Orientador: José Luiz Moraes Vasconcelos / Banca: Rui Machado / Banca: Heraldo César Gonçalves / Resumo: Foram realizados quatro experimentos com o objetivo de avaliar possíveis mecanismos relacionados ao aumento da prenhez com a utilização de ácidos graxos poliinsaturados (PF). No exp. 1 foram utilizadas 51 vacas multíparas Nelore não lactantes, ovuladas, para avaliar se PF alteram a produção de progesterona (P4) e o momento da luteólise. No exp. 2 foram utilizadas 43 vacas multíparas Nelore não lactantes, ovuladas, para avaliar se PF alteram a sensibilidade do corpo lúteo de seis dias a aplicação de prostaglandina (i.m. 12,5mg de dinoprost trometamina, Lutalyse). No exp. 3 foram utilizadas 27 vacas multíparas Nelore, ovuladas, com 30 a 40 dias pós parto para avaliar se PF diminuem a incidência de ciclo curto. Os tratamentos utilizados foram: grupo controle (100g mineral + 100g milho + 100g caolin vaca dia); grupo SF (100g mineral + 100g milho + 100g/vaca/dia de Megalac (7-9% C18:2; Arm&Hammer a Church&Dwight Company, EUA); grupo PF (100g mineral + 100g milho + 100g/vaca/dia de Megalac-E QGN, Brasil: 40-42% de C18:2; 2-3% de C18:3). No exp. 4 (1457 vacas multíparas Nelore) foi avaliado se a suplementação com PF pós IATF por diferentes períodos altera a taxa de prenhez. Nos experimentos 1, 2 e 3 não foi detectado efeito de PF nas concentrações de P4 durante o período avaliado, no momento da luteólise e na incidência de ciclo curto (P>0,1). Ao se agrupar os dados do Exp 1 e 2, a concentração de P4 (P=0,01) no dia 6 foi maior nos animais suplementados com PF em relação ao SF e controle (4,45; 3,25; 3,48 ng/ml, respectivamente; EPM=0,278). No experimento 4 vacas recebendo PF por 21 e 28 dias após a IATF tiveram maior (P<0,05) taxa de prenhez (50,38%) quando comparadas com os outros tratamentos agrupados (42,38%). Não foi detectado diferença entre os tratamentos PF21 (50,99%) e PF28 (49,81). Os resultados em conjunto mostram que apesar de suplementação... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Four experiments were designed to evaluate the possible mechanisms related to the increased pregnancy rates in cows supplemented with PF. In Experiment 1, 51 ovulated non-lactating Nelore multiparous cows were used to evaluate if PF supplementation affects circulating progesterone (P4) concentrations and timing of luteolysis. In Experiment 2, 43 ovulated non-lactating Nelore multiparous cows were used to evaluate if PF supplementation alters the sensitivity of a 6-d corpus luteum to exogenous prostaglandin treatment. In Experiment 3, 27 ovulated postpartum Nelore cows were used to evaluate if PF supplementation affects the incidence of premature luteolysis. Beginning at the d of estrus, the daily treatments in these 3 experiments were: Control (0.1 Kg Mineral + 0.1 Kg corn + 0.1 Kg kaolin); SF (0.1 Kg Mineral + 0.1 Kg Megalac-S® [7-9% linoleic acid] + 0.1 Kg corn), this group was used just in experimental 1 and 2; PF (0.1 Kg Mineral + 0.1 Kg Megalac-E® [40-42% linoleic acid; 2-3% linolenic acid] + 0.1 Kg corn). In Experiment 4, we evaluated if the length of PF supplementation in different times after timed-AI alters the pregnancy rate. No effect was detected on P4 concentration, luteolysis or short cycle (P>0.1), but when the cows of exp. 1 and 2 were grouped had higher (P=0.01) concentration of P4) on cows that were supplemented with PF compared with Sf or control (4.45; 3.25; 3.48 ng/ml, respectively; SEM=0,278. Cows supplemented with PF during 21 (PF21,) or 28 d post- AI (PF28) had higher pregnancy rates (50.38%; P < 0.05) than cows from other treatments (42.38%). There was no difference between PF21 (50,99%) and PF28 (49,81%) treatments. These experiments indicated that the possible mechanism for greater conception with PF supplementation post-AI may be due to effects on embryo development, animals supplemented for more than 21d had greater pregnancy rates. / Mestre

The objective was to investigate the effects of progesterone (P4) on luteinizing hormone (LH) release, follicle development, and oocyte competence in cattle. We tested the general hypotheses that: 1) The suppressive effect of P4 on gonadotrophin releasing hormone (GnRH)-mediated LH release can be overcome by increasing GnRH dose or pre-treatment with estradiol (E2); and 2) a shorter period of P4 exposure during the growing phase of the ovulatory follicle improves oocyte competence and fertility after fixed-time artificial insemination or superstimulation in cattle. <p>In the first experiment, heifers (n=22) were treated with 100 or 200 µg of GnRH or pretreated with E2 prior to administration of GnRH during high or low circulating P4 concentrations to characterize LH release (Chapter 2). Increasing the dose of GnRH did not alter LH secretion; however, E2 pretreatment overcame the suppressive effect of high P4 on LH secretion. Cattle with lower (n=11) P4 concentrations had higher circulating LH concentrations than those with higher P4 concentrations (n=11), and tended to have higher ovulation rates. <p>Two experiments were conducted to determine the effect of the duration of P4 exposure during the ovulatory wave on fertility followed fixed-time artificial insemination or superstimulation. In the first experiment (Chapter 3), the dominant follicle was allowed to grow for 3 days (n=181) or 6 days (n=184). Six days of growth resulted in a larger dominant follicle, but in both groups, ovulatory follicles had similar capacities to ovulate and establish pregnancy. In the second experiment (Chapter 4), multiple follicles were allowed to grow for 3 or 6 days by 8 or 14 injections of FSH (at 12-hour intervals). There was no difference between groups for ovulation rate or total ova/embryo recovery rate. Although the 3-day group had higher embryo quality at slaughter (4 days after insemination), further development (7, 9, and 10 days after insemination) did not differ among groups. The effect of FSH starvation following 4 days of FSH treatment (Chapter 4) resulted in loss of ovulatory capability. Overall, a shorter duration of P4 exposure during ovulatory follicle growth did not improve fertility after fixed-time AI or oocyte competence after superstimulation.

The objective was to investigate the effects of progesterone (P4) on luteinizing hormone (LH) release, follicle development, and oocyte competence in cattle. We tested the general hypotheses that: 1) The suppressive effect of P4 on gonadotrophin releasing hormone (GnRH)-mediated LH release can be overcome by increasing GnRH dose or pre-treatment with estradiol (E2); and 2) a shorter period of P4 exposure during the growing phase of the ovulatory follicle improves oocyte competence and fertility after fixed-time artificial insemination or superstimulation in cattle. <p>In the first experiment, heifers (n=22) were treated with 100 or 200 µg of GnRH or pretreated with E2 prior to administration of GnRH during high or low circulating P4 concentrations to characterize LH release (Chapter 2). Increasing the dose of GnRH did not alter LH secretion; however, E2 pretreatment overcame the suppressive effect of high P4 on LH secretion. Cattle with lower (n=11) P4 concentrations had higher circulating LH concentrations than those with higher P4 concentrations (n=11), and tended to have higher ovulation rates. <p>Two experiments were conducted to determine the effect of the duration of P4 exposure during the ovulatory wave on fertility followed fixed-time artificial insemination or superstimulation. In the first experiment (Chapter 3), the dominant follicle was allowed to grow for 3 days (n=181) or 6 days (n=184). Six days of growth resulted in a larger dominant follicle, but in both groups, ovulatory follicles had similar capacities to ovulate and establish pregnancy. In the second experiment (Chapter 4), multiple follicles were allowed to grow for 3 or 6 days by 8 or 14 injections of FSH (at 12-hour intervals). There was no difference between groups for ovulation rate or total ova/embryo recovery rate. Although the 3-day group had higher embryo quality at slaughter (4 days after insemination), further development (7, 9, and 10 days after insemination) did not differ among groups. The effect of FSH starvation following 4 days of FSH treatment (Chapter 4) resulted in loss of ovulatory capability. Overall, a shorter duration of P4 exposure during ovulatory follicle growth did not improve fertility after fixed-time AI or oocyte competence after superstimulation.

MSCAGR (Animal Science) / Department of Animal Science / The aims of the study were to evaluate the effects of dilution and chilled storage time on the quality of semen, and of different artificial insemination techniques on fertility in artificially inseminated indigenous Venda does. Fresh semen was collected using an artificial vagina from three Boer bucks aged 4±1.55 years once every four days during July and August 2016. Semen was pooled and samples were divided into two equal parts, which were extended using Biladyl® extender at ratios of 1:5 and 1:10 v/v (semen to extender), before refrigeration for 120 hours at 5 °C. The fresh undiluted semen and freshly extended semen were evaluated in six replicates for sperm motility, live-dead and sperm morphology using the Sperm Class Analyzer (SCA). Extended semen continued to be evaluated at 24 hour intervals for 120 hours. Ninety indigenous Venda does were obtained from different flocks in the Vhembe district and kept intensively in one 10 m x 40 m pen at the University of Venda experimental farm in the goat feedlot. The does were fed and watered ad libitum. After acclimatization for 14 days, estrus was synchronized using a controlled internal drug release (CIDR) containing 0.3 g of progesterone. Upon removal of the CIDR, does were injected 10 mg of PGF2α (Lutalyse® dinoprost tromethamine) Sterile Solution. At 24 hours after the removal of the CIDR, the does were injected intramuscularly with 300 international units (IU) of equine chorionic gonadotrophin (eCG). Forty eight hours after the removal of the progesterone, freshly collected and diluted (1:5 ratio ~150x106 sperm/ml), five day-stored semen were used to inseminate the does using cervical (CAI), trans-cervical (TAI), and laparoscopic artificial (LAI) insemination methods in a complete randomized design (CRD) with a 2 X 3 factorial arrangement of the treatments with 15 replications per treatment. The does were tested for pregnancy after 30 days using ultrasonography. Analyses of variance was performed on the pregnancy, kidding rates and on prolificacy using the GLM procedure of Minitab (Minitab 2013). Significant differences in all motility parameters were observed between the extension ratios and storage time (P<0.01). There were significant interactions between the extension ratio and storage time (P<0.05) on the sub-population of sperm cells with non-progressive motility (NON-P). Significant (P<0.01) interaction was observed between the semen extension ratio and storage time on medium and slow spermatozoa (P<0.01). The method of insemination did not (P>0.05) affect fertility, though both pregnancy and kidding rates numerically decreased in the order laparoscopic insemination (LAI)≥ trans-cervical insemination (TAI)≥ cervical insemination (CAI). Overall, 71% kidding rate was achieved.

A criopreservação do sêmen resulta em danos à estrutura espermática, sendo nítida a importância da avaliação das partidas de sêmen antes de serem submetidas à inseminação artificial em tempo fixo (IATF). Todavia, nem sempre as avaliações convencionais do sêmen são suficientes para identificar partidas que possam resultar em baixa taxa de prenhez no campo, sendo necessária uma investigação mais profunda e acurada. Neste sentido, este experimento foi realizado com o objetivo de identificar partidas de sêmen que apresentam falhas na fertilidade, mesmo sendo aprovadas pelas avaliações convencionais. Foram realizadas análises convencionais (motilidade, vigor, concentração e morfologia espermática) de 72 partidas de sêmen de 22 touros antes da estação de monta. Destas, 55 partidas de 18 touros foram aprovadas para o uso na IATF, mas somente 28 partidas de 10 touros foram utilizadas. As partidas de sêmen utilizadas na IATF foram submetidas a outras avaliações: análise computadorizada da motilidade espermática (CASA), integridade das membranas plasmática e acrossomal e potencial de membrana mitocondrial (por sondas fluorescentes em microscopia de epifluorescência). Os dados foram analisados pelo Proc Mixed do SAS usando o Test T. As taxas de prenhez das diferentes partidas de sêmen variaram de 71 a 37%, sendo a média e desvio padrão das partidas de 55,57&#177;7,57%. Os dados de fertilidade a campo permitiram a separação das partidas de sêmen como de Alta (&gt;50% de prenhez) e Baixa (50% de prenhez) fertilidade, sendo comparadas quatro partidas de Alta e quatro de Baixa fertilidade. Os dados das características seminais de todas as partidas de sêmen foram submetidos à análise por boxplot e separados em quartis superior e inferior. Quando se comparou as partidas de Alta e Baixa fertilidade notou-se diferença na taxa de prenhez (p&lt;0,01), mas não foi notada diferença para motilidade (p=0,91), vigor (p=0,63), concentração (p=0,27), número de espermatozoides por palheta (NEP, p=0,27), número de espermatozoides móveis e normais por palheta (p=0,18), defeitos maiores (p=0,17), defeitos totais (p=0,43), integridade de membrana plasmática (MPI, p=0,07), alto potencial de membrana mitocondrial (AP, p=0,94), motilidade total (MT, p=0,10), VCL (p=0,80), VSL (p=0,75), VAP (p=0,88), LIN (p=0,78), STR (p=0,71) e BCF (p=0,13). No entanto, foram encontradas diferenças entre os grupos (Alta e Baixa) para defeitos menores (p=0,05), espermatozoides com integridade das membranas plasmática e acrossomal e função mitocondrial (PIAIA)/Palheta (p=0,01), integridade de acrossomo (AI, p=0,03), motilidade progressiva (MP, p&lt;0.01) e rápidos (p&lt;0,01). Quando comparados os quartis superior e inferior das características seminais foram encontradas diferenças para concentração espermática (p&lt;0,01), NEP (p&lt;0,01), número de espermatozoides móveis por palheta (p&lt;0,01), espermatozoides móveis e normais (p&lt;0,01), defeitos maiores (p&lt;0,01), defeitos menores (p=0,01), defeitos totais (p=0,05), MPI (p&lt;0,01), AI (p&lt;0,01), AP (p=0,03), PIAIA (p&lt;0,01), PIAIA/palheta (p&lt;0,01), mas esta divisão de grupos por quartis superior e inferior não apresentaram diferença sobre a fertilidade (p&gt;0,05); sendo que somente. Entretanto, para MT (p&lt;0,01) e MP (p&lt;0,01) além da diferença entre os quartis foi notado efeito da fertilidade (MT, p=0,05 e MP, p=0,01), sendo maior para os de baixa fertilidade. Pode-se concluir que os padrões de qualidade de partidas de sêmen que apresentam fertilidade distinta podem ser semelhantes, além disso, que pode haver diferença na qualidade espermática entre as partidas que não influenciam a fertilidade. Desta forma, são necessárias outras análises mais acuradas para investigar as causas de falha na fertilidade de algumas partidas de sêmen. / Semen cryopreservation results in damage to sperm structure, and it is clear the importance of evaluating the semen batches before submitted it to timed artificial insemination (TAI). However, conventional semen evaluations are not always sufficient to identify batch that may result in reduced pregnancy rate in the field, requiring a more thorough and accurate investigation. Thus, this experiment was conducted in order to identify semen batches that have gaps in fertility, even being adopted by conventional assessments. Conventional analysis (motility, vigor, concentration and morphology) were performed of 72 semen batches from 22 bulls before the breeding season. Of these, 55 batches from 18 bulls were approved for use in TAI, but only 28 batches from 10 bulls were used. Semen batches used in TAI were subjected to further assessment: computer-assisted sperm analysis (CASA), integrity of plasma and acrosomal membranes and mitochondrial membrane potential (by fluorescent probes under epifluorescence microscopy). Data were analyzed by Proc Mixed of the SAS using Test \"T\". Pregnancy rates of different semen batches ranged 71-37%, and the mean and standard deviation of the batches was 55.57 &#177; 7.57%. Field fertility data allowed the separation of the semen batches as \"High\" (&gt;50% pregnancy rate) and \"Low\" (50% pregnancy rate) fertility. It was compared four batches of \"High\" and four batches of \"Low\" fertility. Data of the seminal characteristics of all the semen batches were analyzed by boxplot and separated into upper and lower quartiles. When comparing the batches of \"High\" and \"Low\" fertility was noticed a difference in the pregnancy rate (p&lt;0.01), but was not noticeable difference in motility (p=0.91), vigour (p=0.63), concentration (p=0.27), number of spermatozoa per straw (NEP, p=0.27), number of motile and normal sperm per straw (p=0.18), major defects (p=0.17), total defects (p=0.43) plasma membrane integrity (MPI, p=0.07), high mitochondrial membrane potential (AP, p=0.94), total motility (TM, p=0.10), VCL (p=0.80), VSL (p=0.75), VAP (p=0.88), LIN (p=0.78), STR (p=0.71) and BCF (p=0.13). However, differences were found between the groups (\"High\" and \"Low\") for minor defects (p=0.05), sperm with plasma and acrosomal membranes integrity and high mitochondrial membrane potential (PIAIA)/straw (p=0.01), acrosomal integrity (Al, p=0.03) progressive motility (PM, p &lt;0.01) and rapid (p&lt;0.01). When comparing the upper and lower quartiles of the seminal characteristics differences were found for sperm concentration (p &lt;0.01), NEP (p &lt;0.01), number of motile sperm per straw (p&lt;0.01), motile and normal sperm (p&lt;0.01), major defects (p&lt;0.01), minor defects (p=0.01), total defects (p=0.05), MPI (p&lt;0.01), AI (p&lt;0.01), AP (p=0.03), PIAIA (p&lt;0.01), PIAIA/straw (p&lt;0.01), but this division groups by upper and lower quartiles showed no difference in fertility rate (p&gt;0.05). However, to MT (p&lt;0.01) and MP (p&lt;0.01) the difference between quartiles of fertility effect was noted (MT, p=0.05 and MP, p=0.01), been higher for low fertility. It can be concluded that patterns of semen quality of batches that have distinct fertility may be similar, furthermore, there may be differences in sperm quality among batches that do not affect fertility. Thus, it takes other more accurate analysis to investigate the causes of failure on fertility of some semen batches.

O objetivo deste trabalho foi avaliar mediante testes in vitro as características espermáticas de partidas de sêmen bovino e o impacto sobre a fertilidade quando utilizadas em um programas de IATF. Foram realizados 4 experimentos que serão descritos em forma de artigos científicos. No artigo 1 comparam-se a qualidade do sêmen determinada por métodos subjetivos e objetivos para a análise espermática in vitro de partidas de sêmen bovino. Foram analisadas 80 partidas de sêmen de dois touros, através de análises convencionais (motilidade subjetiva, vigor e morfologia espermática), de acordo com os resultados foram estabelecidos 3 grupos: Alta qualidade (A), Boa qualidade (B) e Qualidade questionável (Q). Posteriormente foram analisados os resultados das análises objetivas usando sondas fluorescentes por microscopia de epifluorescência para determinar a integridade de membranas plasmática, acrossomal e potencial de membrana mitocondrial (%PIAIA); e usando o sistema computadorizado de avaliação da motilidade espermática (CASA), o efeito foi avaliado mediante ANOVA e Tukey 5%. Posteriormente, foram utilizadas 67 partidas para determinar a concordância em determinar a qualidade seminal de acordo com três métodos de avaliação seminal: (1) Análise subjetiva considerando a Motilidade subjetiva, vigor e morfologia espermática (2) Análise pelo CASA levando em consideração a MT, VCL, VSL e VAP e (3) Análise por sondas fluorescentes considerando a %PIAIA. Foram realizadas análises de correlação de Pearson e análise de concordância (significância do Kappa). No artigo 2 foram utilizadas 18 partidas de sêmen classificadas de acordo com o índice de fertilidade de cada touro em dois grupos: de Alta (n=9) e Baixa fertilidade (n=9). A classificação do escore foi proporcionada pela central com base em dados provenientes de 33.198 serviços por IATF. As partidas foram submetidas às análises convencionais, CASA, análise por sondas fluorescentes para avaliar a porcentagem de PIAIA em microscopia de epifluorescência e produção de marcadores mitocondriais por sondas fluorescentes na citometria de fluxo. Foram realizadas análises de correlação de Pearson todas as características espermáticas e os efeitos entre grupos de fertilidade foram avaliados mediante ANOVA e Tukey 5%. No artigo 3 o objetivo determinar a relação entre qualidade espermática medida pela produção de marcadores mitocondriais e integridade das estruturas espermáticas com a taxa de prenhez (TP%) resultante de um programa de inseminação artificial e tempo fixo (IATF). Neste estudo, 29 partidas de sêmen convencional usadas para inseminar 4.795 vacas da raça Nelore submetidas ao mesmo protocolo de IATF, e após o diagnóstico de prenhez foram classificadas em três grupos: Alta fertilidade (A) com TP &ge;60%, Média fertilidade (M) TP entre 53,0 e 59,9% e de Baixa fertilidade (B) TP &lt;52,2%. Doses de sêmen das mesmas partidas foram descongeladas a 37&deg;C durante 30 segundos e submetidas às análises convencionais, CASA, análise por sondas fluorescentes para avaliar a porcentagem de PIAIA em microscopia de epifluorescência e produção de marcadores mitocondriais por sondas fluorescentes na citometria de fluxo. Foram realizadas análises de correlação de Spearman para todas as características espermáticas e os efeitos entre grupos de fertilidade foram avaliados mediante ANOVA e Tukey 5%. No artigo 4 o objetivo foi determinar as subpopulações pela morfometria espermática em partidas de sêmen com diferente escore de fertilidade. Foram utilizadas 13 partidas de sêmen de 6 touros, sendo três touros de alto escore de fertilidade (n=9) e três de baixo escore de fertilidade (n=9). Duas palhetas de cada partida foram descongeladas e uma alíquota foi depositada em formol salino (4%) e posteriormente foi feito a análise por gota húmida para aquisição de 200 imagens da cabeça espermática que foram processadas pelo programa Imagem J para determinação da morfometria, os dados foram submetidos a análise multivariada de agrupamento para formação de subpopulações pelo método de Wards e posteriormente o efeito do grupo de fertilidade foi avaliado por ANOVA e Tukey 5%. Os resultados mostraram que os marcadores da função mitocondrial são bons indicadores da função e qualidade espermática; no entanto a heterogeneidade do sêmen bovino, com subpopulações espermáticas com boa e má qualidade, varia entre touros e entre partidas do mesmo touro e gera confusão na resposta de algumas análises. Determinou-se também, que a presença de subpopulações espermáticas (SBP) com variação na morfometria da cabeça (SBP1, SBP2, SBP3 e SBP4), sendo que a SBP4 foi associada com baixo escore de fertilidade. A conclusão geral é que a qualidade seminal impacta na fertilidade in vivo em bovinos. Sendo que a morfometria da cabeça espermática e funcionalidade das mitocôndrias teve maior impacto. No entanto, todas as características de qualidade espermática devem ser avaliadas em conjunto para determinar o potencial de fertilidade de uma amostra seminal. / The objective of this study was to evaluate the sperm characteristics of bovine semen and the impact on fertility when used in an IATF program. Four experiments were carried out and described in the form of scientific articles. In article 1. We compared the semen quality determined by subjective and objective methods for the in vitro sperm analysis of bovine semen. According to the results, three groups were analyzed: high quality (A), good quality (B) and high quality (A), Questionable quality (Q). Subsequently, the results of objective analyzes using fluorescence probes by epifluorescence microscopy were analyzed to determine the integrity of plasma, acrosomal membrane and mitochondrial membrane potential (% PIAIA); And using the computerized sperm motility evaluation system (CASA), the effect was evaluated using ANOVA and Tukey 5%. Afterwards, 67 matches were used to determine the agreement to determine seminal quality according to three methods of seminal evaluation: (1) Subjective analysis considering subjective motility, vigor and sperm morphology (2) Analysis by CASA taking into consideration the TM, VCL, VSL and VAP and (3) Analysis by fluorescent probes considering % PIAIA. Pearson correlation analysis and concordance analysis (Kappa significance) were performed. In article 2, 18 sets of semen were classified according to the fertility index of each bull in two groups: High fertility (n = 9) and Low fertility (n = 9). The classification of the score was provided by the central office based on data from 33,198 services by IATF. The semen were subjected to conventional analyzes, CASA, fluorescent probe analysis to evaluate the percentage of PIAIA in epifluorescence microscopy and production of mitochondrial markers by fluorescent probes in flow cytometry. Pearson correlation analyzes were performed on all sperm characteristics and effects between fertility groups were evaluated using ANOVA and Tukey 5%. In article 3 the objective was to determine the relationship between sperm quality measured by the production of mitochondrial markers and the integrity of the spermatic structures with the pregnancy rate (TP%) resulting from an artificial insemination and fixed time (IATF) program. In this study, 29 departures of conventional semen used to inseminate 4,795 Nelore cows submitted to the same IATF protocol, and after diagnosis of pregnancy were classified into three groups: High fertility (A) with TP &ge; 60%, Average fertility ( M) TP between 53.0 and 59.9% and Low fertility (B) TP &lt;52.2%. Semen doses of the same departures were thawed at 37 &deg; C for 30 seconds and subjected to conventional, CASA, fluorescent probe analysis to evaluate the percentage of PIAIA in epifluorescence microscopy and production of mitochondrial markers by fluorescent probes in flow cytometry. Spearman correlation analyzes were performed on all sperm characteristics and effects between fertility groups were evaluated using ANOVA and Tukey 5%. In article 3, 18 sets of semen were classified according to the fertility index of each bull in two groups: High (n = 9) and Low fertility (n = 9). The classification of the score was provided by the central office based on data from 33,198 services by IATF. The matches were subjected to conventional analyzes, CASA, fluorescent probe analysis to evaluate the percentage of PIAIA in epifluorescence microscopy and production of mitochondrial markers by fluorescent probes in flow cytometry. Pearson correlation analyzes were performed on all sperm characteristics and effects between fertility groups were evaluated using ANOVA and Tukey 5%. In article 4 the objective was to determine subpopulations by sperm morphometry in semen matches with different fertility scores. Seventeen sets of six bulls were used, three bulls with a high fertility score (n = 9) and three with a low fertility score (n = 9). Two straws of each set were thawed and an aliquot was deposited in saline formaldehyde (4%) and then the wet analysis was performed to acquire 200 images of the spermatic head that were processed by the Image J program to determine morphometry, the data were submitted to a multivariate cluster analysis for clusters by the Ward\'s method and later the effect of the fertility group was evaluated by ANOVA and Tukey 5%. The results showed that markers of mitochondrial function are good indicators of sperm function and quality; However, the heterogeneity of bovine semen, with good and poor quality sperm subpopulations, varies between bulls and between straws and creates confusion in the response of some analyzes. It was also determined that the presence of sperm subpopulations (SBP) with variation in head morphometry (SBP1, SBP2, SBP3 and SBP4), and SBP4 was associated with a low fertility score. The overall conclusion is that seminal quality influences in vivo fertility in cattle. Being that the morphometry of the spermatic head and functionality of the mitochondria had greater impact. However, all sperm quality characteristics should be evaluated together to determine the fertility potential of a seminal sample.

A criopreservação do sêmen resulta em danos à estrutura espermática, sendo nítida a importância da avaliação das partidas de sêmen antes de serem submetidas à inseminação artificial em tempo fixo (IATF). Todavia, nem sempre as avaliações convencionais do sêmen são suficientes para identificar partidas que possam resultar em baixa taxa de prenhez no campo, sendo necessária uma investigação mais profunda e acurada. Neste sentido, este experimento foi realizado com o objetivo de identificar partidas de sêmen que apresentam falhas na fertilidade, mesmo sendo aprovadas pelas avaliações convencionais. Foram realizadas análises convencionais (motilidade, vigor, concentração e morfologia espermática) de 72 partidas de sêmen de 22 touros antes da estação de monta. Destas, 55 partidas de 18 touros foram aprovadas para o uso na IATF, mas somente 28 partidas de 10 touros foram utilizadas. As partidas de sêmen utilizadas na IATF foram submetidas a outras avaliações: análise computadorizada da motilidade espermática (CASA), integridade das membranas plasmática e acrossomal e potencial de membrana mitocondrial (por sondas fluorescentes em microscopia de epifluorescência). Os dados foram analisados pelo Proc Mixed do SAS usando o Test T. As taxas de prenhez das diferentes partidas de sêmen variaram de 71 a 37%, sendo a média e desvio padrão das partidas de 55,57&#177;7,57%. Os dados de fertilidade a campo permitiram a separação das partidas de sêmen como de Alta (&gt;50% de prenhez) e Baixa (50% de prenhez) fertilidade, sendo comparadas quatro partidas de Alta e quatro de Baixa fertilidade. Os dados das características seminais de todas as partidas de sêmen foram submetidos à análise por boxplot e separados em quartis superior e inferior. Quando se comparou as partidas de Alta e Baixa fertilidade notou-se diferença na taxa de prenhez (p&lt;0,01), mas não foi notada diferença para motilidade (p=0,91), vigor (p=0,63), concentração (p=0,27), número de espermatozoides por palheta (NEP, p=0,27), número de espermatozoides móveis e normais por palheta (p=0,18), defeitos maiores (p=0,17), defeitos totais (p=0,43), integridade de membrana plasmática (MPI, p=0,07), alto potencial de membrana mitocondrial (AP, p=0,94), motilidade total (MT, p=0,10), VCL (p=0,80), VSL (p=0,75), VAP (p=0,88), LIN (p=0,78), STR (p=0,71) e BCF (p=0,13). No entanto, foram encontradas diferenças entre os grupos (Alta e Baixa) para defeitos menores (p=0,05), espermatozoides com integridade das membranas plasmática e acrossomal e função mitocondrial (PIAIA)/Palheta (p=0,01), integridade de acrossomo (AI, p=0,03), motilidade progressiva (MP, p&lt;0.01) e rápidos (p&lt;0,01). Quando comparados os quartis superior e inferior das características seminais foram encontradas diferenças para concentração espermática (p&lt;0,01), NEP (p&lt;0,01), número de espermatozoides móveis por palheta (p&lt;0,01), espermatozoides móveis e normais (p&lt;0,01), defeitos maiores (p&lt;0,01), defeitos menores (p=0,01), defeitos totais (p=0,05), MPI (p&lt;0,01), AI (p&lt;0,01), AP (p=0,03), PIAIA (p&lt;0,01), PIAIA/palheta (p&lt;0,01), mas esta divisão de grupos por quartis superior e inferior não apresentaram diferença sobre a fertilidade (p&gt;0,05); sendo que somente. Entretanto, para MT (p&lt;0,01) e MP (p&lt;0,01) além da diferença entre os quartis foi notado efeito da fertilidade (MT, p=0,05 e MP, p=0,01), sendo maior para os de baixa fertilidade. Pode-se concluir que os padrões de qualidade de partidas de sêmen que apresentam fertilidade distinta podem ser semelhantes, além disso, que pode haver diferença na qualidade espermática entre as partidas que não influenciam a fertilidade. Desta forma, são necessárias outras análises mais acuradas para investigar as causas de falha na fertilidade de algumas partidas de sêmen. / Semen cryopreservation results in damage to sperm structure, and it is clear the importance of evaluating the semen batches before submitted it to timed artificial insemination (TAI). However, conventional semen evaluations are not always sufficient to identify batch that may result in reduced pregnancy rate in the field, requiring a more thorough and accurate investigation. Thus, this experiment was conducted in order to identify semen batches that have gaps in fertility, even being adopted by conventional assessments. Conventional analysis (motility, vigor, concentration and morphology) were performed of 72 semen batches from 22 bulls before the breeding season. Of these, 55 batches from 18 bulls were approved for use in TAI, but only 28 batches from 10 bulls were used. Semen batches used in TAI were subjected to further assessment: computer-assisted sperm analysis (CASA), integrity of plasma and acrosomal membranes and mitochondrial membrane potential (by fluorescent probes under epifluorescence microscopy). Data were analyzed by Proc Mixed of the SAS using Test \"T\". Pregnancy rates of different semen batches ranged 71-37%, and the mean and standard deviation of the batches was 55.57 &#177; 7.57%. Field fertility data allowed the separation of the semen batches as \"High\" (&gt;50% pregnancy rate) and \"Low\" (50% pregnancy rate) fertility. It was compared four batches of \"High\" and four batches of \"Low\" fertility. Data of the seminal characteristics of all the semen batches were analyzed by boxplot and separated into upper and lower quartiles. When comparing the batches of \"High\" and \"Low\" fertility was noticed a difference in the pregnancy rate (p&lt;0.01), but was not noticeable difference in motility (p=0.91), vigour (p=0.63), concentration (p=0.27), number of spermatozoa per straw (NEP, p=0.27), number of motile and normal sperm per straw (p=0.18), major defects (p=0.17), total defects (p=0.43) plasma membrane integrity (MPI, p=0.07), high mitochondrial membrane potential (AP, p=0.94), total motility (TM, p=0.10), VCL (p=0.80), VSL (p=0.75), VAP (p=0.88), LIN (p=0.78), STR (p=0.71) and BCF (p=0.13). However, differences were found between the groups (\"High\" and \"Low\") for minor defects (p=0.05), sperm with plasma and acrosomal membranes integrity and high mitochondrial membrane potential (PIAIA)/straw (p=0.01), acrosomal integrity (Al, p=0.03) progressive motility (PM, p &lt;0.01) and rapid (p&lt;0.01). When comparing the upper and lower quartiles of the seminal characteristics differences were found for sperm concentration (p &lt;0.01), NEP (p &lt;0.01), number of motile sperm per straw (p&lt;0.01), motile and normal sperm (p&lt;0.01), major defects (p&lt;0.01), minor defects (p=0.01), total defects (p=0.05), MPI (p&lt;0.01), AI (p&lt;0.01), AP (p=0.03), PIAIA (p&lt;0.01), PIAIA/straw (p&lt;0.01), but this division groups by upper and lower quartiles showed no difference in fertility rate (p&gt;0.05). However, to MT (p&lt;0.01) and MP (p&lt;0.01) the difference between quartiles of fertility effect was noted (MT, p=0.05 and MP, p=0.01), been higher for low fertility. It can be concluded that patterns of semen quality of batches that have distinct fertility may be similar, furthermore, there may be differences in sperm quality among batches that do not affect fertility. Thus, it takes other more accurate analysis to investigate the causes of failure on fertility of some semen batches.

Made available in DSpace on 2014-06-11T19:27:43Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-08-18Bitstream added on 2014-06-13T18:56:24Z : No. of bitstreams: 1 santos_bfs_me_botfmvz.pdf: 331028 bytes, checksum: 6c1ae786a0d61ec16ca32bf05a56936e (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Universidade Estadual Paulista (UNESP) / A idade ao desmame e o estado nutricional das ovelhas no período peri-parto são aspectos determinantes para o incremento da eficiência reprodutiva e produtiva dos rebanhos ovinos. O presente trabalho teve por objetivo submeter ovelhas mestiças da raça Ile de France a dois manejos alimentares distintos (suplementadas e não suplementadas) durante o terço final da gestação e lactação, e quatro idades de desmama (60, 75, 90 e 105 dias), a fim de avaliar a variação do peso corporal e o desempenho reprodutivo das matrizes. Um total de 434 ovelhas foi avaliado quanto ao peso corporal (kg) no momento do parto (PP) e ao desmame (PD), escores de condição corporal ao parto (ECCP) e ao desmame (ECCD) e taxa de prenhez (TP). Observou-se que ovelhas suplementadas apresentaram melhor condição corporal ao parto e ao desmame (3,55 e 3,39, respectivamente) quando comparadas às não suplementadas (3,45 e 3,32, respectivamente) (P<0,05). O maior peso ao desmame observado em ovelhas suplementadas (53,2 kg) confirma um menor desgaste, pois perderam menos peso, em relação às não suplementadas (45,3 kg) (P<0,05). Durante o primeiro ano, observou-se que o manejo nutricional e a idade ao desmame influenciaram positivamente a taxa de prenhez em ovelhas que desmamaram aos 60 e 75 dias (71,88 % e 67,86 %, respectivamente) (p<0,05). Durante todo o período do ano 2 não foi observada diferença significativa na taxa de prenhez. Entretanto, o escore de condição corporal ao desmame das ovelhas do ano 1 estavam significativamente menores que o das ovelhas do ano 2 (3,30 e 3,48, respectivamente). Dessa forma, podemos indicar que a suplementação alimentar teve efeito quando as ovelhas estavam com condição corporal mais baixa, tornando-se importante a utilização de suplementação alimentar para ovelhas em terço final de gestação e durante a lactação, sendo altamente benéfica dentro do sistema de produção.

Made available in DSpace on 2014-06-11T19:27:43Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-11-09Bitstream added on 2014-06-13T19:56:26Z : No. of bitstreams: 1 lopes_cn_me_botfmvz.pdf: 505921 bytes, checksum: 14aad0f6b2df70711c09f6e1b5b4acc9 (MD5) / Universidade Estadual Paulista (UNESP) / Foram realizados quatro experimentos com o objetivo de avaliar possíveis mecanismos relacionados ao aumento da prenhez com a utilização de ácidos graxos poliinsaturados (PF). No exp. 1 foram utilizadas 51 vacas multíparas Nelore não lactantes, ovuladas, para avaliar se PF alteram a produção de progesterona (P4) e o momento da luteólise. No exp. 2 foram utilizadas 43 vacas multíparas Nelore não lactantes, ovuladas, para avaliar se PF alteram a sensibilidade do corpo lúteo de seis dias a aplicação de prostaglandina (i.m. 12,5mg de dinoprost trometamina, Lutalyse). No exp. 3 foram utilizadas 27 vacas multíparas Nelore, ovuladas, com 30 a 40 dias pós parto para avaliar se PF diminuem a incidência de ciclo curto. Os tratamentos utilizados foram: grupo controle (100g mineral + 100g milho + 100g caolin vaca dia); grupo SF (100g mineral + 100g milho + 100g/vaca/dia de Megalac (7-9% C18:2; Arm&Hammer a Church&Dwight Company, EUA); grupo PF (100g mineral + 100g milho + 100g/vaca/dia de Megalac-E QGN, Brasil: 40-42% de C18:2; 2-3% de C18:3). No exp. 4 (1457 vacas multíparas Nelore) foi avaliado se a suplementação com PF pós IATF por diferentes períodos altera a taxa de prenhez. Nos experimentos 1, 2 e 3 não foi detectado efeito de PF nas concentrações de P4 durante o período avaliado, no momento da luteólise e na incidência de ciclo curto (P>0,1). Ao se agrupar os dados do Exp 1 e 2, a concentração de P4 (P=0,01) no dia 6 foi maior nos animais suplementados com PF em relação ao SF e controle (4,45; 3,25; 3,48 ng/ml, respectivamente; EPM=0,278). No experimento 4 vacas recebendo PF por 21 e 28 dias após a IATF tiveram maior (P<0,05) taxa de prenhez (50,38%) quando comparadas com os outros tratamentos agrupados (42,38%). Não foi detectado diferença entre os tratamentos PF21 (50,99%) e PF28 (49,81). Os resultados em conjunto mostram que apesar de suplementação... / Four experiments were designed to evaluate the possible mechanisms related to the increased pregnancy rates in cows supplemented with PF. In Experiment 1, 51 ovulated non-lactating Nelore multiparous cows were used to evaluate if PF supplementation affects circulating progesterone (P4) concentrations and timing of luteolysis. In Experiment 2, 43 ovulated non-lactating Nelore multiparous cows were used to evaluate if PF supplementation alters the sensitivity of a 6-d corpus luteum to exogenous prostaglandin treatment. In Experiment 3, 27 ovulated postpartum Nelore cows were used to evaluate if PF supplementation affects the incidence of premature luteolysis. Beginning at the d of estrus, the daily treatments in these 3 experiments were: Control (0.1 Kg Mineral + 0.1 Kg corn + 0.1 Kg kaolin); SF (0.1 Kg Mineral + 0.1 Kg Megalac-S® [7-9% linoleic acid] + 0.1 Kg corn), this group was used just in experimental 1 and 2; PF (0.1 Kg Mineral + 0.1 Kg Megalac-E® [40-42% linoleic acid; 2-3% linolenic acid] + 0.1 Kg corn). In Experiment 4, we evaluated if the length of PF supplementation in different times after timed-AI alters the pregnancy rate. No effect was detected on P4 concentration, luteolysis or short cycle (P>0.1), but when the cows of exp. 1 and 2 were grouped had higher (P=0.01) concentration of P4) on cows that were supplemented with PF compared with Sf or control (4.45; 3.25; 3.48 ng/ml, respectively; SEM=0,278. Cows supplemented with PF during 21 (PF21,) or 28 d post- AI (PF28) had higher pregnancy rates (50.38%; P < 0.05) than cows from other treatments (42.38%). There was no difference between PF21 (50,99%) and PF28 (49,81%) treatments. These experiments indicated that the possible mechanism for greater conception with PF supplementation post-AI may be due to effects on embryo development, animals supplemented for more than 21d had greater pregnancy rates.

Orientador: Edson Ramos de Siqueira / Banca: Ivanete Susin / Banca: João Ricardo Alves Pereira / Abstract: Weaning age and nutritional status of ewes during peri-partum period are import for increasing reproduction and production performance. The aim of this study was to investigate the effect of two different nutrition management (supplemented and unsupplemented) during late pregnancy and lactation, and four ages of weaning (60, 75, 90 and 105 days) on growth and reproductive performance of ewes. A flock of 434 Ile de France cross bred ewes was evaluated for body weight (Kg) and body condition score (ECC) at lambing (PP and ECCP) and weaning (PD and ECCD), and pregnancy rate (TP). Results showed better body condition score in supplemented group at lambing and weaning (3.55 and 3.39, respectively) than that of ewes unsupplemented (3.45 and 3.32, respectively) (p<0.05). In addition, supplemented ewes showed a higher body live weight than unsupplemented ewes (53.2 Kg and 45.3 Kg, respectively) because the supplemented ewes might not get down fast (p<0.05). During the first year of this experiment, the pregnancy rate was positively affected by nutrition and weaning age at 60 and 75 days (71.88 % and 67.86 %, respectively) (p<0.05). It was not observed significant statistical difference in pregnancy rate during the second year of this experiment. However, the body condition score of ewes at weaning in the first year was lower than that of ewes in the second year (3.30 and 3.48, respectively). Therefore, it suggests that nutrition had a great effect on ewes with low body condition. In conclusion, it is important for great results on intensive production system to supplement ewes during late gestation and weaning period. / Resumo: A idade ao desmame e o estado nutricional das ovelhas no período peri-parto são aspectos determinantes para o incremento da eficiência reprodutiva e produtiva dos rebanhos ovinos. O presente trabalho teve por objetivo submeter ovelhas mestiças da raça Ile de France a dois manejos alimentares distintos (suplementadas e não suplementadas) durante o terço final da gestação e lactação, e quatro idades de desmama (60, 75, 90 e 105 dias), a fim de avaliar a variação do peso corporal e o desempenho reprodutivo das matrizes. Um total de 434 ovelhas foi avaliado quanto ao peso corporal (kg) no momento do parto (PP) e ao desmame (PD), escores de condição corporal ao parto (ECCP) e ao desmame (ECCD) e taxa de prenhez (TP). Observou-se que ovelhas suplementadas apresentaram melhor condição corporal ao parto e ao desmame (3,55 e 3,39, respectivamente) quando comparadas às não suplementadas (3,45 e 3,32, respectivamente) (P<0,05). O maior peso ao desmame observado em ovelhas suplementadas (53,2 kg) confirma um menor desgaste, pois perderam menos peso, em relação às não suplementadas (45,3 kg) (P<0,05). Durante o primeiro ano, observou-se que o manejo nutricional e a idade ao desmame influenciaram positivamente a taxa de prenhez em ovelhas que desmamaram aos 60 e 75 dias (71,88 % e 67,86 %, respectivamente) (p<0,05). Durante todo o período do ano 2 não foi observada diferença significativa na taxa de prenhez. Entretanto, o escore de condição corporal ao desmame das ovelhas do ano 1 estavam significativamente menores que o das ovelhas do ano 2 (3,30 e 3,48, respectivamente). Dessa forma, podemos indicar que a suplementação alimentar teve efeito quando as ovelhas estavam com condição corporal mais baixa, tornando-se importante a utilização de suplementação alimentar para ovelhas em terço final de gestação e durante a lactação, sendo altamente benéfica dentro do sistema de produção. / Mestre