Evaluation of the cell population of the seminiferous epithelium and spermatic indexes of the bat Sturnira lilium (Chiroptera: Phyllostomidae).

Due to the scarcity of information about patterns of spermatogenesis in bats, this study aimed to provide information on the testicular activity of the bat Sturnira lilium along the annual seasons. Thus, a series of morphometrical and stereological analyses were made using the testes of adult S. lilium in order to achieve a better understanding of the sperm production dynamics. Light and transmission electron microscopy analyses were performed in testicular fragments of animals captured during dry and rainy seasons. The testes followed the pattern of organization described for other mammals, and there were no morphological differences between organs collected either in dry or in rainy seasons. Each tubular cross-section in stage 1 was made of 0.5 type-A spermatogonia, 4.4 primary spermatocytes in preleptotene/leptotene, 3.7 in zygotene, 11.9 in pachytene, 35.6 round spermatids and 8.5 Sertoli cells. The mitotic and meiotic indexes were 15.4 and 2.9 cells, respectively, while the spermatogenesis yield was 68.7 cells. The testicular sperm reserves was 37.61×10(6) cells, and daily sperm production per gram of testis averaged 209.68×10(6) cells, both highest averages occurring in the rainy season. S. lilium male bats have a continuous reproductive pattern, high spermatogenesis yield and low support capacity by the Sertoli cells.

Histomorphometric characterization of the intertubular compartment in the testes of the bat Sturnira lilium.

Due to the scarcity of information about the reproduction of bats, it is necessary to perform studies on different species to identify the occurrence of annual variations in their reproductive process. Therefore, the aims of this study were to describe and quantify the intertubular components within the testes of the bat Sturnira lilium and to verify whether seasonality takes place in spermatogenesis of this species. The animals were collected in different seasons and time of collection was grouped into dry and rainy seasons. Testicular fragments were routinely processed for light and transmission electron microscopies and blood samples from each animal were collected for quantification of plasma testosterone. Overall, the Leydig cells (LC) were the main intertubular component (83.2%), with abundant lipid droplets in their cytoplasm. Ultrastructural analysis indicated collagen fibers in the connective tissue and lymphatic spaces, with thin walls, surrounding the seminiferous tubules. The proportion and volume of each intertubular component did not vary significantly between seasons. On average, testosterone concentrations did not vary between rainy and dry seasons (21ng/mL). The total number of LC in the testis (50.0×10(5)) and per gram of testis (11×10(7)) did not vary nor did the Leydigosomatic Index (0.03%). Therefore, it is concluded that S. lilium had significant investment in intertubular tissues, especially in LC. Most of the variables that were assessed did not vary with season of the year, which leads to the assumption that S. lilium has a continuous reproductive cycle in southeastern Brazil.

Histomorphometric evaluation of the Molossus molossus (Chiroptera, Molossidae) testis: the tubular compartment and indices of sperm production.

Insectivorous bats play a very important role in the regulation of tropical ecosystems, but information about their reproductive cycle is lacking. Thus, male Molossus molossus were captured over the four seasons, and morphometric analyses of their testes were conducted to infer on the gonadal dynamics and the reproductive capacity of the species. Testes were immersed in Karnovsky fixative, and fragments were embedded in methacrylate and paraplast for morphometric and TUNEL assay respectively. The least gonadosomatic index (0.3%), tubulesomatic index (0.2%) and tubular diameter (133.2µm) occurred in summer. An adult M. molossus showed a total average of 48.9m of seminiferous tubules per gram of testis. Primary spermatocytes were observed in the zygotene at Stage 1 of the seminiferous epithelium cycle. The greatest meiotic index was obtained in winter (3.8 cells), and the general yield of spermatogenesis was higher in winter (64.5 cells) than in summer (19.1 cells). There was no difference in the apoptotic cells count among seasons. The Sertoli cell index was less in summer (5.9) than in fall (11.6), while the number of Sertoli cells per gram of testis did not vary significantly among the seasons (28.0×10(7)). The spermatic reserve per gram of testis was greater in the fall (63.9×10(7)) and winter (69.8×10(7)) than summer (37.1×10(7)). We conclude that M. molossus males show a continuous reproductive cycle, featuring greater spermatogenic activity during the fall and winter, a tubular length above the average of other mammals and a less support capacity of the Sertoli cells.

Comparação entre duas concentrações de glicerol para a criopreservação de sêmen de suçuarana (Puma concolor) / Comparison between two glycerol concentrations to cryopreservation of semen of mountain lions (Puma concolor)

O desenvolvimento de biotécnicas de reprodução é uma importante ferramenta para a conservação de animais silvestres ameaçados de extinção. Procedimentos de reprodução assistida em suçuarana, no entanto, são escassos na literatura, em especial aqueles relacionados à criopreservação de sêmen. Neste sentido, o presente trabalho objetivou avaliar a congelabilidade do sêmen de suçuaranas adultas mantidas em cativeiro, por meio da comparação entre duas concentrações de glicerol no meio de congelamento. Foram usados cinco machos adultos de suçuarana, mantidos no Centro de Reabilitação de Animais Silvestres do Mato Grosso do Sul (CRAS/MS). As amostras foram coletadas por eletroejaculação e avaliadas quanto ao seu aspecto físico, volume, vigor, motilidade, concentração e índice espermático. De cada ejaculado duas alíquotas foram diluídas em meio Tris-citrato-gema de ovo, em concentrações finais de 5 e 7,5% de glicerol, resfriadas a uma taxa de -0,55ºC/min e congeladas a uma taxa de -5,8ºC/min. Depois de descongeladas, as amostras foram reavaliadas e submetidas aos testes de termorresistência e hiposmótico. O protocolo de criopreservação e descongelamento de sêmen proposto se mostrou eficiente em ambas as concentrações de glicerol testadas, não havendo diferença (p>0,05) entre estas.

The development of biotechnologies of reproduction is an important tool for the conservation of wild animals threatened with extinction. Assisted reproduction procedures in mountain lions, however, are scarce, especially those related to sperm cryopreservation. In this context, this study aimed to evaluate the freezing capacity of semen from adult mountain lions in captivity through the comparison of two concentrations of glycerol in the freezing media. Five adult male mountain lions were used, held at the Rehabilitation Center for Wild Animals of Mato Grosso do Sul (CRAS/MS). Samples were collected by electroejaculation and evaluated for physical appearance, volume, sperm progressive status, sperm motility, sperm concentration and sperm motility index. Each ejaculate was spliced into two aliquots and diluted in Tris-citrate-half egg yolk, at final concentrations of 5 and 7.5% glycerol, cooled at a rate of -0.55ºC/min and frozen at a rate of -5.8ºC/min. Once thawed, the samples were re-evaluated and tested for thermoresistance and hypoosmotic swelling. The suggested protocol for cryopreservation and thawing of semen is efficient in both glycerol concentrations tested, with no difference (p>0.05) between them.

Abdominal and pelvic ultrasound study of the maned wolf (Chrysocyon brachyurus) / Estudo ultrassonográfico abdominal e pélvico de lobo-guará (Chrysocyon brachyurus)

The aim of the present study was the ultrasound characterization of the abdominal and pelvic regions of five maned wolves kept in captivity at the Triage Center of Wild Animals of the Federal University of Viçosa (Centro de Triagem de Animais Silvestres, Universidade Federal de Viçosa). This characterization included descriptions of ultrasonographic aspects and measurements of various structures using B-mode ultrasound. Biometric data were collected to assess the existence of significant linear correlations between these measurements and the measurements obtained by ultrasound. Additionally, hematological and serum biochemistry evaluations of the animals were performed. The ultrasound findings were similar to those available in the literature on domestic dogs, which were used for comparison as a result of the lack of published data regarding maned wolves. The latter species showed characteristics closely resembling those of the former, differing in the spleen and left renal cortex echogenicities, in the appearance of the prostatic and testicular regions and in the hepatic portal vein morphology. In the current study, the biometric values were similar to those previously published; however, no data regarding thoracic perimeter, modified crown-rump length or thoracic depth were found in the literature for this Canidae species. Statistical analysis showed the existence of a strong negative correlation between the modified crown-rump length and left renal length, between the modified crown-rump length and the right renal volume, between the thoracic perimeter and the height at the cranial pole of the left adrenal gland and between the thoracic perimeter and the height at the caudal pole of the left adrenal gland. Laboratory findings, including segmented neutrophil, eosinophil, monocyte and lymphocyte counts and the serum levels of glucose, ALT, alkaline phosphatase, urea, total protein, globulin, creatine phosphokinase, triglyceride, sodium, phosphate, potassium and chloride, were inconsistent with values found by other authors. The ultrasound is a diagnostic imaging method that must be further explored in the medicine of wild animals; therefore, additional research in this area is required.

O presente trabalho objetivou a caracterização ultrassonográfica das regiões abdominal e pélvica de cinco lobos-guarás mantidos em cativeiro no Centro de Triagem de Animais Silvestres da Universidade Federal de Viçosa, com descrição dos aspectos ultrassonográficos e obtenção de medidas de suas diversas estruturas, por meio de ultrassonografia modo-B. Coletaram-se dados de biometria corporal com a finalidade de se verificar a existência de correlação linear significativa entre estas medidas e as mensurações obtidas ultrassonograficamente. Adicionalmente, efetuou-se avaliação hematológica e bioquímica sérica dos animais. Os achados ultrassonográficos foram equiparados aos existentes na literatura de cães domésticos devido à inexistência de dados publicados referentes a lobos-guarás. Esta espécie apresentou características muito semelhantes às do cão, diferindo nas ecogenicidades do baço e córtex renal esquerdo, no aspecto da região prostática, testicular e na morfologia da veia porta hepática. No presente estudo, os valores de biometria corporal foram semelhantes aos anteriormente publicados, porém não se encontraram dados na literatura referentes a perímetro torácico, comprimento vértice-caudal modificado e profundidade torácica para esta espécie de canídeo. A análise estatística revelou a existência de forte correlação negativa entre o comprimento vértice caudal modificado e o comprimento renal esquerdo, entre o comprimento vértice caudal modificado e o volume renal direito, entre o perímetro torácico e a altura do polo cranial da glândula adrenal esquerda e entre o perímetro torácico e a altura do polo caudal da glândula adrenal esquerda. Laboratorialmente, a contagem de neutrófilos segmentados, eosinófilos, monócitos e linfócitos e os níveis séricos de glicose, ALT, fosfatase alcalina, ureia, proteína total, globulina, creatina fosfoquinase, triglicérides, sódio, fosfato, potássio e cloreto foram discordantes de valores obtidos por outros autores. A ultrassonografia é um método de diagnóstico por imagem que necessita ser mais explorado na medicina de animais silvestres, sendo, portanto necessário o incremento de pesquisas nesta área.

Stages and duration of the seminiferous epithelium cycle in the bat Sturnira lilium.

Knowledge of the stages that compose the seminiferous epithelium cycle (SEC) and determination of the duration of spermatogenic processes are fundamental for the accurate quantification of the dynamics of spermatogenesis. The aim of this study was to characterize the stages that compose the SEC of the bat Sturnira lilium, including evaluation of the average frequency of each of these stages throughout the year and calculation of the duration of the spermatogenic process. An ultrastructural characterization of the formation of the acrosomal cap was also performed. Testicular fragments were processed for morphological and immunohistochemical analysis as well as ultrastructural analysis using transmission electron microscopy. According to the tubular morphology method, the SEC in S. lilium is divided into eight stages, following the pattern found in other mammals. Primary spermatocytes were found at zygotene in stage 1 of the cycle. There was no variation in frequency of each of the stages over the seasons, with stage 1 being the most frequent, and stage 7 the least frequent. The duration of one seminiferous epithelium cycle was 3.45 days, and approximately 15.52 days were required for the development of sperm from spermatogonia. Ultrastructural characterization allowed the formation of the acrosomal cap in round spermatids to be monitored. In conclusion, the stages that compose the SEC in S. lilium are generally similar to those described for other mammals, but the duration of the spermatogenic process is shorter than previously recorded for mammals. The presence of primary spermatocytes at zygotene in stage 1 of the cycle is probably due to the longer duration of this stage.

Kinetics of spermatogenesis in the white-lipped peccary (Tayassu pecari).

This study aimed to characterize the stages of the seminiferous epithelium cycle by the tubular morphology method, and to determine the number of differentiated spermatogonia generations in the adult white-lipped peccary. Twenty adult white-lipped peccaries, obtained from commercial slaughterhouse, were used. Fragments of the testicular parenchyma were fixed in 3% glutaraldehyde and embedded into a methacrylate resin. The number of germ and Sertoli cells was estimated by the analysis of cell populations in 50 transversal sections of seminiferous tubules in different stages of the cycle. The tubular morphology method allowed the identification of cellular associations characteristic of the eight stages of the seminiferous epithelium cycle in white-lipped peccaries. The results showed the presence of six generations of differentiated spermatogonia in white-lipped peccaries, and that the cell composition of the eight stages of the seminiferous epithelium cycle in this species is very similar to that described for collared peccaries.

Intrinsic rate of spermatogenesis in free-ranging feral pigs (Sus scrofa sp).

The aim of this research was to evaluate the intrinsic rate of spermatogenesis in adult free-ranging feral pigs. Twelve adult male free-ranging feral pigs were captured, sedated, and orchidectomized, and then were released and observed to complete recovery and return to their natural environment. Fragments of the testes were embedded in plastic resin and used to prepare slides for histometric analysis. Characteristics investigated included cell populations in the seminiferous epithelium in stage 1 of the cycle of the seminiferous epithelium, intrinsic rate of spermatogenesis and Sertoli cell index. The efficiency coefficient of spermatogonial mitosis was 7.59, the meiotic index was 3.03, the overall yield of spermatogenesis was 23.97 and the cell loss ratio during the meiotic prophase was 1.04. Each Sertoli cell supported an average of 0.92 type A spermatogonia, 7.01 primary spermatocytes in preleptotene/leptotene, 7.30 primary spermatocytes in pachytene and 22.16 round spermatids. In conclusion, the results of the present study indicate that the supporting capacity of Sertoli cells in free-ranging feral pigs is among the greatest values reported for most domestic animals, and the overall yield of spermatogenesis is comparable to that reported in wild boars.

The effects of Tynnanthus fasciculatus (Bignoniaceae) infusion on testicular parenchyma of adult Wistar rats.

Traditional medicine provides strong guidance for scientific experiments involving plant products used by the Brazilian people. The species "cipó-cravo" (Tynnanthus fasciculatus) is a plant commonly used either to combat indigestion and stomach aches, or as a general stimulant and aphrodisiac. In this study, the effects of "cipó-cravo" infusion were investigated within the testicular parenchyma of adult Wistar rats. Rats were divided into 3 groups: a control (distilled water) and two treated groups, which received the plant infusion (100 and 200mg/animal/day). The 200mg dose promoted a significant increase of the testicular parenchyma weight and of the volume and total length of the seminiferous tubules, as well as in total daily sperm production and sperm production per gram of testis.

The effects of Tynnanthus fasciculatus (Bignoniaceae) infusion on testicular parenchyma of adult Wistar rats

Traditional medicine provides strong guidance for scientific experiments involving plant products used by the Brazilian people. The species "cipó-cravo" (Tynnanthus fasciculatus) is a plant commonly used either to combat indigestion and stomachaches, or as a general stimulant and aphrodisiac. In this study, the effects of "cipó-cravo" infusion were investigated within the testicular parenchyma of adult Wistar rats. Rats were divided into 3 groups: a control (distilled water) and two treated groups, which received the plant infusion (100 and 200mg/animal/day). The 200mg dose promoted a significant increase of the testicular parenchyma weight and of the volume and total length of the seminiferous tubules, as well as in total daily sperm production and sperm production per gram of testis.

Spermatogenesis in white-lipped peccaries (Tayassu pecari).

We describe here morphological and functional analyses of the spermatogenic process in sexually mature white-lipped peccaries. Ten sexually mature male animals, weighing approximately 39 kg were studied. Characteristics investigated included the gonadosomatic index (GSI), relative frequency of stages of the cycle of seminiferous epithelium (CSE), cell populations present in the seminiferous epithelium in stage 1 of CSE, intrinsic rate of spermatogenesis, Sertoli cell index, height of seminiferous epithelium and diameter of seminiferous tubules, volumetric proportion of components of the testicular parenchyma and length of seminiferous tubules per testis and per gram of testis. The GSI was 0.19%, relative frequencies of pre-meiotic, meiotic and post-meiotic phases were, respectively 43.6%, 13.8% and 42.6%, general rate of spermatogenesis was 25.8, each Sertoli cell supported an average 18.4 germinative cells, height of seminiferous epithelium and diameter of seminiferous tubules were, respectively, 78.4 microm and 225.6 microm, testicular parenchyma was composed by 75.8% seminiferous tubules and 24.2% intertubular tissue, and length of seminiferous tubules per gram of testis was 15.8m. These results show that, except for overall rate of spermatogenesis, the spermatogenic process in white-lipped peccaries is very similar to that of collared peccaries, and that Sertoli cells have a greater capacity to support germinative cells than most domestic mammals.