一起传染病暴发中肠道病毒血清型鉴定和ECHO30基因特征分析

Serotype Analysis of Enterovirus in an Outbreak and Genetic Characterization of Echovirus 30 Isolated from This Outbreak

2003年5～9月,山东省泰安市发生了由肠道病毒(Enterovirus,EV)感染所致的传染病暴发,临床症状以手足口病(HFMD)为主,同时有心肌炎和无菌性脑膜炎等中枢神经系统症状患者也占较大比例。131份病人(粪便、咽拭子、脑脊液)标本中共分离到EV62株,其中ECHO1939株,EV716株,ECHO304株,其它肠道病毒13株。4株ECHO30病毒中的2株分离自2个患者的粪便标本,但用WHO肠道组合血清中和试验未能定出型别。另外2株分离自同一患者的粪便和脑脊液标本。病原学分析表明,ECHO30是引起该患者无菌性脑膜炎的病原。抗E-CHO30标准株的血清中和这4株病毒的滴度低于标准株5～20倍。VP1区全基因序列测定和同源性比较分析表明,4株ECHO30分离株病毒核苷酸同源性在98.0%～98.5%,氨基酸同源性在98.9%～99.3%,提示这4株病毒来源于同一传播链,2003年5-9月ECHO30在该地区可能有局部流行。系统进化树分析表明,ECHO30病毒可以划分为6个基因型,其中基因型1～5为GenBank中已发表的ECHO30分离株,山东分离株与其它5个基因型成员核苷酸差异分别在9.4%～24.4%,在进化树上形成了较独立的分支,是一个新基因型,将其划分为第6基因型。

During May to September in 2003,an outbreak of enterovirus infection occurred in Taian City,Shandong Province.The main clinical symptom is that of hand-foot-and-month disease(HFMD) ,and a number of patients suffered from myocarditis and some central nervous system symptoms such as aseptic meningitis.62 enterovirus strains were isolated from a total of 131 patients' samples in our laboratory. Among these viruses, 39 strains (62.9%) were Echovirus type 19(ECHO19),6 strains (9.68 %) were enterovirus type 71 (EV 71), 4 strains(6.45% )were Echovirus type 30(ECHO30)and 13 strains were other unidentified enteroviruses. Among the 4 strains of ECHO30,2 strains were isolated from the stool specimens and could not be typed by neutralization test using RIVM enterovirus typing panel sera; and the other 2 strains were isolated from the stool specimen and cerebrospinal fluid(CSF) of the same patient. Etiology analysis showed that ECHO30 was one of the pathogens of aseptic meningitis of these patients. Neutralizing antibody assay proved all of the four ECHO30 isolates had lower neutralization titers than the prototype strain Bastianni and suggested there were some differences a-mong their antigenic epitopes.The complete VP1 gene sequence and identity analysis suggested that there were 98.0 % ~ 98.8 % of nucleotide homology and 98.9 % ~ 99.3 % of amino acid homology among these 4 ECHO30 viruses, which indicated that these 4 viruses may derive from the same transmitting chain and had local epidemic during May to September in 2003. Phylogenetic tree based on complete VP1 sequence showed the ECHO30 viruses can be divided into six genotypes, genotypes 1,2,3,4 and 5 include the strains that already have released in GeneBank. The nucleotide diversity of Shandong strains compared with that of the members which belongs to other 5 genotypes ranged from 9.4% to 24.4 % . They formed a separate branch in phylogenetic tree and designed as new genotype 6.