采用高通量测序技术研究抗生素对小鼠肠道菌群的影响

Impact of antibiotics on the murine gut microbiota determined by high-throughput sequencing

目的探讨不同浓度抗生素对小鼠肠道菌群多样性和结构的影响,并预测相关功能变化。方法 15只SPF级ICR小鼠随机分为正常组、低浓度抗生素组和高浓度抗生素组,连续灌胃5 d后,采集小鼠新鲜粪便样本。利用Illumina MiSeq测序平台,对细菌的16S rRNA V3-V4区进行高通量测序,并对测序的结果进行生物信息学分析。结果高、低浓度抗生素组小鼠肠道菌群组成与正常组存在明显差异。与正常组相比,高剂量组小鼠肠道肠球菌属、志贺埃希菌属相对丰度显著升高(t=-2.71,P=0.026;t=-2.30,P<0.05);分节丝状菌属、拟普雷沃菌属相对丰度显著降低(t=2.88,P=0.020;t=2.49,P=0.037),理研菌属极显著降低(t=3.79,P=0.005)。低剂量组小鼠肠道菌群变形菌纲成为优势菌,芽胞杆菌属、粪球菌2、苏黎世杆菌属、普雷沃菌属2、普雷沃菌属7、志贺埃希菌属、沙雷菌属和放线菌属等相对丰度显著升高(均P<0.05);梭杆菌属、泛菌属极显著升高(t=-3.19,P=0.013;t=-3.50,P=0.008);分节丝状菌属、理研菌属相对丰度显著降低(t=2.69,P=0.028;t=2.33,P=0.048)。PICRUSt功能预测分析显示,抗生素组显著增加人类疾病、细胞过程和环境信息处理功能层的基因拷贝数,显著降低有机系统、遗传信息处理和代谢功能层的基因拷贝数。结论广谱抗生素能破坏小鼠肠道的微生态平衡,有必要深入研究抗生素对心血管、免疫性、感染性及神经退行性疾病发展的潜在作用。

Objective To investigate the impact of antibiotics at different concentrations on gut microbial composition in mice,and identify the related functional changes.Methods Fifteen SPF mice were randomly divided into control group,low-dose antibiotic group[Gentamycin sulfate 0.75 g/(kg·d)+Cefotaxime sodium 3.00 g/(kg·d)]or high-dose antibiotic group(Gentamycin sulfate 2.25 g/(kg·d)+Cefotaxime sodium 9.00 g/(kg·d))After 5 days of administration,15 fecal samples were collected and total DNA was extracted from the faecal samples.The primers were designed on bacterial 16 S rRNA V3-V4 region sequences and Illumina Miseq platform was used for high-throughput sequencing.The obtained data were analyzed using bioinformatics.Results Compared with the control group,the relative abundances of Enterococcus and EscherichiaShigella in high-dose antibiotic group significantly increased(all P<0.05),while those of CandidatusArthromitus,Alloprevotella and other genera significantly decreased(all P<0.05),and that of Alistpes extremely significantly decreased(P<0.01).Compared with the control group,Proteobacteria in the low-dose group became dominant;the relative abundances of Bacillus,Coprococcus2,Turicibacter,Prevotella2,Prevotella7,EscherichiaShigella,Serratia and Actinomyces significantly increased(all P<0.05),those of Fusobacterium and Pantoea extremely significantly increased(all P<0.01)while those of CandidatusArthromitus and Alistpes significantly decreased(all P<0.05).PICRUSt analysis showed that the metabolic pathways were potentially affected by antibiotics use.Conclusion Broad-spectrum antibiotics may disrupt gut microbiota,which warrants further studies on the potential role of antibiotics in the development of cardiovascular,immune system,infectious and neurodegenerative disorders.