Livelli di inquinanti organici persistenti in stenelle (<i>Stenella coeruleoalba</i>) spiaggiate nel nord Sardegna = Concentration of persistent organic pollutants in striped dolphin (<i>Stenella coeruleoalba</i>) stranded in North Sardinia

Long-lived apex predators, such as marine mammals, are particularly at risk from effects of persistent organic pollutants (POPs), e.g. polychlorinated biphenyls (PCBs) and dichlorodiphenylethanes (e.g. DDT), due to bioaccumulation and biomagnification. POPs are lipophilic compounds that tend to accumulate in the lipid-rich blubber. In marine mammals, POPs enter the body almost exclusively through the diet. In the present study, the levels of PCBs and DDTs in the blubber of striped dolphin stranded in North Sardinia were determined. Our results showed that pollutant concentration are related to age and sex of the individuals. Considering the well known harmful consequences of bioaccumulation of POPs in marine mammals, further studies are needed to determine which POPs might be linked to effects on health

Ejaculate collection efficiency and post-thaw semen quality in wild-caught Griffon vultures from the Sardinian population

This study aimed to test the feasibility of a programme of semen collection and cryopreservation in Griffon vultures. Four wild-caught individuals kept in captivity because of unrecoverable traumas were used. Semen collection attempts were made twice a week during three consecutive reproductive seasons (December – March) using the abdominal massage method. Ejaculation was successfully induced between late January and late February. Semen collection efficiency was rather low (27.9%) and it did not vary among individuals (p > 0.05). No differences were found in ejaculate volumes (12.5 +/- 9.1 μl), spermatozoa concentration (28.4 +/- 30.9 million cells/ml) and viability (61.3 +/- 13.9%) among the 4 vultures. ATP values differed among the four vultures (p < 0.001); B showed higher nucleotide concentration than both C and D, while it did not differ form A, whose values were higher compared with D. After freezing and thawing, semen in vitro viability, DNA integrity and ATP intracellular concentration were determined. Spermatozoa viability after thawing did not differ among the four individuals (52.6 +/- 5.8 in A, 53.4 +/- 4.6 in B, 50.4 +/- 3.2 in C, 42.5 +/- 2.7 in D), but it decreased significantly compared to fresh semen (p < 0.05). During 4 hrs in vitro culture, spermatozoa collected from B maintained over time a higher viability in vitro when compared to A, C and D. As evaluated by the comet assay method, DNA fragmentation after freezing and thawing did not differ in the 4 vultures. ATP concentration in frozen/thawed semen was significantly lower than in fresh semen (p < 0.0001). This study indicates that semen cryopreservation can be considered as a useful tool in the conservation of Griffon vulture genetic resources, but further studies are needed to optimize this technique

Morphofunctional aspects of in vitro matured oocytes from prepubertal and adult sheep

The aim of this study was to analyze the morphofunctional aspects of prepubertal and adult sheep oocytes subjected to in vitro maturation (IVM). The structural and ultrastructural morphology was examined by light and transmission electron microscopy (LM and TEM) while the metabolic competence, as determined by the distribution of active mitochondria, was assessed by confocal microscopy (CM). Cumulus-oocyte complexes underwent IVM in standard condition for 24 h. Half of the mature metaphase II (MII) stage oocytes were processed for LM and TEM observations. The other half was subjected to immunostaining with MitoTracker Red (to stain mitochondria with functionally active membrane potential), SNARF-1 (a pH sensitive fluoroprobe), Hoechst 33342 (to label DNA) and analyzed by CM. Immature germinal vesicle (GV) stage oocytes, retrieved at 0 h IVM, were used as controls. By LM and TEM all the oocytes were regularly rounded, covered by microvilli and surrounded by an intact zona pellucida. Numerous rounded, oval or hooded mitochondria appeared either isolated or grouped in the ooplasm. The GV was usually rounded in prepubertal oocytes. In the adult, the GV often appeared flattened against the oolemma, with a crescent-shaped outline (an early sign of meiotic resumption). Scattered cortical granules (CGs) were rarely found in the ooplasm of both prepubertal and adult GV oocytes. After 24 h of IVM, CGs became abundant and distributed in a single row under the oolemma, particularly in adult oocytes. CM showed a homogeneous fine-to-granular mitochondrial distribution in prepubertal GV and MII oocytes. In adult GV oocytes, the mitochondrial distribution pattern was granular while in MII oocytes mitochondria were arranged in heterogeneous clusters. Thus, both prepubertal and adult oocytes completed maturation after 24 h in culture and showed an overall good preservation after IVM. However, the diverse distribution patterns of mitochondria in prepubertal oocytes reflect their low developmental competence

Use of a neuroleptic in assisted reproduction of the critically endangered Mohor gazelle (Gazella dama mhorr)

Abstract Stress is a limiting factor in assisted reproduction in wild animals maintained in captivity and measures to reduce it should improve reproductive success. The effect of the long-acting neuroleptic (LAN) perphenazine enanthate was assessed on ovarian stimulation for the recovery of immature oocytes from Mohor gazelle (Gazella dama mhorr) and their subsequent in vitro maturation, fertilization and embryo culture. The viability of embryos after transfer was also examined. Perphenazine enanthate decreased activity levels and facilitated handling of treated animals when compared to controls. LAN-treated animals showed a more regular pattern of respiratory and heart rates and body temperature than controls; no major differences were found in hematological and biochemical parameters between groups. Perphenazine-treated females had lower plasma cortisol levels during the days of intense handling. No significant differences were found in the number of punctured follicles and recovered oocytes between groups. The percentage of mature oocytes per female was significantly higher in the LAN-group. Fertilization and cleavage rates were not significantly different between groups. Embryos developed in culture but none reached the blastocyst stage, and those transferred to the oviduct of synchronized recipients did not develop to term. In conclusion, treatment of females with perphenazine enanthate during ovarian stimulation did not have negative effects on maturation, fertilization and embryo development in vitro. Moreover, an increase in oocyte maturation rate per female was observed. Thus, the use of LANs could be useful to alleviate the effects of handling-stress during assisted reproductive procedures in wild ungulates.

Use of Propylene-Glycol as a Cosolvent for GnRH in Synchronization of Estrus and Ovulation in Sheep

The foreseen shortage of eCG for estrus synchronization in sheep makes necessary the development of alternative protocols. The aim of the present work was to evaluate the reproductive response of sheep in breeding season to the administration of GnRH using propylene-glycol as a cosolvent and the subcutaneous route for slowing and extending the release of GnRH, as well as the most adequate timing for such administration. In the present study, protocols based on a short-term CIDR treatment and a single subcutaneous dose of GnRH in propylene-glycol at 36 h after CIDR removal induced a similar ovarian response to protocols based on administration of eCG at CIDR removal or intramuscular GnRH in distilled water at 56 h after. In such protocol, 80% of the animals developed estrus in a narrow timing (75% between 36 and 48 h after CIDR removal), and all of them also ovulated in a narrow window (87.5% between 72 and 76 h after CIDR removal, with 62.5% between 72 and 76 h) and showed a similar ovulation rate and plasma progesterone concentrations at the induced estrous cycle. Hence, administration of GnRH in propylene-glycol may constitute an alternative to traditional protocols based on the administration of eCG

Does the timing of pasture allocation affect rumen and plasma metabolites and ghrelin, insulin and cortisol profile in dairy ewes?

A study was undertaken to assess the impact of the timing of grazing on rumen and plasma metabolites and some metabolic hormones in lactating dairy sheep allocated to an Italian ryegrass (Lolium multiflorum Lam) pasture in spring for 4 h/d. Twenty-four mid lactation Sarda ewes stratified for milk yield, body weight, and body condition score, were divided into four homogeneous groups randomly allocated to the treatments (2 replicate groups per treatment). Treatments were morning (AM, from 08:00 to 12:00) and afternoon pasture allocation (PM, from 15:30 to 19:30). Samples of rumen liquor (day 39) and blood plasma (days 17 and 34 of the experimental period) were collected before and after the grazing sessions. Moreover, on days 11 and 35, grazing time was assessed by direct observation and herbage intake measured by the double weighing procedure. Grazing time was longer in PM than AM ewes (P < 0.001) but herbage intake was undifferentiated between groups. The intake of water-soluble carbohydrates at pasture was higher in PM than AM ewes (P < 0.05). The post-grazing propionic and butyric acid concentration, as measured on day 39, were higher in PM than AM ewes (P < 0.05). The basal level of glucose on day 34 and insulin (on both sampling days) were higher in PM than AM (P < 0.05). The opposite trend was detected for non-esterified fatty acids (P < 0.05, day 34) and urea (both days). Pasture allocation in the afternoon rather than in the morning decreased plasma concentration of ghrelin (P < 0.001) and cortisol (P < 0.001), with a smoothed trend on day 34 in the latter variable. To conclude, postponing the pasture allocation to afternoon increased the intake of WSC, favoring a glucogenic pattern of rumen fermentation and a rise of glucose and insulin levels in blood, although these effects were not consistent across the whole experimental period. Moreover, the afternoon grazing decreased the level of cortisol and ghrelin, suggesting a higher satiation-relaxing effect

Selection of young ewe lambs according to their antral follicular count: response to exogenous hormonal stimulation and fertility at first breeding season

Anti-Mullerian Hormone (AMH), Antral Follicular Count (AFC) and the response to exogenous hormonal stimulation have been used, in adults, as suitable markers to determine the ovarian reserve (1-4), to predict oocyte quality (5,6) and a wide variety of fertility indices (6-9). This investigation aims to evaluate if animals selected according to their High or Low AFC at an early prepubertal age show different responses, in the number of follicles and AMH plasma levels, to exogenous hormonal stimulation; to verify whether differences are maintained over time until puberty; and to observe possible variations on fertility at first breeding season