Dissertations / Theses on the topic 'Domestic animals – Reproduction'

Feeds often contain molybdenum (Mo) and sulfur (S) in excess of recommended allowances for adequate copper (Cu) absorption by ruminant livestock. Two randomized-block experiments were conducted using lambs given a cereal-based diet (90% of dry matter (DM) intake), containing 8 mg Cu, 0.7 mg Mo and 2.1g S per kg DM, that was either unsupplemented or supplemented with ammonium molybdate or sodium sulfate alone or in combination, to determine the effects of Mo, S and Mo+S on growth, hematology, serum Cu and Mo concentrations and reproductive function. Short term intake (4 weeks) by rams, aged 18 and 20 weeks, of 26 mg Mo alone or in combination with 2g S per kg DM, had no effect on growth, hematology or the concentration of total Cu in serum (TCu). However, supplemented groups had lower (P<0.05) concentrations of serum Cu soluble in trichloroacetic acid (TCA-Cu), but the group given Mo+S had the highest (P<0.05) concentration of serum residual Cu (RCu). Luteinizing hormone (LH) peak amplitude was affected by age*diet interaction (P<0.05), because of high amplitude LH peaks in the serum of lambs, aged 24 wks, given Mo alone. Testosterone serum secretory profiles did not differ among diet groups, but testosterone peak frequency was higher (P<0.05) for older ram lambs. Long term intake (32-39 wks) by ewe, ram and wether lambs (gonadal influence) of 12 mg Mo, or 2g S alone or combined per kg DM also had no effect on hematology or TCu. Food intake and liver weights were higher (P<0.05), but TCA-Cu was lower (P<0.05) for groups given S. Serum concentrations of total Mo (TMo) were higher (P<0.05) for Mo-supplemented groups, but RCu was highest only for the Mo+S group. Growth was affected by Mo*S*gonadal influence*time interaction (P<0.05); until autumn, the body weight of Mo-supplemented groups were higher than those of ram and wether lambs given Mo+S. Ovarian or testicular functions were delayed more severely for Mo- than Mo+S-supplemented groups. Mean concentrations of LH were affected by Mo*S*gonadal influence*time interaction (P<0.05); the mean LH values of Mo-supplemented ram and ewe lambs were higher than the Mo+S-supplemented group, whereas a reverse trend was observed for wether lambs. The amplitude of LH peaks was affected by Mo*S*gonadal influence*time interaction (P<0.05); for ram lambs, LH peak amplitude was higher for Mo than Mo+S-supplemented groups, whereas for wether and ewe lambs the reverse trend was noted. Mean concentrations of serum Cortisol and Cortisol peak amplitude were affected by Mo*S interaction (P<0.05); the mean Cortisol concentration and peak amplitude for the Mo-supplemented group were higher than those for the Mo+S-supplemented group. The frequency of LH and Cortisol peaks did not differ (P>0.10) among diet groups. In conclusion, Mo supplementation of cereal-based diets containing a high concentration of Cu did not adversely affect lamb growth. However, the effect of high dietary Mo on reproductive function appears to depend on the dietary level of S and the induction of high serum RCu (thiomolybdate). Further investigations on the effects of Mo and thiomolybdate on endocrine function may provide a nutritional basis for improving reproductive efficiency in ruminants.
Land and Food Systems, Faculty of
Graduate

Orientador: Maria Denise Lopes
Banca: Frederico Ozanan Papa
Banca: Jussara Tebet
Resumo: A análise dos efeitos da refrigeração sobre a congelabilidade da célula espermática da cauda do epidídimo de gatos domésticos foram avaliados nesse experimento. Após prévia refrigeração a 5O C por 24 horas alíquotas de sêmen foram submetidas a criopreservação e posteriormente descongeladas para análise morfofuncional. 15 animais foram submetidos a orquiectomia de conveniência e seus epidídimos foram manipulados para obtenção de amostras que foram suspensas em meio TE (TRIS/Equex/Gema de Ovo), analisadas, refrigeradas e congeladas. Amostras referentes ao grupo controle foram congeladas imediatamente após a colheita. A analise morfofuncional consistiu em motilidade, vigor, morfologia espermática e Integridade de membrana. Após a descongelação os resultados dos grupos controle e trabalho foram analisados estatisticamente e observou-se que não existe diferença estatística para nenhuma das variáveis analisadas. Embora tenha sido observada diferença estatística entre os momentos pós-colheita e pós-refrigeração por 24 horas para as variáveis motilidade, vigor, integridade de membrana e defeitos primários, nenhuma variável diferiu entre as amostras congeladas pós-colheita e as amostras congeladas pós-refrigeração prolongada. Tais resultados demonstram que um transporte refrigerado a 5O C por 24 horas não interfere sobre a congelabilidade de sêmen do epidídimo de gatos domésticos. Do ponto de vista da conservação tais resultados abrem portas para um melhor fluxo de gametas de espécies selvagens para centrais de biotecnologia permitindo um melhor aproveitamento e conservação deste material.
Abstract: The present study analyzed the effect of cooling on sperm cells obtained from the epididymis tail from domestic cats in respect to their capability for freezing. The treatment group had their samples submitted to freezing after being cooled at 5° C during 24 hours, and th~ control group samples were frozen immediately after collection, Samples from both groups were then thawed and submitted to mo rphofunctiona I analysis. Fifteen animais were submitted to a convenient routine orchiectomy and their epididymis were processed to obtain a semen samples which were suspended in TE medium (TRIS/Equexl egg yolk). The samples were then analyzed, coo,led and frozen. The morphofunctional analysis consisted of sperm motility, vigor, morphology and membrane integrity. After thawing, the semen samples from treatment and control groups were analyzed and no statistic difference was found among ali variables described above.
Mestre

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
A análise dos efeitos da refrigeração sobre a congelabilidade da célula espermática da cauda do epidídimo de gatos domésticos foram avaliados nesse experimento. Após prévia refrigeração a 5O C por 24 horas alíquotas de sêmen foram submetidas a criopreservação e posteriormente descongeladas para análise morfofuncional. 15 animais foram submetidos a orquiectomia de conveniência e seus epidídimos foram manipulados para obtenção de amostras que foram suspensas em meio TE (TRIS/Equex/Gema de Ovo), analisadas, refrigeradas e congeladas. Amostras referentes ao grupo controle foram congeladas imediatamente após a colheita. A analise morfofuncional consistiu em motilidade, vigor, morfologia espermática e Integridade de membrana. Após a descongelação os resultados dos grupos controle e trabalho foram analisados estatisticamente e observou-se que não existe diferença estatística para nenhuma das variáveis analisadas. Embora tenha sido observada diferença estatística entre os momentos pós-colheita e pós-refrigeração por 24 horas para as variáveis motilidade, vigor, integridade de membrana e defeitos primários, nenhuma variável diferiu entre as amostras congeladas pós-colheita e as amostras congeladas pós-refrigeração prolongada. Tais resultados demonstram que um transporte refrigerado a 5O C por 24 horas não interfere sobre a congelabilidade de sêmen do epidídimo de gatos domésticos. Do ponto de vista da conservação tais resultados abrem portas para um melhor fluxo de gametas de espécies selvagens para centrais de biotecnologia permitindo um melhor aproveitamento e conservação deste material.
The present study analyzed the effect of cooling on sperm cells obtained from the epididymis tail from domestic cats in respect to their capability for freezing. The treatment group had their samples submitted to freezing after being cooled at 5° C during 24 hours, and th~ control group samples were frozen immediately after collection, Samples from both groups were then thawed and submitted to mo rphofunctiona I analysis. Fifteen animais were submitted to a convenient routine orchiectomy and their epididymis were processed to obtain a semen samples which were suspended in TE medium (TRIS/Equexl egg yolk). The samples were then analyzed, coo,led and frozen. The morphofunctional analysis consisted of sperm motility, vigor, morphology and membrane integrity. After thawing, the semen samples from treatment and control groups were analyzed and no statistic difference was found among ali variables described above.

Active immunisation against GnRH inhibits reproductive function by inducing a hypogonadotropic condition associated with gonadal atrophy. Despite economic, ethical and environmental advantages of GnRH immunisation in cattle over conventional castration methods, the technology has not yet been commercially adopted. Primarily because of the requirement for numerous booster vaccinations because of the reversibility of physiological effects, the commercial efficacy of immunocastration is currently poor. However, neonatal GnRH immunisation in sheep can result in a permanent suppression of reproduction (Brown et al., 1994; 1995; Clarke et al., 1998). These findings and a study in pigs (Molenaar et al., 1993) indicate that, the hypothalamic/pituitary gland unit (HPU) may be particularly susceptible to GnRH antibodies during a specific window of development in the pre-pubertal animal, but no long-term studies in cattle have been conducted. Therefore the primary objective of this project was to determine the effect of neonatal immunisation against GnRH in cattle. Beef cross bull (n=9; Chapter 3) and heifer calves (n=9; Chapter 4) were vaccinated against a newly developed (Pfizer®) GnRH construct vaccine at -2, 6 and 13 weeks of age. Nine calves of each sex served as negative controls, receiving saline injections only. The GnRH vaccine had proved effective (Dr. A.R. Peters, personnel communication 2000) in inducing immune responses and reducing variation between animals in unpublished industrial studies, compared to earlier vaccines, and hence was reasoned to be capable of raising GnRH antibodies despite the relative immaturity of the neonatal immune system. Following vaccination, circulating GnRH antibodies and reproductive hormones, such as FSH (Chapters 3 and 4), testosterone (Chapter 3), progesterone (to assess onset of puberty) and oestradiol (Chapter 4) were measured and additional intensive serial bleeds were carried out to assess LH parameters up to and beyond puberty (puberty defined by testes circumference in bulls). Gonadal (antral follicles and testes growth) and accessory gland development was quantified throughout the trial using ultrasound scanning. Sexual behaviour (Chapter 3) was studied from 38 weeks of age, while an assessment of sperm quality (Chapter 3), and anabolic response to vaccination was also performed post-mortem (Chapters 3 and 4). GnRH immunisation in neonatal calves did not permanently impair reproduction. A temporary suppression in reproductive function was evident through the disruption of pituitary gland function, as indicated by a reduction of LH pulse amplitude and mean plasma LH concentrations (Chapters 3 and 4). In addition, a reduction in medium- sized follicle numbers, testes growth, plasma testosterone concentration, vesicular gland length and juvenile aggression occurred. Some beneficial anabolic effects were observed e.g., carcass composition grades. Changes all occurred subsequent to increased GnRH antibody titres in immunised cattle. Despite some evidence of prolonged effects on LH amplitude and circulating testosterone after anti-GnRH titres had dissipated, all inhibited parameters, except carcass quality, returned to levels comparable to control animals by 72 weeks of age. No treatment effects on FSH concentrations, large follicle numbers, reproductive tracts (post mortem) or peri- and post-pubertal behaviours were observed following treatment. Sperm morphological abnormalities tended to be more prevalent in GnRH immunised bulls. A significant increase in GnRH antibody titres occurred at -23 weeks of age (Chapter 4), this may have been a rebound in antibody titre, possibly caused by an anti-idiotype immune response (antibody response to GnRH antibodies), or due to significant maturational changes in immune function at this time causing a delayed response to vaccination. Alternatively a novel "auto-immune" response may have been detected, which if confirmed/repeatable might be incorporated into an immunisation protocol to act as a "self-booster". However, no previous reports of such an event have been published and further investigation is urgently required. A more prolonged or permanent suppression of reproductive function may be possible following an earlier, greater and more sustained elevation of antibody titres during the neonatal period. Further development of GnRH vaccines and/or protocols (prime-boost, cytokine modulation vaccines, concomitant passive and active immunisation and pregnant cow GnRH vaccination), and studies of performance and GnRH antibody mechanism(s) of action in cattle are required. Chapters 3 and 4 provide a comprehensive study on pubertal development and neonatal GnRH vaccination, thus contributing significantly to knowledge in these fields. Currently, the vaccine used in this trial may be used to delay puberty in older calves or transiently suppress reproductive function to aid management. The economical viability of animal production systems such as beef and lamb are closely related to rates of reproduction. The Fec B gene in ewes increases ovulation rate and litter size, possibly through the development of precocious follicles, which can switch their primary dependence from FSH to LH. As a result, more follicles are selected to continue growth to an ovulatory size. The precise mechanisms by which these processes occur have recently been shown to involve oocyte follicle interactions (see section 1.1.5). Follicle development is modulated by GHIIGF and inhibin, however attempts to increase follicular development and ovulation through active inhibin immunisation alone have been variable and hence not commercially attractive. To develop successful protocols to induce twin ovulations in cows· and ewes, without superovulation, a clearer, more details understanding of follicullogenesis is required. The objective of the current study was to better understand these mechanisms through investigating interactions of GH/GF and inhibin in the ovary, follicle development, steroidogenesis, and receptor populations using an anoestrous sheep model. Spring born Mule x Charolais ewe lambs were actively immunised (n=8) against porcine inhibin α-C 1-26 peptide conjugated to KLH in NUFCA (primary and 3 boosters (NUFA», while 8 served as negative controls. Seven days following the final booster, the ewes were subdivided to give four groups: (1) controls + saline (n=4); (2) controls + rbGH (4ml s.c; 1mg. mr1; n=4); (3) inhibin immunised + saline (n=4); and (4) immunised + rbGH (n=4). Recombinant bovine growth hormone (rbGH) was given (Lm.) for 6 days. On day 4 GnRH (Receptal®; 1 ml) was injected s.c, to all animals to initiate the beginning of a new follicular wave. Blood samples were collected fortnightly to measure inhibin antibody titres, IGF-I, FSH and steroids. On the seventh day ensuing slaughter serum antibodies and ovaries were harvested. Left ovaries were intended for ISH (mRNA for P450arom) and/or immunohistochemical analysis. Follicles from right ovaries were dissected out, counted, measured and cultured in M199 at 37°C for 2 hours. Culture media was then assayed for oestradiol. Follicle shells were stored at -180°C for LH receptor binding studies. This work reports on the influence of different treatments on follicle populations. All immunised animals produced antibodies, which bound to 1251-inhibin. Using ANOVA to compare treatments it was observed that, Inhibin immunisation significantly (P3.5mm in diameter, but did not affect the smaller <3.5mm population. In contrast, rbGH administration led to a significant (P3.5mm follicle numbers. These findings are in agreement with previous research. The molecular studies of left ovaries are not presented herein as due to time constraints the work was not completed and is currently on going. In conclusion, additional results of this study are required to meet the objectives of the experiment. Further research is required on dominant follicle selection if superovulatory programmes in both livestock and humans are to be more precisely controlled and readily accepted.

On sait que chez les oiseaux chanteurs le chant est le principal caractère sexuel secondaire et qu’il a une influence majeure sur la reproduction. Le chant peut influencer une femelle dans le choix de son partenaire et agir sur la façon dont elle s’investira pour optimiser chaque tentative de reproduction. Le propos général de cette thèse est d’étudier l’impact du chant sur la reproduction chez le canari. Dans une première partie, nous avons étudié l’importance de ce chant par rapport à d’autres stimuli susceptibles d’influer sur le comportement sexuel des femelles. Nous avons montré que dans le choix de leur partenaire, les femelles prennent en compte la relation de dominance entre les mâles quand elle se manifeste par le chant et non pas par des interactions physiques entre eux. Dans une seconde partie, nous nous sommes demandés si le chant d’un mâle pouvait permettre à la femelle d’apprécier son degré de fertilité. Nous avons montré que meilleure était la qualité du chant (longueur et rythme d’émission), meilleure était la qualité du sperme, suggérant que le chant pouvait être un indicateur de fertilité. Dans la dernière partie, nous avons étudié l’impact du chant sur le système reproducteur de la femelle. Nous avons trouvé que des phrases moyennement attractives chantées par un mâle induisaient un dépôt plus important de testostérone dans l’œuf par la femelle, alors que les mêmes phrases produites par ordinateur, donc en l’absence de mâle, n’avaient pas d’effet. Cela suggère que si le chant n’est que moyennement attractif, des stimuli supplémentaires sont requis pour entraîner une augmentation du dépôt de testostérone dans l’œuf. Enfin, nous avons présenté le projet de recherche en cours. Il s’agira d’étudier l’activation des différentes régions du cerveau en réponse au chant. Ce projet pourrait permettre d’identifier les voies neuronales impliquées dans le dépôt différentiel de testostérone dans l’œuf en réponse à ce stimulus
In songbirds, song is known to be the primary sexually selected trait with extensive influences on reproduction. Song can influence a female in her choice for a mate and can also influence how she adjusts her own investment in order to optimize each reproductive attempt. The general aim of this thesis was to investigate the impact of song on reproduction in canary. In the first section, we investigated the importance of song compared to other cues for females to direct their sexual behavior. We showed that females pay attention, in order to direct this sexual behavior, to dominance manifested via song and not via physical interactions. In a second part of this thesis, we wanted to know if song could be a cue for the female that would allow her to judge the fertilization capacities of a male. We showed that the better the song quality was (song length and emission rate) the better the quality of sperm was, suggesting that song could signal male fertility. In the last section, we investigated the impact of song on the female reproductive system. We found that mildly attractive song phrases sung by a live male induced the female to deposit more testosterone in her eggs. However, these same songs, but computer generated, (i. E. Without the presence of the male), did not induce females to deposit more T. This suggests that other cues, in addition to mildly attractive song, may be necessary to increase T deposit. Finally, we present an on-going project, investigating the activation of different brain regions in response to song. This project could help to elucidate a neuronal pathway involved in the differential testosterone deposit in eggs in response to song

Accurately examining animal endocrine profiles pose unique challenges due to possible human interaction influencing basal values. Standard methods of gathering information about an animals endocrine status are often dependent upon restraint and use of invasive methodologies. However to accurately monitor the influence management practices, blood sampling sometimes requires that hormone measurements be observed from animals in a relaxed state. To this end, methods for non-invasive monitoring (NIM) are greatly needed to obtain basal endocrine measurements. Such methods include fecal collections followed by hormone extraction, and remote sampling technologies for obtaining blood samples without handling. The overall objective of this study was to use NIM techniques to effectively collect and monitor hormone profiles from domestic and non-domestic species in an effort to more completely understand stress responses and reproductive cyclicity in animals in which handling may not be possible or desired.

My PhD project addressed the paternal and maternal contribution associated with correct early embryonic development, focusing on microRNAs (miRNAs, short non-coding RNAs regulating gene expression at post-transcriptional level) involved in controlling reproductive function in cattle. To this end, next generation sequencing (NGS) methods and bioinformatic analysis pipelines were set up and validated using low quantities of starting materials, e.g. blastocysts and oocytes. The role of miRNAs carried by spermatozoa during early embryogenesis was investigated by assessing in vitro produced blastocysts from semen of bulls with high and low blastocyst rates. Moreover, follicular fluid and oocyte miRNAs were analyzed in order to unravel molecular mechanisms involved in poor fertility cows. In particular, a model based on low and high antral follicle count ovaries was used to study the influence of ovarian function on oocyte developmental competence. The results indicated that: 1) sperm miRNAs may impact embryo developmental competence affecting the expression of gene networks involved in several cellular processes including cell adhesion, communication and metabolism of the blastocyst; 2) the different oocyte quality is associated with a different miRNA blueprint in both follicular fluids and oocytes through the mis-regulation of biological processes critical for oocyte competence. In conclusion, these studies contributed to improve our knowledge on the function of miRNAs in the sperm, oocytes and pre-implantation embryos. Several miRNAs were identified as potential biomarkers of blastocyst and oocyte competences, which will be valuable to optimize Reproductive Biotechnologies.

Animals that live in a social group are often organized in a hierarchy with rank determining access to resources. Maintaining a dominant position requires a high rate of energetically expensive aggressive displays and physical exertion. Lab based winnerloser studies, often conducted with individuals from non-social species, have shown that subordinates have higher stress hormone levels than dominant individuals (subordinatestress hypothesis). However, in carnivorous animals that are cooperative breeders, displays of aggression are associated with elevated stress hormone levels (dominancestress hypothesis). The effect of reproductive state on dominance and stress is not addressed within either of these hypotheses. The purpose of this study was to examine stress level in relation to dominance rank and reproductive state in a non-cooperative breeder and herbivore, the domestic horse. As rank and reproductive state can affect behavior, I examined activity budgets, behavioral patterns, and social interactions, as well as the proximity and identity of neighbors in the social group. At two facilities in Kentucky, I recorded the social interactions and measured fecal glucocorticoids in pastured, female horses that were either lactating or non-lactating. While fecal glucocorticoid level did not differ between reproductive state and rank, activity behavior demonstrated significant differences between reproductive states. Higher energetic requirements of lactation were reflected in significantly longer bouts of eating and significantly less time spent alert and socializing. The non-cooperative social nature of horses does not limit their reproduction or resource acquisition based upon rank, and therefore does not fit with the dominance-stress hypothesis or subordinate-stress hypothesis and instead offers the alternative of an independent-stress hypothesi

Thesis (Msc Food Sc (Food Science))--University of Stellenbosch, 2006.
The aim of this study is to determine the effect of crossbreeding Zimbabwean Blue (ZB) and South African Black (SAB) ostriches on the morphological, physical, chemical and sensory quality of the meat. However, it is also necessary to determine the reproductive performance of these genotypes to scientifically support decisions made in the ostrich industry. In relation to reproductive traits and body measurements influencing these traits, results from the study suggested that ZB birds are between 9 and 15% heavier than their SAB contemporaries. Regarding SAB females, egg production was 47% higher, levels of shell deaths were lower, percentage of eggs not incubated was lower (P<0.01) and 84% more (P<0.01) chicks were produced in a season compared to their ZB contemporaries. Mates of SAB males produced a higher (P≤0.05) percentage of eggs not incubated and higher shell death percentages than the mates of ZB males. It has to be conceded that ZB females had a lower reproduction than SAB females, limiting the application of this genotype as a dam line in crossbreeding systems. With regard to morphological properties, the pure Blue genotype in comparison to the pure Black genotype differed significantly (P≤0.05), with 16 kg for live weight, 8.3 kg for carcass weight and 3.5 kg for leg weight. However, when comparing carcass yields (expressed as %) there were no significant differences (P>0.05) between genotypes. The M. gastrocnemius, M. femorotibialis accessorius, M. iliotibialis cranialis, M iliotibialis lateralis, M. iliofibularis and M. iliofemoralis showed significant genotype differences (P≤0.05) for individual muscle weight. When comparing the physical meat quality characteristics between the pure Blue genotype and the pure Black genotype, 70% of the muscles were higher (P≤0.05) in pH24, 50% of the muscles were redder (P≤0.05) and significantly less (P≤0.05) saturated in colour, 67% of the muscles had a lower (P≤0.05) percentage drip loss and 50% of the muscles had a lower (P≤0.05) percentage cooking loss. No significant (P>0.05) genotype differences were observed regarding the sensory quality of the meat. Regarding chemical meat quality characteristics, the percentage of moisture was higher and the percentage of lipid was lower for eight of the ten muscle groups from the pure Blue genotype. No significant differences (P>0.05) were found between genotypes or between muscles regarding the percentage of protein present in the meat. The highest (P>0.05) content of soluble collagen, myoglobin and cholesterol was found in the Blue x Black genotype, whereas the lowest percentage of the latter constituents was found in the pure Blue genotype. For the pure Black genotype the concentration of saturated fatty acids in the meat was lower (P≤0.05), the concentrations of total unsaturated fatty acids and desirable fatty acids in the M. illiofibularis were the highest (P≤0.05), while the concentration of monounsaturated fatty acids was also higher (P≤0.05) in both muscles of this genotype compared to the other two genotypes. Regarding both fat depots, the pure Black genotype had a lower (P≤0.05) concentration of saturated fatty acids, a higher (P≤0.05) concentration of monounsaturated fatty acids and total unsaturated fatty acids and a higher (P≤0.05) polyunsaturated:saturated fatty acid ratio. The percentage of desirable fatty acids in the abdominal fat depot was significantly higher (P≤0.05) for the pure Black and the Blue x Black genotype. In conclusion, crossbreeding between SAB and ZB ostriches seems to be a viable option to produce larger birds with more meat, without negatively affecting the overall quality of the meat.

[Truncated abstract] Despite the costs associated with mating, females of many taxa solicit multiple mates during a single reproductive event (polyandry). Polyandry is clearly adaptive when females gain direct benefits from males at mating. However, polyandry has also been shown to increase female fitness in the absence of direct benefits. Thus, a number of genetic benefit hypotheses have been developed to account for the origin of this behaviour. Although not mutually exclusive, a distinction lays between genetic benefits that propose defense against reproductive failure (nonadditive genetic effects), and those that propose benefits from intrinsic sire effects (additive genetic effects). Nonadditive genetic benefits of polyandry have been documented in a number of species; by soliciting multiple mates females can avoid inbreeding and other forms of incompatibility between parental genotypes. Polyandry may also increase female reproductive success when genetically superior males have greater success in sperm competition, and produce better quality offspring. An inevitable consequence of polyandry is that sperm from rival males will overlap in the female reproductive tract and compete to fertilise the ova. The outcome of sperm competition is typically determined by bias in sperm use by the females, interactions between parental genotypes, and ejaculate characteristics that provide a fertilisation advantage. Thus, sperm competition is recognised as a persuasive force in the evolution of male reproductive traits. Comparative analyses across species, and competitive mating trials within species have suggested that sperm competition can influence the evolution of testis size and sperm production, and both sperm form and sperm function. ... After six generations of selection I observed phenotypic divergence in litter size - litter size increased in the polyandrous lines but not in the monandrous lines. This result was not attributable to inbreeding depression, or environmental/maternal effects associated with mating regime. Genetic benefits associated with polyandry could account for this result if increased litter size were attributable to increased embryo survival. However, males from the polyandrous lineages were subject to sperm competition, and evolved ejaculates with more sperm, suggesting that evolutionary increases in litter size may in part be due to improved male fertility. Finally, Chapter Five is an investigation of the natural variation in levels of polyandry in the wild, and the potential for sperm competition to drive macroevolutionary changes in male reproductive traits among geographically isolated island populations of house mice. I sampled seven island populations of house mice along the coast of Western Australia and, by genotyping pregnant females and their offspring, determined the frequency of multiply sired litters within each population. I applied the frequency of multiple paternity as an index of the risk of sperm competition, and looked for selective responses in testis size and ejaculate traits. I found that the risk of sperm competition predicted testis size across the seven island populations. However, variation in sperm traits was not explained by the risk of sperm competition. I discuss these results in relation to sperm competition theory, and extrinsic factors that influence ejaculate quality.

CAPES
O presente trabalho teve como objetivo estimar parâmetros genéticos, em uma população de bovinos da raça Nelore, para as características probabilidade de parto até os 39 meses de idade (PC39), probabilidade de permanência no rebanho (stayability), considerando três partos até 5,2 (STAY5,2) e três partos até 6,2 anos (STAY6,2), e os quatro primeiros intervalos entre partos (IEP1, IEP2, IEP3 e IEP4), tratando- as como variáveis categóricas, e para os perímetros escrotais à desmama (PED) e ao sobreano (PES), tratando-as como variáveis contínuas. Foram usados procedimentos bayesianos, em análises unicaracterística e bicaracterística, por meio dos programas THRGIBBS1F90 e GIBBS2F90. Os dados utilizados pertencem ao programa de melhoramento genético Geneplus - Embrapa. As herdabilidades para PC39, STAY5,2, STAY6,2 , PED e PES em análise unicaracterística foram 0,15, 0,20, 0,23, 0,28 e 0,36, respectivamente. Para o intervalo entre partos as estimativas de herdabilidade, obtidas em análise unicaracterística, para IEP1, IEP2, IEP3 e IEP4 foram 0,13 0,13, 0,12 e 0,17, respectivamente. Estes valores indicam que os ganhos por seleção direta em quaisquer um dos intervalos seriam pequenos.Na análise bicaracterística as herdabilidades para PC39 e PES foram 0,14 e 0,38,respectivamente. As estimativas obtidas foram de baixa magnitude, indicando que os ganhos por seleção direta em qualquer uma das características mensuradas nas fêmeas seriam baixos. Já o PED e PES apresentaram maiores valores de herdabilidades. A correlação entre PES e PC39 foi positiva e de baixa magnitude (0,25), sugerindo que a seleção para o aumento do PES de touros possivelmente levaria a pequenas mudanças na PC39. De modo geral as estimativas de herdabilidade para as características medidas nas fêmeas apresentaram baixos valores, enquanto que para as características medidas nos machos se apresentaram de moderada a baixa, de maneira semelhante à maioria dos estudos com estas características. Entretanto, o modelo de limiar foi capaz de detectar variância genética aditiva para as características em estudo, permitindo promover ganhos, embora pequenos, por seleção direta.
In the present study were obtained, in a population of Nellore cattle, estimates of genetic parameters for the characteristics probability of calving up to 39 months at age (PC39), stayability, considering three calving until 5.2 (STAY5,2) and trhee calving until 6.2 years old (STAY6,2), and the first four calving interval (CI1, CI2, CI3 and CI4), treated as categorical traits, and scrotal perimeter at weaning (EPD) and at 18 months of age (EPS), treated as continuos trait. Were used Baeysian procedures, in single and two trait analysis, by using THRGIBBS1F90 and GIBBS2F90 programs. Information from Nellore animals belonging to the Geneplus-Embrapa program was used. The heritabilities for PC39, STAY3,1, STAY3,2, EPD and EPS in single trait analysis were 0.14, 0.20, 0.23, 0.28 and 0.36, respectively. For the calving interval estimates heritabilities for CI1, CI2, CI3 and CI4 were 0.13, 0.13, 0.12 and 0.017, respectively. These values indicate that genetic gains by direct selection in any of the intervals would be very small. In the two trait analysis the heritabilities for PC39 and EPS were 0.15 and 0.38, respectively. The low values of estimates showed that small gains would be obtained by direct selection in any of traits measured in females. EPD and EPS had higher heritabilities. The correlation between EPS and PC39 was positive with low magnitude (0.25), suggesting that the selection for the increase of EPS of sires could possibly lead to small changes in the PC39. In general, estimates of heritability for traits measured in dams were low and for traits measured in males were moderate to low. This result is likely those reported in most studies with the same traits. However, the threshold model was able to detect additive genetic variance for the traits under study and can permit to promote gains, albeit small, by direct selection.

Porcine Reproductive and Respiratory Syndrome virus (PRRSV) is a worldwide pathogen and it is isolated in 1990 at Lelystad Laboratories (Holland). This virus has a tropism for swine respiratory and reproductive apparatus; the symptoms are described in piglets (cough, fever, anorexia), finishing pigs (cough, fever, lympho nodes iperplasia) and in sows (abortions, infertility). PRRS is a disease with a high impact on the economic losses in swine industry. Nowadays there is not an efficacy vaccine able to protect animals from the disease and the main problem is the poor knowledges about host-virus interaction. The aim of this study was to evaluate the time-course of the immune response to a field Porcine Respiratory and Reproductive Syndrome virus isolate in PRRS-naïve, untreated pigs, as well as in three groups of age and breed-matched pigs injected with a live attenuated PRRS vaccine, its adjuvant and an inactivated PRRS vaccine, respectively. Humoral response was measured by IgG evaluation (ELISA test) and it took place earlier in pigs injected with the live attenuated vaccine, which also developed a much stronger serum-neutralizing antibody response to the vaccine strain. On the other hand no clear protection was evidenced in terms of viremia against field isolate, which showed 11,1% nucleotide divergence in ORF7 from the vaccine strain. Cellular-mediated immunity was evaluated by in vitro test and the interferon (IFN)-g response to PRRSV was almost absent on PVD 60 in all groups under study, the same was evidenced for interleukin (IL)-10 responses to PRRSV. Results indicate that distinct patterns of immune response to a field PRRSV strain can be recognized in PRRS-vaccinated and naïve pigs, which probably underlies fundamental differences in the development and differentiation of PRRSV-specific immune effector cells.

AQPs are a class of small, hydrophobic, integral membrane proteins that are able to make the cell membrane 10 to 100 fold more permeable than the membranes lacking such channels. Based on studies with AQP1, it appears that AQPs exist in the plasma membrane as homotetramers. Each AQP monomer contains 2 hemi-pores, which fold together to form a water channel. Different aqps have different patterns of glycosylation. In the case of AQP1 the peptide backbone is roughly 28 kDa and the glycosylated forms range from 40 to 60 kDa in mass. Water transport activity of AQP1 is inhibited by mercurial ion, and the site of mercurial inhibition is the cysteine in the E loop. I have personally verified that using the bovine ovary granulosa cell swelling and shrinkage as a model for mercurial inhibition. The shortest pathway for water to reach the follicle antral cavity is the transcellular pathway directly through the cytoplasm of granulosa cells. In this first experiment we have considered the functional presence of Hg2+-sensitive Aqps in isolated granulosa cells assessing changes in cell volume in presence or absence of HgCl2. Each of the AQPs has an essentially unique pattern of expression among tissues and during development. In humans 13 AQPs have been identified to date although this number pales in comparison to the greater than 100 related proteins found in plants and bacteria. The landmark discovery of AQPs starts the era of molecular water channels, which has permeated virtually every branch of biology and medicine. A number of recent studies have revealed the importance of mammalian AQPs in both physiology and pathophysiology and have suggested that pharmacological modulation of aquaporin expression and activity may provide new tools for the treatment of variety of human disorders, such as brain edema, glaucoma, tumour growth, fertility and obesity in which water and small solute transport may be involved. Since multiple isoforms of AQPs are differentially expressed in cells and tissues, their localization to specific cells and membrane domains is critical to understanding the role of each AQP in the transfer of water and small solutes. Balancement of water concentration within the uterine environment is important in every reproductive moment of a female mammal life. Rapid transcellular water movements are facilitated by AQPs that increase plasma membrane permeability. The results from my research showed that AQP-mediated water movement between the intraluminal, interstitial, and capillary compartments is crucial in the uterine imbibition mechanism in the bitch, including periodic stromal oedema in preparation for embryo implantation, with evidence that the expression of different AQPs can be regulated by steroid sex hormones. Also, the importance of the AQP-mediated fluid regulation in uterine environment in the peri-implantation period is known in the woman and laboratory mammals, to assure the successful implantation of blastocyst and its survival. The establishment of pregnancy and its maintenance is dependent on strict synchronization of uterine receptivity with embryonic maturation. Water homeostasis during foetal development is of crucial physiologic importance. Amniotic fluid (AF) provides the fluid-filled compartment that is essential for normal foetal growth, movement and development. Placental water flux is an important factor in determining AF volume and fetal hydration, too. AF changes during gestation may be dependent on modulations in solute as well as water permeability and changes in AQPs during gestation suggest their role in mediating placental and membrane water flow. AQPs are channel proteins expressed in cell membranes of many epithelia involved in fluid transport and several AQPs have been identified in adult rat excurrent ducts, where considerable fluid reabsorption occurs. Concerning the morphological changes allowed by the AQP influence in the male reproductive tract I considered first of all the role of AQPs in the epididymis of a domestic species, the cat, and two animal models, the rat and the lamb. Ultrastructural features of the epithelium lining the efferent ducts (ED) in the cat, as in other mammalian species, are strongly indicative of an absorptive activity taking place towards the intraluminal fluids. It is well-known that more than 95% of the fluid leaving the testis is reabsorbed by the ED, but the cell structures involved in the reabsorption processes are still a matter of debate. The purpose of the present work was to study the absorptive pathways in the ED of adult cats by means of 1) the immunohistochemical localization of different isoformes of the AQPs and 2) the localization and the carbohydrate characterization of the endocytotic apparatus by means of the lectin histochemistry. AQP-mediated trans-cellular route together with fluid phase glycocalix-mediated endocytosis. Maternal undernutrition and, under certain circumstances, overnutrition, during pregnancy or during early postnatal life can alter reproductive function of the offspring. It is known that epididymal morphology in adult rats is affected by undernutrition during fetal to prepubertal life. For this study I tested the influence of pre- and postnatal undernutrition on AQP-expression in the adult male genital tract. Exposure to environmental endocrine disrupting chemicals (EDCs) during pregnancy in animals and humans is associated with reduced sperm counts and increased incidences of testicular cancer and reproductive tract abnormalities. Although male fetuses from pregnant ewes exposed to EDCs contained in sewage sludge fertiliser (a “real life” exposure model) exhibit fewer testicular Sertoli and Leydig cells, the epididymis has not been investigated. The movement of fluids and small solutes in the epididymis is critical for adult sperm maturation and this process is regulated, in part, by the AQPs. I examined the effects of sewage sludge exposure on the expression of AQPs in the late gestation fetal epididymis. The results I obtained suggest that exposure to environmental chemicals may affect the capacity of efferent ducts to transport water and small solutes thus influencing fluid resorption, protein synthesis and secretion. Ultimately, this may impact on sperm viability in the adult. Another research item was the study of some factors which control the growth of the gubernaculum testis and regression of the cranial suspensory ligament during foetal development, resulting in transabdominal descent of the testes. INSL3, also known as Leydig insulin-like peptide or relaxin-like factor, is supposed to induce the growth of the gubernaculum in male foetuses, thus being directly responsible for the testicular descent in humans and rodents. This is believed to happen with the LGR8 receptor intervention. A study led during my PhD provides evidence of the immunolocalization of INSL3 in the Leydig cells of dog male foetuses and of LGR8 receptor in different tissues of the gubernaculums testis of the same foetuses. AQPs are also involved in the water imbibition of gubernaculum testis: dog foetuses gubernacula showed a high expression of AQP1, AQP3, AQP4 and AQP9 respectively in blood vessels, smooth muscles, connective tissue and fat. As well known AQPs are ubiquitarly distributed in the body maintaining all the same the specific function as channels to permeate water and small solutes. I collaborated with the University of Perugia regarding the immunolocalization of AQP5 in the sheep salivary glands in relation to the pasture vegetative cycle. AQP5 is widely expressed in exocrine glands; in particular, in the rat salivary glands AQP5 is highly expressed in the apical plasma membrane of serous acinar cells, in the secretory canaliculi and in the intercalated duct cells, while it is lacking in mucous acinar cells and in striated ducts, indicating its important role in the saliva production (further supported by the observation of markedly depressed rates of salivary secretion in APQ5-deficient mice. I studied the presence of AQP5 in sheep parotid and mandibular glands, and its expression in the different stages of pasture vegetative cycle. AQP5 is expressed in parotid gland and the reactivity was higher when animals were fed on grasslands at the end of vegetative cycle than on grasslands at the maximum of vegetative development. Ultimately I collaborated to a study created by the Reproduction Unit of the University of Milan regarding the relationship between nutrition and reproductive efficiency in the mare. Leptin, the hormone product of ob gene expression, is an important endocrine indicator of adipose mass and nutritional status, as well as an important regulator of various aspects of feed intake, growth, metabolism and reproduction. Leptin amount in the blood is proportional to body energy stores and/or body mass, so, inadequate nutrition might impair reproductive function leading, for example, to the delayed onset of puberty. Indeed, the onset of puberty in humans and animals is associated with an increase in fat and consequent increase in circulating leptin, suggesting that leptin may be required for normal growth and development of reproductive organs. In the mare, besides many reports quantifying the correlation of circulating concentration of leptin with body condition scores1,2, only few informations exist about the presence of leptin (Ob) and leptin receptor (Ob-R) in the ovary or in the oocyte. The results support the hypothesis that in the horse leptin is differently localized during oocyte IVM showing different immunoreaction intensity related either to the horse breed or to the reproductive puberal development. The presence of leptin in the ovary could suggest a possible involvement in oocyte maturation, angiogenesis, follicle rupture or subsequent corpus luteum formation. Moreover, leptin may be involved in the determination of the animal pole of the oocyte and in the establishment of the inner cell mass and trophoblast in the embryo.

The aim of this thesis was to investigate cellular processes critical to equine maturation, fertilization and embryo development in vitro. Equine compact cumulus oocytes (CCO) in vitro have low competence, so we evaluated the efficacy of a pre-maturation step in improving the coordination of cytoplasmic and nuclear maturation of CCO by the addition of Roscovitine and our results showed partial improving of the competence of these oocytes. Moreover, we carried out a study to examine the immunolocalization of leptin (Ob) and leptin receptor (Ob-R) in CCO recovered from fillies and from mares of light vs heavy body weight breeds after slaughtering and to further improve the maturation rate of CCO we supplemented the maturation medium with leptin and investigated its effect in expanded and compact cumulus oocytes on maturation, fertilization and embryo cleavage rates after intra cytoplasmic sperm injection (ICSI). The results supported the hypothesis that, in the horse, leptin is differently localized during oocyte IVM showing different immunoreaction intensity related either to the horse breed or to the reproductive puberal development. In vitro, the presence of leptin during oocyte maturation improves the progression of meiosis and the rate of fertilization after ICSI but, by contrast to other species, in our horse model no differences were detected in the subsequent development rates of embryos: probably species-differences exist in embryos with regard to sensitivity to leptin.

Fertility preservation has received unprecedented attention nowadays. In addition to cryopreservation and re-implantation of embryos, oocytes, and ovarian tissue pieces, in vitro culture system for follicles/oocytes has been considered an alternative strategy for fertility preservation. Reproduction strategies based on the recovery of oocytes' population from antral follicles are unsatisfactory, and the success of this approach has not exceeded 35% of embryos produced in vitro for over 30 years. The possibility of accessing the reserve of smaller follicles (primordial, secondary, and up to the preantral stage) would amplify the number of gametes available for increasing reproductive potential. Furthermore, this would open enormous prospects for the rescue of fertility in various conditions in the human clinic and genetic rescue in animal breeding and biodiversity preservation programs. However, this would require developing protocols capable of growing immature oocytes to the stage in which they can be matured and fertilized in vitro. Culture systems to achieve in vitro growth (IVG) of immature oocytes to maturity and subsequent fertilization in vitro (IVF) have been the subject of research for almost 40 years. Several systems that support the growth of later stages of follicle development from rodents have been developed, with some reporting the production of live young, but they are still at an experimental stage, and further research is required before the protocols could be clinically applied. One of the significant limitations is identifying growth factors, hormones, and nutrients necessary for each specific follicle development stage. This evidence has led to hypothesize the development of culture systems consisting of a step-by-step approach, although no reliable protocols have been developed so far. The oocyte culture at the early stages of development represents an alternative to maximize the potential source of gamete used for fertility preservation. Several attempts have been made to recreate these conditions in vitro, but no reliable protocols have been developed to date. The lack of knowledge in the mechanisms involved in the early development of the oocyte and this passage from growing to fully grown stage be one of the most critical steps during oocyte development, these still represent the significant limiting factor for this technology. The studies conducted during the doctorate program led to defining a physiological culture system that successfully differentiated growing bovine oocytes. This study used parameters predictive of oocyte differentiation to evaluate the current technique's efficiency and efficacy. Based on previous observations from our laboratory, we initially hypothesized that zinc plays a role during the latest stages of oocyte growth and differentiation, particularly in controlling transcription during the final stage of oocyte growth. This first study demonstrated that zinc supplementation improves the meiotic competence of growing oocytes, affects the global transcription activity and the global DNA methylation. This information was used in the next part to better define a culture system for growing oocytes. The subsequent study provided a 5-days protocol named L-IVCO (long in vitro culture of oocytes) to promote growing oocyte differentiation until the acquisition of meiotic and embryonic developmental competencies in a significantly higher proportion of the published protocols. This study demonstrated that a physiological medium could support a gradual transition of the oocyte from immature to mature stage, thus generating suitably quality blastocysts after fertilization. In conclusion, our studies provide an improved protocol that can increase the source of fertilizable gametes in preservation programs and gives a prospective approach in human clinics, animal breeding programs, and salvage intervention of threatened species. Moreover, our studies defined a model to perform in-depth studies of the cellular and molecular processes that regulate the acquisition of meiotic and developmental competence during oocyte differentiation.

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In this study a total of 48 embryos were used in the compacts morulae, initial blastocysts and blastocysts stages in the degrees I and II, obtained from 14 adult domestic cats. Two animals presented natural manifestation of the oestrus and 12 animals were artificially induced to ovarian activity and ovulation. They received one only application of 150 IU of gonadotropins equine corionic (eCG) and 84h after, one application of 100 IU of gonadotropins human corionic (hCG), and all the animals were coupled naturally. Six days after the first mating, all cats were submitted to laparotomy, and through an adapted transcornual technique the embryos were collected and classified in agreement with IETS (1998). 27 of these were cultivated in a TCM 199 modified with a controlled CO2 atmosphere for 24 hours. The embryos were classified by development stage, quality and blastomeres number, observing a development index until expanded blastocysts in degree I of 77.7% with an average of 161±15,8 blastomeres was reached. In 21 embryos the freezing technique was tested in a commercial medium with 10% of glycerol associated with 0,1molL-1 of sucrose. After the unfreezing, in 37oC water with the same cultivation conditions, a developmental index of 38.1%, was observed until the stage of expanded blastocysts in degree I, with average number of 139.1±11,8 blastomeres. Although the freezing and unfreezing protocols, had reduced the developmental index of feline embryos by 51% and reduce the blastomeres number (P<0.01), it was considerate a viable technique in domestic and wild felines.
Foram utilizados um total de 48 embriões nos estágios de mórula compacta, blastocisto inicial e blastocisto, nos graus I e II, obtidos de 14 gatas domésticas adultas. Dois animais apresentaram manifestação natural do cio e 12 animais foram induzidos artificialmente à atividade ovariana e ovulação. Para tal receberam uma única aplicação de 150 UI de gonadotrofina coriônica eqüina (eCG) e 84h após, uma única aplicação de 100 UI de gonadotrofina coriônica humana (hCG). Todos os animais foram acasalados naturalmente e seis dias após a primeira cópula, todas as gatas foram submetidas a laparotomia, e por meio da técnica de lavagem transcornual adaptada, foram coletados os embriões, os quais foram classificados de acordo com IETS (1998). Vinte e sete foram cultivados em meio TCM 199 modificado, em estufa de CO2, por 24 horas, sendo então avaliados quanto ao seu estágio de desenvolvimento, qualidade e número de blastômeros, observando-se uma taxa de desenvolvimento até o estágio de blastocisto expandido grau I de 77,7%, com um número médio de 161±15,8 blastômeros. Em 21 embriões foi testada a congelabilidade em meio comercial a base de 10% glicerol associado a 0,1molL-1 de sacarose. Após o descongelamento, em banho maria a 37oC, nas mesmas condições de cultivo, observou-se uma taxa de desenvolvimento, até o estágio de blastocisto expandido grau I, de 38,1% com um número médio de 139,1±11,8 blastômeros. Desta forma, embora o protocolo de congelamento e descongelamento tenha reduzido em 51% a taxa de desenvolvimento de embriões felinos, com uma queda (P< 0,01) do número de blastômeros, mostrou-se viável, com promissor uso in vivo em felinos domésticos e silvestres.

Several studies indicate that in utero and peri-natal exposure to some classes of endocrine disruptors induces adverse reproductive effects, but it remains unclear whether such effects may be transmitted to subsequent generations. The present study examined the effects in mice of exposure to di(2-ethyl-hexyl) phthalate (DEHP) or to polychlorinated biphenyls (PCBs) throughout pregnancy and lactation on reproductive health in male and female offspring, at adult age, over three generations. Groups of two to three dams were exposed to increasing doses of contaminants with the diet from gestational day 0.5 until the end of lactation. The doses employed were within the range of environmental exposure levels in humans (DEHP: 0, 0.05, 5, and 500 mg/kg/day; PCBs 101+118: 0, 1, 10, 100 μg/kg/day). In DEHP experiments, treatment of pregnant F0 dams with the 500 mg dose caused complete pregnancy failure, while a slight reduction in litter size in the 5 mg was observed. Male and female F1 offspring born from dams treated with 0.05 and 5 DEHP doses showed, once they reach adult age, significant morphological and functional alterations of the reproductive system. Specifically: i) lower body weight; ii) altered gonad weight (i.e.: lighter testis and heavier ovary) and morphology; iii) reduced germ cells quality; iv) low expression of steroidogenesis and gonadotropin-receptor genes in the gonads; and v) up-regulated gonadotropin subunits gene expression in the pituitary. DEHP exposure altered morphological and reproductive indices in male offspring only in the first generation. Conversely, F2 and F3 female offspring exhibited altered gonad weight and morphology, concomitantly with poor embryo quality, similarly to what observed in F1, thus showing a transgenerational transmission of reproductive adverse effects. Interestingly, also disregulation of selected ovarian and embryonic genes was maintained up to the third generation. In PCBs experiments, treatment did not affect F0 dams’ reproductive outcome. Nevertheless, whole-body PCB burden increased in a dose-dependent manner confirming the effectiveness of the treatment. Furthermore, concentrations at all doses investigated were greater in the offspring than in the dams, confirming that the progeny were exposed as a result of maternal exposure. Pre- and peri-natal exposure to PCBs resulted in male and female offspring showing significant reproductive abnormalities, at adult age. Specifically, compared to controls, they showed reductions in: i) testis weight and seminiferous tubule diameter; ii) sperm viability and developmental capacity; iii) ovary weight; iv) oocyte developmental capacity. Furthermore, F1 ovaries showed a dose-dependent increase in follicular atresia, associated with down-regulation of cyp19a1 and pten mRNA levels. PCBs adverse reproductive effects in females were limited to F1 generation. In contrast, male offspring exhibited reduced sperm viability and altered seminiferous tubule distribution up to the third generation. These results evidence that maternal exposure to PCBs can affects reproductive health in multiple generations. In conclusion, our data indicate that exposure to the endocrine disruptors DEHP or PCBs, at the time of gonadal sex determination, perturbed significantly the reproductive indices of male and female adult offspring. Furthermore, some of the reproductive deficiencies observed upon direct exposure have been observed up to the third generation. These findings have significant implications for reproductive health and fertility of animals and humans.

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Conselho Nacional de Desenvolvimento Científico e Tecnológico
The aim of this study was to genetically evaluate litter traits in Landrace pigs using bi-trait, multi-trait and repeatability models and to estimate the dominance effect with and without the inclusion of polygenic effect for growth and carcass traits in a F2 pig population. For Landrace pigs, the following traits were evaluated: number of piglets born at first (NLN1), second (NLN2) and third (NLN3) farrowings; number of piglets weaned at first (NLD1), second (NLD2) and third (NLD3) farrowings; litter weight at birth at first (PLN1), second (PLN2) and third (PLN3) farrowings; litter weight at weaning at first (PLD1), second (PLD2) and third (PLD3) farrowings; average piglets weight at birth at first (PMLN1), second (PMLN2) and third (PMLN3) farrowings; average piglets weight at weaning at first (PMLD1), second (PMLD2) and third (PMLD3) farrowings. The analysis among farrowings using multi-trait and repeatability models showed that the additive genetic variance and heritability for the first farrowing were lower than those estimated in the other farrowings, indicating that the expression of the studied traits at first farrowing is propably controlled by different genes or differents genes combinations from those that regulate the expression of the second and third farrowings. The genetics correlations among the three farrowings, using multi-trait model for average piglets weight at birth (0.9809 to 0.9970) and average piglets weight at weaning (0.9610 to 0.9800), were high, but different from the unit. The other traits showed correlations with lower magnitude. These results are insufficient to consider these traits in the three farrowing orders as being genetically the same trait. We conclude that the multi-trait model is recommended for genetic evaluation of these traits in different farrowings, considering different farrowings as different traits. The traits were also analyzed within each farrowing with a bi-trait animal model. These analyses showed that the genetic correlations between birth and weaning traits were positive and high in all farrowings, which indicates that selection for improving the traits at birth results in genetic gain of the same traits at weaning. However, these traits at birth should be included in the analyses due to negative genetic correlations between litter size and average piglet weight at birth and at weaning, number of piglets weaned and average piglet weight at birth and at weaning and between litter weight at birth and number of weaned piglets. We conclude that methods that consider the correlations among these traits in this population should be used for genetic evaluation of the traits. In another approach, the combination of genomic data and pedigree was used to study the importance of additive and dominance genetic variances of growth and carcass traits in a F2 pig population. Two GBLUP models were used, a model without inclusion of the polygenic effect (ADM) and a model with the polygenic effect (ADMP). Additive effects showed a greater contribution towards the control of growth and carcass traits. Moreover, the dominance effect was important for all traits, showing a more relevant role in backfat thickness. The narrow-sense and broad-sense heritability estimates for growth (0.06 to 0.42; 0.10 to 0.51, respectively) and carcass (0.07 to 0.37; 0.10 to 0.76, respectively) traits showed wide variation. The inclusion of the polygenic effect in the ADMP model changed the broad-sense heritability estimates only for birth weight and weight at 21 days of age.
O objetivo deste estudo foi avaliar geneticamente características de leitegada em suínos da raça Landrace e estimar via análise genômica o efeito de dominância com e sem a inclusão do efeito poligênico em suínos F2. Foram avaliadas as seguintes características em animais da raça Landrace, utilizando modelos bicaracteristico, multicaracterístico e de repetibilidade: número de leitões nascidos no primeiro (NLN1), segundo (NLN2) e terceiro (NLN3) partos; número de leitões desmamados no primeiro (NLD1), segundo (NLD2) e terceiro (NLD3) partos; peso da leitegada ao nascimento no primeiro (PLN1), segundo (PLN2) e terceiro (PLN3) partos; peso da leitegada ao desmame no primeiro (PLD1), segundo (PLD2) e terceiro (PLD3) partos; peso médio dos leitões ao nascimento no primeiro (PMLN1), segundo (PMLN2) e terceiro (PMLN3) partos; peso médio dos leitões ao desmame no primeiro (PMLD1), segundo (PMLD2) e terceiro (PMLD3) partos. As análises entre partos com os modelos multicaracterístico e de repetibilidade, via REML, mostraram que a variância genética aditiva e a herdabilidade para a primeira ordem de parto foram menores que as estimadas nas demais ordens de parto, o que indica que, provavelmente, a expressão dessas características estudadas no primeiro parto é controlada por genes ou combinações gênicas diferentes daquelas que regulam a expressão das mesmas no segundo e no terceiro parto. correlações genéticas entre as três ordens de parto no As modelo multicaracterístico para as características peso médio do leitão ao nascimento (PMLN) e peso médio do leitão ao desmame (PMLD) foram altas variando de 0,9809 a 0,9970 para PMLN e de 0,9610 a 0,9800 para PMLD, mas diferente da unidade, as demais características apresentaram correlações de menor magnitude. Esses resultados são insuficientes para considerar estas características nas três ordens de parto como sendo, geneticamente, a mesma característica. Conclui-se que o modelo multicaracterístico é recomendado para avaliações genéticas das características número de leitões nascidos e desmamados, peso da leitegada e peso médio do leitão ao nascimento e a desmama nas diferentes parições, tratando diferentes parições como características diferentes. As características foram analisadas também dentro de cada parto com o modelo animal bicaracterístico. Nestas análises observou- se que as correlações genéticas entre as características de nascimento e de desmama foram positivas e altas no primeiro, segundo e terceiro parto para a maioria das características, o que indica que a seleção para aumento número de leitões nascidos, peso da leitegada e peso médio do leitão ao nascimento resultará em ganho genético das mesmas características a desmama, porém as três características devem ser incluídas nas análises devido às correlações genéticas negativas entre número de leitões nascidos e peso médio do leitão ao nascimento e ao desmame, número de leitões desmamados e peso médio do leitão ao nascimento e ao desmame e entre peso da leitegada ao nascimento e número de leitões desmamados. Conclui-se que devem ser utilizadas para avaliação genéticas destas características metodologias que não desprezem as correlações existentes entre elas na população em questão. Foi utilizada a combinação de dados genômicos e pedigree para estudar a importância das variâncias genéticas aditivas e de dominância de características de crescimento e de carcaça em uma população de suínos F2. Foram utilizados dois modelos GBLUP, um modelo sem a inclusão do efeito poligênico (ADM) e um modelo com o efeito poligênico (ADMP). Efeitos aditivos apresentaram maior contribuição para o controle de características de crescimento e de carcaça. Além disso, o efeito de dominância foi importante para todas as características, mostrando um papel mais relevante na espessura de toucinho. As herdabilidades no sentido restrito e no sentido amplo para as características de crescimento variaram de 0,06 a 0,42 e de 0,10 a 0,51, respectivamente e para as características de carcaça de 0,07 a 0,37 no sentido restrito e de 0,10 a 0,76 no sentido amplo, mostrando grande variação. A inclusão do efeito poligênico no modelo ADMP mudou as estimativas de herdabilidade no sentido amplo apenas para peso ao nascimento e peso aos 21 dias de idade.

Pendant de nombreuses années, les femelles ont été décrites comme passives dans les interactions mâles-femelles et ont parfois été négligées dans les études sur les comportements reproducteurs et la communication animale. Pourtant, il est de plus en plus évident que les comportements des femelles influencent ceux des mâles et que les mâles peuvent ajuster leurs comportements de parades aux comportements des femelles. L’objectif principal de cette thèse est de mieux comprendre les signaux que les femelles émettent lors d’interactions avec un mâle dans un contexte de reproduction chez le canari domestique. L’ensemble de mes résultats montre tout d’abord que les femelles utilisent des signaux de parades visuels et acoustiques, c’est-à-dire des postures de sollicitation à l’accouplement et des trilles-spécifiques de femelles, comme une invitation à s’accoupler, mais qu’elles peuvent aussi les utiliser pour inciter le mâle à parader et l’aider à échantillonner la qualité de partenaires potentiels. D’autre part, ces deux signaux n’auraient pas la même efficacité selon le contexte d’émission. Ensuite, il semblerait que les signaux véhiculés par la modalité visuelle pourraient jouer un rôle plus important qu’on ne le pensait jusque-là dans les interactions intersexuelles chez cette espèce. Enfin, cette thèse a aussi pu apporter de nouveaux éléments concernant les préférences des femelles pour les chants de mâles et a montré que les méthodes utilisées en laboratoire pour tester les préférences des femelles étaient fiables et congruentes. Cette thèse permet de contribuer aux recherches grandissantes mettant en évidence le rôle important de la femelle dans les interactions intersexuelles
Much studies on reproduction and animal communication have considered the female as the passive sex; the role of the female during male-female interactions have often been overlooked. However, there is growing evidence that female behaviours can affect those of the males and that males can adjust their courtships to female behaviours. The aim of this thesis is to understand the signals produced by females during interactions with a male in a reproductive context in the domestic canary. Overall, my results show that females not only use their visual and acoustic signals, the copulation solicitation display and the female-specific trills, as an invitation to copulate but also to incite male to sing as an aid to sample potential mates. Then, these two signals could not have the same efficacy in different contexts of transmission. Moreover, the visual components of the communication seem to be more important than previously thought during intersexual interactions in this species. Finally, this thesis provides new elements about the female preferences for male songs and shows that methods used to test female preferences in laboratory are reliable and congruent. This thesis contributes to the growing number of researches showing that females play an active role in intersexual interactions

La sub-fertilità rappresenta uno dei principali problemi che l'industria lattiero-casearia si trova ad affrontare attualmente. Si tratta di un problema multifattoriale che dipende da diversi aspetti della funzione riproduttiva. La qualità dell'ovocita, determinata durante la follicologenesi, è sicuramente cruciale per ottenere risultati riproduttivi ottimali. Per migliorare la fertilità femminile nel bovino è fondamentale definire i fattori e i meccanismi che determinano la qualità del gamete femminile, in quanto il declino della funzione riproduttiva in questa specie è dovuto principalmente alla scarsa qualità degli ovociti. In particolare, è importante conoscere quali proteine regolano la meiosi, la fecondabilità e lo sviluppo embrionale dell'ovocita. In questo progetto, abbiamo considerato principalmente il ruolo del Progesterone Receptor Membrane Component-1 (PGRMC1) sia nell'ovocita che nel compartimento follicolare, considerato che il dialogo tra compartimento germinale e compartimento somatico è uno dei fattori chiave coinvolti nell'acquisizione della competenza allo sviluppo del gamete femminile. Il PGRMC1 è una proteina multifunzionale ed altamente conservata in specie anche filogeneticamente distanti. Nei mammiferi è espressa in diversi sistemi, compresi gli organi riproduttivi, e localizza in diversi compartimenti subcellulari. Tuttavia, il suo ruolo e il preciso meccanismo d'azione in ogni sistema e/o compartimento subcellulare non sono ancora del tutto conosciuti. La parte più considerevole di questo progetto di dottorato riguarda gli studi effettuati sul ruolo del PGRMC1 nella divisione cellulare, data la sua localizzazione a livello di fuso mitotico e meiotico. Abbiamo dimostrato che la deplezione del PGRMC1 determina difetti nella divisione cellulare e la sua interazione con l'Aurora chinasi B (AURKB) indica che potrebbe svolgere la sua azione durante la citodieresi, l'ultima fase della divisione. Durante la maturazione meiotica, abbiamo valutato anche il suo ruolo nel mediare l'azione del progesterone (P4) confrontando il suo effetto con quello del recettore nucleare del progesterone (nPGR). L'inibizione di entrambi i tipi di recettore determina lo stesso effetto sull'organizzazione della piastra metafasica e sulla competenza allo sviluppo dell'ovocita, ma in fasi differenti. Inoltre, abbiamo testato l'ipotesi che il PGRMC1 possa modulare la funzione del nucleolo. Studi di immunofluorescenza hanno confermato la presenza del PGRMC1 nel nucleolo di cellule della granulosa (bGC) e negli ovociti di bovino, nonchè la colocalizzazione con la nucleolina, la proteina nucleolare più abbondante che svolge importanti funzioni in questo compartimento subcellulare. Inoltre, la down-regolazione del PGRMC1 determina uno spostamento della nucleolina dal nucleolo al nucleoplasma suggerendo un'associazione funzionale tra queste due proteine. Questa interazione è probabilmente mediata dalla presenza di ulteriori molecole in quanto successivi esperimenti di "in-situ proximity ligation" mostrano che le due proteine non interagiscono direttamente. Oltre al tema principale di questo progetto, la divisione cellulare è uno dei processi più importanti anche nel cancro e l'espressione del PGRMC1 è elevata in molti tipi di tumore nell'uomo. Per questo abbiamo intrapreso uno studio preliminare per caratterizzare la sua espressione nei diversi tipi di tumore mammario di cane. In base ai nostri risultati, l'espressione del PGRMC1 diminuisce con l'aumentare della malignità del tumore e le diverse isoforme di PGRMC1 sono presenti sia nel tessuto sano che in quello neoplastico. In conclusione, i nostri risultati suggeriscono che il PGRMC1 può avere un ruolo nel compartimento follicolare, in particolare sia nella mitosi che nella meiosi. La sua funzione potrebbe essere legata alla sua localizzazione nucleolare, nonchè alla sua capacità di mediare l'azione del P4 durante la meiosi dell'ovocita. Il suo ruolo come regolatore della divisione cellulare può essere rilevante anche in alcune patologie quali il cancro. Ulteriori studi saranno diretti a valutare i meccanismi di azione molecolari tramite i quali il PGRMC1 agisce in questi processi al fine di ampliare la nostra conoscenza di questa proteina poliedrica.
Subfertility is one of the major problems that dairy industry is facing nowadays. Subfertility is a multifactorial issue depending on different aspects of reproductive function. Oocyte quality, which is determined during folliculogenesis, is certainly crucial for optimal reproductive outcome. Since declining fertility of cattle is mainly due to the poor quality of the oocytes, defining the factors and mechanisms that affect oocyte quality is essential to improve female fertility. In particular, basic knowledge of which proteins within the oocyte regulate meiosis, oocyte fertilizability and early embryonic development would be advantageous. In this project, we primarily considered the role of Progesterone Receptor Membrane Component-1 (PGRMC1) in both the oocyte and follicular compartment since the dialogue between the two compartments is one of the key factors involved in oocyte competence acquisition. PGRMC1 is a highly conserved and multifunctional protein that is found expressed in multiple systems, including reproductive organs, and localizes in multiple sub-cellular compartments. However, its role(s) and precise mechanism(s) of action in each systems and/or subcellular compartment are not yet fully understood. The more considerable part of this PhD project consists of studies on PGRMC1's involvement as a cell division regulator, according to its localization to the mitotic and meiotic spindle. We showed that PGRMC1 depletion leads to defective cell division in both oocyte and somatic cells and its action could be exerted during cytokinesis, the very last mitotic phase, as demonstrated by its association with Aurora kinase B (AURKB). During meiotic maturation, we also evaluated PGRMC1 involvement as a possible mediator of progesterone (P4) action, by comparing the effects of inhibiting its function with that of nuclear progesterone receptor's (nPGR) inhibition. Our data suggests that both receptors have an effect on meiotic progression but possibly at different stages of oocyte maturation. Moreover, we tested the hypothesis that PGRMC1 might modulate the function of the nucleolus. Immunofluorescence experiments confirmed PGRMC1 nucleolar localization in bovine granulosa cells (bGC) and bovine oocytes and its co-localization with nucleolin, the most abundant nucleolar protein exerting important functions in this subcellular compartment. Moreover, a PGRMC1/nucleolin functional association is suggested by PGRMC1's downregulation determining nucleolin shift from the nucleolus to the nucleoplasm. However, in situ proximity ligation assay did not detect a direct interaction between these two proteins, suggesting the involvement of additional molecules that could mediate PGRMC1/nucleolin interaction. Besides the main theme of this project, cell division is one of the most important processes in cancer biology and PGRMC1 is known to be overexpressed in many types of tumors in humans. Therefore, we studied its expression in different types of canine mammary tumors. Our preliminary results showed PGRMC1 expression decreases with the malignancy of the tumor and different PGRMC1 isoforms are present both in normal and tumoral tissue. To conclude, these findings suggest a role of PGRMC1 in the follicular compartment, being implicated in both meiotic and mitotic process of the germinal and somatic compartment, respectively. This action it is likely mediated by PGRMC1 located at a particular site of the spindle, i.e the midzone and /or the midbody of dividing cells. PGRMC1 also localizes at the nucleolus of both oocytes and granulosa cells where it likely exerts additional function(s) in mediating cellular stress and/or other nucleolar-relates process. Clearly these findings have major implication in the overall process of folliculogenesis. Moreover, PGRMC1's role as a regulator of cell proliferation could be relevant also in some diseases such as cancer. Further investigations will aim at investigating PGRMC1's molecular mechanisms of action in these processes to widen our knowledge on this multifaceted protein.