基于高通量测序的猪鞭虫雌、雄虫差异基因表达分析

doi:10.11843/j.issn.0366-6964.2016.09.021

摘要/Abstract

摘要：

为了探明猪鞭虫（Trichuris suis）雌、雄虫的转录组差异，丰富T.suis转录组数据信息，利用Illumina Hiseq 2000对临床采集的T.suis雌、雄虫进行高通量测序，将组装得到的Unigenes在NR、GO、COG和KEGG数据库中比对注释，并进行差异表达基因分析。结果，T.suis 雌、雄虫分别获得59 657 854条和48 414 184条高质量测序数据，分别组装获得21 026个和28 886个Unigenes。注释到NR、GO、COG和KEGG数据库的雌虫Unigenes数量分别为11 700、2 287、1 650和9 690个，雄虫Unigenes数量分别为12 902、2 414、1 791和10 377个，在NR数据库比对显示84.90%以上信息均来源于猪鞭虫。T.suis 雌、雄虫共有4 320个差异表达基因。以雌虫为对照，雄虫中共有3 496个基因表达上调，有824个基因表达下调。共有906个差异表达基因获得160个KEGG通路富集，其中显著富集通路7个。在T.suis转录组中发现了许多重要基因的表达，包括与T.suis繁殖相关的主要精子蛋白、组蛋白H2A/H2B及卵黄蛋白原，与T.suis致病性、免疫及炎症调节相关的蛋白酶、丝氨酸蛋白酶抑制分子和半胱氨酸蛋白酶抑制分子，以及众多未知功能的基因。本研究揭示了T.suis雌、雄虫差异表达基因的数量，获得了这些差异表达基因的功能、分类和代谢通路注释，为丰富T.suis转录组信息，揭示鞭虫与宿主互作特点及抗鞭虫靶点药物研发奠定基础。

Abstract:

To find transcriptome difference of female and male Trichuris suis，and to enrich pig whipworm transcriptome data，the transcriptomes were analyzed by Illumina Hiseq 2000 high-throughput RNA sequencing.All of the assembled Unigenes were annotated using BLAST search against NR，GO，COG and KEGG databases.The differentially expressed genes were enriched in gene ontology and KEGG pathway.The results showed that 21 026 and 28 886 unigenes were got from female and male T.suis transcriptomes which were assembled from 59 657 854 and 48 414 184 clean reads respectively.The annotated Unigenes of female T.suis to NR，GO，COG and KEGG databases were 11 700，2 287，1 650 and 9 690 respectively，while the data of male T.suis were 12 902，2 414，1 791 and 10 377.There were 4 320 differentially expressed genes found in female and male T.suis，and more than 84.90% of the information were derived from pig whipworm in NR database.Take female T.suis for reference，3 496 genes were up-regulated and 824 genes were down-regulated in male T.suis.There were 906 differentially expressed genes enriched in KEGG pathways，and 7 of them were significantly enriched.Many important genes associated with reproduction including Major sperm protein，Histone H2A/H2B and Vitellogenin，and genes related to pathogenisis，immunity and inflammatory regulation such as protease，Serpin and Cystatin were found in transcriptome of T.suis，in addition to these，there were many genes of unknown functions.This study revealed the number of differentially expressed genes in female and male T.suis，and obtained the function，classification and metabolic pathways of the differentially expressed genes，which enriched T.suis transcriptome information，and provide a solid basis for the study on T.suis-host interactions and developing anti-whipworm drugs.

中图分类号:

S852.73

周作勇，邵骜骏，徐婷婷，陈家磊，胡世君. 基于高通量测序的猪鞭虫雌、雄虫差异基因表达分析[J]. 畜牧兽医学报, 2016, 47(9): 1914-1923.

ZHOU Zuo-yong，SHAO Ao-jun，XU Ting-ting，CHEN Jia-lei，HU Shi-jun. Differential Expression Analysis of Female and Male Trichuris suis by a High-throughput Transcriptome Sequencing Method[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(9): 1914-1923.

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参考文献

［1］NISSEN S，POULSEN I H，NEJSUM P，et al.Prevalence of gastrointestinal nematodes in growing pigs in Kabale District in Uganda［J］.Trop Anim Health Prod，2011，43(3)：567-572.
［2］MATSUBAYASHI M，KITA T，NARUSHIMA T，et al.Coprological survey of parasitic infections in pigs and cattle in slaughterhouse in Osaka，Japan［J］.J Vet Med Sci，2009，71(8)：1079-1083.
［3］LAI M，ZHOU R Q，HUANG H C，et al.Prevalence and risk factors associated with intestinal parasites in pigs in Chongqing，China［J］.Res Vet Sci，2011，91(3)：e121-e124.
［4］WENG Y B，HU Y J，LI Y，et al.Survey of intestinal parasites in pigs from intensive farms in Guangdong Province，People’s Republic of China［J］.Vet Parasitol，2005，127(3-4)：333-336.
［5］ROEPSTORFF A，MEJER H，NEJSUM P，et al.Helminth parasites in pigs：new challenges in pig production and current research highlights［J］.Vet Parasitol，2011，180(1-2)：72-81.
［6］XIAO S H，UTZINGER J，TANNER M，et al.Advances with the Chinese anthelminthic drug tribendimidine in clinical trials and laboratory investigations［J］.Acta Trop，2013，126(2)：115-126.
［7］VERCRUYSSE J，SCHETTERS T P，KNOX D P，et al.Control of parasitic disease using vaccines：an answer to drug resistance?［J］.Rev Sci Tech，2007，26(1)：105-115.
［8］BOES J，HELWIGH A B.Animal models of intestinal nematode infections of humans［J］. Parasitology，2000，121 Suppl：S97-S111.
［9］HEWITSON J P，GRAINGER J R，MAIZELS R M.Helminth immunoregulation：the role of parasite secreted proteins in modulating host immunity［J］.Mol Biochem Parasitol，2009，167(1)：1-11.
［10］HIEMSTRA I H，KLAVER E J，VRIJLAND K，et al.Excreted/secreted Trichuris suis products reduce barrier function and suppress inflammatory cytokine production of intestinal epithelial cells［J］.Mol Immunol. 2014，60(1)：1-7.
［11］SCHÖLMERICH J.Trichuris suis ova in inflammatory bowel disease［J］. Dig Dis，2013，31(3-4)：391-395.
［12］SUMMERS R W，ELLIOTT D E，URBAN J J，et al. Trichuris suis therapy for active ulcerative colitis：a randomized controlled trial［J］.Gastroenterology，2005，128(4)：825-832.
［13］JEX A R，NEJSUM P，SCHWARZ E M，et al.Genome and transcriptome of the porcine whipworm Trichuris suis［J］.Nat Genet，2014，46(7)：701-706.
［14］祁云霞，刘永斌，荣威恒.转录组研究新技术：RNA-Seq及其应用［J］.遗传.2011，33(11)：1191-1202.
QI Y X，LIU Y B，RONG W H.RNA-Seq and its applications：a new technology for transcriptomics［J］.Hereditas，2011，33(11)：1191-1202.(in Chinese)
［15］LIU G H，XU M J，SONG H Q，et al.De novo assembly and characterization of the transcriptome of the pancreatic fluke Eurytrema pancreaticum (trematoda：Dicrocoeliidae) using Illumina paired-end sequencing［J］.Gene，2016，576(1 Pt 2)：333-338.
［16］POMAZNOY M Y，LOGACHEVA M D，YOUNG N D，et al.Whole transcriptome profiling of adult and infective stages of the trematode Opisthorchis felineus［J］.Parasitol Int，2016，65(1)：12-19.
［17］WANG X，XU X，LU X，et al.Transcriptome Bioinformatical Analysis of Vertebrate Stages of Schistosoma japonicum Reveals Alternative Splicing Events［J］.PLoS One，2015，10(9)：e138470.
［18］WALKER R A，SHARMAN P A，MILLER C M，et al.RNA Seq analysis of the Eimeria tenella gametocyte transcriptome reveals clues about the molecular basis for sexual reproduction and oocyst biogenesis［J］. BMC Genomics，2015，16(1)：94.
［19］HACARIZ O，AKGUN M，KAVAK P，et al.Comparative transcriptome profiling approach to glean virulence and immunomodulation-related genes of Fasciola hepatica［J］.BMC Genomics，2015，16(1)：366.
［20］ZHU X Q，KORHONEN P K，CAI H，et al.Genetic blueprint of the zoonotic pathogen Toxocara canis［J］.Nat Commun，2015，6(2)：6145.
［21］MORTAZAVI A，WILLIAMS B A，MCCUE K，et al.Mapping and quantifying mammalian transcriptomes by RNA-Seq［J］.Nat Methods，2008，5(7)：621-628.
［22］GOZ＇DZIK K，ENGSTRÖM A，HÖGLUND J.Optimization of in-house ELISA based on recombinant major sperm protein (rMSP) of Dictyocaulus viviparus for the detection of lungworm infection in cattle［J］.Res Vet Sci，2012，93(2)：813-818.
［23］RAMOS I，MARTIN-BENITO J，FINN R，et al.Nucleoplasmin binds histone H2A-H2B dimers through its distal face［J］. J Biol Chem，2010，285(44)：33771-33778.
［24］鲁爽.精子发生过程中组蛋白H2A.H2B编码及功能研究［D］.北京：中国协和医科大学，2009.
LU S.The research on histone H2A.H2B protein coding and function of spermatogenesis［D］.Beijing：Peking Union Medical College，2009.(in Chinese)
［25］李兆杰，杨丽君，王静，等.卵黄蛋白原的研究进展［J］.生命科学.2010，22(3)：284-290.
LI Z J，YANG L J，WANG J，et al.The progress in studies on vitellogenin［J］.Chinese Bulletin of Life Sciences，2010，22(3)：284-290.(in Chinese)
［26］TILNEY L G，CONNELLY P S，GUILD G M，et al.Adaptation of a nematode parasite to living within the mammalian epithelium［J］.J Exp Zool A Comp Exp Biol，2005，303(11)：927-945.
［27］TOUBARRO D，LUCENA-ROBLES M，NASCIMENTO G，et al.Serine protease-mediated host invasion by the parasitic nematode Steinernema carpocapsae［J］.J Biol Chem，2010，285(40)：30666-30675.
［28］BALASUBRAMANIAN N，TOUBARRO D，SIM?ES N.Biochemical study and in vitro insect immune suppression by a trypsin-like secreted protease from the nematode Steinernema carpocapsae［J］.Parasite Immunol，2010，32(3)：165-175.
［29］TOUBARRO D，LUCENA-ROBLES M，NASCIMENTO G，et al.An apoptosis-inducing serine protease secreted by the entomopathogenic nematode Steinernema carpocapsae［J］.Int J Parasitol，2009，39(12)：1319-1330.
［30］HEWITSON J P，GRAINGER J R，MAIZELS R M.Helminth immunoregulation：the role of parasite secreted proteins in modulating host immunity［J］.Mol Biochem Parasitol，2009，167(1)：1-11.
［31］KLAVER E J，KUIJK L M，LAAN L C，et al.Trichuris suis-induced modulation of human dendritic cell function is glycan-mediated［J］. Int J Parasitol，2013，43(3-4)：191-200.
［32］VERMEIRE J J，CHO Y，LOLIS E，et al.Orthologs of macrophage migration inhibitory factor from parasitic nematodes［J］.Trends Parasitol，2008，24(8)：355-363.
［33］SELKIRK M E，LAZARI O，HUSSEIN A S，et al.Nematode acetylcholinesterases are encoded by multiple genes and perform non-overlapping functions［J］.Chem Biol Interact，2005，157-158：263-268.
［34］NOZAKI T，ALI V，TOKORO M.Sulfur-containing amino acid metabolism in parasitic protozoa［J］.Adv Parasitol，2005，60：1-99.
［35］MORGAN C T，NOBLE D，KIMBLE J.Mitosis-meiosis and sperm-oocyte fate decisions are separable regulatory events［J］.Proc Natl Acad Sci U S A，2013，110(9)：3411-3416.
［36］RADTKE F，MACDONALD H R，TACCHINI-COTTIER F.Regulation of innate and adaptive immunity by Notch［J］.Nat Rev Immunol，2013，13(6)：427-437.
［37］FORTINI M E.Notch signaling：the core pathway and its posttranslational regulation［J］.Dev Cell，2009，16(5)：633-647.

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