应用Illumina NovaSeq测序技术对骨癌患犬肠道菌群的分析

Analyzing the gut microbiome of canine with osteocarcinoma using the high-throughput sequencing technology Illumina Nova

通过分析骨癌患犬与健康犬肠道菌群组成与基因功能预测,筛选出作为犬骨癌评价依据的细菌,为下一步犬益生菌制剂的开发提供理论依据。本研究采集骨癌患犬与健康犬的新鲜粪便并提取其DNA进行Illumina NovaSeq高通量测序。结果显示:2组犬肠道菌群的Shannon、Simpson、Chao1指数无显著差异(P>0.05),骨癌组犬的ACE和Observed_species指数显著低于健康组(P<0.05)。在门水平上,犬骨癌组肠道中梭杆菌门(Fusobacteriota)和拟杆菌门(Bateroidota)的平均相对丰度显著高于健康组(P<0.05),而变形菌门(Proteobacteria)显著低于健康组(P<0.05);在属水平上,犬骨癌组肠道中梭杆菌属(Fusobacterium)和拟杆菌属(Bacteroides)的平均相对丰度显著高于健康组(P<0.05),而布劳特氏菌属(Blautia)、乳杆菌属(Lactobacillus)、链球菌属(Streptococcus)和Turicibacter属显著低于健康组(P<0.05)。PCoA分析表明2组的肠道菌群存在显著差异(P<0.05)。LDA差异贡献分析发现骨癌组的丰度增加并具有统计学差异(P<0.05)的物种有13个,且造成显著影响力最大的5个物种正好位于同一个进化树上。Tax4Fun功能预测分析显示,骨癌组的碳水化合物代谢、能量代谢、聚糖生物合成与代谢、辅助因子和维生素的代谢通路显著高于健康组(P<0.05),但膜转运、信号转导、细胞群落-原核生物、转录显著低于健康组(P<0.05)。本研究结果丰富了对骨癌患犬肠道菌群变化的认识,肠道优势菌(梭杆菌属和拟杆菌属)有可能作为一种生物标志物用于犬骨癌的早期诊断。

This experiment was conducted to compare and analyze the gut microflora composition and gene function prediction of canines with osteocarcinoma,and to screen out the bacteria as the basis for osteocarcinoma evaluation,so as to provide theoretical basis for development of probiotics for canines in the future.The fresh feces were collected from canines infected with osteocarcinoma and from healthy canines,and the DNA was extracted.The composition and function of the gut microbiome of the canines were analyzed using the high-throughput sequencing technology Illumina Nova.The results showed that the flora of the two groups had no significant difference in the Shannon,Simpson,Chao1 indexes(P>0.05),but the ACE index and observed species index were significantly lower in the osteocarcinomainflicted group than those in the healthy group(P<0.05).At the phylum level,the relative abundance of Fusobacteriota and Bateroidota in the osteocarcinomainfected group were higher than that in healthy group,and the relative abundance of Proteobacteria was lower than that in healthy group(P<0.05).At the genus level,the relative abundances of Fusobacterium and Bacteroides in the osteocarcinoma infected group were higher than those in the healthy group,the relative abundances of Blautia,Lactobacillus,Streptococcus and Turicibacter were lower in osteocarcinoma infected group than those in the healthy group(P<0.05).PCoA showed that there were significant different in the gut microbiome between the two groups of canines.The LDA different contribution analysis showed that 15 species were significantly higher in the osteocarcinoma-infected group than those in the healthy group,and the five most important species were just located in one evolutionary tree.In terms of the function prediction,the main contents of carbohydrate metabolism,energy metabolism,glycan biosynthesis and metabolism,and metabolism of cofactors and vitamins in the osteocarcinoma-infected group were significantly higher those in the healthy group(P<0.05);however,the main contents of membrane transport,signal transduction,cellular community-prokaryotes and transcription were significantly lower in the osteocarcinoma-infected group than that in the healthy group(P<0.05).In summary,these results enriched our knowledge of the change in the gut microbiome in canines with osteocarcinoma.As the dominant microbiomes,Fusobacterium and Bacteroides might be used as biomarkers for early diagnosis of primary osteocarcinoma in canines.