Introduction: The aim of the study was the application and evaluation of real-time PCRs based on the fluorescence of SYBR Green I intercalating dye for the detection of three Bacillus anthracis genes in contaminated liver and blood samples. The goals for detection were rpoB gene as a chromosomal marker, pag gene located on plasmid pXO1, and capC gene located on plasmid pXO2.

Introduction: The aim of the study was the application and evaluation of real-time PCRs based on the fluorescence of SYBR Green I intercalating dye for the detection of three Bacillus anthracis genes in contaminated liver and blood samples. The goals for detection were rpoB gene as a chromosomal marker, pag gene located on plasmid pXO1, and capC gene located on plasmid pXO2. Material and Methods: Five B. anthracis strains were used for the experiments. Additionally, single strains of other species of the genus Bacillus, i.e. B. cereus, B. brevis, B. subtilis, and B. megaterium, and strains of six other species were used for evaluation of the specificity of the tests. Three SYBR Green I real-time PCRs were conducted allowing confirmation of B. anthracis in the biological samples. Results: The observation of amplification curves in real-time PCRs enabled the detection of the chromosomally encoded rpoB gene, pag gene, and capC gene of B. anthracis. The specificity of the tests was confirmed by estimation of the melting temperature of the PCR products. The sensitivity and linearity of the reactions were determined using regression coefficients. Strains of other microbial species did not reveal real-time PCR products. Conclusion: All real-time PCRs for the detection of B. anthracis in biological samples demonstrated a significant sensitivity and high specificity.

Introduction: Avian reovirus (ARV) infections in poultry populations are reported worldwide. The reovirus belongs to the genus Orthoreovirus, family Reoviridae. The aim of the study was to evaluate the incidence of ARV infections in the poultry population based on diagnostic tests performed in 2010–2017.

Introduction: Avian reovirus (ARV) infections in poultry populations are reported worldwide. The reovirus belongs to the genus Orthoreovirus, family Reoviridae. The aim of the study was to evaluate the incidence of ARV infections in the poultry population based on diagnostic tests performed in 2010–2017. Material and Methods: Samples of the liver and spleen were collected from sick birds suspected of ARV infection and sent for diagnostics. Isolation was performed in 5–7-day-old SPF chicken embryos infected into the yolk sac with homogenates of internal organs of sick birds. Four primer pairs were used to detect the σNS, σC, σA, and µA ARV RNA gene fragments. A nested PCR was used for the detection of the σNS and σC genes. Results: In 2010–2017, ARV infection was found in birds from 81 flocks of broiler chickens and/or layers, 8 flocks of slaughter turkeys, and in 4 hatchery embryos at 17–20 days of incubation. The primers used in RT-PCR and nested PCR did not allow effective detection of ARV RNA in all virus-positive samples. Conclusion: The problem of ARV infections in the poultry population in Poland still persist. The primers used for various ARV segments in RT-PCR and nested PCR did not allow effective detection of RNA in the visceral organs of sick birds. The presented results confirm the necessity of using classical diagnostic methods (isolation in chicken embryos, AGID).

Electrical cardioversion is a therapeutic procedure used to convert various types of arrhythmias back to sinus rhythm. It is used to restore the sinus rhythm in dogs with atrial fibrillation. The effect of the electrical energy used during cardioversion on red blood cells (RBC) is not fully understood. Studies on humans reported lysis of RBC following electrical cardioversion. Similar studies have not been carried out on dogs. The aim of the study was to assess the effect of electrical cardioversion on chosen RBC parameters.

Electrical cardioversion is a therapeutic procedure used to convert various types of arrhythmias back to sinus rhythm. It is used to restore the sinus rhythm in dogs with atrial fibrillation. The effect of the electrical energy used during cardioversion on red blood cells (RBC) is not fully understood. Studies on humans reported lysis of RBC following electrical cardioversion. Similar studies have not been carried out on dogs. The aim of the study was to assess the effect of electrical cardioversion on chosen RBC parameters. The study was carried out on 14 large and giant breed dogs weighing from 30 to 84 kg with lone atrial fibrillation (lone AF). Electrical cardioversion was carried out under general anaesthesia by biphasic shock with 70–360 J of energy. Blood was collected at T0 – during atrial fibrillation, prior to cardioversion, and at T1 – 30 min after electrical cardioversion. Complete blood counts as well as total and direct bilirubin concentrations were evaluated. A maximum output of 360 J was used. In all cases, electrical cardioversion was effective, and no significant changes in the number of RBC and RBC indices were noted. Similarly, there were no statistically significant differences in the levels of total and direct bilirubin. Electrical cardioversion in dogs led neither to statistically nor clinically significant RBC lysis.

Introduction: Traditionally, evolutionary analysis of equine influenza virus (EIV) is based on the HA gene. However, the specificity of the influenza virus enables the classification of viral strains into different phylogenetic groups, depending on the gene being analysed. The aim of the study was to analyse phylogenetic paths of EIV based on M gene with reference to the HA gene.

Introduction: Traditionally, evolutionary analysis of equine influenza virus (EIV) is based on the HA gene. However, the specificity of the influenza virus enables the classification of viral strains into different phylogenetic groups, depending on the gene being analysed. The aim of the study was to analyse phylogenetic paths of EIV based on M gene with reference to the HA gene. Material and Methods: M gene of Polish isolates has been sequenced and analysed along with all M sequences of EIV available in GenBank database. Phylogenetic analysis was performed using BioEdit, ClustalW, and MEGA7 softwares. Results: The clustering of the strains isolated not only from Asia but also from Europe into one common Asian-like group of EIV was observed. Twelve nucleotide substitutions in the M gene of strains from the Asian-like group were crucial for the evolutionary analysis. We also observed homology in the M gene of the Asian-like and H7N7 strains. Conclusions: M gene specific for the Asian-like group is present in strains recently isolated in Europe and Asia, which were classified previously in the Florida 2 clade based on HA. Therefore, Asian-like group does not seem to be assigned to a specific geographical region. Traces of H7N7 strains in more conservative genes like M of some contemporary EIV strains may indicate the link between the old phylogenetic group and recent H3N8 strains. Analysis of conservative genes may be more useful in tracking the direction of virus evolution than in the genes where the high variability rate may blur the original relationships.

Veterinarians use flumequine (FLU) widely but its toxicological effects are still unclear.

Veterinarians use flumequine (FLU) widely but its toxicological effects are still unclear. FLU doses of 53, 200, or 750 mg/kg were administered orally for six weeks to pubertal male rats for evaluation of their toxicity. Weight gain was poorer after seven days of exposure to FLU 750, but relative weights of the brain, adrenal and thyroid glands, and testes were notably higher. Haematological and lipid profile parameters, cardiac markers, and inorganic phosphate significantly increased in the FLU 750 group. Blood glucose, oestradiol and serum concentrations of immunoglobulins G (IgG) and E (IgE) significantly decreased after treatment. The levels of interleukins 10 (IL-10) and 6 (IL-6) fell significantly in the FLU 200 and FLU 750 groups. Cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and cyclooxygenase-2 (Cox-2) expression amplified after treatment. Serum levels of free triiodothyronine (fT3) and free thyroxine (fT4) reduced in the FLU 200 and FLU 750 groups without changes in total T3 or T4 level. All doses of FLU significantly depressed concentrations of thyroid-stimulating hormone (TSH) and testosterone. Histopathology of thyroid glands from rats treated with FLU 750 showed degeneration and depletion of thyroid follicular epithelial cells. Expression of 8-hydroxydeoxyguanosine (8-OHdG) was increased in a dose-dependent manner in the brain, but decreased in the testes. Expression of CYP1A1 increased in the adrenal and pituitary glands. The results of this study suggest that the toxicity of FLU in rats is an effect of its disruptive influence on the pituitary-thyroid hormonal system and on the dysfunction of the immune system.

Amitraz is a formamide exhibiting both acaricidal and insecticidal activity and is frequently used by beekeepers to protect honeybee colonies against Varroa destructor mites. The aim of this apiary trial was to evaluate the impact of honeybee colony fumigation with amitraz on the level of contamination of honey stored in combs.

Amitraz is a formamide exhibiting both acaricidal and insecticidal activity and is frequently used by beekeepers to protect honeybee colonies against Varroa destructor mites. The aim of this apiary trial was to evaluate the impact of honeybee colony fumigation with amitraz on the level of contamination of honey stored in combs. Experimental colonies were fumigated four times every four days with one tablet of Apiwarol per treatment. Honey was sampled from combs of brood chambers and combs of supers one day after each amitraz application and from harvested honey. Amitraz marker residues (as a total of amitraz and metabolites containing parts of molecules with properties specific to the 2,4-DMA group, expressed as amitraz) were evaluated in honey. All analysed samples were contaminated with amitraz metabolites. 2,4-DMA and DMPF were the most frequently determined compounds. The average concentration of amitraz marker residue in honey from groups where a smouldering tablet was located directly in beehives was significantly higher than that of residue in honey from groups with indirect smoke generation. No significant effect on the honey contamination deriving from the place where it was exposed to smoke (combs of brood chambers and supers) was noted. Amitraz marker residues exceeded the MRL in 10% of honey samples from combs. Fumigation of beehives with amitraz results in contamination of honey stored in combs.

Salicylic acid is a derivative of benzoic acid and occurs in nature. The main target of this study was to develop the liquid chromatography coupled with tandem mass spectrometry technique as a method for determination of salicylic acid in feed materials and compound feed.

Salicylic acid is a derivative of benzoic acid and occurs in nature. The main target of this study was to develop the liquid chromatography coupled with tandem mass spectrometry technique as a method for determination of salicylic acid in feed materials and compound feed. Salicylic acid was extracted from feed with 0.1% hydrochloric acid in methanol. Separation was achieved in 8 min in a gradient elution using 0.1% formic acid and acetonitrile. The analyte was detected using negative electrospray tandem mass spectrometry. The procedure was validated to the specifications of the European Commission Decision No. 2002/657/EC. The validation results showed the repeatability of the method, which was evaluated at three levels (0.25, 0.5, and 1.0 mg/kg). Calibration curves for the working ranges were linear (R² 0.9911 to 0.9936), and recoveries ranged from 98.3% to 101%. The LOD and LOQ for compound feed were 0.02 and 0.05 mg/kg, respectively. Salicylic acid was found mostly in corn, and its concentrations differed depending on whether it was young or fully grown (5.30–12.8 mg/kg and 0.13–1.01 mg/kg, respectively). A sensitive and reliable method for the determination of salicylic acid in feed and compound feed using LC-MS/MS was developed.

The aim of this study was examination of honey samples collected from apiaries situated in all Polish provinces for occurrence of Clostridium spp., especially C. perfringens.

The aim of this study was examination of honey samples collected from apiaries situated in all Polish provinces for occurrence of Clostridium spp., especially C. perfringens. The study was carried out on 240 honey samples (15 samples/province). Estimation of Clostridium titre, its cultures and C. perfringens isolate characterisation were performed according to the standard PN-R-64791:1994. A multiplex PCR method for detection of genes coding cpa (α toxin), cpb (β), cpb2 (β2), etx (ε), iap (ι), and cpe (enterotoxin) toxins was used. Clostridium spp. was noticed in 56% (136/240) of samples, and its titres ranged between 0.1 g and 0.001 g. Clostridium perfringens occurrence was evidenced in 27.5% (66/240) of samples. All isolates were classified to toxinotype A. Evidence of a high number of positive samples with occurrence of Clostridium spp. indicates a potential risk to consumers’ health. The infective number of Clostridium spp. is unknown; however, the obtained results have shown that a risk assessment on the entire honey harvesting process should be made in order to ensure microbiological safety. Moreover, a detailed study should be undertaken on the antibiotic resistance of C. perfringens isolates from honey samples.

Introduction:Clostridium perfringens is commonly found in the gastrointestinal tract of animals and humans and continues to cause one of the most prevalent foodborne diseases in man.

Introduction: Clostridium perfringens is commonly found in the gastrointestinal tract of animals and humans and continues to cause one of the most prevalent foodborne diseases in man. Material and Methods: A total of 355 samples were examined for the occurrence of C. perfringens: rectal swabs from cattle, sheep, and goats, fresh stool samples from diarrhoea sufferers having been in contact with these animals, irrigation water and soil samples from the husbandry sites, and preharvesting fresh produce from farms irrigated with the sampled water. All samples were collected from Cairo and Giza governorates, Egypt. PCR analysis was carried out with positive isolates using the α-toxin gene. Sequence analysis of the gene of C. perfringens isolates was performed using the neighbour-joining approach. Bootstrap analysis was executed with 1,000 resamplings. Results: 174 C. perfringens strains were isolated with a 49.01% prevalence. The highest prevalence of C. perfringens in apparently healthy animals was found in sheep (65.45%) followed by goats (58%), buffaloes (55%), and cattle (47.1%). Its prevalence in humans being in contact with these animals was 47.5%. The bacterium’s isolation from the soil and irrigation water was achieved in 40% and 31.7% of samples, respectively, posing a risk, particularly when the water and soil contact food in the field, shown by the fresh produce isolation of 40%. A significant relationship between the prevalence of C. perfringens in animal and environmental samples was identified (P < 0.05). A significant relationship was identified neither between animal species and C. perfringens prevalence, nor between the environmental source and C. perfringens prevalence (P > 0.05). All isolates were positive for the α-toxin gene by PCR. The sequence analysis and the phylogenetic relationship of the α-toxin genes from different samples revealed that C. perfringens from faeces of apparently healthy cattle, buffaloes, sheep, and goats is a significant threat in places where it can contaminate the soil and water. In addition, the sequence of C. perfringens from humans suffering from diarrhoea was found in the same cluster with the sequence from cows, goats, and sheep. Conclusion: The role of apparently healthy animals in transmitting C. perfringens to humans, either through being in direct or indirect contact via water or soil in the cultivation of vegetables and fruits, was demonstrated.

Introduction: Highly pathogenic Asian H5-subtype avian influenza viruses have been found in poultry and wild birds worldwide since they were first detected in southern China in 1996. Extensive control efforts have not eradicated them. Vaccination prevents such viruses infecting poultry and reduces the number lost to compulsory slaughter. The study showed the efficacy of inactivated H5 vaccine from the H5N8 virus against highly pathogenic H5N8 and H5N6 avian influenza viruses in chickens.

Introduction: Highly pathogenic Asian H5-subtype avian influenza viruses have been found in poultry and wild birds worldwide since they were first detected in southern China in 1996. Extensive control efforts have not eradicated them. Vaccination prevents such viruses infecting poultry and reduces the number lost to compulsory slaughter. The study showed the efficacy of inactivated H5 vaccine from the H5N8 virus against highly pathogenic H5N8 and H5N6 avian influenza viruses in chickens. Material and Methods: Reverse genetics constructed an H5 vaccine virus using the HA gene of the 2014 H5N8 avian influenza virus and the rest of the genes from A/PR/8/34 (H1N1). The vaccine viruses were grown in fertilised eggs, partially purified through a sucrose gradient, and inactivated with formalin. Chickens were immunised i.m. with 1 µg of oil-adjuvanted inactivated H5 antigens. Results: Single dose H5 vaccine recipients were completely protected from lethal infections by homologous H5N8 avian influenza virus and shed no virus from the respiratory or intestinal tracts but were not protected from lethal infections by heterologous H5N6. When chickens were immunised with two doses and challenged with homologous H5N8 or heterologous H5N6, all survived and shed no virus. Conclusion: Our results indicate that two-dose immunisations of chickens with H5 antigens with oil adjuvant are needed to provide broad protection against different highly pathogenic H5 avian influenza viruses.