Sperm motility of Prochilodus lineatus in relation to dilution rate and temperature of the activating medium

The objective of this study was to assess the effects of an activating solution on the sperm motility duration (SMD) of 'curimbatá', Prochilodus lineatus through of the definition of qualitative and quantitative parameters of the semen pool used in the experiment; evaluation of the effects of different ratios of semen dilution corresponding to 1-:-1, 1-:-2, 1-:-20, 1-:-200, 1-:-2000, 1-:-20-000 and 1-:-100-000 semen:dilute solution on the SMD and, assessment of the effects of different temperatures of the activating solution (5, 10, 15, 20, 25, 30, 35, 40, 45 and 50°C) on the SMD. The results of SMD were directly proportional to the dilution (P<0.05), starting from the dilution of 1-:-2 (semen:water), with 23.04-s. Were used three replicates of the semen pool for each test. Two-year-old brookstock were maintained in ponds culture conditions. In November-December 2006, twelve mature males broodfish were selected (mean weight and length of 405.8±134.2-g and 25.6±3.1-cm, respectively). The males released that semen under slight pressure of the urogenital papilla were selected for the experiment. The SMD increased proportionally to the increase in dilution, until it reached a maximum of 28.83-s for the ratio 1-:-100-000 semen: dilute solution. The results of SMD in relation to the temperature of the activating solution exhibited a quadratic behavior (P<0.05) with a maximum theoretical performance in terms of sperm motility duration of 21.36-s at a temperature of 17.3°C. Thus, for the species considered, the increase in the dilution ratio proved favorable for the rise in motility duration until the maximum value studied of 1-:-100-000 semen:dilute solution. As for the temperature of the activating solution, the best results of SMD were obtained at the temperature of 17.3°C. At higher temperatures used in the experiment (25, 30, 35, 40, 45 and 50°C), a decrease in motility duration. © 2010 Blackwell Verlag, Berlin.

Reproductive technology in domestic carnivorous

Background: The delay in development of artificial reproduction techniques on carnivorous could be due to countless reasons, but the lack of commercial interest is probably the most important one. The majority of canines are small structures, canidae are extremely fertile and a great number of species are adapted to domestication or captivity. Finally, the canine gamete physiology presents a difficult adaptation of technology knowledge obtained from other species. Furthermore, domestic felines are animals of company and there is no interest in reproducing them in a large scale, as it has been observed in other domestic animals, however, besides of being a valuable model for the development of in vitro techniques, the domestic cat is also used as an embryo receptor for different species of small wild felines due to physiological similarities among them, in vitro embrionary development, Review: It was reviewed the main insights about the reproductive physiology in female dogs, in vitro oocytary maturation (IVM), pregnancy and conception rate with dogs' frozen/unfrozen semen and PIV in domestic cats. The majority of mammal oocytes restart meiosis spontaneously after ovulation and reaches MII in artificial environment; in an in vitro maturation system in bovines, around 90% of oocytes complete their maturation, although its development capacity can be reduced subsequently. The success of IVM in canidae have been limited, with maturation rate varying from 0 to 58%, usually around 20%. The greatest difficulties include oocyte quality, hormonal environment, protein supplementation, cumulus / oocyte cell interaction, donor breed and age, culture systems, oxygen tension, amino acids, growth factor and sequential means. The freezing process reduces the quality of the semen, firstly because it reduces the number of living sperms and secondly because freezing produces cell modifications that could alter the sperm motility, longevity, integrity of membranes and its fertilizing capacity. Conclusion: Nowadays, several researches are being performed with the aim of increasing viability after dogs' and cats' semen is unfrozen, using extenders, cryoprotectors, freezing and unfreezing curves, addition of antioxidant substances. The aim of this text is to inform about the improvements obtained on the artificial reproduction techniques, emphasizing the oocytary maturation in female dogs, semen cryopreservation and artificial insemination in domestic dogs and cats.

Bactérias e resposta inflamatória no útero de cadelas, após inseminação artificial

The present study aimed to obtain information about the uterine inflammatory response (number of polymorphonuclear neutrophilic granulocytes - PMNs) in bitches after artificial insemination (AI) and identify the uterine microflora present after the following treatments: insemination using semen with extender (n=6), insemination with fresh semen (n=6) and no inseminated (n=6). The percentage of PMNs on the endometrial surface and within histological sections was evaluated together with the presence of aerobic bacteria in the uterine lumen. For endometrial cytology, there was no significative difference on the number of inflammatory cells between bitches not inseminated (3.05 ± 1.74 PMNs) and those inseminated with fresh semen (3.55 ± 1.51 PMNs); There was a significative difference in both groups compared to the inseminated with semen plus extender (7.80 ± 1.67 PMNs) (p<0.05). Histology showed that there was no significative difference on the number of inflammatory cells between bitches not inseminated (87.72 ± 35.2 PMNs) and those inseminated with fresh semen (122.97 ± 43.31 PMNs); however, it was observed differences in both groups compared to those inseminated with semen plus extender (171.94 ± 42.74 PMNs) (p<0.05). Eight animals, randomly distributed in the groups, showed the presence of Staphylococcus sp and Proteus sp., in the microbiological exam. The extender for semen, with Tris, is a potent inducer of uterine inflammation, and positive uterine cultures may be obtained during estrus without inflammation or uterine infection.

Influência da raça do touro (Bos indicus x Bos taurus) na tolerância ao estresse térmico calórico de embriões bovinos produzidos in vitro

To better understand the differences related to HS resistance between Bos indicus and Bos taurus, we aim to verify if the HS tolerance is due mostly to the genetic contribution from the oocyte, spermatozoa or both. Oocytes from Nelore and crossbreed Holstein cows (cHST) were collected, matured and fertilized with semen from Nelore (N), Angus (An), Brahman (Bra) and Gir (Gir) bulls. Nine six hours post insemination (hpi), ≥ 16 cells embryos were separated in two groups: control and HS. In control group, embryos were cultured at 39°C, whereas in the HS group, embryos were subjected to 41°C for 12 h, and then returned to 39°C. There was no effect of HS on blastocyst and hatched blastocyst rates in all breeds analyzed. The percentage of oocytes that cleaved and reached morula stage was significantly lower (p < 0.05) in cHST x Gir as compared to the other breeds. Additionally, blastocyst rates was higher in cHST x N than in cHST x An and cHST x Gir (p < 0.05). It was concluded that the oocyte is more important than the spermatozoa for the development of thermotolerance, since the breed of the bull did not influence embryo development after HS.

Análise espermática de perdizes (Rhynchotus rufescens) criadas em cativeiro e suplementadas com selênio

Due to the commercial importance of the red-winged tinamou (Rhynchotus rufescens), for the past few years, the employment of reproductive biotechnologies has been attempted. Thirty animals were randomly assigned into two groups: control group (no selenium) and treatment group (supplemented with 0,2 a 0,8 mg selenium/ 1000 kg ration). Animals were allocated at the FCAV - UNESP/Jaboticabal (2007-2008). Semen collections were performed by digital manipulation and divided in pools of at least 150 μL. After the immediate evaluation of motility, vigour, concentration and morphology, an aliquot of 20 μL was diluted in 300 μL of physiologic solution in order to test acrosome and membrane integrities, which were performed by counting 200 cells for each test. Cells were evaluated as follows: 1) Intact acrosome: lilac acrosome; Non-intact acrosome: pink acrosome; 2) Live cells: non stained; Dead: stained. Data was statistically analysed using the SAS System for Windows. No differences were found between treatment and control groups for volume, motility, vigour, mean number of spermatozoa per animal, concentration, Intact acrosome, Intact membrane. The difference found on midpiece sperm defect (Se = 1,33 ± 0,53 and control = 3,78 ± 0,69, p = 0.0107) may be due to the damages caused by the selenium deficiency to the architecture of the midpiece, which compromises sperm mobility and fertilization capacity.

Idade masculina: Qualidade do sêmen e fertilidade: Idade masculina X qualidade seminal

The man's age seems to affect reproductive function but to a lower degree compared with the woman's age. In fact, the effect of paternal age on semen quality has been discussed, but the results are contradictory. Considering the differences in the results of studies, this review aimed to analyze the literature regarding the effect of paternal age on semen parameters (volume, concentration, motility and morphology) in sperm DnA damage in assisted reproduction results and genetic diseases.

Congelação de espermatozoides epididimários de gatos utilizando o diluidor Botu-crio® após refrigeração por 24 h em contêiner de transporte de sêmen Botu-tainer®

The possibilities of using the sperm collected from the epididymis have been widely used because the fertilizing capacity sperm preservation and the possibility of using it for wild cats. But in the process of cryopreservation, some studies show a decrease in the quality of the sperm when left under cooling before frozen for some time. This study aimed to assess the quality of the epididymal sperm obtained from domestic cats after cryopreservation using a diluent based on egg yolk and glycerol (Botu-crio®), comparing the morphofunctional characteristics after cooling for 24 hours in a container of semen transport (Botu-tainer®). We use eight cats submitted to elective orchiectomy, aging from eight months, without racial determination, and good nutritional status. These sperm characteristics were: motility, vigor, concentration, membrane integrity and morphology. It has been found, after statistical analysis, that the container of semen was able to maintain sperm viability, even for 24h. We also observed a significant decrease on all parameters after frozen, consequential, probably to thermal stress that occurs in processing. However, the percentage of membrane integrity after thawing shows good employability of the Botu-crio®, which viability is possible to perform in vitro fertilization, requiring higher ratings.

Possible mechanism by which zinc protects the testicular function of rats exposed to cigarette smoke

Background: The aim of this study was to evaluate the changes in testicular function of rats due to cigarette smoke exposure and the possible mechanism by which zinc protects against these alterations. Methods: MaleWistar rats (60 days old) were randomly divided into 3 groups: control (G1, n = 10); exposed to cigarette smoke (G2, n = 10; 20 cigarettes/day/9 weeks) and exposed to cigarette smoke and supplemented with zinc (G3, n = 8; 20 cigarettes/day/9 weeks; 20 mg/kg zinc chloride daily for 9 weeks, by gavage). After the treatment period, the animals were euthanized, and materials were collected for analyses. Results: G2 rats showed a reduction in body mass; impaired sperm concentration, motility, morphology and vitality; and increased malonaldehyde and thiol group levels and superoxide dismutase activity as compared to G1. Zinc prevented the reduction of sperm concentration and the excessive increase of lipid peroxidation and induced an increase in plasma testosterone levels, wet weight of testis and thiol group concentration. Conclusions: Exposure to cigarette smoke led to harmful effects on testicular function at least partially due to the exacerbation of oxidative stress. Supplementary zinc had an important modulator/protector effect on certain parameters. The mechanism of zinc protection can be through an increase of SH concentration. Thus, zinc supplementation may be a promising addition to conventional treatments for male infertility related to smoking. Copyright © 2012 by Institute of Pharmacology Polish Academy of Sciences.

Improvement of Matrinxã, Brycon amazonicus, Larviculture by Exposing Eggs to Triiodothyronine

This work evaluated the effect of triiodothyronine (T3) on larviculture of matrinxã, Brycon amazonicus. Oocytes of three females were pooled, fertilized with pooled semen of two males and separated in four batches that were immersed in triiodothyronine solutions as follows: M1 (control - water); M2 (0.01mg/L T3); M3 (0.05mg/L T3); and M4 (0.1mg/L T3). Triiodothyronine did not affect fertilization rate and number of hatched larvae. Weight of hatched larvae was significantly higher in treatments M3 and M4, as well as among larvae sampled at Day 12 in all treatments. After 12d of rearing, biomass gain was higher in the hormone treatments (M1 688±569mg; M2 2436±562mg; M3 3572±569mg; and M4 4129±770mg). In general, coefficients of variation of weight (CVw) and length (CVl) did not differ among treatments and cannibalism was registered between 36 and 72hours post-hatching (h.p.h.) without differences among treatments. Larval survival increased in the hormone treatments (M1 26.5%; M2 37.6%; M3 40.6%; and M4 40.8%). The results indicate that the immersion of matrinxã eggs in triiodothyronine can promote beneficial effects to its larviculture and indicate promising perspectives for culture of this tropical species. © by the World Aquaculture Society 2013.

Brucella ovis REO 198 natural and experimental infection in Santa Inês rams from Brazil

B. ovis pathogenicity was evaluated in experimentally inoculated and naturally infected rams. Ten animals were submitted to simultaneous conjunctival and intrapreputial inoculation with 2x109 CFU/ mL of B. ovis REO 198. After that, animals underwent physical examination and blood samples were collected for serology every week. Positive serology results started to be observed in the 3rd week, with fluctuations in titers. Clinical changes began in the 5th week after inoculation and were associated with positive serology in the acute phase of the disease. Presence of B. ovis in semen and urine culture was intermittent. Three non-inoculated animals showed natural infection. B. ovis was shed twice in semen of one serology-negative animal. The study underscored the pathogenic characteristics of B. ovis REO 198 in Santa Inês rams, as well as the importance of animals as potential sources of infection.

Sexual transmission of Toxoplasma gondii in sheep

Male sheep of reproductive age were distributed into three groups: GI, a sheep inoculated (oral) with 2.0×105 oocysts of the P strain of Toxoplasma gondii; GII, a sheep infected (subcutaneous) with 1.0×106 tachyzoites of the RH strain of T. gondii; and GIII, a sheep kept as a control (not infected). After the inoculation of the males, 12 breeding ewes, which were not pregnant and which were serologically negative for reproductive diseases (particularly toxoplasmosis), were distributed into three groups, synchronized, and subsequently exposed to natural mating with previously inoculated males. The distribution was as follows: five ewes that underwent natural mating with the GI male, five ewes that were exposed to natural mating with the GII male, and two ewes that were mated with the non-infected male (control). Serum samples of all the ewes were collected on days -30, -14, -7, -1, and 0 (days before natural mating) and on days 1, 3, 5, 7, 11, 14, and weekly until birth; the presence of serum antibodies against T. gondii was assessed by IFAT. Using a bioassay and PCR, T. gondii was isolated from the semen of the infected reproducing sheep before mating. Following natural mating, 5 of the 12 females displayed antibodies specific for T. gondii; of these animals, two of the ewes underwent natural mating with the male inoculated with oocysts (GI) and three with the male infected with tachyzoites (GII). One of the females that displayed antibodies specific to this coccidian and that underwent natural mating with the GII sheep had a macerated fetus on the 70th day following coverage. Using a bioassay after the birth, it was possible to isolate T. gondii from samples of the pool of tissues from the five females that seroconverted after natural mating and from their respective lambs. Using PCR, the DNA of T. gondii was isolated from the pool of tissues from one and two females exposed to natural mating with the reproductive males infected with the oocysts and tachyzoites, respectively. Using this technique, it was also possible to diagnose the presence of the parasite in the pool of tissues from the lambs of one female that underwent natural mating with the male sheep infected with oocysts. These results demonstrated the sexual transmission of T. gondii in the sheep species with consequent vertical transmission to their lambs. © 2013 Elsevier B.V.

Evidence of sexual transmission of Toxoplasma gondii in goats

Male goats of reproductive age that were serologically negative for Toxoplasma gondii were selected and distributed according to the following arrangement: (A) one goat infected orally with 2.0 × 105 oocysts; (B) one goat infected subcutaneously with 1.0 × 106 tachyzoites; and (C) one uninfected goat kept as a control. After T. gondii inoculation, 12 non-pregnant female breeder goats that were serologically negative for the main reproductive diseases, especially toxoplasmosis, were synchronized and then exposed to natural mating by the males that had previously been inoculated: five females exposed to natural mating by male A (group GI); five females exposed to natural mating by male B (group GII); and two females exposed to natural mating by the uninfected male C (group GIII). In serum samples obtained from all the female goats before and after natural mating, the presence of antibodies against T. gondii was investigated using the ELISA test. PCR was performed on semen samples, on females and fetal tissues and placenta. Ten out of the 12 females showed specific antibodies against T. gondii after natural mating: five in GI and five in GII. On several dates on which natural mating occurred, T. gondii was identified in semen samples from the infected males, using PCR. Subsequently, after the females had been sacrificed, it was also possible to identify T. gondii in tissue samples from the infected females and from their fetuses, stillbirths and offspring, using PCR. Therefore, these results prove, for the first time, that T. gondii infection can be transmitted sexually from male to female goats. © 2013 Elsevier B.V. All rights reserved.

The relationships between scrotal surface temperature, age and sperm quality in stallions

In horses, spermatogenesis normally occurs at an average intratesticular temperature of 35. °C; therefore, mechanisms for testicular thermoregulation are essential. Measuring the scrotal surface temperature by thermography is one of the methodologies used to evaluate the effectiveness of testicular thermoregulation. The objective of this study was to determine the relationship between the control of scrotal surface temperature and sperm quality in horses of different ages. In total, 24 Quarter Horse stallions were divided into three groups: YS (young stallions), AS (adult stallions) and OS (old stallions). Initially, we calculated the testicular volume (TV) and evaluated various aspects of the semen (sperm kinetics, plasma membrane integrity and sperm morphology) for all the animals. We also evaluated rectal temperature (RT), body surface temperature (BST,) and average scrotal surface temperature in the testicular region (SST) before (M0) and after sun exposure (M1). Differences were observed (p<0.05) between the RT and BST before and after sun exposure in all three groups. However, there were no differences (p>0.05) in the SST values at these two time points, thus demonstrating the efficiency of the mechanisms for testicular thermoregulation. The SST was similar (p>0.05) among all three groups. Based on these results, we conclude that fertile stallions of different age groups are able to maintain SST and measuring the heat radiating from the scrotum using a digital infrared thermographer. We can also conclude that measuring the heat radiating from the scrotum using a digital infrared thermographer is a practical and efficient tool for monitoring SST in horses. © 2013 Elsevier B.V.

Involvement of pituitary gonadotropins, gonadal steroids and breeding season in sex change of protogynous dusky grouper, Epinephelus marginatus (Teleostei: Serranidae), induced by a non-steroidal aromatase inhibitor

Two experiments were performed using the aromatase inhibitor (AI) letrozole (100mg/kg) to promote sex change, from female-to-male, in protogynous dusky grouper. One experiment was performed during the breeding season (spring) and the other at the end of the breeding season (summer). During the spring, AI promoted sex change after 9weeks and the sperm produced was able to fertilize grouper oocytes. During the summer, the sex change was incomplete; intersex individuals were present and sperm was not released by any of the animals. Sex changed gonads had a lamellar architecture; cysts of spermatocytes and spermatozoa in the lumen of the germinal compartment. In the spring, after 4weeks, 11ketotestosterone (11KT) levels were higher in the AI than in control fish, and after 9weeks, coincident with semen release, testosterone levels increased in the AI group, while 11KT returned to the initial levels. Estradiol (E2) levels remained unchanged during the experimental period. Instead of decreasing throughout the period, as in control group, 17 α-OH progesterone levels did not change in the AI-treated fish, resulting in higher values after 9weeks when compared with control fish. fshβ and lhβ gene expression in the AI animals were lower compared with control fish after 9weeks. The use of AI was effective to obtain functional males during the breeding season. The increase in androgens, modulated by gonadotropins, triggered the sex change, enabling the development of male germ cells, whereas a decrease in E2 levels was not required to change sex in dusky grouper. © 2013 Elsevier Inc.

Sperm motility parameters for Steindachneridion parahybae based on open-source software

Summary: The objective of this work was to evaluate the sperm motility of 13 Steindachneridion parahybae males using open-source software (ImageJ/CASA plugin). The sperm activation procedure and image capture were initiated after semen collection. Four experimental phases were defined from the videos captured of each male as follows: (i) standardization of a dialogue box generated by the CASA plugin within ImageJ; (ii) frame numbers used to perform the analysis; (iii) post-activation motility between 10 and 20 s with analysis at each 1 s; and (iv) post-activation motility between 10 and 50 s with analysis at each 10 s. The settings used in the CASA dialogue box were satisfactory, and the results were consistent. These analyses should be performed using 50 frames immediately after sperm activation because spermatozoa quickly lose their vigor. At 10 s post-activation, 89.1% motile sperm was observed with 107.2 μm s-1 curvilinear velocity, 83.6 μm s-1 average path velocity, 77.1 μm s-1 straight line velocity; 91.6% were of straightness and 77.1% of wobble. The CASA plugin within ImageJ can be applied in sperm analysis of the study species by using the established settings. © 2013 Blackwell Verlag GmbH.