Cryopreservation of jaguar (Panthera onca) sperm cells using different cryoprotectants and different thawing temperatures

The cryopreservation of jaguar semen must be improved to produce high-quality biobanking doses. Until now, the rare studies of semen freezing in the species have only evaluated glycerol, always with a significant reduction in sperm quality in thawed semen. The purpose of this study was to assess the efficacy of three cryoprotectants, dimethylsulfoxide (DMSO), glycerol (GLY), and methanol (MET), in the cryopreservation of jaguar semen in an LDL-based extender, as well as the effect of thawing temperature on dosage quality. Five mature males with a history of reproduction were used. On the males, an infrared thermal image (IRT) was captured, the spicules and testes were analyzed, and the CASA system was used to evaluate the quality of fresh and thawed sperm. The superficial IRT was 4.6 ± 1.2 °C cooler than the anal sphincter, and the semen measured between 27.3 and 28.7 °C shortly after exiting the urethra. The total motility of fresh sperm was 55.3 ± 22.6%, and progressive motility was 36.3 ± 18%. The total motility of thawed sperm was 5.28 ± 2.51%, 4.49 ± %2.49, and 0.51 ± 0.62% for DMSO, GLY, and MET, respectively. DMSO and GLY performed better than MET, and there was no difference in thawing temperature (37°C 30 s vs. 50°C 12 s). All animals exhibit a considerable level of morphological changes in sperm. Low amounts of total and progressive motility were found in the thawed sperm. Males with a high level of sperm morphological changes were found to be fertile, but the lone male with normospermia was infertile. Thus, we contest the applicability of the commonly used morphological classification for bovines to felid species.(AU)

Viability of Toxoplasma gondii in cattle semen cryopreserved with different concentrations of cryoprotectant / Viabilidade do Toxoplasma gondii em sêmen de bovino criopreservado em diferentes concentrações de crioprotetores

ABSTRACT: Toxoplasma gondii can be eliminated in bovine semen. Cryopreserved semen is often used due to the fact that artificial insemination in dairy and beef cattle provides benefits in terms of production. However, little is known regarding the viability and infectivity of T. gondii tachyzoites in cryopreserved bovine semen. In the present study, cattle semen negative for T. gondii were contaminated with 1 x 106 tachyzoites (RH strain) and cryopreserved with and without different cryoprotectants, such as DMSO (concentrations of 2.5%, 5.0%, 7.5%, 8.0% and 10.0%) and glycerol (2.25%, 2.5%, 3.0%, 5.0%, 7.5% and 10.0%), followed by freezing in liquid nitrogen (-196°C). After 24 hours, the samples were thawed and inoculated in 10 mice per cryoprotectant concentration. The mice were evaluated for clinical signs of toxoplasmosis (rough coat, diarrhea, hypoactivity and sudden death) as well as serum titers of IgM and IgG and the presence of tachyzoites in the peritoneal lavage. The results revealed that T. gondii remained infective in all samples. Clinical signs of toxoplasmosis were observed in the mice beginning with the 6th day post-inoculation (DPI) and 100% lethality was found between the 7th and 9th DPI. Viable tachyzoites were recovered from peritoneal exudate of dead mice (except for the control group), with higher mean of tachyzoite counts in the intraperitoneal lavage for 5% DMSO (±3.32 x 106), 8% DMSO (±3.53 x 106), 3% glycerol (±4.75 x 106), 7.5% glycerol (±6.26 x 106) and the absence of cryoprotectant (±3.11 x 106). Seroconversion occurred in the treated groups, with titers of IgG from 1:16 to 1:128 and IgM from 1:16 to 1:512. T. gondii viability and infectivity were maintained in cattle semen during 24 hours of cryopreservation at -196°C with and without cryoprotectant. However, further studies are necessary to determine whether cryopreserved semen contributes to the spread of toxoplasmosis through artificial insemination.

RESUMO: Sabe-se que Toxoplasma gondii pode ser eliminado no sêmen bovino. A inseminação artificial em bovinos leiteiros e de corte proporcionou avanços e benefícios nas produções e para isso o sêmen criopreservado é frequentemente utilizado. No entanto, pouco se sabe sobre a viabilidade e infectividade dos taquizoítos de T. gondii em sêmen bovino criopreservado. Para isso o sêmen bovino, negativo para T. gondii, foi contaminado com 1x106 taquizoítos (cepa RH), criopreservados com ou sem diferentes crioprotetores como DMSO (2.5%, 5.0%, 7.5%, 8.0% e 10.0%) e Glicerol (2.25%, 2.5%, 3.0%, 5.0%, 7.5% e 10.0%) e congelados em nitrogênio líquido (-196°C). Após 24 horas, essas amostras foram descongeladas e inoculadas em 10 camundongos por diluente de concentração de crioprotetor. Os camundongos foram avaliados quanto a sinais clínicos de toxoplasmose (pele áspera, diarreia, hipoatividade e morte súbita), títulos séricos de IgM e IgG e presença de taquizoítos no lavado peritoneal. Os resultados mostraram que T. gondii se manteve infectante em todas as amostras, inclusive naquelas sem crioprotetor. Sinais clínicos de toxoplasmose foram observados nos camundongos a partir do 6º dia pós-inoculação (DPI) e 100% de letalidade foi verificada entre o 7º ao 9º DPI. Nos camundongos mortos, exceto no grupo controle, taquizoítos viáveis foram recuperados do exsudato peritoneal, com maior média de taquizoítos quantificados na lavagem intraperitoneal para DMSO a 5% (±3.32x106), 8% (±3.53x106) e glicerol 3% (±4.75x106), 7,5% (±6.26x106) e livre de crioprotetor (±3.11x106). A soroconversão ocorreu nos grupos tratados com títulos de IgG (1:16 a 1:128) e IgM (1:16 a 1:512). A viabilidade e infectividade do T. gondii no sêmen bovino durante as 24 horas de criopreservação a -196°C foram mantidas com ou sem crioprotetor. No entanto, mais estudos são necessários para verificar se o sêmen criopreservado contribui para a disseminação da toxoplasmose na inseminação artificial.

Glyceryl guaiacol ether for sedation of horses undergoing orchiectomy when standing / Éter gliceril-guaiacol para sedação de equinos submetidos à orquiectomia em pé

A study was designed to compare two sedation protocols to be used in horses undergoing orchiectomy when standing. In both protocols, the induction to the sedative state was performed with bolus detomidine at a dose of 10 µg/kg, intravenously (IV). In the first protocol (eight horses), the sedative state was maintained with 1% glyceryl guaiacol ether (GGE) in continuous infusion, at a dose of 1ml/kg/hour IV. In the second protocol (eight horses) this sedative effect was maintained with detomidine in continuous infusion at the same dose and induction route. Orchiectomy was performed on all animals. The two protocols allowed the surgeries to be performed when standing. However, horses kept under sedation by the GGE showed greater relaxation and a more intense degree of sedation. It should be noted that the use of GGE to maintain the sedative state in horses is unprecedented in the literature. The drug is used in anesthetic protocols in the species, but only in pre-anesthetic medication in general anesthesia. It was concluded that the two sedation protocols allowed the performance of orchiectomy with the horses when standing. However, the protocol in which 1% GGE was used showed more profound sedation, without adverse effects.

Foi realizado um estudo para comparar dois protocolos de sedação a serem utilizados em equinos submetidos à orquiectomia em estação. Em ambos os protocolos, a indução ao estado sedativo foi realizada com bolus de detomidina, na dose de 10µg/kg, por via intravenosa (IV). No primeiro protocolo (oito cavalos), o estado sedativo foi mantido com 1% de éter gliceril guaiacol (GGE) em infusão contínua, na dose de 1mL/kg/hora IV. No segundo protocolo (oito cavalos), esse efeito sedativo foi mantido com detomidina, em infusão contínua, na mesma dose e via de indução. Foi realizada orquiectomia em todos os animais. Os dois protocolos permitiram que as cirurgias fossem realizadas em estação No entanto, os cavalos mantidos sob sedação pelo GGE apresentaram maior relaxamento e grau de sedação mais intenso. Ressalta-se que o uso de GGE para manutenção do estado sedativo em equinos é inédito na literatura. O fármaco é utilizado em protocolos anestésicos na espécie, mas apenas na medicação pré-anestésica em anestesia geral. Concluiu-se que os dois protocolos de sedação permitiram a realização da orquiectomia com os cavalos em estação. No entanto, o protocolo em que foi utilizado GGE 1% apresentou sedação mais profunda, sem efeitos adversos.

DGAT and LEP gene polymorphisms and their association with carcass characteristics and the lipid profile of meat from Nellore cattle / Polimorfismo dos genes DGAT e LEP e sua associação com características de carcaça e perfil lipídico da carne de bovinos nelore

The objective of this study was to evaluate the association of polymorphisms in the diacylglycerol acyltransferase (DGAT) and leptin (LEP) genes with the performance, carcass characteristics, meat quality, and lipid profile of Nellore cattle. A total of 100 intact male Nelore cattle were used to analyze the performance, carcass, physicochemical and centesimal composition, and fatty acid profile of beef. To identify the polymorphisms, the PCR­single-strand conformation polymorphism (SSCP) technique was applied to genomic DNA extracted from muscle tissue. The SSCP technique revealed the presence of four band patterns for the DGAT gene (AC, AD, AE and BB) with five alleles (A, B, C, D and E). For the LEP gene, five band patterns (AA, AB, AC, BB and BC) with three alleles (A, B and C) were observed. For the LEP gene, the AB genotype was associated with higher backfat thickness and ribs weight, while the BB genotype was associated with lower ribs yield; higher hindquarter yield was associated with AC and BB genotypes. Higher contents of C17:0, C18:0 and lower contents of C18:2ω6C, total polyunsaturated fatty acids, total ω6 and ratio of polyunsaturated and saturated fatty acids (PUFA/SFA) were verified for the AC genotype of the LEP gene. The AC and AA genotypes of the LEP gene were associated with higher means of C15:0 and C18:1ω9t. For the DGAT gene, the highest C24:0 content was associated with the AE genotype and the lowest with the AD and BB genotypes. Polymorphisms in the DGAT and LEP genes influence carcass parameters and the lipid profile of the meat of Nellore cattle.

Objetivou-se com este estudo avaliar a associação do polimorfismo dos genes da Diacilglicerol Aciltransferase (DGAT) e Leptina ­ (LEP) em relação ao desempenho, características de carcaça, qualidade de carne e perfil lipídico de bovinos Nelore. Para o estudo, foram utilizados um total de 100 bovinos nelores machos inteiros e avaliado os parâmetros de desempenho, composição da carcaça, composição físico-química, centesimal e perfil lipídico da carne. Para identificação dos polimorfismos foi utilizada a técnica de PCR-SSCP a partir da extração do DNA genômico do tecido muscular. A técnica de SSCP revelou a presença de quatro padrões de banda para o gene DGAT (AC, AD, AE e BB) com cinco alelos (A, B, C, D e E) e, para o gene LEP foram verificados cinco padrões de bandas (AA, AB, AC, BB e BC) com três alelos (A, B e C). Para o gene LEP, o genótipo AB foi associado a maior espessura de gordura subcutânea e peso do corte ponta de agulha, enquanto o genótipo BB foi associado a menor rendimento do corte ponta de agulha; maior rendimento do corte traseiro foi associado aos genótipos AC e BB. Maiores teores de C17:0, C18:0 e menores de C18: 2ω6C, total de ácidos graxos poli-insaturados, total de ω6 e a relação de ácidos graxos poli-insaturados e saturados (POL/SAT) foram verificados para o genótipo AC do gene LEP. Os genótipos AC e AA do gene LEP foram associados a maiores médias de C15:0 e C18:1ω9t. Para o gene DGAT, os maiores teores de C24:0 foram associados o genótipo AE e o menores aos genótipos AD e BB. A ocorrência de polimorfismo nos genes DGAT e LEP revelaram influência destes sobre parâmetros de carcaça e perfil lipídico da carne de bovinos Nelore.

Effect of glycerol addition time on the cryopreserved Korean native brindle cattle (Chikso) sperm quality

Although cryopreservation is an efficient method for maintaining the biological and genetic resources of sperm, the sperm damage during the cryopreservation process cannot be ignored. It should be possible to obtain the most effective cryopreservation performance by accurately grasping the effects of various factors on the cryopreservation of sperm. The previous study demonstrated that a suitable standard protocol for cryopreservation of Korean native brindled cattle (Chikso) does not exist, based on the methods for semen cryopreservation of Chikso differ in each research center. The most obvious difference between most of protocols is the addition of glycerol before and after cooling during the Chikso cryopreserved semen process. Therefore we focused on the effects of glycerol addition time on the quality of cryopreserved Chikso sperm. In the present study, 27 individual Chikso samples were collected by transrectal massage and divided into two parts: the "cryopreservation method A" group (adding glycerol before cooling) and the "cryopreservation method B" group (adding glycerol after cooling). Meanwhile, the values of various sperm parameters were derived from each group, including sperm motility, kinematics, capacitation status, cell viability, and intracellular ATP levels, which we used to compare and evaluate sperm function. The results of this study indicated that during the semen cryopreservation process of the Chikso, the addition of glycerol after cooling yielded superior results in a variety of sperm parameters, such as sperm motility, progressive motility, rapid motility, VCL, VSL, VAP, ALH, capacitation status, viability, and intracellular ATP level after freezing and thawing. Our study is suggested that the glycerol addition time during the cryopreservation process for Chikso should be considered. In addition, our results may be provided reference to develop suitable the cryopreservation procedure of the Chikso sperm.(AU)

Comparação entre diferentes técnicas empregadas na conservação e manutenção de peças anatômicas / Comparison between different techniques used in the conservation and maintenance of anatomical parts

Dentre as diversas formas de obter a conservação de peças anatômicas para estudo, a formolização é a mais utilizada. Esta técnica altera a coloração e textura, dificultando o processo de ensino-aprendizagem. Além disso, proporciona um ambiente insalubre que pode comprometer a saúde de indivíduos expostos constantemente ao formol. Existem técnicas alternativas como a glicerinação, criodesidratação e corrosão, as quais reduzem os riscos à saúde ocasionados pela exposição aos materiais, proporcionam um ambiente livre de odores desagradáveis e facilitam a manipulação e o armazenamento das peças, quando comparadas com a formolização. O presente trabalho visa à apresentação de técnicas alternativas para a conservação e manutenção de peças anatômicas e seus devidos procedimentos, buscando a substituição da utilização do formol como forma principal de conservação. A técnica de glicerinação apresenta grande eficiência quanto à preservação das características de coloração e formato das vísceras, além de ser uma técnica de baixo custo e não apresentar riscos à saúde, assim como a técnica de criodesidratação, a criodesidratação difere em relação à tonalidade das vísceras, entretanto demonstra facilidade quanto à conservação, armazenamento e manutenção das mesmas. Já a injeção e corrosão de órgãos facilitam a visualização da arquitetura interna de órgãos ocos, além de permitir fazer o modelo de vascularização de órgãos. No entanto, esta pode ser considerada uma técnica cara a depender do polímero utilizado.

Among the several ways to obtain the conservation of anatomical parts for study, formalization is the most used. This technique changes the color and texture, making the teaching-learning process difficult. In addition, it provides an unhealthy environment that may compromise the health of individuals constantly exposed to formaldehyde. There are alternative techniques such as glycerination, cryodehydration, and corrosion, which reduce health risks caused by exposure to materials, provide an environment free of unpleasant odors and facilitate handling and storage of parts, when compared to formalization. This work aims to present alternative techniques for the conservation and maintenance of anatomical parts and their proper procedures, seeking to replace the use of formaldehyde the main form of conservation. The glycerination technique presents great efficiency in preserving the viscera's color and shape characteristics, in addition to being a low-cost technique and not presenting health risks, as well as the cryodehydration technique. The cryodehydration differs in relation to the tonality of the viscera; however, it demonstrates convenience in terms of their conservation, storage, and maintenance. On the other hand, the injection and corrosion of organs facilitate the visualization of the internal architecture of hollow organs, in addition to allowing the model of organ vascularization. However, it can be considered an expensive technique depending on the polymer used.

Glycerin supplementation strategies for three or seven days affects oxidative stress, follicle dynamics and ovulatory response in Morada Nova sheep

The Ziwuling black goat is an indigenously in China, their offspring are frequently affected by congenital cryptorchidism. The extracellular matrix (ECM) contains cytokines and growth factors that regulate the development of the testis, and component changes often result in pathological changes. Cryptorchidism is closely related to structural changes in ECM. In this study, the histochemical staining, immunohistochemical, immunofluorescence and Western blot combined with semi-quantitative analysis was used to describe the distribution of the important ECM components Collagen type IV (Col IV), laminin (LN)and heparan sulfate proteoglycans (HSPG) in the normal and cryptorchid testes of Ziwuling black goats. Results showed that: The histochemical staining showed that the dysplasia of seminiferous tubules and decreased number of Sertoli cells in cryptorchidism, as well as sparse collagen fiber. Meanwhile, the distribution of reticular fibers is relatively rich. Furthermore, the PAS and AB staining in the interstitial vessels and lamina propria of seminiferous tubules is weak. The immunohistochemical and immunofluorescence revealed that Col IV, LN was strongly expressed in Leydig, Sertoli cells of normal testes and moderately positive in the spermatogonia and spermatids, but HSPG was not expressed in the spermatogonia. However, cryptorchidism, the expression of Col IV, LN and HPSG in Leydig, Sertoli cells significantly decreased, as well as the expression of Col IV and LN in capillary endothelial cells, but HSPG was moderately expressed in spermatogonia. Based on these data, the underdevelopment of spermatogenic epithelium, decreased synthesis function of collagen fibers and Leydig cells develop usually in the cryptorchidism were shown to be closely related to the abnormal metabolism of Col IV and LN. The positive expressed of HSPG in the spermatogonia of cryptorchid testes is related to the compensatory development of spermatogonia.(AU)

Comparison of three different strategies to treat sciatic nerve regeneration: an experimental study

Purpose: To compare the effect of vein conduit filled with adipose tissue stem cells (ASC) on peripheral nerve injury regeneration. Methods: We analyzed 30 male Wistar rats surgically submitted to a 5-mm gap on the sciatic nerve. Then, the animals were divided into three groups: nerve autografting (AG, n=10), autogenous inverted glycerol-conserved vein (VG, n=10), and autogenous inverted glycerol-conserved vein + ASC (VASCG, n=10). The study endpoints were neuromotor functional analysis, gastrocnemius muscle weight, and sciatic nerve graft histomorphometry analysis. In the histologic analysis, we added a control group (naïve nerve). Results: Regarding functional analysis (Walking tract- score), the findings at week 3 showed a difference between the AG and the VG (-96.6 vs. -59.6, p=0.01, respectively) and between the VG and the inverted vein + VASCG (-59.9 vs. -88.92, p=0.02). At week 12, this study showed a difference between the AG and the VG (-64.8 vs. -47.3, p=0.004, respectively), and also a difference between the VG and the VASCG (-47.3 vs. -57.4, p=0.02, respectively). There was no difference in the histomorphometry analysis (nerve diameter, Schwann cells counting). The gastrocnemius muscles on the intervention side were more atrophic when compared to the gastrocnemius muscles on the control side. Conclusions: Our results suggested better functional recovery in the inverted vein group when compared to control group, and inverted vein + ASC group.

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 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)

Propolis in the control of bacterial bovine mastitis: a tool for the production of organic milk / Própolis no controle de mastite bovina bacteriana: uma ferramenta para a produção de leite orgânico

Bovine mastitis is one of the main causes of economic damage in dairy farms. Therefore, the control and prevention of microorganisms involved in this disease, mainly Escherichia coli, Staphylococcus aureus, and Streptococcus agalactiae, are essential. One of the most important steps for the prevention of the disease is the use of antiseptic products before and after the milking process to avoid bacteria from infecting the udder of the animal. Currently, the most used antiseptic product in dairy farms is iodine-based, and organic dairy farms, which follow several strict regulations, including the use of natural products whenever possible, are often forced to adopt non-natural antiseptic products, such as iodine-based ones, because of the lack of natural alternatives. Propolis, a natural substance produced by honeybees, has been extensively studied for its various properties, one of which is antimicrobial activity. Therefore, a new natural antiseptic product containing 1% propolis in 10% hydroalcoholic solution for the pre-dipping, and 10% glycerol solution added with 0.2% citronella oil for the post-dipping was analyzed for its capacity to reduce bacteria in vivo in order to prevent bovine mastitis, allowing its use on organic dairy farms. A total of 128 samples were analyzed in terms of bacterial growth for Enterobacteriaceae and Staphylococcus spp. using the spreadplate technique. The reduction in the bacterial concentration after the application of the products was compared between two antiseptic solutions, an iodine-based solution as the control and a propolis-based one as the natural alternative. The results obtained show a similar efficiency for both products in terms of total bacterial reduction, indicating considerable antimicrobial activity against bacteria most commonly associated with bovine mastitis. Molecular analysis was carried out for the identification of Streptococcus agalactiae; the PCR results were negative for the presence of S. agalactiae in all samples, indicating that the animals most likely did not have any form of the disease. The efficiency of the natural antiseptic was satisfactory, indicating an important find facilitating organic milk production worldwide, showcasing a natural antiseptic solution with efficient antimicrobial activity.(AU)

A mastite bovina é uma das principais causas de prejuízo econômico na indústria leiteira, portanto o controle e prevenção de microrganismos envolvidos nessa doença, principalmente Escherichia coli, Staphylococcus aureus e Streptococcus agalactiae é essencial. Uma das principais etapas na prevenção dessa doença é o uso de produtos antissépticos antes e depois do processo de ordenha, a fim de evitar contaminação bacteriana no úbere do animal. Atualmente, o produto antisséptico mais utilizado na indústria leiteira é a base de iodo, e fazendas produtoras de leite orgânico, que precisam seguir uma série de regulações estritas, incluindo o uso de produtos naturais sempre que possível, são frequentemente forçadas a adotar antissépticos não-naturais, como os a base de iodo por falta de alternativas naturais. Própolis, uma substância natural produzidas por abelhas, tem sido extensivamente estudada por suas várias propriedades, sendo uma delas antimicrobiana. Portanto, um novo produto antisséptico natural contendo própolis à 1% em solução hidroalcóolica 10% para o pré-dipping, e glicerinada 10% adicionada de óleo de citronela à 0,2% para o pós-dipping, foi avaliada quanto a sua capacidade de reduzir bactérias in vivo e prevenir a mastite bovina, além de poder ser utilizado na indústria leiteira orgânica. Um total de 128 amostras foram analisadas em termos de crescimento bacteriano para Enterobacteriaceae e Staphylococcus spp. utilizando a técnica de plaqueamento em superfície, a redução da concentração bacteriana após a aplicação dos produtos foi comparada entre duas soluções antissépticas, uma solução a base de iodo servindo como controle, e uma solução a base de própolis como a alternativa natural. Os resultados obtidos mostraram uma eficiência similar entre os produtos à base de iodo e própolis em termos de redução bacteriana total, indicando uma grande atividade antibacteriana contra as bactérias mais comumente associadas com a mastite bovina. Realizou-se análise molecular para a identificação de Streptococcus agalactiae, os resultados da PCR foram negativos para a presença de S. agalactiae em todas as amostras, indicando que os animais provavelmente não possuíam nenhuma forma da doença. A eficiência do antisséptico natural foi satisfatória, indicando um achado importante para auxiliar o crescimento da indústria de leite orgânico de forma mundial, mostrando uma solução antisséptica natural com atividade antimicrobiana eficiente.(AU)

Avanços na andrologia de animais selvagens / Advances in wild animal andrology

As tecnologias de reprodução assistida (TRA) são de fundamental importância para a conexão de indivíduos em diferentes localidades, facilitando assim o intercâmbio genético e favorecendo a variabilidade genética de uma espécie. Por esta razão, as TRAS podem ser ferramentas importantes para a conservação de espécies ameaçadas de extinção. Apesar dos esforços nas últimas décadas, o avanço no desenvolvimento de tais tecnologias está aquém à urgência de reverter processos de baixa variabilidade genética em algumas espécies. A necessidade de refinamento das técnicas para as particularidades fisiológicas e comportamentais de cada espécie, somada à raridade de acesso aos animais são os principais fatores relacionados as dificuldades em se avançar com as TRAS. As técnicas mais recentes desenvolvidas para a recuperação de espermatozoides em animais selvagens são a colheita farmacológica, com uso de alfa-2-agonistas e a criopreservação / vitrificação testicular com posterior cultivo. Pouco de avançou, no entanto, em relação aos métodos de criopreservação, prevalecendo associação clássica de TRIS-gema-glicerol. Discutimos, então os métodos usados para acesso ao gameta masculino em espécies selvagens e suas aplicações na conservação animal.(AU)

Assisted reproduction technologies (ART) are of fundamental importance for connecting individuals in different locations, thus facilitating genetic exchange and favoring the genetic variability of a species. For this reason, TRAS can be important tools for the conservation of endangered species. Despite efforts in recent decades, the advance in the development of such technologies is short of the urgency of reversing processes of low genetic variability in some species. The need to refine the techniques for the physiological and behavioral particularities of each species, added to the rarity of access to animals, are the main factors related to the difficulties in advancing with TRAS. The most recent techniques developed for sperm collection in wild animals are pharmacological collection, with the use of alpha-2-agonists and testicular cryopreservation / vitrification with subsequent cultivation. Little progress has been made, however, in relation to cryopreservation methods, prevailing the classic association of TRIS-yolk-glycerol. We therefore discuss the methods used to access the male gamete in wild species and their applications in animal conservation.(AU)

Effects of glycerol and sodium pentaborate containing new formulation on sleeve gastrectomy model in rats

ABSTRACT Purpose: The development of cutting surface leakage and postoperative peritoneal adhesions (PPA) after sleeve gastrectomy (SG) are the most serious operative complications. We investigated the effectiveness of the newly developed glycerol and sodium pentaborate containing formulation on the prevention of these complications. Methods: Sixteen Sprague Dawley rats (mean weight 310 ± 50 g, mean age 3 months old) were divided into two groups, consisting of eight rats in each. SG and a double-layer suture technique were performed for each group. In study group, there was the mixture of 2 mL 3% glycerol plus 3% sodium pentaborate formulation, and in the control group 2 mL 0.9% NaCl was injected into the peritoneal cavity. Rats were sacrificed after 30 days, then macroscopic adhesion grade scoring and histopathological evaluations were assessed. Results: Macroscopic PPA scores in the control and study groups were 2.75 ± 0.16 and 1.50 ± 0.327, respectively (p = 0.004). Histopatologic fibrosis scores in the control and study groups were 0.87 ± 0.125 and 2.00 ± 0.26, respectively (p = 0.002). Conclusions: In SG operation model, glycerol plus sodium pentaborate compound decreased PPA formation and also increased stomach cut surface line fibrosis. This new formulation is hopeful for more safe SG operations.

Plasticizer types and whey protein coatings on internal quality and shelf life of eggs stored for 42 days

ABSTRACT: Effects of plasticizer types and whey protein concentrate (WPC) as coatings were evaluated on internal quality and shelf life of eggs stored for 42 days. Eggs were coated with WPC at 8 % solution combined with the plasticizers glycerol (GLY), sorbitol (SOR), and propylene glycol (PRO). The eggs were stored at 20 °C. Weight loss, Haugh Unit (HU), albumen and yolk pH, and yolk index (YI) were evaluated weekly from day 0 to 42 days. After, electron microscopy of the eggshell structure was performed. The data was submitted to the analysis of variance and the effects of treatment, storage time, and the interaction between these factors were evaluated. There was significant interaction between factors (treatment and periods) for weight loss (p 0.0001), HU (p 0.0001), albumen (p 0.0001), and yolk pH (p 0.0001), and YI (p 0.0001). After 42 days, uncoated eggs showed greater weight loss (5.4 %), compared to WPC+GLY, (3.8 %), WPC+SOR (3.3 %) and WPC+PRO (3.9 %). Similar results were verified for HU at 42 days of storage. Uncoated eggs showed HU of 58.46 (B), while coated eggs showed higher values: WPC+GLY 66.58 (A), WPC+SOR 68.79 (A), and WPC + PRO 71.53 (A). The plasticizers GLY, SOR and PRO, associated with WPC, demonstrated effectiveness in maintaining the quality of eggs throughout the 42 days of storage. However, WPC+SOR showed superiority in preserving quality integrity of eggs. This result can be related to the chemical structure of SOR, making the combination more efficient for storage.

Seed and seedling morphology of Bauhinia scandens L / Morfologia de semente e plântula de Bauhinia scandens L

Bauhinia scandens has potential importance as an ornamental and medicinal plant. Researchers have isolated and identified 1-O-alkylglycerol in the leaves of the B. scandens plant, and established antitumor properties using the Brine Shrimp toxicity test, an internationally accepted bioassay. Although this species has high potential, little is known about the viability of seedling production and the morphology of these plants, particularly in terms of seed characteristics and initial stages of germination. The objective of this study was to characterize the seed morphology, germination, and seedlings of B. scandens. Seed water content, weight, and coloration were evaluated. This study also included a description of seed biometrics, external and internal structures, germination, and seedling morphology. Internal seed morphology was evaluated by the anatomical sectioning and X-ray methods. The morphology data obtained were subjected to descriptive statistical analysis and germination data were determined using Cramér's V. B. scandens seeds have a coloration ranging from very dark grayish-red to dark reddish-brown, flat oblong shapes, and rounded bases and apexes with full or slightly undulating margins. Healthy seedlings are produced mainly by seeds with well-formed internal structures. The reddish petiole of the seedling leaves is a taxonomic character for B. scandens identification. The non-domestication and genetic variability of this species reflect on the seed and seedling color and size variation.(AU)

Bauhinia scandens tem potencial como planta ornamental e importância medicinal. Nas folhas da planta de B. scandens, foi relatado pelos pesquisadores, o isolamento e identificação de glicerol 1-O-alquilo, os mesmos estabeleceram a propriedade antitumoral por um bioensaio, aceito internacionalmente denominado teste de toxicidade de Brine Shrimp. Por ser ainda pouco conhecida e com alto potencial exploratório, a viabilidade da produção de mudas e maior conhecimento sobre sua morfologia se faz necessária. Faltam informações sobre as características das sementes e estágios iniciais de germinação. O objetivo deste estudo foi caracterizar o tipo morfológico de sementes, germinação e plântulas de B. scandens. Foram avaliados o teor de água, o peso de mil sementes e a coloração das sementes. Este estudo também incluiu a descrição e biometria da semente, estrutura (externa e interna), germinação e morfologia das plântulas. A morfologia interna das sementes foi avaliada por corte anatômico e método de raios-X. Os dados morfológicos obtidos foram submetidos à análise estatística descritiva e os dados de germinação foram testados utilizando Cramér's V. B. scandens apresentam coloração que varia de vermelho acinzentado muito escuro a marrom avermelhado escuro e o tipo de semente é de forma oblonga plana, base e ápice são arredondados e cheios ou margens ligeiramente onduladas. Plãntulas normais são produzidas majoritariamente por sementes com uma estrutura interna bem formada. O pecíolo avermelhado das folhas da plântula é um caráter taxonômico para identificação de B. scandens. A não domesticação e a variabilidade genética desta espécie refletem na variação da cor e tamanho das sementes e plântulas.(AU)

Uso da galactose na vitrificação de sêmen ovino em palhetas / Use of galactose to vitrify ram semen in straws

A vitrificação de espermatozoides é uma técnica que apresenta grande potencial para criopreservação de material genético, e sua eficácia tem sido superior aos métodos convencionais em algumas espécies. No entanto, existem poucos estudos sobre sua eficiência com sêmen ejaculado de carneiros e o uso da galactose como crioprotetor extracelular durante a vitrificação. Objetivou-se com este estudo avaliar o efeito da galactose (0,01 M), associada ou não ao glicerol (3% e 7%), em meio comercial (Steridyl® - controle), na criopreservação de espermatozoides de carneiros pelo método de palhetas, comparando o método clássico de congelação e a vitrificação. Ejaculados de seis carneiros da raça Dorper em idade reprodutiva foram coletados com vagina artificial, aliquotados, diluídos individualmente (100 × 106 espermatozoides/mL) nos meios testados, envasados em palhetas de 0,25 mL e submetidos à congelação clássica ou vitrificação. Foram analisadas a cinemática, morfologia, morfometria, viabilidade, integridade física e funcional da membrana espermática. A congelação clássica obteve melhores resultados de motilidade total e progressiva do que a vitrificação nos quatro extensores testados, uma vez que as amostras vitrificadas não apresentaram motilidade pós-reaquecimento (p < 0,05). A adição de galactose ou glicerol ao meio comercial não trouxe efeito benéfico tanto para a vitrificação quanto congelação clássica.

Sperm vitrification is a technique with great potential for cryopreservation of genetic material, with superior effectiveness compared to conventional methods in some species. However, few studies have shown its efficiency with ejaculated sperm of rams and the use of galactose as an extracellular cryoprotectant during vitrification. This study aimed to evaluate the effect of galactose (0.01 M), with or without glycerol addition (3% and 7%) in a commercial extender (Steridyl® - control) for ram sperm cryopreservation in straws, comparing the classic freezing method and vitrification. Ejaculates from six breeding soundness Dorper rams were collected with an artificial vagina, aliquoted, individually diluted (100 × 106 sperm/mL) on extenders tested, loaded into 0.25 mL straws, and subjected to a classic freezing method or vitrification. Sperm kinematics, morphology, morphometry, viability, and physical and functional integrity of the sperm membrane were evaluated. The classic freezing method resulted in higher total and progressive motility than vitrification, as no motility was detected in vitrified samples after rewarming (p<0.05). The addition of galactose or glycerol to the commercial medium did not benefit both vitrification and the classic freezing method.

Plasticizer types and whey protein coatings on internal quality and shelf life of eggs stored for 42 days

Effects of plasticizer types and whey protein concentrate (WPC) as coatings were evaluated on internal quality and shelf life of eggs stored for 42 days. Eggs were coated with WPC at 8 % solution combined with the plasticizers glycerol (GLY), sorbitol (SOR), and propylene glycol (PRO). The eggs were stored at 20 °C. Weight loss, Haugh Unit (HU), albumen and yolk pH, and yolk index (YI) were evaluated weekly from day 0 to 42 days. After, electron microscopy of the eggshell structure was performed. The data was submitted to the analysis of variance and the effects of treatment, storage time, and the interaction between these factors were evaluated. There was significant interaction between factors (treatment and periods) for weight loss (p < 0.0001), HU (p < 0.0001), albumen (p < 0.0001), and yolk pH (p < 0.0001), and YI (p < 0.0001). After 42 days, uncoated eggs showed greater weight loss (5.4 %), compared to WPC+GLY, (3.8 %), WPC+SOR (3.3 %) and WPC+PRO (3.9 %). Similar results were verified for HU at 42 days of storage. Uncoated eggs showed HU of 58.46 (B), while coated eggs showed higher values: WPC+GLY – 66.58 (A), WPC+SOR – 68.79 (A), and WPC + PRO – 71.53 (A). The plasticizers GLY, SOR and PRO, associated with WPC, demonstrated effectiveness in maintaining the quality of eggs throughout the 42 days of storage. However, WPC+SOR showed superiority in preserving quality integrity of eggs. This result can be related to the chemical structure of SOR, making the combination more efficient for storage.

Plasticizer types and whey protein coatings on internal quality and shelf life of eggs stored for 42 days

Effects of plasticizer types and whey protein concentrate (WPC) as coatings were evaluated on internal quality and shelf life of eggs stored for 42 days. Eggs were coated with WPC at 8 % solution combined with the plasticizers glycerol (GLY), sorbitol (SOR), and propylene glycol (PRO). The eggs were stored at 20 °C. Weight loss, Haugh Unit (HU), albumen and yolk pH, and yolk index (YI) were evaluated weekly from day 0 to 42 days. After, electron microscopy of the eggshell structure was performed. The data was submitted to the analysis of variance and the effects of treatment, storage time, and the interaction between these factors were evaluated. There was significant interaction between factors (treatment and periods) for weight loss (p < 0.0001), HU (p < 0.0001), albumen (p < 0.0001), and yolk pH (p < 0.0001), and YI (p < 0.0001). After 42 days, uncoated eggs showed greater weight loss (5.4 %), compared to WPC+GLY, (3.8 %), WPC+SOR (3.3 %) and WPC+PRO (3.9 %). Similar results were verified for HU at 42 days of storage. Uncoated eggs showed HU of 58.46 (B), while coated eggs showed higher values: WPC+GLY 66.58 (A), WPC+SOR 68.79 (A), and WPC + PRO 71.53 (A). The plasticizers GLY, SOR and PRO, associated with WPC, demonstrated effectiveness in maintaining the quality of eggs throughout the 42 days of storage. However, WPC+SOR showed superiority in preserving quality integrity of eggs. This result can be related to the chemical structure of SOR, making the combination more efficient for storage.(AU)

Uso da túnica vaginal canina conservada em glicerina a 98% como curativo biológico no tratamento de feridas cutâneas produzidas em ratos Wistar / Use of canine vaginal tunic preserved in 98% glycerin as biological curative in the treatment of skin produced in wound Wistar rats

Na procura por material alternativo no tratamento de feridas cutâneas, buscou-se com este estudo avaliar o comportamento da túnica vaginal canina conservada em glicerina a 98% como curativo biológico no tratamento de feridas cutâneas provocadas experimentalmente em ratos. Foram utilizados 16 ratos Wistar, nos quais foram induzidas experimentalmente duas lesões cutâneas na região dorsal, sendo uma cranial e a outra caudal. Uma ferida serviu de controle, a qual não recebeu tratamento, a outra recebeu a túnica vaginal recobrindo toda extensão da lesão. A seleção de qual das feridas, cranial ou caudal, iria receber a membrana foi feita de forma aleatória. Os animais foram distribuídos em quatro grupos de igual número para análises clínicas, macroscópicas e histológicas decorridos três, sete, 14 e 21 dias de pós-operatório. Na avaliação clínica e macroscópica pode-se observar que o processo de reparação teve evolução semelhante tanto no grupo controle quanto no grupo membrana. Na análise histopatológica foi constatada grande quantidade de infiltrado inflamatório nos períodos iniciais, no entanto, decorridos 21 dias, a epiderme já se encontrava reparada, e em todos os momentos foi observada maior presença de tecido conjuntivo no grupo membrana, sugerindo maior precocidade na cicatrização. Deste modo, concluiu-se que a túnica vaginal canina pode ser utilizada como nova alternativa no tratamento de feridas cutâneas.(AU)

In the search for alternative material in the treatment of cutaneous wounds, this study aimed to evaluate the behavior of canine vaginal tunic conserved in glycerin 98% as a biological dressing in the treatment of cutaneous wounds provoked experimentally in rats. Sixteen animals were used in which two cutaneous lesions were caused in the dorsal region, one cranial and the other caudal. One wound served as a control which did not receive treatment, the other received the vaginal tunic covering all extension of the lesion. The selection of which of the wounds, cranial or caudal, would receive the membrane was made at random. The animals were divided into four groups of equal numbers for clinical, macroscopic, and histological analyzes after three, seven, 14 and 21 postoperative days. In the clinical and macroscopic evaluation, it can be observed that the healing process had similar evolution in both the control group and the membrane group. In the histopathological analysis, a large amount of inflammatory infiltrate was observed in the initial periods; however, after 21 days the epidermis was already healed, and at all times a greater presence of connective tissue was observed in the membrane group, suggesting a greater precocity in healing. Thus, it was concluded that the canine vaginal tunic can be used as a new alternative in the treatment of cutaneous wounds.(AU)

Uso da túnica vaginal canina conservada em glicerina a 98% como curativo biológico no tratamento de feridas cutâneas produzidas em ratos Wistar / Use of canine vaginal tunic preserved in 98% glycerin as biological curative in the treatment of skin produced in wound Wistar rats

Na procura por material alternativo no tratamento de feridas cutâneas, buscou-se com este estudo avaliar o comportamento da túnica vaginal canina conservada em glicerina a 98% como curativo biológico no tratamento de feridas cutâneas provocadas experimentalmente em ratos. Foram utilizados 16 ratos Wistar, nos quais foram induzidas experimentalmente duas lesões cutâneas na região dorsal, sendo uma cranial e a outra caudal. Uma ferida serviu de controle, a qual não recebeu tratamento, a outra recebeu a túnica vaginal recobrindo toda extensão da lesão. A seleção de qual das feridas, cranial ou caudal, iria receber a membrana foi feita de forma aleatória. Os animais foram distribuídos em quatro grupos de igual número para análises clínicas, macroscópicas e histológicas decorridos três, sete, 14 e 21 dias de pós-operatório. Na avaliação clínica e macroscópica pode-se observar que o processo de reparação teve evolução semelhante tanto no grupo controle quanto no grupo membrana. Na análise histopatológica foi constatada grande quantidade de infiltrado inflamatório nos períodos iniciais, no entanto, decorridos 21 dias, a epiderme já se encontrava reparada, e em todos os momentos foi observada maior presença de tecido conjuntivo no grupo membrana, sugerindo maior precocidade na cicatrização. Deste modo, concluiu-se que a túnica vaginal canina pode ser utilizada como nova alternativa no tratamento de feridas cutâneas.

In the search for alternative material in the treatment of cutaneous wounds, this study aimed to evaluate the behavior of canine vaginal tunic conserved in glycerin 98% as a biological dressing in the treatment of cutaneous wounds provoked experimentally in rats. Sixteen animals were used in which two cutaneous lesions were caused in the dorsal region, one cranial and the other caudal. One wound served as a control which did not receive treatment, the other received the vaginal tunic covering all extension of the lesion. The selection of which of the wounds, cranial or caudal, would receive the membrane was made at random. The animals were divided into four groups of equal numbers for clinical, macroscopic, and histological analyzes after three, seven, 14 and 21 postoperative days. In the clinical and macroscopic evaluation, it can be observed that the healing process had similar evolution in both the control group and the membrane group. In the histopathological analysis, a large amount of inflammatory infiltrate was observed in the initial periods; however, after 21 days the epidermis was already healed, and at all times a greater presence of connective tissue was observed in the membrane group, suggesting a greater precocity in healing. Thus, it was concluded that the canine vaginal tunic can be used as a new alternative in the treatment of cutaneous wounds.