RESUMO
Infectious bronchitis (IB) is an acute, highly contagious contact respiratory disease of chickens caused by infectious bronchitis virus (IBV). IBV is very prone to mutation, which brings great difficulties to the prevention and control of the disease. Therefore, there is a pressing need for a method that is fast, sensitive, specific, and convenient for detecting IBV. In this study, a real-time fluorescence-based recombinase-aided amplification (RF-RAA) method was established. Primers and probe were designed based on the conserved regions of the IBV M gene and the reaction concentrations were optimized, then the specificity, sensitivity, and reproducibility of this assay were tested. The results showed that the RF-RAA method could be completed at 39â within 20â¯min, during which the results could be interpreted visually in real-time. The RF-RAA method had good specificity, no cross-reaction with common poultry pathogens, and it detected a minimum concentration of template of 2 copies/µL for IBV. Besides, its reproducibility was stable. A total of 144 clinical samples were tested by RF-RAA and real-time quantitative PCR (qPCR), 132 samples of which were positive and 12 samples were negative, and the coincidence rate of the two methods was 100â¯%. In conclusion, the developed RF-RAA detection method is rapid, specific, sensitive, reproducible, and convenient, which can be utilized for laboratory detection and clinical diagnosis of IBV.
Assuntos
Galinhas , Infecções por Coronavirus , Vírus da Bronquite Infecciosa , Técnicas de Amplificação de Ácido Nucleico , Doenças das Aves Domésticas , Recombinases , Sensibilidade e Especificidade , Vírus da Bronquite Infecciosa/genética , Vírus da Bronquite Infecciosa/isolamento & purificação , Animais , Galinhas/virologia , Doenças das Aves Domésticas/virologia , Doenças das Aves Domésticas/diagnóstico , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Recombinases/metabolismo , Recombinases/genética , Reprodutibilidade dos Testes , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Amplificação de Ácido Nucleico/veterinária , Primers do DNA/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Fluorescência , Técnicas de Diagnóstico Molecular/métodosRESUMO
BACKGROUND: Infectious bronchitis virus (IBV) leads to huge economic losses in the poultry industry worldwide. The high levels of mutations of IBV render vaccines partially protective. Therefore, it is urgent to explore an effective antiviral drug or agent. The present study aimed to investigate the in vivo anti-IBV activity of a mixture of plant essential oils (PEO) of cinnamaldehyde (CA) and glycerol monolaurate (GML), designated as Jin-Jing-Zi. RESULTS: The antiviral effects were evaluated by clinical signs, viral loads, immune organ indices, antibody levels, and cytokine levels. The infection rates in the PEO-M (middle dose) and PEO-H (high dose) groups were significantly lower than those in the prevention, positive drug, and PEO-L (low dose) groups. The cure rates in the PEO-M and PEO-H groups were significantly higher than those in the prevention, positive drug, and PEO-L groups, and the PEO-M group had the highest cure rate of 92.31%. The symptom scores and IBV mRNA expression levels were significantly reduced in the PEO-M group. PEO significantly improved the immune organ indices and IBV-specific antibody titers of infected chickens. The anti-inflammatory factor levels of IL-4 and IFN-γ in the PEO-M group maintained high concentrations for a long time. The IL-6 levels in the PEO-M group were lower than those in prevention, positive drug, and PEO-L groups. CONCLUSION: The PEO had remarkable inhibition against IBV and the PEO acts by inhibiting virus multiplication and promoting immune function, suggesting that the PEO has great potential as a novel anti-IBV agent for inhibiting IBV infection.
