Development of Mutiplex Polymerase Chain Reaction for the Detection and Differentiation of Enteric Adenoviruses in Stool Samples

To find a technique of detecting and differentiating enteric adenoviruses (EAds) in clinical samples, a novel PCR approach was developed. EAds were able to be detected by use of a pair of subgroup F general primers (P1 and P2), and they were also be able to be differentiated from each other in the presence of another adenovirus type 40 (Ad40) specific primer (P3) in the same tube. Our results showed that there was one band for Ad41 and two bands for Ad40, respectively, on running-gel after PCR performance. PCR was performed on 40 specimens in parallel directly with dot-hybridization assay on the same diluted stool samples. 20 of 40 specimens were positive by hybridization (of them 12 were Ad41 and 8 were Ad40), whereas 26 were positive by PCR performance on the same samples with Ad41 18 and Ad40 8 positive as well. Our study indicated that this novel method could be used in clinical laboratory or in epidemic investigation for Eads

The Molecular Epidemiological Study on Enteric Adenovirus in Stool Specimens Collected from Wuhan Area by Using Digoxigenin Labeled DNA Probes

A summer-autumn (1994) molecular epidemiological study of enteric adenoviruses (EAds) in stool specimens collected in Wuhan area was conducted by using Digoxigenin-labelled DNA probes specific to EAd40, and EAd41, respectively.44 of 602 specimens were positive, among which 23 cases were identified as EAd40, 14 were EAd41, infection and 7 were dual infection. The ratio of males to females for the positive specimens was 1. 44. The infection rate of EAd40 and EAd41 each displayed no marked difference in seasons (summer and autumn) and similar age distribution was found between them. All of the two types of EAds in-fections predominated in patients with diarrhea under 3 years old- The results indicated that the Digoxigenin probe could detect DNA quantities as low as 1 pgwith satis factory specificity and the technique can be used for both clinical and ex-perimental purposes.

深圳市婴幼儿腹泻中肠道腺病毒感染的流行特点调查

目的 了解深圳市婴幼儿腹泻患儿肠道腺病毒的感染情况。方法 应用聚合酶链反应技术 (PCR)对 195份婴幼儿腹泻标本进行腺病毒DNA检测 ,并使用RsaⅠ内切酶对扩增产物进行限制性内切酶图谱分析 ,以鉴别肠道腺病毒型别。结果 婴幼儿腹泻标本腺病毒DNA检测的阳性率为13.33% (2 6 / 195 )。肠道腺病毒 40型的感染率为 2 .5 6 % (5 / 195 ) ,肠道腺病毒 41型的感染率为8.2 1% (16 / 195 ) ,其他型别腺病毒的感染率为 2 .5 6 % (5 / 195 ) ,发热是腺病毒感染的特征性的临床表现。结论 肠道腺病毒 40型和 41型是引起深圳市婴幼儿腹泻的重要病原 ,其中又以肠道腺病毒 41型感染为主。

人星状病毒、肠道腺病毒双重实时荧光RT-PCR方法的建立及应用

人星状病毒(Human astrovirus,HAstV)、肠道腺病毒(Enteric adenovirus,EAdV)是引起急性胃肠炎的两种常见病原体,目前尚未实现这两种病原体荧光定量RT-PCR的单管双重检测。为建立一种灵敏特异的双重荧光定量RT-PCR检测方法并应用于实验室样本检测,本研究选择HAstV、EAdV特异性引物和探针,优化反应体系和反应条件,并对该方法的灵敏性、特异性和稳定性进行评价。结果显示,该方法针对HAstV和EAdV的检出限分别可以达到522拷贝/μL和53.5拷贝/μL;与多种常见和罕见的人腹泻病毒没有交叉反应;批内和批间等重复性实验变异系数均小于5%;灵敏度高于常规PCR。本研究提示,建立的HAstV、EAdV双重荧光定量RT-PCR检测方法具有较好的灵敏度、特异性和稳定性,可用于实验室HAstV和EAdV的快速筛查。