The Distribution of Boars Spermatozoa in Morphometrically Distinct Subpopulations after In Vitro Exposure to Radiofrequency Electromagnetic Radiation at 2500 MHz and Their Motility.

Anthropogenic radiofrequency electromagnetic radiation (RF-EMR) from wireless technologies has increased dramatically. The boar semen used for artificial insemination is essential in sustaining the pig industry, and additionally it is also exposed to the effects of the RF-EMR of wireless technologies. Furthermore, there are no data on the effects of RF-EMR on semen quality, and this is the first analysis of sperm's morphometric parameters for assessing the effect of RF-EMR on the spermatozoa subpopulations of boars. This study investigated the effect of RF-EMR on in vitro exposed breeding boar semen spermatozoa motility and the proportions of spermatozoa subpopulations according to their morphometric head and tail parameters. The semen samples of 12 boars were divided into control and experimental groups. The samples in the experimental group were exposed in a gigahertz transverse electromagnetic chamber at a frequency of 2500 MHz (the frequency band used in 5G technology) and an electric field strength of 10 Vm-1 for two hours. After exposure, the spermatozoa motility was evaluated for both groups. A morphometric analysis of the semen smears was performed using SFORM software (Version 1.0; VAMS, Zagreb, Croatia). The progressive spermatozoa motility was significantly reduced in the experimental group (74.7% vs. 85.7%). PC analysis and cluster analysis revealed two spermatozoa subpopulations: S1, spermatozoa with a more regular head shape and a smaller midpiece outline, and S2, spermatozoa with a more elongated head shape and a larger midpiece outline. The experimental semen samples had a greater proportion of the S1 spermatozoa subpopulation (68.2% vs. 64.4%). The effect of RF-EMR at 2500 MHz on the in vitro exposed boar semen resulted in decreased progressive spermatozoa motility and a lower proportion of the spermatozoa subpopulation with a higher fertilizing potential.

A Survey Study of Veterinary Student Opinions and Knowledge about Pet Reptiles and Their Welfare.

Exotic pet medicine is rapidly evolving, with reptiles becoming increasingly popular pet animals. Yet, there are only a few literature reports on veterinary perception of reptiles kept as pets. The aim of the study was to assess opinions and knowledge of the Croatian veterinarians-to-be about pet reptiles and their welfare. The questionnaire survey was conducted in the academic year 2019-2020 and included students of all six years of the integrated undergraduate and graduate study at the Faculty of Veterinary Medicine, University of Zagreb. First-year students were surveyed twice, before and after having attended the compulsory course on animal welfare. Questionnaire statements were 5-point Likert scale questions, requiring the students to express their opinions about turtles, lizards and snakes as pets, issues related to their welfare, risks they pose to the health and safety of humans, other animals and the environment, and their self-reported knowledge about pet reptiles. Although expressing higher opinions after having attended the course on animal welfare, first-year student responses remained neutral to most of the statements. Such a trend continued until the end of the study. Student responses revealed that they were uncertain about their knowledge of reptiles as pets, considering different educational areas observed. Study results emphasised the need of alterations in veterinary curriculum and additional student education in reptile medicine. The results obtained have broad implications involving not only the welfare of pet reptiles in clinical practice and elsewhere but also the health and safety of humans and other animals, as well as environmental protection.

Effects of Dietary Vibroactivated Clinoptilolite Supplementation on the Intramammary Microbiological Findings in Dairy Cows.

: The aim of this study was to determine the effects of dietary vibroactivated clinoptilolite supplementation on the intramammary microbiological findings in dairy cows, causative agents of potential intramammary infection, and their sensitivity to antibiotics. Cows (n = 78) were randomly divided into two groups: CPL-treated group that received clinoptilolite (CPL) in-feed (n = 38), i.e., 50 g natural powdered zeolite CPL, twice daily from the seventh month of pregnancy to 75 days after calving, and the control group (CON) of untreated cows (n = 40). Milk samples were taken from each cow on days 7, 25, 45, and 75 postpartum. The following causative pathogens were isolated in 86 udder quarters: Staphylococcus aureus in 5.81% of positive samples, Staphylococcus spp. 9.32%, coagulase-negative Staphylococcus (CNS) 22.09%, Streptococcus uberis 13.95%, Streptococcus agalactiae 1.16%, Streptococcus sp. 3.49%, Escherichia coli 8.13%, Enterococcus spp. 6.98%, Corynebacterium spp. 11.63%, Pasteurella sp. 10.47%, Serratia spp. 2.33%, and Arcanobacterium pyogenes, Citrobacter sp., Prototheca sp., and yeasts each in 1.16% of samples. Additionally, 3.87% of environmental microflora samples (n = 47) and 89.06% of udder samples (n = 1083) were bacteriologically negative. The most effective antibiotics were cefoperazone and amoxicillin-clavulanic acid, while cloxacillin and tetracycline were the least effective antibiotics in both groups. In the clinoptilolite supplemented (CPL) group (n = 38) of 14 cows, nine causative agents of mastitis were isolated in 27 quarters, while in the control (CON) group (n = 40) of 24 cows, 13 causative agents of mastitis were isolated in 59 quarters. Cows from the CON group had a 1.96 times higher risk of intramammary infection than cows from the CPL group during the observation period (odds ratio = 1.96, p = 0.0031; 95% CI = 1.2570-3.0770).

Effects of in-feed clinoptilolite treatment on serum metabolic and antioxidative biomarkers and acute phase response in dairy cows during pregnancy and early lactation.

The objective of this study was to evaluate the effects of in-feed clinoptilolite (CPL) on serum metabolic and antioxidative biomarkers, acute phase proteins and reproductive performance in cows during pregnancy and lactation. A total of 78 Holstein-Friesian cows were randomly assigned into two groups: the treatment group, cows fed CPL (n = 38) which received 50 g of powdered CPL twice a day from day 180 before parturition to day 60 postpartum; and the control group (n = 40). Blood samples were taken on days 180, 90, 60, 30 and 10 before parturition, on day of calving and on days 5, 12, 19, 26, 33, 40 and 60 postpartum, and were analysed for metabolic biomarkers: glucose, triglycerides, total cholesterol, high density lipoprotein cholesterol, non-esterified fatty acids, beta-hydroxybutyrate (BHB), antioxidative biomarkers and acute phase proteins: paraoxonase-1 (PON1), apolipoprotein A-I, haptoglobin (Hp) and serum amyloid A (SAA). CPL supplementation increased concentration of glucose and significantly decreased (P < .05) level of BHB during puerperium. The SAA concentration in CPL-fed cows was significantly decreased (P < .05) on days 33, 40 and 60 postpartum as well as Hp concentration on days 0 and 12 postpartum. The results of this study suggest that the CPL-fed cows may have improved metabolic status due to the tendency of greater glucose levels and decreased BHB values during early lactation. In addition, acute phase response was lower (P < .05) in CPL-fed cows. Such an outcome might be attributed to the effect of dietary CPL on intensity and severity of the negative energy balance and inflammatory response in dairy cows.

Influence of season, rainfall and air temperature on the reproductive efficiency in Romanov sheep in Croatia.

The aim of this study was to investigate the influence of season, rainfall and air temperature on the reproductive efficiency in the Romanov breed of sheep in continental part of Croatia during five consecutive years (2012-2016). During this period, 5379 matings resulted in 5046 successful conceptions, i.e. lambings at eight medium-scale Romanov breed sheep farms. The conception rate was 93.81%, fecundity was 195% and average preweaning mortality until 90 days of age was 12.41%. The seasonal distribution of lambings was 47.64% for ewes that delivered in winter (n = 2422), 23.37% in spring (n = 1179), 18.82% in summer (n = 950) and 9.81% in autumn (n = 495). Sexual activity was lowest during spring and early summer when air temperatures were above average (very and extremely warm), and sexual activity peaked from August to September, especially during extremely wet and very wet seasons. Litter size was greater during winter than in other seasons (1.70 vs. 1.54) and was significantly different as compared to each of selected years of the study period. There was a statistically significant difference in the number of pregnant ewes between mating seasons. Most female Romanov lambs born during winter and early spring mated in late summer or autumn and delivered at the age of 1 year or earlier. The seasonal distribution of matings and lambing was not uniform throughout the seasons over five consecutive years. Thus, it can be assumed that air temperature and rainfall during different seasons could affect the reproductive efficiency in Romanov breed of sheep in continental part of Croatia.

Exogenous melatonin influences distribution of French Alpine buck spermatozoa in morphometrically distinct subpopulations during the non-breeding season.

The aim of this study was to establish subpopulations of spermatozoa in bucks using the principal component (PC) and cluster analysis according to morphometric head and tail variables, and to determine differences in proportions of subpopulations between exogenous melatonin-treated and control bucks. The bucks (n = 12) were assigned to two groups comprising six bucks each. By the end of March, four melatonin implants were inserted in the bucks in the experimental group. Semen was collected weekly using an artificial vagina from March to May (the non-breeding season). Analyses were performed in stained smears by SFORM computer-assisted program for eight head and five tail variables. The PC analysis revealed four components with the most important value for each (head outline, head ellipticity, mid-piece length and width). Cluster analysis indicated there were three subpopulations (average-sized spermatozoa- C_1; small and less-elliptic - C_2; big and elliptic - C_3). Melatonin-treated bucks had a greater proportion of C_1 spermatozoa and a lesser proportion of C_2 spermatozoa during May (P < 0.05). To the best of our knowledge, this is the first study where PC and cluster analyses were performed on buck semen with head and tail variables analyzed together, in the same analysis, to evaluate spermatozoa population. Also, this is the first analysis of morphometric variables for assessing the influence of melatonin on spermatozoa subpopulations. The positive effect of melatonin on the proportions of spermatozoa in subpopulations could have been a consequence of the decreasing the proportion of the subpopulation with the least head and tail sizes and ellipticity.

Antioxidant protection and lipid peroxidation in testes and different parts of epididymis in boars.

The antioxidative ability of testes and epididymis to protect spermatozoa from detrimental effects of oxidation processes induced by excessive generation of reactive oxygen species has not been previously studied in detail in boar reproductive tissues. The aim of this study was to investigate differences in level of antioxidative protection, intensity of lipid peroxidation, and values of biochemical parameters in testes and different parts of epididymis in sexually mature boars. The study was performed on five Swedish landrace boars from the same litter aged 10 months kept under same ambient conditions. After slaughtering performed at the end of November; tissue samples of testes and the head, body and tail of epididymis were taken. The activity of glutathione peroxidase, glutathione reductase, superoxide dismutase, γ-glutamyltransferase, acid phosphatase, and lactate dehydrogenase, and concentrations of triacylglycerol, phospholipids, cholesterol, free fatty acids were determined in obtained supernatants from homogenized tissues spectrophotometrically; the concentration of malondialdehyde was determined by high-performance liquid chromatography. Significantly higher activities of glutathione peroxidase and glutathione reductase (P < 0.05) were found in testes compared with epididymis. In testes, a significantly higher activity of superoxide dismutase was found than in the head and tail of epididymis (P < 0.05). Malondialdehyde concentration in head of epididymis was significantly higher than in testes, or the body and tail of epididymis (P < 0.05). Simultaneously, malondialdehyde concentration in testes was significantly higher than in body and tail of epididymis (P < 0.05). In tail of epididymis, significantly higher activities of γ-glutamyltransferase, acid phosphatase, and lactate dehydrogenase were recorded than in testes. Significantly lower concentrations of triacylglycerol and free fatty acids were recorded in epididymis tail in comparison to epididymis head (P < 0.05). It could be concluded that high activities of antioxidative enzymes in testes of boars are essential for the appropriate protection of spermatozoa and cells of testes tissue against oxidative damages. The tissues of testes and epididymis head in boars were more susceptible to lipid peroxidation. Results of the present study indicated physiological importance of antioxidative enzymes in reproductive system in boars, and thus may serve for better understanding the mechanisms of male infertility.

Influence of different periods of the year and age on the parameters of antioxidative status and oxidative stress in the blood serum of breeding bulls.

The sources of variations that may cause physiological differences between blood serum biochemistry parameters of bulls have not been investigated in detail. Aim of the present study was to establish influence of different periods of the year and the age of breeding bulls on parameters of antioxidative status and oxidative stress in their serum and to correlate these monitored variables. Research was performed on two groups, each comprising 9 Simmental bulls: a younger group (YB) (aged 2-4 years) and older one (OB) (aged 5-10 years). Blood samples for biochemical analyses were collected from jugular vein in cold (CP) and warm periods (WP) of the year. Reduced glutathione (GSH), uric acid (UA), total protein (TP), albumin (ALB), thiobarbituric acid reactive substance (TBARS), and protein carbonyl content (PCC) serum concentration were determined, as well as activities of selenium-dependent glutathione peroxidase (Se-GSH-Px), total superoxide dismutase (TSOD), manganese superoxide dismutase (MnSOD), copper-zinc superoxide dismutase (CuZnSOD) and catalase (CAT). Serum values of SeGSH-Px, MnSOD, UA and TP in OB were significantly higher compared to those in YB during CP of the year. Significantly higher PCC concentration in serum of YB and OB were established in CP of the year than in WP. TBARS serum concentration in YB was significantly higher in comparison to that in OB during CP of the year. It can be concluded that both OB and YB show a great sensitivity to climate condition alterations during CP in comparison to WP of the year and that YB show even greater sensitivity.

Influence of boar breeds or hybrid genetic composition on semen quality and seminal plasma biochemical variables.

The enzyme concentrations of seminal plasma are important for spermatozoa metabolism and function in boars. The need has arisen for introducing a biochemical evaluation of semen, along with the usual standard semen analyses. There are no data on the influence of boar breeds on the seminal plasma biochemical variables investigated in this study. Therefore, the objective was to determine the influence of breed and hybrid genetic composition of boars on semen quality and seminal plasma biochemical variables. Semen samples of 27 boars (Swedish Landrace, German Landrace, Large White, Pietrain and Pig Improvement Company hybrid-PIC-hybrid), aged between 1.5 and 3 years, were collected. After evaluation of semen quality, the seminal plasma was separated from the spermatozoa by centrifugation of semen. The seminal plasma was subjected to spectrophotometric analysis to determine alkaline phosphatase (ALP), acid phosphatase (ACP), γ-glutamyltransferase (GGT), creatine kinase (CK) and lactate dehydrogenase (LDH) and to atomic absorption spectrophotometric analysis to measure the concentration of calcium and magnesium. Conventional semen quality variables differed depending on breed and PIC-hybrid genetic composition, though these differences were typically insignificant. In the seminal plasma, significant differences were determined in enzyme activity (ALP, GGT, CK and LDH) and in calcium concentration among boars of different breeds. There are, therefore, differences in semen quality and significant differences in the seminal plasma biochemical variables among boars of different breeds and PIC-hybrid genetic composition. The data and differences in semen variables detected in the present study provide knowledge for enhancing evaluation and monitoring of boar reproductive potential, semen quality and explain the potential causes of boar infertility.