为了建立禽腺病毒4型(FAdV-4)的环介导等温扩增(LAMP)检测方法,本研究以FAdV-4 hexon基因高度保守的区域设计LAMP引物,并对反应体系和反应条件进行优化,同时评价该方法的特异性、灵敏度及对临床样品的适用性.结果显示,本研究建立的LAMP方法的最佳反应条件为63℃扩增65min;反应体系中内外引物最佳比例为8 ∶1,MgSO4和dNTP浓度分别为60 mmol/L和10mmol/L;与FAdV-10、FAdV-8b、FAdV-11、NDV、IBDV、AIV-H9、IBV、ILTV、ORT、CIAV、ALV等均无交叉反应,具有良好的特异性;本方法可检测的DNA最低浓度为7.5×10-8ng/μL,灵敏度是传统PCR法的100倍.用本研究建立的LAMP法和PCR法对80份临床样品进行检测,结果相符率为90.5％.上述结果表明,本研究建立的FAdV-4 LAMP检测方法具有准确性高、特异性强、灵敏度高和临床应用可行性强的优点,且结果易于判定,便于现场检测,为FAdV-4提供了一种新的现场诊断方法.

The primary flight feathers, secondary flight feathers, tail feathers and down feathers of Grus nigricollis and Grus grus were microscopically imaged, and the morphological data were determined and compared by statistical analysis. The quantitative indexes of morphological structure were statistically analyzed by single factor analysis and sample t test, and the Fisher discriminant function of characteristic indexes was established. The results were showed as following: the length of the barb in the inner web of remiges feathers, the angle between the barb in the inner web of tail feathers and the area of the accessory pinna of a primary flight feather, are remarkably different between G. nigricollis and G. grus in the terms of macrostructure; the number of cilia couples and ventral teeth from distal barbules in secondary flight feathers, the number of ventral teeth in proximal barbules of remiges feathers, the number of hamuli from distal barbules in tail feathers, the distance between nodular and diameters of nodular between the 2 kinds of Grus in the terms of microstructure also showed significantly different;macroscopic and microscopic characteristics of the Grus feathers were quantitatively analyzed, and Fisher discriminant function based on the difference index was constructed. The accuracy rate of using back substitution method to test the discriminant equation is 80.2%. It suggested that Grus of primary and secondary flight feathers and tail feathers of sample classification can be achieved by the number of hamuli, cilia couples, ventral teeth from distal barbules, the number of ventral teeth in proximal barbules, the angle between the barb in the inner web, the length of the barb in the inner web of remiges feathers, the area of the accessory pinna and Length of a feather shaft. It further enriches the identification methods or approaches of rare crane species in China.