9种瘿蚜的Wolbachia分子检测和系统发育分析
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摘要
[目的]以五倍子蚜、杨树瘿蚜和黄连木瘿蚜等9种瘿蚜为实验材料,探讨瘿蚜Wolbachia与其它自由生活蚜虫携带的Wolbachia的株系差异及亲缘关系,明确五倍子蚜、杨树瘿蚜和黄连木瘿蚜等9种瘿蚜的Wolbachia感染情况、株系及系统发育关系。[方法]从田间采集新鲜虫瘿,将蚜虫转移到离心管内,100%乙醇-20℃保存。采用Wolbachia的16S rRNA、wsp、fts Z、gro E和glt A基因引物,以瘿蚜的总DNA为模板进行扩增、测序和分析,联合NCBI中Wolbachia 16S rRNA相关基因序列进行系统发育分析。[结果]在检测的9种瘿蚜中,有3种瘿蚜共3个种群感染有Wolbachia,其中滇叶瘿绵蚜昆明种群的Wolbachia属于B超群,肚倍蚜昆明种群和角倍蚜峨眉种群的Wolbachia可能属于新发现的O超群。[结论]与自由生活的蚜虫相比,瘿蚜的Wolbachia感染率偏低,且株系间的差异较大。
[Objective]To determine the infection and phylogenetic relationships of Wolbachia in nine gall aphids hosted on Rhus spp.,Populus yunnanensis and Pistacia chinensis. [Method]Fresh aphid galls were collected in the field and transferred the aphids to an Eppendorf tube,and placed in 100% ethanol then deposited in-20℃. The genomic DNA was amplified and sequenced using the primers of 16 S rRNA,wsp,fts Z,gro E and glt A genes. The phylogenetic relationships were analysis using Wolbachia 16 S rRNA gene sequences and relevant gene sequences from NCBI. [Result]Three species among the nine gall aphids species were infected with Wolbachia. The strain from Pemphigus yunnanensis of Kunming population belongs to B supergroup,and strain from Kaburagia rhusicola of Kunming population and Schlechtendalia chinensis of Emei population belong to O supergroup. [Conclusion]Compared with the free-living aphids,Wolbachia infected in gall aphids have low infection rates and significant differences among strains.
[Objective]To determine the infection and phylogenetic relationships of Wolbachia in nine gall aphids hosted on Rhus spp.,Populus yunnanensis and Pistacia chinensis. [Method]Fresh aphid galls were collected in the field and transferred the aphids to an Eppendorf tube,and placed in 100% ethanol then deposited in-20℃. The genomic DNA was amplified and sequenced using the primers of 16 S rRNA,wsp,fts Z,gro E and glt A genes. The phylogenetic relationships were analysis using Wolbachia 16 S rRNA gene sequences and relevant gene sequences from NCBI. [Result]Three species among the nine gall aphids species were infected with Wolbachia. The strain from Pemphigus yunnanensis of Kunming population belongs to B supergroup,and strain from Kaburagia rhusicola of Kunming population and Schlechtendalia chinensis of Emei population belong to O supergroup. [Conclusion]Compared with the free-living aphids,Wolbachia infected in gall aphids have low infection rates and significant differences among strains.
引文
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[26]Sintupachee S,Milne J R,Poonchaisri S,et al.Closely related Wolbachia strains within the pumpkin arthropod community and the potential for horizontal transmission via the plant[J].Microbial E-cology,2006,51(3):294-301.
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[29]江幸福,王蕾,张蕾,等.蔬菜蚜虫感染沃尔巴克氏体(Wolbachia)的分子检测[J].植物保护,2009,35(4):63-65.
[30]Duron O,Bouchon D,Boutin S,et al.The diversity of reproductive parasites among arthropods:Wolbachia do not walk alone[J].BMC Biology,2008,6(1):1-12.
[31]West S A,Cook J M,Werren J H,et al.Wolbachia in two insect host-parasitoid communities[J].Molecular Ecology,1998,7(11):1457-1465.
[32]Casiraghi M,Bordenstein S R,Baldo L,et al.Phylogeny of Wolbachia pipientis based on glt A,gro EL and fts Z gene sequences:clustering of arthropod and nematode symbionts in the F supergroup,and evidence for further diversity in the Wolbachia tree[J].Microbiology,2005,151(12):4015-4022.
[2]Augustinos A A,Santosgarcia D,Dionyssopoulou E,et al.Detection and characterization of Wolbachia infections in natural populations of aphids is the hidden diversity fully unraveled[J].PLo SONE,2011,6(12):e28695.
[3]Wang Z,Su X M,Wen J,et al.Widespread infection and diverse infection patterns of Wolbachia in Chinese aphids[J].Insect Science,2014,21(3):313-325.
[4]Bing X L,Xia W Q,Gui J D,et al.Diversity and evolution of the Wolbachia endosymbionts of Bemisia(Hemiptera:Aleyrodidae)whiteflies[J].Ecology and Evolution,2014,13(4):2714-2737.
[5]Baldo L,Dunning Hotopp J C,Jolley K A,et al.Multilocus sequence typing system for the endosymbiont Wolbachia pipientis[J].Applied and Environmental Microbiology,2006,72(11):7098-7110.
[6]Paraskevopoulos C,Bordenstein S R,Wernegreen J J,et al.Toward a Wolbachia multilocus sequence typing system:discrimination of Wolbachia strains present in drosophila species[J].Current Microbiology,2006,53(5):388-395.
[7]Werren J H,Baldo L,Clark M E.Wolbachia:master manipulators of invertebrate biology[J].Nature Reviews Microbiology,2008,6(10):741-751.
[8]Stouthamer R,Breeuwert J A,Luck R F,et al.Molecular identification of microorganisms associated with parthenogenesis[J].Nature,1993,361(6407):66-68.
[9]Yen J H,Barr A R.New hypothesis of the cause of cytoplasmic incompatibility in Culex pipiens L[J].Nature,1971,232(5313):657-658.
[10]Gdd H,Men M,Walker L E.The importance of cytoplasmic male killing elements in natural populations of the two spot ladybird,Adalia bipunctata(Linnaeus)(Coleoptera:Coccinellidae)[J].Biological Journal of the Linnean Society,1993,49(2):195-202.
[11]Rigaud T,Souty-Grosset C,Raimond R,et al.Feminizing endocytobiosis in the terrestrial crustacean Armadillidium vulgare Latr.(Isopoda):Recent acquisitions[J].Endocytobiosis and Cell Research,1991,7(3):259-273.
[12]Moreira L A,Iturbe-Ormaetxe I,Jeffery J A,et al.Wolbachia symbiont in Aedes aegypti limits infection with dengue,Chikungunya,and Plasmodium[J].Cell,2009,7(139):1268-1278.
[13]Mc Meniman C J,Lane R V,Cass B N,et al.Stable introduction of a life-shortening Wolbachia infection into the mosquito Aedes aegypti[J].Science,2009,323(5910):141-144.
[14]Hedges L M,Brownlie J C,O’Neill S L,et al.Wolbachia and virus protection in insects[J].Science,2008,322(5902):702.
[15]Hosokawa T,Koga R,Kikuchi Y,et al.Wolbachia as a bacteriocyte-associated nutritional mutualist[J].Proceedings of the National Academy of Sciences of the United States of America,2010,2(107):769-774.
[16]Fast E M,Toomey M E,Panaram K,et al.Wolbachia enhance Drosophila stem cell proliferation and target the germline stem cell niche[J].Science,2011,334(6058):990-992.
[17]Bourtzis K.Wolbachia-based technologies for insect pest population control[J].Advances in Experimental Medicine and Biology,2008,627(1):104-113.
[18]Saridaki A,Bourtzis K.Wolbachia:more than just a bug in insects genitals[J].Current Opinion in Microbiology,2010,13(1):67-72.
[19]杨子祥.五倍子高产培育技术[M].北京:中国林业出版社,2011.
[20]Werren J H,Windsor D M.Wolbachia infection frequencies in insects:evidence of a global equilibrium[J].Proceeding of the Royal Society London B:Biological Sciences,2000,267(1450):1277-1285.
[21]Zhou W,Rousset F,O’Neill S.Phylogeny and PCR-based classification of Wolbachia strains using wsp gene sequences[J].Proceeding of the Royal Society London B:Biological Sciences,1998,265(1395):509-515.
[22]Gottlieb Y,Ghanim M,Chiel E,et al.Identification and localization of a Rickettsia sp.in Bemisia tabaci(Homoptera:Aleyrodidae)[J].Applied&Environmental Microbiology,2006,72(5):3646-3652.
[23]Caspi F A,Inbar M,Steinberg S,et al.Characterization of the symbiont Rickettsia in the mirid bug Nesidiocoris tenuis(Reuter)(Heteroptera:Miridae)[J].Bulletin of Entomological Research,2014,104(6):681-688.
[24]Stouthamer R,Breeuwert J A,Luck R F,et al.Molecular identification of microorganisms associated with parthenogenesis[J].Nature,1993,361(6407):66-68.
[25]Breeuwer J A,Stouthamer R,Barns S M,et al.Phylogeny of cytoplasmic incompatibility microorganisms in the parasitoid wasp genus Nasonia(Hymenoptera:Pteromalidae)based on 16S ribosomal DNA sequences[J].Insect Molecular Biology,2010,1(1):25-36.
[26]Sintupachee S,Milne J R,Poonchaisri S,et al.Closely related Wolbachia strains within the pumpkin arthropod community and the potential for horizontal transmission via the plant[J].Microbial E-cology,2006,51(3):294-301.
[27]Wang Z,Shen Z R,Song Y D,et al.Distribution and diversity of Wolbachia in different populations of the wheat aphid Sitobion miscanthi(Hemiptera:Aphididae)in China[J].European Journal of Entomology,2009,106(1):49-55.
[28]Jeyaprakash A,Hoy M A.Long PCR improves Wolbachia DNA amplification:wsp sequences found in 76%of sixty-three arthropod species[J].Insect Molecular Biology,2000,9(4):393-405.
[29]江幸福,王蕾,张蕾,等.蔬菜蚜虫感染沃尔巴克氏体(Wolbachia)的分子检测[J].植物保护,2009,35(4):63-65.
[30]Duron O,Bouchon D,Boutin S,et al.The diversity of reproductive parasites among arthropods:Wolbachia do not walk alone[J].BMC Biology,2008,6(1):1-12.
[31]West S A,Cook J M,Werren J H,et al.Wolbachia in two insect host-parasitoid communities[J].Molecular Ecology,1998,7(11):1457-1465.
[32]Casiraghi M,Bordenstein S R,Baldo L,et al.Phylogeny of Wolbachia pipientis based on glt A,gro EL and fts Z gene sequences:clustering of arthropod and nematode symbionts in the F supergroup,and evidence for further diversity in the Wolbachia tree[J].Microbiology,2005,151(12):4015-4022.