岩藻多糖对幽门螺杆菌感染后小鼠肠道菌群紊乱的调节作用

REGULATORY EFFECT OF FUCOIDAN ON INTESTINAL FLORA DISTURBANCE IN MICE AFTER HELICOBACTER PYLORI INFECTION

目的探讨岩藻多糖对幽门螺杆菌(HP)感染后小鼠肠道菌群紊乱的调节作用。方法将22只8周龄C57BL/6品系小鼠,随机分为空白对照组(A组):小鼠每天灌服双蒸水,持续5周;单纯感染组(B组):小鼠每天灌服HP菌悬液,持续3周,再灌服2周双蒸水;单纯干预组(C组):单纯岩藻多糖干预2周,再灌服双蒸水3周;预防干预组(D组):岩藻多糖预干预2周,再感染HP 3周;治疗干预组(E组):感染HP 3周后,岩藻多糖干预2周。采集实验结束当天小鼠的粪便样品进行基因测序分析,采用Illumina HiSeq测序平台进行OTU(operational taxonomic units)聚类与注释;利用QIIME软件生成不同分类水平上的物种丰度表,再利用R语言工具绘制成样品各分类学水平下的物种分布柱状图;使用QIIME软件,通过NMDS分析对各组肠道菌群的β多样性进行分析;通过LEfSe技术分析组间差异显著性,寻找组间具有统计学差异的物种;最后通过Metastats分析,根据P值筛选出导致两组样品组成差异的肠道菌群。结果α多样性分析结果中的Shannon指数显示,E组生物多样性较B组升高;β多样性的NMDS分析结果显示E组菌群结构近似A组;粪便菌群LEfSe分析显示共有67个菌属在A、B、C、D和E组间差异具有显著意义(P<0.05,LDA值>2);进一步的Metastats分析显示岩藻多糖干预减轻了HP感染后有害细菌如脱硫弧菌(Desulfovibrio)以及泽泻属(Alistipe)的相对丰度,增加了肠道有益菌如双歧杆菌(Bifidobacterium)、粪便杆菌(Faecalibacterium)的相对丰度。结论岩藻多糖可提高肠道部分有益菌的相对丰度,降低部分有害细菌的相对丰度,并可增加肠道微生物多样性,对HP感染后小鼠肠道菌群紊乱具有调节作用。

Objective To investigate the regulatory effect of fucoidan on intestinal flora disturbance in mice after Helicobacter pylori(HP) infection. Methods A total of 22 C57BL/6 mice, aged 8 weeks, were randomly divided into blank control group(group A, treated with double-distilled water daily by gavage for 5 consecutive weeks), infection group(group B, treated with HP bacterial suspension daily by gavage for 3 consecutive weeks, followed by double-distilled water by gavage for 2 weeks), intervention group(group C, intervention with fucoidan alone for 2 weeks, followed by double-distilled water by gavage for 3 weeks), prevention+intervention group(group D, intervention with fucoidan for 2 weeks, followed by HP infection for 3 weeks), and treatment+intervention group(group E, HP infection for 3 weeks, followed by intervention with fucoidan for 2 weeks). Fecal samples were collected for gene sequencing analysis on the day of finishing the experiment;the Illumina HiSeq sequencing platform was used to perform clustering and annotation of operational taxonomic units(OTUs);QIIME software was used to generate species abundance tables at different taxonomic levels, and R language was used to plot species distribution histograms at each taxonomic level;NMDS analysis was performed using QIIME software to analyze the beta-diversity of intestinal flora;the LEfSe technique was used to analyze significant differences between groups and identify the species with significant difference;finally Me-tastats analysis was performed, and P value was used to screen out the intestinal flora leading to the difference in composition between the two groups of samples. Results Shannon index in alpha-diversity analysis showed that group E had higher biodiversity than group B, and the beta-diversity analysis based on NMDS showed that group E had a similar composition of intestinal flora to group A. LEfSe analysis of fecal flora showed that there were significant differences in 67 bacterial genera between groups A, B, C, D, and E(P<0.05, LDA>2). Further Metastats analysis showed that fucoidan intervention reduced the relative abundance of harmful bacteria(Desulfovibrio and Alistipe) and increased the relative abundance of probiotic bacteria(Bifidobacterium and Faecalibacterium) after HP infection. Conclusion Fucoidan can increase the relative abundance of several probiotic bacteria and reduce the relative abundance of harmful bacteria in the intestinal tract, and meanwhile, it can increase the diversity of intestinal flora. Therefore, it can exert a regulatory effect on intestinal flora distur-bance in mice after HP infection.