志贺菌CRISPR的检测及其与耐药的关系

Detection of CRISPR and its relationship to drug resistance in Shigella

摘要:

摘要:【目的】检测志贺菌成簇规律间隔的短回文重复序列(Clustered regularly interspaced short palindromic repeats，CRISPR)，并分析其与志贺菌耐药的关系。【方法】根据CRISPR DB数据库公布的志贺菌确定的CRISPR结构序列CRISPR-S2、CRISPR-S4和可能的CRISPR结构序列CRISPR-S1、CRISPR-S3设计四对引物，对60株志贺菌进行PCR扩增。采用CRISPR Finder分析CRISPR，采用改良K-B药敏纸片法检测志贺菌耐药情况，并分析CRISPＲ-S4 与耐药的关系。【结果】确定的CRISPR结构的总阳性率为95%，四个CRISPR位点组成12种CRISPR谱型(A-L)，除K型外均含确定的CRISPR结构，新发现1种重复序列和12种间隔序列。60株志贺菌的多重耐药率为53.33%。CRISPR-S4阳性菌株与阴性菌株之间，耐药的分布差异无统计学意义，但多重耐药菌株和耐TE菌株CRISPR-S4的重复序列多为R4.1，其3'末端缺失碱基AC;多重耐药菌株CRISPR-S4的间隔序列多为Sp5.1、Sp6.1和Sp7。【结论】CRISPR在志贺菌中广泛分布。CRISPR重复序列的变异和间隔序列的多样性可能与志贺菌耐药有关。

Abstract:

Abstract:［Objective］To detect clustered regularly interspaced short palindromic repeats (CRISPR) in Shigella，and to analyze its relationship to drug resistance．［Methods］ Four pairs of primers were used for the detection of convincing CRISPR structures CRISPR-S2 and CRISPR-S4，questionable CRISPR structures CRISPR-S1 and CRISPRＲ-S3 in 60 Shigella strains．All primers were designed using sequences in CRISPR database． CRISPR Finder was used to analyze CRISPＲ and susceptibilities of Shigella strains were tested by agar diffusion method. Furthermore，we analyzed the relationship between drug resistance and CRISPR-S4.［Results］The positive rate of convincing CRISPR structures was 95%．The four CRISPR loci formed 12 spectral patterns (A-L)，all of which contained convincing CRISPR structures except type K．We found one new repeat and 12 new spacers. The multi-drug resistance rate was 53.33%．We found no significant difference between CRISPR-S4 and drug resistant．However，the repeat sequence of CRISPR-S4 in multi-or TE-resistance strains was mainly R4.1 with AC deletions in the 3' end，and the spacer sequences of CＲISPＲ-S4 in multidrug resistance strains were mainly Sp5.1，Sp6.1 and Sp7．［Conclusion］CRISPR was common in Shigella．Variations of repeat sequences and diversities of spacer sequences might be related to drug resistance in Shigella.