Introduction: The study aimed to clarify the changes in the concentration of inflammatory mediators, proteases, and cartilage degradation biomarkers in the synovial fluid of joints in an equine osteoarthritis model.

Introduction: The study aimed to clarify the changes in the concentration of inflammatory mediators, proteases, and cartilage degradation biomarkers in the synovial fluid of joints in an equine osteoarthritis model. Material and Methods: Osteoarthritis was induced in eight Mongolian horses by a sterile intra-articular injection of amphotericin B, which was injected into the left carpal joint in a dose of 2 mL (25 mg/mL). The control group comprised five horses which were injected with an equal dose of sterile physiological saline into the left carpal joint. Synovial fluid was obtained at baseline and every week after injection. Test methods were based on ELISA. Results: In the course of the osteoarthritis, the concentration of biomarkers in joint synovial fluid showed an increasing trend. IL-1, IL-6, MMP-9, MMP-13, ADAMTS-5, CS846, GAG, HA, CTX-II, and COMP concentrations sharply increased before the onset of significant symptoms of lameness, whereas TNF-α, MMP-2, and MMP-3 concentrations rose sharply after the occurrence of such symptoms. Conclusion: The results obtained confirm that the concentrations of IL-1, IL-6, MMP-9, MMP-13, ADAMTS-5, CS846, GAG, HA, CTX-II and COMP increase substantially in equine osteoarthritis, which provides a theoretical basis for the rapid diagnosis of the disease.

Introduction: Intentional and accidental poisoning of animals is often caused by readily available commercial pesticides, such as the molluscicide metaldehyde. A retrospective analysis of suspected metaldehyde poisonings between 2014 and 2016 in Italy was conducted. Material and Methods: Biological matrices were collected for toxicological analyses in the course of routine Institute activity. A total of 183 organs from dogs and cats and 49 pieces of bait, here specificall y poisoned food used to lure animals, were analysed and the presence of metaldehyde was confirmed by gas chromatography coupled to mass spectrometry (GC/MS). Results: A high content of metaldehyde was demonstrated in the analysed samples from dogs and cats: 50 (27.3%) animals were found positive for metaldehyde intoxication together with 17 pieces of bait. Conclusion: The study emphasised the need for the control of metaldehyde use by the veterinary service.

Introduction: Intentional and accidental poisoning of animals is often caused by readily available commercial pesticides, such as the molluscicide metaldehyde. A retrospective analysis of suspected metaldehyde poisonings between 2014 and 2016 in Italy was conducted. Material and Methods: Biological matrices were collected for toxicological analyses in the course of routine Institute activity. A total of 183 organs from dogs and cats and 49 pieces of bait, here specificall y poisoned food used to lure animals, were analysed and the presence of metaldehyde was confirmed by gas chromatography coupled to mass spectrometry (GC/MS). Results: A high content of metaldehyde was demonstrated in the analysed samples from dogs and cats: 50 (27.3%) animals were found positive for metaldehyde intoxication together with 17 pieces of bait. Conclusion: The study emphasised the need for the control of metaldehyde use by the veterinary service.

Introduction: The purpose of the current study was to evaluate the blood glutathione peroxidase (GSH-Px) and malondialdehyde (MDA) levels under seasonal variations in dairy cows during transition period, and to assess the relationship between chosen reproductive parameters, GSH-Px, and MDA.

Introduction: The purpose of the current study was to evaluate the blood glutathione peroxidase (GSH-Px) and malondialdehyde (MDA) levels under seasonal variations in dairy cows during transition period, and to assess the relationship between chosen reproductive parameters, GSH-Px, and MDA. Material and Methods: Holstein cows calving in January were assigned into winter group (n = 42), while cows calving in August were assigned into summer group (n = 42). Blood samples were collected from the jugular vein 21, 14, and 7 days before calving, at calving (0 day), and 7, 14, and 21 days after calving. Reproductive parameters obtained from farm records were evaluated. Results: In both groups of cows, GSH-Px activity decreased from 21 days before calving to day 0, and it gradually continued to increase until 21 days after calving. GSH-Px activity was higher in winter group compared to summer group during the transition period (P < 0.05). MDA levels in both groups increased over time starting from 21 days before calving to 0 day, but it gradually decreased thereafter. MDA levels were higher in summer group compared to winter group during the transition periods (P < 0.05). Summer group of cows showed higher intervals of calving-to-oestrus, calving-to-conception, and higher insemination index (P < 0.01). Negative correlation was recorded between GSH-Px and MDA during all examination days (P < 0.01). MDA levels correlated with calving to conception interval on day 21 before calving and day 0 (P < 0.01) and insemination index on day 0 and 21 days after calving (P < 0.01). GSH-Px activity was negatively correlated with calving to conception interval on day 21 before calving, day 0, and 21 days (P < 0.01) after calving. Negative correlation on day 21 before calving and day 0 was also determined between GSH-Px and insemination index (P < 0.01). Conclusion: This study showed that blood oxidant and antioxidant levels have affected the fertility parameters in cows under seasonal variations.

Introduction: The main problem in determination of chloramphenicol in food of animal origin is a large number of matrices. The main target of this study was to create a method for determination and confirmation of chloramphenicol in products and food of animal origin. Material and Methods: Each 5 g matrix sample was mixed with 5 mL of water and 10 mL of acetonitrile/ethyl acetate, homogenised, and centrifuged. The organic layer was evaporated and redissolved in 6 mL of 4% NaCl. The extract was cleaned up by SPE technique. Chloramphenicol was analysed by LC-MS/MS in electrospray mode. Results: The procedure was validated according to the Commission Decision No. 2002/657/EC. The apparent recoveries were in the range of 92.1% to 107.1% with a repeatability less than 11.0% (4.4%-11.0%) and within-laboratory reproducibility below 13.6% (4.7%-13.6%). Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of chloramphenicol in more than 20 different matrices.

Introduction: The main problem in determination of chloramphenicol in food of animal origin is a large number of matrices. The main target of this study was to create a method for determination and confirmation of chloramphenicol in products and food of animal origin. Material and Methods: Each 5 g matrix sample was mixed with 5 mL of water and 10 mL of acetonitrile/ethyl acetate, homogenised, and centrifuged. The organic layer was evaporated and redissolved in 6 mL of 4% NaCl. The extract was cleaned up by SPE technique. Chloramphenicol was analysed by LC-MS/MS in electrospray mode. Results: The procedure was validated according to the Commission Decision No. 2002/657/EC. The apparent recoveries were in the range of 92.1% to 107.1% with a repeatability less than 11.0% (4.4%-11.0%) and within-laboratory reproducibility below 13.6% (4.7%-13.6%). Conclusion: The method was successfully validated and proved to be efficient, precise, and useful for quantification of chloramphenicol in more than 20 different matrices.

Introduction: The paper presents the method of simultaneous determination of 10 illegal azo dyes in feed by ultra-high performance liquid chromatography coupled with tandem mass spectrometry technique. Material and Methods: The dyes were extracted with hexane, evaporated to dryness, and analysed. Separation was achieved in 7 min in a gradient elution using acetonitrile (A) and 0.1% formic acid (B) as a mobile phase. Results: The validation results showed the repeatability of the method, which was evaluated at three levels (50, 500, and 5,000 μg/kg). All the matrix calibration curves for the working ranges were linear (R² 0.9904 to 1.0), the repeatability was between 2.1% and 24%, and recoveries ranged from 77.9% to 120%. The LOD and LOQ were at 1-2 and 5-10 μg/kg for different dyes, respectively. Furthermore, the method was applied in the homogeneity tests of the in-house prepared feed containing Sudan I at the levels of 0.5, 5, and 50 mg/kg. Conclusions: A sensitive, selective, and fast multiresidue method was successfully developed and validated. Its robustness was confirmed by the analysis of an experimental feed containing Sudan I.

Introduction: The paper presents the method of simultaneous determination of 10 illegal azo dyes in feed by ultra-high performance liquid chromatography coupled with tandem mass spectrometry technique. Material and Methods: The dyes were extracted with hexane, evaporated to dryness, and analysed. Separation was achieved in 7 min in a gradient elution using acetonitrile (A) and 0.1% formic acid (B) as a mobile phase. Results: The validation results showed the repeatability of the method, which was evaluated at three levels (50, 500, and 5,000 μg/kg). All the matrix calibration curves for the working ranges were linear (R2 0.9904 to 1.0), the repeatability was between 2.1% and 24%, and recoveries ranged from 77.9% to 120%. The LOD and LOQ were at 1-2 and 5-10 μg/kg for different dyes, respectively. Furthermore, the method was applied in the homogeneity tests of the in-house prepared feed containing Sudan I at the levels of 0.5, 5, and 50 mg/kg. Conclusions: A sensitive, selective, and fast multiresidue method was successfully developed and validated. Its robustness was confirmed by the analysis of an experimental feed containing Sudan I.

Introduction: Tuberculosis is a highly infectious disease affecting humans and animals. It is caused by the Mycobacterium tuberculosis complex (MTBC) – Mycobacterium bovis and Mycobacterium caprae, which are aetiological factors of bovine tuberculosis (bTB). In Poland, the bTB eradication programme exists. Animals diagnosed with tuberculosis are in the majority of cases not treated, but removed from their herd and then sanitary slaughtered. Material and Methods: In total, 134 MTBC strains isolated from cattle in Poland were subjected to microbiological analysis. The resistance phenotype was tested for first-line antimycobacterial drugs used in tuberculosis treatment in humans: streptomycin, isoniazid, rifampicin, ethambutol, and pyrazinamide. The strains were isolated from tissues collected post mortem, so the test for drug resistance fulfilled only epidemiological criterion. Results: The analysis of drug-resistance of MTBC strains revealed that strains classified as M. bovis were susceptible to 4 antimycobacterial drugs: isoniazid, rifampicin, streptomycin, and ethambutol, and resistant to pyrazynamide. The strains classified as M. caprae were sensitive to all tested drugs. Conclusion: The results indicate that despite enormously dynamic changes in mycobacterial phenotype, Polish strains of MTBC isolated from cattle have not acquired environmental resistance. The strains classified as M. bovis are characterised by natural resistance to pyrazinamide, which is typical for this species.

Introduction: Tuberculosis is a highly infectious disease affecting humans and animals. It is caused by the Mycobacterium tuberculosis complex (MTBC) – Mycobacterium bovis and Mycobacterium caprae, which are aetiological factors of bovine tuberculosis (bTB). In Poland, the bTB eradication programme exists. Animals diagnosed with tuberculosis are in the majority of cases not treated, but removed from their herd and then sanitary slaughtered.

Introduction: Infections with bovine foamy virus (BFV) were found in many countries but there is a lack of large-scale surveys on the prevalence of BFV among dairy cattle. The aim of this study was to develop and validate the recombinant Gag protein-based ELISA and to estimate the prevalence of antibodies against BFV. Material and Methods: Gag coding region from BFV was cloned into expression vector pT7Arg-STOP, which expressed a high level of recombinant Gag protein from E.coli. The ELISA was standardised, and the cut-off value and sensitivity and specificity of the test were calculated using a receiver operating characteristic and Bayesian estimation. Results: A total of 3,051 serum samples were tested by ELISA and 939 (30.8%) sera were recognised as positive. When Bayesian approach was used, the overall true BFV prevalence was 29.7% (95% CI: 25.9-33.4%). Conclusion: Expressed Gag protein of BFV has been used successfully as an antigen for ELISA. Eventually, this study provides basic information about the epidemiological status of infection with BFV in dairy cattle in Poland, which can be used for further studies on dissemination and transmission of BFV infection.

Introduction: Infections with bovine foamy virus (BFV) were found in many countries but there is a lack of large-scale surveys on the prevalence of BFV among dairy cattle. The aim of this study was to develop and validate the recombinant Gag protein-based ELISA and to estimate the prevalence of antibodies against BFV. Material and Methods: Gag coding region from BFV was cloned into expression vector pT7Arg-STOP, which expressed a high level of recombinant Gag protein from E.coli. The ELISA was standardised, and the cut-off value and sensitivity and specificity of the test were calculated using a receiver operating characteristic and Bayesian estimation. Results: A total of 3,051 serum samples were tested by ELISA and 939 (30.8%) sera were recognised as positive. When Bayesian approach was used, the overall true BFV prevalence was 29.7% (95% CI: 25.9-33.4%). Conclusion: Expressed Gag protein of BFV has been used successfully as an antigen for ELISA. Eventually, this study provides basic information about the epidemiological status of infection with BFV in dairy cattle in Poland, which can be used for further studies on dissemination and transmission of BFV infection.

Introduction: The prevention and control of Actinobacillus pleuropneumoniae in commercial production settings is based on serological monitoring. Enzyme-linked immunosorbent assays (ELISAs) have been developed to detect specific antibodies against a variety of A. pleuropneumoniae antigens, including long-chain lipopolysaccharides (LPS) and the ApxIV toxin, a repeats-in-toxin (RTX) exotoxin unique to A. pleuropneumoniae and produced by all serovars. The objective of this study was to describe ApxIV antibody responses in serum and oral fluid of pigs.Material and Methods: Four groups of pigs (six pigs per group) were inoculated with A. pleuropneumoniae serovars 1, 5, 7, or 12. Weekly serum samples and daily oral fluid samples were collected from individual pigs for 56 days post inoculation (DPI) and tested by LPS and ApxIV ELISAs. The ApxIV ELISA was run in three formats to detect immunlgobulins M, G, and A (IgM, IgG and IgA) while the LPS ELISA detected only IgG.Results: All pigs inoculated with A. pleuropneumoniae serovars 1 and 7 were LPS ELISA serum antibody positive from DPI 14 to 56. A transient and weak LPS ELISA antibody response was observed in pigs inoculated with serovar 5 and a single antibody positive pig was observed in serovar 12 at ≥35 DPI. Notably, ApxIV serum and oral fluid antibody responses in pig inoculated with serovars 1 and 7 reflected the patterns observed for LPS antibody, albeit with a 14 to 21 day delay.Conclusion: This work suggests that ELISAs based on ApxIV antibody detection in oral fluid samples could be effective in population monitoring for A. pleuropneumoniae.

Introduction: The prevention and control of Actinobacillus pleuropneumoniae in commercial production settings is based on serological monitoring. Enzyme-linked immunosorbent assays (ELISAs) have been developed to detect specific antibodies against a variety of A. pleuropneumoniae antigens, including long-chain lipopolysaccharides (LPS) and the ApxIV toxin, a repeats-in-toxin (RTX) exotoxin unique to A. pleuropneumoniae and produced by all serovars. The objective of this study was to describe ApxIV antibody responses in serum and oral fluid of pigs.

Introduction: Aujeszky’s disease virus (ADV) infects a wide range of animals, including members of the Suidae family, i.e. domestic and wild pigs, as well as wild boar. Since wild boar are a potential ADV reservoir and a source of infection for domestic pigs, the aim of the study was to evaluate ADV antibody prevalence in the Polish wild boar population, during the years 2011 to 2014. Material and Methods: Wild boar blood samples were collected during three consecutive hunting seasons; i.e. 2011/2012, 2012/2013, and 2013/2014, and tested for ADV antibodies by ELISA. Results: ADV antibodies were detected in samples from all tested voivodships. The average seroprevalence reached 32.2%. Seroprevalence, over the examined hunting seasons, was 27.4% in 2011/2012, 32.4% in 2012/2013, and 35.5% in 2013/2014. The highest percentage of seroreagents was detected in four voivodships, situated along the western border of Poland, i.e. Zachodnio-Pomorskie (ZP), Lubuskie (LB), Dolnośląskie (DS), and Opolskie (OP). This area is positively correlated with the highest density of the wild boar population and the highest wild boar hunting bag. Conclusion: The results of this study confirm that the wild boar population may still pose a threat to domestic pigs, which is of special importance at the final stage of Aujeszky’s disease eradication programme in Poland.

Introduction: Aujeszky’s disease virus (ADV) infects a wide range of animals, including members of the Suidae family, i.e. domestic and wild pigs, as well as wild boar. Since wild boar are a potential ADV reservoir and a source of infection for domestic pigs, the aim of the study was to evaluate ADV antibody prevalence in the Polish wild boar population, during the years 2011 to 2014.