Hair cortisol and dehydroepiandrosterone sulfate concentrations in healthy beef calves from birth to 6 months of age.

Cortisol (C) and dehydroepiandrosterone (DHEA) are recognized as the main fetal steroids, and they are likely to influence fetal development and have long-term effects on newborn hypothalamic-pituitary-adrenal axis (HPA) function. DHEA is often measured as its sulfates and expressed as DHEA-S. Hair analysis represents a promising methodological approach for the non-invasive measurement of steroids, allowing for a retrospective analysis of the total exposure to steroids over time, and avoiding the influence of acute events or circadian fluctuations. Hair cortisol and DHEA concentrations have been investigated in cows, but no studies have been performed on calves. The object of this study was to evaluate hair cortisol (HC) and hair DHEA-S (HDHEA-S) concentrations in beef calves from birth to six months of age. Hair samples of 12 beef calves (seven males, five females) were firstly collected at birth (T1) and then every three weeks up to six months of age (T2-T10), collecting only the re-grown hair. HC and HDHEA-S were analyzed by radioimmunoassay (RIA). Calves sex, weight and APGAR score were registered immediately after birth. Statistical analysis revealed that both HC and HDHEA-S were influenced by sampling time (P < 0.001). HC concentrations were higher at T1 compared to all subsequent samplings (T2-T10, P < 0.01); HC concentrations were higher at T2 compared to T4-T10 (P < 0.01), while no further changes were detected from T3 onward. Higher HDHEA-S concentrations were registered at T1, T2 and T3 compared to all the other samplings (P < 0.01). No correlation was found between hair concentrations of both steroids and calf sex or birthweight. APGAR score was negatively correlated only with HC at birth (P < 0.05). These data demonstrate that C and DHEA-S are quantifiable in the hair of calves and are influenced by their age. The higher HC detected at birth (T1) probably reflects the high serum C concentrations present late in pregnancy and increased by the fetal HPA axis, by which parturition is initiated in cows. The highest HDHEA-S at birth (T1) in calves indicates that the largest amounts of DHEA and its sulfates are produced during fetal development. Moreover, the findings of higher HC at three weeks after birth and of higher HDHEA-S until six weeks after birth, suggest that C and DHEA secretion continues also beyond birth, and that these steroids could be involved in the events occurring during the challenging first weeks of age in the calf.

Metabolic and hormonal control of energy utilization and partitioning from early to mid lactation in Sarda ewes and Saanen goats.

In a recent study, we observed that starch-rich diets used in mid lactation induced lower milk production persistency and higher body fat accumulation in dairy ewes compared with dairy goats. Because these species differences could be linked to hormonal mechanisms that drive energy partitioning, in the same experiment, we explored the evolution of metabolic and hormonal status during lactation to test this hypothesis. Twenty mature Sarda dairy ewes and 20 mature Saanen goats [15-134 ± 11 d in milk (DIM), mean ± SD] were compared simultaneously. In early lactation, each species was allocated to one dietary treatment: high-starch diet [HS: 20.4% starch, on dry matter (DM) basis], whereas from 92 ± 11 DIM, each species was allocated to 1 of 2 dietary treatments: HS (20.0% starch, on DM basis) and low-starch (LS: 7.8% starch, on DM basis) diets. Blood samples were collected in the morning to analyze glucose, nonesterified fatty acids (NEFA), growth hormone (GH), insulin, and insulin-like growth factor I (IGF-I). Data were analyzed using the PROC MIXED procedure of SAS with repeated measurements (SAS Version 9.0). The HS and LS diets applied in mid lactation did not affect metabolic status of the animal within species; thus, only a comparison between species was carried out. From early to mid lactation, plasma glucose concentration was higher in ewes than in goats (54.57 vs. 48.35 ± 1.18 mg/dL), whereas plasma NEFA concentration was greater in goats than in ewes (0.31 vs. 0.25 ± 0.03 mmol/L). Goats had higher plasma GH concentration and lower plasma insulin content than ewes (4.78 vs. 1.31 ng/mL ± 0.47; 0.11 vs. 0.26 µg/L ± 0.02). Plasma IGF-I concentration did not vary between species. The comparison of metabolic and hormonal status of lactating Sarda dairy ewes and Saanen goats, carried out by studying simultaneously the 2 species in the same stage of lactation and experimental conditions, suggests that the higher insulin and glucose concentration observed in Sarda ewes explains why they partitioned more energy toward body reserves than to the mammary gland, especially in mid lactation. This can justify the negative effect of high-starch diets in mid-lactating Sarda ewes. Conversely, the highest GH and NEFA concentration observed in Saanen goats explain why they partitioned more energy of starch diets toward the mammary gland than to body reserves and justify the positive effect of high-starch diet in mid lactation. Together, these different responses contribute to explain why specialized dairy goats, such as the Saanen breed, have a higher milk production persistency than specialized dairy sheep breeds, such as the Sarda.

Comparison of AlphaLISA and RIA assays for measurement of wool cortisol concentrations.

Radioimmunoassay (RIA) methods have always represented a technique of choice for the determination of steroids in biological samples. The Amplified Luminescent Proximity Homogenous Assay-Linked Immunosorbent Assay (AlphaLISA) is now emerging as the new-generation immunoassay technology that does not require washing/separation steps. The aim of this study was to adapt the Perkin-Elmer's AlphaLISA kit for wool cortisol and compare it with a RIA wool cortisol assay. Wool from lambs, 35 at birth (A0) and 54 at two months old (A2), was collected and each extract was evaluated for wool cortisol concentrations (HCC) both by RIA and AlphaLISA immunoassay. The two methods showed good precision, sensitivity and specificity for determining HCC. Both methods were able to detect significant differences between the high and the low HCC assessed in lambs at A0 and A2 (P < 0.01). The HCC assessed with RIA were significantly higher than those assessed with AlphaLISA (P < 0.01). Moreover, the correlation between HCC measured using the AlphaLISA and RIA methods was strong (r = 0.878). The regression analyses show a constant and not proportional error. This could be due to the diversity in the dosage steps and to the diversity of the molecules used in the two methods. Results support the validity of using AlphaLISA as an alternative method to RIA for the quantification of cortisol in sheep wool and considering the performances showed it has a great potential to be further applied as an excellent tool to evaluate HCC in samples derived from animal species.

Effect of breed body-size on leptin amniotic fluid concentrations at term pregnancy in dogs.

Because of the need to improve the knowledge about canine perinatology, and given the major role of fetal fluids in sustaining the course of pregnancy and fetal development, an in-depth analysis to better understand the role of some hormones in these compartments is essential. Among all, leptin is recognized to play a key role not only on the energetic homeostasis, but also at multiple levels, influencing the control of reproduction, food assumption and metabolism. Even if in humans and other species it is reported the presence of leptin receptors during fetal development, very little is known about the canine species, in which the role of leptin still needs to be fully understood. The present study aimed to assess the amniotic fluid leptin (AFL) concentrations at term pregnancy in healthy dogs, and to evaluate the possible influence played by breed body-size (after assessment of correlation with maternal bodyweight and placental weight), or other maternal (age, parity, and the so-called "litter effect") and neonatal (gender, birth weight, litter size) parameters on AFL concentrations, analyzed by ELISA test. The study was performed on 90 healthy, viable and normal weighted puppies, 39 small-sized (adult body weight < 10 kg) and 51 large-sized (adult body weight > 25 kg), born by 29 purebred, healthy bitches, submitted to elective Caesarean section because of breed-related or individual high risk for dystocia. The results showed that the mean AFL concentration in the small-sized puppies was significantly (p < 0.05) higher in comparison to large-sized puppies (867.48 vs 698.42 pg/ml), while all the other studied parameters did not show to influence AFL concentrations. In conclusions, the present study showed significant higher at term AFL concentrations in small-sized as compared to large-sized breeds, suggesting an influence of breed body-size on fetal metabolism, as previously reported for NEFA and IGF-I.

Perinatal concentrations of 17ß-estradiol and testosterone in the toe claws of female and male dogs from birth until 60 days of age.

This study was conducted to assess concentrations of 17ß-estradiol (E2) and testosterone (T) in toe claws of puppies collected at birth, at 30 and at 60 days of age, evaluating changes relating to age and effect of puppy sex, Apgar score, bodyweight at birth, "litter effect", litter size, and maternal age. Puppies (n = 89), 46 males and 43 females, with normal weight and without malformations, were assigned for the study. Within 12 h of birth tips of toe claws were clipped, and the re-growth tissue of the claws was collected at 30 and 60 days of age. Steroid quantifications occurred using a radioimmunoassay. The results indicated there were lesser concentrations (P < 0.001) of both hormones at 30 and 60 days of age than at birth and that concentrations were similar at 30-60 days of age. There were greater (P < 0.001) T concentrations in males than females, with there being an interaction between sex and sampling time (P < 0.01). The Apgar score was positively (P < 0.001) related to T concentrations in toe claws at birth. The bodyweight was positively correlated (P < 0.05) with T concentrations, with an interaction among puppy sex, bodyweight and sampling time (P < 0.05). Results of the present study confirmed the usefulness of toe claws as a matrix for study of hormonal changes in perinatology of dogs. Results of the study also indicate there are greater E2 and T concentrations at birth compared with 30 and 60 days of age that could be the result of these prenatal steroids affecting fetal development.

Luteal activity and effect of dietary energy restriction on follicular development in lactating cows.

The aim of this research has been to evaluate the presence of anomalies in the ovarian cycle activity during postpartum and to verify whether 72-hr dietary fasting during the dominance phase, the phase before ovulation, might modify the ovarian follicle population. The presence of anomalies in ovarian cycle activity has been evaluated in 30 Italian Friesian cows starting from 20 days postpartum until 211 days of lactation. Long oestrus and brief dioestrus or scarce luteal activity have been the main anomalies found through measuring progesterone concentrations in the whey. Until 100 days of lactation, the BCS values of the problematic animals have been significantly lower than those in animals with normal ovarian activity. After 100 days of lactation, the ovarian anomalies continued to appear despite the fact that all the animals have reached comparable BCS values. Starting from the results of this trial, the effect of 72-hr dietary fasting on dominant follicles has been studied in six cows. Ultrasonography revealed that the diameter of the follicles at 71 days postpartum has been significantly lower than at 181 days. A 72-hr dietary restriction at 101 and 211 days postpartum did not affect the size of the dominant follicle. However, at 101 days postpartum, half of the animals presented follicular cysts. The effect of fasting differed if the animal has been in early postpartum or 211 days of lactation. Further researches are necessary to understand how different metabolic conditions can modify the follicular population but on the other hand the study shows the utility for farmers and field veterinarians of monitoring the resumption of the ovarian cycle postpartum through the whey progesterone concentrations.

Cortisol fetal fluid concentrations in term pregnancy of small-sized purebred dogs and its preliminary relation to first 24 hours survival of newborns.

Along the perinatal period, in mammals, cortisol (C) plays a pivotal role in the final intrauterine fetal maturation and in the early neonatal adaptation. Because of the scarce knowledge about canine perinatology, the present study was aimed to assess the C concentrations in amniotic and allantoic fluids collected, without invasiveness, from small-sized, purebred newborn puppies born by elective cesarean section, at term of pregnancy. Possible correlations between fetal fluid C concentrations and maternal parity, litter size, birth weight, Apgar score, were evaluated. In addition, the possible effect of fetal fluid C concentrations on newborn survival at 24 hours of age, and the effect of the litter or the newborn gender on fetal fluid C concentrations were also assessed. The results, obtained from 50 born alive, normal-weight puppies, without gross physical malformation, showed that C concentration was higher in allantoic than in amniotic fluid (P < 0.01), even if a strong positive correlation between the two fluids C concentration was found (P < 0.0001; R = 0.83). Neither amniotic nor allantoic C concentrations were correlated to maternal parity, litter size, birth weight, and Apgar score. Interestingly, higher amniotic (P < 0.05), but not allantoic, C concentrations were found in puppies not surviving at 24 hours after birth. Therefore, it could be suggested that this parameter may be useful for the recognition, at birth, of puppies needing special surveillance during the first day of age. A significant (P < 0.001) effect of the litter in both amniotic and allantoic C concentrations was found. In conclusion, the present results showed that in small-sized purebred puppies, born at term by elective cesarean section, the exact fetal, maternal, or placental source contributing to fetal fluid C concentrations remains to be clarified. From a clinical perspective, however, the evaluation of amniotic C concentration at birth seems useful for the detection of puppies that need special surveillance during the first 24 hours of age, and should be coupled to the early newborn evaluation by Apgar score. However, the small total number of newborns, and especially of the dead puppies enrolled in the present study, suggests that further, more-focused investigations on a large number of subjects are needed before the method could be considered for application in the clinical practices.

Coat and claws as new matrices for noninvasive long-term cortisol assessment in dogs from birth up to 30 days of age.

The last stage of fetal development and the neonatal period represent the most critical phases for the mammals' offspring. In the dog, the knowledge about the final intrauterine fetal development and biology, as well as about the neonatal physiology, remains scarce. Hormonal changes occurring in the last intrauterine fetal phase and during the early neonatal age are still not completely clear, probably because of the invasiveness related to the collection of the more common biological matrix, represented by circulating blood. Toward term of pregnancy, during parturition, and after birth, the hypothalamic-pituitary-adrenal axis is a key system regulating several physiological processes, and its activity was previously investigated by blood analysis, considered an invasive procedure providing a single-point measurement. In respect to animal welfare, and for a more correct long-term retrospective investigation, noninvasive hormonal studies were performed firstly on the hair of humans and coat of animals and, more recently, in the nails of human beings. This study was aimed to assess cortisol (COR) in coat and claws of newborn puppies and to evaluate the possible influence of the newborn gender, breed body size, and age on coat and claws COR concentrations. The results obtained from 165 newborn puppies evidenced that coat and claws COR levels were highly correlated each other (P < 0.0001), although the COR accumulation in the two matrices was different in relation to the class of age. Moreover, the puppies age influenced both coat and claws COR concentrations (P < 0.05), with premature puppies showing higher values when compared to term born-dead puppies or puppies dead between 1 and 30 days of age. The present study reported that COR is quantifiable in coat and claws of newborn dogs. Moreover, both matrices appear as useful tools for new, noninvasive, long-term perinatal and neonatal researches also in canine species.

Study of progesterone and cortisol concentrations in the Italian Friesian claw.

The present research was conducted to study progesterone and cortisol concentrations in the claw of cattle and to verify whether the cattle claw could be considered an efficient matrix to provide retrospective information regarding progesterone and cortisol concentrations related to pregnancy and peripartum periods. These 2 steroids are involved in hoof growth. The study was performed on 32 calves and 24 pregnant milking cows of the Holstein breed, which were clinically healthy and lacking any claw disorders. Samples of at least 0.5cm in thickness were taken from the sole. Progesterone and cortisol concentrations were determined by RIA. The cortisol concentration in the horny shoe of calves from 0 to 30 d of age was significantly higher than the concentration at 31 to 60 and 61 to 120 d of age. Conversely, the progesterone concentration showed no statistically significant difference in relation to age. The horn progesterone concentrations recorded in the milking dairy cows at 7 mo of pregnancy showed high variability in the horizontal sections of the sole (the individual coefficient of variation ranged between 0.09 and 1.11). In 6 cows, genuine extreme values (genuine outliers) of the progesterone level were found. Moreover, significant differences existed among the progesterone concentrations of the sole's transverse sections. We detected a significant positive correlation between the weight of the horn samples after freeze-drying and their weight after hydration. The cortisol and progesterone levels in soaked horn samples were found to be significantly lower than in the same dry samples. These results show that cortisol and progesterone can be measured in the cattle claw horn. The claws of mature dairy cows could not be used as a matrix to provide a retrospective measure of cumulative hormone secretion, whereas the analysis of the calves' claw horns showed retrospective hormonal information similar to hair samples.

Short communication: hair cortisol concentrations in Holstein-Friesian and crossbreed F1 heifers.

The aim of this study was to evaluate the hypothalamic-pituitary-adrenal (HPA) axis activity of Holstein-Friesian and crossbreed F1 heifers by analysis of the cortisol concentrations in hair samples. Cortisol, the primary hormone of the HPA axis, is the biological endpoint for the investigation of the HPA response. The study was conducted on 290 prepubertal heifers; 142 heifers were pure Holstein-Friesian and 148 were crossbreed F1 heifers obtained from the 3-way rotational system with Swedish Red and Montbéliarde breeds. Extraction was performed on the hair using methanol, and cortisol concentrations were determined by a radioimmunoassay method. Cortisol concentrations measured in regrown hair of crossbreed F1 heifers were significantly lower than those in hair of Holstein-Friesian heifers. This result helps us to better understand the differences in HPA activity and allostatic load between Holstein-Friesian and crossbreed F1 heifers and allows us to better assess the adaptability of these animals to the environment and the importance of crossbreed traits for profitability in dairy farming.

Candidate gene marker associations with fatty acid profiles in heavy pigs.

Heavy pigs are used in Italy to produce fresh meat, dry-cured ham, salami and lard. Fatty acid profile determines both the dietary and organoleptic qualities of these products. The objective was to study the polymorphisms of two genes that code for enzymes of the fatty acid metabolism, namely, Stearoyl-CoA desaturase (SCD) and Fatty acid desaturase 2 (FADS2). We also investigated the polymorphism of the Sterol regulatory element binding protein gene (SREBF1) as it regulates SCD and FADS2 transcription. Significant associations of SCD were found with the ratio of oleic to stearic acid; with the concentration (g/100g of fat) of stearic and oleic acids. The concentrations of arachidonic and linoleic acids and the ratio of dihomo-gamma-linolenic to linoleic acid were significantly associated with FADS2 polymorphisms. SREBF1 polymorphism was associated with oleic acid concentrations and the ratio of arachidonic to linoleic acids. Our findings suggest that the SCD, FADS2 and SREBF1 polymorphisms are associated with dietary quality of heavy pig meat products.

Insulin-like growth factor I: could it be a marker of prematurity in the foal?

Insulin-like growth factor (IGF)-I represents one of the most important growth regulators, playing a central role in fetal and neonatal growth. Plasma IGF-I levels increase rapidly after birth, and they are influenced by numerous factors, including sex, age, nutritional state, and premature birth. The aims of this study were: (1) to evaluate the IGF-I plasma profile in healthy newborn foals during the first 2 weeks of life; (2) to assess the possible influence of sex and birth weight on this hormone; (3) to analyze the percentage increment of IGF-I values in healthy foals; (4) to evaluate the influence of prematurity on IGF-I profile; (5) to verify the role of IGF-I as a diagnostic marker of prematurity; and (6) to analyze the percentage increment of IGF-I in premature foals. Thirty-four healthy term foals were enrolled as the control group and from each foal plasma was collected within 6 hours from birth, at 12 hours, daily from Day 1 to Day 7, and at Days 10 and 14 after birth. Eleven foals aged younger than 1 week and diagnosed as premature and hospitalized at a Equine Perinatology Unit were also enrolled; from each foal plasma was collected daily from the day of admission to discharge or death. Insulin-like growth factor I was analyzed by RIA. In the control group, an increasing trend of IGF-I concentrations was found, with higher values from Day 4 to 10 compared with data obtained at less than 6 hours of life, and from Day 5 to 10 compared with 12 and 24 hours and 3 days. No differences were found in healthy foals analyzed in relation to birth weight and sex. In premature foals an increasing trend was observed but no statistical differences were found among sampling times, and no differences were found between healthy and premature foals. The IGF value in premature foals at admission was always higher compared with the lowest recorded level in healthy age-matched foals, thus this parameter does not seem to have a diagnostic role for prematurity in foals. Finally, the evaluation of the percentage increment of IGF-I concentrations showed a significant increase in full-term foals on Day 5, 6, 7, and 10 compared with 12 and 24 hours, and no differences were observed in premature foals. In conclusion, prematurity in newborn foals seems to affect only partially IGF-I plasma concentrations and it does not seem to be a reliable marker for this pathological condition.

Corpus luteum development and function and relationship to pregnancy during the breeding season in the Mediterranean buffalo.

The aim of this study was to ascertain corpus luteum (CL) development and function in buffaloes synchronized and mated by artificial insemination (AI) during the breeding season. Italian Mediterranean buffalo cows (n = 43) at 86.5 ± 2.7 days postpartum were synchronized by the Ovsynch-TAI Program and inseminated using frozen thawed semen at 20 and 44 h after the second injection of GnRH. The CL dimensions (diameter and area) and blood flow were examined on Days 5, 10, 15, 20, and 25 after AI by realtime B-mode/colour-Doppler ultrasonography. The resistive index (RI), pulsatility index (PI) and time average medium velocity (TAMV) were recorded at each time, together with CL dimensions. Blood samples were taken on the days of ultrasonography for progesterone (P4) assay by RIA. Data were grouped into pregnant or non-pregnant and retrospectively analyzed by repeated measure ANOVA and correlation analyses. Dimensions of the CL on Days 10, 20, and 25 after AI were greater (P < 0.01) in buffaloes pregnant on Day 45 (n = 18) compared with non-pregnant buffaloes (n = 25). The former buffaloes also showed a greater (P < 0.01) rate of CL growth between Days 5 and 10 after AI. Blood flow to the CL on Day 10 after AI showed a higher TAMV (P < 0.01) and lower RI (P < 0.05) in pregnant buffaloes compared with non-pregnant buffaloes. Negative correlations were observed on Day 10 after AI between CL diameter and RI (r = -0.61; P < 0.01) and PI (r = -0.60; P < 0.01); P4 concentrations and RI (r = -0.46; P < 0.02); and RI and pregnancy (r = 0.45; P < 0.02). Positive correlations were observed between pregnancy and CL size (r = 0.54; P < 0.01), ΔCL diameter between Days 5 and 10 (r = 0.52; P < 0.01), ΔCL area between Days 5 and 10 (r = 0.48; P < 0.015), and ΔP4 between Days 5 and 10 (r = 0.50; P < 0.01). Based on these findings it is concluded that the period between Day 5 and 10 is very important for CL growth and crucial in evaluating pregnancy. Accordingly, the assessment of CL parameters during the period from Day 5 to Day 10 after AI might be used to predict the likelihood of an ongoing pregnancy.

Growth, metabolic status and ovarian function in buffalo (Bubalus bubalis) heifers fed a low energy or high energy diet.

The aim was to establish the capacity of buffalo heifers to adapt their metabolic requirements to a low energy diet. Murrah buffalo (Bubalus bubalis) heifers undergoing regular estrous cycles were randomly assigned by age, live weight (LW) and body condition score (BCS) to a high energy group (HE, 5.8 milk forage units (MFU)/day, n=6) or low energy group (LE, 3.6 MFU/day, n=6). Circulating concentrations of metabolic substrates, metabolic hormones and reproductive hormones were determined weekly for 19 weeks. Ovarian follicular characteristics and oocyte parameters were also ascertained weekly. Heifers fed the LE diet had a better dry matter conversion than heifers fed the HE diet and the calculated daily energy provision was negative for heifers fed the LE diet (-0.248 MFU) and positive for heifers fed the HE diet (5.4 MFU). Heifers fed the HE diet had an increase in 50 kg LW over the duration of the study whereas LW remained constant for heifers fed the LE diet. The BCS of heifers fed the HE diet (4.2) was greater (P<0.05) than the BCS for heifers fed the LE diet (3.4). Heifers fed the HE diet had greater (P<0.05) circulating concentrations of metabolic substrates (glucose, total cholesterol and HDL cholesterol) and metabolic hormones (insulin, glucagon, leptin and T3) compared with heifers fed the LE diet. There were no significant differences in circulating reproductive hormones between the two groups of heifers. Ovarian follicular characteristics were similar for the two groups of heifers while heifers fed the LE diet tended to have oocytes of reduced quality compared with heifers fed the HE diet. The most notable finding was that heifers fed the LE diet had a negative calculated daily energy provision but were able to maintain LW and reproductive activity. It was concluded that buffalo heifers may potentially have the capacity to undergo metabolic adjustment and reduce their energy requirements when dietary energy is limiting. This adaptive capacity would explain why buffaloes remain productive in environments that are limiting to other ruminants.

Candidate gene markers involved in San Daniele ham quality.

San Daniele dry-cured hams (also known as prosciutto) are produced in the Northeastern region of Italy. This high value product requires high quality fresh meat to avoid processing problems. The Sterol Regulatory Element Binding Protein-1 (SREBF1) is a transcription factor involved in the regulation of fatty acid synthesis in muscle and adipose tissues. The SREBF1 gene, its regulating genes SCAP and MBTPS1, and one of its target genes, SCD, were investigated for associations with several meat quality traits of San Daniele hams. Significant associations of some gene markers were found with carcass weight, lean percentage, backfat thickness, ham green weight, ham fat cover thickness, shear force (WBSF), salting losses and instrumental colour of both lean and fat. These findings provide initial evidences that SNPs in SREBF1, SCAP, MBTPS1 and SCD are associated with San Daniele ham quality and may be considered as markers for selective breeding programs.

Corpus luteum function and pregnancy outcome in buffaloes during the transition period from breeding to non-breeding season.

The aim in this study was to investigate corpus luteum function and embryonic loss in buffaloes mated by artificial inseminations (AI) during the transitional period from breeding to non-breeding season. The study was carried out using 288 multiparous Italian Mediterranean Buffalo cows at 110 ± 4 days in milk. The buffaloes were mated by AI after synchronization of ovulation by the Ovsynch-TAI protocol 25 days after AI buffaloes underwent trans-rectal ultrasonography to assess embryonic development. Pregnancy diagnosis was confirmed on Days 45 and 70 after AI by rectal palpation. Buffaloes pregnant on Day 25 but not on Day 45 were considered to have undergone late embryonic mortality (LEM), whilst buffaloes pregnant on Day 45 but not on Day 70 were considered to have undergone foetal mortality (FM). Corpus luteum size and blood flow were determined by real-time B-mode/colour-Doppler on day 10 after AI in 122 buffaloes. The resistive index (RI) and pulsatility index (PI) were recorded at the time. Milk samples were collected on Days 10, 20 and 25 after AI in all inseminated buffaloes for the assay of whey P4 concentrations. Data were analysed by anova. Pregnancy rate on Day 25 after AI was 48.6% (140/288) and declined to 35.4% (102/288) and 30.6% (88/288) by Day 45 and Day 70 respectively. The incidences of LEM and FM were respectively 27.1% (38/140) and 13.7% (14/102). Pregnant buffaloes had greater (p < 0.01) whey concentrations of P4 from Day 20 onwards than buffaloes which showed LEM, whilst P4 in buffaloes that showed FM did not differ from the other two groups on Day 10 and Day 20. Corpus luteum blood flow on Day 10 after AI showed higher RI (p < 0.05) and PI (p = 0.07) values in buffaloes that subsequently were not pregnant on Day 25 compared with pregnant buffaloes. Buffaloes that were not pregnant on Day 45 also had a higher (p = 0.02) RI value on Day 10 than pregnant buffaloes, whilst PI values on Day 10 did not differ for the two groups of buffaloes. It was concluded that blood flow to the corpus luteum on Day 10 after AI influences corpus luteum function as judged by P4 secretion and also embryonic development and attachment in buffaloes.

Is a delayed treatment with GnRH, HCG or progesterone beneficial for reducing embryonic mortality in buffaloes?

The aims of this study were to verify the efficacy of delayed hormonal treatments performed on day 25 post-insemination on pregnancy rate at 45 and 70 days in buffalo. The trial was performed on 385 buffaloes synchronized by the Ovsynch/TAI protocol and submitted to artificial insemination (AI). Twenty-five days after AI, pregnant animals were assigned to four treatments: (1) GnRH agonist (n = 52), 12 microg of buserelin acetate; (2) hCG (n = 51), 1500 IU of human chorionic gonadotrophin; (3) Progesterone (n = 47), 341 mg of P4 intramuscular (im) every 4 days for three times; (4) Control (n = 54), treatment with physiological saline (0.9% NaCl). Milk samples were collected on days 10, 20 and 25 after AI in all buffaloes to determine progesterone concentration in whey by radioimmunoassay method. Statistical analysis was performed by anova. Pregnancy rate on day 25 after AI was 52.9%, but declined to 41.8% by day 45, indicating an embryonic mortality (EM) of 21%. If only control group is considered, the incidence of EM was 38.9%. Pregnant buffaloes had higher (p < 0.01) progesterone concentrations on day 20 and 25 after AI than both non-pregnant buffaloes and buffaloes that showed EM. The treatments on day 25 increased (p < 0.01) pregnancy rate, although in buffaloes with a low whey progesterone concentration on day 20 and 25 after AI (n = 22); all treatments were ineffective to reduce EM.

Plasma and follicular fluid fatty acid profiles in dairy cows.

Composition of follicular fluid to which the preovulatory follicle is exposed may be one of the major factors determining subsequent fertility, as fatty acids are a precursor of hormones involved in dominance, ovulation and atresia mechanisms. The objective of this paper is to observe fatty acid profiles in various lipid classes according to estrogenic activity of follicles. For each of the 18 cows, we analysed plasma and follicular fluid fatty acid profiles of phospholipids, non-esterified fatty acid (NEFA), cholesteryl esters and triglycerides fractions. Follicles were classified as active (ratio oestrogen to progesterone E2/P4 > 1) and inactive (E2/P4 < 1). For seven cows, we get both types of follicles, six had only one active follicle and five cows had only one inactive follicle. The NEFA profile for palmitic acid, stearic acid, oleic acid, linoleic acid, C20:3n6, arachidonic acid and docosapentaenoic acid (DPA; p < 0.001) is different between inactive and active follicles and plasma. Compared with active follicular fluid and plasma, follicular fluid of inactive follicles showed lower stearic acid, higher oleic acid, arachidonic acid and DPA (p < 0.05) in phospholipids. No significant differences were observed in the cholesteryl ester fraction, which is composted mainly of linoleic acid. Triglyceride concentrations were too low to get reliable results. This study suggests that follicles have a specific fatty acid metabolism depending on oestrogen activity.

Delayed treatment with GnRH agonist, hCG and progesterone and reduced embryonic mortality in buffaloes.

The present study examined the effect of delayed treatment with tropic hormones and progesterone (P4) on embryonic mortality in buffaloes. Buffaloes with a conceptus on Day 25 after AI were assigned to the following treatments: Control (n=41), i.m. physiological saline; GnRH agonist (n=36), i.m. 12 microg buserelin acetate; hCG (n=33), i.m. 1500 IU hCG; P4 (n=38), i.m. 341 mg P4 every 4 days on three occasions. Control buffaloes had an embryonic mortality of 41.4% (17/41) between Days 25 and 45, and this was reduced (P<0.01) by treatment with GnRH agonist (11.1%, 4/36), hCG (9.0%, 3/33) and P4 (13.1%, 5/38). On Day 45, buffaloes treated with hCG and which ovulated had greater (P<0.05) concentrations of P(4) in whey (453 +/- 41 pg/ml) than buffaloes in the same treatment that did not ovulate (297 +/- 32 pg/ml). A similar but non-significant trend was observed for buffaloes treated with GnRH agonist. It was concluded from the findings that the treatment of buffaloes on Day 25 after AI with tropic hormones or P4 is beneficial to processes associated with embryonic implantation.