Polish Journal of Veterinary Sciences

This study was carried out to evaluate the potential effects of 90 days-long dietary supple- mentation of probiotic and yeast culture on immunity condition of lambs. Fifteen Rahmani growing male lambs (about 5 months old and 23.21±2.75 kg body weight) were randomly allo- cated to three equal groups consisting of 5 animals each. The animals in the first group, served as a control (group C), were fed a basal diet without any supplementation. The lambs in the second and third group were fed the basal diet supplemented with probiotic (group Y) or yeast culture (group YC), respectively. The probiotic consisted of live yeast (Saccharomyces cerevisae) alone, while the yeast culture was composed of Saccharomyces cerevisiae and the media on which it was grown. In group Y and YC, each lamb was supplemented daily with 0.5 g and 7.0 g of live yeast and yeast culture, respectively. Blood samples were collected before feeding the supplements and then every 15 days until the day 90th. Total and differential leucocytic counts, total protein, albumin, IgA, IgG and IgM levels were measured in blood. There were insignificant (p>0.05) variations in the levels of total and differential leucocytic counts and total protein among the groups throughout the experiment. However, significant differences (p<0.05) were found in globulin, IgA, IgG and IgM in both (Y) and (YC) groups, but the effect of yeast culture seems to be better than that of the probiotic. In conclusions, the obtained results indicate that the tested probiotic and yeast culture improve the immunological status of lambs.

In the last decade, atmospheric plasma has been used to treating bacterial infections in humans due to its bactericidal effects; however, its efficacy in dogs is unclear. This study evaluated the in vitro bactericidal efficacy of atmospheric plasma on Staphylococcus pseudinter- medius and Pseudomonas aeruginosa, two of the most important bacterial agents isolated from canine pyodermas. Three isolates each of S. pseudintermedius and P. aeruginosa obtained from dogs with pyoderma were subjected to atmospheric plasma. The isolates from the control group were not exposed to plasma, while those from the treatment groups were exposed to plasma for 15 (7.5 J/cm2), 30 (15 J/cm2), 60 (30 J/cm2), or 90 (45 J/cm2) seconds. After each treatment, a reduction in colony formation was observed. Bacterial viability was evaluated using the LIVE/ DEAD® BacLight™ Bacterial Viability Kit. The antibacterial effects were evaluated with Image J software and significance was assessed statistically in comparison to the control group. The bactericidal effect of atmospheric plasma against both bacteria increased significantly in a time-dependent manner. These results demonstrate the bactericidal capacity of atmospheric plasma, and suggest that it could serve as an alternative treatment method for canine pyoderma. Further studies are needed to evaluate the safety and efficacy of atmospheric plasma in dogs.

The present study attempted to elucidate possible routes leading to the achievement of sero- positive results, among young (aged ≤1 year) wild boar population. In the years 2017-2018, the National Reference Laboratory (NRL) for African swine fever (ASF) in Poland examined nearly 27-thousand wild boar blood samples, collected during an active surveillance of ASF risk zones, for the presence of viral DNA and anti-ASFV antibodies. Out of all the examined samples, 420 were positive. However, in more than half of them (292 samples) antibodies against African swine fever virus (ASFV) were detected, while ASFV DNA was not detected in blood. Out of all 292 seropositive/PCR-negative samples, 126 belonged to young wild boars (aged ≤1 year). For this reason, the NRL in Poland has examined 10 selected seropositive wild boar carcasses to confirm or exclude post-mortem lesions for ASF as well as to investigate the presence of viral DNA in the internal organs. Neither pathological lesions for ASF nor the presence of genetic material of ASFV were found in the examined wild boars. To elucidate this outcomes, following hypotheses about possible reasons of the obtained results were drawn: the presence of convalescent animals, infection of low-virulent ASFV isolate and the vertical transmission of antibodies through the colostrum.

The aim of the present study was to investigate blood parameters and subsequent fertility in cows with or without increased postpartum polymorphonuclear neutrophil activity. The study was conducted with 15 Brown Swiss cows between 1-3 lactations. Polymorphonuclear neutrophil activities were assessed at 10±4 days before and after parturition. The cows which maintained their phagocytic and oxidative burst activites compared to the prepartum period were classified as control (CON), and cows which increased phagocytic and oxidative burst activites were defined as increased cellular immune response (ICIR) cows. Energy, protein metabolism markers, hepatic enzymes, blood mineral levels and body condition scores were measured at -10±4, 3±2, 10±4 days relative to parturition. Pregnancy rates, the number of inseminations, and calving to pregnancy intervals were evaluated. The mean non-esterified fatty acid (NEFA) and beta -hydroxybutyric acid (BHB) concentrations were lower in ICIR cows. Mean serum calcium (Ca) concentrations were in subclinical hypocalcemia level at day 3±2, 10±4 days postpartum in CON cows. Postpartum immune cell functions and NEFA, BHB concentrations were negatively cor- related. The calving to pregnancy interval were longer in the control cows. However, total preg- nancy rates and the number of insemination in both groups were similar. In conclusion, postpar- tum polymorphonuclear neutrophil activity is affected by periparturient metabolic status. Postpartum energy metabolites negatively affected the postpartum cellular immune response. The increased postpartum polymorphonuclear neutrophil activity at early postpartum period is positively related with subsequent fertility in dairy cows.

The European rabbit (Oryctolagus cuniculus) is a good model in biomedicine used in research on several human diseases. The reference values of B and T cells and their subpopu- lations are very important to understand how the adaptive immune system is responding to infectious agents. The aim of this study was to determine values of B and T cells and their subpopulations in Polish mixed-breed rabbits, considering seasons of the year and sex. The study was performed on 200 Polish mixed-breed rabbits and the percentage of B and T lymphocytes was measured cytometrically using mouse anti-rabbit antibodies. The study revealed that the season of the year and sex of the animals affected the percentage of B- and T-cells and their subpopulations in peripheral blood. Statistically significant values of CD19+ B-cells in spring and autumn, of T CD5+ cells in spring and winter, of T CD4+ in spring, summer, autumn and winter, of T CD8+ in winter and of T CD25+ in spring were noted. Generally the highest values were found mainly in warm part of the year, while the lowest in colder months. A statistical significance was also observed between males and females – changes were found in T CD4+ and T CD25+ lymphocytes in spring, T CD8+ cells in winter and higher percentage was generally obtained in females than in males. The only exception was the T CD5+ subpopulation in which no differences were observed between the sexes and throughout the year. This is the first paper on adaptive immune system cell values in the European rabbit of domestic breeds.

To date, only a few studies on the azithromycin (AZM) pharmacokinetics in ornamental birds have been published. In the current study AZM concentrations in domestic pigeon (Columba livia domestica) plasma samples were analyzed using a validated ultra-high performance liquid chromatography tandem mass spectrometry method. The aim of the current study was to carry out an analysis of the pharmacokinetics and pharmacodynamics after administration of a single oral dose of a sustained-release AZM formulation and to conduct a simulation of treatment based on selected minimal inhibitory values. The study was performed with 12 healthy adult pigeons, both sexes. The pigeons tolerated AZM very well and no adverse effects were observed in any animal during the study. Based on the observed characteristics of the pharmacokinetics/ /pharmaco dynamics profiles of AZM in pigeons, it should be noted that 35 mg/kg per os as a single starting dose and 25 mg/kg every 24 h are recommended for treatment of both suscep- tible and less susceptible pathogens.

From the regulatory point of view a strong link between an animal model and human pharmacodynamics of biological drugs is very important to qualify the model as “relevant”. Consistent changes in cell population between human physiology and animal model gain value of this model which then can be pharmacodynamically “relevant” from the regulatory point of view. Consequently, the aim of this study was to determine how similar to human observations is the effect of selected biological drugs on blood cells in a pig model. The study was to carry out a comparative analysis of the variability of selected biochemical and hematological parameters of the blood after administration of five different human therapeutic monoclonal antibodies (mAbs) after a single subcutaneous (SC) dose in breeding pigs. The tested drugs were siltuximab (Syl- vant®), omalizumab (Xolair®), infliximab (Inflectra®), pembrolizumab (Keytruda®), and vedoli- zumab (Entyvio®) given in a single 1 mg/kg SC injection. Each of the tested drugs exerted a sig- nificant effect on at least two of the tested parameters three weeks after the administration. Siltuximab significantly influenced 9 of the analyzed parameters. Vedolizumab significantly influenced 8 of the analyzed parameters. Infliximab had the lowest impact of all the tested drugs, as it significantly influenced only two of the analyzed parameters. The study has proved that the impact of mAbs on the analyzed parameters can be significantly extended over time. This requires the monitoring of hematological parameters in the pig model even many weeks af- ter administration of a drug in a relatively small dose.

Anestrus is essential to an unsuccessful pregnancy in dairy cows. One of the many factors that influences anestrus is the inactive ovary. To characterize in detail the plasma metabolic pro- file, anestrus cows suffering from inactive ovaries were compared with those with natural estrus. The Holstein cows 60 to 90 day postpartum in an intensive dairy farm were assigned into inactive ovaries groups (IO, n=20) and natural estrus group (CON, n=22) according to estrus signs and rectal palpation of ovaries. Plasma samples from two groups of cows were collected from the tail vein to screen differential metabolites using gas chromatography/mass spectrometry (GC/MS) techniques and multivariate statistical analysis and pathways. The results showed that 106 compounds were screened by GC/MS and 14 compounds in the IO group were decreased by analyzing important variables in the projection values and p values of MSA.Through pathway analysis, 14 compounds, mainly associated with carbohydrate metabolism and amino acid meta- bolism, were identified to results in IO, which may seriously affect follicular growth. Metabolo- mics profiling, together with MSA and pathway analysis, showed that follicular growth and development in dairy cows is related to carbohydrate and amino acid metabolism by a single or multiple pathway(s).

The objective of this study was to evaluate the effects of dietary zeolite clinoptilolite on re- productive performance, serum progesterone and insulin-like growth factor 1 (IGF-1) concentra- tions in 78 Holstein Friesian (HF) cows during pregnancy and lactation. The cows were divided into two groups comprising 40 (control group; CON) and 38 (CPL group) cows. To assess repro- ductive performance of HF cows the following variables were registered: the interval from calv- ing to first insemination (days open to first service, DFS), the interval from calving to pregnancy (days open to pregnancy, DOP), and the number of services per pregnancy (NSP). The average values of progesterone (5.64±0.59 ng/mL vs. 5.16±0.64 ng/mL) were not statistically different (p<0.05) and IGF-1 levels (400.17±17.72 ng/mL vs. 348.36±20.39 ng/mL) were higher in the CON than in the CPL group which received 50 g of clinoptilolite twice a day. However, in the CPL group ovarian cyclity resumed on days 40 and 60 postpartum. In addition, DFS (p<0.05) and DOP (p>0.05) were shorter in the CPL than in the CON group (115.1±19.9 and 137.5±36.3 days vs. 124.2±17.3 and 143.8±33.5 days, respectively). During 305 days of lactation, milk production was higher in the CPL vs. CON (8325.5±628.8 kg vs. 8050±586.8 kg). The NSP was lower in the CPL than in CON group (1.91 vs. 2.14). The dietary clinoptilolite supplement had a positive in- fluence on milk yield, exhibited modulating effects on endocrine status of dairy cows, and im- proved reproductive performance, with the decreased NSP, and fewer DFS and DOP.

Fumonisins are highly toxic metabolites produced by Fusarium proliferatum and Fusarium verticillioides. Little is known about the effects of a chronic low level of fumonisins on intestinal structure and innervation in monogastric animals, even though the intestine is the first organ exposed to fumonisins. The influence of the most prevalent strains of fumonisins, FB1 and FB2, on intestinal and liver morphology, the enteric nervous system and intestinal epithelial cell prolif- eration was investigated in an experimental rat model of fumonisin intoxication. Adolescent (5-weeks-old), male Wistar rats were randomly divided into a control group (C group) not treated with fumonisins or intoxicated with fumonisins (FB group). FB1 together with FB2 were daily administered intragastrically at a dose of 90 mg/kg body weight for 21 days. The damaging effect was assessed by determination of the activity of ALAT and AspAT. Samples from the small intes- tine and liver were taken and blood samples were collected to determine the activity of gamma-glutamyl transferase (GGT) and amylase. The exposure to FBs resulted in histopathological degenerative alterations in hepatocytes, including mild vacuolar degeneration and ballooning. FB exposure was also toxic in the duodenum and jejunum, where significant changes in morphology, cell proliferation, collagen wall fibres and innervation were observed. Taken together, the results obtained strengthen the hypothesis that chronic exposure to FBs could induce intestinal damage, including damage to the enteric nervous system and may have consequences for general health.

This study set out to investigate, for the first time, the distribution and colocalization pattern of cocaine-and amphetamine-regulated transcript (CART) and one of the calcium binding-proteins: parvalbumin (PV) in the chinchilla’s hippocampus proper (HP). HP, consisting of Ammon’s horn (CA) and the dentate gyrus (DG), is an important component of the limbic system, involved in learning and memory processes. CA showed a higher immunoreactivity of CART (-IR) compared to DG. CART-IR neurons were mainly observed in the molecular layer of DG and in the pyramidal layer of CA. CART-IR fibers were present in the granular layer; in the hilus numerous mossy fibers were detected, while in the molecular layer CART-IR fibers were not found. In all CA fields (CA1-CA3), CART-IR fibers were only present in the lacuno- sum-molecular layer. Immunofluorescence with double- labeling showed that only CART-IR cells stained positive for PV, whereas in CART-IR fibers there was no PV-positive reaction. Our research supplements missing knowledge about the distribution and colocalization pattern of CART with PV in the chinchilla’s hippocampus, and also provides a better understanding of the similarities and differences among individuals of the same species and also with other mammals.

The aim of this study was to evaluate the effect of high doses of calcium bentonite on the blood parameters, anticoccidial activity and intestinal histology of broiler chickens. Three undred and sixty one-day old broilers were distributed into three treatments (T+VE, T-VE, TB )with three replicates. Amprolium was added to the feed of the positive control group, calcium bentonite powder was added to the TB group, and nothing was added to the feed of the T-VE group. Coccidiosis was induced on day 14, the birds were kept until day 49, measurements of the diffe- rent variables started from week 3, blood samples were collected via wing vein, and fecal oocysts were counted from the intestinal contents of each individual bird using the McMaster techni- que. A decrease in feed consumption, body weight gain and conversion ratio was noticed in the calcium bentonite group. Broilers in the calcium bentonite group (TB ) and negative control group (T-VE ) showed clinical signs of coccidiosis (blood in feces) and the number of oocysts in feces increased with time. Histopathological examinations of the affected caeca also demonstrated excessive tissue damage, hemorrhage, the presence of clusters of large schizonts and merozoites in the tissue, and coccidian oocysts in the lumen. Feed conversion was highest in the T+VE group.

Injection of lymphokine activated killer (LAK) cells is known as useful for activation of cellular immune system. Although the effect of LAK cells has been clarified in human or mice, this effect on function of immune cells has not been examined in calves. Healthy ten Holstein calves were injected with the LAK cells 2 days after birth (LAK Group), and another eight calves were observed as controls (Control Group). All calves received the colostrum formulation on the day of birth, and then, were inoculated with a live attenuated vaccine of bovine herpesvirus (BHV)-1 at 2 (the first vaccination) and 6 (the second vaccination) weeks after birth. Peripheral blood of their dam obtained 3 weeks before calving was used for preparation of LAK cells. Blood samples were taken prior to vaccine inoculation and 3 days after the first inoculation, as well as 3 and 6 days after the second vaccination from all calves. Numbers of CD8+ and CD21+ cells increased significantly after the second vaccination in the LAK Group compared with Control Group. The present study suggested the improved effect of injecting LAK cells originated from dams on immune cells function of young calves after BHV-1 live vaccine.

To explore the role of Toll-like receptors (TLRs) and interferon (IFN) in the innate immunity against porcine epidemic diarrhea virus (PEDV), we detected the expression of TLR genes in PEDV-infected IPEC-J2 cells by real-time PCR. We also detected the level of interferon α (IFN-α) and interferon γ (IFN-γ) by enzyme-linked immunosorbent assay (ELISA). Results showed that IPEC-J2 cells exhibited a clear pathological change after PEDV infection at 24 h. In addition, TLR7, TLR9 and TLR10 expressions were significantly upregulated in PEDV-infected IPEC-J2 cells at 24 h. Interestingly, the expression patterns of TLR2 and TLR4 were consistent at different stages of PEDV infection. The expression level of TLR3 decreased significantly with the increase of infection time, but the expression levels of TLR5 and TLR8 genes at 6 h and 12 h were significantly lower than those in the control group (p<0.01). There were significant correlations among the expression levels of TLR genes (p<0.05). Cytokine detection showed that the secretion level of IFN-α in the PEDV-infected group was significantly higher than that in the control group (p<0.01), and IFN-γ at 6 h and 12 h after PEDV infection was significantly higher than that in control group (p<0.01). Therefore, our results suggest that PEDV infection can induce innate immune responses in intestinal porcine jejunum epithelial cells, leading to changes in the expression of Toll-like receptors, and can regulate the resistance to virus infection by affecting the release levels of downstream cytokines.

Introduction: Effective and safe anesthesia for rodents has long been a leading concern among biomedical researchers. Intraperitoneal injection constitutes an alternative to inhalant anesthesia.

Purpose: The aim of this study was to identify a safe, reliable, and effective anesthesia and postoperative analgesia protocol for laboratory rats exposed to painful procedures.

Material and methods: Twenty-seven female Wistar rats in an ongoing study that required surgery were randomized into groups for three different intraperitoneal anesthesia protocols and three different analgesia regimens. The anesthesia groups were (1) medetomidine + ketamine (MK), (2) ketamine + xylacine (KX), and (3) fentanyl + medetomidine (FM). Three analgesia groups were equally distributed among the anesthesia groups: (1) local mepivacaine + oral ibuprofen (MI), (2) oral tramadol + oral ibuprofen (TI), and (3) local tramadol + oral tramadol + oral ibuprofen (TTI). A core was assigned to measure anesthesia (0-3) and analgesia (0-2) effectiveness; the lower the score, the more effective the treatment.

Results: The mean MK score was 0.44 versus 2.00 for FM and 2.33 for KX. Mean score for analgesia on the first postoperative day was TTI (4.66) TI (9.13), and MI (10.14). Mean score 48 hours after surgery was TTI (3.4), TI (6.71), and MI (9.5). These differences were statistically significant.

Conclusion: MK was shown to be a reliable, safe, and effective method of anesthesia. The TTI analgesia regimen is strongly recommended in light of these results.

The objectives of this study were to determine the genetic relatedness, antibiogram and virulence factors of Staphylococcus aureus (S. aureus) isolated from bovine mastitis, associated farm workers, dairy cow farm veterinarians (private veterinarians), veterinary students, and non-veterinary university students. A total of 84 S. aureus isolates (27 from mastitis, 11 from farm workers, 9 from private veterinarians, 22 from veterinary students, and 16 from non-veterinary university students) were used to determine antimicrobial sensitivity patterns using disk diffusion test, virulence factors using PCR technique and phylogenic analysis using pulsed field gel electrophoresis. All S. aureus isolates were resistant to 2 or more commonly used antibiotics. All isolates from mastitis, farm workers, and veterinary students carried the genes encoding coagulase and thermonuclease factors while isolates from non-veterinary university students carried the genes encoding coagulase, clumping, and thermonuclease factors. The mecA gene was detected in 22.2%, 81.8%, 100%, 95.5% and 100% of isolates from mastitis, farm workers, private veterinarians, veterinary students, and non-veterinary university students, respectively. In the phylogenic analysis, 10 (45.5%), 6 (66.7%) and 8 (72.7%) isolates from veterinary students, private veterinarians and farm workers, respectively were more than 80% similar to isolates from mastitis. Results of this study indicate that S. aureus isolates from mastitis milk and those from related dairy cow personnel and veterinarians share similar antimicrobial sensitivity patterns and virulence factors, therefore a common source of bacteria may exist. Furthermore, possible transmission of S. aureus between cows and cow-related personnel and vice versa could also exist.

Enterotoxigenic Escherichia coli (ETEC) is the causative agent of a wide range of diseases, which are the important cause of illness and mortality in piglets. ETEC strains expressing F4 fimbriae are frequently associated with post-weaning diarrhea (PWD) and lead to great economic losses in swine production industry worldwide. The aim of this study was to establish a rapid and effective isothermal amplification method for detection of F4 fimbriae. Loop-mediated isothermal amplification (LAMP), Polymerase spiral reaction (PSR) and cross-priming ampli- fication (CPA) were used to develop and optimize the detection method first time. Subsequently, the specificity and sensitivity of these methods were evaluated, and the clinical samples were detected with these methods. All the F4-positive samples could produce ladder-like amplifica- tions products and lead the chromogenic substrate SYBR Green I produce green fluorescence, while in blank control and negative samples lack of this pattern or remained orange. The sensi- tivity of LAMP and CPA were 10 times higher than PSR method. Meanwhile, these three methods were validated with clinical samples, 7 were found positive, while 125 samples were negative, the testing results were consisted with the real-time PCR method. These findings suggested that the isothermal amplification based on the F4 fimbriae is a rapid, effective and sensitive method under resource constrains.

The aim of the present study was to investigate inline lactate dehydrogenase (LDH) dynamic changes based on different cow factors – different number and stages of lactation, milk yield, and the status of reproduction in clinically healthy dairy cows.

In the Herd Navigator system, LDH activity levels (μmol/min per litre) were measured using dry-stick technology. A total of 378 cows were selected. According to their reproductive status, the cows were classified as belonging to the following groups: Fresh (1 – 44 days after calving); Open (45 – 65 days after calving); Inseminated (1 – 35 days after insemination); Pregnant (35 – 60 days after insemination and pregnant). According to their productivity, the cows were classified into the following groups: <15 kg/day, 15 – 25 kg/day, 25 – 35 kg/day and >35 kg/day. The cows were milked with a DeLaval milking robot (DeLaval Inc. Tumba Sweden) in combination with a Herd Navigator analyser (Lattec I/S. Hillerød Denmark).

In conclusion inline dynamic changes in the milk LDH concentration may increase together with the rise in the lactation period frequency. The highest LDH level determinated in the group of the fresh cows ranged from 5 to 10 DIM, while the highest LDH concentration level was found in the fresh cow milk. Thus, there was a positive relationship between the milk concentration of LDH and the milk yield.

The aim of the study was to develop a reliable and cost-effective method for detection of nonsense mutation in APAF1 gene causing lethal effect called HH1 (Holstein Haplotype1) and to evaluate its prevalence in a sample of Polish Holstein-Friesian bulls. One hundred seventy eight bulls born between 1996 and 2017 were included in the analysis. They were kept in four artificial insemination centers and have in the pedigree the known carrier of HH1. All bulls were diagnosed by novel PCR-SSCP technique. Specific amplicons of 261 bp APAF1 gene fragment were used to detect changes in single stranded conformation (SSCP) caused by nonsense mutation C/T responsible for HH1. Each new carrier was used to trace another potential carriers among their offspring available in Polish Holstein Bull Repository Database. Among 178 bulls, 85 HH1 carriers were found. Our results show that nonsense mutation in APAF1 gene is already transmitted and segregating in Polish Holstein-Friesian cattle and its frequency may increase if no action will be undertaken against actual carriers.

The in vitro anti-Babesia canis activities of nine essential oils were investigated. Among the tested essential oils Achillea millefolium, Eugenia caryophyllus and Citrus grandis were the most active (IC50 values of 51.0, 60.3 and 61.3 μg/mL, respectively). The oils from Abies sibirica, Rosmarinus officinalis, Eucalyptus globulus, Cinnamonum zeylanicum, Mentha piperita and Pinus sylvestris were less active (IC50 values of 134.3, 237.3, 239.3, 367.9, 837.5 and 907.3 μg/mL, respectively). The results support the concept that some essential oil constituents may be useful in the clinical management of babesiosis.