Effect of different concentrations of Trolox® in association with docosahexaenoic acid on equine semen freezing.

The aim of this study was to evaluate the association of different concentrations of Trolox® and the addition of a fixed concentration of DHA in the freezing of semen of Mangalarga Marchador stallions. To that end, 16 ejaculates were frozen in the following extenders: E1) BotuCrio® (BC; Control); E2) BC + 50 ngml-1 DHA + 30 µM Trolox® (BCDHA30T); E3) BC + 50 ngml-1 DHA + 40 µM Trolox® (BCDHA40T); E4) BC + 50 ngml-1 DHA + 50 µM Trolox® (BCDHA50T). All the tested extenders were similar in preserving different kinematic parameters, cell functional integrity, compacted DNA, and high and intermediate mitochondrial activity (P>0.05). However, sperm cryopreserved in BCDHA40T showed higher velocities than sperm frozen in the control extender (P<0.05). The 30 µM concentration of Trolox® was worse for sperm motility and the 50 µM concentration of Trolox® did not adequately preserve the structural integrity of the membranes in an extender containing DHA when compared to the BotuCrio® (P<0.05) extender. The use of Trolox® in freezing extenders containing DHA did not maximize the effect of BotuCrio®, except for in the case of sperm velocity parameters when at a concentration of 40 µM.

Testicular morphometry as a tool to evaluate the efficiency of immunocastration in lambs.

This study aimed to evaluate the efficiency of immunocastration in lambs using testicular morphometry. Thirty lambs were randomly divided into two treatments (subcutaneous administration of 1.0 mL and 0.5 mL of an anti-GnRH vaccine) and a control group (1.0 mL saline solution). The animals were vaccinated at four months of age, received a second dose 30 days later, and were slaughtered 90 days after the first vaccine dose. After slaughter, testicles were collected, and samples were removed for histological processing and evaluation of testicular morphometric parameters. Analysis of variance, Tukey's test, and Kruskal-Wallis test were performed, with a 5% level of significance. There was a reduction in testicular weight, gonadosomatic index, seminiferous tubule diameter, germinal epithelium height, leydigosomatic index, and total tubule length. The total length per testicular gram increased in the immunocastrated group. Intrinsic spermatogenesis yield, Sertoli cell indices, and estimates of sperm and Sertoli cell production were reduced in the immunized groups (P < 0.05). The anti-GnRH vaccine in lambs at doses of 1.0 mL and 0.5 mL is sufficient to promote immunocastration, verified through severe changes in testicular morphometry from animals.

Flaxseed added to the diet of Alpine goats affects the nutrients intake and blood parameters.

The flaxseed is a nutraceutical food used as a source of α-linolenic acid, which can bring benefits to the health of mammals. This study was carried out to examine the effect of flaxseed inclusion in the diets on the intake of nutrients, body weight, and blood parameters of Alpine goats. Twenty-one adult females with an initial average weight of 41.06 ± 1.84 kg were used in a completely randomized design, with four experimental treatments (0, 5, 10, and 15% of flaxseed in the total diet) and five replications per treatment. The intake of the dry matter presented a decreasing linear effect (P < 0.001), with a reduction of 53.5% between the control diet (0% of flaxseed) and 15% of flaxseed, but no effect was observed on weight gain (P > 0.05). Inclusion of flaxseed from 0 to 15% linearly decreased the intakes of organic matter, crude protein, non-fibrous carbohydrates, and neutral detergent fiber, but increased the ether extract intake (P < 0.001). Regarding plasma concentration traits, increasing the flaxseed levels from 0 to 15% had linear positive effect on LDL, VLDL, and triglycerides (P < 0.05), but no effect on cholesterol, HDL, glucose, creatinine, and urea (P > 0.05). The inclusion of flaxseed in the diet for 80 days changes the nutrients intake and blood parameters but shows no impact on body weight. However, further studies are required to determine the impact of flaxseed on the goat's health in the long term since there has been an increase in the concentration of triglycerides, LDL, and VLDL.

Docosahexaenoic acid in diluent for goat semen cryopreservation.

The effects of docosahexaenoic acid (DHA) in the diluent for cryopreservation of goat semen on seminal quality and the optimal levels to be used were evaluated. After collection, semen was pooled and physically evaluated, then divided into four aliquots with different DHA levels in the diluent: 0, 10, 20, and 30 ng mL-1. The semen was cryopreserved in a TK 3000® freezing machine and then thawed for assessment at 37 °C. Sperm motility and vigor, membrane integrity, acrosomal integrity, mitochondrial activity, and sperm chromatin compaction were evaluated after thawing. A completely randomized design was used. For normally distributed variables, ANOVA and regression analysis were used to test for differences between treatments, and for non-parametric data, the Kruskal Wallis test was used at the 5% significance level. There were no differences among groups in terms of membrane integrity, acrosomal integrity, or chromatin compaction. There was a decrease in class I mitochondrial activity with increasing DHA level (P<0.05), but no differences in classes II, III, and IV (P>0.05). The inclusion of 10 to 30 ng mL-1 of DHA in the diluent did not result in improvements in seminal quality parameters after thawing, with some impairment observed in the mitochondrial activity of the sperm cells.

Reproductive parameters of lambs immunocastrated with anti-GnRH vaccine.

The objective of this study was to evaluate the testicular biometric, seminal, and plasma testosterone levels in lambs subjected to an anti-GnRH vaccine as a method of castration. Thirty entire, crossbred Santa Inês male lambs were randomly distributed into three treatment (T): T1 was the control group, with the administration of 1 mL of saline solution subcutaneously (SC); 1.0 and 0.5 mL of an anti-GnRH vaccine were administered SC in T2 and T3, respectively. Testicular biometric variables, physical and morphological variables of semen, and plasma testosterone concentrations were evaluated. At D60, there was a reduction in testicular length, width, thickness, and scrotal circumference of the immunocastrated animals regardless of the vaccine dose used (P < 0.05). A reduction in semen physical variables at both dosages (P < 0.05) was observed, with azoospermia, in 80% and 70% of animals in the T2 and T3 groups, respectively. At D60, the immunocastrated animals also showed an increase in spermatozoa defects (P < 0.05), whereas plasma testosterone concentration decreased (P < 0.05). Immunocastration of lambs using the Bopriva vaccine at doses of 1.0 and 0.5 mL is efficient in inducing azoospermia in up to 80% of animals, although two doses in a 30-day interval are necessary for it to be an effective and safe method. Efficacy was demonstrated through a reduction in serum testosterone levels, testicular biometry, and seminal fluid analysis. Considering the efficacy of both doses in this study, we recommend using the lower dose (0.5 mL), which will allow for a 50% reduction in vaccine costs.

Effects of equine chorionic gonadotropin administered via the Baihui acupoint on follicular ovarian dynamics and the luteal function of cattle during an ovulation synchronization treatment regimen for fixed-time artificial insemination.

Objectives were to evaluate effects of a smaller than typically used dose of equine chorionic gonadotropin (eCG) during a fixed-time artificial insemination (FTAI) treatment regimen. Transrectal ultrasonic (US) examinations were conducted on dairy cows on Day 0 (D0) and the treatment regimen was initiated with administrations of an intravaginal progesterone (P4) implant, estradiol benzoate (im), and prostaglandin F2α (PGF2α; im). On D8, the P4 implant was removed and PGF2α and estradiol cypionate were administered to all animals. Subsequently, cows were randomly assigned to three groups and eCG was administered to Groups 1, 2, and 3 in doses of 300 (im); 100 (im); and 100 (Baihui acupoint) IUs, respectively. The B-mode and power-flow US cineloops were performed to assess follicular dynamics and evaluate various morphological and vascular characteristics of the corpus luteum. Blood samples were collected to quantify serum P4 concentrations. There were no differences between the ovulation synchronization treatment regimens for all follicular dynamic variables tested; however, cows in Group 3 differed from Group 2 having a larger follicle diameter (FD) on D10 (P = 0.06) and larger preovulatory FD (P = 0.09), as well as a blood perfusion area of the dominant follicle wall on D8 (P = 0.07). There were no differences in responses to the ovulation synchronization treatment regimens for the luteal variables evaluated subsequent to ovulation. In conclusion, the Baihui acupoint was effective as an alternative route for eCG dose reduction when FTAI treatment regimens were imposed without detrimentally affecting values for reproductive variables.

Subdose of human chorionic gonadotropin applied at the Hou Hai acupoint on follicular dynamics and luteal development in donkeys.

The objective of this study was to evaluate the effects of an hCG subdose applied at the Hou Hai acupoint as an ovulation inducer in donkeys. Eleven donkeys were distributed in randomized blocks in T1 = application of 1,500 IU of hCG intravenous (IV); T2 = 450 IU of hCG applied at the false acupoint (IV), and T3 = 450 IU of hCG applied at the Hou Hai acupoint. There was no difference (P > 0.05) between the treatments regarding the mean diameter of the pre-ovulatory follicle (34.5 ± 1.3 mm), the ovulation rate (96.97%), the interval between induction and ovulation (58.07 ± 16.82 h), the mean diameter of the CL (D0 = 23.0 ± 0.6; D2 = 27.7 ± 1.9 and D8 = 28.2 ± 0.8mm), and serum P4 concentrations (10.50 ± 2.99 ng.mL-1). The application of 450 IU of hCG at the Hou Hai acupoint increased ovulation rate (72.73%) more than 48 h after induction (P = 0.03) and a larger diameter of the CL on D4 (30.7 ± 5.1 mm) (P = 0.04). The vascularization area of the CL on D8, obtained by minimum number of colored pixel (NCP), was greater (P < 0.05) in the donkeys that received 1,500 IU of IV hCG (T1, 41.91 ± 1.17), and we found a positive correlation (P < 0.05) between mean NCP and P4 concentration in the donkeys that received 450 IU of hCG IV at the false acupoint (T2) or at the Hou Hai acupoint (T3). The application of 450 IU of hCG by IV route at the false acupoint or the Hou Hai acupoint was sufficient to induce ovulation in donkeys, demonstrating that the average dosage commonly used for this species is too high.

Immunocastration of goats using anti-gonadotrophin releasing hormone vaccine.

The aim of this study was to evaluate the immuno-sterilizing action of anti-gonadotrophin-releasing hormone (anti-GnRH) vaccine in goats. Eighteen male goats were randomly distributed to receive three treatments: T1 (control) - whole animals, and T2 and T3 - application of 0.5 and 1.0 mL of anti-GnRH vaccine, respectively, with six replicates and one goat per experimental unit. Vaccine was administered at 8 months of age and 30 days after the first immunization. Testicular biometry was evaluated monthly, along with seminal collections, for the physical and morphological evaluation of semen. At the time of slaughter, the testicle were collected, and fragments were measured and removed for histological evaluation. The data were evaluated for normality by the Shapiro-Wilk test, followed by appropriate statistical tests for each variable. A reduction in width and length of the right and left testicles was observed and, consequently, the scrotal circumference of the immunized animals reduced after the second vaccine application (P < 0.05). Thirty-days after the first vaccine application, there was a negative effect on seminal production and quality; and 60 days after the second application, a pronounced reduction was observed in all seminal parameters in the vaccinated animals, including azoospermia (83.33% of animals; P < 0.05). Vaccine application reduced testicular weight, seminiferous tubule diameter, and gonadosomatic and tubulosomatic index (P < 0.05), but did not influence the proportion of testicular parenchyma components (P > 0.05). Two applications of the anti-GnRH conjugate are effective for the immunological castration of goats, and the 0.5 mL dose is recommended for use in crossbred goats.

Follicular response and oocyte production following variations in ovarian stimulation in goats.

In this study, goats were subjected to ovarian stimulation protocols to evaluate possible differences in ovarian follicular responses and oocyte production. Two experiments were conducted to assess the effects of hormonal protocol duration (seven or twelve days) and number of follicle stimulating hormone (FSH) applications (one or five doses). All animals received intravaginal sponges saturated with 60 mg medroxyprogesterone acetate and an application of 125 µg cloprostenol 72 h before the sponges were removed. For ovarian stimulation, 120 mg FSH was applied in a single dose 36 h before laparoscopic follicular aspiration (LOPU) or in five doses (30, 30, 20, 20, and 20 mg) at 12 h intervals, with the last dose applied 36 h before LOPU. In the first experimental phase, ultrasonography was performed to monitor follicular number and diameter, and in the second phase, the animals received LOPU to count the follicles and cumulus-oocyte complexes (COCs) and for morphological classification. There was no significant effect (P > 0.05) of any variable or combination of variables on follicle number on Day (D) 0 or D3/D8 (day of LOPU). However, evaluation at D6/D11 revealed an effect (P < 0.05) from the protocol duration with the highest number of small follicles resulting from the short protocol. There was also an effect (P < 0.05) of FSH dose number on the resulting number of medium and large follicles, with more medium follicles recovered after a single dose and more large follicles after multiple doses. There was no effect of any variable (P > 0.05) on the diameters of the large, medium, and small follicles, except for the D4/D9 evaluation, which showed a combination effect for large follicles (P < 0.05). In the second phase, no variable had an effect on the number of follicles visualized or aspirated, number of COCs recovered, recovery rate, morphological quality of COCs in grades 1, 2, 3, and 4, or rate of viable COCs. Thus, all four protocols efficiently stimulated the ovarian response and oocyte production of goats, although the short protocol (7 days) with a single dose of FSH was most efficacious based upon the greater number of medium-sized follicles available for aspiration, the shorter time, and greater practicality of execution.

Aspectos clínico-macroscópicos da agenesia pulmonar unilateral esquerda em filhote de jabuti (Chelonoidis carbonarius) / Clinical and macroscopic aspects of left unilateral pulmonary agenesis in tortoise hatchling (Chelonoidis carbonarius)

A agenesia pulmonar é uma anomalia congênita extremamente rara, representada pela falha no desenvolvimento dos brotamentos pulmonares, que geralmente acontece unilateralmente. O objetivo do trabalho foi relatar aspectos clínico-macroscópicos da ocorrência de agenesia pulmonar unilateral em filhote de Chelonoidis carbonarius. Ovos de jabutis (n = 83) foram incubados artificialmente a 29ºC durante 131 dias, sendo que somente 10 eclodiram. No momento da eclosão, todos os filhotes apresentavam aspecto morfológico normal. Os filhotes foram alimentados diariamente com frutas e verduras até completarem aproximadamente dois meses de idade, quando foram eutanasiados. Durante este período, observou-se que um dos filhotes apresentou como sinais clínicos respiração ofegante e pela boca, dificuldade em se alimentar e pouco deslocamento dentro da caixa, ficando isolado dos demais filhotes. À necropsia, este apresentou ausência do pulmão esquerdo, baixo desenvolvimento de órgãos e dilatação cardíaca. O caso traz como relevância a ocorrência da agenesia pulmonar unilateral à esquerda, sem associação com outras malformações.

Pulmonary agenesis is an extremely rare congenital anomaly, represented by the failure in the development of lung buds, which usually occurs unilaterally. The aim of study is present a report of clinical and macroscopical aspects and consequences for body development in the event of unilateral lung agenesis in tortoise's hatchlings (Chelonoidis carbonarius). The eggs of tortoises were incubated artificially at 29 °C for 131 days. Eighty-three eggs were incubated and only 10 hatched. At hatching, all hatchlings had presented normal morphological appearance. The hatchlings were fed daily with fruits and vegetables for two months, when they were euthanized. We observed a case in which the tortoise showed signs of pulmonary agenesis of the left side, without association with other malformations. Clinical signs included wheezing, breathing through the mouth, difficulty in feeding and little movement within the box. At necropsy, we observed the absence of the left lung, reduced size of viscera and cardiac dilatation. The case brings relevance to the occurrence of unilateral lung agenesis at left in tortoise's hatchling, with description of the clinical signs and macroscopic findings during necropsy.