食源性单核细胞增生李斯特菌CRISPR结构的研究

Analysis of CRISPR Regions in Food-borne Listeria monocytogenes

摘要:

为探明河北省2177份零售食品样本中检出的18株谱系I（血清型1/2b，4b）菌株的CRISPR序列结构，用PCR方法扩增了其CRISPR序列，采用生物信息学方法进行序列同源性分析，并根据在18菌株中CRISPR结构的阵列排布进行聚类分型研究。结果表明：18株菌共检出三类CRISPR序列排布（CRSIPR array：locus1，2，3）。CRISPR locus1是一退化残基，repeat序列保守性差，spacer数量少。locus2和locus3是新掺入的，结构完整，活力旺盛，repeats序列保守，spacer数量增长迅速。locus1最普遍（13/18）；有5株serotype1/2b的菌株检出至少两个活性loci（locus2和locus3）中的一个，在serotype 4b菌株中未检出。总计，基因组中具有3个CRISPR loci的有2株，具有2个的有3株，仅有1个的有8株，无该结构的有5株。根据CRISPR array的排布共将18株菌聚类分型为5簇，4个亚簇，能较好的对1/2b血清型的菌株进行分型研究。该免疫防御系统的插入，增强了高致病性谱系I菌株对环境的适应能力，增大了消除食品加工销售过程中该类菌株的难度，给食品安全造成更大威胁，因此相关部门应加强对该特征菌株的监管力度。

Abstract:

In order to find out CRISPRs in 18 lineage I (serotype1/2b, 4b) strains of food-borne Listeria monocytogens isolated from 2177 retail food samples in Hebei province, the CRISPR sequences were obtained by PCR amplification, and homology analysis was predicted using bioinformatic methods, then cluster typed by using a CRISPR-based approach. We detected three CRISPR loci in all studied strains. Loucus1 was most popular (13/18); locus2 and locus3, as two activity loci , were only detected at least one in 5 serotype 1/2b strains. Locus1, a putative remnant of a functional CRISPR ancestor, was beared unconserved repeats (DR1) and a few of spacers; locus2 and locus3, relative new functional structures, were beared conserved repeats (DR2, DR3) and fast growing number of spacers. In all, 2 strains containerd all three loci, 3 strains contained the first 2 of the three loci, as well as 8 strains only contained the first one locus, and 5 strains did not find any CRISPR structure. As the diversity of CRISPR arrays, 18 strains typed to 5 clusters (A~F), and 4 subset clusters (A1and A2, B1 and B2). CRISPR typing can be good used in differentiating serotype 1/2b strains. In other hands, the CRISPR structures, especially that beared more spacers, enhance the host's environmental adaptability, enlarged the clear difficulty in food processing, alarm the food safety. The government should reinforce the control of these kinds of L. monocytogenes.