CAJ | 학술논문

目的：了解我国不同来源非O157产志贺毒素大肠埃希菌（ Shiga toxin-producing Esche-richia coli， STEC）分离株携带的eae基因型别。方法 PCR扩增eae全长基因，扩增产物经测序并拼接后在美国国立生物技术信息中心（ NCBI）中进行BLASTn比对，判定其eae基因型别，并与GenBank中已公布的30种 eae 不同型别的序列及产志贺毒素大肠埃希菌主要流行血清型（ O157∶H7， O26∶H11， O103∶H2， O111∶H8， O145∶H28， O45∶H2， O121∶H19）代表性基因组测序菌株的eae序列用Neighbor-Joining法构建系统发生树，分析其进化关系。根据mlst．ucc．ie数据库提供的大肠埃希菌多位点序列分型（ multilocus sequence typing， MLST）方案对菌株进行MLST分型分析，确定菌株等位基因型及序列类型，并用BioNumerics软件构建最小生成树（ minimum spanning tree， MST），分析携带eae基因的非O157 STEC分离株与溶血性尿毒综合征相关大肠埃希菌（ HUS-associated en-terohemorrhagic E．coli，HUSEC）、E．coli MLST数据库中所有人源流行血清群O26、O103、O111、O121、O145、O157 STEC菌株的进化关系。结果10株不同来源的非O157产志贺毒素大肠埃希菌分离株经扩增拼接后均获得了约2．8 kb的eae基因全长序列，分为3种已知的eae基因型别，其中以β1型为主（6／10），2株θ型，2株γ1型。某些eae序列与主要流行血清型STEC的eae序列完全一致。10株菌分为7个ST型，与HUSEC、人源流行血清群STEC菌株具有相近的进化关系。结论我国非O157产志贺毒素大肠埃希菌分离株eae基因存在一定程度的多样性，eae阳性菌株具有潜在的高致病性。
목적：료해아국불동래원비O157산지하독소대장애희균（ Shiga toxin-producing Esche-richia coli， STEC）분리주휴대적eae기인형별。방법 PCR확증eae전장기인，확증산물경측서병병접후재미국국립생물기술신식중심（ NCBI）중진행BLASTn비대，판정기eae기인형별，병여GenBank중이공포적30충 eae 불동형별적서렬급산지하독소대장애희균주요류행혈청형（ O157∶H7， O26∶H11， O103∶H2， O111∶H8， O145∶H28， O45∶H2， O121∶H19）대표성기인조측서균주적eae서렬용Neighbor-Joining법구건계통발생수，분석기진화관계。근거mlst．ucc．ie수거고제공적대장애희균다위점서렬분형（ multilocus sequence typing， MLST）방안대균주진행MLST분형분석，학정균주등위기인형급서렬류형，병용BioNumerics연건구건최소생성수（ minimum spanning tree， MST），분석휴대eae기인적비O157 STEC분리주여용혈성뇨독종합정상관대장애희균（ HUS-associated en-terohemorrhagic E．coli，HUSEC）、E．coli MLST수거고중소유인원류행혈청군O26、O103、O111、O121、O145、O157 STEC균주적진화관계。결과10주불동래원적비O157산지하독소대장애희균분리주경확증병접후균획득료약2．8 kb적eae기인전장서렬，분위3충이지적eae기인형별，기중이β1형위주（6／10），2주θ형，2주γ1형。모사eae서렬여주요류행혈청형STEC적eae서렬완전일치。10주균분위7개ST형，여HUSEC、인원류행혈청군STEC균주구유상근적진화관계。결론아국비O157산지하독소대장애희균분리주eae기인존재일정정도적다양성，eae양성균주구유잠재적고치병성。
Objective To analyze the subtypes of eae genes in various non-O157 Shiga toxin-pro-ducing Escherichia coli ( STEC) strains isolated in China.Methods The complete nucleotide sequences of 10 eae genes were amplified by PCR and sequenced.The BLASTn software was used to analyze the se-quences for eae gene subtyping.A phylogenetic tree was constructed based on the10 ea e gene sequences to-gether with the gene sequences of 30 different subtypes in GenBank and those of STEC strains of 7 prevalent serotypes (O157 ∶H7, O26 ∶H11, O103 ∶H2, O111 ∶H8, O145 ∶H28, O45 ∶H2 and O121 ∶H19) using MEGA 5.0.Multilocus sequence typing (MLST) was performed on the 10 STEC strains with reference to the Escherichia coli ( E.coli) MLST website ( http://mlst.warwick.ac.uk/mlst/dbs/Ecoli) for the typing of multiple loci.A minimum spanning tree ( MST) was constructed using the BioNumerics software to inves-tigate the phylogenetic relationships between the 10 eae gene-positive STEC strains in this study and hemolyt-ic uremic syndrome-associated enterohemorrhagic E.coli ( HUSEC) strains as well as all human STEC strains of O157, O26, O45, O103, O111, O121 and O145 serotypes submitted to the E.coli MLST website data-base.Results The complete nucleotide sequences of eae genes in 10 non-O157 STEC strains were 2.8 kb in length and belonged to 3 known subtypes.The predominant subtype wasβ1, accounting for 60%of the 10 STEC strains (6/10), followed byθandγ1 subtypes with two strains in each type.The eae gene sequences in certain strains were identical to those of the prevalent serotypes.Seven sequence types ( STs) were identi-fied from the 10 STEC strains carrying eae gene.Conclusion The eae genes harbored by the non-O157 STEC strains isolated from different specimens in China were diverse and had close phylogenetic relationships with the highly pathogenic and prevalent STEC strains.This study implied that the STEC strains harboring eae gene had high pathogenic potential.