Ornithobacterium rhinotracheale (ORT) causes significant economic losses to the poultry industry around the world. The bacterium often affects poultry as part of multiple infections causing very serious clinical signs that are usually not limited only to the respiratory system. This study’s main objective was the retrospective detection and identification of ORT in turkey flocks. ORT identification was performed in 6,225 samples taken from 133 different flocks between 2015 and 2020. Molecular methods were used, specifically real-time PCR and traditional PCR. We focused on partial 16S rRNA gene sequences of isolates, which were compared with sequences obtained from GenBank. The reaction products were analysed phylogenetically. Molecular methods indicating secondary infections was carried out, and the bacterial composition of the upper respiratory tract was 16S metasequenced for selected flocks to identify any other pathogens. The presence of ORT was detected in 30.83% of samples by real-time PCR and 28.57% by PCR. Phylogenetic analysis of the PCR products from the turkeys samples showed that their sequences resolved into two main genetic groups. Tests for the occurrence of secondary infections showed the presence of Mycoplasma gallisepticum and M. synoviae in some samples but the total absence of Bordetella avium. The upper respiratory tract in turkeys was dominated by two major phyla Firmicutes and Proteobacteria. At the genus level, the genera Ornithobacterium, Mycoplasma, Gallibacterium, Avibacterium, and Escherichia-Shigella were found which may include pathogenic bacteria that can cause clinical symptoms. The results of the analysis of multiple infection carried out in flocks with respiratory signs are probably associated with outbreaks of ornithobacteriosis in turkey flocks in Poland.

Ornithobacterium rhinotracheale (ORT) causes significant economic losses to the poultry industry around the world. The bacterium often affects poultry as part of multiple infections causing very serious clinical signs that are usually not limited only to the respiratory system. This study’s main objective was the retrospective detection and identification of ORT in turkey flocks.

Bisphenols, as endocrine disruptors, may cause a wide range of health problems in humans, but so far, not all of them have been confirmed in animals, including pigs. Since animals are also exposed to bisphenols, we hypothesised that these substances may have an effect on uterine contractility in pigs. Therefore, the aim of the study was to investigate the effect of the most-used bisphenol, bisphenol A (BPA), and a selected analogue, bisphenol F (BPF), on the contractile activity of the pig uterus. The investigation utilised smooth muscles from immature pigs (n = 6), cyclic pigs on days 12–14 of the oestrous cycle (n = 6) or early pregnant pigs on days 12–16 of pregnancy (n = 6). Strips of the myometrium were exposed to BPA and BPF at concentrations of 10⁻¹³–10⁻¹ M. Smooth muscle contractility was determined with equipment for measuring isometric contractions. BPA caused a significant decrease in contraction amplitude, and frequency and in myometrial tension in all groups examined. BPF caused a decrease in the amplitude and frequency of contractions in all groups and in myometrial tension in the early pregnant group. The obtained results indicate that both BPA and BPF relaxed the porcine myometrium, but these changes, especially in the amplitude and frequency of contractions, were more evident after BPF treatment. The extent of relaxation is dependent on the physiological status of the animals.

Bisphenols, as endocrine disruptors, may cause a wide range of health problems in humans, but so far, not all of them have been confirmed in animals, including pigs. Since animals are also exposed to bisphenols, we hypothesised that these substances may have an effect on uterine contractility in pigs. Therefore, the aim of the study was to investigate the effect of the most-used bisphenol, bisphenol A (BPA), and a selected analogue, bisphenol F (BPF), on the contractile activity of the pig uterus.

Enhancer of zeste homologue 2 (EZH2) is the human homologue of Drosophila zeste gene enhancer. The aim of this study was to determine the expression of EZH2 in canine mammary carcinomas (CMCs) and its relationship with clinicopathological features. The expression of EZH2 mRNA and protein in 53 CMC tissue and 8 normal mammary gland tissue samples was measured by quantitative real-time PCR and immunohistochemical staining assay, respectively. The relationship between EZH2 protein expression and clinicopathological features was analysed by χ2 test to further explore the clinical significance of EZH2 in CMCs. Compared with normal mammary gland tissues, EZH2 mRNA expressions were significantly increased in CMC tissues (P < 0.01). Moreover, normal mammary glands did not express the EZH2 protein but carcinomic glands did, and expression increased in CMCs with high histological grades, especially in histological grade II (P < 0.05). However, EZH2 expression was not related to age, tumour size, or metastasis (P > 0.05). The expression of EZH2 in one type of CMC was not significantly different from the expression in any other type (P > 0.05). EZH2 is highly expressed in CMCs, indicating that it can be used as a molecular marker for early diagnosis, prognosis, or therapy of CMCs.

Enhancer of zeste homologue 2 (EZH2) is the human homologue of Drosophila zeste gene enhancer. The aim of this study was to determine the expression of EZH2 in canine mammary carcinomas (CMCs) and its relationship with clinicopathological features.

Pyrrolizidine alkaloids (PAs) and tropane alkaloids (TAs) are natural contaminants of honey and respectively hepatoxic and neurotoxic compounds. Because honey is a popular constituent of the human diet, it is relevant to warrant the safety of the product. For that reason, a method for simultaneous determination of PAs and TAs in honey based on liquid chromatography– mass spectrometry was developed. The analytical protocol used sulphuric acid extraction and solid-phase extraction purification. The developed procedure was subjected to validation in terms of linearity, selectivity, repeatability, reproducibility, limits of quantification and determination, matrix effect and uncertainty. A total of 29 honey samples were analysed for the determination of PAs and TAs. All the evaluated validation parameters fulfilled the requirements of European Commission Decision 2002/657/EC. At least one of the monitored alkaloids was determined in 52% of the samples. Among the most abundant alkaloids were echimidine, intermedine and lycopsamine. The total PA concentrations ranged from 2.2 to 147.0 μg kg⁻¹. Contrastingly, none of the monitored TAs was detected in the analysed samples. An assessment of the dietary exposure to PAs from the consumption of the contaminated honeys showed that three of them would pose a risk to consumers, especially if they were children. A sensitive method suitable for simultaneous determination of PAs and TAs in honey was developed and validated. The analysis of 29 honey samples for PAs and TAs revealed that honey destined for retail could pose a risk to consumers.

Pyrrolizidine alkaloids (PAs) and tropane alkaloids (TAs) are natural contaminants of honey and respectively hepatoxic and neurotoxic compounds. Because honey is a popular constituent of the human diet, it is relevant to warrant the safety of the product. For that reason, a method for simultaneous determination of PAs and TAs in honey based on liquid chromatography– mass spectrometry was developed.

Contagious agalactia (CA) is a disease affecting small ruminants with worldwide distribution and caused by several mycoplasmas, especially M. agalactiae. The main option for systematic diagnosis under monitoring control programmes is the enzyme-linked immunosorbent assay (ELISA) test. This study was designed to appraise the performance of two commercial indirect ELISA tests using M. agalactiae p48 protein and one using total protein, for antibody detection in small ruminants after natural infection with different M. agalactiae strains. We carried out the test evaluation using sera of confirmed M. agalactiae-positive goats with clinical signs. In addition, test agreement was assessed by kappa between the three commercial ELISA tests. All three ELISA tests showed high validity scores (Youden’s J: 72.9–84%). The sensitivity values for the P48 protein-based tests were 76.9% and 84.6%, and was 79% for the total protein-based test. The specificity of all tests was 100%. In addition, between the total protein-based ELISA test and the other two ELISA tests based on the P48 protein, the agreement was substantial (kappa: 0.762–0.763) and the agreement between the latter two tests was almost perfect (kappa: 0.93). The validity parameters for all tests allowed their application for diagnostic purposes in lactating goats excreting M. agalactiae in milk and presenting clinical signs. The agreements show that any of these ELISA tests could be equally well used for diagnosis in programmes against CA.

© 2022 A. Sánchez et al. This document is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/ This document is the Accepted version of a Published Work that appeared in final form in Journal of Veterinary Research. To access the final edited and published work see https://doi.org/10.2478/jvetres-2022-0010 | Introduction: Contagious agalactia (CA) is a disease affecting small ruminants with worldwide distribution and caused by several mycoplasmas, especially M. agalactiae. The main option for systematic diagnosis under monitoring control programmes is the enzyme-linked immunosorbent assay (ELISA) test. Material and Methods: This study was designed to appraise the performance of two commercial indirect ELISA tests using M. agalactiae p48 protein and one using total protein, for antibody detection in small ruminants after natural infection with different M. agalactiae strains. We carried out the test evaluation using sera of confirmed M. agalactiae-positive goats with clinical signs. In addition, test agreement was assessed by kappa between the three commercial ELISA tests. Results: All three ELISA tests showed high validity scores (Youden’s J: 72.9–84%). The sensitivity values for the P48 protein-based tests were 76.9% and 84.6%, and was 79% for the total protein-based test. The specificity of all tests was 100%. In addition, between the total protein-based ELISA test and the other two ELISA tests based on the P48 protein, the agreement was substantial (kappa: 0.762–0.763) and the agreement between the latter two tests was almost perfect (kappa: 0.93). Conclusion: The validity parameters for all tests allowed their application for diagnostic purposes in lactating goats excreting M. agalactiae in milk and presenting clinical signs.

Contagious agalactia (CA) is a disease affecting small ruminants with worldwide distribution and caused by several mycoplasmas, especially M. agalactiae. The main option for systematic diagnosis under monitoring control programmes is the enzyme-linked immunosorbent assay (ELISA) test.

Multi-class and multi-residue analyses are very complex procedures because of the physico-chemical properties of veterinary drug residues and other contaminants. The purpose of the study was to develop an analytical method for the sensitive determination of 69 analytes in bovine milk by liquid chromatography electrospray ionisation–tandem mass spectrometry. Antimicrobial, anabolic hormone, lactone, β-agonist, mycotoxin and pesticide residues were analysed in 120 raw milk samples from different dairy farms in North Macedonia. Stable isotopically labelled internal standards were used to facilitate effective quantification of the analytes. The linear regression coefficients were higher than 0.99, the limits of detection ranged from 0.0036 to 47.94 μg/L, and the limits of quantification ranged from 0.053 to 59.43 μg/L. The decision limit values ranged from 0.062 to 211.32 μg/L and the detection capability from 0.080 to 233.71 μg/L. Average recoveries of the analytes spiked in raw milk were in the range of 70.83% to 109%, intra-day coefficient of variation (CV) values from 2.41% to 22.29%, and inter-day CV values from 3.48% to 23.91%. The method was successfully applied in the testing of bovine milk samples. In five samples residues were detected. They were sulfadimethoxine (in two samples), enrofloxacin, tetracycline and oxytetracycline and were at concentrations below the EU maximum residue limit. The method is useful for routine testing for this group of chemical hazards in bovine milk.

Multi-class and multi-residue analyses are very complex procedures because of the physico-chemical properties of veterinary drug residues and other contaminants. The purpose of the study was to develop an analytical method for the sensitive determination of 69 analytes in bovine milk by liquid chromatography electrospray ionisation–tandem mass spectrometry.

It has been suggested that coagulase-negative staphylococci can serve as reservoirs of virulence genes for other bacteria. This study assessed the presence of such genes in selected isolates recovered from meat of the giant African snail (Achatina achatina). Virulence genes were detected using a polymerase chain reaction targeting specific primers. Two representative isolates were identified using 16S rRNA gene sequencing. The results showed that the staphylococcal enterotoxin A gene (sea) was present in five out of the eight isolates studied. The isolates expressed resistance mainly to three antibiotics: chloramphenicol, norfloxacin and cloxacillin in descending order of incidence. Most importantly, the Staphylococcus sciuri isolate NEDU 181, in addition to being resistant to the three aforementioned antibiotics, also harboured the sea gene. Our findings demonstrate, for the first time, the presence of toxigenic and antibiotic-resistant coagulase-negative Staphylococcus spp. in commercially-available fresh snail meat. With staphylococcal enterotoxin A known to survive cooking temperature, this presents a food safety concern.

It has been suggested that coagulase-negative staphylococci can serve as reservoirs of virulence genes for other bacteria. This study assessed the presence of such genes in selected isolates recovered from meat of the giant African snail (Achatina achatina).

Akabane virus (AKAV) has been detected in a variety of host species in China, but there are only limited records of its occurrence in goats. However, more attention needs to be paid to understanding the diversity of viruses in this species. The aim of the study was to explore the genotype characteristics and variation trend of AKAV and their relationship with virulence in Yunnan, China. Blood samples were collected from goats during routine surveillance of goat diseases in Yunnan province in 2019. The AKAV CX-01 strain was isolated using BHK-21 cells. To understand pathogenicity, the virus was intraperitoneally (IP) and intracerebrally (IC) inoculated into suckling mice and tissue samples were subsequently analysed histopathologically and immunohistochemically. Akabane virus CX-01 strain induced encephalitis and impairment of the central nervous system with fatal consequences. Phylogenetic analysis based on the ORF sequences of the small segments indicated that the AKAV isolate used was most closely related to the GD18134/2018 Chinese midge and bovine NM BS/1strains, while phylogenetic analysis based on the medium segments showed a close relationship between CX-01 and the Chinese GLXCH01 strain. The CX-01 isolate was related to AKAV genogroup Ia and probably originated from a recombination of different strains.

Akabane virus (AKAV) has been detected in a variety of host species in China, but there are only limited records of its occurrence in goats. However, more attention needs to be paid to understanding the diversity of viruses in this species. The aim of the study was to explore the genotype characteristics and variation trend of AKAV and their relationship with virulence in Yunnan, China.

Ochratoxin A (OTA) is a mycotoxin notably produced by Aspergillus and Penicillium spp. Bacillus subtilis fermentation extract (BSFE) contains specific enzymes which hydrolyse OTA. This study evaluated the efficiency of BSFE in ameliorating the immunotoxic and nephrotoxic effects of OTA in broiler chickens. Day-old broiler chicks were divided equally into four groups of ten: control, OTA (0.5 mg/kg feed), BSFE product (1 mL/L water) and OTA + BSFE at the same concentrations. The chicks were vaccinated against avian influenza, Newcastle disease, and infectious bronchitis, and lymphoproliferation was induced in all birds by phytohaemagglutinin-P (PHA-P). Serum samples were taken before sacrifice and organ tissue samples were taken after, in which renal function biomarkers were assayed and the presence of OTA residue was evaluated by high-performance thin-layer chromatography. Protein markers of apoptosis were determined by qPCR, and tissue lesions were examined histopathologically. Exposure to OTA significantly decreased the antibody response to the vaccines and the lymphoproliferative response to PHA-P, and significantly elevated the renal function indicators: serum urea, uric acid and creatinine. It also induced oxidative stress (reduced catalase activity and glutathione concentration), lipid peroxidation (increased malondialdehyde content), apoptosis (increased Bax and Caspase-3 and decreased Bcl-2 gene levels) and pathological lesions in kidney, bursa of Fabricius, spleen and thymus tissue. Residues of OTA were detected in the serum and tissue. BSFE mitigated most of these toxic effects. BSFE counters OTA-induced immunotoxicity and nephrotoxicity because of its content of carboxypeptidase and protease enzymes.

Ochratoxin A (OTA) is a mycotoxin notably produced by Aspergillus and Penicillium spp. Bacillus subtilis fermentation extract (BSFE) contains specific enzymes which hydrolyse OTA. This study evaluated the efficiency of BSFE in ameliorating the immunotoxic and nephrotoxic effects of OTA in broiler chickens.