Connecting the Dots: Livestock Animals as Missing Links in the Chain of Microplastic Contamination and Human Health.

Plastic pollution is a global diffuse threat, especially considering its fragmentation into microplastics (MPs) and nanoplastics (NPs). Since the contamination of the aquatic environment is already well studied, most studies have now focused on the soil. Moreover, the number of studies on the exposure routes and toxic effects of MNPs in humans is continuously increasing. Although MNPs can cause inflammation, cytotoxicity, genotoxicity and immune toxicity in livestock animals, which can accumulate ingested/inhaled plastic particles and transfer them to humans through the food chain, research on this topic is still lacking. In considering farm animals as the missing link between soil/plant contamination and human health effects, this paper aims to describe their importance as carriers and vectors of MNP contamination. As research on this topic is in its early stages, there is no standard method to quantify the amount and the characteristics of MNPs in different matrices. Therefore, the creation of a common database where researchers can report data on MNP characteristics and quantification methods could be helpful for both method standardization and the future training of an AI tool for predicting the most abundant/dangerous polymer(s), thus supporting policy decisions to reduce plastic pollution and perfectly fitting with One Health principles.

Self-assembly of glycoprotein nanostructured filaments for modulating extracellular networks at long range.

The intriguing capability of branched glycoprotein filaments to change their hierarchical organization, mediated by external biophysical stimuli, continues to expand understanding of self-assembling strategies that can dynamically rearrange networks at long range. Previous research has explored the corresponding biological, physiological and genetic mechanisms, focusing on protein assemblies within a limited range of nanometric units. Using direct microscopy bio-imaging, we have determined the morpho-structural changes of self-assembled filament networks of the zona pellucida, revealing controlled levels of structured organizations to join distinct evolved stages of the oocyte (Immature, Mature, and Fertilized). This natural soft network reorganizes its corresponding hierarchical network to generate symmetric, asymmetric, and ultimately a state with the lowest asymmetry of the outer surface roughness, and internal pores reversibly changed from elliptical to circular configurations at the corresponding stages. These elusive morpho-structural changes are regulated by the nanostructured polymorphisms of the branched filaments by self-extension/-contraction/-bending processes, modulated by determinate theoretical angles among repetitive filament units. Controlling the nanoscale self-assembling properties by delivering a minimum number of activation bio-signals may be triggered by these specific nanostructured polymorphic organizations. Finally, this research aims to guide this soft biomaterial into a desired state to protect oocytes, eggs, and embryos during development, to favour/prevent the fertilization/polyspermy processes and eventually to impact interactions with bacteria/virus at multiscale levels.

Impact of Microplastics and Nanoplastics on Livestock Health: An Emerging Risk for Reproductive Efficiency.

Pollution due to microplastics and nanoplastics is one of the major environmental issues of the last decade and represents a growing threat to human and animal health. In aquatic species, there is a large amount of information regarding the perturbation of marine organisms; instead, there are only a few studies focusing on the pathophysiological consequences of an acute and chronic exposure to micro- and nanoplastics in mammalian systems, especially on the reproductive system. There are several studies that have described the damage caused by plastic particles, including oxidative stress, apoptosis, inflammatory response, dysregulation of the endocrine system and accumulation in various organs. In addition to this, microplastics have recently been found to influence the evolution of microbial communities and increase the gene exchange, including antibiotic and metal resistance genes. Special attention must be paid to farm animals, because they produce food such as milk, eggs and meat, with the consequent risk of biological amplification along the food chain. The results of several studies indicate that there is an accumulation of microplastics and nanoplastics in human and animal tissues, with several negative effects, but all the effects in the body have not been ascertained, especially considering the long-term consequences. This review provides an overview of the possible adverse effects of the exposure of livestock to micro- and nanoplastics and assesses the potential risks for the disruption of reproductive physiological functions.

Carotenoids in female and male reproduction.

Carotenoids are among the best-known pigments in nature, confer color to plants and animals, and are mainly derived from photosynthetic bacteria, fungi, algae, plants. Mammals cannot synthesize carotenoids. Carotenoids' source is only alimentary and after their assumption, they are mainly converted in retinal, retinol and retinoic acid, collectively known also as pro-vitamins and vitamin A, which play an essential role in tissue growth and regulate different aspects of the reproductive functions. However, their mechanisms of action and potential therapeutic effects are still unclear. This review aims to clarify the role of carotenoids in the male and female reproductive functions in species of veterinary interest. In female, carotenoids and their derivatives regulate not only folliculogenesis and oogenesis but also steroidogenesis. Moreover, they improve fertility by decreasing the risk of embryonic mortality. In male, retinol and retinoic acids activate molecular pathways related to spermatogenesis. Deficiencies of these vitamins have been correlated with degeneration of testis parenchyma with consequent absence of the mature sperm. Carotenoids have also been considered anti-antioxidants as they ameliorate the effect of free radicals. The mechanisms of action seem to be exerted by activating Kit and Stra8 pathways in both female and male. In conclusion, carotenoids have potentially beneficial effects for ameliorating ovarian and testes function.

Environmental Factors Affecting the Reproductive Efficiency of Italian Simmental Young Bulls.

The objectives of the study aimed to evaluate the effect of weather conditions and scrotal circumference (SC) on standard semen characteristics of Italian Simmental young bulls (n = 577), all raised in the same performance station and sampled by the artificial vagina (AV) method. Considering the increasing SC, the results showed a significant increase in quantitative semen parameters (p < 0.05 and p < 0.0001, for sperm concentration and total number of spermatozoa, respectively); for every extra centimeter of SC, 17.5 × 106 spermatozoa/mL and 0.102 × 109 of total spermatozoa were produced. The age of the animal at semen collection (395 and 465 days) had similar positive effects. The effect of the average temperature humidity index (THI limits ≤ 40 and ≥70) in the previous 60, 30, and 10 days before the semen collection was also considered. Sixty days before the semen collection, the increasing THI increased both primary (p < 0.0001) and secondary (p < 0.0001) abnormalities while the percentage of morphologically normal spermatozoa decreased (p < 0.0001). Thirty and ten days before collection, the same effect on morphological traits was maintained, but total and progressive motility was also influenced (p < 0.01) with an unexpected increasing pattern. Thus, environmental conditions can influence semen quality during the entire spermatogenesis and results can guide future research on this breed.

Delayed First Milking in Unassisted Overnight Calving Did Not Affect the Quality of Colostrum but Influenced Serum Brix Refractometry in Holstein Calves at Two Days of Life.

Timely administration of good-quality colostrum represents the first farm strategy to avoid the failure of passive transfer (FPT). However, calves born during the night are likely to be fed later than recommended. Our aim was to evaluate whether night-occurring calving and delayed first milking affected colostrum quality and immune passive transfer. The dataset included 463 calvings. Four liters of colostrum were administered by an esophageal tube feeder. The mean Brix% of colostrum was 27.43%, while serum Brix% at two days of life in calves was 10.19%. According to the Generalized Linear Model, parity ≥ 4, calving months of March, April, and from September to November positively influenced the quality of colostrum. Dams carrying a male calf produced lower quality colostrum compared with those carrying a female calf (−2.78 ± 1.04 Brix%, p = 0.008); heavier female calves were associated with greater colostrum quality (0.29 ± 0.05 for each kg increase, p < 0.001). Night- or day-calving had no effect on the quality of colostrum. The only factor influencing the serum Brix% of female Holstein calves at two days of life was the day- or night-occurring birth (−0.386 ± 0.188 Brix% in calves born during the night, p = 0.04). Our results showed that calves born overnight and fed the day after had decreased serum Total Protein concentrations as indicated by reduced Brix refractometer readings, compared with calves born during the day and fed quickly after birth. However, the administration of 4 L of high-quality colostrum likely improved their serum Brix% at two days of life. Alternatively, where the prevalence of good-quality colostrum is lower, improving calving supervision and ensuring timely feeding are important to reduce the risk of FPT.

Effect of Culling Management Practices on the Seroprevalence of Johne's Disease in Holstein Dairy Cattle in Central Italy.

A study was performed in Umbria, central Italy, to find out whether different culling strategies adopted by farms to control Johne's disease (JD) infection exerted effects on the seroprevalence in dairy cattle. Fifty Fresian dairy herds in the Perugia and Assisi districts were visited and an audit of herd management was conducted. Among the 50 herds, 20 were selected for the consistency of management practices and, according to the culling strategy, two groups were created: group A (aggressive culling protocol, with average herd productive life <1100 days) and group B (lower culling rate, with productive life greater than 1500 days). The presence of antibodies to Mycobacterium avium subspecies paratuberculosis (Map) in the serum was determined using a commercial enzyme-linked immunosorbent assay (ELISA) kit. It was found that 3.3% (n = 14) of the cows of group B (n = 422, from 17 herds) were positive for Map antibodies, in comparison with 5.7% (n = 21) of the cows from group A (n = 366, from three herds). The odds ratio from multiple logistic regression (adjusted odds ratio 2.446, 95% confidence interval 0.412 to 14.525) showed that Johne's disease prevalence in herds with a greater productive life was not higher than in herds with typical modern management characterized by more aggressive culling. This is a significant finding, indicating that aggressive culling may not be necessary. Current JD control recommendations are derived from data obtained in high-prevalence paratuberculosis areas (northern Europe, including northern Italy), while methods of information transfer to dairy farms in low-prevalence areas should be reassessed to ensure that the correct measures, including basic calving management and calf-rearing practices, are thoroughly implemented. Using the manufacturer's suggested cut-off for a positive ELISA test and the sensitivity and specificity claimed, the overall true prevalence in Umbria dairy cattle was calculated as 7% (95% confidence interval 5.2% to 8.8%).

How to Predict Parturition in Cattle? A Literature Review of Automatic Devices and Technologies for Remote Monitoring and Calving Prediction.

Cattle farming is facing an increase in number of animals that farmers must care for, together with decreasing time for observation of the single animal. Remote monitoring systems are needed in order to optimize workload and animal welfare. Where the presence of personnel is constant, for example in dairy farms with great number of lactating cows or with three milking/day, calving monitoring systems which send alerts during the prodromal stage of labor (stage I) could be beneficial. On the contrary, where the presence of farm personnel is not guaranteed, for example in smaller farms, systems which alert at the beginning of labor (stage II) could be preferred. In this case, time spent observing periparturient animals is reduced. The reliability of each calving alarm should also be considered: automatic sensors for body temperature and activity are characterized by a time interval of 6-12 h between the alarm and calving. Promising results have been shown by devices which could be placed within the vaginal canal, thus identifying the beginning of fetal expulsion and optimizing the timing of calving assistance. However, some cases of non-optimal local tolerability and cow welfare issues are reported. Future research should be aimed to improve Sensitivity (Se), Specificity (Sp) and Positive Predictive Value (PPV) of calving alert devices in order to decrease the number of false positive alarms and focusing on easy-to-apply, re-usable and well tolerated products.

In Vitro Folliculogenesis in Mammalian Models: A Computational Biology Study.

In vitro folliculogenesis (ivF) has been proposed as an emerging technology to support follicle growth and oocyte development. It holds a great deal of attraction from preserving human fertility to improving animal reproductive biotechnology. Despite the mice model, where live offspring have been achieved,in medium-sized mammals, ivF has not been validated yet. Thus, the employment of a network theory approach has been proposed for interpreting the large amount of ivF information collected to date in different mammalian models in order to identify the controllers of the in vitro system. The WoS-derived data generated a scale-free network, easily navigable including 641 nodes and 2089 links. A limited number of controllers (7.2%) are responsible for network robustness by preserving it against random damage. The network nodes were stratified in a coherent biological manner on three layers: the input was composed of systemic hormones and somatic-oocyte paracrine factors; the intermediate one recognized mainly key signaling molecules such as PI3K, KL, JAK-STAT, SMAD4, and cAMP; and the output layer molecules were related to functional ivF endpoints such as the FSH receptor and steroidogenesis. Notably, the phenotypes of knock-out mice previously developed for hub.BN indirectly corroborate their biological relevance in early folliculogenesis. Finally, taking advantage of the STRING analysis approach, further controllers belonging to the metabolic axis backbone were identified, such as mTOR/FOXO, FOXO3/SIRT1, and VEGF, which have been poorly considered in ivF to date. Overall, this in silico study identifies new metabolic sensor molecules controlling ivF serving as a basis for designing innovative diagnostic and treatment methods to preserve female fertility.

Assessment of Sensitivity and Profitability of an Intravaginal Sensor for Remote Calving Prediction in Dairy Cattle.

One critical point of dairy farm management is calving and neonatal first care. Timely calving assistance is associated with the reduction of calf mortality and postpartum uterine disease, and with improved fertility in dairy cattle. This study aimed to evaluate the performance and profitability of an intravaginal sensor for the prediction of stage II of labor in dairy farms, thus allowing proper calving assistance. Seventy-three late-gestating Italian Holstein cows were submitted to the insertion of an intravaginal device, equipped with light and temperature sensors, connected with a Central Unit for the commutation of a radio-signal into a cell phone alert. The remote calving alarm correctly identified the beginning of the expulsive phase of labor in 86.3% of the monitored cows. The mean interval from alarm to complete expulsion of the fetus was 71.56 ± 52.98 min, with a greater range in cows with dystocia (p = 0.012). The sensor worked correctly in both cold and warm weather conditions, and during day- or night-time. The intravaginal probe was well tolerated, as any cow showed lesions to the vaginal mucosa after calving. Using sex-sorted semen in heifers and beef bull semen in cows at their last lactation, the economic estimation performed through PrecisionTree™ software led to an income improvement of 119 € and 123 €/monitored delivery in primiparous and pluriparous cows, respectively. Remote calving alarm devices are key components of "precision farming" management and proven to improve animal welfare, to reduce calf losses and to increase farm incomes.

Equine Chorionic Gonadotropin as an Effective FSH Replacement for In Vitro Ovine Follicle and Oocyte Development.

The use of assisted reproductive technologies (ART) still requires strategies through which to maximize individual fertility chances. In vitro folliculogenesis (ivF) may represent a valid option to convey the large source of immature oocytes in ART. Several efforts have been made to set up ivF cultural protocols in medium-sized mammals, starting with the identification of the most suitable gonadotropic stimulus. In this study, Equine Chorionic Gonadotropin (eCG) is proposed as an alternative to Follicle Stimulating Hormone (FSH) based on its long superovulation use, trans-species validation, long half-life, and low costs. The use of 3D ivF on single-ovine preantral (PA) follicles allowed us to compare the hormonal effects and to validate their influence under two different cultural conditions. The use of eCG helped to stimulate the in vitro growth of ovine PA follicles by maximizing its influence under FBS-free medium. Higher performance of follicular growth, antrum formation, steroidogenic activity and gap junction marker expression were recorded. In addition, eCG, promoted a positive effect on the germinal compartment, leading to a higher incidence of meiotic competent oocytes. These findings should help to widen the use of eCG to ivF as a valid and largely available hormonal support enabling a synchronized in vitro follicle and oocyte development.

Differential Expression Pattern of Retroviral Envelope Gene in the Equine Placenta.

Endogenous retroviruses (ERVs) are proviral phases of exogenous retroviruses, which have coevolved with vertebrate genomes for millions of years. The conservation of ERV genes throughout evolution suggests their beneficial effects on their hosts' survival. An example of such positive selection is demonstrated by the syncytin gene, which encodes a protein with affinity for various mammalian placentas that is involved in the formation of syncytiotrophoblasts. Although the horse has an epitheliochorial placenta, in which the fetal trophoblasts are simply apposed to the intact uterine epithelium, we have previously demonstrated that the equine ERV (EqERV) env RNA is unexpectedly expressed in placental tissue. In the present study, we investigated the mRNA expression pattern of the EqERV env gene in different parts of the equine placenta, to gain more insight into its putative role in the fetal-maternal relationship. To this end, we used reverse transcription-quantitative PCR (RT-qPCR) and in situ hybridization assays to analyze different target areas of the equine placenta. The retroviral env gene is expressed in the equine placenta, even though there is no syncytium or erosion of the uterine endometrium. The gene is also expressed in all the sampled areas, although with some quantitative differences. We suggest that these differences are attributable to variations in the density, height, and degree of morphological complexity of the chorionic villi forming the microcotyledons. The involvement of the EqERV env gene in different functional pathways affecting the fetus-mother relationship can be hypothesized.

Remote monitoring system as a tool for calving management in Mediterranean Buffalo heifers (Bubalus bubalis).

Buffalo breeding is common in Southern Italy. Dystocia compromises dam's and newborn health and welfare. Difficult parturition could be solved through prompt calving assistance, even if the identification of the beginning of delivery is challenging. Herein, we aimed to evaluate a remote calving alarm system in 15 Mediterranean buffalo heifers. An intravaginal probe was placed close to the external cervical os once premonitory signs of delivery were observed. No vaginal discharge nor signs of discomfort were notified in the days following the insertion of the probe. Heifers calved from 48 to 72 hr after the alarm was activated. The system correctly warned the farm personnel at the beginning of stage II of parturition, except for 2 cases. In the former, the intravaginal probe was expelled but the poor carrier network coverage negatively affected phone's signal quality; in the latter, recurrent vaginal prolapse was responsible for non-retention of the probe. Overall median expulsive phase was 68 ± 8 min, while the expulsion of a female calf took 54 ± 22.0 min and 90 ± 34.0 min in males, with significant difference (p =.02). Deliveries were homogeneously distributed across a 24-hr interval. No retention of foetal membranes nor metritis was identified at postpartum clinical examination. The calving alarm system used in this work was well tolerated in buffalo heifers. The introduction of smart technology in buffalo farming could contribute to the overall farm net return by reducing calf losses, especially for calves born from sexed-sorted semen, and by increasing animal welfare through quick resolution of dystocia. Further studies will be necessary to evaluate the net return in buffalo farms which will implement a remote calving alarm system on a wider population.

Estimating the net return of a remote calving alarm system in a dairy farm.

The aim of the study was to evaluate the net return of the implementation of a remote calving monitoring system for obstetrical and neonatal assistance on the herd economy in a dairy farm model. A total of 680 parturitions over a 7-yr period were evaluated. Age at first calving was restricted from 23 to 27 mo for primiparous cows to be included. Among groups of cows that were ready to calve in a 15-d interval, primiparous and multiparous were randomly assigned to the experimental group and monitored through a calving alarm system, whereas the others accounted for controls. Final parturition groups were as follows: control primiparous (CPP, n = 218), control multiparous (CM, n = 345), monitored primiparous (MPP, n = 56), and monitored multiparous (MM, n = 61). Monitored groups received prompt calving assistance and first neonatal care, whereas the presence of farm personnel was discontinuous for controls. A biological model was built considering significant differences in calf loss, early culling, milk production, and days open between groups. Then, a partial budget model was used to estimate costs and net return on a simulated herd of 100 lactating cows. Incidence of calf death was greater in control groups (11.06% and 10.73% in CPP and CM, respectively) compared with monitored cows (0.00% and 1.69% in MPP and MM, respectively). Multiparous cows with calf loss had increased relative risk (relative risk = 3.487) for early culling compared with multiparous counterparts with no neonatal loss. Daily milk production in the first 2 mo was 3.79 kg greater in multiparous cows with no dead calf, compared with their counterparts. A significant difference in median days open was found in MPP and CPP (118 and 148 d, respectively). In the final economic model, different simulations were analyzed. They were created assuming different prices or hypothesizing calving monitoring only in primiparous animals. The model estimated different, but always positive, net return. In conclusion, implementing a calving alarm system led to a net return from €37 to 90 per cow per year (€1 = US$1.15 at the time of the study). However, the device alone is not sufficient: it must be supported by qualified calving monitoring and assistance. Optimized personnel presence in the calving area at the right time leads to prompt calving and neonatal calf assistance and colostrum feeding within the first hours of life, thus reducing calf death and days open, and increasing milk production.

Systemic profiling of ectopic fat deposits in the reproductive tract of dairy cows.

During the transition period, high-yielding dairy cows suffer from negative energy balance, intense lipomobilization and impaired lipid metabolism; this metabolic condition can lead to overburdened triglycerides accumulation in the liver, known as liver lipidosis, which has been associated to impaired fertility in dairy cows. The mechanisms of this impairment can be in principle correlated with the presence and the extent of ectopic fat depots. However, current methods for evaluating fat accumulation in liver and in the reproductive tract suffer from low resolution, sensitivity, and specificity. Confocal microscopes are equipped with Gallium arsenide phosphide detectors, thus enabling the acquisition of intense signals from tissue biopsies. This method could differentiate whether fat deposition occurred without requiring sample sectioning. Here, we examined with this technique liver, uterine and ovarian samples of heifers and regularly slaughtered repeat breeder and overconditioned dairy cows, to quantify lipid droplets and depots at a submicrometer scale with high specificity. With the aid of this technique, we found lipid depots in uterine and ovarian specimens. Moreover, we found that the size and number of depots increased with the degree of liver lipidosis. Further studies are needed to elucidate the relationship between the severity and extent of these deposits and the fertility of lactating dairy cows. Since tissues other than liver display different characteristic lipid droplet distributions, this technique can be potentially employed to shed new light on the pathogenesis of lipidosis and to assess new risk factors for infertility.

Validation of the Italian version of the UNESP-Botucatu unidimensional composite pain scale for the assessment of postoperative pain in cattle.

OBJECTIVE: The aim of this study was to validate the Italian version of the UNESP-Botucatu Unidimensional Composite Pain Scale (UCPS-IV) for assessing postoperative pain in cattle. STUDY DESIGN: Video analysis and psychometric testing. ANIMALS: A total of 40 Nellore cattle, age 2-3 years, weighing 365±51 kg. METHODS: An English version of the scale was translated to Italian and back translated to English to ensure semantic equivalence. A total of four Italian observers and the researcher who developed the original scale (considered as the gold standard) analysed videos of 40 cattle previously subjected to orchiectomy (recorded for each cattle at four predetermined timepoints during the perioperative period) unaware of the videos' timepoint. They assigned a pain score [ranging from no pain (0) to severe pain (10)] using the Italian version of the scale and verified the need for analgesic treatment for each animal. The obtained scores were subjected to psychometric validity, responsiveness and reliability tests. RESULTS: The factor analysis of the scale resulted in a one-factor solution. Significant changes in pain scores in response to surgery and analgesics confirmed not only the content and construct validity, but also responsiveness. The agreement between the researcher who developed the original scale and the blinded observers and the correlation between the pain scores determined by the scale and those determined by three classical unidimensional pain scales confirmed criterion and concurrent validity. Internal consistency of the scale was excellent. Inter- and intrarater reliability ranged from moderate to good and from moderate to very good for all scale items, respectively, supporting reproducibility and stability. The cut-off point for rescue analgesia calculated by the receiver operating characteristic curve was > 3. CONCLUSIONS AND CLINICAL RELEVANCE: The results confirm that the UNESP-Botucatu UCPS-IV appears to be a valid and reliable tool for the assessment of postorchiectomy pain in the bovine species. The determination of a cut-off point for rescue analgesia is also helpful for guiding analgesic therapy.

Influence of lipoproteins at dry-off on metabolism of dairy cows during transition period and on postpartum reproductive outcomes.

High-yielding dairy cows are metabolically challenged during transition, when intense mobilization and hepatic oxidation of lipids is achieved, thus leading to fatty infiltration, ketosis and generalized inflammation. The condition is associated to periparturient diseases and poor fertility. The aim of this study was to assess whether serum lipoprotein concentrations in the dry period could influence the occurrence of postpartum diseases and reproductive performance in dairy cows. The study was carried out on 30 multiparous Holstein Friesian cows. Blood samples were collected at dry-off (-60 days), 30 days after dry-off and within 12 h after parturition for biochemical and serum lipoprotein assays. From 10 to 60 days after parturition milk was collected twice weekly after feeding, for milk whey progesterone assay. The Optimal Cutpoint package identified a threshold of 89% for serum High Density Lipoprotein (HDL) concentration at the beginning of the dry period with 95% of confidence interval. Cows with serum HDL greater than 89% (High group, n = 10) showed better reproductive performance when compared to those with low values (Low group, n = 18). The odds ratio for reproductive disorders in High group was 0.6875, however, differences were not significant probably due to both the reduced number of animals per group and overall low incidence of postpartum reproductive disease. First postpartum luteal activity occurred around day 23, while the second one between days 40 and 48. The average calving to first AI interval was 64.00 ± 3.95 days and 94.50 ± 12.32 days in High and Low group, respectively (P < 0.05). The calving-conception interval was 129.86 ± 24.42 days and 199.18 ± 24.73 days in High and Low groups, respectively (P < 0.05). Low group displayed an increase in liver markers, that is total bilirubin, with 0.46 ± 0.09 mg/dL and 0.23 ± 0.08 mg/dL, in Low and High group respectively (P < 0.05), and NEFA/cholesterol ratio, with 0.30 ± 0.06 and 0.14 ± 0.03, in Low and High groups, respectively (P < 0.05), at parturition. Concentrations of HDL >89% at dry-off could be suggestive of improved liver adaptation to the transition, and probably of enhanced fertility in High group.