患者血液和温泉水中阿尔卑斯浴者菌的分离及鉴定

Isolation and identification of Balneatrix alpica from patient's blood and hot spring water

目的:通过对患者血液来源的阿尔卑斯浴者菌( Balneatrix alpica)分离株X117和该患者所在小区天然温泉水的分离株GN-1进行菌种鉴定与微生物学特征分析,为该菌作为罕见病原体引起临床相关感染的病原学诊断提供实验依据。 方法:以阿尔卑斯浴者菌DSM 16621 T作为参考,对分离株X117和GN-1进行生化表型鉴定、基质辅助激光解吸电离飞行时间质谱鉴定、16S rRNA基因测序和系统发育分析、单核苷酸多态性(SNP)距离以及全基因组相关指数分析,以准确确定其分类学地位和同源性;微量肉汤稀释法进行药敏试验;VFDB毒力因子数据库进行毒力基因注释和分析。 结果:分离株X117和GN-1在哥伦比亚血琼脂平板和巧克力平板上培养4 d可见菌落呈淡黄色或棕黄色,菌落表面呈斑驳状,革兰染色阴性杆菌,氧化酶和吲哚试验阳性,与阿尔卑斯浴者菌DSM 16621 T特征一致。16S rRNA系统发育分析表明分离株X117和GN-1均为阿尔卑斯浴者菌,其与阿尔卑斯浴者菌DSM 16621 T平均核苷酸一致性(ANI)值为98.44%和98.41%,基因组DNA-DNA杂交值(dDDH)均为87.1%。SNP距离差异为13,显示X117和GN-1可能为同一克隆株。药敏试验显示阿尔卑斯浴者菌对临床常用的抗革兰阴性杆菌药物均敏感。3株阿尔卑斯浴者菌毒力相关基因主要涉及黏附、侵袭、鞭毛及生物膜的形成等方面。 结论:本研究确认了一起与天然温泉水密切相关的阿尔卑斯浴者菌引起的血流感染,天然温泉水可能是阿尔卑斯浴者菌临床感染的重要来源。

Objective To identify and characterize two Balneatrix alpica strains isolated from a patient's blood sample(strain X117)and the natural hot spring water in the patient's residential district(strain GN-1),and to provide experimental evidence for the pathogenic diagnosis of clinical infection caused by this rare pathogen.Methods Biochemical phenotypic identification,matrix-assisted laser desorption ionization time-of-flight mass spectrometry(MALDI-TOF MS),16S rRNA gene sequencing,phylogenetic analysis,single-nucleotide polymorphism(SNP)analysis,and genome-wide analysis were performed to accurately determine the taxonomic status of the isolates X117 and GN-1 by using Balneatrix alpica DSM 16621'as a reference.Microdilution broth method was used to test their antimicrobial susceptibility.The virulence genes carried by them were annotated and analyzed using the virulence factor database(VFDB).Results Strains X117 and GN-1 formed light yellow or tan colonies with mottled surfaces on Columbia blood agar and chocolate agar plates after 4 d of culture.They were Gram-negative rods and positive for oxidase and indole tests,which were consistent with the characteristics of Balneatrix alpica DSM 16621T.The phylogenetic analysis based on the 16S rRNA gene showed that the isolates X1i7 and GN-1 were both Balneatrix alpaca.The average nucleotide identity(ANI)values between the two isolates and Balneatrix alpica DSM 16621T were 98.44%and 98.41%,respectively,and the digital DNA-DNA hybridization(dDDH)values were both 87.1%.The SNP distance between the two strains was 13,indicating that X117 and GN-1 might belong to the same clone.The antibiotic susceptibility testing showed that all of the three Balneatrix alpica strains were sensitive to the commonly used antibiotics against Gram-negative rods.The virulence genes carried by the three Balneatrix alpica strains were mainly involved in adhesion,invasion,flagella and biofilm formation.Conclusions This study identified a case of bloodstream infection caused by Balneatrix alpica which was closely related to natural hot spring water.Natural hot spring water migh be an important source of clinical infections caused by this species.