Utilización de la herramienta chroma key para el estudio de la morfología y la fisiología espermática en entornos virtuales / Use of the chroma key to study sperm morphology and physiology in virtual environments

RESUMEN: Uno de los retos en el uso de nuevas metodologías y tecnologías durante la crisis sanitaria causada por el virus SARS-CoV-2 ha sido mantener la motivación del alumnado en entornos virtuales. Por ello, el objetivo de este trabajo fue valorar la utilidad de materiales audiovisuales creados con chroma key en la metodología flipped classroom para impartir algunos conceptos teóricos en la asignatura de Biología del Desarrollo en el Grado en Biología de la Universidad de Alicante. Para ello, el profesorado de la asignatura elaboró vídeos utilizando la tecnología chroma key, los cuales fueron visualizados por parte del alumnado antes de las sesiones teóricas online. Durante dichas sesiones, el alumnado puso en práctica los conceptos comentados en los vídeos a través de la realización de actividades. La percepción del estudiantado sobre la metodología empleada se obtuvo mediante un cuestionario de opinión, en el cual el 90 % de los encuestados/as manifestaron que el uso combinado del flipped classroom con chroma key facilitaba el aprendizaje al adaptarse al ritmo y necesidades educativas de cada estudiante. Asimismo, destacaron que el uso de escenografía virtual con chroma key hizo más amena y atrayente la docencia online. En conclusión, el chroma key constituye una herramienta eficaz para realizar materiales educativos en flipped classroom que, además, resulta llamativo y motivador para el alumnado.

SUMMARY: One of the challenges in the use of new methodologies and technologies during the health crisis caused by the SARS-CoV-2 virus has been to keep students motivated in virtual environments. Therefore, the objective of this work was to assess the usefulness of audiovisual materials created with chroma key in the flipped classroom methodology to teach some theoretical concepts in the subject of Developmental Biology in the Degree in Biology at the University of Alicante. For this, the teaching staff of the subject produced videos using chroma key technology, which were viewed by the students before the online theoretical sessions. During these sessions, the students put into practice the concepts discussed in the videos by carrying out activities. The students' perception of the methodology used was obtained through an opinion questionnaire, in which 90 % of the respondents stated that the combined use of the flipped classroom with chroma key facilitated learning by adapting to the rhythm and educational needs of each student. They also highlighted that the use of virtual scenery with chroma key made online teaching more enjoyable and attractive. In conclusion, the chroma key is an effective tool for creating educational materials in the flipped classroom that is also attractive and motivating for students.

Efecto de la suplementación del medio de incubación con extracto de Berberis microphylla (calafate) sobre la funcionalidad espermática en semen bovino descongelado / Effect of supplementation of incubation medium with Berberis microphylla (calafate) extract on sperm functionality in thawed bovine semen

RESUMEN: En el semen criopreservado, los procesos de congelación/descongelación y posterior manipulación, dañan las células espermáticas provocando disminución de la capacidad fecundante de los espermatozoides descongelados. Estos procesos han sido asociados con el estado de estrés oxidativo (EO) inducido por altos niveles de especies reactivas de oxígeno (EROS), causando daño a la función y estructura espermática. Los espermatozoides descongelados pueden ser protegidos de este daño, con la adición de antioxidantes (AO) al medio de incubación. El fruto de Calafate (Berberis microphylla G. Forst.) posee una alta capacidad antioxidante, lo que hace interesante investigar el efecto de sus componentes antioxidantes en estos procesos biotecnológicos especialmente postdescongelación. El objetivo de este estudio fue determinar el efecto de la suplementación de extracto liofilizado de fruto de Calafate (ELC), sobre la calidad espermática post-descongelación. Previamente se caracterizó el ELC, determinando la actividad antioxidante y metabolitos como fenoles y antocianinas; posteriormente, espermatozoides de bovino descongelados fueron incubados en un medio base suplementado con diferentes concentraciones de ELC. Post-incubación se evaluó la motilidad progresiva; la viabilidad e integridad de la membrana plasmática (SYBR14- PI) y acrosomal (FITC-PNA/PI) y la peroxidación lipídica (BODIPY) por citometría de flujo. La caracterización de ELC demostró que tanto la actividad antioxidante como los fenoles y antocianinas incrementan concomitante con el aumento de la concentración de ELC. La adición de ELC al medio de incubación, dependiendo de la concentración y tiempo de incubación, sería eficaz en proteger la motilidad, viabilidad e integridad de la membrana plasmática y disminuir la lipoperoxidación en los espermatozoides de bovino descongelados.

SUMMARY: In cryopreserved semen, the freezing/thawing process following of manipulation, damage the sperm cell, decreasing the fertilizing capacity of the thawed sperm; being one of the main factors of this damage the oxidative stress. The sperm once thawed can be protected from this damage, with the addition of antioxidants to the incubation medium. The Calafate fruit (Berberis microphylla G. Forst.) has a high antioxidant capacity, making it an interesting resource for investigating the effect of its antioxidant components on biotechnological processes. The objective of this study was to determine the effect of supplementation of Calafate fruit lyophilized extract (ELC) on sperm quality. The lyophilized extract of the Calafate fruit was characterized, determining the antioxidant activity and metabolites such as phenols and anthocyanins; subsequently, thawed bovine sperm were incubated in a medium supplemented with different concentrations of ELC. Post-incubation, progressive motility was evaluated. By flow cytometry, the viability and integrity of the plasma (SYBR14-PI), and acrosomal (FITC-PNA / PI), as well as lipid peroxidation (BODIPY), was determined. The characterization of Calafate fruits lyophilized extract indicated that antioxidant activity, phenols and anthocyanins increased concomitantly with the increase of dose extract used. The addition of ELC to the incubation medium, depending on the concentration and incubation time, would be effective to protect motility, viability and integrity of the plasma membrane and decreased lipid peroxidation in thawed bovine sperm.

Semen parameters in men recovered from COVID-19 / 亚洲男科学杂志(英文版)

The novel coronavirus disease (COVID-19) pandemic is emerging as a global health threat and shows a higher risk for men than women. Thus far, the studies on andrological consequences of COVID-19 are limited. To ascertain the consequences of COVID-19 on sperm parameters after recovery, we recruited 41 reproductive-aged male patients who had recovered from COVID-19, and analyzed their semen parameters and serum sex hormones at a median time of 56 days after hospital discharge. For longitudinal analysis, a second sampling was obtained from 22 of the 41 patients after a median time interval of 29 days from first sampling. Compared with controls who had not suffered from COVID-19, the total sperm count, sperm concentration, and percentages of motile and progressively motile spermatozoa in the patients were significantly lower at first sampling, while sperm vitality and morphology were not affected. The total sperm count, sperm concentration, and number of motile spermatozoa per ejaculate were significantly increased and the percentage of morphologically abnormal sperm was reduced at the second sampling compared with those at first in the 22 patients examined. Though there were higher prolactin and lower progesterone levels in patients at first sampling than those in controls, no significant alterations were detected for any sex hormones examined over time following COVID-19 recovery in the 22 patients. Although it should be interpreted carefully, these findings indicate an adverse but potentially reversible consequence of COVID-19 on sperm quality.

Efecto de la borra de café sobre la biológica espermática: aproximación in vitro / Effect of spent coffee grounds on sperm biology: in vitro approach

OBJETIVO: El objetivo de este trabajo fue establecer el efecto de la borra de café sobre la movilidad y los parámetros funcionales de los espermatozoides humanos in vitro. MATERIALES Y MÉTODOS: La borra de café, un subproducto obtenido en establecimientos especializados en la preparación de café soluble a base de grano, se diluyo en tampón fosfato salino y se mezcló en proporciones iguales con las muestras de semen de 16 voluntarios aparentemente sanos. A cada muestra se le determinó el efecto sobre la movilidad espermática en función del tiempo (30, 60, 90 y 120 minutos, n=16) y sobre los parámetros funcionales (n=6) por medio de citometría de flujo: potencial de membrana mitocondrial, producción de especies reactivas de oxígeno y lipoperoxidación de la membrana espermática. RESULTADOS: La incubación de los espermatozoides con la borra de café evidencio un cambio positivo en la movilidad espermática. Adicionalmente, la incubación con la borra de café incremento significativamente el potencial de membrana mitocondrial en los espermatozoides. CONCLUSIÓN: La borra de café, seguramente debido a los compuestos antioxidantes, afecta positivamente la movilidad espermática aumentando el potencial de membrana mitocondrial. Por lo tanto, esto es un paso inicial en la búsqueda de un suplemento de origen natural que aumente la calidad seminal.

OBJECTIVE: The objective of this work is to establish the effect of spent coffee grounds on the motility and functional parameters of human spermatozoa, in vitro. MATERIALS AND METHODS: Spent coffee grounds, a by-product obtained in specialized establishments in the preparation of soluble coffee based on grain, was diluted in saline phosphate buffer and mixed in equal proportions with semen samples from 16 apparently healthy volunteers. Each sample was determined the effect on sperm motility as a function of time (30, 60, 90 and 120 minutes, n=16) and on functional parameters (n=6) by means of flow cytometry: mitochondrial membrane potential, reactive oxygen species production and membrane lipoperoxidation. RESULTS: The incubation of the spermatozoa with the spent coffee grounds showed a positive change in sperm motility. Additionally, incubation with spent coffee grounds significantly increased the mitochondrial membrane potential in human sperm cells. CONCLUSION: Spent coffee grounds, probably due to antioxidant compounds, positively affects sperm motility by increasing mitochondrial membrane potential. Therefore, this is an initial step in the search for a supplement of natural origin that increases seminal quality.

Quantitative and qualitative variables of semen from surubim do Iguaçu, Steindachneridion melanodermatum Garavello, 2005 (Siluriformes: Pimelodidae) / Variáveis quantitativas e qualitativas de sêmen de Surubim do Iguaçu Steindachneridion melanodermatum Garavello, 2005 (Siluriformes: Pimelodidae)

Abstract The objective of the present work was to assess the qualitative and quantitative characteristics of semen from the surubim do Iguaçu (Steindachneridion melanodermatum). Induced spermiation was achieved in eleven males with mean weight of 1.76 ± 0.48 kg and average age of two years and semen was collected by stripping. The average volume was 1.34 ± 0.73 mL. The duration of sperm motility was 154.4 ± 72.6 and 149.0 ± 77.5 seconds after activation with hatchery water and distilled water, respectively. The sperm concentration estimated by hemocytometer was 5.423 ± 2.155 x 1010 spermatozoa/mL. The results indicate that S. melanodermatum semen is easily obtained during the spawning season and the seminal characteristics are adequate insemination and subsequent in vitro fertilization.

Resumo O objetivo do presente trabalho foi avaliar características qualitativas e quantitativas do sêmen de surubim do Iguaçu (Steindachneridion melanodermatum). Para as análises, utilizaram-se onze machos com peso médio de 1,76±0,48 kg e idade média de dois anos, submetidos ao tratamento de hipofisação. O volume médio de sêmen coletado foi de 1,34 ± 0,73 mL. O tempo médio de motilidade dos espermatozoides após a coleta foi de 154,4 ± 72,6 e 149,0 ± 77,5 segundos para ativação com água do tanque e água destilada, respectivamente. Para a concentração espermática, a média foi de 5,423 ± 2,155 x1010 espermatozoides/mL. Os resultados indicam que o sêmen de S. melanodermatum é facilmente obtido no período reprodutivo analisado, e que as características seminais avaliadas são adequadas para reprodução induzida.

Semen bovino sexado congelado-descongelado en producción de embriones in vitro / In vitro embryo production with frozen-thawed bovine semen sexed

El objetivo de este trabajo fue evaluar la fertilidad in vitro del semen bovino sexado (SX) vs. no sexado (NS) congelado-descongelado de dos toros Holstein, cada uno de la misma partida. Determinar el sexo de las crías significa un avance importante para la producción. El citómetro de flujo separa los espermatozoides X e Y por diferencia de ADN (4 % mayor en X), con 90 % de efectividad. Los complejos ovocito-cúmulus (COC) se obtuvieron de folículos de 2 a 8 mm de ovarios de frigorífico, se cultivaron para maduración 22 h en TCM-199 + 5 % de SFB + 10 % licor folicular bovino (LFB), en gotas de 100 µl, cubiertos con aceite mineral, en incubadora (38,5 C, 5 % CO2 y 95 % de humedad). Posmaduración, se formaron al azar 4 grupos de COC los cuales fueron inseminados con NS y SX de los toros 1 y 2. Los COC se incluyeron en gotas de 100 ml a razón de 10 COC por gota de semen capacitado a una concentración de 2x106 espermatozoides/ml en todos los grupos, incubados durante 6 h. Posteriormente se cultivaron en CR1aa + 5 % SFB, en incubadora. A las 48 h se evaluó el clivaje y al día 7 el desarrollo embrionario. Los resultados fueron analizados con el Test de c2. Se encontró diferencias significativas en el toro 1 en el desarrollo embrionario a favor del NS (p<0,05). En el toro 2 no se encontró diferencias significativas en el clivaje ni en el desarrollo (p<0,05).

The objective of this study was to evaluate the in vitro fertility of sexed (SX) vs. non-sexed (NS), frozen-thawed bovine semen from two Holstein bull, from the same batch each one. Offspring sexing represents an important advance for livestock production. Flow cytometry separates X and Y spermatozoa by difference in DNA (4 % greater in X) with 90 % effectiveness. Cumulus-oocytes complexes (COC) were obtained from follicles measuring between 2 and 8 mm collected from slaughterhouse ovaries; they were then cultured 22 h for maturation in TCM-199 + 5 % BFS + 10 % bovine follicular fluid (BFF) in 100 µl drops with mineral oil, in incubator (38.5 C, 5% CO2 and 95 % humidity). Postmaturation, 4 groups were randomly formed and inseminated with NS and SX of the 1 and 2 bulls, including them in 100 µl drops at 10 COC per drop of capacitated semen diluted to a concentration of 2x106 sperms/ml in all groups, incubated during 6 h. They were then cultured in CR1aa + 5% BFS in an incubator. At 48 h cleavage and at day 7 embryonic development, were assessed. Results were analyzed with c2 square Test. There were significant differences (P<0.05) in the embryonic development in bull 1, grater in NS. In bull 2 there were not significant differences in cleavage neither in embryo development.

A bull breeding soundness evaluation system with greater emphasis on scrotal circumference

Bull breeding soundness evaluation (BBSE) is a method applied to reduce the risk of using subfertile bulls in herds. There are currently two BBSE systems, those of the Society for Theriogenology (SFT) and the Western Canadian Association of Bovine Practitioners (WCABP). Scrotal circumference (SC), sperm motility (SM) and normal sperm (NS) of 454 bulls aged between 12 and 15 months of a Spanish beef breed were used to compare both systems, and since there is no agreement on that BBSE system must be applied in Spain, a single one was proposed for its consideration. SC was adjusted to 15 months (SC15) and the mean of the BBSE traits was: SC15 (34.2±2.4cm), SM (76.6±14.6%) and NS (76.8±12.3%). In the PROPOSED system, the SM and NS thresholds were those defined by the WCABP system, while the SC15 thresholds were set by combining the SFT threshold and SC15±1SD in order to establish four classification categories, the three proposed by the WCABP system: unsatisfactory, questionable and satisfactory, and other category, called superior, for bulls with SM≥60%, NS≥70% and SC15≥Mean+1SD. The PROPOSED system scored fewer bulls as unsatisfactory than the SFT and the WCABP systems (8.6%, 23.6% and 22.5%, respectively; P<0.01), while the percentage of bulls from worst to best in the other three categories under the PROPOSED system was: 26.0%, 54.2% and 11.2%, respectively. In conclusion, the PROPOSED system gives more emphasis to SC, sets differences between bulls classified as satisfactory by the other systems and can be considered a good system for Spain and for other countries that have no defined their own system...

Validación de sybr-14 y 6-cfda para evaluar la viabilidad e integridad de la membrana plasmática en espermatozoides caninos de raza chihuahua / Validation of sybr-14 and 6-cfda to evaluate the viability and plasma membrane integrity in sperm of chihuahua canine breed

En la aplicación de técnicas reproductivas es importante determinar in vitro la capacidad fecundante de los espermatozoides, para ello se utilizan combinaciones de tinciones para evaluar los diferentes parámetros de función espermática, aumentando así la precisión de la estimación de la muestra. El objetivo del estudio fue comparar la efectividad de la utilización de los fluorocromos 6-CFDA y SYBR-14 combinados con PI para determinar la viabilidad e integridad de la membrana plasmática por citometría de flujo. Se utilizó semen fresco de caninos (n=5) de raza Chihuahua, con una concentración espermática >150x106 esp/ml y motilidad progresiva >80%. Tres protocolos fueron ensayados: grupo 1: SYBR-14/PI, grupo 2: 6-CFDA/PI y grupo 3: PI. La integridad de la membrana plasmática de los espermatozoides fue similar entre grupos 1 y 2, independiente del fluorocromo utilizado (37,26±13,9 y 33,8±14,6, respectivamente; p=0,4601). Asimismo, la viabilidad espermática entre los grupos 1, 2 y 3 (62,7±13,9, 66,1±14,6 y 66,4±13,3, respectivamente; p=0,8987). En conclusión, no se evidenció diferencias en la efectividad para determinar la viabilidad e integridad de la membrana plasmática mediante la utilización de SYBR-14 y 6-CFDA, ambas tinciones pueden ser incorporadas al análisis de rutina de semen canino de raza Chihuahua.

In applying reproductive techniques in vitro it is important to determine the fertilizing capacity of the sperm, for this a combination of dyes were used to assess different parameters of sperm function, thereby increasing the accuracy of the estimation of the sample. In dogs (Canis lupus familiaris) Chihuahua breed there is no precedent for evaluating sperm function parameters. The aim was to assess the viability and plasmatic membrane integrity, basic parameters of sperm function. Propidium iodide (PI) was used, a fluorescent dye-specific DNA, which combined with fluorochromes permeable acts as marker of the sperm membrane integrity. The effectiveness of the use of 6-CFDA and SYBR-14 fluorochromes combined with PI was also compared to determine viability and sperm membrane integrity using flow cytometry. Fresh semen of dogs (n=5) Chihuahua breed was used with a concentration of >200x106 sp/ml and progressive motility >80%. Three protocols were performed: group 1: SYBR-14/PI, group 2: 6-CFDA/PI and group 3: PI. The plasma membrane integrity of sperm was similar, independent of the fluorophore used between groups 1 and 2 (13.9±37.26 and 33.8±14.6, respectively, p=0.4601). This also applied to sperm viability between groups 1, 2 and 3 (62.7±13.9, 66.1±14.6 and 66.4±13.3, respectively, p=0.8987). No difference was demonstrated in effectiveness to determine the viability and integrity of the sperm membrane using SYBR-14 and 6-CFDA, both dyes can be incorporated in to routine analysis of semen in canine Chihuahua breed.

Reproduction of the Brazilian snapper, Lutjanus alexandrei Moura & Lindeman, 2007 (Perciformes: Lutjanidae), off the northern coast of Pernambuco, Brazil

Reproductive aspects of the Brazilian snapper Lutjanus alexandrei, were characterized, including a description of the development of oocytes and spermatogenic cells, size at first sexual maturity, and fecundity. A total of 540 fish were analyzed with 250 having their gonads sectioned to allow microscopic evaluation. Six maturity stages were identified for females and males: immature, maturing, mature, spawning, spawned, and resting. Fish standard length (SL) varied from 13.0 to 28.3 cm and sex ratio was 1.6 males: 1.0 females. Monthly distributions of mean Gonadosomatic Index (GSI) and maturity stages suggest that spawning occurs mainly in a protracted period, during the warmer months, from November to March. The size of first sexual maturity was estimated at 17.1 cm SL for females and 16.8 cm SL for males. Oocyte development suggests that L. alexandrei exhibits a multiple batch spawning behavior and batch fecundity varied from 34,000 to 324,000 oocytes.

Os aspectos reprodutivos da baúna-de-fogo Lutjanus alexandrei foram caracterizados, incluindo a descrição do desenvolvimento dos ovócitos e células espermatogênicas, do tamanho de primeira maturação sexual, e da fecundidade. Um total de 540 peixes foi analisado, dos quais 250 tiveram as suas gônadas seccionadas para avaliação microscópica. Seis estágios de maturidade sexual foram determinados para fêmeas e machos: imaturo, em maturação, maduro, desovando, desovado e repouso. O comprimento padrão (CP) dos peixes variou de 13,0 a 28,3 cm e a proporção sexual foi de 1,6 machos: 1,0 fêmeas. As distribuições mensais dos valores médios do Índice Gonadosomático (IGS) e dos estágios de maturidade sexual sugerem a ocorrência de desovas em um período prolongado, principalmente nos meses de temperaturas mais quentes, entre novembro e março. O tamanho médio de primeira maturação sexual foi estimado em 17,1 cm CP para as fêmeas e 16,9 cm CP para os machos. O padrão de desenvolvimento dos ovócitos sugere que L. alexandrei exibe comportamento de múltiplas desovas por lote, e a fecundidade variou entre 34.000 a 324.000 ovócitos.

Age-related changes in quality and fertility of porcine semen

The aim of this study was to investigate the effect of boar age on quality traits and fertility of liquid-stored semen. Boars were allocated into 3 age groups: 7-10 months (young), 18-33 months (mature), 51-61 months (old). Ejaculates of > 200x10(6) sperm/ml and 85% total motile sperm were extended to 30x10(6) sperm/ml, stored at 17-18 °C and used within 12-24 h for artificial insemination (AI) of 2062 multiparous sows. After 24 h of storage, aliquots of diluted semen were assessed for sperm progressive motility (SPM), incidence of sperm chromatin instability (SCI), proportion of live morphologically normal sperm (LMNS) and head morphometry of LMNS. The results showed that young boars had higher percentages of SCI and lower proportions of LMNS than those of the mature (p < 0.05) and old (p < 0.001) boars, respectively. Sperm head dimensions of young and old boars were greater (p < 0.03-0.001) than those of mature boars. The farrowing rate of young boars (65%) was significantly lower (p < 0.001; χ2= 30-61) than those of the mature (87.2%) and old (84.7%) boars. The relationship between sperm head dimensions and boar fertility was non-significant. In conclusion, boar age is an important physiological factor contributing to the success of swine AI.

Evaluación y estandarización de la calidad del espermiograma: nuevos límites inferiores de referencia / Evaluation and standardization of the quality of semen: new lower reference limit

El análisis seminal o espermiograma es uno de los parámetros más usados en la evaluación de la fertilidad masculina. La OMS (WHO, 2010), presentó el 5 Manual para el examen y procesamiento del semen humano, documento que fue analizado durante el primer taller de estandarización del análisis seminal (PLEAS), realizado en Santiago de Chile, mayo del 2010. Posteriormente se aplicaron los nuevos valores indicados como "límite de referencia inferior" (LIR), en el estudio del análisis seminal realizados por varios autores (2003 al 2010). Los resultados obtenidos indican que el 81 por ciento de los investigadores latino americanos creen que el nuevo manual estandariza mejor la concentración espermática, un 96 por ciento está de acuerdo con la nueva subclasificación en la motilidad espermática en progresiva (A), no progresiva (B) e inmóviles (C). El 68 por ciento estima que el mejor instrumental de recuento es la cámara de Neubauer. Respecto a los controles de calidad solo el 18 por ciento realiza controles de calidad externa. El 100 por ciento de los investigadores estima conveniente realizar continuos talleres de estandarización. Respecto a la aplicación de los LIR en las poblaciones en estudio, todos ellos cumplirían con los estándares actuales para ser considerada una población con capacidad de fertilidad. Sin embargo varios autores opinan que una nueva versión del manual OMS, debe realizarse urgentemente para estandarizar mejor la concentración espermática (15 millones por mL) y la morfología según criterios estrictos (4 por ciento), valores de referencia que consideran muy bajos.

Spermogram or semen analysis is one of the most used parameters in the evaluation of male fertility. WHO (2010) presented the 5th Manual for review and processing of human semen, a document that was discussed during the first workshop of standardization of semen analysis (PLEAS), held in Santiago de Chile, May 2010. Subsequently applied the new values expressed in "lower referencelimit" (LRL) for semen in several analysis studies conducted by various authors (2003 to 2010). The results indicate that 81 percent of Latin American researchers believe the new manual standardizes best sperm concentration, 96 percent agree with the new subclassification in progressive sperm motility (A), non-progressive (B) and immobile (C). 68 percent determined that the best instruments for the sperm countis the Neubauer haemocytometerchamber. Regarding quality control only 18 percent performed external quality control. 100 percent of researchers believe it is appropriate to conducton going standardization workshops. Regarding the application of LRL in the study populations (2003-2010), 100 percent comply with the standards to be considered a population with fertility capacity. However, several authors argue that a new version of the WHO manual, must be re-done urgently to better standardize sperm concentration (15 million/mL) and morphology according to strict criteria (4 percent), reference values considered very low.

Rat dorsal prostate is necessary for vaginal adhesion of the seminal plug and sperm motility in the uterine horns

The rat prostate comprises dorsal, ventral and lateral lobes that are morphologically and biochemically distinct. Lesions to these structures are expected to affect the quality of the ejaculate and male fertility. In experiment 1, we analyzed ejaculate parameters of males that had chemical lesions of the dorsal or ventral lobes. At pre-lesion and at 5 and 20 days post-lesion males were mated, and after ejaculation, seminal fluid and seminal plug were obtained from the mated females. In experiment 2, the ventral lobes were ablated, and the ejaculate was analyzed. In experiment 3, the fertility of males with chemically-lesioned dorsal lobes or ablation of the ventral lobes was evaluated. Chemical lesion of the dorsal lobe prevented the adhesion of the seminal plug to vaginal walls. When these males were tested at 5-days postlesion, no sperm were found in uterus, and at 20-days post-lesion, the few sperm encountered showed slow progressive motility. None of the females that mated with dorsal lobe-lesioned males became pregnant. However, chemical lesion or ablation of the ventral lobes did not affect ejaculate or fertility. Our results indicate that the dorsal prostatic lobes are indispensable for reproductive success in males, and define parameters of ejaculate with which fertility can be estimated.

Análisis de las variables del espermiograma en jóvenes sanos en Arica-Chile / Spermograms of healthy young subjects living in Arica, Chile

Background: Semen analysis is one of the parameters used to predict male fertility. Semen can be altered by environmental pollutants; therefore it could be used as a biological marker of exposure in contaminated areas. Aim: To analyze the spermogram values in a sample of healthy young males, residing in Arica, Chile. Material and methods: One hundred and two healthy university students volunteers aged 18 to 30 years answered a questionnaire about fertility, habits and andrologic diseases and provided a semen sample. Within three hours after ejaculation, semen volume, pH, sperm concentration, motility and morphology were analyzed. Results: Six percent of volunteers had offspring, 1 percent declared to be infertile, 32 percent smoked and 78 percent con-sumed alcohol. Semen pH was 7.6 ± 0.5, volume, 2.9 ± 1.6 ml, sperm concentration, 62.8 ± 62.3 x 10(6)/ml, normal morphology, 15.0 ± 7.9 percent, overall motility, 42.2 ± 23.2 percent and grade A motility, 19.2 ± 18.6 percent. The percentage of subjects that had normal semen values was 82 percent for total sperm count, 76 percent for sperm concentration, 72 percent for volume, 64 percent for vitality, 63 percent for pH, 57 percent for morphology, 38 percent for overall motility and 26 percent for grade A motility. Conclusions: This sample of healthy young males had a normal sperm count in comparison with international reports. However it is necessary to characterize the spermogram in uncontaminated areas of Chile to ensure that our results are within the expected values for the country.

Nuevos conceptos en la transmisión sexual del HIV / New advances on sexual transmission of HIV

La transmisión sexual es la vía más importante de diseminación de la infección por HIV-1. Para lograr transmitirse por esta vía, el virus debe atravesar la barrera epitelial y alcanzar a las células blanco de la infección en el subepitelio: los linfocitos T CD4, los macrófagos y las células dendríticas. Los mecanismos subyacentes a la transmisión no son aún comprendidos con claridad. En los últimos años se ha resaltado la importancia de la interacción entre el HIV y las células dendríticas durante la transmisión y en la diseminación inicial del virus en la mucosa. El HIV, mediante complejos mecanismos, utiliza a las células dendríticas tanto como para favorecer su diseminación, como también para impedir la apropiada activación de la respuesta inmune adaptativa. Recientes hallazgos demuestran que durante la transmisión sexual, diferentes componentes del semen son capaces de modular esta interacción entre las células dendríticas y el HIV.

Unprotected sexual intercourse between discordant couples is by far the most common mode of human immunodeficiency virus type 1 (HIV-1) transmission. After deposition of HIV-1 on the recipient mucosa, infectious virus must cross the mucosal epithelium and interact with T CD4 lymphocytes, macrophages, and dendritic cells (DCs), which are the initial targets of infection. The mechanisms through which HIV-1 cross the mucosal epithelium and gain access to target cells are still not well characterized. The capacity of HIV-1 to hijack DCs appears to be crucial for viral transmission and early HIV-1 pathogenesis. HIV-1 subverts DC function favouring local dissemination of the virus and also interfering with the development of adaptive immune response. Recent findings show that semen modulates the interaction between dendritic cells and HIV during sexual transmission.

Age-related ultrasonography, cytology, and microbiologic exam of canine prostate / Ultrassonografia, citologia e exame microbiológico da próstata de cães em diferentes idades

The physiological parameters that could be reference for trustful diagnosis and prognosis of prostate disorders in dogs were obtained. Thirty six intact male dogs without clinical signs of neither prostatic nor reproductive disorders were allocated according the age in three groups. These animals were submitted to semen manual collection for microbiological exams; transabdominal ultrasonography to evaluate dimensions, ecogenicity, and texture of prostatic parenchyma and aspirative puncture with fine needle for cytological and microbiological analyses. Ultrasonography revealed that the predominant prostatic shape was round with regular surface. Dimensions varied according to age, being small in young animals and large in old ones. There was a positive correlation between prostatic dimensions and body weight. Microbiological exams detected microorganisms on seminal plasma from 11 dogs and prostate tissue aspirated from 10 animals, although they were healthy. Cytology did not reveal any inflammatory, proliferative, or neoplasic alteration in young and middle age dogs, but in three older dogs signs of hyperplasia/hypertrophy was found. It was observed positive correlation between age and cellular area but a negative correlation was observed between nucleus:cytoplasm ratio and craniocaudal dimension.

Obtiveram-se parâmetros fisiológicos que pudessem ser utilizados como referência para diagnóstico e prognóstico confiáveis de doença prostática em cães. Trinta e seis cães, sem sinais clínicos de doença prostática ou distúrbios reprodutivos, foram distribuídos em três grupos de acordo com a idade.Os animais foram submetidos à colheita manual de sêmen para exames microbiológicos, à ultrassonografia transabdominal, para avaliar as dimensões, a ecogenicidade e a ecotextura prostática, e à punção aspirativa com agulha fina, para análise citológica e microbiológica. A ultrassonografia revelou que a forma predominante da próstata foi globosa, com superfície de contorno regular. As dimensões variaram de acordo com a idade, sendo pequena em animais jovens e grande nos animais idosos. Houve correlação positiva entre as dimensões prostáticas e o peso corporal. Os exames microbiológicos detectaram microrganismos no plasma seminal de 11 cães e no tecido prostático aspirado de 10 animais, embora eles fossem saudáveis. A citologia não revelou nenhuma alteração inflamatória, proliferativa ou neoplásica nos cães jovens e de meia idade, mas, em três cães idosos foram encontrados sinais de hiperplasia/hipertrofia. Foi observada correlação positiva entre a idade e a área celular e correlação negativa entre a relação núcleo:citoplasma e a dimensão craniocaudal.

Características andrológicas e do sêmen de touros do composto Red Norte (Nelore x Tabapuã x Red Angus x Sinepol) / Andrological and semen characteristics of cross-bred Red Norte (Nelore x Tabapuã x Red Angus x Sinepol) young bulls

Avaliaram-se as características andrológicas do sêmen de touros jovens do composto Red Norte (Nelore x Tabapuã x Red Angus x Sinepol), com idade média de 13,9±0,8 meses, com o objetivo de estimar o advento da puberdade e a qualidade do sêmen. Foram avaliados o perímetro escrotal (PE), o peso e as características seminais de 70 tourinhos, classificados em três grupos, de acordo com o PE: GI=27-33cm (n=24), GII=33-35cm (n=24) e GIII=35-43cm (n=22). As médias de peso e a idade de cada grupo (G) foram, respectivamente: GI=411,2±37,4kg e 13,8±1,0 meses, GII=426,9±31,5kg e 14,0±0,7 meses e GIII=438,4±38,3kg e 14,0±0,6 meses. As características seminais para cada grupo foram, volume 4,2±3,1mL, 5,3±2,6mL e 4,5±2,1mL; motilidade 31,3±24,1 por cento, 44,2±23,9 por cento e 43,9±21,5 por cento e vigor 2,8±1,6, 3,5±1,3 e 3,5±1,3, respectivamente. O espermiograma apresentou valores médios de concentração de 130,5±266,2x10(6)/mL, 289,5±390,2x10(6)/mL e 333,9±523,7x10(6)/mL, defeitos totais de 81,4±15,9 por cento, 73,8±15,4 por cento e 67,9±19,0 por cento; defeitos maiores de 87,3±26,2 por cento, 66,8±24,9 por cento e 56,7±17,1 por cento e defeitos menores de 16,6±14,9 por cento, 33,2±24,9 por cento e 43,3±17,1 por cento, respectivamente. Dos setenta animais examinados, sete (10 por cento) foram considerados aptos à reprodução. Os resultados mostraram que a patologia espermática diminuiu em razão do aumento do PE.

Reproductive traits of cross-breed Red Norte (Nelore x Tabapuã x Red Angus x Sinepol) young bulls averaging of 13.9±0.8 month-old were evaluated, in order to determine the puberty onset and semen quality in these animals. Scrotal circumference (SC), body weight (BW), and semen parameters of 70 bulls were measured. Animals were allotted in three groups (G) according to their SC: GI=27-33cm (n=24), GII=33-35cm (n=24), and GIII=35-43cm (n=22). BW and age of each group were, respectively: GI=411.2±37.4kg and 13.8±1.0 month-old, GII=426.9±31.5kg and 14.0±0.7 month-old, and GIII=438.4±38.3kg and 14.0±0.6 month-old. Seminal physical characteristics for same order of groups were: volume 4.2±3.1mL, 5.3±2.6mL, and 4.5±2.1mL; motility 31.3±24.1 percent, 44.2±23.9 percent, and 43.9±21.5 percent; and vigor 2.8±1.6, 3.5±1.3, and 3.5±1.3. The spermiogram presented concentration values of 130.5±266.2x10(6)/mL, 289.5±390.2x10(6)/mL, and 333.9±523.7x10(6)/mL; total defects of 81.4±15.9 percent, 73.8±15.4 percent, and 67.9±19.0 percent; major defects of 87.3±26.2 percent, 66.8±24.9 percent and 56.7±17.1 percent; and minor defects of 16.6±14.9 percent, 33.2±24.9 percent, and 43.3±17.1 percent, for same order of groups. Seven out of 70 bulls were considered satisfactory potential breeders. Results showed that semen pathology progressively decreased when SC increased.

Efeitos do estresse de trabalho sobre parâmetros seminais de cães da raça Rottweiler / Evaluation of work stress effects on Rottweilers male dog´s fertility

O presente trabalho teve como objetivo avaliar os efeitos do estresse de trabalho sobre a fertilidade de cães machos. Foram utilizados 18 cães da raça rottweiler, férteis, com idade média de 4 anos, distribuídos aleatoriamente em dois grupos: repouso (controle) e trabalho (tratamento). O tratamento era composto por 5 fases: 1-adaptação, 2-adestramento básico, 3-adestramento militar e condicionamento, 4-acampamento, 5-repouso. Durante todo o período experimental, foram feitas duas coletas de sêmen semanais, para avaliação do ejaculado. No final de cada fase foi realizada coleta de sangue para dosagem dos níveis plasmáticos de cortisol. Os dados foram analisados pelo SAS, á =5%. Observou-se efeito negativo do estresse sobre os parâmetros motilidade (72,63 vs. 57,62, p<0,0001) e vigor espermático (3,06 vs. 2,52, p<0,0001), porcentagem de defeitos maiores(16,01 vs. 26,80, p< 0,0001) e totais (29,61 vs. 40,34, p<0,0001).Observou-se interação (p<0,0001) entre tempo e tratamento sobre a variável cortisol, indicando que o tipo de agente estressante (fase) interfere no nível plasmático de cortisol. Com base nesses resultados,é possível concluir que o estresse de trabalho interferiu de maneira negativa na fertilidade dos cães.

The present work aimed to evaluate the effect of work stress male dogs fertility. Eighteen fertile male Rottweiler dogs, mean aged 4 years old, were randomly allocate into two groups: resting (control) and working (treatment). Treatment consisted of 5 working stages: 1– adaptation, 2- basic obedience training, 3 - physical conditioning and military training, 4 – camping and 5 – resting. During the experimental period, semen was collected and evaluated twice a week.At the end of each stage, blood collections were performed in order to evaluate plasmatic levels of cortisol. Data were analyzed using the statistical software SAS (a=5%). A significant negative effect of stress was observed on sperm motility (resting=72.63 vs. working=57.62,p<0.001), progressive motility (resting=3.06 vs. working=2.52, p<0.0001), major defects (resting=16.01 vs. working=26.80,p<0.0001) and total defects (resting=29.61 vs. working=40.34,p<0.0001). Furthermore, a significant interaction (P<0.0001) was observed between treatment and working stage periods for the plasmatic levels of cortisol, suggesting that the level of stress may vary according to the type of work. Results indicated that work stress may negatively influence fertility in male dogs.

Interferência da idade sobre a qualidade seminal / Interference of age on semen quality

OBJETIVO: avaliar a influência da idade sobre a qualidade seminal de homens com quadro de infertilidade conjugal submetidos a análises espermáticas em um serviço de reprodução humana. MÉTODOS: estudo retrospectivo, no qual foram avaliados os espermogramas de todos os homens em processo de investigação para infertilidade conjugal no período de Setembro de 2002 a Dezembro de 2004, em um serviço de reprodução assistida do nordeste do Brasil. Foram incluídos 531 indivíduos submetidos a 531 avaliações espermáticas. Foram analisados os seguintes parâmetros: volume, concentração, motilidade e morfologia espermática. O total de investigados foi dividido em grupos, de acordo com resultados obtidos de cada variável estudada. Os grupos referentes ao volume seminal foram: hipoespermia, normoespermia e hiperespermia. Os grupos referentes à concentração espermática foram: azoospermia, oligoospermia, normospermia e poliespermia. Os grupos referentes à motilidade foram: motilidade normal e astenospermia. Os grupos referentes à morfologia foram: morfologia normal e teratospermia. As médias das idades dos pacientes entre os grupos com parâmetros normais foram comparadas com as de grupos alterados, sendo utilizado o teste t. Para análise estatística, foi utilizado o programa XLSTAT (p<0,05). RESULTADOS: os indivíduos estudados tinham média de idade de 37±7,9 anos, com um volume seminal médio de 3±1,4 mL, uma concentração espermática de 61,4±66,4 espermatozóides por mL de sêmen, motilidade progressiva de 44,7±19,4% do total de espermatozóides e morfologia normal de 11,2±6,6% dos espermatozóides. As médias de idade entre os grupos foram semelhantes, exceto a média dos indivíduos com hipoespermia, que foi significativamente maior que a dos homens com normoespermia (39,6±10,3 versus 36,5±7,3, p=0,001). CONCLUSÕES: a idade interfere, de forma inversamente proporcional, sobre o volume do ejaculado, porém não exerce influência na concentração, motilidade e morfologia espermática.

PURPOSE: to evaluate the influence of age on the quality of semen in men submitted to spermatic analysis in a human reproduction service, in cases of conjugal infertility. METHODS: a retrospective study in which the spermiograms of all men in process of investigation for conjugal infertility in a service of assisted reproduction in the Northeast of Brazil were evaluated from September 2002 to December 2004. A number of 531 individuals submitted to 531 spermatic evaluations were included in the study. The following parameters have been analyzed: spermatic volume, concentration, motility and morphology. The men under investigation have been divided in groups, according to the results obtained in each of the variables studied. Seminal volume groups were divided in: hypospermia, normospermia and hyperspermia. Spermatic concentration groups were divided in: azoospermia, oligospermia, normospermia and polyspermia. Motility groups were divided in: normal motility and asthenospermia. Morphology groups were divided in: normal morphology and teratospermia. The t test has been used to compare the average age of patients in groups with normal and in groups with altered parameters. The program XLSTAT (p<0.05) has been used for the statistical analysis. RESULTS: the individuals studied presented an average of 37±7.9 years old, with an average of seminal volume of 3±1.4 mL, a spermatic concentration of 61.4±66.4 spermatozoids by mL of semen, a progressive motility of 44.7±19.4% of the total of spermatozoids and normal morphology of 11.2±6.6% of the spermatozoids. Average age among groups were similar, except for that of individuals with hypospermia, which was significantly higher than the one from men with normospermia (39.6±10.3 versus 36.5±7.3, p=0.001). CONCLUSIONS: age interferes in an inversely proportional way on the ejaculated volume, but does not influence spermatic concentration, motility and morphology.

Qualitative parameters of the piapara semen (Leporinus elongatus Valenciennes, 1850) / Parâmetros qualitativos do sêmen de piapara (Leporinus elongatus Valenciennes, 1850)

Qualitative parameters of piapara semen (Leporinus elongatus) were evaluated before and after hormonal induction with carp pituitary extract at 2.5 mg.kg-1 of live weight. The progressive motility, the spermatic vigor and the lifetime of the spermatozoa were higher before the hormonal induction (P > 0.05). The percentage of normal spermatozoa and spermatozoa with secondary pathologies did not differ (P > 0.05) between treatments: before induction (44.0 and 44.4 percent, respectively) and after-induction (44.3 and 46.7 percent, respectively). However, the percentage of primary pathologies was higher (P < 0.05) for the semen collected before induction than for the semen collected after induction; the estimates were 12.2 and 8.0 percent, respectively. The most frequent pathologies were the taillessness with the frequencies of 27.4 and 36.3 percent followed by the headlessness for which the estimates were 10.1 and 3.9 percent, before and after induction respectively. The semen collected before the hormonal induction presented better qualitative parameters.

Os parâmetros qualitativos do sêmen de piaparas (Leporinus elongatus) foram avaliados antes e após a indução hormonal com o extrato de hipófise de carpa na dosagem 2,5 mg.kg-1 de peso vivo. A motilidade progressiva, o vigor espermático e o tempo de vida dos espermatozóides apresentaram valores superiores (P < 0,05) no sêmen coletado antes da indução hormonal. Já a estimativa da porcentagem de espermatozóides normais ou com patologias leves, na pré-indução (44,0 e 44,4 por cento, respectivamente) e na pós-indução hormonal (44,3 e 46,7 por cento, respectivamente) não diferiram (P > 0,05) entre os tratamentos. Porém a estimativa de patologias graves foi maior (P < 0,05) no sêmen de pré-indução do que no pós-induzido com 12,2 e 8 por cento, respectivamente. A patologia mais freqüente foi cabeça solta com 27,4 e 36,3 por cento seguida por cauda solta com 10,1 e 3,9 por cento, antes e após a indução hormonal respectivamente. De acordo com os resultados, o sêmen de piapara coletado na pré-indução hormonal apresentou os melhores parâmetros qualitativos.