呼吸道RNA病毒,如传染性支气管炎病毒(IBV)和禽偏肺病毒(aMPV),其特征是产生呼吸损伤和对生殖器官的不利影响,由于死亡率高,鸡蛋产量和质量下降,经济上会影响家禽生产。特别是,aMPV有三种基因型,在鸡中报道频率更高:aMPV-A,aMPV-B,和aMPV-C本研究提出了一种多重RT-qPCR检测方法的设计,用于同时诊断aMPV和IBV的3种感兴趣的基因型,随后对200个有呼吸道症状的接种疫苗鸡的气管样本进行测试,最后对发现的序列进行系统发育分析。该测定检测到每个病毒基因组的多达1个拷贝。标准曲线显示在多重测定中的效率在90和100%之间,测定间和测定内的变异系数分别为0.363和0.459,测定间和测定内的变异系数分别为0.363和0.459。69.5%的样本单独或合并感染呈阳性。114例IBV阳性,13用于aMPV-A,25用于aMPV-B。未检测到aMPV-C的RNA。最常见的组合是6个样本中的aMPV-B和IBV,在2个样本的合并感染中最不常见的是aMPV-A和aMPV-B。该测定法专门用于扩增所研究的呼吸道病毒的基因组(IBV,aMPV-A,aMPV-B,aMPV-C),因为其他病毒基因组没有显示扩增(ChPV,CAstV,ANV,和FAdV)或来自阴性对照。部分基因组Sanger测序能够鉴定IBV的循环疫苗来源和野生型毒株以及aMPV-B的疫苗和疫苗来源毒株。总之,这种新开发的多重RT-qPCR被证明能够检测所研究的呼吸道病毒中的个体感染以及共感染.它被证明是快速和安全地诊断这些感染的可靠和有效的工具。此外,这项研究代表了厄瓜多尔家禽中aMPV菌株的首次报道,并证明了aMPV-A的循环,aMPV-B,和GI-13IBV毒株在该国未接种疫苗的鸡群中。因此,它强调了在厄瓜多尔更详细和定期同时识别这些病原体的重要性。
Respiratory RNA viruses such as Infectious bronchitis virus (IBV) and Avian metapneumovirus (aMPV), which are characterized by generating both respiratory damage and adverse effects on reproductive organs, affect poultry production economically due to high mortality rate and decrease in egg production and quality. Particularly, aMPV has three genotypes that have been reported with greater frequency in chickens: aMPV-A, aMPV-B, and aMPV-C. The present study proposes the design of a
multiplex RT-qPCR assay for the simultaneous diagnosis of the 3 genotypes of interest of aMPV and IBV, followed by testing of 200 tracheal samples of vaccinated chickens with respiratory symptoms and finally a phylogenetic analysis of the sequences found. The assay detected up to 1 copy of each viral genome. The standard curves showed an efficiency between 90 and 100% in the
multiplex assay and inter- and intra-assay coefficients of variation of 0.363 and 0.459, respectively and inter- and intra-assay coefficients of variation of 0.363 and 0.459, respectively. 69.5% of samples were found positive alone or in coinfection. 114 samples were positive for IBV, 13 for aMPV-A and 25 for aMPV-B. RNA of aMPV-C was no detected. The most commonly found combination was aMPV-B and IBV within 6 samples, and the least common was aMPV-A and aMPV-B in coinfection in 2 samples. The assay was specific for amplification of the genomes of the studied respiratory viruses (IBV, aMPV-A, aMPV-B, aMPV-C) as no amplification was shown from other viral genomes (ChPV, CAstV, ANV, and FAdV) or from the negative controls. Partial genomic Sanger sequencing enabled to identify circulating vaccine-derived and wild-type strains of IBV and vaccine and vaccine-derived strains of aMPV-B. In conclusion, this newly developed
multiplex RT-qPCR was shown to be able to detect individual infections as well as co-infections among the respiratory viruses investigated. It was demonstrated to be a reliable and efficient tool for rapidly and safely diagnosing these infections. Furthermore, this study represents the first report of aMPV strains in Ecuadorian poultry and demonstrates the circulation of aMPV-A, aMPV-B, and GI-13 IBV strains in unvaccinated chicken populations in the country. Thus, it highlights the importance of simultaneously identifying these pathogens in greater detail and on a regular basis in Ecuador.