捻转血矛线虫伊维菌素敏感虫株与耐药虫株差异miRNA的转录组学分析  

Transcriptomic analysis of miRNAs between ivermectin sensitive and resistant strains of Haemonchus contortus

作　　者：温海峰 张艳敏 张海龙 石雅琴 毛晓伟 寇慧琳 于宇 李峥 陈昕迪 王腾宇 高娃 王文龙 刘春霞[4] WEN Hai-feng;ZHANG Yan-min;ZHANG Hai-long;SHI Ya-qin;MAO Xiao-wei;KOU Hui-lin;YU Yu;LI Zheng;CHEN Xin-di;WANG Teng-yu;GAO Wa;WANG Wen-long;LIU Chun-xia(Ministry of Agriculture Key Laboratory of Clinical Diagnosis and Treatment Technology in Animal Disease,College of Veterinary/Medicine,Inner Mongolia Agricultural University,Hohhot 010018,China;Liangcheng County Animal Husbandry Workstation,Ulanqab 013750,China;Inner Mongolia Key Laboratory of Tick-borne Zoonotic Infectious Disease,Department of Medicine/Hetao College,Bayannur 015000,China;College of Life Science,Inner Mongolia Agricultural University,Hohhot 010018,China)

机构地区：[1]内蒙古农业大学兽医学院/农业部动物疾病临床诊疗技术重点实验室,内蒙古呼和浩特010018 [2]内蒙古乌兰察布市凉城县畜牧工作站,内蒙古乌兰察布013750 [3]河套学院医学系/内蒙古自治区蜱媒人畜共患传染病重点实验室,内蒙古巴彦淖尔015000 [4]内蒙古农业大学生命科学学院,内蒙古呼和浩特010018

出　　处：《中国预防兽医学报》2023年第3期245-252,共8页Chinese Journal of Preventive Veterinary Medicine

基　　金：内蒙古自然科学基金项目(2020MS03012)。

摘　　要：为探究捻转血矛线虫的伊维菌素(IVM)敏感虫株与耐药虫株显著差异的microRNA(miRNA)转录谱,发掘其耐药相关的miRNA,本实验采用Illumina Hiseq2000平台分别对捻转血矛线虫IVM敏感虫株与耐药虫株测序,构建cDNA文库,通过质控后采用生物信息学软件筛选捻转血矛线虫IVM敏感与耐药虫株的差异mi RNA。结果显示,捻转血矛线虫的IVM敏感与耐药虫株筛选出转录显著差异的miRNA共375个,其中253个转录上调,122个转录下调。利用RNAhybrid(V6.01)、Miranda(V3.3a)、TargetScan(V7.0)软件对筛选到的mi RNA靶基因进行预测,并取交集作为mi RNA靶基因预测结果,将筛选出显著差异的靶基因通过perl脚本与显著差异的miRNA进行关联分析,结果显示共关联到2106对miRNA-mRNA为负调控关系,其中涉及到477个差异的m RNA和603个差异的miRNA。对显著差异的miRNA靶基因进行GO富集分析,结果显示其中有492个GO terms,包括生物过程(BP)322个、细胞组分(CC)47个、分子功能(MF)123个。按照q值由小到大的顺序排列,被显著富集的有肌细胞发育分化及内肽酶活性等功能;按照富集基因数目由多到少排列,被显著富集的为有机物代谢、细胞代谢等功能。对显著差异的miRNA靶基因进行KEGG信号通路富集分析,按照q值由小到大的顺序排列,被显著富集到的有抗原处理与递呈等信号通路,其中显著差异靶基因也被富集到药物代谢-其他酶等与药物代谢相关的信号通路,以及一些在捻转血矛线虫耐IVM中起重要作用的通路,如:药物代谢细胞色素P450、细胞色素P450对外源药物代谢通路、ABC转运蛋白等。随机从差异miRNA中选10个进行RT-q PCR的验证,结果显示,相对转录水平结果与测序结果一致。本研究上述结果首次证实,捻转血矛线虫对IVM耐药性的产生与其代谢过程、肌细胞的分化发育过程及免疫过程中的某个或多个环节有一定的联系,本研究为进一步探究捻转In order to explore the microRNA(miRNA)transcription profiles of ivermectin sensitive and resistant strains of Haemonchus contortus,and to identify the miRNA associated with drug resistance.The Illumina Hiseq2000 platform was used to sequence the ivermectin sensitive and resistant strains of H.contortus.The cDNA library was constructed,and after quality control,the different miRNAs of ivermectin sensitive and resistant strains of H.contortus were screened by using bioinformatics software.The results showed that 375 miRNAs with significant transcriptional differences between ivermectin sensitive and resistant strains of H.contortus were screened,of which 253 were up-regulated and 122 were down-regulated.RNAhybrid(V6.01),Miranda(V3.3a)and TargetScan(V7.0)software was used to predict miRNA target genes,and the intersection was used as the prediction result of miRNA target genes.The correlation analysis between the target genes with significant differences and the miRNA with significant differences was conducted by perl script.The results showed that a total of 2106 miRNA-mRNA pairs were associated with negative regulation,including 477 differentially transcripted mRNAs and 603 differentially encoded miRNAs.GO enrichment analysis of significantly different miRNA target genes showed that 492 of them had GO terms,including 322 biological processes(BP),47 cell components(CC),and 123 molecular functions(MF).According to the order of q value from small to large,were significantly enriched muscle cell development and differentiation,endopeptidase activity,in accordance with the number of enrichment genes from more to less,was significantly enriched in organic substance metabolic process,cell metabolism.KEGG pathway enrichment was performed on miRNA target genes with significant differences,which were arranged in the order of q value from small to large,including antigen processing and presentation,among which significant difference target genes were also enriched in drug metabolism-other enzymes and other pathways related

关 键 词：捻转血矛线虫 伊维菌素 耐药性 转录组 mi RNA 

分 类 号：S852.7[农业科学—基础兽医学]