广东省2株白喉棒状杆菌的鉴定与分析

Identification and analysis of 2 Corynebacterium diphtheria strains in Guangdong Province

目的对广东省2株C.diphtheriae白喉棒状杆菌进行精准鉴定与分析。方法通过API Coryne棒状杆菌生化卡对2010年广州和2020年珠海分离的C.diphtheriae进行生化及质谱鉴定;利用Illumina平台进行测序,CLC软件拼接,JSpeciesWS线上工具进行平均核苷酸同源性识别;BLASTN进行narKGHIJ和tox基因检测,使用MEGA-X构建wgSNP系统发育进化树。结果GD-Guangzhou-2010为Belfanti型,GD-Zhuhai-2020为Gravis型。棒状杆菌种间ANI值计算结果显示,GD-Guangzhou-2010与C.belfantii间ANIb=99.61%,GD-Zhuhai-2020与C.diphtheriae间ANIb=97.64%。BLASTN结果显示,GD-Guangzhou-2010硝酸盐还原基因narKGHIJ均为阴性且未携带tox基因,GD-Zhuhai-2020硝酸盐还原基因narKGHIJ均为阳性,未携带tox基因。C.diphtheriae的wgSNP进化树显示两个主要进化分支,第一个分支包含所有Mitis型和Gravis型以及GD-Zhuhai-2020,第二个分支包含所有的C.belfantii、C.diphtheriae subsp.lausannense分离株及GD-Guangzhou-2010。结论广东省成功分离鉴定出两株非产毒性C.diphtheriae,系统发生树提示GD-Guangzhou-2010和GD-Zhuhai-2020位于两个不同的进化分支。

Objective To identify and analyze two strains of C.diphtheriae in Guangdong Province by combining whole genome sequencing with traditional detection methods.Methods The C.diphtheriae was isolated from Guangzhou in 2010 and Zhuhai in 2020 respectively.Isolates were identified by API Coryne strips and MALDI-TOF-MS.Genomic DNA was sequenced by using Illumina.The assembly was performed for each strain using CLC software.J Species WS online tool was used for average nucleoside homology identification,then narKGHIJ and tox gene were detected by NCBI online analysis tool BLSATN.MEGA-X was used to build a wgSNP phylogenetic tree.Results GD-Guangzhou-2010 was Belfanti and GD-Zuhai-2020 was Gravis.ANIb between GD-Guangzhou-2010 and C.belfantii was 99.61%.ANI between GD-Zhuhai-2020 and C.diphtheriae was 97.64%.BLASTN results showed that the nitrate reduction gene narKGHIJ and tox gene of GD-Guangzhou-2010 was negative,while GD-Zhuhai-2020 nitrate reduction gene narKGHIJ was positive.There were two obvious clades in wgSNP phylogenetic tree.The first clades included all Mitis and Gravis types strains as well as GD-Zhuhai-2020.The second clades contained all isolates of C.belfantii,C.diphtheriae subsp.lausannense and GD-guangzhou-2010.Conclusion Two non-toxic C.diphtheriae strains are successfully isolated and identified.The phylogenetic tree suggests that GD-Guangzhou-2010 and GD-Zhuhai-2020 are located in two different evolutionary branches.