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利福平耐药与敏感肺结核患者痰液样本微生物菌群多样性对比分析OA北大核心CSTPCD

Comparison of microbiota diversity in the sputum of pulmonary tuberculosis patients with rifampicin resistance or sensitivity

中文摘要英文摘要

目的:利用16S高通量测序技术,探索利福平耐药与利福平敏感肺结核患者呼吸道菌群的差异.方法:收集北京市疾病预防控制中心(70份)和湖南省胸科医院(10份)确诊的肺结核患者的痰液(共80份)进行前瞻性研究,通过痰液样本GeneXpert MTB/RIF检测、靶向rpoB高通量测序(tNGS测序)和利福平药物敏感性试验,将80例患者分为利福平耐药组(RFP-R组,41例;利福平耐药决定区突变)和利福平敏感组(RFP-S组,39例;利福平耐药决定区无突变).对痰液微生物16S V4区域进行高通量测序,测序结果进行生物信息学分析及统计分析.结果:RFP-R组和RFP-S组共获得41 950 224个原始16S rRNA序列.RFP-R组的平均操作分类单元(operational taxonomic units,OTU)为 344 个,RFP-S 组的平均 OTU 为 342 个.厚壁菌门(47.72%)、拟杆菌门(22.36%)、变形菌门(14.67%)、放线菌门(7.57%)、梭杆菌门(6.46%)是肺结核患者痰液中检出的主要细菌门,占微生物群落的98.78%.与RFP-S组相比,RFP-R组梭杆菌门(7.531%vs.5.167%;t=2.218,P=0.030)和螺旋体门(0.596%vs.0.246%;t=2.128,P=0.038)相对丰度显著增加.与RFP-S组相比,RFP-R组颤杆菌克属(0.690%vs.0.326%;t=3.112,P=0.003)、产线菌属(0.135%vs.0.030%;t=2.171,P=0.035)、Allisonella菌属(0.046%vs.0.006%;t=3.237,P=0.002)、假单胞菌属(0.205%vs.0.054%;t=2.040,P=0.046)和密螺旋体属(0.601%vs.0.242%;t=2.155,P=0.036)相对丰度显著增加;二氧化碳噬纤维菌属(0.351%vs.0.772%;t=-2.044,P=0.046)和 Pauljensenia 菌属(2.314%vs.3.706%;t=-2.660,P=0.010)相对丰度显著减少.结论:利福平耐药与敏感结核分枝杆菌感染不影响肺结核患者呼吸道菌群的整体结构,但利福平耐药与敏感患者在2个门(梭杆菌门、螺旋体门)和7个属(颤杆菌克属、产线菌属、Allisonella菌属、假单胞菌属、密螺旋体属、二氧化碳噬纤维菌属、Pauljensenia菌属)的丰度中存在差异,其中二氧化碳噬纤维菌属和Pauljensenia菌属在利福平敏感患者中丰度较高,其余在利福平耐药患者中丰度较高.

Objective:To explore the structural differences in the respiratory tract flora community in pulmonary tuberculosis patients with rifampicin(RFP)resistance or sensitivity using 16S high-throughput sequencing technology.Methods:Sputum samples(80 patients)from Beij ing Center for Disease Control and Prevention(70 patients)and Hunan Chest Hospital(10 patients)were included in this prospective study.Using Xpert assay,high-throughput targeted rpoB sequencing and RFP susceptibility assay on these sputum samples,the 80 patients were divided into RFP-resistant group(RFP-R,with RFP resistance based on the RFP-resistance determining region(RRDR)mutation;41 patients)and RFP-sensitive group(RFP-S,no RRDR mutation;39 patients).High-throughput sequencing of the 16S V4 region,bioinformatics analysis,and statistical analysis of the sequencing results were performed.Results:In total,41 950 224 original 16S rRNA reads were obtained from the RFP-R and RFP-S groups,with 344 and 342 average operational classification units(OTUs)for the RFP-R group and RFP-S group,respectively.Firmicutes(47.72%),Bacteroidota(22.36%),Proteobacteria(14.67%),Actinobacteriota(7.57%),and Fusobacteriota(6.46%)were the main bacterial phyla detected,accounting for 98.78%of the microbial community.Compared with the RFP-S group,the relative abundances of the Fusobacteriota(7.531%vs.5.167%;t=2.218,P=0.030)and Spirochaeta(0.596%vs.0.246%;t=2.128,P=0.038)in the RFP-R group were significantly higher,the same as for Oribacterium(0.690%vs.0.326%;t=3.112,P=0.003),Filifactor(0.135%vs.0.030%;t=2.171,P=0.035),Allisonella(0.046%vs.0.006%;t=3.237,P=0.002),Pseudomonas(0.205%vs.0.054%;t=2.040,P=0.046),and Treponema(0.601%vs.0.242%;t=2.155,P=0.036),whereas Capnocytophaga(0.351%vs.0.772%;t=-2.044,P=0.046)and Pauljensenia(2.314%vs.3.706%;t=-2.660,P=0.010)in the RFP-R group showed decreased relative abundances compared with those in the RFP-S group.Conclusion:RFP resistant and sensitive Mycobacterium tuberculosis infection did not affect the overall structure of the respiratory tract flora in pulmonary tuberculosis patients.However,significant differences were observed in the relative abundance of two Phyla(Fusobacteriota,Spirochaeta)and seven Genera(Oribacterium,Filifactor,Allisonella,Pseudomonas,Treponemas,Capnocytophaga,Pauljensenia)between the RFP-resistant and sensitive groups.Specifically,Capnocytophaga and Pauljensenia exhibited higher abundance in the RFP-sensitive group while the remaining taxa showed higher abundance in the RFP-resistant group.

张泓泰;任怡宣;胡培磊;王嫩寒;李洁;田丽丽;赵琰枫;陈双双;李传友

北京市疾病预防控制中心结核病实验室,北京 100035湖南省胸科医院检验科,长沙 410000

临床医学

结核,肺利福平抗药性生物多样性细菌

Tuberculosis,pulmonarySputumRifampinDrug resistanceBiodiversityBacteria

《中国防痨杂志》 2024 (006)

625-633 / 9

北京市自然科学基金(7242078);首都卫生发展科研专项(2024-2G-3017;2022-1G-3012) National Natural Science Foundation of Beijing(7242078);Capital's Funds for Health Improvement and Research(2024-2G-3017;2022-1G-3012)

10.19982/j.issn.1000-6621.20240047

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