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目的研究昆明地区2002~2004年轮状病毒肠炎患儿的病毒分子流行特征。方法收集昆明医学院第一附属医院2002、2003、2004年9~12月儿科住院和门诊轮状病毒肠炎患儿的粪便标本计210份。采用聚丙烯酰胺凝胶电泳法检测轮状病毒基因组,用逆转录-聚合酶链反应(RT-PCR)以及巢式-PCR(net-PCR)对5种主要的VP7的血清型(G1、G2、G3、G4和G9)进行分型,并采用RT-PCR对VP7阳性标本进行NSP4(非结构蛋白4)的基因扩增,选取30份标本用net-PCR法扩增出NSP4CDNA500bp送去测序。结果210份标本中轮状病毒基因阳性143份(68·1%),其中长型143份,未发现短型和混合型。143份阳性标本行RT-PCR扩增VP7全基因,134份获得阳性产物,进一步用net-PCR分型显示G3型73例(73/134,54·5%),G1型38例(28·4%),未分型11例(8·2%),混合型8例(6%),G4型4例(3%),未发现G2和G9型。所有VP7阳性标本均可以扩增出NSP4的全长,其中30份标本送去测序显示有变异。结论A组轮状病毒是目前昆明地区婴幼儿腹泻的主要致病原,在不同的3年中,轮状病毒电泳型以长型为主,血清型以G3、G1型为主,G4型少见,未见G2、G9型。3年监测结果显示NSP4存在变异,但未发现轮状病毒肠炎临床症状与NSP4的变异有关。
Objective To study the molecular epidemiology of children with rotavirus enteritis in Kunming from 2002 to 2004. Methods Totally 210 stool samples from pediatric inpatients and outpatients with rotavirus enteritis in the first affiliated hospital of Kunming Medical College from 2002 to 2003 and from 2004 to 2004 were collected. The genome of rotavirus was detected by polyacrylamide gel electrophoresis. The serotypes of five major VP7s (G1, G2 , G3, G4 and G9). RT-PCR was used to amplify NSP4 (nonstructural protein 4) gene in VP7 positive samples. NSP4 cDNA was amplified by net-PCR and sequenced. Results Among the 210 specimens, 143 rotavirus genes (68.1%) were positive, 143 of them were long, and no short form or mixed form was found. In 143 positive samples, all the VP7 genes were amplified by RT-PCR and 134 were positive by net-PCR. 73 cases (73/134, 54.5%) of G3 and 38 cases of G1 4%), 11 (8.2%), 8 (6%), and 4 (3%) G4. No G2 or G9 was found. All VP7 positive samples can amplify the full length of NSP4, of which 30 samples sent to the sequencing showed variation. Conclusions Group A rotavirus is the major cause of diarrhea in infants and young children in Kunming. In the three years, the rotavirus electrophoresis type is mainly of long type, serotypes of G3 and G1 type are predominant, G4 type is rare , No G2, G9 type. Three years of monitoring showed that there was variation in NSP4, but no clinical evidence of rotavirus enteritis was found to be associated with variation in NSP4.