中国产志贺毒素大肠杆菌(STEC)O157:H7的单核苷酸多态性（SNP）分型研究
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摘要
产志贺毒素大肠杆菌(Shiga toxin-producing Escherichia coli, STEC)O157:H7是一类重要的人畜共患病病原,可引起人水样腹泻、出血性肠炎以及溶血性尿毒综合征等。单核苷酸多态性(Single nucleotide polymorphisms, SNPs)分析是近年来用于揭示0157：H7群体基因组水平多样性、流行病学关系以及详细了解0157：H7演化过程的一种基因分型方法。我们对于中国0157：H7菌株之间以及中国菌株与其他国家菌株之间的遗传关系所知甚少。
本研究采用了Manning等人以及Clawson等人建立的SNPs分型体系,共计64个SNP位点,将中国325株STECO157:H7菌株分为5个SNP基因型(SG-1～SG-5)。最常见的基因型为SG-5型(79.69%)及SG-1型(14.46%)。从患者分离的菌株集中在SG-1型及SG-5型,仅一株存在于SG-3型中。47株SG-1型菌株,通过Xuzhou21基因组特异SNPs位点可分为两个亚型(SG-1.1和SG-1.2)。SG-1.1型菌株是导致1999苏皖疫情相关菌株。1999年HUS/HC从患者分离的菌株集中在SG-1.1。
根据MLST分型结果,136株中国STEC O157:H7菌株分为4个ST型别：ST23、ST24、 ST96和ST97。 MLST分型结果与SNP分型结果有较好的一致性且SNP分型能力较强。ST24型菌株包括SG-2及SG-3两个基因型,同时,ST23型菌株包括SG-4和SG-5基因型。ST96和ST97属于相同的基因型SG-1，SG-1.1亚型菌株为ST96型,SG-1.2亚型菌株为ST97型。
中国STEC O157:H7菌株有5个基因型,且在Manning等人划分的Clade以及Clawson等人区分的分支中,均显示较远的遗传关系,表明这些基因型菌株在我国是平行传播的。
我们对5个SG型别的36株菌进行重测序,做基因组间SNP分析,结果显示,36株0157：H7菌株共计有2486个SNPs,其中包括同义突变位点809个和非同义突变位点1195个,基因间的SNP位点475个,终止突变位点7个。基于全基因组SNP位点的分型结果,SG-5菌株呈分散状态。另外,SG-2、 SG-3的3株菌为ST24型菌株,呈现出较分散的3个分支,提示SG-2、 SG-3存在较大差异。可通过这些SNPs位点区分出SG-1型菌株中和1999年苏皖疫情相关的SG-1.1型菌株。
Shiga toxin-producing Escherichia coli O157:H7is a well-known zoonotic pathogen as a cause of diarrhea, hemorrhagic colitis (HC), and hemolytic uremic syndrome (HUS). Single nucleotide polymorphisms (SNPs) have been widely used to determine genetic relatedness, explore the population and evolution and epidemiological relationship of O157:H7. There are few reports of genetic diversity of Chinese O157:H7isolates and their relationships with global isolates.
The minimum sets of32SNPs each from Manning et al. and Clawson et al. were used to type325Chinese O157:H7isolates. The total64SNPs divided the Chinese O157:H7isolates into5SNP genotypes (SG-1to SG-5). The most common SGs were SG-5(79.69%) and SG-1(14.46%). Human isolates concentrated in SG-1and SG-5, and there is only1human isolates in SG-3. The47isolates in SG-1were further divided into2subtypes (SG-1.1and SG-1.2) by an additional SNP sourced from Xuzhou21genome. Strains in SG-1.1caused the1999Xuzhou deadly outbreak.
Of325Chinese isotates,136isolates were typed by MLST into4types:ST23, ST24, ST96and ST97. MLST results are consistent with SNP typing with the latter offering slightly higher resolution. ST24is divided into SG-2and SG-3while ST23is divided into SG-4and SG-5. However ST96and ST97belonged to the same SG (SG-1). All SG-1.1isolates are ST96while all SG-1.2isolates belong to ST97.
Chinese isolates have been found to belong to a limited number of SNP genotypes and are represented by distantly related clades in Manning et al. and lineages in Claswon et al., suggesting parallel spread of these SNP genotypes in China.
Thirty-six isolates representing different SGs were included for sequencing and comparative analysis. All the isolates has a total of2486SNPs, including809synonymous mutations and1195non-synonymous mutations,475inter-gene SNPs,7stop mutations.Phylogenetic tree based on SNP concatenation could separate all SG-5strains suggesting the high resolution of SNPs of Chinese O157:H7isolates to capture the genetic diversity of Chinese O157:H7isolates, In addition,3ST24isolates of SG-2and SG-3were divided into the three branches, revealing substantial genomic differences. The outbreak related strains in SG-1.lcan be separated from SG-1using these SNPs.
本研究采用了Manning等人以及Clawson等人建立的SNPs分型体系,共计64个SNP位点,将中国325株STECO157:H7菌株分为5个SNP基因型(SG-1～SG-5)。最常见的基因型为SG-5型(79.69%)及SG-1型(14.46%)。从患者分离的菌株集中在SG-1型及SG-5型,仅一株存在于SG-3型中。47株SG-1型菌株,通过Xuzhou21基因组特异SNPs位点可分为两个亚型(SG-1.1和SG-1.2)。SG-1.1型菌株是导致1999苏皖疫情相关菌株。1999年HUS/HC从患者分离的菌株集中在SG-1.1。
根据MLST分型结果,136株中国STEC O157:H7菌株分为4个ST型别：ST23、ST24、 ST96和ST97。 MLST分型结果与SNP分型结果有较好的一致性且SNP分型能力较强。ST24型菌株包括SG-2及SG-3两个基因型,同时,ST23型菌株包括SG-4和SG-5基因型。ST96和ST97属于相同的基因型SG-1，SG-1.1亚型菌株为ST96型,SG-1.2亚型菌株为ST97型。
中国STEC O157:H7菌株有5个基因型,且在Manning等人划分的Clade以及Clawson等人区分的分支中,均显示较远的遗传关系,表明这些基因型菌株在我国是平行传播的。
我们对5个SG型别的36株菌进行重测序,做基因组间SNP分析,结果显示,36株0157：H7菌株共计有2486个SNPs,其中包括同义突变位点809个和非同义突变位点1195个,基因间的SNP位点475个,终止突变位点7个。基于全基因组SNP位点的分型结果,SG-5菌株呈分散状态。另外,SG-2、 SG-3的3株菌为ST24型菌株,呈现出较分散的3个分支,提示SG-2、 SG-3存在较大差异。可通过这些SNPs位点区分出SG-1型菌株中和1999年苏皖疫情相关的SG-1.1型菌株。
Shiga toxin-producing Escherichia coli O157:H7is a well-known zoonotic pathogen as a cause of diarrhea, hemorrhagic colitis (HC), and hemolytic uremic syndrome (HUS). Single nucleotide polymorphisms (SNPs) have been widely used to determine genetic relatedness, explore the population and evolution and epidemiological relationship of O157:H7. There are few reports of genetic diversity of Chinese O157:H7isolates and their relationships with global isolates.
The minimum sets of32SNPs each from Manning et al. and Clawson et al. were used to type325Chinese O157:H7isolates. The total64SNPs divided the Chinese O157:H7isolates into5SNP genotypes (SG-1to SG-5). The most common SGs were SG-5(79.69%) and SG-1(14.46%). Human isolates concentrated in SG-1and SG-5, and there is only1human isolates in SG-3. The47isolates in SG-1were further divided into2subtypes (SG-1.1and SG-1.2) by an additional SNP sourced from Xuzhou21genome. Strains in SG-1.1caused the1999Xuzhou deadly outbreak.
Of325Chinese isotates,136isolates were typed by MLST into4types:ST23, ST24, ST96and ST97. MLST results are consistent with SNP typing with the latter offering slightly higher resolution. ST24is divided into SG-2and SG-3while ST23is divided into SG-4and SG-5. However ST96and ST97belonged to the same SG (SG-1). All SG-1.1isolates are ST96while all SG-1.2isolates belong to ST97.
Chinese isolates have been found to belong to a limited number of SNP genotypes and are represented by distantly related clades in Manning et al. and lineages in Claswon et al., suggesting parallel spread of these SNP genotypes in China.
Thirty-six isolates representing different SGs were included for sequencing and comparative analysis. All the isolates has a total of2486SNPs, including809synonymous mutations and1195non-synonymous mutations,475inter-gene SNPs,7stop mutations.Phylogenetic tree based on SNP concatenation could separate all SG-5strains suggesting the high resolution of SNPs of Chinese O157:H7isolates to capture the genetic diversity of Chinese O157:H7isolates, In addition,3ST24isolates of SG-2and SG-3were divided into the three branches, revealing substantial genomic differences. The outbreak related strains in SG-1.lcan be separated from SG-1using these SNPs.
引文
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