Effect of Different Vitrification Techniques on Viability and Apoptotic Index of Domestic Cat Testicular Tissue Cells.

Vitrification is essential for successful tissue cryopreservation and biobanking in wild cats. This study aimed to compare different methods of vitrification (Ovarian Tissue Cryosystem-OTC, Straws-STW, and Solid Surface vitrification-SSV) for testicular fragment vitrification in tom cats. Testicular fragments were recovered from five adult tom cats and subjected to equilibrium vitrification using different cryovials and methods under the same conditions of vitrification solutions and cryoprotectants. The efficiencies of the methods were evaluated using histological analysis of spermatogonia and Sertoli cell nuclei, seminiferous tubular basement membrane detachment, and the gonadal epithelium shrinkage score scale. Cell viability was assessed using Hoechst PI and Terminal deoxynucleotidyl transferase nick end labeling (TUNEL) assay. The results showed that OTC is an effective vitrification method for maintaining the distinction between spermatogonia and Sertoli cells. OTC was similar to the control for basal membrane detachment parameters (p = 0.05). Epithelial shrinkage was low in the SSV group, which showed the highest percentage of viable cells among the vitrified groups (p = 0.0023). The OTC and SSV vitrification methods were statistically similar in terms of the percentage of TUNEL-positive cells (p = 0.05). Therefore, OTC and SSV provide favorable conditions for maintaining viable cat testicular tissue cells after vitrification.

Casuistry of xenarthrans treated at the Veterinary Hospital of the Federal University of Pará, Brazilian Amazon / Casuística de Xenarthras atendidos no Hospital Veterinário da Universidade Federal do Pará, Amazônia brasileira

ABSTRACT: The present study aimed to analyze the casuistry of xenarthrans treated at the veterinary hospital of the Federal University of Pará to quantify the most important cases for clinical care and identify the main species, sex, age group, origin, and destination of the animals admitted. The data were collected from January 2013 to August 2020. The variables analyzed were the number of animals by species, sex, age group, weight (kilograms), form of admission (rescue or guardianship), person responsible for the admission of the animal, origin, circumstances of admission, length of stay (days), and evolution or destination of the animal (release, captivity, death, euthanasia, or escape). Fisher's exact test was used to determine whether disease incidence was related to sex and whether animal mortality was influenced by sex and disease. The chi-square test was used to determine whether mortality differed by age group. Ninety-two (92) xenarthrans were included: 66 Bradypus variegatus, 10 Choloepus didactylus, 15 Tamandua tetradactyla, and one Dasypus novemcinctus. The animals were mainly from the municipality of Castanhal (70%) and were acquired through rescue (88%, B. variegatus; 80%, C. didactylus; 100%, T. tetradactyla), a significant proportion of which was per natural persons (88%, B. variegatus; 80%, C. didactylus; 100%, T. tetradactyla). The most important circumstances that led to the admission of the animals were as follows: animals for examination (42%, n=39), young animals at risk (9%, n=8), hit by a vehicle (8%, n=7), respiratory disease (7%, n=6), fall from a tree (5%, n=5), and electrotrauma (5%, n=5). Release was the most common destination of animals after hospitalization (59% B. variegatus, 50% C. didactylus, and 66% T. tetradactyla). The sex of the animals did not affect the incidence of disease or mortality in any species. In contrast, mortality of B. variegatus was significantly higher in animals admitted with disease (p=0.000). Mortality in cubs of this species was higher than that in juveniles and adults (p=0.003).

RESUMO: O objetivo deste trabalho foi analisar a casuística dos Xenarthras atendidos no Hospital Veterinário da Universidade Federal do Pará (UFPA), a fim de quantificar as principais ocorrências para atendimento clínico, identificar as principais espécies, o sexo, a faixa etária, a procedência e destinação dos animais recebidos. Os dados foram obtidos entre janeiro de 2013 a agosto de 2020. As variáveis analisadas foram número de animais por espécie, sexo, faixa etária, peso (quilogramas), forma de recebimento (resgate, tutela), responsável pelo encaminhamento do animal ao hospital, procedência, circunstância de admissão, tempo de internação (dias) e evolução ou destinação do animal recebido (soltura, cativeiro, morte, eutanásia, fuga). O teste de Fisher foi utilizado para avaliar se a ocorrência de doenças estava relacionada ao sexo, bem como se a mortalidade animal foi influenciada pelo sexo e por enfermidades. O teste Qui-quadrado foi utilizado para avaliar se a mortalidade diferiu em relação à faixa etária. Um total de 92 animais Xenarthras foi admitido, composto por 66 Bradypus variegatus, 10 Choloepus didactylus, 15 Tamandua tetradactyla e um Dasypus novemcinctus. Os animais foram procedentes principalmente do município de Castanhal (70%), sendo o resgate a principal forma de aquisição dos animais (88%, B. Variegatus; 80%, C. Didactylus; 100%, T. tetradactyla), realizado em maior parte por pessoa física (88%, B. Variegatus; 80%, C. Didactylus; 100%, T. tetradactyla). O sexo dos animais não influenciou a ocorrência de doenças ou mortalidade em nenhuma espécie. Por outro lado, a mortalidade de B. variegatus foi significativamente maior naqueles animais admitidos com alguma enfermidade (p=0,000). A mortalidade em filhotes desta espécie foi maior quando comparada com as categorias jovem e adulta (p=0,003).

Epididymal tail solid-surface vitrification as an effective method for domestic cat sperm cryobanking.

This study aimed to describe the viability of domestic feline spermatozoa after epididymal tail vitrification. For this, 10 pairs of testis-epididymis complexes were used. The epididymal tails were vitrified using the solid-surface vitrification (SSV) method, in which two vitrification media containing ethylene glycol (EG) 40% or glycerol (GLY) 40% were tested. Vitrification with the presence of EG resulted in better results for all sperm motility parameters (motility, vigour and CASA) compared with GLY (P < 0.05). There were no statistical differences for sperm viability and acrosome integrity, plasma membrane integrity, or overall health of morphologically normal sperm before or after vitrification among experimental groups. In conclusion, epididymal tail vitrification appears to be a suitable method for long-term storage of cat sperm, especially if the procedure is performed with EG as the cryoprotectant.

Micromorphological and ultrastructural description of spermatozoa from squirrel monkeys (Saimiri collinsi Osgood, 1916).

Saimiri collinsi is used as an animal model in biotechnology research for conservation of species from the genus Saimiri. However, the development of biotechnologies depends on a proper knowledge of the sperm morphology to understand the basic aspects of sperm physiology, as potential male fertility depends on different cellular sperm structures. With this purpose, this study characterized the micromorphological and ultrastructural characteristics of squirrel monkeys (Saimiri collinsi) sperm using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). SEM electromyography revealed that a normal Saimiri collinsi sperm measures 71.7 ± 0.7 µm with lateral tail insertion, a paddle-shaped flattened head and an acrosome occupying most of the head. TEM also showed that the middle piece is characterized by a central 9 + 2 microtubule axoneme surrounded by nine dense fibres, and that the mitochondria were juxtaposed, forming the mitochondrial sheath. Here we provide the first micromorphological and ultrastructure description of S. collinsi sperm.

Single injection of eCG/hCG leads to successful estrous synchronization in the collared peccary (Pecari tajacu Linnaeus, 1758).

The establishment of protocols for the control of the ovarian function of collared peccaries is recommended for the development of assisted reproductive techniques. The goals were to (1) compare a gonadotropin combination with prostaglandin analogue to synchronize timing of onset of estrus among animals, and (2) elucidate the effects of the most desirable protocol for performing an artificial insemination study and macroscopic evaluation of the ovaries. Three of five females treated with a double administration of 120 µg prostaglandin (cloprostenol) at a 9-day interval expressed symptoms of estrus 9 days after the second injection. One female presented estrus after 6 days, whereas other did not respond to the treatment. All females (5/5) treated with a single dose containing 400 IU eCG and 200 IU hCG manifested estrus 6 days after the hormone injection. In a second experiment, ten females that were estrous synchronized using eCG/hCG, were artificially inseminated with fresh semen and monitored for pregnancy every 30 days. Although there was no detection of fetuses by ultrasonic examination, seven females (7/10) had greater than basal progesterone values for 60 days after the treatments were imposed. Ovaries from two females treated with eCG/hCG were collected 6 days post-injection. There was confirmation of an ovarian stimulation as a result of the presence of 88 and 25 antral follicles, as well as three and eight hemorrhagic structures in ovaries of each female, respectively. It, therefore, is proposed that eCG/hCG can be used as an effective treatment for estrous synchronization in collared peccaries.

B-mode ultrasonographic evaluation of long bones in Falconiformes and Strigiformes birds.

Bone fractures in birds are usually diagnosed with the aid of traditional radiography. However, this technique remains limited because of the difficulties associating this examination with real-time procedures. The aim of this study was to describe the use of B-mode ultrasound to assess the long bones of two avian orders. For the study, we used carcases of birds from the orders Falconiformes (n = 9) and Strigiformes (n = 12), with weights ranging from 108 to 1020 g. An ultrasound device with a 5-12 MHz linear probe was employed to produce images of the long bones (humerus, radius, ulna, femur and tibiotarsus). Ultrasound (US) measurements and physical measurements using a caliper were applied to compare the diameter of the bones. Images were also recorded from the US examination performed in two live patients attending the hospital with suspected bone fractures. No statistical difference was found between the two methods of measurement in carcases weighing up to 267 g (P > 0.01). The US examination provided relevant clinical information about the bone cortex and assisted in real-time surgical procedures. RESEARCH HIGHLIGHTS Long bones of Falconiformes and Strigiformes birds were assessed with B-mode ultrasound. Ultrasound analysis is a relevant tool in clinical orthopaedics for avian species. Ultrasound of the bone might be applied for monitoring of healing processes.

Vitrification of domestic cat (Felis catus) ovarian tissue: Effects of three different sugars.

We aimed to evaluate the effect of three extracellular cryoprotectants on the morphology of vitrified feline preantral follicles. Feline ovarian fragments (0.5 × 2 × 2 mm) collected from five domestic adult cats subjected to ovariohysterectomy for routine castration were vitrified with ethylene glycol (EG) 40% combined or not with sucrose (0.1 or 0.5 M), trehalose (0.1 or 0.5 M), or raffinose (0.1 M). After vitrification using the solid-surface method and warming of the tissues, cryoprotectants were washed out of the ovarian tissues, which were fixed for histological analysis. The percentages of normal follicles were similar to the control (fresh) (62.9 ± 4.1%) only for tissues exposed and cryopreserved with EG + trehalose at concentrations of 0.1 (35.8 ± 8.3%) and 0.5 M (33.4 ± 5.4%). All the other sugars decreased the percentages of morphologically normal follicles as compared to the control group and the trehalose groups. Based on the results of the present study, we recommend the use of trehalose as the extracellular cryoprotectant for the vitrification of feline ovarian tissue.

Development of fresh and vitrified agouti ovarian tissue after xenografting to ovariectomised severe combined immunodeficiency (SCID) mice.

The aim of the present study was to evaluate the development of fresh and vitrified agouti ovarian tissue after xenografting to C57Bl/6 severe combined immunodeficiency (SCID) female mice. Ovaries were obtained from five female agoutis and divided into 16 fragments. Five fragments were transplanted immediately to ovariectomised SCID mice and the others were vitrified, stored for 2 weeks and transplanted only after rewarming. Tissue fragments were transplanted under the kidney capsule in recipients. The return of ovarian activity in recipients was monitored by the observation of external signs of oestrus and vaginal cytology over a period of 40 days after transplantation, after which the grafts were removed and evaluated for morphology, cell proliferation and the occurrence of DNA fragmentation. Ovarian activity returned in four of five mice that received fresh ovarian tissue from agoutis and in one of six mice that had received vitrified tissue a mean (±s.e.m.) 20.6±8.6 days after xenotransplantation. After graft removal, a predominance of primordial and primary follicles was observed in all grafts. Vitrification reduced cell proliferation and increased the occurrence of DNA fragmentation in grafted agouti ovarian tissue. In conclusion, the present study demonstrates that xenografted agouti ovarian tissue, fresh or vitrified, is able to promote the return of ovarian activity in ovariectomised SCID C57B1/6 mice. However, improvements to vitrification protocols for agouti ovarian tissue are necessary.

Population estimate and morphometry of ovarian preantral follicles from three recently recognized squirrel monkey species: a comparative study.

We describe morphological and morphometrical characteristics of preantral ovarian follicles from three recently recognized Saimiri species: S. macrodon, S. cassiquiarensis and S. vanzolinii; the last one a threatened species. Ovaries from four adult monkeys were evaluated: one pair from a pregnant S. macrodon, two ovarian pairs from S. cassiquiarensis females (one of them pregnant), and one left ovary from a senile S. vanzolinii, applying classical histology. Follicular preantral population was quantified and morphology and morphometry of primordial, primary and secondary follicles were evaluated. Follicular preantral population varied among species, being 347,153 in the ovaries of the S. macrodon, 270,342 and 278,376 in the ovaries of both adult non-pregnant and pregnant S. cassiquiarensis females, and 28,149 in the ovary from a senile S. vanzolinii. Most follicles were at primordial or transition stages, except for the senile S. vanzolinii female, which presented the lowest percentages of primordial and transition follicles when compared with primary and secondary ones. Most preantral follicles (>70%) were morphologically normal in the ovaries from all studied S. macrodon and S. cassiquiarensis females, but the ovary of the senile S. vanzolinii female presented a significant decrease in the percentage of normal follicles (primordial: 61%, transition: 52%, primary: 54%, and secondary: 48%). In general, follicular diameter increased significantly from primordial to transition, and subsequently from primary to secondary follicles.

Extender supplementation with catalase maintains the integrity of sperm plasma membrane after freezing-thawing of semen from capuchin monkey.

We aimed to evaluate the effect of supplementation of ACP-118® extender with the antioxidant catalase (10 and 50 µg/ml) on Sapajus apella sperm motility, vigour, and plasma membrane integrity during the processes of seminal liquefaction, cooling, and freezing. Catalase did not affect any of the evaluated parameters after semen dilution or cooling. Cryopreserved sperm in the presence of 50 µg/ml catalase presented a plasma membrane integrity similar to that fresh sperm, however.

Unilateral ovarian absence in a Black-headed Squirrel Monkey (Saimiri vanzolinii Ayres, 1985), a threatened neotropical primate species.

Ovarian agenesis is an unusual anomaly with traumatic or congenital origin. In the present case report, we describe our findings in a senile S. vanzolinii female. As this neotropical primate species is listed as vulnerable, with limited geographic distribution in the Brazilian Amazonia, ovarian agenesis may be an important finding to be reported.

Morphologic analysis of sperm from two neotropical primate species: comparisons between the squirrel monkeys Saimiri collinsi and Saimiri vanzolinii.

Sperm morphometry can be applied to identify different animal groups and species and to evaluate sperm quality. Furthermore, knowledge on species-specific differences will help to enhance biological information, as well as to develop efficient reproductive technologies. The aims in the present study were to describe sperm morphometry from the recently characterized species S. collinsi and S. vanzolinii, to verify if the morphometric sperm patterns are similar or different between both species, and to determine if the sperm morphometry is affected by the levels of sperm defects using the S. collinsi as a model. Semen was collected from S. collinsi (n = 10) and S. vanzolinii (n = 2) monkeys, and sperm was submitted to morphological analysis. From the 10 samples from S. collinsi, five presented sperm of poor quality and two subgroups were formed for this species, i.e. high and poor quality sperm. Data on sperm motility and vigour were analysed, as well morphometric parameters on sperm head and tail. It was observed the normal morphometry was correlated with high quality sperm. Poor quality sperm presented smaller and 7% more ellipticity in their head, when compared with high quality sperm. Sperm from S. vanzolinii presented larger head than those from S. collinsi, but tail lengths were similar. Sperm morphometry can be used as a complementary tool to predict sperm motility and vigour for the S. collinsi species, and S. collinsi appear as a suitable model for S. vanzolinii.

Detrimental Effect of Phenol Red on the Vitrification of Cat (Felis catus) Ovarian Tissue.

The aim of this study was to investigate the effects of different media with or without phenol red or the antioxidant trolox on the successful vitrification of feline ovarian tissue. In a first experiment, ovarian cortical pieces from three cats were vitrified in solutions of Roswell Park Memorial Institute (RPMI)-1640 medium, Minimum Essential Medium, Dulbecco's modified Eagle's medium, or Tissue Culture Medium 199 as basic medium, supplemented or not with 50 µM of trolox, all containing 40% ethylene glycol (EG) and 1 M of sucrose. RPMI-1640 (phenol red-free) without trolox was the only medium that preserved the percentage of morphologically normal preantral follicles similar to control (80%). The main difference between RPMI-1640 and the other media was the absence of phenol red and CaCl2. In a second experiment, ovarian cortical pieces from three cats were vitrified in a solution containing RPMI-1640 as basic medium, 40% EG, 1 M of sucrose, supplemented or not with phenol red or CaCl2 alone, or in combination. It was observed that phenol red supplementation led to follicular degeneration. Finally, to evaluate the interaction between phenol red and the cryoprotectant agent (i.e., EG), ovarian tissue was exposed to RPMI-1640 supplemented with phenol red and EG at different concentrations (10%, 20%, or 40%). There was an inverse relationship between EG concentration and free phenol red in the medium after exposure. It is suggested that vitrification of feline ovarian tissue should be performed in a phenol red-free medium. Medium supplementation with 50 µM of trolox was deleterious for follicular morphology.

Ovarian and uterine periovulatory Doppler ultrasonography in bitches / Ultrassonografia Doppler ovariana e uterina em cadelas

This paper aims to describe the uterine and ovarian ultrasonographic characteristics and Doppler velocimetric features of their arteries in bitches during the periovulatory period. Fifteen estrous cycles in 10 animals were evaluated. The ultrasonographic characteristics, resistance indices (RI) and pulsatility indices (PI) of the uterus and ovaries in each animal were recorded 5 days before and after ovulation (D0). The data were statistically analyzed, and the results were expressed as the mean ± standard error of mean (P<0.05). In results the ultrasonographic features of the uterus were the same on all of the cycles and evaluated days. The uterus had an average diameter of 0.85±0.02cm. An increase in the volume of the ovaries and the diameter of the ovarian follicles were measured. Ovaries had a volume of 0.64±0.06cm³, and the follicles cavities had a diameter of 0.46 ± 0.01 cm on the day of ovulation. After ovulation, it was observed that some follicles not collapse in some cycles. Two days prior to ovulation, the uterine blood perfusion decreased. This decrease remained unchanged until ovulation. Following ovulation, we measured a gradual increase in the uterine perfusion and in the ovarian artery. This artery directed blood flow to the ovaries and increased the intra-ovarian perfusion on the day after ovulation. In conclusion, specific features are observed in the uterus and ovarian ultrasound image and Doppler values of their arteries presented on the periovulatory days and when associated allow to estimate more accurately the date of ovulation.(AU)

Este trabalho teve como objetivo descrever as características ultrassonográficas uterinas e ovarianas, e dopplervelocimétricas das suas artérias nos dias periovulatórios em cadelas. Quinze ciclos estrais em 10 animais foram avaliados. As características ultrassonográficas, índices de resistência (IR) e índices de pulsatilidade (IP) do útero e dos ovários em cada animal foram registrados 5 dias antes e depois da ovulação (D0). Os dados foram analisados estatisticamente e os resultados foram expressos em média ± erro padrão da média (P<0,05). Como resultados as características ultrassonográficas uterinas foram semelhantes em todos os ciclos e dias avaliados, tendo o corpo uterino um diâmetro médio de 0,85±0,02cm. Foi observado um aumento no volume dos ovários e nos diâmetros dos folículos, tendo os ovários um volume de 0,64±0,06cm³, e a cavidade dos folículos um diâmetro de 0,46±0,01cm no dia da ovulação. Após a ovulação, foi observado colapso dos folículos somente em alguns ciclos. A perfusão sanguínea uterina diminuiu dois dias antes da ovulação e permaneceu inalterada até a ovulação. Após a ovulação, houve um aumento gradual na perfusão das artérias uterinas e ovarianas, direcionando o fluxo de sangue para os ovários e aumentando a perfusão da artéria intra-ovariana um dia após a ovulação. Conclui-se que características específicas são observadas na imagem ultrassonográfica uterina e ovariana, e dopplervelocimétricas de suas artérias nos dias periovulatórios, e quando associadas permitem estimar com mais precisão o dia da ovulação.(AU)

Ovarian and uterine periovulatory Doppler ultrasonography in bitches / Ultrassonografia Doppler ovariana e uterina em cadelas

This paper aims to describe the uterine and ovarian ultrasonographic characteristics and Doppler velocimetric features of their arteries in bitches during the periovulatory period. Fifteen estrous cycles in 10 animals were evaluated. The ultrasonographic characteristics, resistance indices (RI) and pulsatility indices (PI) of the uterus and ovaries in each animal were recorded 5 days before and after ovulation (D0). The data were statistically analyzed, and the results were expressed as the mean ± standard error of mean (P<0.05). In results the ultrasonographic features of the uterus were the same on all of the cycles and evaluated days. The uterus had an average diameter of 0.85±0.02cm. An increase in the volume of the ovaries and the diameter of the ovarian follicles were measured. Ovaries had a volume of 0.64±0.06cm³, and the follicles cavities had a diameter of 0.46 ± 0.01 cm on the day of ovulation. After ovulation, it was observed that some follicles not collapse in some cycles. Two days prior to ovulation, the uterine blood perfusion decreased. This decrease remained unchanged until ovulation. Following ovulation, we measured a gradual increase in the uterine perfusion and in the ovarian artery. This artery directed blood flow to the ovaries and increased the intra-ovarian perfusion on the day after ovulation. In conclusion, specific features are observed in the uterus and ovarian ultrasound image and Doppler values of their arteries presented on the periovulatory days and when associated allow to estimate more accurately the date of ovulation.

Este trabalho teve como objetivo descrever as características ultrassonográficas uterinas e ovarianas, e dopplervelocimétricas das suas artérias nos dias periovulatórios em cadelas. Quinze ciclos estrais em 10 animais foram avaliados. As características ultrassonográficas, índices de resistência (IR) e índices de pulsatilidade (IP) do útero e dos ovários em cada animal foram registrados 5 dias antes e depois da ovulação (D0). Os dados foram analisados estatisticamente e os resultados foram expressos em média ± erro padrão da média (P<0,05). Como resultados as características ultrassonográficas uterinas foram semelhantes em todos os ciclos e dias avaliados, tendo o corpo uterino um diâmetro médio de 0,85±0,02cm. Foi observado um aumento no volume dos ovários e nos diâmetros dos folículos, tendo os ovários um volume de 0,64±0,06cm³, e a cavidade dos folículos um diâmetro de 0,46±0,01cm no dia da ovulação. Após a ovulação, foi observado colapso dos folículos somente em alguns ciclos. A perfusão sanguínea uterina diminuiu dois dias antes da ovulação e permaneceu inalterada até a ovulação. Após a ovulação, houve um aumento gradual na perfusão das artérias uterinas e ovarianas, direcionando o fluxo de sangue para os ovários e aumentando a perfusão da artéria intra-ovariana um dia após a ovulação. Conclui-se que características específicas são observadas na imagem ultrassonográfica uterina e ovariana, e dopplervelocimétricas de suas artérias nos dias periovulatórios, e quando associadas permitem estimar com mais precisão o dia da ovulação.

Adaptation of a trap door technique for the recovery of ovarian cortical biopsies from Cebus apella (capuchin monkey).

There is a paucity of efficient cryopreservation protocols for primordial follicles enclosed in the ovarian tissue from non-human primates (NHP), in special New World primates. Our objective was to establish an optimal procedure for the recovery of ovarian biopsies from capuchin monkeys. To this end, we adapted a trap door biopsy method. Follicular density and quality of the biopsies were evaluated and ultrasound analysis was performed before and continuously after surgery to assess ovarian structure. Ovarian tissue biopsies recovered by the trap door technique allowed the successful harvesting of primordial follicles from capuchin monkeys, and no complication was recorded. The female cycle was not affected by surgery and no adherence was found thereafter. In conclusion, the adaptation of a trap door biopsy method is a safe procedure and allows recovery of healthy primordial follicles.

Embryo production by parthenogenetic activation and fertilization of in vitro matured oocytes from Cebus apella.

The efficiency of in vitro fertilization (IVF) depends on the viability of spermatozoa. For capuchin monkeys (Cebus apella), in vitro capacitation of spermatozoa is challenging because of their unique seminal coagulum. Motile spermatozoa can be obtained after liquefaction of the semen coagulum in coconut water-based solution. The objective of the present study was to establish an optimal in vitro maturation (IVM) protocol for capuchin monkeys and to observe the effect of follicle stimulating hormone (FSH) and luteinising hormone (LH) on IVF and parthenogenetic activation (PA) of oocytes collected from unstimulated females. We assessed spermatozoa quality after recovery from seminal coagulum using the solution ACP-118® as an extender. Oocytes were matured in vitro for 36 or 40 h and subjected to IVF or PA by applying ionomycin combined either with 6-dimethylaminopurine (6-DMAP) or roscovitine. In total, 87% of oocytes reached metaphase II (MII) after 40 IVM and 4-cell embryo production was obtained after IVF and parthenogenesis using ionomycin/6-DMAP. ACP-118® was used successfully to harvest viable spermatozoa from semen coagulum and in the preservation of spermatozoa, which were able to fertilize oocytes in vitro.

Vitrification of ovarian tissue from primates and domestic ruminants: an overview.

In the last decade, vitrification protocols to preserve human ovarian tissue have been regularly reported, even more often than the protocols developed for large mammals, such as ruminants and nonhuman primates. In order to facilitate the use of domestic ruminants (cows, goats, and sheep) and nonhuman primates as animal models, application of similar protocols as used for human material is performed. Next to it, the addition of indispensable or exclusion of avoidable compounds in the vitrification of human ovarian tissue should be tested in such experiments with animal models. The objective of this mini-review is to summarize the current protocols used for the vitrification of ovarian tissue and to evaluate the vitrification methods in humans, nonhuman primates, and domestic ruminants.

Unilateral ovarian absence in two capuchin monkeys.

BACKGROUND: Ovarian absence is a rare condition with congenital or traumatic origin. METHODS: Gross and microscopic examination of two capuchin monkeys were performed. RESULTS: Case 1 had absence of the right ovary and distal uterine tube, bilateral absence of fimbriae, and a rudimentary uterus. Case 2 had unilateral ovarian absence, but a functional contralateral ovary, and a normal uterus. CONCLUSIONS: Unilateral ovarian absence was confirmed in two capuchin monkeys for the first time.

Semen coagulum liquefaction, sperm activation and cryopreservation of capuchin monkey (Cebus apella) semen in coconut water solution (CWS) and TES-TRIS.

The objectives of the present study were to test the effect of coconut water solution and TES-TRIS on the seminal coagulum liquefaction, sperm activation in fresh diluted semen, and on the cryopreservation of semen from capuchin monkeys (Cebus apella). Semen was collected from six males by electro-ejaculation, diluted in TES-TRIS or coconut water solution (CWS), and incubated at 35°C until the coagulated fraction of the semen was completely liquefied. In the experiment I, after liquefaction, samples were diluted in TES-TRIS or CWS, plus 6 and 10mM/mL of caffeine. Sperm motility and vigor were evaluated during 5h. For experiment II, after liquefaction, semen samples were extended in TES-TRIS (3.5% glycerol in the final solution) or CWS (2.5% glycerol in the final solution), cryopreserved and stored in liquid nitrogen for 1 week. The seminal coagulum was liquefied in (mean±SDM) 4.5±1.7 and 2.8±1.1h in TES-TRIS and CWS, respectively. Sperm were motile in TES-TRIS and CWS for 5.0±1.4 and 1.0±0.5h, respectively. The mean motility in this period was 38±22% (TES-TRIS) and 22.0±16.0 (CWS). Motility increased after caffeine addition only in samples diluted in CWS containing 6mM (22.5±16.0) or 10mM (28.0±19.0) caffeine. Post-thaw live sperm percentage was 26.2% in TES-TRIS and 13.2% in CWS. For cryopreservation of semen from C. apella TES-TRIS (3.5% glycerol) was more appropriate than CWS (2.5% glycerol). CWS+caffeine potentially increase sperm motility and may be useful in artificial insemination of fresh diluted semen.