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Simple Summary: The placenta serves as the vital interface between maternal and fetal circulation, playing a pivotal role in ensuring the fetus's nutrition and oxygenation. It is, in fact, a highly sophisticated and multifaceted organ, capable of integrating signals from both the mother and the fetus with remarkable efficiency. PAG levels are indicative of the number of fetal cotyledons in the placenta and, consequently, the size of the placenta. This provides insight into placental development and fetal–placental well-being. However, to the best of our knowledge, the effect of maternal undernutrition on PAG secretion has not been extensively investigated in ruminant species. Starting from this premise, this study aimed to determine whether undernutrition might exert an influence on the production of this glycoprotein, with subsequent implications for embryonic and fetal well-being. To the best of our knowledge, no studies in the literature have reported on the relationship between the pregnancy-associated glycoprotein (PAG) concentration and undernutrition during pregnancy in sheep. Therefore, the aim of this study was to investigate the effects of undernutrition on the blood concentration of PAGs in pregnant ewes carrying single and multiple fetuses, undergoing either dietary energy restriction or receiving 100% of their energy requirements during the period of maximal placenta growth. From d 24 to 100 of pregnancy, the ewes were fed ryegrass hay and two different iso-proteic concentrates, fulfilling either 100% of the ewes' energy requirements (control group; n = 30, 14 singleton pregnancies, 16 multiple pregnancies) or only 50% (feed-restricted group; n = 29; 11 singleton pregnancies, 18 multiple pregnancies). Blood samples were collected from all the ewes the day before intravaginal sponge insertion (T0) and, thereafter, every 30 days starting from d 24 of gestation (T1) until d 30 after parturition (T6), to determine the PAG, progesterone, NEFA, total protein, and glucose concentrations. Our results showed that PAG concentrations are not only influenced by the number of fetuses, but also by the ewe's diet. In particular, the PAG marginal means were higher in the feed-restricted group than in the control group (7.8 ± 0.6 and 10.8 ± 0.9 ng/mL for the control and feed-restricted groups, respectively; p = 0.040). This finding confirms that a dietary restriction between d 24 and d 100 of gestation leads to a growth in placental functions as inferred from the increase in the concentration of the PAG levels. Therefore, the measurement of PAG concentrations, in addition to serving as a marker of pregnancy, can also serve as a marker of placental functionality. [ABSTRACT FROM AUTHOR]

Introduction: Although intensive care units (ICUs) are where severe and complicated cases are managed, there is limited evidence on treatment outcomes in Ethiopia. Therefore, this study is aimed at assessing the magnitude and associated factors of mortality among patients with sepsis admitted to ICUs at southern Amhara public hospitals, Ethiopia. Methods: A total of 547 medical records of patients with sepsis admitted to the ICUs at Injibara, Debre Markos, and Debre Tabor hospitals in the past 3 years were retrieved from August 10–31, 2022. Multivariable logistic regression analyses were conducted and adjusted odds ratios (AOR) with a 95% confidence interval (CI) were reported, and a p value < 0.05 was set to declare the significance of the association. Results: In this study, 46.2% (95% CI: 41.7%–50.3%) of patients with sepsis died during their ICU stay. Respiratory, gastrointestinal, and urinary tract infections contributed to 32.3%, 25.8%, and 9.6% of mortality, respectively. Individuals aged 60 and above (AOR: 4.07; 95% CI: 2.23–7.44), those with a Glasgow Coma Scale ≤ 10 at admission (AOR: 11.27; 95% CI: 4.64–27.37), respiratory site of infection (AOR: 5.38; 95% CI: 2.94–9.86), creatinine level > 1.1 mg/dL (AOR: 4.20; 95% CI: 2.33–7.60), vasopressor use (AOR: 3.13; 95% CI:1.66–5.95), initiation of antibiotics 1–3 h after admission (AOR: 2.80; 95% CI: 1.64–4.76), and a hospital stay of more than 20 days (AOR: 3.44; 95% CI: 1.40–8.46) were significantly associated with mortality. Conclusion: Overall, the mortality rate among patients with sepsis admitted to ICUs in southern Amhara public hospitals was high. Mainly, death was attributed to respiratory infections. Elderly patients and those with deteriorated clinical conditions at admission were at higher risk. Therefore, special attention is needed for elderly patients admitted with respiratory infections, antibiotics should be initiated as early as possible, and interventions must be designed to shorten the length of stay in ICUs. [ABSTRACT FROM AUTHOR]

This review aims to analyse the fluctuations of fecal thyroid hormone metabolites (FTMs) related to environmental and individual variables in different species of wild ungulates and provide a collection of assay methods. The great advantage of fecal sampling is being completely non-invasive. A systemic search was conducted from 2019 to 2024, using data sources PubMed, Scopus, Web of Science, and the World Wide Web, and ten studies were found on this topic. Three studies used the radioimmunoassay method for FTMs analysis, while the others used a less expensive enzyme-linked immunosorbent assay. Most of these papers validated the method for the species-specific matrix. Related to the studied variables, some authors analysed FTM fluctuations only concerning individual variables, and others in response to both. Temperature and fecal cortisol metabolites (FCMs) were the most studied environmental and individual variables, respectively. Since FTMs are an integrative measure of plasma thyroid hormones, the information obtained from a non-invasive-assay method regarding wild ungulate physiology is becoming of great interest to the scientific community. [ABSTRACT FROM AUTHOR]

Simple Summary: Antibiotics are emerging contaminants found in several matrices of environmental, animal, and human origin. This high occurrence constitutes a major global threat, which is also responsible for the development of antimicrobial resistance. Therefore, it is vitally important to develop new, simple, and reliable methods by which to quantify them in several matrices. In this work, the mussel matrix was chosen as the sentinel organism; a new analytical method for the determination of two commonly used macrolides in mussels harvested along the Sardinian coasts was developed. None of the tested macrolides were found in the sampling sites at concentrations quantifiable by the developed method. These results can be useful for competent authorities evaluating the food safety of mussels (as concerns the studied antibiotics) and pollution derived from these drugs in the marine environment. Contaminants of emerging concern (CECs) are compounds found in several environmental compartments whose ubiquitous presence can cause toxicity for the entire ecosystem. Several personal care products, including antibiotics, have entered this group of compounds, constituting a major global threat. It is essential to develop simple and reliable methods by which to quantify these contaminants in several matrices. In this work, mussels were chosen as sentinel organisms to assess environmental pollution and the safety of bivalve mollusk consumption according to the "One Health perspective". A liquid chromatographic tandem mass spectrometry method (LC-MS/MS) was developed for the quantification of two macrolides, erythromycin (ERY) and azithromycin (AZI), in mussels. This new method was validated according to international guidelines, showing high selectivity, good recoveries (>60% for both of them), sensitivity, and precision. The method was successfully applied for ERY and AZI research in mussels farmed along the Sardinian coasts (Italy), demonstrating itself to be useful for routine analysis by competent authorities. The tested macrolides were not determined in the analyzed sites at concentrations above the limits of detection (LODs). These results demonstrate the food safety of mussels (as concerns the studied antibiotics) and a negligible amount of pollution derived from these drugs in the studied area. [ABSTRACT FROM AUTHOR]

Sunscreens are topical preparations containing one or more compounds that protect humans from ultraviolet (UV) light. This review focuses on analytical methods, particularly liquid chromatography, with the aim of identifying and determining UV filters (UVFs) in environmental and marine biota matrices. A literature review was conducted using NIH (PubMed and Medline), FDA and EPA databases, Google Scholar, and federal regulations. This retrospective literature review is focused on the last five years. UVF quantification in environmental and biological matrices showed a wide array of methods where liquid chromatography is predominant. The scientific literature identified a large variety of analytical methodologies that are compared in this review to evaluate the better results in terms of limits of quantification and the possibility to identify as many analytes as possible simultaneously. [ABSTRACT FROM AUTHOR]

Simple Summary: The beneficial effect of polyunsaturated fatty acids (PUFA) in different body systems and their function in mammals is well known. The use of linseed oil (LO, rich in alpha-linolenic acid/ALA) as a feed additive has been investigated to improve the reproductive and productive performance of different livestock species. However, their effects on the antioxidative defense systems of female reproductive structures are scarcely documented. The results obtained in the present study demonstrate that dietary supplementation of by-pass LO enhances the antioxidative system in luteal and uterine tissues on maternal recognition of the pregnancy period (on Days 14 and 16 after mating) in ewes. This study analyzed the effects of dietary supplementation with by-pass linseed oil (LO; rich in α-linolenic acid) on maternal antioxidant systems at Days 14 and 16 of pregnancy in Sarda ewes. This trial used sixteen dry ewes. Eight ewes (CT group) were fed with a control diet without LO, and eight ewes (LO group) were fed with a diet supplemented with LO (10.8 g of α-linolenic acid/ewe/day). Both diets had similar crude protein and energy levels. The experiment included 10 days of an adaptation period and 31 days of a supplementation period. This supplementation period was divided into Period −2 (from Day −15 to −8), Period −1 (from Day −7 to −1; before synchronized mating period/Day 0), Period +1 (from Day +1 to + 7 after mating), and Period +2 (from Day +8 to +15 after mating). Estrous synchronization was induced in all the ewes using an intravaginal sponge (45 mg fluorgestone acetate) for 14 days and equine chorionic gonadotropin (350 UI/ewe) at the end of the treatment. On Days 14 (CT, N = 4; LO, N = 4) and 16 (CT, N = 4; LO, N = 4) after mating, the ewes were slaughtered. Samples of plasma, uterine, and luteal tissues were collected. Thiols, total antioxidant activity (TEAC), superoxide dismutase (SOD) activity, and malondialdehyde (MDA) content were measured. On Day 16, thiol and TEAC in luteal tissues were higher in the LO group when compared with the control one (p < 0.05). Moreover, TEAC was higher for the LO group in uterine tissues on Days 14 and 16 (p < 0.05). SOD activity was higher in the LO group in luteal and uterine tissues on Day 14 and Day 16, respectively (p < 0.001). On Day 16, uterine MDA content was lower for the LO group (p < 0.001). No differences were found between groups at the plasmatic level. However, the by-pass LO supplementation enhanced the analyzed antioxidant parameters in luteal and uterine tissues. In conclusion, these results demonstrate that by-pass LO supplementation exerted a positive effect on antioxidative defenses on maternal structures during the embryo-maternal recognition period in ewes. Thus, this could contribute to improving the maternal environment during the embryo-maternal recognition period in mammals. [ABSTRACT FROM AUTHOR]

Simple Summary: The oxidative stress in the cell is the result of an imbalance between oxidants and the antioxidant defense system. In particular, oxidative stress in erythrocytes and spermatozoa of small domestic ruminants can negatively affect their productivity and welfare. This review highlights the assay methods for the quantification of ROS and enzymatic antioxidant activity in these cells, in order to help the researchers to evaluate the intracellular oxidative status and antioxidant defenses. These data can be useful in monitoring the health and reproduction of small domestic ruminants. The present review aims to provide an overview of the assay methods for the quantification of ROS and principal enzymatic antioxidants as biomarkers of oxidative stress in erythrocytes and spermatozoa of small domestic ruminants. A complete literature search was carried out in PubMed, Scopus and the World Wide Web using relevant keywords and focusing on the last five years (2018–2023). Among spectrophotometry, fluorometry and chemiluminescence, the most widely used method for ROS assay is fluorometry, probably because it allows to simultaneously assay several ROS, using different probes, with greater economic advantages. Regarding intracellular antioxidant enzymes, recent literature reports only spectrophotometric methods, many of which use commercial kits. The use of a less sensitive but cheapest method is suitable because both erythrocytes and spermatozoa samples are highly concentrated in domestic ruminant species. All methods considered in this review have been found to be appropriate; in general, the differences are related to their costs and sensitivity. Quantification of ROS and enzymatic antioxidant activity in erythrocytes and spermatozoa may find application in the study of the welfare and health status of small domestic ruminants for monitoring livestock production. [ABSTRACT FROM AUTHOR]

Simple Summary: Polyunsaturated fatty acids (PUFAs) are well known for their beneficial role in different body systems and their function in mammals. The use of PUFA-ω3 as a feed additive has been investigated with the aim of improving the reproductive and productive performance of different livestock species, but their effects on dairy sheep are scarcely documented. The present study demonstrated that in milking ewes, dietary supplementation of bypass linseed oil (LO; rich in alpha-linolenic acid/ALA) modified the lipidic metabolic status, increased milk PUFA composition, and the size of luteal tissue at the onset of maternal recognition of pregnancy (on day 11 after insemination). The present study showed that the use of a bypass LO supplementation during the periconceptional period did not increase the reproductive indexes in ewes. Nevertheless, dietary bypass LO modified the circulating concentrations of lipid metabolites and the milk fatty acid profile, demonstrating its absorption and utilization by body tissues. The aim of the present study was to assess whether the strategic supplementation of bypass LO can enhance reproductive indexes—fertility, lambing rate, and prolificacy—in dairy Sarda ewes at the end of lactation. To assess whether LO supplementation leads to the adsorptions of PUFAs and their subsequent utilization by the body tissues, milk composition and fatty acid content were analyzed. Forty-eight ewes were assigned to the following groups: the control group (CT; N = 24), fed with a control diet without LO; and the treatment group (LO; N = 24), fed with a diet supplemented with LO (10.8 g/ewe/day). Both diets had similar crude protein and energy levels and were offered for 38 days (−21 to +17 days after artificial insemination). The trial included an adaptation period (7 days) followed by a regular supplementation (31 days) period. Estrus synchronization was induced in all the ewes using an intravaginal sponge and equine chorionic gonadotropin. Fifty-five hours after pessaries withdrawal, all ewes were inseminated using the cervical route and fresh semen. Cholesterol (p < 0.01), high-density lipoprotein (p < 0.001), and triglyceride (p < 0.05) levels in plasma were higher in the LO group. Plasmatic levels of non-esterified fatty acids were lower in the LO group after the end of the supplementation period (p < 0.05). Milk unsaturated fatty acids (UFAs), monounsaturated fatty acids (MUFAs), total polyunsaturated fatty acids (PUFAs), PUFAs omega 3 (PUFAs-ω3) and 6 (PUFAs-ω6), and trans fatty acids were higher in the LO group (p < 0.001), while saturated fatty acids (SFAs) were higher in the CT group during the supplementation period (p < 0.001). Three days after the end of the supplementation period, the content of milk UFAs (p < 0.05), PUFAs (p < 0.001), MUFAs, and PUFAs-ω6 (p < 0.01) were still higher in the LO group. whereas SFA was higher in the CT group (p < 0.01). There was no difference between groups in terms of ovulation rate, progesterone levels in plasma, fertility rate, prolificacy, and total reproductive wastage. However, the total area of luteal tissue was higher in the LO group (p < 0.01). Results obtained demonstrated that LO supplementation exerts a positive role in corpus luteum size at the onset of the peri-implantation period in Sarda dairy ewes. Additionally, the results obtained in the present study showed that the use of dietary bypass LO affects lipid metabolites in plasma and milk fatty acid profiles, demonstrating the ALA uptake by body tissues. [ABSTRACT FROM AUTHOR]

The use of bisphenol S (BPS) as a substitute of Bisphenol A is increasing in several products and it can be found in different environmental and biological matrices. Its toxicity has been studied at different levels and one of BPS toxic mechanisms at high concentrations seems to be the induction of oxidative stress through the generation of reactive oxygen species (ROS). This study evaluates the ability of a curcuma and ginger (CG) mixture to exert an antioxidant effect on rat hepatocytes treated with BPS. The effects of the mixture were compared to those of a well-known antioxidant (Trolox). Three different BPS concentrations were used in order to verify ROS production. 70 µg/mL and 150 µg/mL of BPS generated a significant ROS increase (p < 0.01) as compared to control, while CG mixture was able to decrease this ROS production in hepatic cells, as compared to cells treated with 70 µg/ml of BPS (p < 0.01) restoring control levels. BPS 70 µg/mL was tested for total antioxidant capacity (TEAC), superoxide dismutase (SOD) and total thiols. TEAC and SOD significant decreased (p < 0.05 and p < 0.01, respectively) as compared to controls and CG mixture was able to restore control values. Given the widespread BPS use, results obtained in this study can be of high impact for the community, demonstrating the ability of a mixture of natural products to prevent BPS-induced oxidative stress. [ABSTRACT FROM AUTHOR]

Determination of serum or plasma progesterone (P4) concentrations is important to recognize pregnant and non-pregnant ewes, and also to predict the number of carried lambs. The 2 most common methodologies for the detection of plasma P4 are radioimmunoassay (RIA) and enzyme immunoassay (EIA). RIA is very expensive, and not all laboratories are equipped to perform this test; EIA is commercially available for human use, but only a few companies produce species-specific kits, which are expensive. We verified for ovine plasma a less expensive and easily available ELISA kit (DiaMetra) designed to quantify P4 in humans. Pools of ovine and human plasma were used to compare repeatability, accuracy, sensitivity, and stability of P4 measured by the DiaMetra kit. Repeatability data were within 15%, and accuracy values were ~90% for both plasma matrices. Stability data showed a loss of <20% for freeze–thaw and <30% for 30-d storage. All parameters were acceptable under international guidelines for method validation. The human ELISA kit was used successfully to quantify plasma P4 in 26 ewes during pregnancy until delivery. P4 concentrations were also correlated with the number of carried lambs. [ABSTRACT FROM AUTHOR]

Abstract Currently, the assessment of sperm function in a raw or processed semen sample is not able to reliably predict sperm ability to withstand freezing and thawing procedures and in vivo fertility and/or assisted reproductive biotechnologies (ART) outcome. The aim of the present study was to investigate which parameters among a battery of analyses could predict subsequent spermatozoa in vitro fertilization ability and hence blastocyst output in a goat model. Ejaculates were obtained by artificial vagina from 3 adult goats (Capra hircus) aged 2 years (A, B and C). In order to assess the predictive value of viability, computer assisted sperm analyzer (CASA) motility parameters and ATP intracellular concentration before and after thawing and of DNA integrity after thawing on subsequent embryo output after an in vitro fertility test, a logistic regression analysis was used. Individual differences in semen parameters were evident for semen viability after thawing and DNA integrity. Results of IVF test showed that spermatozoa collected from A and B lead to higher cleavage rates (0 < 0.01) and blastocysts output (p < 0.05) compared with C. Logistic regression analysis model explained a deviance of 72% (p < 0.0001), directly related with the mean percentage of rapid spermatozoa in fresh semen (p < 0.01), semen viability after thawing (p < 0.01), and with two of the three comet parameters considered, i.e tail DNA percentage and comet length (p < 0.0001). DNA integrity alone had a high predictive value on IVF outcome with frozen/thawed semen (deviance explained: 57%). The model proposed here represents one of the many possible ways to explain differences found in embryo output following IVF with different semen donors and may represent a useful tool to select the most suitable donors for semen cryopreservation.

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

Microcystins (MCs) are hepatotoxins harmful for animal and human health. The most toxic type between them is MC-LR whose presence has been investigated in different reservoirs all around the world. In this work microcystins were monitored in spring and summer in water and mussels (Mytilus galloprovincialis) of two Sardinia lagoons: Cabras and Calich lagoons. A Solid Phase Extraction method was developed to clean and concentrate samples before the Enzyme Linked Immunosorbent Assay (ELISA) and the following Mass Spectrometry detection. MCs presence was detected using the screening ELISA test in both lagoons. MCs peak was revealed in July for water and mussels belonging to Cabras lagoon (0.75±0.07 ng/L in water and 0.12±0.04 ng/g ww in mussels). In water of Calich lagoon there was a constant trend in the concentration of MCs during the considered months, while there was a MCs peak in July (0.6±0.5 ng/g ww) in mussels. The following LC-MS/MS analysis did not reveal MC-LR presence in all analyzed samples. These results can be useful to enrich knowledge on public health and consumer’s safeguard. [ABSTRACT FROM AUTHOR]

Background: REAC technology (acronym for Radio Electric Asymmetric Conveyor) is a technology platform for neuro and bio modulation. It has already proven to optimize the ions fluxes at the molecular level and the molecular mechanisms driving cellular asymmetry and polarization. Methods: This study was designed to verify whether this technology could extend spermatozoa life-span during liquid storage, while preserving their functions, DNA integrity and oxidative status. At 0, 24, 48, and 72 h. of storage at 4 °C, a battery of analyses was performed to assess spermatozoa viability, motility parameters, acrosome status, and DNA integrity during REAC treatment. Spermatozoa oxidative status was assessed by determining lipid peroxidation, the activity of superoxide dismutase (SOD), and the total antioxidant capacity. Results: During liquid storage REAC treated spermatozoa, while not showing an increased viability nor motility compared to untreated ones, had a higher acrosome (p > 0.001) and DNA integrity (p > 0.01). Moreover, the analysis of the oxidative status indicated that the mean activity of the intracellular superoxide dismutase (SOD) was significantly higher in REAC treated spermatozoa compared to untreated controls (p < 0.05), while the intracellular concentration of malondialdehyde (MDA), an end product of lipid peroxidation, at the end of the REAC treatment was higher in untreated controls (p > 0.05). The REAC efficacy on spermatozoa oxidative status was also evidenced by the higher trolox equivalent antioxidant capacity (TEAC) found in both the cellular extract (p < 0.05) and the storage media of REAC treated spermatozoa compared to untreated controls (p < 0.0001). Conclusion: The present study demonstrated that REAC treatment during liquid storage preserves spermatozoa acrosome membrane and DNA integrity, likely due to the enhancement of sperm antioxidant defenses. These results open new perspective about the extending of spermatozoa functions in vitro and the clinical management of male infertility. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate the potential protective effect of Lycium barbarum polysaccharides (LBP) pretreatment against cadmium (Cd)-induced hepatotoxicity in rats. Wistar rats were divided into control group, LBP group (300 mg/kg orally, once a day, for 30 days), Cd group (CdCl 4 mg/kg i.p. once), and LBP + Cd group (LBP 300 mg/kg orally, once a day, for 30 days + CdCl 4 mg/kg i.p. 24 h after the last treatment). Cd liver injury was examined by morphological/histological changes, transaminases, total protein concentration, and oxidative stress evaluated by MDA, 3NT, GSH, SOD, and TEAC activities. Cd intoxication caused gross morphological changes with hyperemia of the parenchyma, increased volume, and disappearance of the anatomical limits of the lobes associated with an increase of ALT, GSH, and TEAC in plasma and a decrease of MDA, GSH, and TEAC in liver, SOD, and total proteins in plasma. LBP pretreatment caused a slight improvement in the histological architecture and in the 3NT amount together with a significant improvement of hematic parameters. On the basis of the obtained results, we can affirm that LBP pretreatment can ameliorate liver conditions, but further studies are needed to better evaluate the protective antioxidant effects of LBP against Cd-induced toxicity. [ABSTRACT FROM AUTHOR]

Amphetamine designer drugs are central nervous system stimulants that are widely disseminated in the illegal market. Generally, in forensic laboratories, immunoassay methods are the first line of screening for these types of drugs in a biological specimen (typically blood, urine or oral fluid). In this article, we describe the cross-reactivity profiles of 30 new amphetamine designer drugs, using the Neogen® [Amphetamine Specific and Methamphetamine/3,4-Methylenedioxymethamphetamine (MDMA) assays] drug tests. To assess the potential matrix influence on the response, each assay was tested on whole blood, urine and oral fluid. Concentrations of 10,000 ng/mL were not sufficient to produce a positive response for the majority of the analyzed amphetamines. This clearly demonstrates that, although these kits are extremely effective for the target drugs for which they are intended (amphetamine, methamphetamine and MDMA), they cannot be used to reliably identify the tested designer drugs in real cases, as these concentrations greatly exceed those expected to be found in forensic samples. [ABSTRACT FROM AUTHOR]

The presence of microcystin-LR (MC-LR) was monitored in Tapes decussatus harvested in two Sardinian ponds (Cabras and Tortolì, Italy) in spring and summer. After solid phase extraction, samples were analyzed using a screening enzyme-linked immunosorbent assay (ELISA) followed by a liquid chromatographic coupled to tandem mass spectrometer (LC–MS/MS) analysis. Results obtained through the ELISA test showed the presence of microcystins with a maximum concentration in August for Cabras pond (0.55 ng/g) and in September for Tortolì pond (0.85 ng/g). The LC–MS/MS analysis did not confirm the presence of MC-LR suggesting that results obtained with the ELISA technique could be due to the presence of other microcystins. According to the tolerable daily intake suggested by the World Health Organization, these results hint that clams harvested in these ponds are safe for human health. These data can contribute to enrich the knowledge about the healthiness of Sardinian ponds and of their products. [ABSTRACT FROM AUTHOR]

Our aim is to verify if oocyte developmental potential is related to the timing of meiotic progression and to mitochondrial distribution and activity using prepubertal and adult oocytes as models of low and high developmental capacity respectively. Prepubertal and adult oocytes were incorporated in an in vitro maturation system to determine meiotic and developmental competence and to assess at different time points kinetic of meiotic maturation, 2D protein electrophoresis patterns, ATP content and mitochondria distribution. Maturation and fertilization rates did not differ between prepubertal and adult oocytes (95.1% vs 96.7% and 66.73% vs 70.62% respectively for prepubertal and adult oocytes). Compared to adults, prepubertal oocytes showed higher parthenogenesis (17.38% vs 2.08% respectively in prepubertals and adults; P<0.01) and polispermy (14.30% vs 2.21% respectively in prepubertals and adults; P<0.01), lower cleavage rates (60.00% vs 67.08% respectively in prepubertals and adults; P<0.05) and blastocyst output (11.94% vs 34.% respectively in prepubertals and adults; P<0.01). Prepubertal oocytes reached MI stage 1 hr later than adults and this delay grows as the first meiotic division proceeds. Simultaneously, the protein pattern was altered since in prepubertal oocytes it fluctuates, dropping and rising to levels similar to adults only at 24 hrs. In prepubertal oocytes ATP rise is delayed and did not reach levels comparable to adult ones. CLSM observations revealed that at MII, in the majority of prepubertal oocytes, the active mitochondria are homogenously distributed, while in adults they are aggregated in big clusters. Our work demonstrates that mitochondria and their functional aggregation during maturation play an active role to provide energy in terms of ATP. The oocyte ATP content determines the timing of the meiotic cycle and the acquisition of developmental competence. Taken together our data suggest that oocytes with low developmental competence have a slowed down energetic metabolism which delays later development. [ABSTRACT FROM AUTHOR]

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

Simple Summary: Glycerol is widely used as a feed supplement in ruminant nutrition. However, its administration at moderate and high doses results in an increase in plasma osmolality and in an alteration of red blood cell (RBC) indices. The present study aimed at further elucidating the effect of glycerol on RBCs' functionality by evaluating the effect of different glycerol concentrations on RBC homeostasis under in vitro conditions. Obtained results showed that glycerol permeates the RBC membrane and leads to hemolysis when glycerol concentration exceeds 200 mg/dL. However, RBCs' antioxidant defenses appear to protect cell membranes without causing an increase of oxidative stress markers. Moreover, no alteration in RBCs' intracellular Ca2+ ion concentrations and metabolic activity were found. In conclusion, glycerol-based nutritional treatment should be designed in sheep to avoid exceeding 200 mg/dL glycerol circulating concentration to prevent RBC osmotic stress. The use of high doses of glycerol as a livestock feed supplement is followed by a rapid increase in plasma concentrations and consequently in plasma osmolality. Moreover, glycerol is a highly diffusible molecule that can readily permeate the red blood cell (RBC) membrane following a concentration gradient. A rise in glycerol plasma concentrations can thus alter RBC homeostasis. The present study aimed at investigating both glycerol osmotic effects on sheep RBCs and their oxidative response under in vitro conditions. Sheep blood samples were suspended in media supplemented with increasing glycerol concentrations (0, 25, 50, 100, 150, 200, 250, 300, 350, 400 mg/dL), which reflected those found in vivo in previous studies, and incubated at 37 °C for 4h. Thereafter, osmolality and hemolysis were determined in spent media, while cell extracts were used to assay intracellular concentration of glycerol, ATP, Ca2+ ions, oxidative stress markers and reactive oxygen species (ROS).The study confirmed that glycerol intracellular concentrations are directly related with its concentration in the incubation media, as well as hemolysis (p < 0.001) which increased significantly at glycerol concentrations higher form 200 mg/dL. ROS intracellular level increased at all glycerol concentration tested (p < 0.01) and total thiols decreased at the highest concentrations. However, RBCs proved to be able to cope by activating their antioxidant defense system. Superoxide dismutase activity indeed increased at the highest glycerol concentrations (p < 0.001), while total antioxidant capacity and malonyldialdehyde, a typical product of lipid peroxidation by ROS, did not show significant changes. Moreover, no alterations in intracellular Ca2+ ions and ATP concentrations were found. In conclusion, glycerol-induced hemolysis can be related to the induced osmotic stress. In sheep, nutritional treatments should be designed to avoid reaching glycerol circulating concentrations higher than 200 mg/dL. [ABSTRACT FROM AUTHOR]

Simple Summary: Reproductive management of sheep for autumnal lambing often require induction and synchronization of oestrus and ovulation, either for natural mating or artificial insemination, by the use of pharmacological treatments. Such treatments are mostly based on the administration of progesterone followed by a single intramuscular dose of equine chorionic gonadotrophin (eCG) at progesterone withdrawal. However, repeated eCG treatments in consecutive mating seasons can result in the outbreak of resistance with a rise of anti-eCG antibodies. Furthermore, the future use and availability of eCG appears to be strongly challenged by the highly active animal-rights movement because the hormone is obtained from pregnant mares. The present study demonstrated that the administration of glycerol-based formulations to milked ewes is a valid alternative to eCG treatment in reproductive management protocols based on the induction of ovulation with progesterone-releasing devices at the end of anoestrus period. The glucogenic treatment administration to late lactation dairy ewes at the end of the anoestrus period improved their metabolism without harming animal production or animal welfare, thus promoting a sustainable reproductive management of dairy sheep. This study investigated whether the administration of equine chorionic gonadotrophin (eCG) in a protocol to induce and synchronize ovulations before mating could be replaced by the administration of glycerol-based formulations in milked ewes at the end of their seasonal anoestrus. Forty-eight late-lactation dairy ewes of the Sarda breed were synchronized using sponges impregnated with progestogen and then joined with fertile rams (day (D) 0, ram introduction). From D−4 to D−1, the ewes received by gavage either 100 mL of a glucogenic mixture (70% glycerol, 20% propylene glycol and 10% water; GLU group; n = 24) or 100 mL of water (GON group; n = 24) twice daily. Moreover, on the day of sponge withdrawal (D−1), GON ewes received 200 IU of eCG. There were no differences in reproductive performances between groups. GLU ewes showed higher glycemia (p < 0.001), insulinemia (p < 0.05), plasma glycerol (p < 0.001), triglycerides (p < 0.001) and lower cholesterol (p < 0.001), non-esterified fatty acids (NEFA; p < 0.05) and urea (p < 0.001). Plasma osmolality was higher in GLU but only 4 h after dosing (p < 0.001). Milk yield and milk composition were not affected by the treatments with exception of milk glycerol (p < 0.001) and milk urea (p < 0.001), which were higher and lower in GLU than GON ewes, respectively. In conclusion, the administration of the glucogenic mixture to late lactation dairy ewes at the end of anoestrus period resulted in reproductive responses as good as the ones obtained by the eCG treatment, suggesting that the objective of a sustainable reproductive management of dairy sheep can be successfully pursued. [ABSTRACT FROM AUTHOR]

Simple Summary: The present study aimed to determine the blood concentrations of L-arginine, asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA), and L-homoarginine, modulating nitric oxide (NO) synthesis, in single, twin. and triplet pregnancies in ewes undergoing either dietary energy restriction or receiving 100% of their energy requirements. Blood concentrations of L-arginine, of its metabolites. and the ratio between NO synthesis boosters and inhibitors are altered in energy-restricted ewes, these alterations being higher in ewes carrying multiple fetuses. The aim of this study was to investigate the blood concentrations of L-arginine, asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA), and L-homoarginine, which are regulators of nitric oxide (NO) synthesis, in single, twin, and triplet pregnancies in ewes undergoing either a dietary energy restriction or receiving 100% of their energy requirements. From day 24 to 100 of pregnancy, the ewes were fed ryegrass hay and two different iso-proteic concentrates fulfilling either 100% of ewes' energy requirements (control group; n = 30, 14 singleton pregnancies, 12 twin pregnancies, and 4 triplet pregnancies) or only 45% (feed-restricted group; n = 29; 11 singleton pregnancies, 15 twin pregnancies, and 3 triplet pregnancies). Blood samples were collected monthly to measure, by capillary electrophoresis, the circulating concentrations of arginine, ADMA, homoarginine, SDMA, and of other amino acids not involved in NO synthesis to rule out possible direct effects of diet restriction on their concentrations. No differences between groups were observed in the circulating concentrations of most of the amino acids investigated. L-homoarginine increased markedly in both groups during pregnancy (p < 0.001). SDMA (p < 0.01), L-arginine, and ADMA concentrations were higher in feed-restricted ewes than in controls. The L-arginine/ADMA ratio, an indicator of NO production by NOS, decreased towards term without differences between groups. The ADMA/SDMA ratio, an index of the ADMA degrading enzyme activity, was higher in controls than in feed-restricted ewes (p < 0.001). Obtained results show that circulating concentrations of L-arginine, of its metabolites, and the ratio between NO synthesis boosters and inhibitors are altered in energy-restricted ewes, and that these alterations are more marked in ewes carrying multiple fetuses. [ABSTRACT FROM AUTHOR]

The chemical modification of the molecular structure of psychoactive substances is a very common practice in the illicit drugs market, to by-pass current regulations; this lead to the production of compounds, known as "designer drugs", with the same or greater pharmacological effects of the parent drug. The phenomenon is also favored by the fact that the new synthetic compounds are not considered illegal by existing legislation. Amphetamine derivatives represent one of the largest classes of designer drugs. Generally, in toxicological laboratories, rapid screening tests are used for a first monitoring of drugs abuse. However, the available immunoassays for this class of substances are designed for amphetamine, methamphetamine and methylenedioxymethamphetamine, and generally they are unable to detect various amphetamine analogues. This can constitute a disadvantage because it can generate a great number of false-negative results. The present review aims to provide an overview of the cross-reactivity studies carried out on commercially available immunoassays to identify the presence of amphetamine derivatives in biological samples. The knowledge of cross-reactivity data makes it easier to interpret analytical results by demonstrating that a negative result does not always indicate the non-consumption of an amphetamine derivative. This review highlights the great need for more comprehensive screening immunoassays to use when analyzing biological matrices for drugs of abuse search, specifically for the more recent designer drugs.. [Display omitted] • Cross-reactivity of commercial immunoassays for amphetamines were compared. • New amphetamine designer drugs gave negative result to screening tests. • More comprehensive screening immunoassays for recent designer drugs are needed. [ABSTRACT FROM AUTHOR]

The effects of the intra-ruminal dosing of a glucogenic mixture including 70% glycerol, 20% propylene glycol and 10% water was studied on thirty late lactation dairy ewes of Sarda breed. The animals were divided in two homogeneous groups receiving by gavage either 200 mL of water (CTR group; body weight 40.9 ± 1.5 kg) or 200 mL of the above mixture (GLY group; body weight 39.4 ± 1.3 kg) twice daily from d 16 to d 19 of the oestrus cycle, synchronised by “ram effect”. The ewes were then mated and their reproductive responses to the synchronised mating evaluated by scanning on d 50 and at lambing. During the treatment, the ewes were housed in an open hut, machine milked twice daily and fed concentrate and hay to meet their nutrient requirements. During the treatment, concentrate intake was markedly reduced in GLY when compared with CTR (P < 0.001), without any effect on ewe body weight or body condition. The administration of the glucogenic mixture increased plasma osmolarity and blood volume as estimated by serum total protein concentration. Moreover, it increased plasma content of glycerol, glucose (P < 0.001) and insulin (P < 0.01) while decreasing plasma level of NEFA (P < 0.001) and urea (P < 0.05). Milk yield (P < 0.01) and milk lactose content (P < 0.001) were decreased by the glucogenic treatment, whereas milk protein and casein contents were increased (P < 0.001). As for reproductive performance, the glucogenic treatment numerically increased ewe’s conception rate, but the difference was not statistically significant. Prolificacy did not change between groups. In conclusion, the administration of a glucogenic mixture to late lactation dairy ewes caused significant changes both in plasma and in milk composition during the treatment. However, reproductive performances were unaffected by the treatment. [ABSTRACT FROM AUTHOR]