尖音库蚊复合体蚊虫DNA甲基化水平分析  

Analysis on DNA methylation levels in mosquitoes of the Culex pipiens complex

作　　者：郭思含 黄新安 徐寒黎 李春晓 刘康康 邢丹 赵腾 GUO Sihan;HUANG Xin’an;XU Hanli;LI Chunxiao;LIU Kangkang;XING Dan;ZHAO Teng(Artemisinin Research Center,Guangzhou University of Chinese Medicine,Guangzhou 510006,Guangdong,China;State Key Laboratory of Pathogen and Biosecurity,Beijing 100071,China;College of Life Sciences and Bioengineering,School of Physical Science and Engineering,Beijing Jiaotong University,Beijing 100091,China)

机构地区：[1]广州中医药大学青蒿研究中心,广东广州510006 [2]病原微生物生物安全全国重点实验室,北京100071 [3]北京交通大学生命科学与生物工程研究院,北京100091

出　　处：《中国寄生虫学与寄生虫病杂志》2024年第4期502-511,共10页Chinese Journal of Parasitology and Parasitic Diseases

摘　　要：目的分析比较尖音库蚊复合体中淡色库蚊、致倦库蚊、骚扰库蚊的DNA甲基化水平以及致倦库蚊吸血前后DNA甲基化水平。方法采集羽化后5 d且未吸血的3个库蚊亚种和吸血3 d的致倦库蚊,提取DNA,超声切割为约250 bp的片段后进行测序,测序数据与致倦库蚊基因组序列(Taxonomy:ID7176)进行比对,获取全基因组胞嘧啶碱基甲基化信息。从基因组、染色体和染色体元件水平分析3个库蚊亚种的甲基化水平(甲基化水平高于3%定义为高甲基化)。比较未吸血的3个库蚊亚种间以及吸血前后致倦库蚊的甲基化水平差异,P<0.001且甲基化差异绝对值>5的位点记为差异甲基化位点;Q<0.05且甲基化差异绝对值>3的区域记为差异甲基化区域(DMR)。将距离DMR最近的转录起始位点(TSS)所在基因记为DMR相关基因,对其进行基因本体论(GO)富集分析。结果淡色库蚊、骚扰库蚊和致倦库蚊全基因组甲基化水平分别为0.454%~0.672%、0.491%~0.649%和0.499%~0.655%(均低于3%),CHH位点甲基化水平分别为0.631%、0.618%和0.624%,均高于CHG位点的0.567%、0.559%、0.559%(t=7.14、83.43、6.87,均P<0.05)和CG/CpG位点的0.508%、0.505%、0.505%(t=10.59、12.52、13.33,均P<0.05);3个库蚊亚种共有的高甲基化位点有56个、高甲基化区域有11个。淡色库蚊、骚扰库蚊和致倦库蚊之间全基因组甲基化水平差异无统计学意义(F=0.07,P>0.05),1号染色体(0.568%、0.562%、0.565%)、2号染色体(0.573%、0.564%、0.566%)、3号染色体(0.575%、0.566%、0.569%)的甲基化水平差异均无统计学意义(F=0.05、0.11、0.13,均P>0.05),启动子(0.567%、0.552%、0.556%)、外显子(0.562%、0.556%、0.558%)、内含子(0.561%、0.550%、0.555%)和TSS(0.579%、0.506%、0.621%)的甲基化水平差异均无统计学意义(F=0.37、0.06、0.06、0.16,均P>0.05)。淡色库蚊和骚扰库蚊间筛选出178个差异甲基化位点、4个DMR,淡色库蚊和致倦库蚊间筛选出209个差异甲基�Objective To analyze and compare the DNA methylation levels of three subspecies of Culex pipi⁃ens complex,including Cx.p.pallens,Cx.p.molestus and Cx.p.quinquefasciatus,and the DNA methylation levels of Cx.p.quinquefasciatus before and after blood‑feeding.Methods Mosquitoes of the 3 subspecies were collected at 5 days post‑feathering without blood‑feeding and Cx.p.quinquefasciatus were collected at 3 days after blood‑feeding.DNA was extracted and sonicated into fragments of approximately 250 bp.The fragmented DNA was sequenced,and the data were aligned with the reference genome sequence of Cx.p.quinquefasciatus(Taxonomy ID:7176).Methyla‑tion levels of the 3 subspecies were analyzed at the genomic,chromosomal and elemental levels(the methylation level above 3%was considered hypermethylated).The differences in methylation levels among the 3 subspecies mosquitoes without blood‑feeding,and in Cx.p.quinquefasciatus before and after blood‑feeding were compared.The sites with P<0.001 and absolute value of methylation difference>5 were identified as differentially methylated sites.The regions with Q<0.05 and the absolute value of methylation difference>3 were identified as differentially methylated regions(DMR).The genes with the nearest transcription start site(TSS)to DMRs were identified as DMR‑associated genes,which were subjected to gene ontology(GO)enrichment analysis.Results The genome‑wide methylation levels of Cx.p.pallens,Cx.p.molestus and Cx.p.quinquefasciatus were 0.454%-0.672%,0.491%-0.649%and 0.499%-0.655%,respectively,all were below 3%.The methylation levels of CHH of Cx.p.pallens,Cx.p.molestus and Cx.p.quinquefas⁃ciatus were 0.631%,0.618%and 0.624%,respectively,which were higher than CHG(0.567%,0.559%,0.559%)(t=7.14,83.43,6.87,all P<0.05)and CG/CpG(0.508%,0.505%,0.505%)(t=10.59,12.52,13.33,all P<0.05).There were 56 hypermethylated sites and 11 hypermethylated regions present among all 3 subspecies.No significant differ‑ences were found among the 3 subspecies(F=0.07,P>0.05)at genome�

关 键 词：尖音库蚊复合体 DNA甲基化 全基因组亚硫酸氢盐测序技术 

分 类 号：R384.112[医药卫生—医学寄生虫学]