Identification of Apoptotic Bodies in Equine Semen

Apoptosis in the testis is required to ensure an efficient spermatogenesis. However, sometimes, defective germ cells that are marked for elimination during this process escape elimination in the testes, giving rise to ejaculates with increased percentages of abnormal and apoptotic spermatozoa and a high percentage of apoptotic bodies. Apoptosis markers in the ejaculate have been associated with low fertility, either in animals or humans. Therefore, the goal of this study was to investigate whether fresh equine semen contains apoptotic bodies [initially named Merocyanine 540 (M540) bodies] and to study the relationship between the quantity of these bodies and cell concentration, the volume of ejaculate, viability and motility. Moreover, we also studied whether the presence apoptotic bodies in fresh semen was related to the resistance of the stallion spermatozoa to being incubated at 37°C or being frozen and thawed. Fresh equine semen was stained with fluorescent dyes such as M540 and Annexin‐V. Active Caspase 3 was studied in fresh semen through Western blotting and immunofluorescence with a specific antibody. Sperm kinematics was assessed in fresh, incubated and thawed samples using computer‐assisted semen analysis, and viability was evaluated with the LIVE/DEAD Sperm Viability Kit. Overall, our results demonstrate for the first time the presence of apoptotic bodies in equine semen. The quantity of apoptotic bodies was highly variable among stallions and was positively correlated with Caspase 3 activity in fresh samples and negatively correlated with the viability and motility of stallion spermatozoa after the cryopreservation process.     

Colloid Centrifugation Selects Normal Spermatozoa from Polymorphic Bull Ejaculates: A Case Study

Semen from a Western Finncattle bull exhibiting a highly polymorphic spermiogram was processed by colloid centrifugation using Androcoll‐B, a species‐specific silane‐coated silica colloid. In the first experiment, Single Layer Centrifugation (SLC) was used to identify which density colloids were needed to separate different cell populations. Colloids of the two chosen densities were then used in a density gradient resulting in two sperm subpopulations, one containing nearly all normally sized spermatozoa and the other enriched for the macrocephalic spermatozoa. Microcephalic spermatozoa did not appear in either of the selected subpopulations. Using a combination of SLC and DGC with this species‐specific colloid, it was possible to separate the spermatozoa into different subpopulations, that is, a subpopulation containing nearly all normally sized spermatozoa, and another one enriched for the macrocephalic spermatozoa. Thus, colloid centrifugation could be used to select sufficient normal spermatozoa from a highly polymorphic ejaculate for AI, if desired.     

Anti‐Mullerian Hormone Concentration and Antral Ovarian Follicle Population in Murrah Heifers Compared to Holstein and Gyr Kept Under the Same Management

This study was performed to evaluate plasma concentrations of anti‐Mullerian hormone (AMH) and the ovarian antral follicle population (AFP) in different genetic groups. Cyclic heifers (13 Bubalus bubalis [Murrah]; 15 Bos taurus [Holstein] and 10 Bos indicus [Gyr]) were maintained under the same management and were synchronized with two doses of 150 μg IM d‐cloprostenol administered 14 days apart. After the second d‐cloprostenol treatment, heifers had their ovaries scanned daily by ultrasound to define the day of ovulation. On the same day, the AFP was determined and a plasma sample was collected to measure AMH. Murrah heifers had less AFP (25.6 ± 2.1 follicles; p = 0.01) and plasma AMH concentration (0.18 ± 0.03 ng/ml; p < 0.001) than Gyr (60.0 ± 12.2 follicles and 0.60 ± 0.12 ng/ml of AMH); however, data were similar when compared to Holstein (35.9 ± 6.8 follicles and 0.24 ± 0.06 ng/ml of AMH) heifers. Regardless of genetic background, there was a positive relationship between the AFP and plasmatic AMH concentration (Murrah [r = 0.62; p < 0.01], Holstein [r = 0.66; p < 0.001] and Gyr [r = 0.88; p < 0.001]). Also, when heifers were classified according to high‐ or low‐AMH concentration based on the average within each genetic group, high‐AMH heifers had greater (p < 0.0001) AFP than low‐AMH heifers. In conclusion, both Murrah and Holstein heifers presented lower plasma AMH concentration and AFP when compared to Gyr.     

Changes in Bull Sperm Kinematics after Single Layer Centrifugation

The objective of this study was to evaluate bull sperm kinematics after centrifugation through a single layer of a colloid [Single Layer Centrifugation (SLC)]. Ejaculates from 20 bulls were extended and stored at 4–6°C for 24 h during transport to the laboratory for SLC through Androcoll‐B, followed by measurement of sperm kinematics in all samples. Total motility (86% and 88% for uncentrifuged and SLC samples, respectively) and progressive motility (84% for both the groups) were similar (p > 0.05). In contrast, straightness (STR) (0.65 vs 0.69), linearity (LIN) (0.32 vs 0.35) and beat cross frequency (BCF) (22.3 vs 23.6 Hz) were significantly higher in the SLC‐selected samples than in the uncentrifuged samples, whereas velocity of the average path (VAP) (95 vs 90 μm/s), curvilinear velocity (VCL) (192 vs 180 μm/s), amplitude of lateral head deviation (ALH) (7 μm vs 6.5 μm) and hypermotility (49% vs 38%) were significantly decreased. The kinematics of the samples with the poorest motility was improved most by SLC. In conclusion, even though SLC had no direct effect on total and progressive motility, it appeared to have a positive influence on several other kinematic parameters that may be important for fertilization after artificial insemination.     

Resveratrol treatment controls microbial flora, prolongs shelf life, and preserves nutritional quality of fruit

Resveratrol is known as a grapevine secondary metabolite with fungicide activity. Its exogenous application on harvested grapes resulted in the reduction of microbial flora growth, and consequently, prolonged shelf life, without affecting the nutritional quality of the fruit. Resveratrol treatment also resulted in being effective on fruit that normally does not accumulate such metabolites as, for example, tomatoes, apples, avocado pears, and peppers. As a result, all treated fruits maintained their post-harvest quality and health longer than the untreated ones. This study demonstrates the potential use of resveratrol as a natural pesticide to reduce post-harvest fungi development on a broad spectrum of fruit types.     

Fertilisation and weed control effects on yield and weeds in durum wheat grown under rain-fed conditions in a Mediterranean climate

Summary A field study was undertaken to examine the effect of fertiliser type and weed control system on grain yield, yield components and weed population in durum wheat ( Triticum turgidum. var. durum) grown in a 4-year rotation [bare fallow–barley ( Hordeum vulgare )–vetch ( Vicia sativa )–wheat]. Fertilisation treatments were: no fertilisation, organic fertilisation with 2500 kg ha⁻¹ of compost (sheep manure and cereal straw) and chemical fertilisation with 100–60–60 kg ha⁻¹ of NPK. Weed-control treatments were: no control, herbicide, harrowing with long-flex spring tines and strip sowing with inter-row hoeing. Chemical fertilisation increased grain yield with respect to the other treatments, amongst which no significant differences were noted. Weed-control systems afforded no improvement in yield compared with controls; indeed, yields obtained using inter-row hoeing were lower. The weed population consisted of a large variety of species, of which the most important were Convolvulus arvensis and Polygonum aviculare . Application of weed-control systems reduced weed density. Herbicide was the most effective, reducing weed biomass by 80%, whilst harrowing and hoeing reduced weed biomass by 40% and 52% respectively. Inter-row hoeing does not appear to be a viable alternative to herbicides, when used as the sole weed control method in a non-diverse cropping system in Mediterranean climates. The lack of response to compost suggests a need for further long-term research.     

Plasma indicators of muscle damage in a model of nutritional myopathy in weaner sheep

SUMMARY Subclinical nutritional myopathy was induced in 5-month-old sheep by feeding them a diet low in vitamin E and selenium. Subsequently clinical myopathy was induced by dosing with protected polyunsaturated fatty acids. Plasma activities of creatine kinase (CK), pyruvate kinase, aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase and aldolase, enzymes of muscle origin, all remained above their reference ranges in clinically affected sheep, but fluctuated widely. Similar fluctuations occurred in subclinically affected animals, resulting in some activities being within the reference ranges and some above, at different times. Plasma malondialdehyde, an indicator of lipid peroxidation, proved of no diagnostic value. Terminal plasma CK activities were significantly correlated with microscopic damage in the vastus lateralis (VL), but not the vastus intermedius (VI) or the tensor fascia lata (TFL) muscles. AST was the most highly correlated with damage in VI and VL. In two clinically affected sheep successfully treated with an oral dose of α-tocopherol actetate all enzymes decreased steadily to within their reference ranges, at rates probably related to their plasma half-lives. These results suggest that measurement of plasma CK activity would be useful in monitoring recovery of treated sheep.     

Further investigation of lead exposure as a potential threatening process for a scavenging marsupial species

There is a growing recognition of the harmful effects of lead exposure on avian and mammalian scavengers. This can lead to both lethal and non-lethal effects which may negatively impact wildlife populations. Our objective was to assess medium-term lead exposure in wild Tasmanian devils (Sarcophilus harrisii). Frozen liver samples (n = 41), opportunistically collected in 2017–2022, were analysed using inductively coupled plasma mass spectrometry (ICP-MS) to determine liver lead concentrations. These results were then used to calculate the proportion of animals with elevated lead levels (>5 mg/kg dry weight) and examine the role of explanatory variables that may have influenced the results. The majority of samples analysed were from the south-east corner of Tasmania, within 50 km of Hobart. No Tasmanian devil samples were found to have elevated lead levels. The median liver lead concentration was 0.17 mg/kg (range 0.05–1.32 mg/kg). Female devils were found to have significantly higher liver lead concentrations than males (P = 0.013), which was likely related to lactation, but other variables (age, location, body mass) were not significant. These results suggest that wild Tasmanian devil populations currently show minimal medium-term evidence of exposure to lead pollution, although samples were concentrated in peri-urban areas. The results provide a baseline level which can be used to assess the impact of any future changes in lead use in Tasmania. Furthermore, these data can be used as a comparison for lead exposure studies in other mammalian scavengers, including other carnivorous marsupial species.