Diferentes concentrações antibióticas adicionadas ao diluente de sêmen suíno / Different antibiotic concentrations added to the swine semen extender

Devido a contaminação bacteriana, utiliza-se antibióticos nos diluentes de sêmen para inibir o crescimento bacteriano durante a sua estocagem. Assim, este trabalho teve por objetivo determinar o limite de toxicidade de diferentes concentrações antibióticas da penicilina/estreptomicina adicionadas ao meio diluente do sêmen e investigar a influência sobre a motilidade espermática. Para isso, o sêmen de cinco reprodutores híbridos comerciais foi coletado uma vez por semana, durante nove semanas, através da técnica da mão enluvada. Cada ejaculado foi distribuído de forma igual entre todos os tratamentos, que consistiram em diferentes concentrações de solução de antibióticos, sendo: T1 = 0,0g/L (controle); T2 = 4,2g/L; e T3 = 12,6g/L. O sêmen diluído em água de coco em pó (ACP-103@) foi conservado por cinco dias e analisado nos dias: D0 (dia da coleta), D2 e D4 (último dia de conservação) quanto ao vigor e à motilidade espermática. Em D0, o sêmen conservado em diluente acrescido de 12,6g de solução antibiótica apresentou menor valor de vigor espermático em relação aos demais tratamentos (p<0,05). Já em D2 e D4, os tratamentos contendo antibióticos apresentaram resultados de vigor aquém dos obtidos no grupo controle (p<0,05). Em relação à motilidade espermática, os maiores percentuais de células móveis foram observados nas amostras de sêmen conservadas em ACP 0,0g/L (controle) quando comparados aos demais tratamentos (p<0,05). Conclui-se que as concentrações antibióticas utilizadas no estudo (4,2 e 12,6 g/L) mostraram efeitos tóxicos logo após a diluição do sêmen suíno, afetando de forma negativa parâmetros seminais durante a conservação a 17 °C.

Due to bacterial contamination, antibiotics are used in semen extenders to inhibit bacterial growth during storage. Thus, the present study aimed to determine the toxicity limit of different antibiotic concentrations of penicillin/streptomycin added to the semen extender and investigate the influence on sperm motility. For this purpose, semen from five commercial hybrid breeders was collected once a week for 9 weeks, using the gloved hand technique. Each ejaculate was distributed equally among all treatments, which consisted of different concentrations of antibiotic solution: T1 = 0.0g/L (control); T2 = 4.2g/L; T3 = 12.6g/L. The semen diluted in powdered coconut water (ACP-103@) was kept for 5 days and analyzed on the following days: D0 (collection day), D2, and D4 (last conservation day) for sperm vigor and motility. In D0, the semen preserved in the diluent added with 12.6 g of antibiotic solution showed a lower sperm vigor value compared to the other treatments (p<0.05). In D2 and D4, treatments containing antibiotics presented vigor results below those obtained in the control group (p<0.05). Regarding sperm motility, higher percentages of mobile cells were observed in semen samples conserved in ACP 0.0 g/L (control) when compared to the other treatments (p<0.05). It is concluded that the antibiotic concentrations used in the study (4.2 and 12.6 g/L) showed toxic effects soon after the swine semen, negatively affecting seminal parameters during storage at 17 °C.

Sêmen refrigerado ­ estado da arte em diferentes espécies / Chilled semen ­ state of the art in different species

Na atual conjuntura da criação artificial de bovinos e bubalinos, o material genético masculino de qualidade superior de reprodutores é explorado ao máximo possível através da inseminação artificial em tempo fixo de um grande número de fêmeas com apenas um único ejaculado. Para isso, é necessário um sêmen de boa qualidade que desempenhe um papel indispensável na melhoria das taxas de fertilidade, independente de qual tipo seja utilizado (fresco, refrigerado e congelado). Porém, o processo de congelação/descongelação causa uma série de injúrias aos espermatozoides, ocasionando resultados inferiores para percentuais de viabilidade espermática, motilidade, membrana plasmática e integridade acrossomal, potencial de membrana mitocondrial, cinemática do esperma, quando comparado ao sêmen refrigerado. Assim, o objetivo desta revisão é disseminar o conhecimento sobre o uso sêmen refrigerado na preservação de germoplasma de reprodutores bovinos e bubalinos para melhorar as taxas de concepção em propriedades. Para isso, serão abordados comentários sobre o armazenamento do sêmen refrigerado, com ênfase nas diferenças entre curvas de refrigeração, suas vantagens e desvantagens relativas para procedimentos de uso na IATF, identificando o método mais indicado por diversos autores, o estado atual da biotécnica, seus méritos e possibilidades futuras.(AU)

In the current conjuncture of the artificial creation of bovines and buffaloes, the male genetic material of superior quality of sires is exploited to the maximum possible through the fixed-time artificial insemination of a large number of females with only a single ejaculate. For this, a good quality semen is needed that plays an indispensable role in improving fertility rates, regardless of which type is used (fresh, chilled and frozen). However, the freezing/thawing process causes a series of injuries to spermatozoa, causing lower results for percentages of sperm viability, motility, plasma membrane and acrosomal integrity, mitochondrial membrane potential, sperm kinematics, when compared to refrigerated semen. Thus, the objective of this review is to disseminate knowledge about the use of chilled semen in the preservation of germplasm of bovine and buffalo breeders to improve conception rates in properties. For this, comments on the storage of refrigerated semen will be addressed, with emphasis on the differences between refrigeration curves, their relative advantages and disadvantages for procedures for use in FTAI, identifying the method most indicated by several authors, the current state of biotechnics, its future merits and possibilities.(AU)

Nutracêuticos no desempenho reprodutivo de garanhões / Nutraceuticals on the stallion breeding performance

O termo nutracêutico vem se tornando comum na reprodução animal e descreve produtos derivados dos alimentos, que podem fornecer benefícios adicionais a saúde, além dos valores básicos encontrados na dieta. Entre os vários nutracêuticos utilizados na reprodução, temos o ômega-3, ômega-6, vitaminas do complexo B, L-carnitina, ß-caroteno e antioxidantes. Portanto, objetivou-se apresentar as principais substâncias utilizadas como nutracêuticos e antioxidantes, seja utilizado de forma oral ou incorporado a diluentes de sêmen, com a finalidade de melhorar a qualidade seminal de garanhões, e consequentemente incrementar taxas de prenhez.(AU)

Nutraceuticals have become remarkably popular in animal reproduction and describe products derived from foods, which may provide additional health benefits, beyond the basic value found in diets. Among the main nutraceuticals used in male reproduction, there are omega-3 and 6, B-complex vitamins, L-carnitine, ß-carotene, and antioxidants. The aim of this review is to present the main substances used as nutraceuticals and antioxidants, either used orally or in combination with semen extenders, focusing on the improvement of seminal quality, and pregnancy rates.(AU)

Addition of orange, pineapple and beet juices as extenders for cryopreservation of ram semen / Adição de sucos de laranja, abacaxi e beterraba em diluidor para criopreservação de sêmen de carneiros

Searching for improvements in semen cryopreservation, natural substances are commonly studied focusing to improve the sperm quality. The aim of this study were evaluated the effect of adding orange, pineapple, and beet juices in different concentrations and combinations to the ram semen cryopreservation extender. Five ejaculates from five adult rams were used. The semen pool was diluted in egg yolk-based extender and mixed with the following 15 treatments (at a final concentration of 400.106 sptz/mL): orange 10% (O10) and 15% (O15); pineapple 10% (P10) and 15% (P15); beet 10% (B10) and 15% (B15); pineapple + orange 10% (PO10) and 15% (PO15); pineapple + beet 10% (PB10) and 15% (PB15); beet + orange 10% (BO10) and 15% (BO15); pineapple + beet + orange 10% (PBO10) and 15% (PBO15); and the control group (CON). Post-thaw in 0.25 mL straws semen quality analysis of cryopreserved semen was performed by CASA and flow cytometry. Analysis of variance (PROC GLM) was carried out and the averages were compared using the SNK test. Pearson's correlation test was also performed. No effect was noted in the addition of juices to the semen extender prior to cryopreservation. Post-thawed, although, statistically similar to the control group, the total motility of the B10 group reached acceptable standards of total motility. In addition, B10 group showed the highest values (p< 0.05) of progressive motility than control group or other treatments. The addition of 10% beet juice to the ram semen extender can improve the cryopreservation of sperm motility.

Em busca de melhorias na criopreservação do sêmen, substâncias naturais são comumente estudadas com o objetivo de melhorar a qualidade do sêmen. O objetivo deste estudo foi avaliar o efeito da adição de sucos de laranja, abacaxi e beterraba em diferentes concentrações e combinações ao diluidor de criopreservação de sêmen ovino. Foram utilizados cinco ejaculados de cinco carneiros adultos. O pool de sêmen foi diluído em diluente à base de gema de ovo e misturado com os seguintes 15 tratamentos (na concentração final de 400x106 sptz/ml): laranja 10% (O10) e 15% (O15); abacaxi 10% (P10) e 15% (P15); beterraba 10% (B10) e 15% (B15); abacaxi + laranja 10% (PO10) e 15% (PO15); abacaxi + beterraba 10% (PB10) e 15% (PB15); beterraba + laranja 10% (BO10) e 15% (BO15); abacaxi + beterraba + laranja 10% (PBO10) e 15% (PBO15); e o grupo controle (CON). Pós-descongelação em palhetas de 0,25 ml a análise da qualidade do sêmen criopreservado foi realizada pelo CASA e citometria de fluxo. A análise de variância foi realizada e as médias comparadas pelo teste SNK. O teste de correlação de Pearson também foi realizado. Nenhum efeito foi observado na adição de sucos ao diluidor de sêmen antes da criopreservação. Após o descongelamento, embora estatisticamente semelhante ao grupo controle, a motilidade total do grupo B10 atingiu padrões aceitáveis de motilidade total. Além disso, o grupo B10 apresentou os maiores valores (p<0,05) de motilidade progressiva que o grupo controle ou os outros tratamentos. A adição de 10% de suco de beterraba ao diluente de sêmen ovino pode melhorar a criopreservação da motilidade espermática.

Parâmetros espermáticos do sêmen suíno conservado a 10 e 17o C, em diluentes comerciais e alternativos / Spermatic parameters of swine semen conserved at 10 and 17o C, in commercial and alternative extenders

O uso de diluentes adequados é importante para o sucesso da inseminação artificial. O objetivo deste estudo foi avaliar a eficiência de diluentes em diferentes temperaturas. Foram utilizados os seguintes diluentes: Beltsville Thwaing Solution (BTS); TRIS (Tris-hidroximetil-aminometano - H2NC(CH2OH)3); TRIS + gema de ovo (TGO) e Leite em pó desnatado (LPD). Cinquenta ejaculados foram analisados quanto ao vigor, à motilidade, à funcionalidade de membrana e à atividade mitocondrial. Os diluentes TGO e LPD mantiveram os maiores valores para vigor e motilidade a 10 °C, em comparação aos demais tratamentos (p<0,05). A 17 °C, esses diluentes apresentaram resultados melhores de vigor e motilidade em relação aos demais, somente até D1 (p<0,05). A partir de D2, o BTS mostrou-se como o melhor diluente para a preservação dessas características. Maior percentual de espermatozoides com membrana funcional foi observado no TGO a 10 °C, seguido do LPD, BTS e TRIS (p<0,05). A 17 °C, o BTS manteve essa característica, exceto em D0, em que o TGO e o LPD apresentaram melhores resultados. Quanto a atividade mitocondrial, maiores valores foram observados a 10 °C no TGO em todos os dias de análise e em D0 à temperatura de 17 °C. O diluente TRIS + gema de ovo mostrou-se como uma alternativa para conservação do sêmen suíno a 10 °C, assim como o LPD. Além disso, esses mesmos diluentes constituem alternativas para conservação a curto prazo a 17 °C.

The use of suitable extenders is important to artificial insemination success. Therefore, the objective of this study was to evaluate the efficiency of extenders at different temperatures. The following extenders were used: Beltsville Thawing Solution (BTS); TRIS (Tris-hidroximetil-aminometano - H2NC(CH2OH)3); TRIS + egg yolk (TGO), and skim powdered milk (SPM). Fifty ejaculates were analyzed regarding sperm vigor, motility, membrane functionality, and mitochondrial activity. The TGO and SPM extenders maintained the highest values for vigor and motility at 10 °C, in comparison to the other treatments (p<0.05). At 17 °C, these extenders showed better results of vigor and motility compared to the others, only until D1 (p<0.05). From D2, BTS proved to be the best extender to preserve these characteristics. A higher percentage of sperm with a functional membrane was observed for TGO at 10 °C, followed by SPM, BTS, and TRIS (p<0.05). At 17 °C, BTS maintained this characteristic, except for DO, which showed better results for TGO and SPM. Regarding the mitochondrial activity, higher values were observed at 10 °C for TGO on all days of analysis and D0 at a temperature of 17 °C. The extender TRIS + egg yolk was shown as an alternative for the conservation of swine semen at 10 °C, as well as the SPM. In addition, these same extenders are alternatives for short-term conservation at 17 °C.

Sperm quality and morphometry characterization of cryopreserved canine sperm in ACP-106c or TRIS

Morphological sperm evaluation supported by the morphometry can be used in the determination of the seminal quality and in the investigation of potential extenders. Although there are studies comparing TRIS and ACP extenders, there are no comparative studies between them for the computerized assisted semen analysis (CASA), sperm viability, membrane functionality and sperm morphometry parameters of cryopreserved canine semen. Hence, we aimed to evaluate the effects of ACP-106c and TRIS on post-freezing canine sperm quality. Five dogs were submitted to semen collection twice with one-week interval. The semen was evaluated within the parameters: total motility, vigor, concentration, viability, plasma membrane functionality, morphology and morphometry. In the morphometric evaluation, the morphologically normal sperm was measured as: length, width, area and perimeter of the head and the midpiece, tail length and total length. The parameters of ellipticity, elongation, regularity and roughness were determined. Then, the semen was divided into two aliquots that were diluted in TRIS or ACP-106c, with the addition of egg yolk and glycerol. The diluted semen was refrigerated and frozen. The thawed samples were evaluated. Total motility, viability, sperm membrane functionality and normal morphology reduced after thawing in both extenders (morphology reduced from 89.60 ± 1.3% to 84.40 ± 1.8 and 84.60 ± 1.1% in TRIS and ACP-106c, respectively). However, it did not differ between TRIS and ACP-106c. In the ACP106c the sperm head defects in cryopreserved semen were higher compared to fresh semen (P < 0.05). For all the morphometric parameters evaluated, there were no differences between fresh and cryopreserved samples (3.70 ± 0.4% vs. 2.30 ± 0.5%). In kinetics, with an interval of one week statistical differences between the extenders were found only in the parameters ALH and LIN (P < 0.05). Regardless of the extender, there were no changes in the morphometric parameters of sperm after thawing.(AU)

Heterologous in vitro fertilization and embryo production for assessment of jaguar (Panthera onca Linnaeus, 1758) frozen-thawed semen in different extenders

Heterologous in vitro fertilization (IVF) is an important tool for assessing fertility of endangered mammals such as the jaguar, considering difficult access to females for artificial insemination and to obtain homologous oocytes. We aimed to evaluate the fertility of jaguar sperm cryopreserved with different extenders, using domestic cat oocytes to assess the development of hybrid embryos. Semen from four captive jaguars was obtained by electroejaculation. Samples were cryopreserved in powdered coconut water (ACP-117c) or Tris extender containing 20% egg yolk and 6% glycerol. Thawed spermatozoa were resuspended (2.0 × 106 spermatozoa/mL) in IVF medium and co-incubated with cat oocytes matured in vitro for 18 h. Presumptive zygotes were cultured for 7 days. After 48 h, cleavage rate was evaluated, and non-cleaved structures were stained for IVF evaluation. On days 5 and 7, the rate of morula and blastocyst formation was assessed. Data were analyzed using the Fisher exact test (p < 0.05). No difference was observed between ACP-117c and Tris extenders, respectively, for oocytes with 2nd polar body (2/51, 3.9 ± 2.9% vs. 2/56, 3.6 ± 3.1%), pronuclear structures (5/51, 9.8 ± 4.7% vs. 8/56, 14.3 ± 8.0%), and total IVF rates (7/36, 19.4 ± 5.0% vs. 10/37, 27.0 ± 13.8%). All the samples fertilized the oocytes, with 22.9 ± 3.2% (16/70) and 16.7 ± 3.6% (12/72) cleavage of mature oocytes for ACP-117c and Tris extenders, respectively. Morula rates of 4.3 ± 2.3% (3/70) and 5.6 ± 2.2% (4/72) were observed for ACP-117c and Tris, respectively. Only the Tris extender demonstrated blastocyst production (2/12, 16.7 ± 1.5% blastocyst/cleavage). We demonstrated that jaguar ejaculates cryopreserved using ACP-117c and Tris were suitable for IVF techniques, with blastocyst production by ejaculates cryopreserved in Tris. This is a first report of embryos produced in vitro using jaguar sperm and domestic cat oocytes through IVF.(AU)

Sperm cell capacitation status of ram semen after cooling

Background: The use of conventional artificial insemination (AI) in sheep production is usually associated with lower fertility rates when frozen semen is used. Cooled ram semen has been an alternative over frozen semen due to the higher viability, seminal quality and fertility rates following AI. The semen preservation process promotes sperm cell modifications similar to capacitation (capacitation-like) that causes cell damage affecting viability and seminal quality, but such effects are unclear for cooled semen. The aim of this study was to determine the status of sperm cell capacitation (CA) and acrosome reaction (AR) during ram semen processing and cooling under different extenders, dilution factors, and aerobiosis conditions as a function of storage time at 5o C. Materials, Methods & Results: Two consecutive ejaculates per day per male were collected from 2 adult rams by artificial vagina at 48-72 h intervals, in three replications. After macro- and microscopic evaluations, semen was segregated into groups under 3 extenders (Tris-egg yolk or TY, citrate-egg yolk or CY, skimmed milk or SM), 2 dilution factors (1 x 109 or Bi, 100 x 106 or Mi cells/mL), and 2 aerobiosis conditions (aerobic or A, semi-anaerobic or SA). Diluted semen was cooled to 5ºC and stored for up to 72 h, with evaluations every 24 h. Aliquots of fresh ejaculates and of each cooled diluted subgroup, according to extender, dilution, and aerobiosis, were collected at times T0 and T72 for determination of acrosome status and membrane integrity by the chlortetracycline (CTC) and trypan blue-Giemsa stainings, respectively. No differences were detected in sperm cell motility (M) and motility vigor (V) between fresh and diluted semen. After cooling, a significant decrease in M was observed after 48 h in CY and SM compared with fresh semen and 0 h of cooling, while V started to decrease after 24 h in CY compared with TY. Likewise, M/V from different dilutions and aerobic conditions decreased more significantly after 48 and 24 h of cooling, respectively. The sperm capacitation status did not show differences in the proportion of non-capacitated (NCA), CA and AR sperm cells between TY, CY, and SM extenders (NCA: 75.0%, 71.3%, 74.0%; CA: 15.7%, 17.2%, 15.9%; AR: 9.3%, 11.5%, 10.2%) or between Bi and Mi dilutions (NCA: 74.0%, 72.9%; CA: 15.9%, 16.6%; AR: 10.1%, 10.5%), respectively. However, differences (P < 0.05) were observed between A and SA aerobic conditions, with CA (17.0% vs. 15.5%) and AR (11.9% vs. 8.7%) rates being higher in A than SA, respectively, with no differences in NCA (71.1% vs. 75.8%), irrespective of the storage time. Sperm cell viability decreased after 48 h, especially in CY (P < 0.05). Discussion: Ram sperm cells can suffer irreversible damage due to thermal shock during cooling. Egg yolk-based extenders provide phospholipids and cholesterol to protect the sperm cell membrane during the thermal shock caused by the change in temperature. In this study, sperm cells had irreversible decreases in M/V, with increase in acrosome and plasma membrane damage after cooling to 5ºC. The largest and smallest decreases in M and V over time were observed in the CY and TY extenders, respectively. In addition to the extender type, the semen preservation method and storage time promoted changes in the capacitation status, AR and in sperm cell viability, which per se were associated with a decrease in semen fertility. In fact, the proportions of CA and/or AR sperm cells gradually increased over time after dilution and storage at 5ºC, with a negative correlation between sperm cell viability and M/V over time. In summary, extender and cooling time affected mostly M/V, while aerobiosis condition and dilution factor were more associated with acrosome status and sperm survival, with the extender having less impact on the acrosome status as a function of time.

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.(AU)

Influência da genética e dos métodos de conservação sobre a qualidade do sêmen suíno / Influence of genetics and conservation methods on the quality of swine semen

A demanda do mercado impulsionou o melhoramento genético na suinocultura caracterizado pelo cruzamento entre linhagens e raças distintas a fim de possibilitar o aproveitamento da heterose. Esse melhoramento acelerado ocorreu em decorrência da inseminação artificial e manipulação do sêmen, associado ao emprego de diluentes e técnicas de refrigeração. Sendo assim, esta revisão tem como objetivo compilar o conhecimento acerca da influência genética e dos métodos de conservação sobre a qualidade do sêmen suíno, além de discutir a composição e eficiência dos principais diluentes e crioprotetores destinados à conservação espermática nesta espécie. Inicialmente, os animais eram selecionados baseados em características produtivas como habilidade materna, qualidade de carcaça e desempenho. Contudo, a fertilidade do reprodutor, caraterizada pela qualidade espermática e libido, é extremamente importante para indústria suinícola uma vez que determina, o potencial produtivo do plantel indiretamente. Evidências demonstram que os parâmetros espermáticos sofrem influência da genética, sendo que cada raça se destaca numa determinada característica seminal. Além disso, o manejo, idade, alimentação, sazonalidade, as características intrínsecas do espermatozoide e os métodos de conservação do ejaculado também determinam sua viabilidade. Apesar de preconizado devido à fácil execução e ótimos resultados, o sêmen refrigerado tem como limitador a produção excessiva de espécies reativas de oxigênio. Da mesma forma, o congelamento do ejaculado promove alterações espermáticas em virtude do choque térmico. Sendo assim, o emprego de diluentes e crioprotetores propícios que possibilitem a manutenção a longo prazo da viabilidade espermática é imprescindível para ambos os processos.

The market demand promoted the genetic improvement in swine farming characterized by the crossing between different strains and breeds to make possible the use of heterosis. This accelerated improvement occurred due to artificial insemination and semen manipulation, associated with the use of extenders and refrigeration techniques. Therefore, this review aims to compile knowledge about the genetic influence and conservation methods on the quality of swine semen, besides to discussing the composition and efficiency of the main extenders and cryoprotectants intended for sperm conservation in this species. Initially, animals were selected based on productive traits such as maternal ability, carcass quality, and performance. However, male fertility, characterized by sperm quality and libido, is extremely important for the swine industry as it determines the productive potential of the herd indirectly. Evidences demonstrates that sperm parameters are influenced by genetics, with each race standing out in certain seminal trait. In addition, the management, age, feeding, seasonality, the intrinsic characteristics of the sperm cell, and the methods of ejaculate conservation also determine its viability. Despite being recommended due to its easy execution and excellent results, refrigerated semen has the excessive production of reactive oxygen species as a limiter. Likewise, the ejaculate freezing promotes sperm changes due to heat shock. Therefore the use of suitable extenders and cryoprotectants that allow the long-term maintenance of sperm viability is essential for both processes.

Reproductive performance of single fixed-time inseminated sows with semen doses submitted to different semen storage times

This study aimed to evaluate the reproductive performance of sows submitted to single fixed-time insemination (SFTAI) using boars according to capacity for liquid in vitro semen preservation, type of extender, and storage time. Boars (n = 12) were classified into two groups based on progressive motility (PM) at 120 h of semen storage: low (PM - 64.5%) - and high-preservation (PM - 83.9%) capacity for semen storage. Weaned sows (n = 397, parity - 1 to 7) were inseminated (1.5×109 sperm cells) in a factorial design: two classes of boars (low- or high-preservation), two types of extenders (short- or long-term), and two semen storage times at insemination (24 or 72 h). An adapted triptorelin acetate protocol was used for SFTAI. Total sperm motility (TM) and PM at insemination were greater in high-preservation boars at 72 h compared with low-preservation boars at 24 or 72 h (P < 0.01). Short- or long-term extender did not affect (P ≥ 0.68) TM and PM in high-preservation boars; however, long-term extender improved these parameters in low-preservation boars (P < 0.01). Pregnancy and farrowing rates were not affected by groups (P > 0.05). Total piglets born (TPB) was reduced (P = 0.05) in low-preservation boars with 72 h of storage (13.6 ± 0.5) compared to high-preservation boars with semen stored for 24 or 72 h (15.2 ± 0.5 and 15.5 ± 0.5, respectively). The low-preservation boars reduced the TPB in sows submitted to SFTAI, and this reduction was greater using semen stored for 72 h.(AU)

Effect of Different Extenders on Sperm Motility and Vitality in Goose Semen Cryopreservation

This study aimed to investigate the usability of different diluents containing 6% Dimethylformamide (DMF) for cryo-preservation of the semen of geese (Anser cygnoides). The diluents of Glucose (G), Tris-Glucose (T), Lactated Ringer's-Glucose (LG), and Lactated Ringer's (L), all of which contained 6% DMF, were used as cryoprotectants. The researchers collected semen samples from four geese, twice a week over a four-week period, by means of abdominal massage; they then calculated how much sperm each goose ejaculated. Next, the semen samples were pooled and their spermatological parameters were determined. Their volume (4x (mL)), concentration (×108/mL), pH, motility (%), and vitality (%) rates were 0.31±0.01, 3.49±0.32, 7.13±1.06, 67.75±1.28, and 70.00±2.03, respectively. Then, these pooled semen samples were equally divided into four groups. Once they were frozen and thawed, the researchers discovered that the diluent L had the highest motility rate: 40.12% ± 1.35. The motility rates of the other diluents were as follows: LG (28.25%± 1.48), G (21.50% ± 1.41), and T (5.12% ± 0.83). Likewise, the vitality rates (%) of the diluents were as follows: L (41.93% ±1.87), LG (31.50%±1.88), G (29.43% ±1.45), and T (10.56%±1.34), respectively. Freezing and thawing appeared to lower each diluent's vitality and motility rates. However, for the Lactated Ringer's (L), this decrease was predictable. Therefore, Lactated Ringer's diluent containing 6% DMF can be used in cryo-preservation of goose semen.(AU)

Equine semen cooled with chemically defined extender containing soy lecithin / Sêmen equino refrigerado com diluidor quimicamente definido contendo lecitina de soja

Equine semen has historically been chilled using milk-based media. However, the use of animal-based components presents several potential concerns, such as variability in formulations, microbial contamination and regulatory issues. We aimed to evaluate the potential of including different concentrations of soy lecithin (LS) in chemically defined Biggers, Whitten and Whittingham (BWW) medium for cooling equine semen to 15°C. Ejaculates were diluted as six different experimental groups: 1) BotuSêmen® (control); 2) BWW; 3) BWW + 1% LS; 4) BWW + 2% LS; 5) BWW + 4% LS and 6) BWW + 6% LS. BWW medium, did not preserve motility, velocity, straightness (STR), linearity (LIN), amplitude of lateral sperm head displacement (ALH), cross flagellar beat frequency (BCF), functional and structural integrity of equine spermatozoa during 24 h of refrigeration when compared to BotuSêmen® (P <0.05). The use of BWW for cooling equine semen was only possible with the addition of LS, being the concentrations equal or higher than 2% better, because they preserved total motility, curvilinear velocity (VCL) and LIN with the same potential of BotuSêmen® (P >0.05). Nevertheless, BotuSêmen® showed superiority in preserving the percentage of sperm progressive motility, average path velocity (VAP), linear progressive velocity (VSL) and BCF during cooling compared to the other extenders (P <0.05). The inclusion of soy lecithin, from 2 to 6% in the BWW medium, allowed maintaining the viability of equine semen cooled at 15ºC for up to 24 hours.

O sêmen equino tem sido historicamente refrigerado usando meios à base de leite. No entanto, o uso de componentes de origem animal causa várias preocupações potenciais, como variabilidade nas formulações, contaminação microbiana e questões regulatórias. Objetivou-se avaliar o potencial de inclusão de diferentes concentrações de lecitina de soja (LS) no meio quimicamente definido BWW - Biggers, Whitten e Whittingham para refrigeração de sêmen equino e armazenamento na temperatura de 15°C. Os ejaculados foram diluídos em seis diferentes grupos experimentais: 1) BotuSêmen® (controle); 2) BWW; 3) BWW + 1% lecitina de soja (LS); 4) BWW + 2% LS; 5) BWW + 4% LS e 6) BWW + 6% LS. O meio BWW, não preservou a motilidade, a velocidade, a retilinearidade (STR), a linearidade (LIN), a amplitude do deslocamento lateral da cabeça (ALH), a frequência de batimento flagelar cruzado (BCF), a integridade funcional e estrutural dos espermatozoides equino durante 24 h de refrigeração quando comparado ao BotuSêmen® (P <0,05). O uso de BWW para refrigeração de sêmen equino só foi possível com adição de lecitina de soja, sendo as concentrações igual ou superior a 2% melhores, pois preservaram a motilidade total, a velocidade curvilinear (VCL) e LIN com mesmo potencial do BotuSêmen® (P >0,05). Ainda assim, o diluidor comercial BotuSêmen® apresentou superioridade em preservar o percentual de espermatozoides progressivamente móveis, a velocidade média da trajetória (VAP), a velocidade linear progressiva (VSL) e a frequência do batimento flagelar cruzado (BCF) durante a refrigeração comparado aos demais diluidores (P <0,05). A inclusão de lecitina de soja, de 2 a 6% no meio BWW, permitiu a manutenção da viabilidade do sêmen equino refrigerado a 15ºC por até 24 horas.

A preliminary study on the effects of E-Z Mixin® and EquiPlus® extenders supplemented with Edible Bird's Nest on the quality of chilled Arabian stallion semen

Abstract The aim of this study was to evaluate the effects of adding different concentrations of edible bird's nest (EBN) which is secreted by swiftlet birds (Aerodramus fuciphagus), into EquiPlus® and E-Z Mixin® extenders on the quality of chilled Arabian stallion semen at various storage times (0, 24 and 48 h). Ten ejaculates were collected from five stallions, and diluted using the two extenders containing 0% (control), 0.12%, 0.24% and 0.24% of EBN + seminal plasma (SP). All the diluted semen samples were then cooled and stored at 5 °C, and examined at 0, 24 and 48 h. Sperm kinetic parameters were assessed using computer assisted sperm analysis (CASA) and viability were assessed using Hoechst33342/PI stain. In both extenders, total motility (TM) and progressive motility (PM) were significantly higher at 0.12% and 0.24% compared to 0.24% + SP at 24 and 48 h. At 0.12%, E-Z mixin® treated semen had significantly higher TM and PM than EquiPlus® at 24 and 48 h. At 0.12% and 0.24%, average path velocity (VAP), straight-line velocity (VSL) and curvilinear velocity (VCL) were significantly higher in E-Z mixin® treated semen compared to EquiPlus® at 24 and 48 h. Comparisons between the two extender types at different concentrations of EBN showed no significant difference in lateral head amplitude (ALH), linearity (LIN), straightness (STR), beat cross frequency (BCF) and viability, irrespective of the storage time. The percentage of viable was significantly higher in E-Z mixin® than EquiPlus® at 0 and 48 h in control and 0.12%. Supplementation of the E-Z mixin® extender with 0.12% and 0.24% EBN concentrations in the absence of SP provided better CASA parameters such as TM, PM, VAP, VSL, and VCL at 24 and 48 h storage time. In conclusion, the results of this study indicated that chilled semen from Arabian stallion that was extended using E-Z mixin® and supplemented with 0.12% and 0.24% EBN concentrations performed better and yielded superior results in sperm kinetic parameters and % viable compared to EquiPlus® at 24 and 48 h storage time.

Influence of antibiotics on bacterial load and sperm parameters during short-term preservation of collared peccary semen

Abstract Studies on semen and sperm cells are critical to develop assisted reproductive technologies for the conservation of the collared peccary. The objective of the study was to compare the effect of different antibiotics on the bacterial load and sperm quality during short-term storage of peccary semen. Fresh semen samples from 10 males were extended in Tris-egg yolk or Tris-Aloe vera supplemented with streptomycin-penicillin (SP; 1 mg/mL - 1000 IU/mL or 2 mg/mL - 2000 IU/mL) or gentamicin (30 µg/mL or 70 µg/mL) before storage at 5°C. Bacterial load and sperm motility, membrane integrity and function, mitochondrial activity, and morphology, were evaluated at different time points for 36 h. The SP and gentamicin treatments concentration inhibited (p < 0.05) bacterial growth for 36 h regardless of the extender. Compared to the other treatments, Tris-egg yolk plus 70 µg/mL gentamicin maintained the sperm parameters for longer, including total motility (41.9 ± 6.1%) at 24 h, and membrane integrity (58.3 ± 2.1%) at 36 h. In contrast, the highest SP concentration in both extenders impaired sperm membrane integrity at 36 h (p < 0.05). For the liquid storage of collared peccary semen, it therefore is recommended to use Tris extender supplemented with egg yolk and gentamicin (70 µg/mL).

A preliminary study on the effects of E-Z Mixin® and EquiPlus® extenders supplemented with Edible Birds Nest on the quality of chilled Arabian stallion semen

The aim of this study was to evaluate the effects of adding different concentrations of edible birds nest (EBN) which is secreted by swiftlet birds (Aerodramus fuciphagus), into EquiPlus® and E-Z Mixin® extenders on the quality of chilled Arabian stallion semen at various storage times (0, 24 and 48 h). Ten ejaculates were collected from five stallions, and diluted using the two extenders containing 0% (control), 0.12%, 0.24% and 0.24% of EBN + seminal plasma (SP). All the diluted semen samples were then cooled and stored at 5 °C, and examined at 0, 24 and 48 h. Sperm kinetic parameters were assessed using computer assisted sperm analysis (CASA) and viability were assessed using Hoechst33342/PI stain. In both extenders, total motility (TM) and progressive motility (PM) were significantly higher at 0.12% and 0.24% compared to 0.24% + SP at 24 and 48 h. At 0.12%, EZ mixin® treated semen had significantly higher TM and PM than EquiPlus® at 24 and 48 h. At 0.12% and 0.24%, average path velocity (VAP), straight-line velocity (VSL) and curvilinear velocity (VCL) were significantly higher in E-Z mixin® treated semen compared to EquiPlus® at 24 and 48 h. Comparisons between the two extender types at different concentrations of EBN showed no significant difference in lateral head amplitude (ALH), linearity (LIN), straightness (STR), beat cross frequency (BCF) and viability, irrespective of the storage time. The percentage of viable was significantly higher in E-Z mixin® than EquiPlus® at 0 and 48 h in control and 0.12%. Supplementation of the E-Z mixin® extender with 0.12% and 0.24% EBN concentrations in the absence of SP provided better CASA parameters such as TM, PM, VAP, VSL, and VCL at 24 and 48 h storage time. In conclusion, the results of this study indicated that chilled semen from Arabian stallion that was extended using E-Z mixin® and supplemented with 0.12% and 0.24% EBN concentrations performed better and yielded superior results in sperm kinetic parameters and % viable compared to EquiPlus® at 24 and 48 h storage time.(AU)

Influence of antibiotics on bacterial load and sperm parameters during short-term preservation of collared peccary semen

Studies on semen and sperm cells are critical to develop assisted reproductive technologies for the conservation of the collared peccary. The objective of the study was to compare the effect of different antibiotics on the bacterial load and sperm quality during short-term storage of peccary semen. Fresh semen samples from 10 males were extended in Tris-egg yolk or Tris-Aloe vera supplemented with streptomycin-penicillin (SP; 1 mg/mL - 1000 IU/mL or 2 mg/mL - 2000 IU/mL) or gentamicin (30 µg/mL or 70 µg/mL) before storage at 5°C. Bacterial load and sperm motility, membrane integrity and function, mitochondrial activity, and morphology, were evaluated at different time points for 36 h. The SP and gentamicin treatments concentration inhibited (p < 0.05) bacterial growth for 36 h regardless of the extender. Compared to the other treatments, Tris-egg yolk plus 70 µg/mL gentamicin maintained the sperm parameters for longer, including total motility (41.9 ± 6.1%) at 24 h, and membrane integrity (58.3 ± 2.1%) at 36 h. In contrast, the highest SP concentration in both extenders impaired sperm membrane integrity at 36 h (p < 0.05). For the liquid storage of collared peccary semen, it therefore is recommended to use Tris extender supplemented with egg yolk and gentamicin (70 µg/mL).(AU)

Conduzindo um garanhão de forma segura para cobertura ou colheita de sêmen / Handling a breeding stallion to in-hand natural service or semen collection

Cavalos de alto valor genético, morfológico e funcional adquirem valorização comercial, tornando-se garanhões que, frequentemente ultrapassam valores superiores a milhões de reais. Várias pesquisas abordam diluentes para sêmen fresco e congelado, melhoria da qualidade seminal, infertilidade, mas pouco se pesquisa e publica a respeito da contenção e condução do garanhão para colheita de sêmen/cobertura. Embora, na maioria das vezes não seja o médico veterinário que conduz esses animais para a sala de colheita de sêmen, é importante ter conhecimento do manejo desses animais, já que acidentes, às vezes fatais, são relatados em diversos lugares do Brasil e do mundo. Portanto, objetivou-se com esta revisão abordar os principais métodos de contenção e condução de um garanhão para a área de cobertura ou colheita de sêmen no intuito de minimizar riscos e acidentes.(AU)

Few horses become stallions, due to that, they have high genetic merit and commercial value; Usually, more than millions of reais. Lots of research has been addressed to extenders for fresh and frozen semen, improvement of seminal quality, infertility; However, not that much has been researched and published about the restraint and handling a breeding stallion to in-hand natural service or semen collection. Although, most of the time it is not the veterinarian who handles these animals to the breeding environment. It is essential to know how to do in a safety way, since accidents, sometimes fatal, have been reported in different places in Brazil and the world. Therefore, this review aimed to address the restraint methods and handle a breeding stallion to a breeding-Shed room.(AU)

Conduzindo um garanhão de forma segura para cobertura ou colheita de sêmen / Handling a breeding stallion to in-hand natural service or semen collection

Cavalos de alto valor genético, morfológico e funcional adquirem valorização comercial, tornando-se garanhões que, frequentemente ultrapassam valores superiores a milhões de reais. Várias pesquisas abordam diluentes para sêmen fresco e congelado, melhoria da qualidade seminal, infertilidade, mas pouco se pesquisa e publica a respeito da contenção e condução do garanhão para colheita de sêmen/cobertura. Embora, na maioria das vezes não seja o médico veterinário que conduz esses animais para a sala de colheita de sêmen, é importante ter conhecimento do manejo desses animais, já que acidentes, às vezes fatais, são relatados em diversos lugares do Brasil e do mundo. Portanto, objetivou-se com esta revisão abordar os principais métodos de contenção e condução de um garanhão para a área de cobertura ou colheita de sêmen no intuito de minimizar riscos e acidentes.

Few horses become stallions, due to that, they have high genetic merit and commercial value; Usually, more than millions of reais. Lots of research has been addressed to extenders for fresh and frozen semen, improvement of seminal quality, infertility; However, not that much has been researched and published about the restraint and handling a breeding stallion to in-hand natural service or semen collection. Although, most of the time it is not the veterinarian who handles these animals to the breeding environment. It is essential to know how to do in a safety way, since accidents, sometimes fatal, have been reported in different places in Brazil and the world. Therefore, this review aimed to address the restraint methods and handle a breeding stallion to a breeding-Shed room.

Nutracêuticos e antioxidantes na qualidade seminal de garanhões – resultados obtidos na última década / Nutraceuticals and antioxidants on the stallion semen quality – results obtained over the last decade

Alguns anos se passaram desde o nascimento do primeiro potro com sêmen congelado. Entretanto, sabe-se que a refrigeração e a criopreservação dos espermatozoides podem induzir danos celulares, que estão associados ao rompimento de membranas lipídicas, resultando em danos às mitocôndrias e perda da integridade de membrana plasmática e acrossomal. Devido a isso, novos diluentes, protocolos de refrigeração e criopreservação de sêmen e vários suplementos nutricionais têm sido utilizados nos últimos anos com o objetivo de melhorar a fertilidade de garanhões. O termo nutracêutico vem se tornando comum na reprodução animal e descreve produtos derivados dos alimentos, que podem fornecer benefícios adicionais a saúde, além dos valores básicos encontrados na dieta. Entre os vários nutracêuticos utilizados na reprodução, temos o ômega-3, ômega-6, vitaminas do complexo B, L-carnitina, β-caroteno e antioxidantes. Portanto, objetivou-se apresentar as principais substâncias utilizadas como nutracêuticos e antioxidantes, seja utilizado de forma oral ou incorporado a diluentes de sêmen, com a finalidade de melhorar a qualidade seminal de garanhões, e consequentemente incrementar taxas de prenhez.

It has been many years since the foaling of the first foal with frozen semen. However, it is known that cooling and freezing semen can induce damage to the cells, which are associated to injuries to spermatic membranes. For this reason, new extenders, cooling and frozen semen protocols and several nutritional supplements have been used in recent years with the aim of improving the stallions' fertility. Nutraceuticals have become remarkably popular in animal reproduction and describe products derived from foods, which may provide additional health benefits, beyond the basic value found in diets. Among the main nutraceuticals used in male reproduction, there are omega-3 and 6, B-complex vitamins, Lcarnitine, β-carotene, and antioxidants. The aim of this review is to present the main substances used as nutraceuticals and antioxidants, either used orally or in combination with semen extenders, focusing on the improvement of seminal quality, and pregnancy rates.