Sperm head morphometry in ejaculates of adult marmosets (Callithrix jacchus): a model for studying sperm subpopulations and among-donor variations

In humans and other mammals, sperm morphology has been considered one of the most important predictive parameters of fertility. The objective was to determine the presence and distribution of sperm head morphometric subpopulations in a nonhuman primate model (Callithrix jacchus), using an objective computer analysis system and principal component analysis (PCA) methods to establish the relationship between the subpopulation distribution observed and among-donor variation. The PCA method revealed a stable number of principal components in all donors studied, that represented more than 85% of the cumulative variance in all cases. After cluster analysis, a variable number (from three to seven) sperm morphometric subpopulations were identified with defined sperm dimensions and shapes. There were differences in the distribution of the sperm morphometric subpopulations (P < 0.001) in all ejaculates among the four donors analyzed. In conclusion, in this study, computerized sperm analysis methods combined with PCA cluster analyses were useful to identify, classify, and characterize various head sperm morphometric subpopulations in nonhuman primates, yielding considerable biological information. In addition, because all individuals were kept in the same conditions, differences in the distribution of these subpopulations were not attributed to external or management factors. Finally, the substantial information derived from subpopulation analyses provided new and relevant biological knowledge which may have a practical use for future studies in human and nonhuman primate ejaculates, including identifying individuals more suitable for assisted reproductive technologies. (c) 2012 Elsevier Inc. All rights reserved.

Identification of sperm head subpopulations with defined pleiomorphic characteristics in ejaculates of captive Goeldi's monkeys (Callimico goeldii)

The aim of this study was to evaluate the incidence of pleiomorphisms and its influence on the distribution of sperm morphometric subpopulations in ejaculates from the vulnerable Goeldi's monkey (Callimico goeldii) by using a combination of computerized analysis system and Principal Component Analysis (PCA) methods Each sperm head was measured for four primary spermatozoal head dimensional parameters (area [A (μm2)], perimeter [P (μm)], length [L (μm)] and width [W (μm)]) and three head shape derived parameters (ellipticity [(L/W)], elongation [(L-W)/(L+W)] and rugosity [(4πA/P2)]) Six separate subpopulations (SPs) were identified: SP1, constituted by very large, narrow and very elliptical spermatozoa (A=16.85±1.56μm2, W=2.75±0.42μm and ellipticity=2.16±0.24); SP2, characterized by average sized, short, wide and round spermatozoa (A=15.00±1.92μm2, L=5.06±0.49μm, W=3.51±0.31μm and ellipticity=1.44±0.15); SP3, represented by small, wide and slightly round spermatozoa (A=14.95±1.75μm2, W=3.47±0.29μm and ellipticity=1.48±0.14); SP4 included very small, short and very round spermatozoa (A=14.15±2.38μm2, L=4.90±0.57μm and elongation=0.18±0.05); SP5 consisted of average sized and slightly elliptical spermatozoa (A=15.14±1.72μm2 and ellipticity=1.49±0.14); and SP6 included large and round spermatozoa (A=16.30±1.62μm2 and elongation=0.19±0.04) There were differences in the sperm subpopulation distribution (P<0.001) among the five donors analyzed In conclusion, the results of the current study confirmed that the use of computer sperm analysis methods combined with PCA cluster analyses are useful methods to identify, classify, and characterize different sperm head morphometric subpopulations in neotropical primates Broadening our knowledge of C goeldii sperm morphometric abnormalities as well as developing reliable techniques for sperm evaluation may be essential for ex situ conservation of this threatened species © 2012 Elsevier B.V.

Differential distribution of sperm subpopulations and incidence of pleiomorphisms in ejaculates of captive howling monkeys (Alouatta caraya)

The aim of this study was to develop an objective method to determine the incidence of pleiomorphisms and its influence on the distribution of sperm morphometric subpopulations in ejaculates of howling monkeys (Alouatta caraya) by using a combination of computerized analysis system (ASMA) and principal component analysis (PCA) methods. Ejaculates were collected by electroejaculation methods on a regular basis from five individuals maintained under identical captive environmental, nutritional, and management conditions. Each sperm head was measured for dimensional parameters (Area [A, (square micrometers)], Perimeter [P, (micrometers)], Length [L, (micrometers)], and Width [W, (micrometers)]) and shape-derived parameters (Ellipticity [(L/W)], Elongation [(L - W)/(L + W)], and Rugosity [(4 pi A/P-2)]). PCA revealed two principal components explaining more than the 96 % of the variance. Clustering methods and discriminant analyzes were performed and seven separate subpopulations were identified. There were differences (P < 0.001) in the distribution of the seven subpopulations as well as in the incidence of abnormal pleiomorphisms (58.6 %, 49.8 %, 35.1 %, 66.4 %, and 55.1 %, P < 0.05) among the five donors tested. Our results indicated that differences among individuals related to the incidence of pleiomorphisms, and sperm subpopulational structure was not related to the captivity conditions or the sperm collection method, since all individuals were studied under identical conditions. In conclusion, the combination of ASMA and PCA is a useful clinical diagnostic resource for detecting deficiencies in sperm morphology and sperm subpopulations in A. caraya ejaculates that could be used in ex situ conservation programs of threatened species in Alouatta genus or even other endangered neotropical primate species.

Assessment of in vitro sperm characteristics and their importance in the prediction of conception rate in a bovine timed-AI program

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Effect of sequence of insemination after simultaneous thawing of multiple semen straws on conception rate to timed AI in suckled multiparous Nelore cows

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Evaluation of semen characteristics of the species Mazama americana in captivity

In recent years the concept of genomic resource banks has grown as a way of maintaining the genetic variability of populations, while the quality of cryopreservation of the gametes determines the effectiveness of such banks. However, the absence of basic knowledge regarding the physiology of species and their semen characteristics hampers the establishment of reproductive biotechnologies. Thus, this paper aimed to determine certain physicochemical (volume, colour, appearance, pH and osmolarity) and microscopic characteristics (mass movement, motility, vigour, concentration, and sperm morphology and morphometry) of semen of the species Mazama americana. To achieve this, five males of the species were used, and three semen samples per buck (electroejaculation) were collected at intervals of 2 weeks. The volume, pH and osmolarity of the ejaculate were 0.39 ± 0.14 mL, 6.90 ± 0.74 and 297.74 ± 19.10 mOsm/kg, respectively, while the values obtained for mass movement, motility, vigour and concentration were 3.33 ± 0.82; 69.6 ± 8.92%; 3.53 ± 0.50, and 244.07 ± 98.65 × 107/mL, respectively. Regarding the colour of the ejaculate, five samples were classified as ivory, two as yellowish, two as whitish and six as white. Regarding appearance, seven samples were considered creamy and eight, milky. Morphology was analysed in a humid chamber under phase contrast microscopy and 73.50 ± 5.57% of cells presented normal morphology, 8.37 ± 3.15% presented major defects and 18.13 ± 6.46% presented minor defects. To determine sperm morphometry, an optical microscope (Leica DM 5000B) and the Leica Qwin image analyser program were used, resulting in 8.09 ± 0.40, 4.65 ± 0.30, 2.81 ± 0.44 and 30.25 ± 3.02 m for length, largest width, smallest width and area, respectively. Copyright © CSIRO 2013.

Análise comparativa de características seminais de Mazama cinzas brasileiros: morfometria espermática e proteômica do plasma seminal

Pós-graduação em Medicina Veterinária - FCAV

Effect of sequence of insemination after simultaneous thawing of multiple semen straws on conception rate to timed AI in suckled multiparous Nelore cows

The objective was to determine the effect of sequence of insemination after simultaneous thawing of multiple 0.5 mL semen straws on conception rate in suckled multiparous Nelore cows. The effect of this thawing procedure on in vitro sperm characteristics was also evaluated. All cows (N = 944) received the same timed AI protocol. Ten straws (0.5 mL) of frozen semen from the same batch were simultaneously thawed at 36 degrees C, for a minimum of 30 sec. One straw per cow was used for timed AI. Frozen semen from three Angus bulls was used. Timed AI records included sequence of insemination (first to tenth) and time of semen removal from thawing bath. For laboratory analyses, the same semen batches used in the field experiment were evaluated. Ten frozen straws from the same batch were thawed simultaneously in a thawing unit identical to that used in the field experiment. The following sperm characteristics were analyzed: sperm motility parameters, sperm thermal resistance, plasma and acrosomal membrane integrity, lipid peroxidation, chromatin structure, and sperm morphometry. Based on logistic regression, there were no significant effects of breeding group, body condition score, AI technician, and sire on conception rate, but there was an interaction between sire and straw group (P = 0.002). Semen from only one bull had decreased (P < 0.05) field fertility for the group of straws associated with the longest interval from thawing to AI. However, the results of the laboratory experiment were unable to explain the findings of the field experiment. Sperm width:length ratio of morphometric analysis was the single sperm characteristic with a significant interaction between sire and straw group (P = 0.02). It was concluded that sequence of insemination after simultaneous thawing of 10 semen straws can differently affect conception rates at timed AI, depending on the sire used. Nevertheless, the effects of this thawing environment on in vitro sperm characteristics, remain to be further investigated. (C) 2012 Elsevier Inc. All rights reserved.

Chromatin and morphometry alterations of bovine sperm head after Percoll (TM) density gradient centrifugation

The aim of this work was to assess the morphometry and chromatin integrity of bovine sperm head after a three layers discontinuous Percoll (TM) density gradient centrifugation. Frozen semen doses were obtained from six bulls of different breeds, including three taurine and three Zebu animals. Three ejaculates per bull were evaluated. The semen doses were thawed and two smears were made from each sample before (control) and after the Percoll (TM) centrifugation (Percoll (TM) group). The smears were stained with toluidine blue and grayscale digital images were captured and processed in Scilab environment software. It was observed that chromatin heterogeneity was reduced (P<0.05) and chromatin decondensation was increased (P<0.05) after the Percollm treatment utilized. In addition, it was observed that sperm head length was higher (P<0.05) and the side symmetry was lower (P<0.05) in centrifuged sperm cells. When analyzed separately by subspecies, it was observed that the decrease (P<0.05) in chromatin heterogeneity and the increase (P<0.05) in chromatin decondensation occurred in Zebu sperm heads. In addition, the length and the width:length ratio of sperm heads was affected by Percoll (TM) centrifugation in Zebu semen. In conclusion, the three layers discontinuous Percoll (TM) centrifugation increased the chromatin decondensation and the morphometric alterations of frozen-thawed bovine semen. However, the real implication of these findings in fertility rates of centrifuged sperm must be investigated. (C) 2011 Elsevier B.V. All rights reserved.

Daily sperm production and testicular morphometry in goats according to external scrotal conformation

The present study aimed to compare testicular histology and the testicular cell population as well as spermatogenic efficiency in goats with different scrotal conformations. Eighteen goats were divided into 3 groups: Group I - goats without bipartition of the scrotum, Group II - animals with bipartition of the scrotum up to 50% of the testicular length, Group III - goats with scrotal bipartition more than 50% of the testicular length. In goats in Groups I, II and III, the values for the volume density of seminiferous epithelium were 68.9 +/- 0.6%, 71.5 +/- 2.8% and 73.4 +/- 4.7% (P < 0.05), the height of the seminiferous epithelium were 60.2 +/- 4.9 mu m, 61.0 +/- 5.0 mu m and 73.1 +/- 6.6 mu m (P < 0.05), total length of seminiferous tubules found for Groups I, II and III were 2091.9 +/- 27 m, 2172.5 +/- 24.1 m, and 2340.1 14 m (P < 0.05), number of Sertoli and Leydig cells were 1.8 +/- 0.4 x 10(9) and 1.4 +/- 0.1 x 10(9), 2.2 +/- 0.4 and 2.2 +/- 0.7 x 10(9), and 2.5 +/- 0.1 10(9) and 2.3 +/- 0.510(9)(P < 0.05)and daily sperm production observed were 2.1 0.3 x 109, 2.8 0.4 x 109, and 3.1 0.7 x 109 (P < 0.05). In conclusion, goats with greater scrotal bipartition have a greater capacity to produce reproductive cells that is reflected in a greater reproductive potential. (C) 2011 Elsevier B.V. All rights reserved.

Assessment of field fertility and several in vitro sperm characteristics following the use of different Angus sires in a timed-AI program with suckled Nelore cows

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Glutamate-induced obesity leads to decreased sperm reserves and acceleration of transit time in the epididymis of adult male rats

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Glutamate-induced obesity leads to decreased sperm reserves and acceleration of transit time in the epididymis of adult male rats

Background: Given the established fact that obesity interferes with male reproductive functions, the present study aimed to evaluate sperm production in the testis and storage in the epididymis in a glutamate-induced model of obesity. Methods: Male rats were treated neonatally with monosodium glutamate (MSG) at doses of 4 mg/kg subcutaneously, or with saline solution (control group), on postnatal days 2, 4, 6, 8 and 10. On day 120, obesity was confirmed by the Lee index in all MSG-treated rats. After this, all animals from the two experimental groups were anesthetized and killed to evaluate body and reproductive organ weights, sperm parameters, plasma hormone levels (FSH, LH and testosterone), testicular and epididymal histo-morphometry and histopathology. Results: Significant reductions in absolute and relative weights of testis, epididymis, prostate and seminal vesicle were noted in MSG-treated animals. In these same animals plasma testosterone and follicle-stimulating hormone (FSH) concentrations were decreased, as well as sperm counts in the testis and epididymis and seminiferous epithelium height and tubular diameter. The sperm transit time was accelerated in obese rats. However, the number of Sertoli cells per seminiferous tubule and stereological findings on the epididymis were not markedly changed by obesity. Conclusions: Neonatal MSG-administered model of obesity lowers sperm production and leads to a reduction in sperm storage in the epididymis of adult male rats. The acceleration of sperm transit time can have implications for the sperm quality of these rats.

Assessment of field fertility and several in vitro sperm characteristics following the use of different Angus sires in a timed-AI program with suckled Nelore cows

In order to provide information that may help researchers to understand the main cause(s) of differences in bull fertility frequently observed in field trials, this study aimed to investigate conception rates as well as several in vitro sperm characteristics of different sires of unknown fertility utilized in a Timed-AI (TAI) program. Suckled Nelore cows submitted to the same TAI protocol were allocated into eight breeding groups of approximately 120 animals each. Frozen semen doses from three Angus bulls and three different batches from each bull were utilized. Approximately 100 doses from each batch were used in TAI. Sires, batches and AI technicians were equally distributed across breeding groups. Cows were examined for pregnancy diagnosis 40 d after TAI. For in vitro sperm analyses, the same thawing procedure was repeated in the laboratory to mimic field conditions. The following in vitro sperm characteristics were assessed: computerized motility, thermal resistance, plasma and acrosomal membrane integrity, lipid peroxidation, morphology, morphometry and chromatin structure. No effect of breeding group, body condition score, AI technician and sire was observed. However, some significant differences among bulls were detected in laboratory analyses. Semen from sire presenting numerically lower (P > 0.05) pregnancy/AI also presented lower (P < 0.05) values in all sperm characteristics analyzed in thermal resistance test at 4 h (Total Motility, Progressive Motility, Average Path Velocity, Straight-Line Velocity, Curvilinear Velocity, Amplitude of Lateral Head Displacement, Beat Cross Frequency, Straightness, Linearity, and Percentage of Rapidly Moving Cells), higher (P < 0.05) Major and Total Defects in sperm morphological test, lower (P < 0.05) Length, Ellipticity and Fourier parameter (Fourier 0) in sperm morphometric analysis as well as higher (P < 0.05) chromatin heterogeneity. It was concluded that, although no bull effect was observed in the field experiment, the sire that presented numerically lower pregnancy/AI also presented lower semen quality according to the laboratory analyses performed. (C) 2012 Elsevier B.V. All rights reserved.