摘要
目的研究简便快速肠道腺病毒基因型分型方法。方法采集137例非腹泻人群的粪便样本,在A549细胞中培养,提取病变细胞的基因组DNA进行PCR扩增或直接提取粪便悬液过滤上清基因组DNA进行巢式PCR扩增,对PCR阳性者进行克隆、测序以及基因型分型。结果 137份样本中,两种方法均有17份样本检出腺病毒,阳性率为12.4%(17/137)。序列分析显示,两种方法检出的腺病毒型别一致。这17份样本分别属于人腺病毒5型(5份)、7型(3份)、12型(3份)及48型(6份)。结论两种方法的敏感性、特异性一样,但方法二是直接提取粪便悬液过滤上清基因组DNA进行巢式PCR扩增,无需方法一需进行3次体外细胞感染实验后从病变细胞中提取基因组DNA进行PCR扩增,节省实验的时间及成本,特别是避免体外细胞感染过程中可能出现的污染,值得推荐。
Aim To compare two genotyping methods for genotyping of intestinal adenovirus. Methods 137 stool samples were collected from non-diarrhea groups by （I） isolating and culturing of 137 stool samples in the A549 ceils, extracting genomic DNA of diseased cells for PCR amplification and sequence analysis; （2） extracting DNA of fecal samples directly, amplified by nested PCR,to determine the viral genotype by sequence analysis. Results The positive rate of two methods were 12.4%（17/137）. Sequence analysis showed that the genotype of the intestinal adenovirus were the same by two methods.The 17 samples belonged to human adenovirus type 5（5 samples）,type 7（3 samples）,type 12（3 samples） and type 48 （6samples）. Conclusion Although the sensitivity and specificity of two methods are same,but method 2 is more convenient as it allow detect extracting genomic DNA from filtered fecal suspension supernatant and amplified by nested- PCR.
出处
《中国热带医学》
CAS
2010年第11期1301-1303,共3页
China Tropical Medicine
基金
国家高技术研究发展计划(863计划)(No.2007AA021002)
广东省自然科学基金资助项目(No.7001521)
广东省科技计划资助项目(No.2008B030301101)
关键词
腺病毒
基因型
病毒分离
流行病学
序列分析
Adenovirus
Genotype
Viral isolation
Epidemiology
Sequence analysis
