RESUMEN
BACKGROUND: Glaesserella parasuis (G. parasuis) is an influential pathogen of the pig, which induces high morbidity and mortality in naive pig populations in the pig industry. Accurate and rapid detection of the agent is important for disease control. In this study, a simple recombinase polymerase amplification (RPA) with a Lateral flow (LF) strip (RPA-LF-GPS) was developed to detect G. parasuis. RESULTS: The RPA-LF-GPS can specifically detect G. parasuis a limit of 100 CFU from other common related pathogens causing arthritis in the pig. The RPA-LF-GPS assay can use boiled synovial fluid samples as a template with the same sensitivity as other DNA extraction methods. In the detection of clinic positive synovial fluid sample, RPA-LF-GPS is equally sensitive (98.1%) compared with that of PCR (90.4%) (P > 0.05). The whole procedure of the RPA-LF-GPS assay could be finished in 1 hour without professional equipment. CONCLUSIONS: RPA-LF-GPS assay is a rapid and simple method for point-of-care diagnostic testing for G. parasuis infection.
Asunto(s)
Infecciones por Haemophilus/veterinaria , Haemophilus parasuis , Recombinasas/metabolismo , Enfermedades de los Porcinos/microbiología , Líquido Sinovial/microbiología , Animales , Infecciones por Haemophilus/microbiología , Técnicas de Amplificación de Ácido Nucleico/veterinaria , Reacción en Cadena de la Polimerasa/veterinaria , Recombinasas/química , Recombinasas/genética , Sensibilidad y Especificidad , Porcinos , Enfermedades de los Porcinos/diagnósticoRESUMEN
Selection of potent cytokine adjuvants is important for the development of Staphylococcus aureus DNA vaccines. Several potential cytokines have been proven to induce enhanced immune responses in animal models and clinical tests. There is still no reported use of IL18 as an adjuvant to design DNA vaccines against S. aureus. In this study, we cloned the main fibronectin binding protein gene (a fragment from clumping factor A, ClfA(221-550)) of S. aureus and bovine interleukin 18 (bIL18). Then recombinant plasmids were constructed based on the eukaryotic expression vector pVAX1 with or without bIL18. Indirect immunofluorescence assays in transfected HeLa cells indicated that the recombinant DNAs (rDNAs) could be expressed correctly and had antigenicity. BALB/c mice were used as experimental models to examine the immunogenicity of rDNAs in vivo. The ClfA(221-550) rDNA provoked antibody production. The bIL18 rDNA induced production of the Th1 type cytokines IL2 and IFNgamma, and ClfA(221-550) and bIL18 synergistically stimulated T-lymphocyte proliferation. The data demonstrated that bIL18 is a potent adjuvant that could be used to enhance cellular immunity.
