Sperm motility decreasing in a semen sample over time is an indirect approach to assess spermatozoa viability and should be related to the ejaculate fertility. The biospeckle (BSL) is an interacting phenomenon between laser light and biological specimens that allows measurement of sperm kinetic activity by means of an index called the inertial moment (IM). The aim of this work was to evaluate sperm motility diminishing in frozen bovine semen with BSL and analyze semen fertility in relation to the motility decreasing behavior patterns showed by different semen samples. Fertility was assessed by beef heifer conception rates after AI. Semen of 6 mature IA donor bulls (Bos taurus indicus) was previously divided in 2 groups (group I: motility ≥50%; group II: motility <50%), each group comprised 3 animals. Semen was thawed at 37°C for 30 sin a water bath. One aliquot of 10 μL was placed in a warmed slide, covered with a slip, and evaluated by light microscopy and BSL just after thawing. Then, samples were kept at room temperature to induce a decrease in sperm motility. Each sample was illuminated 6 times by laser light at intervals of 2 min (n = 36) and IM values were obtained. The semen samples were used in an AI program of beef heifers (n = 166) under the same nutrition and management conditions. All inseminations were done by the same technician, and donor bulls were used consecutively throughout the breeding season. Pregnancy diagnosis was done by ultrasound images (Falco 100, 6 MHz, Pie Medical, Crawley, UK) 28 days after insemination. The SISVAR software was used in variance analysis and comparison of means by the Tukey test. Group I showed greater (P < 0.05) IM mean value (153.5 ± 27.48) compared with group II (107.94 ± 30.61), which means that this group had higher sperm motility during the time of evaluation. In addition, conception rates (0.63 ± 0.49) for group I were also higher (P < 0.05) compared with group II. Apparently semen fertility is related to the sperm kinetics measured by the IM. Mean IM values (134.0 ± 22.65 and 96 ± 22.34, groups I and II, respectively) obtained in the last illumination for each group did not differ (P > 0.05), but a tendency to differ was observed when a greater number of illuminations was done. Our data suggest that the BSL was capable of identifying spermatozoa surviving between high and low motility groups, and the sperm motility decreasing measured by the IM might be an objective approach to evaluate semen fertilization potential. Financial support: FAPEMIG, grant EDT 94/07, CNPq.
The biospeckle (BSL) is based on the incidence of laser light upon active biological materials, followed by analysis and manipulation of secondary images, and then, application to those images of a second-order statistics technique called the inertial moment (IM) that results in information about the activity of the specimen. This technique was previously used to analyze sperm motility. A bull’s fertility could be evaluated by several semen parameters in vitro and more precisely by conception rates of AI heifers. The objective of this work was to evaluate the fertility of bull’s frozen semen showing different levels of activity in an AI program. Semen of 6 mature IA donor bulls (Bos taurus indicus) was previously divided in 2 groups (group I: motility ≥50%; group II: motility <50%), each group comprising 3 animals. Sperm concentration was 30 to 35 × 106 cells per straw. Semen was thawed at 37°C for 30 s in a water bath. Samples were illuminated by nonpolarized He-Ne laser (632 nm, 10 mW) for 40 s, the time needed to take 512 images under a time rate of 0.08 s. Images were acquired by a CCD camera and analyses were conducted by building space-time speckle matrices from center columns of 512 images of the dynamic speckle and then the IM was calculated. Beef heifers (n = 109) under the same nutrition and management conditions underwent AI with the semen samples. All inseminations were done by the same technician and donor bulls were used consecutively throughout the breeding season. Pregnancy diagnosis was done by ultrasound images (Falco 100, 6 MHz, Pie Medical, Crawley, UK) 28 days after insemination. The SISVAR software was used for variance analysis and comparison of means by Tukey test at a nominal level of 5%. The IM (194.13 ± 31.99) obtained from 10 illuminations per sample of each bull and the conception rate (CR) of group I (CR = 0.63 ± 0.49) differed (P < 0.05) compared with group II (IM = 142.03 ± 30.62; CR = 0.43 ± 0.50). Pearson correlation coefficient (P < 0.05) between IM and CR was r = 0.21, showing coherence with IM and CR, despite its low value. These data showed that second-order statistics are correlated to semen fertility and could be used to evaluate ejaculates of the bull. Financial support: FAPEMIG grant EDT 94/07 and CNPq.
Sperm motility is a physical parameter evaluated in semen samples of the bull and is thought to be related to the fertility in the male. Despite being a characteristic of simple and fast evaluation, motility estimates involve subjective components when analyzed by light microscopy (LM) that might restrain their evaluation in some conditions. Moreover, in some species, poor correlations were observed between spermatozoa motility and semen fertility. The incidence of coherent light in a semen sample generates a phenomenon called biospeckle (BSL) that is capable of measuring the kinetics activity of the spermatozoa. In this work the relationship among sperm kinetics variables evaluated by LM and BSL and conception rates in beef heifers was investigated, with the purpose to predict frozen semen fertility when laser light is used. Sixty semen samples of 6 mature AI donor bulls (Bos taurus indicus) herein named A, B, C, D, E, and F were thawed at 37°C for 30 s in a water bath and evaluated by LM and BSL. In LM evaluation, an index (IND) was proposed to group together, in a single mean estimate, the spermatozoa motility (M: % scale) and velocity (V: 1 to 5 scale) according to the equation IND = [V × 20 + M]/2. In the BSL evaluation, each sample was illuminated (n = 10 per bull) by a laser beam (He-Ne, 632 nm and 10 MW) for 40 s and a mean inertial moment (IM) was obtained for each donor bull. These semen samples were used in an AI program of beef heifers (n = 166) clinically examined for reproductive health, body condition, and weight. Pregnancy diagnosis was done by ultrasound images (Falco 100, 6 MHz, Pie Medical, Crawley, UK) 28 days after insemination. Fertility estimates were done by the generalized linear model using logistic regression (stepwise methodology), generating an equation to predict the conception rate of the semen, the variables of which were IM and IND. Results of the predicted conception rates (pCR) using IM and IND and the observed conception rates (oCR) were A (0.5490; 0.57), B (0.6483; 0.68), C (0.7108; 0.71), D (0.4552; 0.28), E (0.4797; 0.54), F (0.3825; 0.47), respectively. Positive correlations (P < 0.05) were observed between pCR and oCR (r = 0.79) showing a similar behavior between in vitro and in vivo estimates. Results of this work showed that there is a high correlation between spermatozoa kinetics and semen fertility in the bull and that BSL motility analysis could be used as an approach to evaluate the fertility of semen samples. Financial support: FAPEMIG grant EDT 94/07, CNPq.
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