【摘要】 禽腺病毒属成员鸭腺病毒3型(duck adenovirus type 3,DAdV-3)和禽腺病毒4型(fowl adenovirus serotype-4,FAdV-4)是养殖场主要流行的病原，对养禽业造成巨大的经济损失。目前尚未见可同时快速检测这2种病原的双重荧光定量PCR检测方法，本研究基于DAdV-3 GDMM10毒株和FAdV-4 GDMZ毒株Fiber2基因序列比对及遗传进化分析，分别在Fiber2基因保守区设计特异性引物，建立了能同时鉴别临床上常见的DAdV-3和FAdV-4双重荧光定量PCR检测方法。结果表明，DAdV-3 GDMM10和FAdV-4 GDMZ Fiber2基因处于两个不同进化分支上，同源性为46.10%。建立的检测DAdV-3和FAdV-4双重荧光定量PCR方法特异性强、灵敏度高，最低可检出45拷贝/μL的DAdV-3样品和17拷贝/μL的FAdV-4样品；且检测结果可直接通过Tm值差异进行判定，简化操作时间。利用该双重荧光定量PCR方法对2022年度临床上采集的34份肝炎-心包积液疑似样品进行检测，DAdV-3和FAdV-4检出率分别为17.65%和2.94%。为进一步开展禽源DAdV-3和FAdV-4的分子流行病学及共感染致病机制奠定了基础。更多还原

【Abstract】 Duck adenovirus type 3(DAdV-3) and fowl adenovirus serotype 4(FAdV-4),members of the genus Aviadenovirus,are the main prevalent pathogens in poultry farms, causing huge economic losses to the poultry industry.Till date, there is no duplex fluorescence real-time quantitative PCR assay that can rapid and simultaneously detect these two pathogens.Based on the homologous sequences alignment and genetic evolution analysis of DAdV-3 and FAdV-4 fiber2 genes, two pairs of specific primers were designed in the conservative regions of fiber2 genes from DAdV-3 GDMM10 and FAdV-4 GDMZ strains, respectively, and a duplex fluorescence real-time quantitative PCR assay for simultaneous detection of DAdV-3 and FAdV-4 was established.The phylogenetic analysis showed that the fiber2 genes of DAdV-3 GDMM10 and FAdV-4 GDMZ strains were in two different evolutionary branches, and the homology was 46.10%.The established dual fluorescence quantitative PCR method exhibited high specificity and sensitivity for both DAdV-3and FAdV-4detection.The sensitivity of this duplex qPCR assay was 45copies/μL and 17copies/μL for DAdV-3and FAdV-4,respectively.A total of 34clinical suspected samples with hepatitises hydropericardium syndrome were collected and tested in 2022,and the positive rates of DAdV-3and FAdV-4were 17.65%and 2.94%,respectively.This study will provide a foundation for further research on the molecular epidemiology and pathogenic mechanism of DAdV-3and FAdV-4co-infections in poultry.更多还原