CAJ | 학술논문

目的研究肺炎克雷伯临床耐药菌株新基因组岛KpGI-2的结构和功能.方法采用PCR方法自一株肺炎克雷伯耐药菌株HS04160扩增得到新的插入序列KpGI-2,通过测序以及生物信息学方法分析其序列结构并推测其生物学功能,采用细菌生长试验证实其生物学功能.结果 phe55 tRNA基因位点处PCR扩增得到的6.4 kb的插入序列KpGI-2,序列分析发现其GC含量为38.03%,明显低于肺炎克雷伯菌株基因组平均的GC含量(57.5%).KpGI-2中含有163 bp的重复序列,可以确定为一个新的基因组岛.KpGI-2携带有5个orf,其中orf5与沙门菌属中的Fic(fllamentation induced by cAMP)蛋白具有高度相似性.肺炎克雷伯临床菌株普遍携带有一个高度保守的fic基因以及一个相对保守的fic基因,携带有KpGI-2的临床菌株HS04160丢失了相对保守的fic基因.通过细菌生长试验分析发现orf5以及KpGI-2在cAMP诱导下对细胞生长具有调节作用,KPGI-2的功能可能与细菌生长的调节相关.结论 KpGI-2是位于肺炎克雷伯临床菌株基因组岛插入热点phe55 tRNA基因位点上的一个新基因组岛,其携带的基因与细胞生长相关.
목적 연구폐염극뢰백림상내약균주신기인조도KpGI-2적결구화공능.방법 채용PCR방법자일주폐염극뢰백내약균주HS04160확증득도신적삽입서렬KpGI-2,통과측서이급생물신식학방법분석기서렬결구병추측기생물학공능,채용세균생장시험증실기생물학공능.결과 phe55 tRNA기인위점처PCR확증득도적6.4 kb적삽입서렬KpGI-2,서렬분석발현기GC함량위38.03%,명현저우폐염극뢰백균주기인조평균적GC함량(57.5%).KpGI-2중함유163 bp적중복서렬,가이학정위일개신적기인조도.KpGI-2휴대유5개orf,기중orf5여사문균속중적Fic(fllamentation induced by cAMP)단백구유고도상사성.폐염극뢰백림상균주보편휴대유일개고도보수적fic기인이급일개상대보수적fic기인,휴대유KpGI-2적림상균주HS04160주실료상대보수적fic기인.통과세균생장시험분석발현orf5이급KpGI-2재cAMP유도하대세포생장구유조절작용,KPGI-2적공능가능여세균생장적조절상관.결론 KpGI-2시위우폐염극뢰백림상균주기인조도삽입열점phe55 tRNA기인위점상적일개신기인조도,기휴대적기인여세포생장상관.
Objective To investigate the construction and function of a hove island KpGI-2 from Klebsiella pneumoniae clinical isolate HS04160.Methods Based on the bioinformatics analysis,PCR assay and viable cell count experiment were employed to investigate the insertion sequence at the possible island insertion hotspet phe55 Trna loeus.Results A 6.4 kb insertion sequence KpGI-2 exhibited difierent GC content with the core sequence of K.pneumoniae strains and took the direct sequences in it.These showed that it was a novel genomic island in K.pneumoniae clinical isolate.There were five orf in KpGI-2.Most of them showed low similarity with the known protein except ORF5 exhibited high similarity with the Fic protein in Salmonella enteria strain.Fic protein was involved in the cell growth regulation induced by cAMP.There ale two fic gene kpn_03747 and kpn_03553 in K.pneumoniae clinical isolates.One clinical isolate HS04160 lost the fic gene kpn_03553 but harbored the KpGI-2 which carried a Fic-like gene.At another hand,ORF5 in KpGI-2 showed high similarity with the Fic protein in S.enteria which was also located in a 14.7 kb genomic island after pheV tRNA locus.It strongly substantiated the idea of a cross-genus acquisition of KpGI-2 and its fic gene.Viable cell count experiment showed that the orfs including orf5 in KpGI-2 affected the cell growth after cAMP incubation.orf2-3 decreased the cell growth whereas orf4 or orf5 increased cell growth after 5 mmol/L cAMP incubation,which hinted that KpGI-2 island might be involved in the cell growth regulation.Conclusion Based on the bioinformatics analysis and the molecular technology analysis,KpGI-2 was indicated to be a novel genomic island at the hotspot phe55 tRNA locus in K.pneumoniae clinical isolates and related to cell growth.