Heritability Estimate and Genetic Correlations of Reproductive Features in Nellore Bulls, Offspring of Super Precocious, Precocious and Normal Cows Under Extensive Farming Conditions

The present work aimed to estimate heritability and genetic correlations of reproductive features of Nellore bulls, offspring of mothers classified as superprecocious (M1), precocious (M2) and normal (M3). Twenty one thousand hundred and eighty-six animals with average age of 21.29 months were used, evaluated through the breeding soundness evaluation from 1999 to 2008. The breeding soundness features included physical semen evaluation (progressive sperm motility and sperm vigour), semen morphology (major, minor and total sperm defects), scrotal circumference (SC), testicular volume (TV) and SC at 18 months of age (SC18). The components of variance, heritability and genetic correlations for and between the features were estimated simultaneously by restricted maximum likelihood, with the use of the vce software system vs 6. The heritability estimates were high for SC18, SC and TV (0.43, 0.63 and 0.54; 0.45, 0.45 and 0.44; 0.42, 0.45 and 0.41, respectively for the categories of mothers M1, M2 and M3) and low for physical and morphological semen aspects. The genetic correlations between SC18 and SC were high, as well as between these variables with TV. High and positive genetic correlations were recorded among SC18, SC and TV with the physical aspects of the semen, although no favourable association was verified with the morphological aspects, for the three categories of mothers. It can be concluded that the mothers sexual precocity did not affect the heritability of their offspring reproduction features.

Mating system parameters at hierarchical levels of fruits, individuals and populations in the Brazilian insect-pollinated tropical tree, Tabebuia roseo-alba (Bignoniaceae)

For many tree species, mating system analyses have indicated potential variations in the selfing rate and paternity correlation among fruits within individuals, among individuals within populations, among populations, and from one flowering event to another. In this study, we used eight microsatellite markers to investigate mating systems at two hierarchical levels (fruits within individuals and individuals within populations) for the insect pollinated Neotropical tree Tabebuia roseo-alba. We found that T. roseo-alba has a mixed mating system with predominantly outcrossed mating. The outcrossing rates at the population level were similar across two T. roseo-alba populations; however, the rates varied considerably among individuals within populations. The correlated paternity results at different hierarchical levels showed that there is a high probability of shared paternal parentage when comparing seeds within fruits and among fruits within plants and full-sibs occur in much higher proportion within fruits than among fruits. Significant levels of fixation index were found in both populations and biparental inbreeding is believed to be the main cause of the observed inbreeding. The number of pollen donors contributing to mating was low. Furthermore, open-pollinated seeds varied according to relatedness, including half-sibs, full-sibs, self-sibs and self- half-sibs. In both populations, the effective population size within a family (seed-tree and its offspring) was lower than expected for panmictic populations. Thus, seeds for ex situ conservation genetics, progeny tests and reforestation must be collected from a large number of seed-trees to guarantee an adequate effective population in the sample.

The role of AT1 receptor-mediated reproductive function in renovascular hypertension in male rats

There is an association between hypertension and reproductive dysfunction. Angiotensin II (Ang II) is involved in the pathogenesis of hypertension and the regulation of reproduction. The present study aimed to determine whether the angiotensinergic system mediates the effects of hypertension on ieproductive function in male rats subjected to a two-kidney, one-clip (2K1C) model. Sexual behavior parameters, gametogenesis and plasma concentrations of Ang II, testosterone, prolactin and corticosterone were evaluated in male rats 28 days after 2K1C or sham surgery and losartan (Los) treatment (a type 1 angiotensin II (All) receptor antagonist) or vehicle (V) treatment. The animals were divided into Sham + V, 2K1C + V. Sham + Los and 2K1C + Los groups. The 2KiC + V group showed a hypertensive response, inhibition of sexual behavior, spermatogenesis dysfunction, and increases in plasma Ang II and prolactin. Conversely, plasma testosterone decreased, and plasma corticosterone remained constant. Losartan treatment normalized blood pressure and prevented the changes in plasma testosterone and prolactin, sexual behavior and spermatogenesis in the 2KiC + Los group. In addition, losartan treatment caused an additional increase in circulating Ang II in both groups (She m + Los arid 2K1C + Los). Together, these results suggest that Ang II, acting through the All receptor, modulates behavioral and endocrine parameters of reproductive function during renovascular hypertension. In addition, the effects of circulating Ang II on plasma testosterone and prolactin seem to contribute to the spermatogenic and sexual dysfunctions in hypertensive rats. (C) 2012 Els.evier Inc. All rights reserved.

Genetic relationship between growth and reproductive traits in Nellore cattle

The objective of this study was to evaluate the genetic relationship between postweaning weight gain (PWG), heifer pregnancy (HP), scrotal circumference (SC) at 18 months of age, stayability at 6 years of age (STAY) and finishing visual score at 18 months of age (PREC), and to determine the potential of these traits as selection criteria for the genetic improvement of growth and reproduction in Nellore cattle. The HP was defined as the observation that a heifer conceived and remained pregnant, which was assessed by rectal palpation at 60 days. The STAY was defined as whether or not a cow calved every year up to the age of 6 years, given that she was provided the opportunity to breed. The Bayesian linear-threshold analysis via the Gibbs sampler was used to estimate the variance and covariance components applying a multitrait model. Posterior mean estimates of direct heritability were 0.15 +/- 0.00, 0.42 +/- 0.02, 0.49 +/- 0.01, 0.11 +/- 0.01 and 0.19 +/- 0.00 for PWG, HP, SC, STAY and PREC, respectively. The genetic correlations between traits ranged from 0.17 to 0.62. The traits studied generally have potential for use as selection criteria in genetic breeding programs. The genetic correlations between all traits show that selection for one of these traits does not imply the loss of the others.

Reproductive biology of Boops boops (Linnaeus, 1758) off Gran Canaria (Canary Islands): a preliminary study

[EN] Preliminary data of the length-weight relationship and reproduction of the bogue Boops boops off Gran Canaria (Canary Islands, Central-east Atlantic) are provided. Two thousand and twenty-one individuals of bogue, ranging from 4 to 34 cm TL, were obtained from purse seine commercial landing. Reproduction parameters as sex determination, duration of spawning season, size at first maturity and GSI variation along time were determinated based on macroscopic evaluation of gonads. The results obtained suggest that bogue is a total spawner, with a long spawning season extending from January to May. Size at first maturity was 16.7 and 17.9 cm TL for males and females, respectively. The length-weight relationship obtained showed a positive allometry in both sexes

(Table 1) Breeding parameters for 60 male Adélie penguins (Pygoscelis adeliae) at station Dumont d'Urville (Antarctica) depending on treatment (control vs. CORT)

Corticosterone, the main stress hormone in birds, mediates resource allocation, allowing animals to adjust their physiology and behaviour to changes in the environment. Incubation is a time and energy-consuming phase of the avian reproductive cycle. It may be terminated prematurely, when the parents' energy stores are depleted or when environmental conditions are severe. In this study, the effects of experimentally elevated baseline corticosterone levels on the parental investment of incubating male Adelie penguins were investigated. Incubation duration and reproductive success of 60 penguins were recorded. The clutches of some birds were replaced by dummy eggs, which recorded egg temperatures and rotation rates, enabling a detailed investigation of incubation behaviour. Corticosterone levels of treated birds were 2.4-fold higher than those of controls 18 days post treatment. Exogenous corticosterone triggered nest desertion in 61% of the treated birds; consequently reducing reproductive success, indicating that corticosterone can reduce or disrupt parental investment. Regarding egg temperatures, hypothermic events became more frequent and more pronounced in treated birds, before these birds eventually abandoned their nest. The treatment also significantly decreased incubation temperatures by 1.3 °C and lengthened the incubation period by 2.1 days. However, the number of chicks at hatching was similar among successful nests, regardless of treatment. Weather conditions appeared to be particularly important in determining the extent to which corticosterone levels affected the behaviour of penguins, as treated penguins were more sensitive to severe weather conditions. This underlines the importance of considering the interactions of organisms with their environment in studies of animal behaviour and ecophysiology.

Biostimulation methods associated with early weaning and reproductive rhythms in primiparous rabbit does

El objetivo general de esta Tesis Doctoral ha sido tratar de mejorar los parámetros reproductivos de las conejas primíparas lactantes, empleando dos métodos de manejo (destete temprano y extensificación del ritmo reproductivo), que están directamente relacionados con su balance energético. Para ello, se diseñaron 2 experimentos en este tipo de hembras. En el primero, se estudió el efecto del destete a 25 días post-parto (dpp) sobre la actividad ovárica y el metabolismo energético de las conejas una semana más tarde (32 dpp). Un total de 34 primíparas lactantes con 8 gazapos fueron distribuidas en tres grupos: 10 conejas se sacrificaron a los 25 dpp (grupo L25), 13 fueron destetadas a los 25 dpp y sacrificadas a los 32 dpp (grupo NL32), y 11 conejas no se destetaron y fueron sacrificadas a los 32 dpp (grupo L32). No se observaron diferencias significativas entre grupos en el peso corporal, el peso del ovario, ni en las concentraciones séricas de ácidos grasos no esterificados y de proteínas totales. A pesar de que el grupo NL32 presentó un bajo consumo de alimento (122 ± 23,5 g / día, p <0,001), su contenido corporal estimado de lípidos (16,9 ± 1,09%, P <0,008), proteínas (19,7 ± 0,07%, P <0,0001), y energía (1147 ± 42,7 MJ / kg, p <0,006) fueron más elevados y las concentraciones séricas de glucosa (158 ± 24,5 mg/dl, p <0,04) más bajas que en los grupos L25 (11,9 ± 1,3%, 18,5 ± 0,08%, 942 ± 51,3 MJ/kg y 212 ± 27,9 mg/dl) y L32 (13,4 ± 1,03%, 18,5 ± 0,1%, 993 ± 40,4 MJ/kg y 259 ± 29,5 mg/dl), respectivamente. En el grupo L25 se observó un menor número medio de folículos ≥ 1 mm en la superficie ovárica en comparación con los grupos NL32 y L32 (12,7 ± 1,5 vs. 18,0 ± 1,45 y 17,6 ± 1,67, p <0,05). La población folicular ovárica en las secciones histológicas y la inmunolocalización de los receptores de prolactina fueron similares en todos los grupos. En el grupo L25, tanto la maduración nuclear de oocitos, medida en términos de tasas alcanzadas de Metafase II (67,0 vs. 79,7 y 78,3%, P <0.05) y la maduración citoplasmática, medida por el porcentaje de gránulos corticales (GC) total o parcialmente migrados en los oocitos, fueron significativamente menores que en los grupos NL32 y L32 (16,0 vs 38,3 y 60,0%, P <0.05). En conclusión, a pesar de que el destete precoz a 25 dpp pareció mejorar las reservas de energía de las conejas primíparas, este hecho no se reflejó claramente a nivel ovárico a los 32 dpp y fue similar independientemente del destete, por lo que éste último podría llevarse a cabo más tarde. En el segundo experimento, se compararon dos ritmos reproductivos. Se utilizaron un total de 48 conejas primíparas lactantes con 8 gazapos que se asignaron al azar en dos grupos experimentales: a) lactantes sacrificadas a comienzos del post-parto (11 dpp) de acuerdo a un ritmo semi-intensivo (n = 24), y b) lactantes sacrificadas al final del período post-parto (25 dpp) de acuerdo con un ritmo más extensivo (n = 24). En ellas, se estudió el peso vivo, la composición corporal estimada, parámetros metabólicos y endocrinos (estradiol y progesterona) y características ováricas como la población folicular y la tasa de atresia, así como la maduración nuclear y citoplásmica de los oocitos. En este estudio, el peso vivo, el contenido de energía corporal, los depósitos grasos y los ácidos grasos no esterificados disminuyeron a lo largo del post-parto con respecto al momento del parto (P <0,05). Las concentraciones séricas de proteínas y glucosa aumentaron en el mismo periodo post-parto (P <0,05). Se observaron similares niveles de estradiol y progesterona en ambos ritmos, así como una población folicular, tasas de maduración nuclear (tasa de oocitos en metafase II) y citoplasmática (porcentaje de oocitos con gránulos corticales migrados), similares en ambos momentos del post-parto. Sin embargo, el número de folículos preovulatorios en la superficie ovárica fue menor (P <0,05) y la tasa de atresia tendió a ser mayor con un porcentaje también menor de folículos sanos (P <0,1) en los ovarios de las hembras sometidas al ritmo extensivo. En conclusión, al final del post-parto (25 días), las conejas primíparas sin destetar muestran un deterioro de sus reservas corporales, de sus parámetros metabólicos séricos y de la calidad de sus oocitos; incluso se ha observado una ligera influencia negativa en el desarrollo de sus folículos ováricos. Por esta razón, se considera que en las conejas primíparas lactantes el manejo reproductivo extensivo (25 dpp) no presenta ninguna ventaja en comparación con el semi-intensivo (11 dpp). A la vista de los resultados de estos dos experimentos, podemos decir que ni el destete temprano, ni la extensificación del ritmo reproductivo han conseguido una mejora en los parámetros reproductivos de una hembra primípara. Por ello, son necesarios más estudios sobre el estado metabólico de la coneja primípara lactante para conseguir métodos o estrategias que lo mejoren y tengan consecuencias directas sobre la actividad reproductiva y sobre su éxito productivo. The general aim of this Thesis was to study two management methods (early weaning and extensive reproductive rhythm) linked to the energy balance of the primiparous rabbit does to improve their reproductive performance. In this sense, 2 experiments were conducted using this kind of females. In the first experiment, the effect of weaning at 25 days post-partum (dpp) on ovarian activity and energetic metabolism one week later (32 dpp) was studied. A total of 34 primiparous lactating rabbit does were used and distributed among three groups: 10 does euthanized at 25 dpp (group L25), 13 does weaned at 25 dpp and euthanized at 32 dpp (group NL32), and 11 non weaned does euthanized at 32 dpp (group L32). No significant differences were observed in live body weight, ovary weight, serum non esterified fatty acids (NEFA) and total protein concentration among groups. Although NL32 does had a low feed intake (122±23.5 g/Day; P < 0.001), their estimated lipids (16.9±1.09%, P < 0.008), protein (19.7±0.07%, P < 0.0001), and energy (1147±42.7 MJ/kg, P < 0.006) body contents were higher and their serum glucose concentrations (158±24.5 mg/dl, P < 0.04) were lower compared to L25 does (11.9±1.3%, 18.5±0.08%, 942±51.3 MJ/kg and 212±27.9 mg/dl) and L32 does (13.4±1.03%, 18.5±0.1%, 993±40.4 MJ/kg and 259±29.5 mg/dl, respectively). A lower number of follicles ≥1mm was observed compared to NL32 and L32 groups (12.7±1.5 vs. 18.0±1.45 and 17.6 ±1.67; P < 0.05) in the ovarian surface of L25 does. Follicular population in the histological ovarian sections and immunolocalization of prolactin receptor were similar in all groups. In group L25, both nuclear maturation of oocytes in terms of Metaphase II rate (67.0 vs. 79.7 and 78.3%; P < 0.05) and cytoplasmic maturation measured by percentage of cortical granules (CG), totally or partially migrated in oocytes were significantly lower than in groups NL32 and L32 (16.0 vs. 38.3 and 60.0%; P < 0.05). Consequently, a higher rate of oocytes with non-migrated CGs was found in group L25 than in groups NL32 and L32 (76.0 vs. 46.8 and 33.3%; P < 0.05). In conclusion, even though early weaning at 25 dpp seemed to improve body energy stored in primiparous does, this fact was not well reflected on the ovarian status at 32 dpp, which was similar regardless of weaning time. In the second experiment, two reproductive rhythms were compared. A total of 48 primiparous Californian x New Zealand White rabbit does suckling 8 kits were randomly allocated in two experimental groups: a) lactating does euthanized at early post-partum period (11 dpp) according to a semi-intensive rhythm (n = 24), and b) lactating does euthanized on later post-partum period (25 dpp) according to a more extensive rhythm (n = 24). Live weight, estimated body composition, serum metabolic and endocrine parameters (oestradiol and progesterone concentrations) and ovarian features like follicle population and atresia rate, and oocyte maturation were studied. Live weight, body energy content, lipid depots and serum non esterified fatty acids (NEFA) concentrations diminished from parturition time to post-partum period (P < 0.05). In addition, serum protein and glucose concentrations increased along postpartum time (P < 0.05). Similar oestradiol and progesterone levels were shown in rhythms as well as similar follicle population and nuclear and cytoplasmic maturation rates measured as metaphase II and cortical granule migration, respectively in both postpartum times. However, number of preovulatory follicles on the ovarian surface was lower (P < 0.05) and atresia rate tended to be higher with also lower percentage of healthy follicles (P < 0.1) in ovaries of females of extensive group. In conclusion, primiparous non-weaned rabbits does at late post-partum time (25 days), Did no show any improvement regarding body reserves, serum metabolic parameters and oocyte quality; even a slight negative influence has been observed in the development of their ovarian follicles. Thus this reproductive management does not present any advantage compared to earlier post-partum (11 days) reproductive rhythm. In summary, according to the obtained results from these two experiments, we can say that the application of early weaning and the extensive rhythms did not achieve an improvement in the reproductive performance of primiparous does. Thus, it is necessary to conduct more studies about the metabolic status of the primiparous lactating doe to achieve strategies in order to improve it and consequently, to improve the reproductive activity and their productive success.

Valutazione della risposta immunitaria cellulare e fenotipizzazione linfocitaria nel suino in corso di infezione naturale da virus della PRRS (“Porcine Reproductive and Respiratory Syndrome”) e dopo stimolazione di PBMC (“Peripheral Blood Mononuclear Cells”) in vitro con Peptide Killer (KP)

SUMMARY The Porcine Reproductive and Respiratory Syndrome (PRRS) virus is one of the most spread pathogens in swine herds all over the world and responsible for a reproductive and respiratory syndrome that causes severe heath and economical problems. This virus emerged in late 1980’s but although about 30 years have passed by, the knowledge about some essential facets related to the features of the virus (pathogenesis, immune response, and epidemiology) seems to be still incomplete. Taking into account that the development of modern vaccines is based on how innate and acquire immunity react, a more and more thorough knowledge on the immune system is needed, in terms of molecular modulation/regulation of the inflammatory and immune response upon PRRSV infection. The present doctoral thesis, which is divided into 3 different studies, is aimed to increase the knowledge about the interaction between the immune system and the PRRS virus upon natural infection. The objective of the first study entitled “Coordinated immune response of memory and cytotoxic T cells together with IFN-γ secreting cells after porcine reproductive and respiratory syndrome virus (PRRSV) natural infection in conventional pigs” was to evaluate the activation and modulation of the immune response in pigs naturally infected by PRRSV compared to an uninfected control group. The course of viremia was evaluated by PCR, the antibody titres by ELISA, the number of IFN-γ secreting cells (IFN- SC) by an ELISPOT assay and the immunophenotyping of some lymphocyte subsets (cytotoxic cells, memory T lymphocytes and cytotoxic T lymphocytes) by flow cytometry. The results showed that the activation of the cell-mediated immune response against PRRSV is delayed upon infection and that however the levels of IFN-γ SC and lymphocyte subsets subsequently increase over time. Furthermore, it was observed that the course of the different immune cell subsets is time-associated with the levels of PRRSV-specific IFN-γ SC and this can be interpreted based on the functional role that such lymphocyte subsets could have in the specific production/secretion of the immunostimulatory cytokine IFN-γ. In addition, these data support the hypothesis that the age of the animals upon the onset of infection or the diverse immunobiological features of the field isolate, as typically hypothesized during PRRSV infection, are critical conditions able to influence the qualitative and quantitative course of the cell-mediated immune response during PRRSV natural infection. The second study entitled “Immune response to PCV2 vaccination in PRRSV viremic piglets” was aimed to evaluate whether PRRSV could interfere with the activation of the immune response to PCV2 vaccination in pigs. In this trial, 200 pigs were divided into 2 groups: PCV2-vaccinated (at 4 weeks of age) and PCV2-unvaccinated (control group). Some piglets of both groups got infected by PRRSV, as determined by PRRSV viremia detection, so that 4 groups were defined as follows: PCV2 vaccinated - PRRSV viremic PCV2 vaccinated - PRRSV non viremic PCV2 unvaccinated - PRRSV viremic PCV2 unvaccinated - PRRSV non viremic The following parameters were evaluated in the 4 groups: number of PCV2-specific IFN-γ secreting cells, antibody titres by ELISA and IPMA. Based on the immunological data analysis, it can be deduced that: 1) The low levels of antibodies against PCV2 in the PCV2-vaccinated – PRRSV-viremic group at vaccination (4 weeks of age) could be related to a reduced colostrum intake influenced by PRRSV viremia. 2) Independently of the viremia status, serological data of the PCV2-vaccinated group by ELISA and IPMA does not show statistically different differences. Consequently, it can be be stated that, under the conditions of the study, PRRSV does not interfere with the antibody response induced by the PCV2 vaccine. 3) The cell-mediated immune response in terms of number of PCV2-specific IFN-γ secreting cells in the PCV2-vaccinated – PRRSV-viremic group seems to be compromised, as demonstrated by the reduction of the number of IFN-γ secreting cells after PCV2 vaccination, compared to the PCV2-vaccinated – PRRSV-non-viremic group. The data highlight and further support the inhibitory role of PRRSV on the development and activation of the immune response and highlight how a natural infection at early age can negatively influence the immune response to other pathogens/antigens. The third study entitled “Phenotypic modulation of porcine CD14+ monocytes, natural killer/natural killer T cells and CD8αβ+ T cell subsets by an antibody-derived killer peptide (KP)” was aimed to determine whether and how the killer peptide (KP) could modulate the immune response in terms of activation of specific lymphocyte subsets. This is a preliminary approach also aimed to subsequently evaluate such KP with a potential antivural role or as adjuvant. In this work, pig peripheral blood mononuclear cells (PBMC) were stimulated with three KP concentrations (10, 20 and 40 g/ml) for three time points (24, 48 and 72 hours). TIME POINTS (hours) KP CONCENTRATIONS (g/ml) 24 0-10-20-40 48 0-10-20-40 72 0-10-20-40 By using flow cytometry, the qualitative and quantitative modulation of the following immune subsets was evaluated upon KP stimulation: monocytes, natural killer (NK) cells, natural killer T (NKT) cells, and CD4+ and CD8α/β+ T lymphocyte subsets. Based on the data, it can be deduced that: 1) KP promotes a dose-dependent activation of monocytes, particularly after 24 hours of stimulation, by inducing a monocyte phenotypic and maturation shift mainly involved in sustaining the innate/inflammatory response. 2) KP induces a strong dose-dependent modulation of NK and NKT cells, characterized by an intense increase of the NKT cell fraction compared to NK cells, both subsets involved in the antibody-dependent cell cytotoxicity (ADCC). The increase is observed especially after 24 hours of stimulation. 3) KP promotes a significant activation of the cytotoxic T lymphocyte subset (CTL). 4) KP can modulate both the T helper and T cytotoxic phenotype, by inducing T helper cells to acquire the CD8α thus becoming doube positive cells (CD4+CD8+) and by inducing CTL (CD4-CD8+high) to acquire the double positive phenotype (CD4+CD8α+high). Therefore, KP may induce several effects on different immune cell subsets. For this reason, further research is needed aimed at characterizing each “effect” of KP and thus identifying the best use of the decapeptide for vaccination practice, therapeutic purposes or as vaccine adjuvant. RIASSUNTO Il virus della PRRS (Porcine Reproductive Respiratory Syndrome) è uno dei più diffusi agenti patogeni negli allevamenti suini di tutto il mondo, responsabile di una sindrome riproduttiva e respiratoria causa di gravi danni ad impatto sanitario ed economico. Questo virus è emerso attorno alla fine degli anni ’80 ma nonostante siano passati circa una trentina di anni, le conoscenze su alcuni punti essenziali che riguardano le caratteristiche del virus (patogenesi, risposta immunitaria, epidemiologia) appaiono ancora spesso incomplete. Considerando che lo sviluppo dei vaccini moderni è basato sui principi dell’immunità innata e acquisita è essenziale una sempre più completa conoscenza del sistema immunitario inteso come modulazione/regolazione molecolare della risposta infiammatoria e immunitaria in corso di tale infezione. Questo lavoro di tesi, suddiviso in tre diversi studi, ha l’intento di contribuire all’aumento delle informazioni riguardo l’interazione del sistema immunitario, con il virus della PRRS in condizioni di infezione naturale. L’obbiettivo del primo studio, intitolato “Associazione di cellule memoria, cellule citotossiche e cellule secernenti IFN- nella risposta immunitaria in corso di infezione naturale da Virus della Sindrome Riproduttiva e Respiratoria del Suino (PRRSV)” è stato di valutare l’attivazione e la modulazione della risposta immunitaria in suini naturalmente infetti da PRRSV rispetto ad un gruppo controllo non infetto. I parametri valutati sono stati la viremia mediante PCR, il titolo anticorpale mediante ELISA, il numero di cellule secernenti IFN- (IFN- SC) mediante tecnica ELISPOT e la fenotipizzazione di alcune sottopopolazioni linfocitarie (Cellule citotossiche, linfociti T memoria e linfociti T citotossici) mediante citofluorimetria a flusso. Dai risultati ottenuti è stato possibile osservare che l’attivazione della risposta immunitaria cellulo-mediata verso PRRSV appare ritardata durante l’infezione e che l’andamento, in termini di IFN- SC e dei cambiamenti delle sottopopolazioni linfocitarie, mostra comunque degli incrementi seppur successivi nel tempo. E’ stato inoltre osservato che gli andamenti delle diverse sottopopolazioni immunitarie cellulari appaiono temporalmente associati ai livelli di IFN- SC PRRSV-specifiche e ciò potrebbe essere interpretato sulla base del ruolo funzionale che tali sottopopolazioni linfocitarie potrebbero avere nella produzione/secrezione specifica della citochina immunoattivatrice IFN-. Questi dati inoltre supportano l’ipotesi che l’età degli animali alla comparsa dell’infezione o, come tipicamente ipotizzato nell’infezione da PRRSV, le differenti caratteristiche immunobiologiche dell’isolato di campo, sia condizioni critiche nell’ influenzare l’andamento qualitativo e quantitativo della risposta cellulo-mediata durante l’infezione naturale da PRRSV. Il secondo studio, dal titolo “Valutazione della risposta immunitaria nei confronti di una vaccinazione contro PCV2 in suini riscontrati PRRSV viremici e non viremici alla vaccinazione” ha avuto lo scopo di valutare se il virus della PRRS potesse andare ad interferire sull’attivazione della risposta immunitaria indotta da vaccinazione contro PCV2 nel suino. In questo lavoro sono stati arruolati 200 animali divisi in due gruppi, PCV2 Vaccinato (a 4 settimane di età) e PCV2 Non Vaccinato (controllo negativo). Alcuni suinetti di entrambi i gruppi, si sono naturalmente infettati con PRRSV, come determinato con l’analisi della viremia da PRRSV, per cui è stato possibile creare quattro sottogruppi, rispettivamente: PCV2 vaccinato - PRRSV viremico PCV2 vaccinato - PRRSV non viremico PCV2 non vaccinato - PRRSV viremico PCV2 non vaccinato - PRRSV non viremico Su questi quattro sottogruppi sono stati valutati i seguenti parametri: numero di cellule secernenti IFN- PCV2 specifiche, ed i titoli anticorpali mediante tecniche ELISA ed IPMA. Dall’analisi dei dati immunologici derivati dalle suddette tecniche è stato possibile dedurre che:  I bassi valori anticorpali nei confronti di PCV2 del gruppo Vaccinato PCV2-PRRSV viremico già al periodo della vaccinazione (4 settimane di età) potrebbero essere messi in relazione ad una ridotta assunzione di colostro legata allo stato di viremia da PRRSV  Indipendentemente dallo stato viremico, i dati sierologici del gruppo vaccinato PCV2 provenienti sia da ELISA sia da IPMA non mostrano differenze statisticamente significative. Di conseguenza è possibile affermare che in questo caso PRRSV non interferisce con la risposta anticorpale promossa dal vaccino PCV2.  La risposta immunitaria cellulo-mediata, intesa come numero di cellule secernenti IFN- PCV2 specifiche nel gruppo PCV2 vaccinato PRRS viremico sembra essere compromessa, come viene infatti dimostrato dalla diminuzione del numero di cellule secernenti IFN- dopo la vaccinazione contro PCV2, comparata con il gruppo PCV2 vaccinato- non viremico. I dati evidenziano ed ulteriormente sostengono il ruolo inibitorio del virus della PRRSV sullo sviluppo ed attivazione della risposta immunitaria e come un infezione naturale ad età precoci possa influenzare negativamente la risposta immunitaria ad altri patogeni/antigeni. Il terzo studio, intitolato “Modulazione fenotipica di: monociti CD14+, cellule natural killer (NK), T natural killer (NKT) e sottopopolazioni linfocitarie T CD4+ e CD8+ durante stimolazione con killer peptide (KP) nella specie suina” ha avuto come scopo quello di stabilire se e come il Peptide Killer (KP) potesse modulare la risposta immunitaria in termini di attivazione di specifiche sottopopolazioni linfocitarie. Si tratta di un approccio preliminare anche ai fini di successivamente valutare tale KP in un potenziale ruolo antivirale o come adiuvante. In questo lavoro, periferal blood mononuclear cells (PBMC) suine sono state stimolate con KP a tre diverse concentrazioni (10, 20 e 40 g/ml) per tre diversi tempi (24, 48 e 72 ore). TEMPI DI STIMOLAZIONE (ore) CONCENTRAZIONE DI KP (g/ml) 24 0-10-20-40 48 0-10-20-40 72 0-10-20-40 Mediante la citometria a flusso è stato dunque possibile analizzare il comportamento qualitativo e quantitativo di alcune sottopopolazioni linfocitarie sotto lo stimolo del KP, tra cui: monociti, cellule Natural Killer (NK), cellule T Natural Killer (NKT) e linfociti T CD4 e CD8+. Dai dati ottenuti è stato possibile dedurre che: 1) KP promuove un’attivazione dei monociti dose-dipendente in particolare dopo 24 ore di stimolazione, inducendo uno “shift” fenotipico e di maturazione monocitaria maggiormente coinvolto nel sostegno della risposta innata/infiammatoria. 2) KP induce una forte modulazione dose-dipendente di cellule NK e NKT con un forte aumento della frazione delle cellule NKT rispetto alle NK, sottopopolazioni entrambe coinvolte nella citotossicità cellulare mediata da anticorpi (ADCC). L’aumento è riscontrabile soprattutto dopo 24 ore di stimolazione. 3) KP promuove una significativa attivazione della sottopopolazione del linfociti T citotossici (CTL). 4) Per quanto riguarda la marcatura CD4+/CD8+ è stato dimostrato che KP ha la capacità di modulare sia il fenotipo T helper che T citotossico, inducendo le cellule T helper ad acquisire CD8 diventando quindi doppio positive (CD4+CD8+) ed inducendo il fenotipo CTL (CD4-CD8+high) ad acquisire il fenotipo doppio positivo (CD4+CD8α+high). Molti dunque potrebbero essere gli effetti che il decapeptide KP potrebbe esercitare sulle diverse sottopopolazioni del sistema immunitario, per questo motivo va evidenziata la necessità di impostare e attuare nuove ricerche che portino alla caratterizzazione di ciascuna “abilità” di KP e che conducano successivamente alla scoperta del migliore utilizzo che si possa fare del decapeptide sia dal punto di vista vaccinale, terapeutico oppure sotto forma di adiuvante vaccinale.

Life history parameters and biocontrol potential of the mealybug parasitoid Coccidoxenoides peregrinus (Timberlake) (Hymenoptera: Encyrtidae): Asexuality, fecundity and ovipositional patterns

Properties relevant to the ovipositional activity and lifetime productivity of Coccidoxenoides peregrinus (Timberlake) were assessed in the laboratory, to determine the potential of this species as a biocontrol agent against the citrus mealybug, Planococcus citri (Risso). In general, this species has not performed well in orchards, except for a few localities on different continents. The mode of reproduction of C peregrinus is almost entirely thelytokous, with males produced sporadically and at low frequency. The females have both pro-ovigenic and synovigenic traits, which raises questions of the utility of this distinction. The females have a high reproductive potential with 10-20 eggs per day available within the first two days (after a short (12 h) pre-oviposition period), and 80-150 eggs per day thereafter until death at about eight days. Mean lifetime fecundity was 239.2 +/- 34.3 eggs. C peregrinus oviposits across a range of P. citri instars, but productivity relies predominantly on second instar hosts. Second stage (N2) hosts received most eggs in choice (about 52%) and no-choice (about 50%) tests. Most eggs deposited into N2 hosts (82%) reached adult stage whereas only a few of those deposited into N1 and N3 (about 5% each) developed successfully. The haemolymph of parasitised reproductive mealybugs contained granular structures and no parasitoid eggs were found 24 h after exposure to ovipositing wasps. Also, no wasps emerged from parasitised adult hosts that were kept alive. Parasitoid eggs deposited into adult hosts were presumed encapsulated and destroyed, as control mealybugs (not exposed to female wasps) had no granular structures in their haemolymph. Wasps exposed to an abundance of hosts soon started ovipositing, but only for a relatively short time each day (about 2.5 h out of a 7 h exposure). They stopped ovipositing despite eggs judged to be mature in their ovaries. The reproductive output of C peregrinus is discussed in relation to the ecological factors that could influence this output, and the implications for biocontrol are discussed. (C) 2003 Elsevier Inc. All rights reserved.

Heterozygote effects in mice with partial truncations in the growth hormone receptor cytoplasmic domain: Assessment of growth parameters and phenotype

The GH receptor (GHR) is essential for normal postnatal growth and development, and the molecular basis of GHR action has been studied intensively. Clinical case studies and more recently mouse models have revealed the extensive phenotype of impaired GH action. We recently reported two new mouse models, possessing cytoplasmic truncations at position 569 (plus Y539/545-F) and 391, which were created to identify functional subdomains within the cytoplasmic signaling domain. In the homozygous state, these animals show progressively impaired postnatal growth coupled with complex changes in gene expression. We describe here an extended phenotype analysis encompassing the heterozygote state to identify whether single copies of these mutant receptors bring about partial or dominant-negative phenotypes. It appears that the retention of the ubiquitin-dependent endocytosis motif the N-terminal cytoplasmic domain permits turnover of these mutant receptors because no dominant-negative phenotype is seen. Nonetheless, we do observe partial impairment of postnatal growth in heterozygotes supporting limited haploinsufficiency. Reproductive function is impaired in these models in a progressive manner, in parallel with loss of signal transducer and activator of transcription-5 activation ability. In summary, we describe a more comprehensive phenotypic analysis of these mouse models, encompassing overall and longitudinal body growth, reproductive function, and hormonal status in both the heterozygote and homozygote state. Our results suggest that patients expressing single copies of similarly mutated GHRs would not display an obvious clinical phenotype.

Factors affecting female reproductive success and the survival of pouch young in the threatened brush-tailed rock-wallaby, Petrogale penicillata

Knowledge of factors affecting the survival of individuals and their reproductive success is essential for threatened species management, but studies assessing these factors are lacking for many threatened rock-wallaby species. In this study we investigated the factors influencing the breeding performance of females and the survival of pouch young in a wild colony of the threatened brush-tailed rock-wallaby. Individuals were trapped between October 2000 and April 2004. More than 50% of the females in the colony were breeding below their full potential and giving birth to only one offspring per year. Most females within the colony bred in synchrony, with a substantial birth peak evident during autumn. Pouch young born in autumn left the pouch during spring and were weaned during summer and autumn when forage was most abundant. Pouch young born during the autumn birth peak or in winter had a substantially higher probability of surviving through to pouch emergence than those born during spring or summer. This study provides demographic parameters that may be used in population models and for comparison with other populations, particularly those that are small and declining. To optimise reproductive success in reintroduction programs, females in good condition and with small pouch young should be released at the end of the wettest season.

Source populations in coastal crabs: parameters affecting egg production

Benthic marine invertebrates may form metapopulations connected via propagule dispersal. Conservation efforts often target potential source coastlines to indirectly benefit areas depending on allochthonous offspring production. Besides population density, adult size structure, sex ratio, brooding frequency and the proportion of breeding individuals may significantly influence the reproductive output of benthic populations, but these effects have seldom been tested. We used rocky shore crabs to assess the spatial variability of such parameters at relevant scales for conservation purposes and to test their consistency over 2 consecutive years; we then used the data to address whether bottom-up processes or biological interactions might explain the patterns observed. We decomposed egg production rates into their components for the 2 most abundant brachyuran species inhabiting the intertidal rocky habitat. Adult density and brooding frequency varied consistently among shores for both species and largely explained the overall spatial trends of egg production. Temporally consistent patterns also included among-shore differences in the size of ovigerous females of the grapsid Pachygrapsus transversus and between-bay differences in the fecundity of the spider crab Epialtus brasiliensis. Sex ratio was remarkably constant in both. We found no positive or negative correlations between adult density and brooding frequency to support either the existence of a component Allee effect (lack of mate encounters) or an effect of intra-specific competition. Likewise, shore-specific potential growth in P. transversus does not negatively correlate with frequency of ovigerous individuals, as would be expected under a critical balance between these 2 processes. The patterns observed suggest that bottom-up drivers may best explain spatial trends in the reproductive output of these species.

Reproductive performance as affected by lactation status and body fat in beef cows.

Seventy-one mature Brangus cows, 38 nonlactating (NL) and 33 in late stage of lactation (L) were fed for 192 days (Phase I) a low energy diet (L). During Phase II (65 days) 19 NL and 17 L cows were fed a high energy diet (H). The other nonlactating (19) and lactating (16) cows remained on the low energy diet. Energy restriction during Phase I did not affect (P> 0.05) cyclic ovarian activity although losses in body weight and condition were substantial. Rapid changes in body weight, condition, and percent empty body lipe (EBLP) during Phase II did not substantially influencefertility, although a five-fold difference in EBLP was observed (NL0H vs. L-L). Treatment groups did not differ (P> 0.05) in conception rate, days from the beginning of the breeding season to breeding and to conception, conception at first service, and number of services per conception. Values observed for these parameters for NL-H, L-H, NL-L, and L-L groups were respectively: 68,4%, `3,.2, 23.3, 36.8% and 1.68; 82,4% 12.7, 19.5, 58.8% and 1.29; 68.4%, 10.2, 17.4, 47.4%, and 1.41; 68.8%, 12.4, 19.5, 43.7%, and 1.50.

Early and late effects of Ibuprofen on mouse sperm parameters, chromatin condensation, and DNA integrity in mice

Background: There are few studies indicating the detrimental effects of ibuprofen on sperm fertility potential and DNA integrity. Objective: To determine the effects of Ibuprofen on sperm parameters, chromatin condensation and DNA integrity of mice. Materials and Methods: In this experimental study, 36 adult male mice with average weight 37 gr were divided into three groups, including control (group I, n=12), normal dosage of ibuprofen (group II, n=12) and high dosage (group III, n=12). Ibuprofen with different doses was dissolved in daily water of animals. After 35, 70 and 105 days, the cauda epididymis of mice were cut and incubated in Ham’s F10 media. Sperm samples were analyzed for parameters (motility, morphology and count), DNA integrity (SCD test) and chromatin condensation (chromomycin A3 and Aniline blue staining). Results: After 35 days, in addition to above mentioned sperm parameters, all of the treated mice showed statistically significant increase in spermatozoa with immature chromatin (P<0.05). However, after 70 days, the rate of sperm DNA fragmentation assessed by SCD was increased in group II (66.5±0.7) and the percentage of immature spermatozoa (AB+ and CMA3+) was higher in group III (77.5±0.7 and 49.5±6.3 respectively) than other groups. After 105 days, the AB+ spermatozoa were increased in both normal dose and high dose groups. Conclusion: Ibuprofen may cause a significant reduction in sperm parameters and sperm chromatin/DNA integrity in mice. It should be noted that these deleterious effects are dose-dependent and can be seen in early and late stage of drug treatments.

Population parameters of the smooth lantern shark, Etmopterus pusillus, in southern Portugal (NE Atlantic)

Etmopterus pusillus is a deep water lantern shark with a widespread global distribution that is caught in large quantities in some areas, but is usually discarded due to the low commercial value. In this work, the population biology was studied and life history parameters determined for the first time in this species. Age was estimated from sections of the second dorsal spine and validated by marginal increment analysis. Males attained a maximum age of 13 years, while 17-year-old females were found. Several growth models were fitted and compared for both size and weight at age data, showing that even though this is a small sized species, it has a relatively slow growth rate. This species matures late and at a relatively large size: at 86.81% and 79.40% of the maximum observed sizes and at 58.02% and 54.40% of the maximum observed ages for males and females, respectively. It has a low fecundity, with a mean ovarian fecundity of 10.44 oocytes per reproductive cycle. The estimated parameters indicate that this species has a vulnerable life cycle, typical of deep water squalid sharks. Given the high fishing pressures that it is suffering in the NE Atlantic, the smooth lantern shark may be in danger of severe declines in the near future. (c) 2007 Elsevier B.V. All rights reserved.