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.

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.

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.

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.

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.

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.

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.

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.

Pregnancy diagnosis in dairy cows by whey progesterone analysis: an ROC approach.

Concentration of progesterone in milk may be used to predict pregnancy status of dairy cattle by the 21st day after insemination. However, the accuracy of this method may be affected by fat-solubility of progesterone and sample storage conditions. After coagulation of a milk sample with rennet, an alternative method is to quantify progesterone concentration in whey with a novel, validated EIA. In this experiment, a receiver operating characteristic (ROC) analysis was performed to estimate the optimal discrimination point for whey progesterone concentration, using a sample of 991 Friesian cows evaluated between the 42nd and 44th day after insemination. Cows also were diagnosed for pregnancy by rectal palpation at this time. The overall conception rate at palpation was 57%. ROC analysis indicated that 259 pg/mL progesterone in whey was the most effective cutoff to discriminate correctly between pregnant and non-pregnant cows. Using this point for prediction, sensitivity was 98.2%, specificity was 70.9% and the area under ROC curve was 0.859, levels generally considered to denote moderate accuracy. The negative likelihood ratio at the cutoff of 259 pg/mL was 0.02, indicating satisfactory performance in detecting negative subjects, while the positive likelihood ratio (+LR=3.37) suggested average performance. In conclusion, EIA of progesterone concentration in whey is a viable method for predicting pregnancy status in cows. However, operators should take management objectives for the herd into account in determining the cutoff point and also considering important influencing variables such as conception rate in the herd. This method can provide diagnostic support for efforts to improve reproductive success, especially in low-fertility herds.

Corpus luteum function and embryonic mortality in buffaloes treated with a GnRH agonist, hCG and progesterone.

The effect of treatment with a GnRH agonist, hCG or progesterone (P(4)) on corpus luteum function and embryonic mortality was investigated in buffaloes inseminated during mid-winter. Italian Mediterranean buffaloes (n=309) were synchronized using the Ovsynch with timed-AI program and mated by AI at 16 h (Day 0) and 40 h after the second injection of GnRH. On Day 5, buffaloes were randomly assigned to four groups: Control (no treatment, n=69), GnRH agonist (buserelin acetate, 12.6 microg, n=73), hCG (1500 IU, n=75) and P(4) (PRID without E(2) for 10 days, n=77). Progesterone (pg/ml) was determined in milk whey on Days 5, 10, 15 and 20 and pregnancy diagnosis was undertaken on Day 26 by ultrasound and Day 40 by rectal palpation. Treatment with buserelin and hCG increased (p<0.05) P(4) on Day 15 compared with controls (456+/-27, 451+/-24 and 346+/-28 pg/ml, respectively). Buffaloes treated with a PRID had intermediate P(4) concentrations (380+/-23 pg/ml). Embryonic mortality between Days 26 and 40 (22.9%) and pregnancies at Day 40 (48.9%) did not differ between treatments. A higher (p<0.01) P(4) concentration was found on Day 20 in pregnant animals compared with non-pregnant and embryonic mortality buffaloes, which did not differ. In summary, buserelin and hCG increased P(4) concentrations on Day 15 but this was not associated with a reduced incidence of embryonic mortality in buffaloes during mid-winter.

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.

Technical note: Direct enzyme immunoassay of progesterone in bovine milk whey.

A simple extraction-free or direct quantitative ELISA for progesterone in bovine milk whey was developed. Whey samples are easy to collect, transport, and store. This method also allows for monitoring progesterone levels in cattle, which is important in reproductive management. The assay was designed to cover the concentration range 0.05 to 2 ng/mL, and the sensitivity of the method was 1.5 pg/mL. The intra- and interassay coefficients of variation were 8 and 12%, respectively. A high correlation (r = 0.90) between ELISA and radioimmunoassay measurements of progesterone in the same milk whey samples was obtained. The method can be easily applied in practice because samples can be stored at room temperature (22 to 26 degrees C) for 4 d. Moreover, because analysis requires milk coagulation, that process can be initiated during transport by standard mail services to the laboratory. Upon arrival at the laboratory, whey can be kept refrigerated for 1 wk before analysis. This tool is useful for monitoring luteal activity of dairy cows.

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.

Relation between myocardial contractility, hypertrophy and pump performance in patients with chronic aortic regurgitation: an echocardiographic study.

The aim of this study was to establish prognostic indices and to detect irreversible left ventricular dysfunction before aortic valve replacement in patients with chronic aortic regurgitation. Therefore, we determined the left ventricular pump/contractility relation (expressed as a ration between ejection fraction and peak systolic pressure/end-systolic volume), afterload mismatch (the relation between the ejection fraction and mean systolic wall stress) and the left ventricular end-diastolic radius/posterior wall thickness ratio in 52 patients with chronic aortic regurgitation by means of M-mode echocardiography. These indices were also calculated in 14 patients with idiopathic dilated cardiomyopathy and in 20 normal controls. The indices allowed separation of normals and patients with mild to moderate aortic insufficiency from patients with idiopathic dilated cardiomyopathy indicating that these indices could distinguish between patients with a reversible afterload mismatch and those with irreversible impaired muscle function. The 38 patients with severe aortic insufficiency showed a large overlap between normal and abnormal indices. Twelve of these patients had a pump/contractility index and a ratio between ejection fraction and wall stress similar to that found in patients with idiopathic dilated cardiomyopathy. All these patients, however, had a left ventricular end-diastolic radius/thickness ratio greater than 4 indicating "inadequate hypertrophy". Of these, 3 patients died perioperatively and 4 developed congestive heart failure within 12 months postoperatively. We conclude that an end-diastolic radius/thickness ratio greater than 4 is a sensitive but not a specific preoperative indicator of irreversible left ventricular damage in patients with chronic aortic regurgitation. This index is readily obtained preoperatively by M-mode echocardiography.

Candidate gene markers associated with somatotropic axis and milk selection.

One of the obstacles to progress in dairy cattle selection is that milk production traits are only expressed after the first calving. However, the use of the quantitative trait loci (QTL) technology will improve the efficiency of dairy industry with a positive image for the consumers. QTL are part of the genome showing a preponderant action and explaining the major part of variation of the trait production. At the present time, the two major strategies developed to detect such QTL are the candidate gene approach and the positional genetics approach. The somatotropic axis contains the most promising candidates in this respect, as it strongly regulates milk production. Then, the identification of favorable QTL associated with the somatotropic axis that are significantly correlated with genetic merits for milk production could lead to more effective selection programs.

Evaluation of the reproductive efficiency of simmental and Friesian dairy cows farmed in Friuli (Northeast Italy) by the analysis of progesterone levels.

The present research was conducted 1) to investigate the usefulness of progesterone profiles for the study of reproductive efficiency, 2) to identify new reproductive indexes and 3) to examine the possibility of modelling changes in the biological status of animals. Two groups of dairy cows were chosen for this investigation: Italian Simmental and Italian Friesian cows. Milk samples were taken from each cow every 3 to 4 d between Day 10 and Day 240 after calving, and whey progesterone was determined by RIA. The cows were classified into the following 4 categories on the basis of their luteinic activity: 1) Acyclic cows with P4 values <100 pg/ml; 2) Cyclic cows with a normal cycle (1 or 2 samples <100 pg/ml, followed by 1 sample >/=120 pg/ml, followed by at least 3 samples >/=200 pg/ml, followed by 1 sample <100 pg/ml) or a short cycle (1 or 2 samples <100 pg/ml, followed by 1 sample >/=120 pg/ml, followed by 2 samples >/= 200 pg/ml, followed by 1 sample <100 pg/ml followed by a normal cycle); 3) pregnant cows with P4 values >120 pg/ml at 21 to 24 d from AI; and 4) cows with atypical luteinic activity and P4 values >120 pg/ml in a single sample. Analysis of the P4 frequency distribution indicated that ovarian activity resumed from Day 15 and Day 18 after calving in the Italian Friesian and Italian Simmental cows, respectively, and the highest percentage of cyclic cows was observed on Day 60 for Italian Friesian cows (45%) and Day 70 for Italian Simmental cows (55%). The acyclic phase was considered to be concluded by Day 164 and Day 120 for Italian Friesian and Italian Simmental cows, respectively. By Day 240, 66% of Italian Friesian cows and 79% of Italian Simmental cows were pregnant. The percentage of cows with atypical luteinic activity was almost constant throughout the experiment (Italian Friesian=5 to 10%, Italian Simmental=10 to 15%). The calving-conception interval was 104 d for Italian Friesian cows and 94 d for Italian Simmental cows. In both groups, the area on the P4 frequency diagram representing the acyclic cows was not very different from that representing the cyclic cows, while the area occupied on the diagram by pregnant animals was larger. These results are derived from an accurate mathematical analysis. The average number of times in the acyclic status for both groups of cows was about 3-fold smaller than the average number in the cyclic status.