Bioengenharia: transplante de células-tronco espermatogoniais em mamíferos / Bioengineering: spermatogonial stem cells transplantation in mammals

O transplante de espermatogônias tronco (SSCs, do inglês Spermatogonial Stem Cell) é uma biotecnologia que consiste na transferência de células tronco testiculares de um doador fértil para um receptor cuja espermatogênese endógena foi suprimida. Essa técnica pode ser aplicada para a produção de machos que gerem uma progênie com características genotípicas do doador selecionado. Especialmente na bovinocultura, tanto de leite como de corte, o transplante de SSCs tem o potencial de substituir a inseminação artificial (IA). Pode-se também, colocar SSCs de um mesmo doador (de genética superior) em mais de um receptor o que aumentaria o número de filhos desse doador. Além disso, possui outras aplicações como a restauração da fertilidade em homens após o tratamento de câncer, conservação de espécies ameaçadas de extinção e tratamento de causas específicas de infertilidade. Assim, com esta revisão tem-se como propósito discorrer acerca de uma biotecnologia da reprodução que permitirá a propagação do valor genético de doadores de sêmen considerados de alto valor zootécnico.

Spermatogonial Stem Cell (SSCs) transplantation is a biotechnology that consists in the transfer of testicular stem cells from a fertile donor to a recipient whose endogenous spermatogenesis has been depleted. This technique can be applied to the production of males that generate a progeny with genotypic characteristics of the selected donor. Especially in beef and dairy cattle, SSCs transplantation has the potential to replace artificial insemination (AI). In addition, it has other applications such as restoring fertility in human species after cancer treatment, conserving endangered species and treating specific causes of infertility. Thus, this aim of this review is to discuss the perspectives of reproductive biotechnology that allows the propagation of the genetic material of high pedigree males.

The role of Rho-associated kinase inhibitor, Y-27632 on primary culture of ovine spermatogonial stem cells

The access to sufficient numbers of spermatogonial stem cells (SSCs) is a prerequisite for the study of their regulation and further biomanipulation. Rho kinase (ROCK) belongs to a family of serine/threonine kinases and involves in a wide range of fundamental cellular functions. The aim of the present study was to study the effect of ROCK inhibitor, Y-27632 (0.1-40 µM), during the primary culture of ovine SSCs. SSCs were collected from 3-5-month-olds lamb testes. The viability of SSCs, the apoptosis assay of SSCs, the intracellular reactive oxygen species (ROS) analysis, and the SSCs markers and apoptosis-related gene expressions were detected by MTT reduction assay, Annexin V–FITC/ Propidium Iodide (PI) dual staining, flow cytometry and real-time-PCR studies, respectively. Morphological analyses indicated that the 5-10 µM Y-27632 had an optimal effect on the number of presumptive SSCs colonies and the area covered by them after a 10 days culture. The cell viability, apoptosis and necrosis of SSCs after 10 days culture were not affected in comparison with the control group, and the 20 µM of Y-27632 resulted in significantly decreased cell viability (P<0.05) and an increased necrosis of cells. On day 10 after culture, the expression of P53 was decreased with an increase from 0 to 10 µM in the Y-27632 dose. In the 20 µM Y-27632 group, the expressions of P53 and Bax were higher and the Bcl-2 was lower than other groups and these values were significantly different from 5 and 10 µM Y-27632 groups (P<0.05). The level of intracellular ROS was decreased with an increase in the Y-27632 dose from 5 to 20 µM in comparison with the control group. In conclusion, the present study demonstrated that Y-27632 at a concentration of 5-10 µM provided optimal culture conditions for the primary culture of ovine SSCs.(AU)

Bioengenharia: transplante de células-tronco espermatogoniais em mamíferos / Bioengineering: spermatogonial stem cells transplantation in mammals

O transplante de espermatogônias tronco (SSCs, do inglês Spermatogonial Stem Cell) é uma biotecnologia que consiste na transferência de células tronco testiculares de um doador fértil para um receptor cuja espermatogênese endógena foi suprimida. Essa técnica pode ser aplicada para a produção de machos que gerem uma progênie com características genotípicas do doador selecionado. Especialmente na bovinocultura, tanto de leite como de corte, o transplante de SSCs tem o potencial de substituir a inseminação artificial (IA). Pode-se também, colocar SSCs de um mesmo doador (de genética superior) em mais de um receptor o que aumentaria o número de filhos desse doador. Além disso, possui outras aplicações como a restauração da fertilidade em homens após o tratamento de câncer, conservação de espécies ameaçadas de extinção e tratamento de causas específicas de infertilidade. Assim, com esta revisão tem-se como propósito discorrer acerca de uma biotecnologia da reprodução que permitirá a propagação do valor genético de doadores de sêmen considerados de alto valor zootécnico.(AU)

Spermatogonial Stem Cell (SSCs) transplantation is a biotechnology that consists in the transfer of testicular stem cells from a fertile donor to a recipient whose endogenous spermatogenesis has been depleted. This technique can be applied to the production of males that generate a progeny with genotypic characteristics of the selected donor. Especially in beef and dairy cattle, SSCs transplantation has the potential to replace artificial insemination (AI). In addition, it has other applications such as restoring fertility in human species after cancer treatment, conserving endangered species and treating specific causes of infertility. Thus, this aim of this review is to discuss the perspectives of reproductive biotechnology that allows the propagation of the genetic material of high pedigree males.(AU)

Diets for Macrobrachium acanthurus breeders: Sperm evaluation / Dietas para reprodutores de Macrobrachium acanthurus: avaliação espermática

The application of biotechnology in animal reproduction has enabled the production of young forms in both quantity and quality. Increasing the number of viable gametes produced by reproducers, among other factors, through an ideal diet, can ensure higher production. Therefore, the aim of this study was to determine the influence of three diets on the sperm survival of Macrobrachium acanthurus. To this end, 24 M. acanthurus males were used, distributed randomly and equally among treatments. Their diets were composed of 100% fresh food (fish and squid muscle - 14% protein), 100% dry feed (commercial feed - 50% protein) and a mixture of these diets containing 30% protein. Spermatophores were extracted through electrical stimulation every 15 days, and the controls consisted of spermatophores obtained directly from nature. No significant difference between diets was observed comparing shrimp and spermatophore weights. The 100% fresh diet provided the best sperm survival performance.

A aplicação de biotecnologias na reprodução animal tem possibilitado produzir formas jovens em quantidade e qualidade. O aumento da quantidade de gametas viáveis produzidos pelos reprodutores, mediante uma dieta ideal, entre outros fatores, pode garantir uma maior produção. Assim, o objetivo deste trabalho foi determinar o efeito de três dietas quanto à sobrevivência espermática de Macrobrachium acanthurus. Para tanto, 24 machos de M. acanthurus foram utilizados, sendo distribuídos ao acaso e de forma igual entre os tratamentos. As dietas foram compostas por 100% de alimento fresco (músculo de peixe e lula - 14% de proteína), 100% de alimento seco (ração comercial - 50% de proteína) e uma mescla dessas dietas contendo 30% de proteína. Os espermatóforos foram extraídos por eletroestimulação a cada 15 dias, sendo o controle aqueles obtidos diretamente da natureza. Não houve diferença significativa entre as dietas quando comparado os pesos dos camarões e dos espermatóforos. A alimentação 100% fresca proporcionou o melhor desempenho para a sobrevivência espermática.

Diets for Macrobrachium acanthurus breeders: Sperm evaluation / Dietas para reprodutores de Macrobrachium acanthurus: avaliação espermática

The application of biotechnology in animal reproduction has enabled the production of young forms in both quantity and quality. Increasing the number of viable gametes produced by reproducers, among other factors, through an ideal diet, can ensure higher production. Therefore, the aim of this study was to determine the influence of three diets on the sperm survival of Macrobrachium acanthurus. To this end, 24 M. acanthurus males were used, distributed randomly and equally among treatments. Their diets were composed of 100% fresh food (fish and squid muscle - 14% protein), 100% dry feed (commercial feed - 50% protein) and a mixture of these diets containing 30% protein. Spermatophores were extracted through electrical stimulation every 15 days, and the controls consisted of spermatophores obtained directly from nature. No significant difference between diets was observed comparing shrimp and spermatophore weights. The 100% fresh diet provided the best sperm survival performance.(AU)

A aplicação de biotecnologias na reprodução animal tem possibilitado produzir formas jovens em quantidade e qualidade. O aumento da quantidade de gametas viáveis produzidos pelos reprodutores, mediante uma dieta ideal, entre outros fatores, pode garantir uma maior produção. Assim, o objetivo deste trabalho foi determinar o efeito de três dietas quanto à sobrevivência espermática de Macrobrachium acanthurus. Para tanto, 24 machos de M. acanthurus foram utilizados, sendo distribuídos ao acaso e de forma igual entre os tratamentos. As dietas foram compostas por 100% de alimento fresco (músculo de peixe e lula - 14% de proteína), 100% de alimento seco (ração comercial - 50% de proteína) e uma mescla dessas dietas contendo 30% de proteína. Os espermatóforos foram extraídos por eletroestimulação a cada 15 dias, sendo o controle aqueles obtidos diretamente da natureza. Não houve diferença significativa entre as dietas quando comparado os pesos dos camarões e dos espermatóforos. A alimentação 100% fresca proporcionou o melhor desempenho para a sobrevivência espermática.(AU)

Biotic potential and reproductive parameters of Spodoptera cosmioides (Walker) (Lepidoptera: Noctuidae) in the laboratory / Potencial biótico e parâmetros reprodutivos de Spodoptera cosmioides (Walker) (Lepidoptera: Noctuidae) em laboratório

In this study we evaluate the biotic potential and reproductive parameters of Spodoptera cosmioides (Walker, 1858) under controlled conditions (25 ± 1 °C, 70 ± 10% RH and 14 hour photophase). Females, on average, (14.433 days) did not live significantly longer than their male counterparts (13.100 days). The mean durations of the pre-oviposition, oviposition and post-oviposition periods were 2.767, 10.600 and 1.067 days, respectively. The mean fecundity of females was 5,073.533 eggs and the mean fertility was 5,021.027 larvae. On average, females copulated 1.167 times. Fecundity was positively correlated with the number of copulations (r = 0.583, P 0.001). The number of copulations, however, was negatively correlated whit the duration of the pre-oviposition (r = -0.560, P = 0.007) and oviposition (r = -0.479, P = 0.048) periods, and overall longevity (r = -0.512, P = 0.031). The biotic potential was estimated at 6.547 x 1021 individuals/female/year. The net reproductive rate (Ro) was 2,193.722 times per generation and the mean generation time (T) was 46.407 days. The intrinsic rate of increase (rm) was 0.166, with a finite rate of increase () of 1.180, per week.(AU)

Neste estudo avaliamos o potencial biótico e parâmetros reprodutivos de Spodoptera cosmioides (Walker, 1858) em condições controladas (25 ± 1 °C, 70 ± 10% UR e 14 horas de fotofase). Em média, as fêmeas (14,433 dias) não viveram significativamente mais do que os machos (13,100 dias). A duração média dos períodos de pre-oviposição, oviposição e pós-oviposição foram 2,767; 10,600 e 1,067 dias, respectivamente. A fecundidade média das fêmeas foi de 5.073,533 ovos e a fertilidade média foi de 5.021,027 larvas. Em média, as fêmeas copularam 1,167 vezes. A fecundidade correlacionou-se positivamente com o número de cópulas (r = 0,583; P 0,001). Entretanto, o número de cópulas correlacionou-se negativamente com a duração dos períodos de pré-oviposição (r = -0,560; P = 0,007), oviposição (r = -0,479; P = 0;048) e a longevidade (r = -0,512; P = 0,031). O potencial biótico foi estimado em 6,547 x 1021 indivíduos/fêmea/ano. A taxa líquida de reprodução (Ro) foi de 2.193,722 vezes por geração e o tempo médio de cada geração (T) foi de 46,407 dias. A taxa intrínseca de crescimento (rm) foi 0,166, com uma taxa finita de aumento () de 1,180, por semana.(AU)

Biotic potential and reproductive parameters of Spodoptera cosmioides (Walker) (Lepidoptera: Noctuidae) in the laboratory

Abstract In this study we evaluate the biotic potential and reproductive parameters of Spodoptera cosmioides (Walker, 1858) under controlled conditions (25 ± 1 °C, 70 ± 10% RH and 14 hour photophase). Females, on average, (14.433 days) did not live significantly longer than their male counterparts (13.100 days). The mean durations of the pre-oviposition, oviposition and post-oviposition periods were 2.767, 10.600 and 1.067 days, respectively. The mean fecundity of females was 5,073.533 eggs and the mean fertility was 5,021.027 larvae. On average, females copulated 1.167 times. Fecundity was positively correlated with the number of copulations (r = 0.583, P 0.001). The number of copulations, however, was negatively correlated whit the duration of the pre-oviposition (r = -0.560, P = 0.007) and oviposition (r = -0.479, P = 0.048) periods, and overall longevity (r = -0.512, P = 0.031). The biotic potential was estimated at 6.547 x 1021 individuals/female/year. The net reproductive rate (Ro) was 2,193.722 times per generation and the mean generation time (T) was 46.407 days. The intrinsic rate of increase (rm) was 0.166, with a finite rate of increase () of 1.180, per week.

Resumo Neste estudo avaliamos o potencial biótico e parâmetros reprodutivos de Spodoptera cosmioides (Walker, 1858) em condições controladas (25 ± 1 °C, 70 ± 10% UR e 14 horas de fotofase). Em média, as fêmeas (14,433 dias) não viveram significativamente mais do que os machos (13,100 dias). A duração média dos períodos de pre-oviposição, oviposição e pós-oviposição foram 2,767; 10,600 e 1,067 dias, respectivamente. A fecundidade média das fêmeas foi de 5.073,533 ovos e a fertilidade média foi de 5.021,027 larvas. Em média, as fêmeas copularam 1,167 vezes. A fecundidade correlacionou-se positivamente com o número de cópulas (r = 0,583; P 0,001). Entretanto, o número de cópulas correlacionou-se negativamente com a duração dos períodos de pré-oviposição (r = -0,560; P = 0,007), oviposição (r = -0,479; P = 0;048) e a longevidade (r = -0,512; P = 0,031). O potencial biótico foi estimado em 6,547 x 1021 indivíduos/fêmea/ano. A taxa líquida de reprodução (Ro) foi de 2.193,722 vezes por geração e o tempo médio de cada geração (T) foi de 46,407 dias. A taxa intrínseca de crescimento (rm) foi 0,166, com uma taxa finita de aumento () de 1,180, por semana.

Biotic potential and reproductive parameters of Spodoptera cosmioides (Walker) (Lepidoptera: Noctuidae) in the laboratory

Abstract In this study we evaluate the biotic potential and reproductive parameters of Spodoptera cosmioides (Walker, 1858) under controlled conditions (25 ± 1 °C, 70 ± 10% RH and 14 hour photophase). Females, on average, (14.433 days) did not live significantly longer than their male counterparts (13.100 days). The mean durations of the pre-oviposition, oviposition and post-oviposition periods were 2.767, 10.600 and 1.067 days, respectively. The mean fecundity of females was 5,073.533 eggs and the mean fertility was 5,021.027 larvae. On average, females copulated 1.167 times. Fecundity was positively correlated with the number of copulations (r = 0.583, P 0.001). The number of copulations, however, was negatively correlated whit the duration of the pre-oviposition (r = -0.560, P = 0.007) and oviposition (r = -0.479, P = 0.048) periods, and overall longevity (r = -0.512, P = 0.031). The biotic potential was estimated at 6.547 x 1021 individuals/female/year. The net reproductive rate (Ro) was 2,193.722 times per generation and the mean generation time (T) was 46.407 days. The intrinsic rate of increase (rm) was 0.166, with a finite rate of increase () of 1.180, per week.

Resumo Neste estudo avaliamos o potencial biótico e parâmetros reprodutivos de Spodoptera cosmioides (Walker, 1858) em condições controladas (25 ± 1 °C, 70 ± 10% UR e 14 horas de fotofase). Em média, as fêmeas (14,433 dias) não viveram significativamente mais do que os machos (13,100 dias). A duração média dos períodos de pre-oviposição, oviposição e pós-oviposição foram 2,767; 10,600 e 1,067 dias, respectivamente. A fecundidade média das fêmeas foi de 5.073,533 ovos e a fertilidade média foi de 5.021,027 larvas. Em média, as fêmeas copularam 1,167 vezes. A fecundidade correlacionou-se positivamente com o número de cópulas (r = 0,583; P 0,001). Entretanto, o número de cópulas correlacionou-se negativamente com a duração dos períodos de pré-oviposição (r = -0,560; P = 0,007), oviposição (r = -0,479; P = 0;048) e a longevidade (r = -0,512; P = 0,031). O potencial biótico foi estimado em 6,547 x 1021 indivíduos/fêmea/ano. A taxa líquida de reprodução (Ro) foi de 2.193,722 vezes por geração e o tempo médio de cada geração (T) foi de 46,407 dias. A taxa intrínseca de crescimento (rm) foi 0,166, com uma taxa finita de aumento () de 1,180, por semana.

Reproductive biology of the cockfish, Callorhinchus callorynchus (Chondrichthyes: Callorhinchidae), in coastal waters of the northern Argentinean Sea

The cockfish, Callorhinchus callorynchus, is a widely distributed holocephalan in the south-western Atlantic and an important resource for Argentinean fisheries. The reproductive characteristics of this species were studied in northern Argentinean coastal waters, where specimens arrive seasonally (winter-spring). Sexual dimorphism in maximum precaudal length (PCL) and total body mass (TM) was found, being females (PCL=630 mm; TM=3330 g; n=167) larger and heavier than males (PCL=482 mm; TM=1630 g; n=19). Maturity size for females was 466.22 mm PCL, representing 74% PCL of the largest female sampled. The mean number of ovarian follicles was 8.37 (± 4.84). The highest values of ovarian follicles diameter, gonadosomatic and oviducalsomatic indices were found in spring. A high proportion of mature females (44%) had atretic ovarian follicles, suggesting that they were in the resting stage of their reproductive cycle. The low number of mature males recorded and the absence of mature females with spermatophore masses or egg cases indicate that the study area would not be a reproductive zone. The seasonal presence of C. callorynchus in this region could be related to trophic movements. These results, obtained in an area with high fishing pressure on chondrichthyans, will be useful for implementing conservation and management measures.(AU)

El pez gallo, Callorhinchus callorynchus, es un holocéfalo con amplia distribución en el Atlántico Sudoccidental y un recurso importante para las pesquerías en Argentina. Las características reproductivas de esta especie fueron estudiadas en aguas costeras al norte del Mar Argentino, adonde se acercan estacionalmente (invierno-primavera). Se observó dimorfismo sexual en los valores máximos de longitud precaudal (LPC) y masa corporal total (MT), siendo las hembras (LPC=630 mm; MT=3330 g; n=167) más grandes y pesadas que los machos (LPC=482 mm; MT=1630 g; n=19). La talla de maduración para las hembras fue de 466,22 mm LPC, representando el 74% de la LPC máxima observada. El número promedio de folículos ováricos fue 8,37 (± 4,84). Los valores más altos del diámetro de los folículos ováricos, índice de la glándula oviductal e índice gonadosomático fueron hallados durante primavera. Una gran proporción de hembras maduras (44%) presentaron folículos ováricos atrésicos, sugiriendo que se encontraban en la etapa de reposo de su ciclo reproductivo. El bajo número de machos maduros registrados y la ausencia de hembras maduras con espermatóforos o cápsulas de huevos indican que las aguas costeras del norte de Argentina no serían una zona reproductiva. La presencia de C. callorynchus en el área de estudio podría estar relacionada con movimientos tróficos. Los resultados, obtenidos en un área con una gran presión pesquera sobre los condrictios, serán útiles para implementar adecuados planes de manejo y conservación de esta especie.(AU)

Reproductive biology of the cockfish, Callorhinchus callorynchus (Chondrichthyes: Callorhinchidae), in coastal waters of the northern Argentinean Sea

The cockfish, Callorhinchus callorynchus, is a widely distributed holocephalan in the south-western Atlantic and an important resource for Argentinean fisheries. The reproductive characteristics of this species were studied in northern Argentinean coastal waters, where specimens arrive seasonally (winter-spring). Sexual dimorphism in maximum precaudal length (PCL) and total body mass (TM) was found, being females (PCL=630 mm; TM=3330 g; n=167) larger and heavier than males (PCL=482 mm; TM=1630 g; n=19). Maturity size for females was 466.22 mm PCL, representing 74% PCL of the largest female sampled. The mean number of ovarian follicles was 8.37 (± 4.84). The highest values of ovarian follicles diameter, gonadosomatic and oviducalsomatic indices were found in spring. A high proportion of mature females (44%) had atretic ovarian follicles, suggesting that they were in the resting stage of their reproductive cycle. The low number of mature males recorded and the absence of mature females with spermatophore masses or egg cases indicate that the study area would not be a reproductive zone. The seasonal presence of C. callorynchus in this region could be related to trophic movements. These results, obtained in an area with high fishing pressure on chondrichthyans, will be useful for implementing conservation and management measures.(AU)

El pez gallo, Callorhinchus callorynchus, es un holocéfalo con amplia distribución en el Atlántico Sudoccidental y un recurso importante para las pesquerías en Argentina. Las características reproductivas de esta especie fueron estudiadas en aguas costeras al norte del Mar Argentino, adonde se acercan estacionalmente (invierno-primavera). Se observó dimorfismo sexual en los valores máximos de longitud precaudal (LPC) y masa corporal total (MT), siendo las hembras (LPC=630 mm; MT=3330 g; n=167) más grandes y pesadas que los machos (LPC=482 mm; MT=1630 g; n=19). La talla de maduración para las hembras fue de 466,22 mm LPC, representando el 74% de la LPC máxima observada. El número promedio de folículos ováricos fue 8,37 (± 4,84). Los valores más altos del diámetro de los folículos ováricos, índice de la glándula oviductal e índice gonadosomático fueron hallados durante primavera. Una gran proporción de hembras maduras (44%) presentaron folículos ováricos atrésicos, sugiriendo que se encontraban en la etapa de reposo de su ciclo reproductivo. El bajo número de machos maduros registrados y la ausencia de hembras maduras con espermatóforos o cápsulas de huevos indican que las aguas costeras del norte de Argentina no serían una zona reproductiva. La presencia de C. callorynchus en el área de estudio podría estar relacionada con movimientos tróficos. Los resultados, obtenidos en un área con una gran presión pesquera sobre los condrictios, serán útiles para implementar adecuados planes de manejo y conservación de esta especie.(AU)

Reproductive potential of Spodoptera eridania (Stoll) (Lepidoptera: Noctuidae) in the laboratory: effect of multiple couples and the size

Abstract This study aimed to evaluate the effect of keeping three couples in the same cage, and the size of adults emerged from small, medium-sized and large pupae (278.67 mg; 333.20 mg and 381.58 mg, respectively), on the reproductive potential of S. eridania (Stoll, 1782) adults, under controlled conditions (25 ± 1 °C, 70% RH and 14 hour photophase). We evaluated the survival, number of copulations, fecundity and fertility of the adult females. The survival of females from these different pupal sizes did not differ statistically, but the survival of males from large pupae was statistically shorter than from small pupae. Fecundity differed significantly and correlated positively with size. The number of effective copulations (espematophores) and fertility did not vary significantly with pupal size. Our results emphasize the importance of indicating the number of copulations and the size of the insects when reproductive parameters are compared.

Resumo Este estudo objetivou avaliar o efeito de confinar três casais em cada gaiola e o tamanho de adultos emergidos de pupas pequenas, medias e grandes (278,67 mg, 333,20 mg e 381,58 mg, respectivamente), sobre o potencial reprodutivo de S. eridania (Stoll, 1782), em condições controladas (25 ± 1 °C, 70% UR e 14 horas de fotofase). Avaliou-se a sobrevivência, o número de cópulas, fecundidade e fertilidade dos adultos. A sobrevivência não diferiu significativamente entre fêmeas provenientes de pupas de diferentes tamanhos, mas os machos oriundos de pupas grandes tiveram sobrevivência significativamente menor que os demais tamanhos. A fecundidade diferiu significativamente e correlacionou-se positivamente com o tamanho. O número de cópulas (espematóforos) e a fertilidade não variaram em função do peso pupal. Os resultados enfatizam a importância de indicar o número de cópulas e o tamanho dos insetos estudados para que comparações entre os parâmetros reprodutivos possam ser efetuadas.

Reproductive potential of Spodoptera eridania (Stoll) (Lepidoptera: Noctuidae) in the laboratory: effect of multiple couples and the size

Abstract This study aimed to evaluate the effect of keeping three couples in the same cage, and the size of adults emerged from small, medium-sized and large pupae (278.67 mg; 333.20 mg and 381.58 mg, respectively), on the reproductive potential of S. eridania (Stoll, 1782) adults, under controlled conditions (25 ± 1 °C, 70% RH and 14 hour photophase). We evaluated the survival, number of copulations, fecundity and fertility of the adult females. The survival of females from these different pupal sizes did not differ statistically, but the survival of males from large pupae was statistically shorter than from small pupae. Fecundity differed significantly and correlated positively with size. The number of effective copulations (espematophores) and fertility did not vary significantly with pupal size. Our results emphasize the importance of indicating the number of copulations and the size of the insects when reproductive parameters are compared.(AU)

Resumo Este estudo objetivou avaliar o efeito de confinar três casais em cada gaiola e o tamanho de adultos emergidos de pupas pequenas, medias e grandes (278,67 mg, 333,20 mg e 381,58 mg, respectivamente), sobre o potencial reprodutivo de S. eridania (Stoll, 1782), em condições controladas (25 ± 1 °C, 70% UR e 14 horas de fotofase). Avaliou-se a sobrevivência, o número de cópulas, fecundidade e fertilidade dos adultos. A sobrevivência não diferiu significativamente entre fêmeas provenientes de pupas de diferentes tamanhos, mas os machos oriundos de pupas grandes tiveram sobrevivência significativamente menor que os demais tamanhos. A fecundidade diferiu significativamente e correlacionou-se positivamente com o tamanho. O número de cópulas (espematóforos) e a fertilidade não variaram em função do peso pupal. Os resultados enfatizam a importância de indicar o número de cópulas e o tamanho dos insetos estudados para que comparações entre os parâmetros reprodutivos possam ser efetuadas.(AU)

VIABILIDADE ESPERMÁTICA DE Macrobrachium amazonicum SUBMETIDOS A DOIS MÉTODOS DE EXTRAÇÃO DE ESPERMATÓFORO

Na aquicultura mundial o cultivo de camarão de água doce tem apresentado um elevado crescimento, sendo uma das espécies mais cultivadas o Macrobrachium amazonicum, que apresenta população de machos adultos podendo ser classificada em quatro morfotipos, diferindo entre si em tamanho, morfologia, fisiologia e comportamento. O objetivo deste trabalho foi verificar a qualidade e viabilidade de espermatóforos e espermatozoides de M. amazonicum submetidos a diferentes formas de extração. Foram utilizados 60 animais machos, entre duas baterias, destes, 30 utilizados para a extração manual e os outros 30 para eletroejaculação. Os animais eram observados diariamente e os parâmetros da água, temperatura e pH monitorados diariamente e feitos a cada dois dias testes de amônia e nitrito. Os espermatóforos extraídos foram então armazenados em microtubos de 1,5 mL contendo solução fisiológica (NaCl a 0,9%) e mantidos em temperatura ambiente. Para a avaliar da porcentagem aparente de espermatozoides vivos e mortos, foram utilizados 50 L de solução composta pelos corantes eosina-nigrosina e adicionada em 50 L de solução espermática. Posteriormente amostras de 25 L da solução total foram levadas a câmara de Neubauer para serem analisadas utilizando microscópio óptico com aumento de 40x. A porcentagem de espermatozoides vivos foi calculada a partir da contagem de 100 espermatozoides. Os dois métodos se mostraram eficientes na extração dos espermatóforos, não apresentando diferença significativa entre eles quanto a taxa de sobrevivência dos espermatozoides (P>0,05). Entretanto, verificou-se diferença (P<0,05) no tamanho e integridade dos espermatóforos extraídos, apresentando os seguintes valores médios finais (±DP) 1,69mm (± 0,53) obtidos manualmente e 3,12mm (± 1,06) eletricamente. Desta forma a extração elétrica se mostrou mais eficiente na obtenção de espermatóforo quando comparada a extração manual.

In aquaculture worldwide the cultivation of freshwater shrimp has shown a high growth, being one of the most cultivated species the Macrobrachium amazonicum, which has a population of adult males and can be superior in four morphotypes, differing in size, morphology, physiology and behavior. The objective of this work was to verify the quality and viability of spermatophores and spermatozoa of M. amazonicum in different forms of extraction. 60 male animals were used, between two batteries, of these, 30 used for manual extraction and the other 30 for electroejaculation. The animals were observed daily and the water, temperature and pH parameters were monitored daily and ammonia and nitrite tests were performed every two days. The extracted spermatophores were then stored in 1.5 mL microtubes containing saline (0.9% NaCl) and collect at room temperature. For the evaluation of the apparent percentage of live and dead sperm, 50 L of solution composed by the dyes eosin-nigrosine and Added in 50 L of sperm solution were used. Subsequently, they removed 25 L of the total solution and were taken to the Neubauer chamber to be analyzed using an optical microscope with a 40x magnification. The percentage of live sperm was calculated from the count of 100 sperm. Both methods improve sperm extraction, they do not increase the difference between them in terms of sperm taxa (P> 0.05). However, there was a difference (P <0.05) in the size and integrity of the extracted spermatophores, these following final mean values (± SD) 1.69mm (± 0.53) manually adjusted and 3.12mm (± 1.06) electrically. Thus, electrical extraction has become more efficient in obtaining spermatophore when compared to manual extraction.

Biotic potential and reproductive parameters of Spodoptera dolichos (Lepidoptera: Noctuidae) in the laboratory

The biotic potential and reproductive parameters of Spodoptera dolichos (Fabricius, 1794) were evaluated under controlled conditions (25 ± 1°C, 70 ± 10% RH and 14 hour photophase). The longevity, pre-, post- and oviposition periods, fecundity, and fertility of 25 couples were evaluated. The longevity of females (12.9 days) was not significantly different than that of males (12.4 days). The mean durations of the pre-, post- and oviposition periods were 3.0, 0.4 and 10.4 days, respectively. The mean fecundity was 4,086.0 eggs per female and mean fertility was 3,557.8 larvae per female. On average, a female copulated 1.4 times. The biotic potential of S. dolichos was estimated at 7.138 x 1018 individuals/female/year. The net reproductive rate (Ro) was 1,711.98 times per generation and the mean generation time (T) was 56.19 days. The intrinsic rate of increase (rm) was 0.133, with a finite rate of increase (l) of 1.142 per day. These results are compared with other species from Spodoptera and their relevance for management strategies of S. dolichos.

Biotic potential and reproductive parameters of Spodoptera dolichos (Lepidoptera: Noctuidae) in the laboratory

The biotic potential and reproductive parameters of Spodoptera dolichos (Fabricius, 1794) were evaluated under controlled conditions (25 ± 1°C, 70 ± 10% RH and 14 hour photophase). The longevity, pre-, post- and oviposition periods, fecundity, and fertility of 25 couples were evaluated. The longevity of females (12.9 days) was not significantly different than that of males (12.4 days). The mean durations of the pre-, post- and oviposition periods were 3.0, 0.4 and 10.4 days, respectively. The mean fecundity was 4,086.0 eggs per female and mean fertility was 3,557.8 larvae per female. On average, a female copulated 1.4 times. The biotic potential of S. dolichos was estimated at 7.138 x 1018 individuals/female/year. The net reproductive rate (Ro) was 1,711.98 times per generation and the mean generation time (T) was 56.19 days. The intrinsic rate of increase (rm) was 0.133, with a finite rate of increase (l) of 1.142 per day. These results are compared with other species from Spodoptera and their relevance for management strategies of S. dolichos.(AU)

Endocrine and paracrine regulation of zebrafish spermatogenesis: the Sertoli cell perspective

Spermatogonial stem cells (SSCs) either self-renew or differentiate into spermatogonia that further develop into spermatozoa. Self-renewal occurs when residing in a specific micro-environment (niche) While displacement from the niche would tip the signalling balance towards differentiation. Considering the cystic type of spermatogenesis in fish, the SSC candidates are single type A undifferentiated (Aund) spermatogonia, enveloped by mostly one niche-forming Sertoli cell. When going through a self-renewal cell cycle, the resulting new single type Aund spermatogonium would have to recruit another Sertoli cell to expand the niche space, while a differentiating germ cell cyle would result in a pair of spermatogonia that remain in contact with their cyst-forming Sertoli cells. In zebrafish, thyroid hormone stimulates the proliferation of Sertoli cells and of type Aund spermatogonia, involving Igf3, a new member of the Igf family. In cystic spermatogenesis, type A und spermatogonia usually do not leave the niche, so that supposedly the signalling in the niche changes when switching from self-renewal to differentiation. Recombinant zebrafish (rz) Fsh down-regulated Sertoli cell anti-müllerian hormone (amh) mRNA levels, and rzAmh inhibited differentiation of type A und spermatogonia as well as Fsh-stimulated steroidogenesis. Thus, for Fsh to efficiently stimulate testis functions, Amh bioactivity should be dampened. We also discovered that Fsh increased Sertoli cell Igf3 gene and protein expression; rzIgf3 stimulated spermatogonial proliferation and Fsh-stimulated spermatogenesis was significantly impaired by inhibiting Igf receptor signaling. We propose that in zebrafish, Fsh is the major regulator of testis functions and, supported by other endocrine systems (e.g. thyroid hormone), regulates Leydig cell steroidogenesis as well as Sertoli cell number and growth factor production to promote spermatogenesis.

Spermatogenesis in the yellowtail tetra Astyanax altiparanae: a histological analyses with emphasis to spermatogonial and spermatid types / Espermatogênese no lambari-do-rabo-amarelo Astyanax altiparanae: uma análise histológica com ênfase aos tipos espermatogoniais e espermático

This study aimed to detailed description of the characteristics of the different germ cell types found in Astyanax altiparanae during spermatogenesis. In this purpose, testes from 25 adult males specimens of A. altiparanae were sampled and submitted to the usual techniques for light microscopy. Based on nuclear shape, chromatin condensation, nucleoli quantity and cell size were identified four spermatogonial types: type A undifferentiated (Aund.*); type A undifferentiated (Aund.); type A differentiated (Adif.); and type B spermatogonia. Spermatocytes were observed in different phases of meiosis (leptotene/zygotene/pachytene and diplotene), metaphase I and II and secondary spermatocytes, being distinguish mainly by their chromosomal organization inside the nucleus. Were also identified three different types of spermatids, which were named as initial, intermediate and final, which can be differentiated by increase in nuclear condensation and spacing among cells inside the cysts, possibly by flagella arise and reduction in nuclear diameter. Thus, this study contribute to a better understand of spermatogenesis in this and other fish species.(AU)

O presente estudo teve como objetivo a descrição detalhada dos diferentes tipos de células germinativas encontradas em Astyanax altiparanae durante a espermatogênese. Deste modo, os testículos de 25 espécimes machos e adultos de A. altiparanae foram coletados e submetidos às técnicas usuais para microscopia de luz. Baseado no formato nuclear, condensação da cromatina, quantidade de nucléolos e tamanho celular foram identificados quatro tipos de espermatogônias: indiferenciadas do tipo A (Aund.*); indiferenciadas do tipo A (Aund.); diferenciadas do tipo A (Adif.); e espermatogônia do tipo B. Os espermatócitos foram observados em diferentes fases da meiose (leptóteno/zigóteno, paquíteno e diplóteno), metáfase I e II e espermatócitos secundários, sendo distinguidos principalmente pela organização dos cromossomos nos núcleos. Também foram identificados três diferentes tipos de espermátides, que foram nomeadas como iniciais, intermediárias e finais, que se diferenciaram pelo aumento da compactação nuclear e espaçamento entre as células no cisto, pelo possível surgimento do flagelo, e pelo diâmetro nuclear. Assim, este estudo contribui para um melhor entendimento da espermatogênese nesta e nas demais espécies de peixes.(AU)

Endocrine and paracrine regulation of zebrafish spermatogenesis: the Sertoli cell perspective

Spermatogonial stem cells (SSCs) either self-renew or differentiate into spermatogonia that further develop into spermatozoa. Self-renewal occurs when residing in a specific micro-environment (niche) While displacement from the niche would tip the signalling balance towards differentiation. Considering the cystic type of spermatogenesis in fish, the SSC candidates are single type A undifferentiated (Aund) spermatogonia, enveloped by mostly one niche-forming Sertoli cell. When going through a self-renewal cell cycle, the resulting new single type Aund spermatogonium would have to recruit another Sertoli cell to expand the niche space, while a differentiating germ cell cyle would result in a pair of spermatogonia that remain in contact with their cyst-forming Sertoli cells. In zebrafish, thyroid hormone stimulates the proliferation of Sertoli cells and of type Aund spermatogonia, involving Igf3, a new member of the Igf family. In cystic spermatogenesis, type A und spermatogonia usually do not leave the niche, so that supposedly the signalling in the niche changes when switching from self-renewal to differentiation. Recombinant zebrafish (rz) Fsh down-regulated Sertoli cell anti-müllerian hormone (amh) mRNA levels, and rzAmh inhibited differentiation of type A und spermatogonia as well as Fsh-stimulated steroidogenesis. Thus, for Fsh to efficiently stimulate testis functions, Amh bioactivity should be dampened. We also discovered that Fsh increased Sertoli cell Igf3 gene and protein expression; rzIgf3 stimulated spermatogonial proliferation and Fsh-stimulated spermatogenesis was significantly impaired by inhibiting Igf receptor signaling. We propose that in zebrafish, Fsh is the major regulator of testis functions and, supported by other endocrine systems (e.g. thyroid hormone), regulates Leydig cell steroidogenesis as well as Sertoli cell number and growth factor production to promote spermatogenesis.(AU)

Spermatogenesis in the yellowtail tetra Astyanax altiparanae: a histological analyses with emphasis to spermatogonial and spermatid types / Espermatogênese no lambari-do-rabo-amarelo Astyanax altiparanae: uma análise histológica com ênfase aos tipos espermatogoniais e espermático

This study aimed to detailed description of the characteristics of the different germ cell types found in Astyanax altiparanae during spermatogenesis. In this purpose, testes from 25 adult males specimens of A. altiparanae were sampled and submitted to the usual techniques for light microscopy. Based on nuclear shape, chromatin condensation, nucleoli quantity and cell size were identified four spermatogonial types: type A undifferentiated (Aund.*); type A undifferentiated (Aund.); type A differentiated (Adif.); and type B spermatogonia. Spermatocytes were observed in different phases of meiosis (leptotene/zygotene/pachytene and diplotene), metaphase I and II and secondary spermatocytes, being distinguish mainly by their chromosomal organization inside the nucleus. Were also identified three different types of spermatids, which were named as initial, intermediate and final, which can be differentiated by increase in nuclear condensation and spacing among cells inside the cysts, possibly by flagella arise and reduction in nuclear diameter. Thus, this study contribute to a better understand of spermatogenesis in this and other fish species.

O presente estudo teve como objetivo a descrição detalhada dos diferentes tipos de células germinativas encontradas em Astyanax altiparanae durante a espermatogênese. Deste modo, os testículos de 25 espécimes machos e adultos de A. altiparanae foram coletados e submetidos às técnicas usuais para microscopia de luz. Baseado no formato nuclear, condensação da cromatina, quantidade de nucléolos e tamanho celular foram identificados quatro tipos de espermatogônias: indiferenciadas do tipo A (Aund.*); indiferenciadas do tipo A (Aund.); diferenciadas do tipo A (Adif.); e espermatogônia do tipo B. Os espermatócitos foram observados em diferentes fases da meiose (leptóteno/zigóteno, paquíteno e diplóteno), metáfase I e II e espermatócitos secundários, sendo distinguidos principalmente pela organização dos cromossomos nos núcleos. Também foram identificados três diferentes tipos de espermátides, que foram nomeadas como iniciais, intermediárias e finais, que se diferenciaram pelo aumento da compactação nuclear e espaçamento entre as células no cisto, pelo possível surgimento do flagelo, e pelo diâmetro nuclear. Assim, este estudo contribui para um melhor entendimento da espermatogênese nesta e nas demais espécies de peixes.

Criopreservação do espermatóforo e da massa espermática do camarão branco Litopenaeus schmitti post-mortem / Post-mortem spermatophore and sperm cryopreservation of the white shrimp Litopenaeus schmitti

This study was carried out in two phases; the first to evaluate the viability of two cooling protocols (A and B) using glycerol (10%) as a cryoprotectant and the second, the efficiency of two cryoprotectants (10% glycerol and DMSO) and times (30, 60 and 90 days of storage in liquid nitrogen) for the cryopreservation of post-mortem sperm mass and spermatophores of the shrimp Litopenaeus schmitti. The protocol A presented a cooling rate of 0.5C min-1 until reaching -32ºC and protocol B, 20ºC min-1 until reach -120ºC, after which the samples were transferred to liquid nitrogen (N2L). The sperm viability was assessed by smearing the semen stained with eosin-nigrosin. The curve resulted in higher mean sperm survival was Protocol A (50.9%). Therefore, for the second experiment, the protocol A was used, however there was no difference between the cryoprotectants for sperm mass, although the influence on survival time. Difference was observed between the cryoprotectants and the influence of time on sperm survival of cryopreserved spermatophores. Glycerol 10% was more efficient for cryopreservation of spermatophore, but for the masses sperm, both can be used.(AU)

O presente trabalho foi realizado em duas etapas; a primeira para avaliar a eficiência de dois protocolos de resfriamento (A e B) utilizando glicerol (10%) como crioprotetor e, a segunda, a eficiência de dois crioprotetores (glicerol e dimetilsulfóxido - DMSO a 10%) e o tempo (30, 60 e 90 dias de estocagem em nitrogênio líquido) para a criopreservação da massa espermática e espermatóforo de camarões post mortem da espécie Litopenaeus schmitti. O protocolo A apresentou velocidade de resfriamento de 0,5ºC min-1 até alcançar -32ºC e o protocolo B, 20ºC min-1 até alcançar -120ºC, sendo os materiais posteriormente transferidos para o nitrogênio líquido (N2L). A viabilidade espermática foi avaliada por meio do esfregaço de sêmen corado com eosina-nigrosina. A curva que resultou na maior média de sobrevivência espermática foi a do protocolo A (50,9%). Portanto, para o segundo experimento foi utilizado o protocolo A; entretanto, neste não houve diferença entre os crioprotetores para a massa espermática, contudo, o tempo influenciou na sobrevivência. Foi observada diferença entre os crioprotetores e a influência do tempo na sobrevivência espermática dos espermatóforos criopreservados. O glicerol a 10% foi mais eficiente para a criopreservação dos espermatóforos, porém, para as massas espermáticas, ambos podem ser utilizados.(AU)