Influence of sperm post-thaw subpopulations of angus and nelore bulls on pregnancy rates by fixed-time artificial insemination.

BACKGROUND: The heterogeneity of ejaculate indicates that fertility is still variable among bulls and that more stringent evaluation methods are needed to identify the ejaculates suitable for AI. OBJECTIVE: To identify and characterize the sperm subpopulations (SP) in thawed semen doses of Nelore and Angus bulls and to evaluate the influence of these sperm subpopulations on pregnancy rate in cows submitted to fixed-time AI (FTAI). MATERIALS AND METHODS: A dose of post-thawed semen from each bull (n=18; consisting of Angus n = 9 and Nelore n = 9) was analyzed for: sperm kinetics; morphology and plasma membrane integrity; and the determination of the sperm subpopulations. Differences between the groups were estimated with the t-test considering a significance level of <5%. RESULTS: There was no influence between breeding bulls for sperm morphology, plasma membrane integrity, and pregnancy rate (P > 0.05). Regarding the kinetic parameters evaluated by the CASA system, Nelore had greater values, for cells with slow velocity (Angus: 16.4 %; Nelore: 21.7%; P = 0.028). In contrast, ANGUS bulls had more static cells (Angus: 27.2%; Nelore: 9.3%; P = 0.048). Based on CASA system data and clustering procedures, four sperm subpopulations were statistically established. In Angus bulls, a higher level of fast and nonlinear spermatozoa were found in SP3 (33.3%), followed by SP1 (32.7%%) with fast and progressive spermatozoa. Whereas, SP1 of Nelore bulls had 33.8% fast and progressive spermatozoa, followed by 32.2% of SP3 with fast and nonlinear spermatozoa. CONCLUSION: Both breeds of bulls presented similar proportions of sperm SP. Consequently, no influence on the pregnancy rates was shown in cows submitted to the IATF programs on a large scale. doi.org/10.54680/fr22310110312.

Evaluation of the Freezability of the Bovine Epididymis Tail Sperm with the Addition of Antioxidants.

BACKGROUND: The cryopreservation and recovery of epididymis tail sperm is an important biotechnology dependent on the composition of the freezing medium. OBJETIVE: To evaluate the effect of melatonin, added to commercial freezing medium extender, on the kinetics and viability of bovine epididymis tail sperm. MATERIAL AND METHODS: Five routines were performed, each consisting of eight epididymis and the structures were sliced onto a glass plate containing a commercial diluting medium for Botubov. The samples were divided into four groups, with 80 x 106 spermatozoa per mL. Group 1: samples diluted in Botubov. Group 2: samples centrifuged (600 g, 10 min), and the pellet re-suspended in Botubov. Group 3, samples diluted in Botubov containing 100 pM melatonin. Group 4: samples centrifuged (600 g, 10 min) and the pellet resuspended in Botubov with 100 pM melatonin. The samples were transferred to 0.5 mL straws at 40 x 106 viable spermatozoa, stabilized at 5º C for 4 h, transferred to liquid nitrogen vapour for 20 min, dipped in liquid nitrogen and stored in a cryogenic cylinder. After thawing (46ºC, 15s), sperm kinetics and viability parameters were evaluated. RESULTS: There was no difference in the parameters of total motility (MT, %), progressive motility (MP, %), progressive linear velocity (VSL, µm/s), curvilinear velocity (VCL, µm/s), linearity (LIN, %), spermatozoa with rapid movement (RAP, %) and level of intact plasma membranes and acrosome (IPMA, %) among the groups studied. However, a difference was observed between the routines performed. CONCLUSION: The protocol for freezing bovine epididymis tail sperm is applicable; however, there is an influence of the epididymis used, for the best efficacy of this biotechnology.

Use of infrared thermography to evaluate the inflammatory reaction in cat testis after intratesticular injection of 0. 9% NaCl or 20% CaCl2 with 1% lidocaine / O uso da termografia infravermelha para avaliar a reação inflamatória após a injeção intratesticular de NaCL 0, 9 % e cloreto de cálcio 20% com lidocaína 1%

Due to the doubts and questions about the inflammatory reaction caused by chemical castration, this study aimed to use infrared thermography to detect, evaluate and monitor the inflammatory reaction caused by the intratesticular injection of calcium chloride (CaCl2) 20% with lidocaine 1%. For this, thermographic measurements were taken before (M0), 10 minutes (M1), 1 and 6 hours (M2 and M3), for 7 consecutive days (M4 to M10), at 15 (M11), 30 (M12) and 60 (M13) days after intratesticular injection. Additionally, changes to testicular tissue and effects over spermatogenesis were evaluated by andrological exam before (M0) and 60 days (M13) after intratesticular injection. All cats were orchiectomized at M13, and testicles were submitted to histological analysis. CaCl2 (20%) with lidocaine (1%) administration produced testicular tissue damage and interfered with the spermatogenesis in 70% of treated cats without exacerbating the inflammatory reaction or impairing the cat's welfare. It was concluded that thermographic evaluation is a useful, efficient, easy and quick method to diagnose and monitor cat testicular inflammatory reactions.(AU)

Devido a constantes dúvidas e questionamentos sobre a reação inflamatória ocasionada pela castração química, este estudo objetivou o uso da termografia infravermelha para detectar, avaliar e monitorar a reação inflamatória causada pela injeção intratesticular de cloreto de cálcio (CaCl 2 ) 20% associada à lidocaína 1%. Para isso, medidas termográficas foram aferidas antes (M0), 10 minutos (M1), uma e seis horas (M2 e M3), por sete dias consecutivos (M4 a M10), aos 15 (M11), 30 (M12), e 60 (M13) dias após injeção intratesticular, nos grupos tratado e controle. Todos os gatos foram orquiectomizados no M13, e os testículos submetidos à análise histológica. A injeção CaCl 2 a 20% associada com lidocaína a 1% produziu lesão testicular e interferiu na espermatogênese de 70% dos gatos tratados, sem exacerbar a reação inflamatória ou prejudicar o bem-estar do animal. Foi concluído que a avaliação termográfica é uma ferramenta útil, eficiente, rápida e fácil para o diagnóstico e o monitoramento da reação inflamatória em gatos.(AU)

Oocyte quality and heat shock proteins in oocytes from bovine breeds adapted to the tropics under different conditions of environmental thermal stress.

In order to evaluate the influence of thermal stress on physiological parameters, and the oocyte quality of Girolando (n = 12) and adapted Pantaneira (n = 12) cattle, twelve sessions of ultrasound guided follicular aspiration (OPU) were performed, between January and November 2014 (during dry (May-September) and rainy season (October-April) in Brazil). The recovered cumulus-oocyte complexes (COCs) were selected and classified, according to quality, immediately after OPU. The oocytes were then stored in 3% paraformaldehyd before conducting immunofluorescence analysis under confocal microscopy to identify HSP70 and 90 proteins. Before each OPU session, the rectal temperature (RT) and respiratory frequency (RF) of each animal were measured. The black globe humidity index (BGHI) was calculated on the day of the OPUs and 90 days before each OPU session, and related to the thermal stress of the animals. The quality of oocytes from Girolando cattle, but not Pantaneira, showed a negative relationship with BGHI of 90 days before OPU. RT of both breeds did not exceed normal values for cattle below BGHI 95. BGHI variation on the day of OPU did not affect RF of the adapted Pantaneira breed (p = 0.3221). On the other hand, Girolando cattle showed a positive relationship between RF and BGHI (p = 0.0103). With increasing BGHI, the amount of HSP70 increased in Girolando oocytes, however, decreased in the Pantaneira breed. We have not observed a relationship between HSP 90 and BGHI, however Girolando cattle produced a greater amount of this protein in relation to the Pantaneira breed. In conclusion, higher BGHIs, 90 days before OPU session, negatively affect oocyte quality of Girolando cattle and positively affect oocyte quality of the Pantaneira breed. Higher BGHIs on the day of the OPU session negatively affected the respiratory frequency of the Girolando breed, and lead to a higher recruitment of HSP70 to protect oocyte maturation. The opposite pattern was observed for Pantaneira. In addition, Pantaneira cattle produced twice as much as HSP70 as Girolando cattle, suggesting that a natural higher production of this protein could be involved in the mechanisms of adaptation to heat conditions.

Aflatoxins ingestion and canine mammary tumors: There is an association?

The aim of this study was to determine the presence of mycotoxins on dogs feed and to explore the potential association between mycotoxins exposure and the chance of mamary tumors in a case-control study. The study included 256 female dogs from a hospital population, 85 with mammary tumors (case group) and 171 without mammary tumors (control group). An epidemiological questionnaire was applied to both groups, and the data were analyzed by the EpiInfo statistical package. For the study, 168 samples of the feed offered to dogs were analyzed for the presence of aflatoxins, fumonisins and zearalenone by high-performance liquid chromatography. Mycotoxins were found in 79 samples (100%) in the case group and 87/89 (97.8%) in the control group. Mycotoxins were detected in all types of feed, regardless feed quality. Level of aflatoxin B1 (p = 0.0356, OR = 2.74, 95%, CI 1.13 to 6.60), aflatoxin G1 (AFG1) (p = 0.00007, OR = 4.60, 95%, CI = 2.16 to 9.79), and aflatoxin G2 (AFG2) (p = 0.0133, OR = 9.91, 95%, CI 1.21 to 81.15) were statistically higher in case of mammary cancer. In contrast, neutering was a protective factor for mammary cancer (p = 0.0004, OR = 0.32, 95%, CI = 0.17 to 0.60).

Population estimate of the preantral follicles and frequency of multioocyte follicles in prepubertal and adult bitches.

Oocytes from preantral follicles could be an alternative for in vitro maturation because most follicles are at the preantral stage. There are few studies that have sought to estimate the number of preantral follicles in bitches. Therefore, the aims of this study were to estimate the population of preantral follicles in the ovaries of small- and medium-sized prepubertal and adult bitches and compare the population of preantral follicles between the right and left ovaries and evaluate the frequency of multioocyte follicles (MOF). Eighty ovaries were collected by elective ovariohysterectomy from 40 healthy bitches. The bitches were divided into four groups: small-size prepubertal bitches (<10 kg, n = 20), medium-size prepubertal bitches (10-20 kg, n = 20), small-size adult bitches (<10 kg, n = 20), and medium-size adult bitches (10-20 kg, n = 20). Immediately after surgery, the ovaries were fixed in Bouin's solution and processed for histology. For each specimen, 70 histologic sections were cut and mounted on slides; then, the number of preantral follicles was estimated using a correction factor. The preantral follicles were classified according to the developmental stage. The data were analyzed using the Kruskal-Wallis test followed by Dunn's test for comparison between groups, and Fisher's exact test was used to evaluate the frequency of MOF (P ≤ 0.05). Considering the population of preantral follicles from the pair of ovaries, medium-size prepubertal bitches had the highest (P < 0.05) population of preantral follicles compared with the small and medium-size adult groups. There was a large variation in the numbers of preantral follicles among individuals of the same weight and within each group. There were differences between medium-size prepubertal and adult bitches regarding the population of preantral follicles in the right ovaries (145,482 ± 110,712 vs. 49,500 ± 44,821; P = 0.02); however, no differences were observed between the groups on the basis of comparisons of the number of preantral follicles in the left ovaries (P > 0.05). The prevalence of primordial MOF was higher in prepubertal bitches (47% vs. 28%), whereas adult bitches had a higher frequency of secondary MOF (49% vs. 25%; P < 0.05). We conclude that medium-size prepubertal bitches had the highest population of preantral follicles compared with small and medium-size adult bitches, and the use of only one ovary per bitch implied contrasting result. The presence of primordial MOF was higher in prepubertal bitches and at the secondary stage in adult bitches.

Prognostic markers of canine pyometra / Marcadores prognósticos na piometra canina

The pyometra is a disease that affects middle age and elderly female dogs during diestrus. Hormonal, microbiological, biochemical and hematological aspects are well described. However, few studies have evaluated the role of each in the prognosis of canine pyometra. The aim of this study was to identify markers associated with clinical worsening of dogs with pyometra. We prospectively evaluated 80 dogs with pyometra treated surgically. Group 1 consisted of dogs that were discharged within 48 hours after surgery and Group 2 consisted of those who required prolonged hospitalization or died. The findings of hematological, biochemical and blood lactate levels were compared between groups and variables such as bacterial multidrug resistance, systemic inflammatory response syndrome (SIRS), hyperlactatemia and increased creatinine were analyzed through the dispersion of frequencies between groups. Among the variables studied, the presence of SIRS and elevated serum creatinine >2.5mg/mL were effective in predicting the worsening of the disease and can be used as prognostic markers of canine pyometra...

A piometra é doença que acomete as fêmeas da espécie canina de meia-idade a idosa durante o diestro. Aspectos hormonais, microbiológicos, hematológicos e bioquímicos são bem descritos. Entretanto, são poucos os estudos que avaliam o papel de cada um desses no prognóstico da piometra canina. O objetivo deste estudo foi identificar marcadores associados ao agravamento clínico de cadelas com piometra. Foram avaliadas prospectivamente 80 cadelas com piometra tratadas cirurgicamente. O grupo 1 foi composto por cadelas que receberam alta até 48 horas de pós-operatório, e o grupo 2 por aquelas que necessitaram de internamento prolongado ou morreram. Os achados hematológicos, bioquímicos e os níveis do lactato sanguíneo foram comparados entre os grupos, e variáveis como multirresistência bacteriana, síndrome da resposta inflamatória sistêmica (SIRS), hiperlactatemia e aumento da creatinina foram analisadas por meio da dispersão de frequências entre os grupos. Entre as variáveis estudadas, a presença de SIRS e a elevação da creatinina sérica >2,5mg/mL foram eficazes em predizer o agravamento da doença e podem ser utilizadas como marcadores prognósticos da piometra canina...

Osteopontin in seminal plasma and sperm membrane of dogs.

The objective of this work was to describe the presence of osteopontin (OPN) in canine seminal plasma and sperm membranes. A pool of seminal plasma and sperm membrane extract from 30 dogs was used. Polyacrylamide electrophoresis gels were performed and the bands were transferred to nitrocellulose paper and Western blot was undertaken using an antibody anti-OPN. Two and 12 bands were marked in the seminal plasma (77.2 and 15.6 kDa) and sperm membrane extracts (70.6-26.6 kDa), respectively. However, from 12 marked bands in the sperm membrane extract, only three (46.4, 37.7 and 36.5 kDa) were strongly marked. We conclude that, seminal plasma and sperm membranes from dogs contain different isoforms of OPN; yet, further studies will be necessary to determine their function in this species.

Fertilizing capacity of frozen epididymal sperm collected from dogs.

The collection of epididymal sperm may be a valuable tool for canine reproduction especially since it can enable collection of cells after death of a valuable dog. The aim of the present study was to evaluate the viability of epididymal sperm after freeze-thawing. Epididymides were obtained from four adult dogs by elective orchiectomy. The caudal portion of the epididymides and part of the deferential ducts were squeezed by means of an anatomic clamp into a Petri dish containing either 0.9% saline solution (Group 1) or Ringer solution without lactate (Group 2). Samples were centrifuged at 800 x g for 10 min, the supernatant was removed and the pellet was diluted in one step with a Tris/citric acid/OEP (Orvus Es Paste) extender containing 7% glycerol and subjected to semen freezing. Oocytes were obtained from canine ovaries, after ovariohysterectomy. Only oocytes that were approximately 100 mum in diameter, with a dark ooplasm surrounded by three- or four-well formed cumulus cell layers were used for sperm testing. Frozen semen samples were thawed in a water bath at 70 degrees C for 8 s and analysed at room temperature for sperm motility and velocity. Oocytes were incubated with spermatozoa in humidified atmosphere containing 5% CO(2) at 38 degrees C for 18 h. Morphological and functional characteristics of spermatozoa were similar in both groups. However, the percentage of sperm cells bound to oocytes was significantly higher in Group 2 than in Group 1. This result suggests that the Ringer solution without lactate was a more suitable medium for collecting epididymal canine sperm than 0.9% saline.

Cryopreservation effects on domestic cat epididymal versus electroejaculated spermatozoa.

Frozen-thawed epididymal spermatozoa have already been successfully used in artificial insemination in the domestic cat, proving to be a valuable resource for the reproduction of felid species, which are threatened with extinction. The aim of this study was to compare the effects of freezing and thawing on domestic cat semen collected by electroejaculation (EL) and from the epididymides (EP) and vasa deferentia. Ten adult cats were anesthetized, electroejaculated and immediately thereafter, orchiectomized. Epididymal spermatozoa were collected through the compression of caudae epididymidis and vasa deferentia. Spermatozoa were frozen-thawed following a single protocol. Sperm motility, sperm progressive status (0-5), plasma membrane integrity and morphology (light and transmission electron microscope) were assessed on two occasions, immediately after collection and after freezing and thawing. There were no significant differences between the electroejaculated and epididymal fresh or frozen-thawed spermatozoa for any of the variables. However, the incidence of acrosome defects after freezing and thawing increased by 19% based on light microscopy, whereas ultrastructural images revealed acrosome damages in most sperm cells. Since these acrosomal changes are known to affect sperm fertilising capacity, further studies are needed to optimize cryopreservation techniques for epididymal as well as electroejaculated domestic cat spermatozoa.

Alteraçöes produzidas a nível de testículo e sêmen de cäes submetidos à administraçäo de sulfato de vincristina / Resulting alterations at the level of testes and semen in dogs submitted to vincristine sulphate administration

Foram estudados 14 animais da espécie canina, sem raça definida, machos, adultos, com o objetivo de analisar, laboratorialmente, o sêmen e os aspectos histológicos dos testículos antes e após a administraçäo do sulfato de vincristina. Para tanto, os animais foram divididos em 2 grupos. No grupo I (8 animais), submeteu-se os animais à orquiectomia unilateral e um fragmento do testículo e epidídimo foram analisados histologicamente. No grupo II (6 animais), inicialmente, condicionou-se os animais para colheita de sêmen. Após obtençäo do material fecundante, este foi analisado laboratorialmente. No grupo I, todos os animais apresentaram degeneraçäo testicular em grau leve ou moderada. No grupo II, observaram-se alteraçöes espermáticas, principamente ao nível de peça intermediária e cauda. Entretanto, considerando que em termos de média, as características físicas e morfológicas do sêmen de cäes sofreram alteraçöes que säo reversíveis à utilizaçäo do sulfato de vincristina, como recurso terapêutico, pode ser instituído nesta espécie animal