acrosome assessed by HOS-G test (25.33, range: 17.48-40.27) were significantly (P<0.001) lower than the functional plasmalemma in the tail assessed by HOS test (39.80, range: 27.85-54.67). The number of sperm bound to zona had significant correlations with the mitochondrial membrane potential (r=0.90, P<0.01) and plasmalemma integrity (fluorogenic, r=0.74 and HOS, r=0.79, P<0.05) and HOS-G, r=0.87, P<0.01). The cleavage rate had significant (P<0.05) correlations with the mitochondrial membrane potential (r=0.70) and plasmalemma integrity measured by HOS-G test (r=0.68). The present study indicates that these attributes could represent important determinants of buffalo sperm quality influencing cleavage rate.