食线虫真菌洛斯里被毛孢线粒体基因组的再分析
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摘要
【目的】鉴定洛斯里被毛孢OWVT-1菌株的线粒体基因组,验证公布的USA-87-5菌株线粒体基因组中的错误,对洛斯里被毛孢正确的线粒体基因组序列进行注释并开展不同被毛孢物种间的比较线粒体基因组学分析。【方法】借助DNA高通量测序数据并通过必要的Sanger测序组装OWVT-1的线粒体基因组。通过PCR验证OWVT-1与公布的USA-87-5线粒体基因组序列差异的真实性。利用多种生物信息方法分析和注释洛斯里被毛孢的线粒体基因组。【结果】公布的洛斯里被毛孢USA-87-5菌株的线粒体基因组存在几处序列错误,包括3处长片段的插入缺失和多处短片段的插入缺失。实际上,洛斯里被毛孢USA-87-5与OWVT-1菌株的线粒体基因组序列完全相同。该菌的线粒体基因组全长62949 bp,在7个基因中共插入13个内含子,部分内含子和基因间区显现出序列退化的特征。洛斯里被毛孢、明尼苏达被毛孢、线虫被毛孢的线粒体基因组具有较强的共线性关系。除一些独立的ORF外,核心蛋白编码基因、rRNA基因和tRNA基因的排列顺序非常保守。基因间区的长短是影响3种被毛孢线粒体基因组大小最主要的因素。【结论】公布的洛斯里被毛孢USA-87-5菌株线粒体基因组中存在序列错误。本文新报道了OWVT-1菌株的线粒体基因组,并进行注释和比较线粒体基因组学分析。
[Objective] To identify the mitochondrial genome(mitogenome) of Hirsutella rhossiliensis OWVT-1 strain, verify erroneous sequences present in the published USA-87-5 mitogenome, and perform comparative mitogenomic analyses of three Hirsutella species. [Methods] The OWVT-1 mitogenome was assembled based on high-throughput DNA sequencing data and Sanger sequencing. PCR assays were performed to verify 3 long varying regions observed during alignment between the published USA-87-5 mitogenome and OWVT-1 mitogenome assembled in this study. Bioinformatics analyses were used to annotate the mitogenome sequence of OWVT-1. [Results] We detected several sequence errors in the published USA-87-5 mitogenome, including long sequence insertions/deletions and small indels. The mitogenomes of OWVT-1 and USA-87-5 were identical without any nucleotide difference. The complete mitogenome of H. rhossiliensis was 62949 bp in length, with 13 introns in seven genes. Several introns and intergenic regions seemed to have degenerated. The mitogenomes of H. rhossiliensis, H. vermicola, and H. minnesotensis showed a high synteny. Except free-standing ORFs, the gene order of core protein-encoding genes, rRNA genes, and tRNA genes were highly conserved among the three Hirsutella mitogenomes. The length at intergenic regions was a main factor affecting mitogenome sizes of different Hirsutella species. [Conclusion] There were erroneous sequences in the published USA-87-5 mitogenome. We reported the authentic mitogenome sequence of H. rhossiliensis using the highly virulent strain OWVT-1.
[Objective] To identify the mitochondrial genome(mitogenome) of Hirsutella rhossiliensis OWVT-1 strain, verify erroneous sequences present in the published USA-87-5 mitogenome, and perform comparative mitogenomic analyses of three Hirsutella species. [Methods] The OWVT-1 mitogenome was assembled based on high-throughput DNA sequencing data and Sanger sequencing. PCR assays were performed to verify 3 long varying regions observed during alignment between the published USA-87-5 mitogenome and OWVT-1 mitogenome assembled in this study. Bioinformatics analyses were used to annotate the mitogenome sequence of OWVT-1. [Results] We detected several sequence errors in the published USA-87-5 mitogenome, including long sequence insertions/deletions and small indels. The mitogenomes of OWVT-1 and USA-87-5 were identical without any nucleotide difference. The complete mitogenome of H. rhossiliensis was 62949 bp in length, with 13 introns in seven genes. Several introns and intergenic regions seemed to have degenerated. The mitogenomes of H. rhossiliensis, H. vermicola, and H. minnesotensis showed a high synteny. Except free-standing ORFs, the gene order of core protein-encoding genes, rRNA genes, and tRNA genes were highly conserved among the three Hirsutella mitogenomes. The length at intergenic regions was a main factor affecting mitogenome sizes of different Hirsutella species. [Conclusion] There were erroneous sequences in the published USA-87-5 mitogenome. We reported the authentic mitogenome sequence of H. rhossiliensis using the highly virulent strain OWVT-1.
引文
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[2]Desmond E,Brochier-Armanet C,Forterre P,Gribaldo S.On the last common ancestor and early evolution of eukaryotes:reconstructing the history of mitochondrial ribosomes.Research in Microbiology,2011,162(1):53-70.
[3]Zhang YJ,Zhao YX,Zhang S,Chen L,Liu XZ.Reanalysis of the mitochondrial genome of the pneumocandin-producing fungus Glarea lozoyensis.Acta Microbiologica Sinica,2017,57(5):724-736.
[4]Ciancio A,Colagiero M,Rosso LC,Gutierrez SNM,Grasso G.Phylogeny and morphology of Hirsutella tunicata sp.nov.(Ophiocordycipitaceae),a novel mite parasite from Peru.Mycoscience,2013,54(5):378-386.
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[6]Quandt CA,Kepler RM,Gams W,Araújo JPM,Ban S,Evans HC,Hughes D,Humber R,Hywel-Jones N,Li ZZ,Luangsa-Ard JJ,Rehner SA,Sanjuan T,Sato H,Shrestha B,Sung GH,Yao YJ,Zare R,Spatafora JW.Phylogenetic-based nomenclatural proposals for Ophiocordycipitaceae(Hypocreales)with new combinations in Tolypocladium.IMAFungus,2014,5(1):121-134.
[7]Xiang MC,Yang EC,Xiao QM,Liu XZ,Shen SY.Hirsutella vermicola sp.nov.,a new species parasitizing bacteria-feeding nematodes.Fungal Diversity,2006,22:255-265.
[8]Zhang YJ,Zhang HY,Liu XZ,Zhang S.Mitochondrial genome of the nematode endoparasitic fungus Hirsutella vermicola reveals a high level of synteny in the family Ophiocordycipitaceae.Applied Microbiology and Biotechnology,2017,101(8):3295-3304.
[9]Wang NN,Zhang YJ,Hussain M,Li K,Xiang MC,Liu XZ.The mitochondrial genome of the nematode endoparasitic fungus Hirsutella rhossiliensis.Mitochondrial DNA Part B,Resources,2016,1(1):114-115.
[10]Zhang YJ,Zhang S,Liu XZ.The complete mitochondrial genome of the nematode endoparasitic fungus Hirsutella minnesotensis.Mitochondrial DNA Part A,DNA Mapping,Sequencing,and Analysis,2016,27(4):2693-2694.
[11]Zhao XH,Xu YL.Advance on Hirsutella sp.of biological control.System Sciences and Comprehensive Studies in Agriculture,2011,27(3):376-381.(in Chinese)赵晓晖,许艳丽.生防真菌被毛孢的研究进展.农业系统科学与综合研究,2011,27(3):376-381.
[12]Liu SF,Chen SY.Efficacy of the fungi Hirsutella minnesotensis and H.rhossiliensis from liquid culture for control of the soybean cyst nematode Heterodera glycines.Nematology,2005,7(1):149-157.
[13]Chen SY,Liu XZ.Control of the soybean cyst nematode by the fungi Hirsutella rhossiliensis and Hirsutella minnesotensis in greenhouse studies.Biological Control,2005,32(2):208-219.
[14]Wang NN,Zhang YJ,Jiang XZ,Shu C,Hamid MI,Hussain M,Chen SY,Xu JP,Xiang MC,Liu XZ.Population genetics of Hirsutella rhossiliensis,a dominant parasite of cyst nematode juveniles on a continental scale.Applied and Environmental Microbiology,2016,82(21):6317-6325.
[15]Liu XZ,Chen SY.Screening isolates of Hirsutella species for biocontrol of Heterodera glycines.Biocontrol Science and Technology,2001,11(1):151-160.
[16]Xiang MC,Yang XH,Wang ZX,Liu XZ,Chen SY,Xiao QM.Variability of morphology,parasitism,and nucleotide sequences among isolates and species of nematophagous Hirsutella.Biological Control,2007,41(1):110-119.
[17]Liu XZ,Chen SY.Nutritional requirements of the nematophagous fungus Hirsutella rhossiliensis.Biocontrol Science and Technology,2002,12(3):381-393.
[18]Sun MH,Liu XZ.Carbon requirements of some nematophagous,entomopathogenic and mycoparasitic Hyphomycetes as fungal biocontrol agents.Mycopathologia,2006,161(5):295-305.
[19]Wang B,Liu XY,Wu WP,Liu XZ,Li SD.Purification,characterization,and gene cloning of an alkaline serine protease from a highly virulent strain of the nematode-endoparasitic fungus Hirsutella rhossiliensis.Microbiological Research,2009,164(6):665-673.
[20]Wang B,Wu WP,Liu XZ.Purification and characterization of a neutral serine protease with nematicidal activity from Hirsutella rhossiliensis.Mycopathologia,2007,163(3):169-176.
[21]Zhang LM,Yang EC,Xiang MC,Liu XZ,Chen SY.Population dynamics and biocontrol efficacy of the nematophagous fungus Hirsutella rhossiliensis as affected by stage of the soybean cyst nematode.Biological Control,2008,47(2):244-249.
[22]Zhang LM,Liu XZ,Zhu SF,Chen SY.Detection of the nematophagous fungus Hirsutella rhossiliensis in soil by real-time PCR and parasitism bioassay.Biological Control,2006,36(3):316-323.
[23]Shu C,Lai YL,Yang EC,Chen SY,Xiang MC,Liu XZ.Functional response of the fungus Hirsutella rhossiliensis to the nematode,Heterodera glycines.Science China Life Sciences,2015,58(7):704-712.
[24]赖屹玲.被毛孢寄生大豆孢囊线虫的分子机制以及杀线虫化合物高通量筛选方法的建立.北京:中国科学院大学博士学位论文,2014.
[25]Chen SY.Infection of Heterodera glycines by Hirsutella rhossiliensis in a Minnesota soybean field.Journal of Nematology,1997,29:573.
[26]Cai WJ,Xu DB,Lan X,Xie HH,Wei JG.A new method for the extraction of fungal genomic DNA.Agricultural Research and Application,2014,(3):1-5.(in Chinese)蔡文娇,徐大彬,蓝霞,谢红辉,韦继光.一种提取真菌基因组DNA的新方法.农业研究与应用,2014,(3):1-5.
[27]Darling AE,Mau B,Perna NT.ProgressiveMauve:multiple genome alignment with gene gain,loss and rearrangement.PLoS One,2010,5(6):e11147.
[28]Hahn C,Bachmann L,Chevreux B.Reconstructing mitochondrial genomes directly from genomic next-generation sequencing reads-a baiting and iterative mapping approach.Nucleic Acids Research,2013,41(13):e129.
[29]Lowe TM,Chan PP.tRNAscan-SE On-line:integrating search and context for analysis of transfer RNA genes.Nucleic Acids Research,2016,44(W1):W54-W57.
[30]Xiao SJ,Nguyen DT,Wu BJ,Hao WL.Genetic drift and indel mutation in the evolution of yeast mitochondrial genome size.Genome Biology and Evolution,2017,9(11):3088-3099.
[2]Desmond E,Brochier-Armanet C,Forterre P,Gribaldo S.On the last common ancestor and early evolution of eukaryotes:reconstructing the history of mitochondrial ribosomes.Research in Microbiology,2011,162(1):53-70.
[3]Zhang YJ,Zhao YX,Zhang S,Chen L,Liu XZ.Reanalysis of the mitochondrial genome of the pneumocandin-producing fungus Glarea lozoyensis.Acta Microbiologica Sinica,2017,57(5):724-736.
[4]Ciancio A,Colagiero M,Rosso LC,Gutierrez SNM,Grasso G.Phylogeny and morphology of Hirsutella tunicata sp.nov.(Ophiocordycipitaceae),a novel mite parasite from Peru.Mycoscience,2013,54(5):378-386.
[5]Simmons DR,Kepler RM,Rehner SA,Groden E.Phylogeny of Hirsutella species(Ophiocordycipitaceae)from the USA:remedying the paucity of Hirsutella sequence data.IMAFungus,2015,6(2):345-356.
[6]Quandt CA,Kepler RM,Gams W,Araújo JPM,Ban S,Evans HC,Hughes D,Humber R,Hywel-Jones N,Li ZZ,Luangsa-Ard JJ,Rehner SA,Sanjuan T,Sato H,Shrestha B,Sung GH,Yao YJ,Zare R,Spatafora JW.Phylogenetic-based nomenclatural proposals for Ophiocordycipitaceae(Hypocreales)with new combinations in Tolypocladium.IMAFungus,2014,5(1):121-134.
[7]Xiang MC,Yang EC,Xiao QM,Liu XZ,Shen SY.Hirsutella vermicola sp.nov.,a new species parasitizing bacteria-feeding nematodes.Fungal Diversity,2006,22:255-265.
[8]Zhang YJ,Zhang HY,Liu XZ,Zhang S.Mitochondrial genome of the nematode endoparasitic fungus Hirsutella vermicola reveals a high level of synteny in the family Ophiocordycipitaceae.Applied Microbiology and Biotechnology,2017,101(8):3295-3304.
[9]Wang NN,Zhang YJ,Hussain M,Li K,Xiang MC,Liu XZ.The mitochondrial genome of the nematode endoparasitic fungus Hirsutella rhossiliensis.Mitochondrial DNA Part B,Resources,2016,1(1):114-115.
[10]Zhang YJ,Zhang S,Liu XZ.The complete mitochondrial genome of the nematode endoparasitic fungus Hirsutella minnesotensis.Mitochondrial DNA Part A,DNA Mapping,Sequencing,and Analysis,2016,27(4):2693-2694.
[11]Zhao XH,Xu YL.Advance on Hirsutella sp.of biological control.System Sciences and Comprehensive Studies in Agriculture,2011,27(3):376-381.(in Chinese)赵晓晖,许艳丽.生防真菌被毛孢的研究进展.农业系统科学与综合研究,2011,27(3):376-381.
[12]Liu SF,Chen SY.Efficacy of the fungi Hirsutella minnesotensis and H.rhossiliensis from liquid culture for control of the soybean cyst nematode Heterodera glycines.Nematology,2005,7(1):149-157.
[13]Chen SY,Liu XZ.Control of the soybean cyst nematode by the fungi Hirsutella rhossiliensis and Hirsutella minnesotensis in greenhouse studies.Biological Control,2005,32(2):208-219.
[14]Wang NN,Zhang YJ,Jiang XZ,Shu C,Hamid MI,Hussain M,Chen SY,Xu JP,Xiang MC,Liu XZ.Population genetics of Hirsutella rhossiliensis,a dominant parasite of cyst nematode juveniles on a continental scale.Applied and Environmental Microbiology,2016,82(21):6317-6325.
[15]Liu XZ,Chen SY.Screening isolates of Hirsutella species for biocontrol of Heterodera glycines.Biocontrol Science and Technology,2001,11(1):151-160.
[16]Xiang MC,Yang XH,Wang ZX,Liu XZ,Chen SY,Xiao QM.Variability of morphology,parasitism,and nucleotide sequences among isolates and species of nematophagous Hirsutella.Biological Control,2007,41(1):110-119.
[17]Liu XZ,Chen SY.Nutritional requirements of the nematophagous fungus Hirsutella rhossiliensis.Biocontrol Science and Technology,2002,12(3):381-393.
[18]Sun MH,Liu XZ.Carbon requirements of some nematophagous,entomopathogenic and mycoparasitic Hyphomycetes as fungal biocontrol agents.Mycopathologia,2006,161(5):295-305.
[19]Wang B,Liu XY,Wu WP,Liu XZ,Li SD.Purification,characterization,and gene cloning of an alkaline serine protease from a highly virulent strain of the nematode-endoparasitic fungus Hirsutella rhossiliensis.Microbiological Research,2009,164(6):665-673.
[20]Wang B,Wu WP,Liu XZ.Purification and characterization of a neutral serine protease with nematicidal activity from Hirsutella rhossiliensis.Mycopathologia,2007,163(3):169-176.
[21]Zhang LM,Yang EC,Xiang MC,Liu XZ,Chen SY.Population dynamics and biocontrol efficacy of the nematophagous fungus Hirsutella rhossiliensis as affected by stage of the soybean cyst nematode.Biological Control,2008,47(2):244-249.
[22]Zhang LM,Liu XZ,Zhu SF,Chen SY.Detection of the nematophagous fungus Hirsutella rhossiliensis in soil by real-time PCR and parasitism bioassay.Biological Control,2006,36(3):316-323.
[23]Shu C,Lai YL,Yang EC,Chen SY,Xiang MC,Liu XZ.Functional response of the fungus Hirsutella rhossiliensis to the nematode,Heterodera glycines.Science China Life Sciences,2015,58(7):704-712.
[24]赖屹玲.被毛孢寄生大豆孢囊线虫的分子机制以及杀线虫化合物高通量筛选方法的建立.北京:中国科学院大学博士学位论文,2014.
[25]Chen SY.Infection of Heterodera glycines by Hirsutella rhossiliensis in a Minnesota soybean field.Journal of Nematology,1997,29:573.
[26]Cai WJ,Xu DB,Lan X,Xie HH,Wei JG.A new method for the extraction of fungal genomic DNA.Agricultural Research and Application,2014,(3):1-5.(in Chinese)蔡文娇,徐大彬,蓝霞,谢红辉,韦继光.一种提取真菌基因组DNA的新方法.农业研究与应用,2014,(3):1-5.
[27]Darling AE,Mau B,Perna NT.ProgressiveMauve:multiple genome alignment with gene gain,loss and rearrangement.PLoS One,2010,5(6):e11147.
[28]Hahn C,Bachmann L,Chevreux B.Reconstructing mitochondrial genomes directly from genomic next-generation sequencing reads-a baiting and iterative mapping approach.Nucleic Acids Research,2013,41(13):e129.
[29]Lowe TM,Chan PP.tRNAscan-SE On-line:integrating search and context for analysis of transfer RNA genes.Nucleic Acids Research,2016,44(W1):W54-W57.
[30]Xiao SJ,Nguyen DT,Wu BJ,Hao WL.Genetic drift and indel mutation in the evolution of yeast mitochondrial genome size.Genome Biology and Evolution,2017,9(11):3088-3099.
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