Viral infections are the greatest threat to waterfowl and cause significant economic losses. Diagnosis and differentiation of three goose viruses is difficult in the field and often requires laboratory confirmation. Therefore, the aim of the study was to develop a triplex PCR and optimise its parameters for simultaneous detection of DNA of goose parvovirus (GPV), goose polyomavirus (GHPV), and goose circovirus (GoCV). The DNA of viruses isolated from field cases from the National Veterinary Research Institute’s own collection was used for the study. The primer attachment temperature, the number of reaction cycles, and the Taq DNA polymerase and Mg²⁺ concentrations were optimised. The sensitivity and specificity of this triplex PCR was also determined. Based on the obtained results, triplex PCR parameters were optimised for simultaneous detection of DNA of GPV, GHPV, and GoCV in one sample. The following PCR products of the expected size were obtained: GPV DNA of 806 bp, GoCV DNA of 571 bp, and GHPV DNA of 180 bp. The developed triplex PCR method proved to be useful for simultaneous detection of infections with three waterfowl viruses and will be used in relevant laboratory diagnostics.

Viral infections are the greatest threat to waterfowl and cause significant economic losses. Diagnosis and differentiation of three goose viruses is difficult in the field and often requires laboratory confirmation. Therefore, the aim of the study was to develop a triplex PCR and optimise its parameters for simultaneous detection of DNA of goose parvovirus (GPV), goose polyomavirus (GHPV), and goose circovirus (GoCV).

Introduction: Numerous mutations in the bovine tumour necrosis factor receptor type two (TNF-RII) gene have been identified, but their biological consequences remain poorly understood. The aim of this study was to determine whether polymorphism in the analysed loci of the bovine TNF-RII gene is linked with the size of cell subpopulations naturally infected with bovine leukaemia virus (BLV) which serve important immune functions in the host. Material and Methods: Samples originated from 78 cows. Polymorphisms in the studied gene were determined by PCR-RFLP and DNA sequencing by capillary electrophoresis. BLV infection was diagnosed by the immunofluorescence (IMF) technique and nested PCR. Cell subpopulations were immunophenotyped with IMF. Results: Similar and non-significant differences in the average percentages of TNFα+, IgM+TNFα+, and CD11b+TNFα+ cells infected with BLV were noted in individuals with various genotypes in the polymorphic sites g.-1646T > G and g.16534T > C of the TNF-RII gene, and significant differences in the percentages of these subpopulations were observed between selected microsatellite genotypes (g.16512CA(n)). Conclusion: STR polymorphism and the number of CA dinucleotide repeats in intron 1 of the TNF-RII gene influence the frequency of TNF+, CD11b+TNF+, and IgM+TNF+ subpopulations naturally infected with BLV. Polymorphism in the gene’s other two sites do not affect the size of these cell subpopulations.

Introduction: Numerous mutations in the bovine tumour necrosis factor receptor type two (TNF-RII) gene have been identified, but their biological consequences remain poorly understood. The aim of this study was to determine whether polymorphism in the analysed loci of the bovine TNF-RII gene is linked with the size of cell subpopulations naturally infected with bovine leukaemia virus (BLV) which serve important immune functions in the host. Material and Methods: Samples originated from 78 cows. Polymorphisms in the studied gene were determined by PCR-RFLP and DNA sequencing by capillary electrophoresis. BLV infection was diagnosed by the immunofluorescence (IMF) technique and nested PCR. Cell subpopulations were immunophenotyped with IMF. Results: Similar and non-significant differences in the average percentages of TNFα+, IgM+TNFα+, and CD11b+TNFα+ cells infected with BLV were noted in individuals with various genotypes in the polymorphic sites g.-1646T > G and g.16534T > C of the TNF-RII gene, and significant differences in the percentages of these subpopulations were observed between selected microsatellite genotypes (g.16512CA(n)). Conclusion: STR polymorphism and the number of CA dinucleotide repeats in intron 1 of the TNF-RII gene influence the frequency of TNF+, CD11b+TNF+, and IgM+TNF+ subpopulations naturally infected with BLV. Polymorphism in the gene’s other two sites do not affect the size of these cell subpopulations.

Introduction: Peroxiredoxin 6 (Prdx6) is a bifunctional protein with glutathione peroxidase activity and phospholipase A2 activity. Previous studies have shown a significant positive correlation between the intracellular survival ability of Brucella and Prdx6. Here, the Prdx6 enzyme with a single activity was constructed to facilitate study of the relationship between the single function of Prdx6 and Brucella infection. Material and Methods: The target open reading frame (ORF) DNAs of Prdx6 with a single active centre were prepared using gene splicing by overlap extension PCR (SOE-PCR), and the recombinant eukaryotic expression plasmids inserted by Prdx6 with the single activity centre were constructed and transfected into murine Raw264.7 macrophages. The glutathione peroxidase activity and phospholipase A2 activity of the constructed Prdx6 were examined. Results: The core centres (Ser³² and Cys⁴⁷) of Prdx6 were successfully mutated by changing the 94ᵗʰ nucleotide from T to G and the 140ᵗʰ nucleotide from G to C in the two enzyme activity cores, respectively. The constructed recombinant plasmids of Prdx6 with the single active centre were transfected into murine macrophages showing the expected single functional enzyme activity, which MJ33 or mercaptosuccinate inhibitors were able to inhibit. Conclusion: The constructed mutants of Prdx6 with the single activity cores will be a benefit to further study of the biological function of Prdx6 with different enzyme activity.

Introduction: Peroxiredoxin 6 (Prdx6) is a bifunctional protein with glutathione peroxidase activity and phospholipase A2 activity. Previous studies have shown a significant positive correlation between the intracellular survival ability of Brucella and Prdx6. Here, the Prdx6 enzyme with a single activity was constructed to facilitate study of the relationship between the single function of Prdx6 and Brucella infection.

Introduction: Laryngeal swab samples collected from three waterfowl slaughterhouses in central Taiwan were cultured and suspected isolates of Riemerella anatipestifer were identified by API 20NE and 16S rDNA PCR. Material and Methods: Serum agglutination was used for serotyping, and antimicrobial susceptibility was tested. Results: Seventy-six R. anatipestifer isolates were detected, and the prevalences in the ducks and geese were 12.3% (46/375) and 8.0% (30/375), respectively. The positive isolation rates were 65.6% for all arriving waterfowl, 76.0% for birds in the holding area, 1.6% for defeathered carcasses, but zero for degummed carcasses. A PCR examination detected R. anatipestifer in the slaughtering area frequently. Serotype B was dominant in both duck (34.8%) and goose (46.7%) isolates, but the wide serotype distribution may very well impede vaccination development. All isolates were resistant to colistin, and 79.7% were resistant to more than three common antibiotics. Conclusion: The results proved that most ducks had encountered antibiotic-resistant R. anatipestifer in rearing, which suggests that the bacterium circulates in asymptomatic waterfowl. It is worth noting that most waterfowl farms were found to harbour R. anatipestifer, and contaminated slaughterhouses are a major risk factor in its spread. Effective prevention and containment measures should be established there to interrupt the transmission chain of R. anatipestifer.

Introduction: Laryngeal swab samples collected from three waterfowl slaughterhouses in central Taiwan were cultured and suspected isolates of Riemerella anatipestifer were identified by API 20NE and 16S rDNA PCR.

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.

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.

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).