长角血蜱功能基因的筛选、克隆、表达及免疫功能的研究
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摘要
长角血蜱(Haemaphysalis longicornis)作为一种吸血性外寄生虫,是人和动物多种病原的重要传播媒介,可传播包括细菌、病毒、寄生虫、立克次氏体等引起的多种疫病,给我国畜牧业的发展带来严重威胁。目前对蜱的防控方法主要是化学药物防治,但随着蜱耐药性的产生以及药物残留、环境污染等公共卫生问题的出现,迫使人们寻求新的更为安全的防治措施。近年来免疫学防治方法成为人们的研究热点。而蜱功能基因的筛选、克隆及表达是免疫学防治的前提。
本实验用兔抗长角血蜱雌性成蜱阳性血清、兔抗长角血蜱幼蜱阳性血清、兔抗长角血蜱卵阳性血清对已构建好的长角血蜱饥饿雌蜱全蜱cDNA表达文库进行免疫学筛选,共获得11个有用的cDNA序列,依次编号为HL01-HL11。核酸序列以及氨基酸序列分析显示HL07与长角血蜱卵黄蛋白原的相似性高达99%,其余均为长角血蜱新基因,编码蛋白与青海血蜱Hq05、青海血蜱肌原蛋白T、肌球蛋白碱性轻链分子、微小牛蜱副肌球蛋白、肩突硬蜱表皮蛋白以及多种蜱的线粒体蛋白等具有相似性。目前已有5个序列登录到GenBank上,登陆号依次为:GU932666、GU932667、GU932668、GU932669、GU932670。选用大肠杆菌BL21(DE3)pLysE作为表达菌株,对已获得的8个阳性克隆(HL02、HL04、HL05、HL06、HL07、HL09、HL10、HL11)的重组质粒进行原核表达。结果显示,除HL10外,其他基因均表达。分别以兔抗长角血蜱雌性成蜱阳性血清、兔抗长角血蜱幼蜱阳性血清为一抗,利用Western-blot方法检测这些重组蛋白的反应性,结果发现HL02、HL04、HL05、HL06均表现出良好的反应原性。这为抗蜱疫苗的研究提供了更多的候选基因。
采用5’RACE技术成功获得了HL05 5’端未知序列,拼接后获得其全长。该基因全长1053bp,ORF含540个碱基,预测编码蛋白大小为20.01ku。生物信息学分析显示,该基因具备一个糖基化位点,有信号肽,属于分泌型蛋白,无跨膜结构,抗原指数高。将该基因全序列提交至GenBank,获得登录号为GQ499841。将重组质粒pSCREEN/HL05在大肠杆菌中成功进行大量表达,利用亲和层析法纯化重组蛋白,免疫动物,间接ELISA法检测抗体水平表明免疫效果良好,Western-blot方法检测蜱各阶段天然蛋白HL05的存在情况,结果显示该蛋白在幼蜱、若蜱、成蜱中均有表达,在卵中不存在。天然蛋白HL05可能具有3个亚型,大小分别是21ku、23 ku、55 ku。攻蜱实验表明该蛋白能够降低雌蜱的饱血体重,说明该基因能够诱导动物产生一定的抗蜱保护效应,可以作为抗蜱疫苗进行深入的研究。
Haemaphysalis longicornis are blood sucking ectoparasites and served as one of the most important vectors transmitting bacteria, viruses, parasites and rickettsia. It has been posing a serious threat to the animal husbandry in China. At present, although the principal tick control measure is chemical acaricides method, the development of alternative methods for tick control is important and urgent due to the appearance of acaricide-resistant ticks, environmental and food chain contamination. The use of tick vaccines has been shown to be the most promising alternative tick control method. The success of this method depends on the identification, cloning and in vitro expression of tick molecules which play a key physiological role in the survival of ticks and spread of disease.
In this study, a cDNA expression library of H. longicornis unfed female tick was screened with anti-H. longicornis unfed female tick serum, anti-H. longicornis larva serum and anti-H. longicornis egg serum produced from rabbit, respectively. Eleven genes encoding immunodominant antigens were obtained, and named for HL01 to HL11. The nucleotide and deduced amino acid sequence analysis showed that HL07 shared 99% similarity with H. longicornis vitellogenin, and the others including were novel gene of H. longicornis, which had significant homology with the H. qinghaiensis Hq05 gene, H. qinghaiensis myogenic protein T, H. qinghaiensis myosin alkali light chain molecules, Boophylus microplus paramyosin, Ixodes scapulars surface protein and mitochondria of a variety of tick, respectively. Five genes were submitted to GenBank, accession numbers were GU932666, GU932667, GU932668, GU932669, GU932670, respectively. Eight genes including HL02, HL04, HL05, HL06, HL07, HL09, HL10, HL11 were expressed in the Escherichia coli BL21(DE3)pLysE. All of them except HL10 were expressed successfully. The expression products of HL02, HL04, HL05 and HL06 gene revealed good reactogenicity by Western-blot with anti-H. longicornis unfed female tick serum and rabbit anti-H. longicornis larva serum. These will provide candidates for further screening of protective antigen genes of H. longicornis.
The HL05 gene was amplified by 5’rapid amplification of cDNA ends (5’RACE), and the full-length cDNA was obtained by splicing. The full-length of HL05 gene was 1053bp and contained an open reading frame (ORF) of 540 bp that codes for 179 amino acid residues with a coding capacity of 20.01ku. Bioinformatics analysis indicated that HL05 protein has a glycosylation site, it was secreted, no transmembrane domain and more potential antigenic determinants. The full-length sequence was submitted to GenBank and accession number was GQ499841. The recombinant plasmid pSCREEN/HL05 was expressed in E. coli BL21 (DE3), and expression products were 55 ku. Affinity chromatography was used to purify the recombinant protein, and the protein HL05 had good immunogenicity by indirect ELISA. Native HL05 was detected in ticks larvae, nymphs and adult except egg successfully, and it may have two isoforms which were 21ku, 23 ku. Vaccination of rabbit with rHL05 resulted in reduction of engorged weight of female ticks compared to the controls, and it conferred that rHL05 is a significant protective immunity. These results revealed that rHL05 could be a candidate vaccine molecule for the control of H. longicornis.
本实验用兔抗长角血蜱雌性成蜱阳性血清、兔抗长角血蜱幼蜱阳性血清、兔抗长角血蜱卵阳性血清对已构建好的长角血蜱饥饿雌蜱全蜱cDNA表达文库进行免疫学筛选,共获得11个有用的cDNA序列,依次编号为HL01-HL11。核酸序列以及氨基酸序列分析显示HL07与长角血蜱卵黄蛋白原的相似性高达99%,其余均为长角血蜱新基因,编码蛋白与青海血蜱Hq05、青海血蜱肌原蛋白T、肌球蛋白碱性轻链分子、微小牛蜱副肌球蛋白、肩突硬蜱表皮蛋白以及多种蜱的线粒体蛋白等具有相似性。目前已有5个序列登录到GenBank上,登陆号依次为:GU932666、GU932667、GU932668、GU932669、GU932670。选用大肠杆菌BL21(DE3)pLysE作为表达菌株,对已获得的8个阳性克隆(HL02、HL04、HL05、HL06、HL07、HL09、HL10、HL11)的重组质粒进行原核表达。结果显示,除HL10外,其他基因均表达。分别以兔抗长角血蜱雌性成蜱阳性血清、兔抗长角血蜱幼蜱阳性血清为一抗,利用Western-blot方法检测这些重组蛋白的反应性,结果发现HL02、HL04、HL05、HL06均表现出良好的反应原性。这为抗蜱疫苗的研究提供了更多的候选基因。
采用5’RACE技术成功获得了HL05 5’端未知序列,拼接后获得其全长。该基因全长1053bp,ORF含540个碱基,预测编码蛋白大小为20.01ku。生物信息学分析显示,该基因具备一个糖基化位点,有信号肽,属于分泌型蛋白,无跨膜结构,抗原指数高。将该基因全序列提交至GenBank,获得登录号为GQ499841。将重组质粒pSCREEN/HL05在大肠杆菌中成功进行大量表达,利用亲和层析法纯化重组蛋白,免疫动物,间接ELISA法检测抗体水平表明免疫效果良好,Western-blot方法检测蜱各阶段天然蛋白HL05的存在情况,结果显示该蛋白在幼蜱、若蜱、成蜱中均有表达,在卵中不存在。天然蛋白HL05可能具有3个亚型,大小分别是21ku、23 ku、55 ku。攻蜱实验表明该蛋白能够降低雌蜱的饱血体重,说明该基因能够诱导动物产生一定的抗蜱保护效应,可以作为抗蜱疫苗进行深入的研究。
Haemaphysalis longicornis are blood sucking ectoparasites and served as one of the most important vectors transmitting bacteria, viruses, parasites and rickettsia. It has been posing a serious threat to the animal husbandry in China. At present, although the principal tick control measure is chemical acaricides method, the development of alternative methods for tick control is important and urgent due to the appearance of acaricide-resistant ticks, environmental and food chain contamination. The use of tick vaccines has been shown to be the most promising alternative tick control method. The success of this method depends on the identification, cloning and in vitro expression of tick molecules which play a key physiological role in the survival of ticks and spread of disease.
In this study, a cDNA expression library of H. longicornis unfed female tick was screened with anti-H. longicornis unfed female tick serum, anti-H. longicornis larva serum and anti-H. longicornis egg serum produced from rabbit, respectively. Eleven genes encoding immunodominant antigens were obtained, and named for HL01 to HL11. The nucleotide and deduced amino acid sequence analysis showed that HL07 shared 99% similarity with H. longicornis vitellogenin, and the others including were novel gene of H. longicornis, which had significant homology with the H. qinghaiensis Hq05 gene, H. qinghaiensis myogenic protein T, H. qinghaiensis myosin alkali light chain molecules, Boophylus microplus paramyosin, Ixodes scapulars surface protein and mitochondria of a variety of tick, respectively. Five genes were submitted to GenBank, accession numbers were GU932666, GU932667, GU932668, GU932669, GU932670, respectively. Eight genes including HL02, HL04, HL05, HL06, HL07, HL09, HL10, HL11 were expressed in the Escherichia coli BL21(DE3)pLysE. All of them except HL10 were expressed successfully. The expression products of HL02, HL04, HL05 and HL06 gene revealed good reactogenicity by Western-blot with anti-H. longicornis unfed female tick serum and rabbit anti-H. longicornis larva serum. These will provide candidates for further screening of protective antigen genes of H. longicornis.
The HL05 gene was amplified by 5’rapid amplification of cDNA ends (5’RACE), and the full-length cDNA was obtained by splicing. The full-length of HL05 gene was 1053bp and contained an open reading frame (ORF) of 540 bp that codes for 179 amino acid residues with a coding capacity of 20.01ku. Bioinformatics analysis indicated that HL05 protein has a glycosylation site, it was secreted, no transmembrane domain and more potential antigenic determinants. The full-length sequence was submitted to GenBank and accession number was GQ499841. The recombinant plasmid pSCREEN/HL05 was expressed in E. coli BL21 (DE3), and expression products were 55 ku. Affinity chromatography was used to purify the recombinant protein, and the protein HL05 had good immunogenicity by indirect ELISA. Native HL05 was detected in ticks larvae, nymphs and adult except egg successfully, and it may have two isoforms which were 21ku, 23 ku. Vaccination of rabbit with rHL05 resulted in reduction of engorged weight of female ticks compared to the controls, and it conferred that rHL05 is a significant protective immunity. These results revealed that rHL05 could be a candidate vaccine molecule for the control of H. longicornis.
引文
1.白启,刘光远,韩根凤.甘肃张家川牛卵形巴贝斯虫的分离及补充传播试验[J].中国兽医科技,1994,24(9):12-15.
2.白启,刘光远,韩根凤.卵形巴贝斯虫在长角血血蜱饱血雌虫及其卵内的发育形态观察[J].中国农业科学,1996,29 (1):89-92.
3.白启,刘光远,张林.牛血孢子虫单一种的分离技术及其保藏方法的研究[J].卵形巴贝斯虫的传播实验.畜牧兽医学报,1992 ,23 (2):166-171.
4.柴慧萍,刘光远,张林,等.长角血蜱饥饿雌蜱cDNA表达文库的构建及免疫学筛选[J].中国寄生虫与寄生虫病学杂志,2009,27(1):6-10.
5.程天印,周金林,周勇志,等.亚洲璃眼蜱唾液腺差异表达基因文库的构建与分析[J].中国预防兽医学报,2006,3(28):261-266.
6.程远国,吴厚永,李德昌,等.长角血蜱唾液腺中腺苷三磷酸双磷酸酶的纯化及其酶切机制[J].昆虫学报,2002, 45 (3) :307-311.
7.程远国,吴厚永,李德昌,等.长角血蜱唾液腺Apyrase的生物学特性和功能的研究[J].寄生虫与医学昆虫学报,2000,7(3):175-182.
8.程远国,吴厚永,李德昌.长角血蜱唾液腺中胶原特异性血小板聚集抑制因子的纯化及机制的研究[J].中国科学(C辑),1999,29(6):587-591.
9.邓国藩,姜在阶.中国经济昆虫志(第39册) :蜱螨亚纲硬蜱科[M].北京:科学出版社,1991:
2.
10.刘光远,谢俊仁,田占成,等.长角血蜱雌蜱唾液腺cDNA表达文库的免疫筛选与初步分析,甘肃农业大学学报,2008,43(1):18～24.
11.刘建刚,刘光远,田占成,等.长角血蜱雄蜱抑制消减杂交cDNA文库的构建与分析[J].中国兽医科学,2009,39(08):659-664.
12.李有全.我国羊泰勒虫的生物学特性研究[李有全博士学位论文].北京:中国农业科学院,2007.
13.李金杰.青海血蜱cDNA表达文库的免疫筛选及基因克隆的表达[李金杰硕士学位论文] .北京:中国农业科学院,2008.
14.李晓明,张志平,杨小龙,等.长角血蜱卵黄蛋白单克隆抗体的制备及鉴定[J].昆虫学报,2008,51(10):1028-1032.
15.马腾.镰形扇头蜱肌钙蛋白Ⅰ基因的克隆及表达[马腾硕士学位论文].北京:中国农业科学院,2006.
16.邱咏梅,夏庆友.EST规模测序在家蚕功能基因组中的应用[J].蚕学通讯,2002 ,22 (4) :16-19.
17.宋广忠,石君帆.寄生蠕虫副肌球蛋白研究进展[J].中国人兽共患病学报,2006, 22 (10):989-991.
18.萨姆布鲁克J.,拉塞尔D W.分子克隆实验指南,3版(,黄培堂,王嘉玺,朱厚础,等)[M]北京:科学出版社,2002 :1193-1194.
19.谢俊仁,刘光远,田占成,等.长角血蜱雌蜱唾液腺cDNA表达文库的构建[J].甘肃农业大学学报,2007,42(6):13-17.
20.王玉霞,周金林,沈杰.蜱功能基因研究进展[J].动物医学进,2005,26(7):31-35.
21.王光雷.蜱害引起家畜大批死亡的调查报告.草食家畜,1997,2:43
22.温进坤,韩梅.Western印记,医学分子生物学理论与研究技术[M].北京:科学出版社,2002,228-238.
23.吴乃虎.基因工程原理[M].北京:科学出版社,第二版,1998,208.
24.向飞宇,周金林,周勇志,等.镰形扇头蜱抑制消减杂交文库的构建与分析[J].中国农业科学,2006,39(11):2347-2353.
25.杨成君,王军.cDNA文库的构建策略及其应用[J].生物技术通报,2007,1:5-9.
26.闫晓菲,韩红玉,岳城,等.cDNA文库及其在原虫研究中的应用[J].生物技术通报,2008,1:52-55.
27.杨小龙,高志华,胡永红,等.长角血蜱卵黄蛋白的纯化及其性质[J].昆虫学报,2004,47(3):316-319.
28.殷宏,吕文顺,张其才,等.甘肃省牛羊部分蜱传性血液原虫的传播实验[J].中国兽医寄生虫病杂志,2000,8 (1) :17-19.
29.赵忠芳,孔繁瑶,等.兔非特异性免疫力抗长角血蜱幼虫感染能力的研究[J].中国农业大学学报,1998,(增刊):79-81.
30.赵海平.青海血蜱幼蜱cDNA表达文库的构建及其免疫学筛选[赵海平硕士学位论文] .北京:中国农业科学院,2007.
31.张宝.吕氏泰勒虫疫苗候选抗原的免疫筛选及其免疫功能的研究[张宝硕士学位论文].北京:中国农业科学院,2009.
32.张维谊.镰形扇头蜱唾液腺cDNA文库的构建及其表达序列标签的研究[张维谊硕士学位论文].南宁:广西大学, 2004.
33.张弗盈,王文杰.膜融合中的SNAREs及其相关蛋白研究进展[J].医学综述2004,10(12).
34. Alekseev AN and Chunikhin SP. The fractions, functions, methods of isolation and the significance for transmission of causative agents of the saliva of ixodid ticks[J]. Med Parazitol (Mosk) , 1989, (4) : 3～9.
35. Aljamali MN, Sauer JR, Essenberg RC. RNA interference: applicability in tick research[J]. Exp Appl Acarol, 2002, 28(124): 89-96.
36. Alim MA, Tsujia N, Miyoshi T, et al. Characterization of asparaginyl endopeptidase, legumain induced byblood feeding in the ixodid tick Haemaphysalis longicornis[J]. Insect Biochem Mol Biol, 2007, 37: 911–922.
37. Alim MA, Tsuji N, Miyoshi T, et al. HlLgm2, a member of asparaginyl endopeptidases/legumains in the midgut of the ixodid tick Haemaphysalis longicornis, is involved in blood-meal digestion[J]. J Insect Physiol, 2008, 54(3):573–585.
38. Alim MA, Tsuji N, Miyoshi T, et al. Developmental Stage- and Organ-Specific Expression Profilesof Asparaginyl Endopeptidass / Legumains in the Ixodid Tick Haemaphysalis longicornis[J]. J Vet Med Sci, 2008, 70(12): 1363–1366.
39. Alim MA, Tsuji N, Miyoshi T et al. Legumains from the hard tick Haemaphysalis longicornis play modulatory roles in blood feeding and gut cellular remodelling and impact on embryogenesis[J]. Int J Parasitol, 2009, 39(1):97–107.
40. Alim MA, Tsuji N, Miyoshi T, et al. Legumains from the hard tick Haemaphysalis longicornis play modulatory roles in blood feeding and gut cellular remodelling and impact on embryogenesis[J]. Int J Parasitol. 2009, 39: 97-107.
41. Attardo GM, Higgs S, Klingler KA, et al. RNA interference-mediated knockdown of a GATA factor reveals a link to anautogeny in the mosquito Aedes aegypti[J]. Proc Natl Acad Sci USA, 2003, 100, 13374–13379.
42. Bates SM and Weitz JI. The status of new anticoagulants[J]. Br.J. Haematol. 2006, 134: 3–19.
43. Boldbaatar D, Sikalizyo Sikasunge C, Battsetseg B, et al. Molecular cloning and functional characterization of an aspartic protease from the hard tick Haemaphysalis longicornis[J].Insect Biochem. Mol. Biol. 2006, 36:25-36.
44. Boisson B, Jacques J C, Choumet V, et al. Gene silencing in mosquito salivary glands by RNAi[J]. FEBS Lett. 2006, 580, 1988–1992.
45. Boutros M, Kiger AA, Armknecht S, et al. Heidelberg Fly Array Consortium, Genome-wide RNAi analysis of growth and viability in Drosophila cells[J]. Science. 2004, 303, 832–835.
46. Bullard B, Leonard K, Larkins A, et al. Troponin of asynchronous flight muscle[J]. J Mol Biol. 1988, 204, 621–637.
47. Chen HG, Nara T, Zeng XJ, et al. Hirayama vaccination of domestic pig with recombinant paramyosin against Schistosoma japonicum in China[J]. Vaccine, 2000, 18 : 214-26.
48. Ceraul SM, Sonenshine DE, Ratzlaff RE, et al. An art hropod defensin expressed by the hemocytes of the American dog tick, Dermacentor variabilis ( Acari: Ixodidae )[J]. Insect Biochem Mol Biol, 2003, 33 (11): 1099-1103.
49. Chen A and Holmes S P. Molecular cloning and functional expression of a serotonin receptor from the Southern cattle tick, Boophilus microplus (Acari: Ixodidae) [J]. Insect Mol Biol, 2004, 13(1): 45-54.
50. Ceraul S M, SM, Dreher-Lesnick SM, Gillespie JJ, et al. New tick defensin isoform and antimicrobial gene expression in response to Rickettsia montanensis challenge[J]. Infect Immun. 2007, 75:1973–1983.
51. de la Fuente J and Kocan KM. Advances in the identification and characterization of protective antigens for development of recombinant vaccines against tick infestations[J]. Expert Rev Vaccines.2003, 2: 583–593.
52. de la Fuente J and Kocan KM. Strategies for development of vaccines for control of ixodid tick species[J]. Parasite Immunol, 2006, 28(7): 275–283.
53. de la Fuente J, Kocan KM, Blouin EF. Tick Vaccines and the Transmission of Tick-BornePathogens[J]. Vet Res Commun, 2007, 31(1): 85–90.
54. da Silva Vazl Jr, Torino Lermen T, Michelon A, er al.Effect of acaricides on the activity of a Boophilus microplus glutathione S-transferase[J]. Vet Parasitol, 2004, 119 (223): 237-245.
55. Emini EA, Hughes JV, Perlow DS, et al. Induction of hepatitis A virus-neutralizing antibody by a virus specific synthetic peptide[J]. J Virol, 1985, 55(3): 836-839.
56. Estrada-Pe?a A and Jongejan F. Ticks feeding on humans: a review of records on humanbiting Ixodoidea with special reference to pathogen transmission[J]. Exp Appl Acarol, 1999, 23(9): 685–715.
57. Ferreira CA, Barbosa MC, Silveira TC, et al. cDNA cloning, expression and characterization of a Boophilus microplus paramyosin[J]. Parasitology, 2002, 125(3): 265-274.
58. Fogaca AC, Lorenzini DM, Kaku LM, et al. Cysteine-rich antimicrobial peptides of the cattle tick Boophilus microplus: isolation, structural characterization and tissue expression profile[J]. Dev Comp Immunol, 2004, 28(3): 191–200.
59. Francischetti IM, Mather TN , Ribeiro JM. Tick saliva is a potent inhibitor of endothelial cell proliferation and angiogenesis[J]. Thromb Haemost, 2005, 94(1): 167–174.
60. Fyrberg E, Fyrberg CC, Beall C, et al. Drosophila melanogaster troponin-T mutations engender three distinct syndromes of myofibrillar abnormalities. J Mol Biol., 1990, 216(3), 657–675.
61. Gao J, Luo J, Fan R, et al. Molecular characterization of a myosin alkali light chain-like protein, a“concealed”antigen from the hard tick Haemaphysalis qinghaiensis[J]. Vet Parasitol, 2007, 147(1-2):140–149.
62. Gao J, Luo J, Fan R, et al. Characterization of a concealed antigen Hq05 from the hard tick Haemaphysalis qinghaiensis and its effect as a vaccine against tick infestation in sheep[J]. Vaccine, 2009, 27(3):483-490.
63. Gong H, LiaoM, Zhou J, et al. Gene silencing of ribosomal protein P0 is lethal to the tick Haemaphysalis longicornis[J]. Veterinary Parasitology, 2007, 151: (2008) 268–278.
64. Gong H, Umemiya R, Zhou J, et al. Blocking the secretion of saliva by silencing the HlYkt6 gene in the tick Haemaphysalis longicorni[J]s. Insect Biochem Mol Biol, 2009, 39(5-6): 372–381.
65. Graf JF, Gogolewski R, Leach-Bing N, et al. Tick control: an industry point of view[J]. Parasitology, 2004, 129: S427–442.
66. GrunclováL, Horn M, VancováM, et al. Two secreted cystatins of the soft tick Ornithodoros moubata: differential expression pattern and inhibitory specificity[J]. Biol Chem, 2006, 387(12): 1635–1644.
67. Guilfoile PG and Packila M. Identification of four genes expressed by feeding female Ixodes scapularis, including three with sequence similarity to previously recognized genes[J]. Exp Appl Acarol, 2004, 32 (122): 103-110.
68. Hatta T, Kazama K, Miyoshi T, et al. Identification and characterisation of a leucine aminopeptidase from the hard tick Haemaphysalis longicornis[J]. Int J Parasitol, 2006, 36(10-11): 1123–1132.
69. Hatta T, Tsuji N, Miyoshi T, et al. Leucine Aminopeptidase in the Ixodid Tick Haemaphysalis longicornis: Endogenous Expression Profiles in Midgut[J]. J Vet Med Sci, 2009, 71(5): 589–594.
70. Hatta T, Umemiya R, Liao M, et al. RNA interference of cytosolic leucine aminopeptidase reduces fecundity in the hard tick, Haemaphysalis longicornis[J]. Parasitol Res, 2007, 100(4):847–854.
71. Horn M, NussbaumerováM, Sanda M, et al. Hemoglobin digestion in blood-feeding ticks: mapping a multipeptidase pathway by functional proteomics[J]. Chem Biol, 2009, 16(10):1053-1063.
72. Hynes WL, Ceraul SM, Todd SM, et al. A defensin-like gene expressed in the black-legged tick, Ixodes scapularis[J]. Med Vet Entomol, 2005, 19(4): 339–344.
73. Imamura S, da Silva Vaz Junior l, Sugino M, et al. A serine protease inhibitor (serpin) from Haemaphysalis longicornis as an anti-tick vaccine[J]. Vaccine, 2005, 23(10): 1301–1311.
74. Imamura S, da Silva Vaz Jr Itabajara, Konnai S, et al. Effect of vaccination with a recombinant metalloprotease from Haemaphysalis longicornis[J]. Exp Appl Acarol, 2009, 48(4): 345–358.
75. Islam MK, Tsuji N, Miyoshi T, et al. The Kunitz-Like Modulatory Protein Haemangin Is Vital for Hard Tick Blood-Feeding Success[J]. PLoS Pathog. 2009, 5(7):1-13.
76. Jameson BA, Wolf H. The antigenic index: a novel algorithm for prediction antigenic determinants[J]. Comput Appl Biosci, 1988, 4(1):181-186.
77. Jaworski DC. Tick‘‘talk’’: protein release by tick salivary cells[J]. Trends Parasitol. 2003, 19(10): 427–429.
78. Jaworski DC, J asinskas A, Metz CN. Identification and characterization of a homologue of the proinflammatory cytokine macrophage migration inhibitory factor in the tick amblyomma americanum[J]. Insect Mol Biol, 2001, 10 (4): 323-331.
79. Karim S, Miller NJ, Valenzuela J, et al. RNAi-mediated gene silencing to assess the role of synaptobrevin and cystatin in tick blood feeding[J]. Biochem Biophys Res Commu, 2005, 334(4) :1336– 1342.
80. Konnai S, Nakajima C, Imamura S, et al. Suppression of cell proliferation and cytokine expression by HL-p36, a tick salivary gland-derived protein of Haemaphysalis longicornis[J]. Immunology, 2009, 126(2): 209–21.
81. Kotsyfakis M, Sá-Nunes A, Francischetti IM, et al. Anti-inflammatory and immunosuppressive activity of sialostatin L, a salivary cystatin from the tick Ixodes scapularis[J]. J Biol Chem, 2006, 281:26298–26307.
82. Kotsyfakis M, Karim S, Andersen JF, et al. Selective cysteine protease inhibition contributes to blood-feeding success of the tick Ixodes scapularis[J]. J Biol Chem, 2007, 282(40):29256–29263.
83. Kyte J and Doolittle RF. A simple method for displaying the hydropathic character of a protein[J]. J Mol Biol, 1982, 157(1):105-132.
84. Lai R, Lomas LO, Jonczy J, et al. Two novel non-cationic defensin-like antimicrobial peptides from haemolymph of the female tick, Amblyomma hebraeum[J]. Biochem J, 2004, 379(Pt3): 681-685.
85. Levy O. Antimicrobial proteins and peptides of blood: templates for novel antimicrobial agents[J]. Blood, 2000, 96(8):2664–2672.
86. Liao M, Zhou J, Hatta T, et al. Molecular characterization of Rhipicephalus (Boophilus) microplus Bm86 homologue from Haemaphysalis longicornis ticks[J]. Vet Parasitol, 2007, 146 (1-2):148-157.
87. Liao M, Hatta T, Umemiya R, et al. Identification of three protein disulfide isomerase members from Haemaphysalis longicornis tick Insect[J]. Biochem Mol Biol, 2007, 37: 641–654.
88. Liao M, Boldbaatar D, Gong H, et al. Functional analysis of protein disulfide isomerases in blood feeding, viability and oocyte development in Haemaphysalis longicornis ticks[J]. Insect Biochem Mol Bioly, 2008, 38(3): 285–295.
89. Lima CA, Sasaki SD, Tanaka AS. Bmcystatin, a cysteine proteinase inhibitor characterized from the tick Boophilus microplus[J]. Biochem Biophys Res Commun, 2006, 347(1): 44–50.
90. Meiringer CT, Auffarth K, Hou H, et al. Depalmitoylation of Ykt6 prevents its entry into the multivesicular body pathway[J]. Traffic, 2008, 9(9): 1510–1521.
91. Martín Hernández R, Cuellar del Hoyo C, Olmeda Garcia AS, et al. Analysis of stage-specific and shared antigens derived from Rhipicephalus sanguineus by electrophoresis and western blotting[J]. Med Vet Entomol, 1995, 9: 358~364.
92. Marden JH, Fitzhugh GH, Girgenrath M, et al. Alternative splicing, muscle contraction and intraspecific variation: associations between troponin T transcripts, Ca(2+) sensitivity and the force and power output of dragonfly flight muscles during oscillatory contraction[J]. J Exp Biol, 2001, 204(20), 3457–3470.
93. Marden JH, Fitzhugh GH, Wolf MR, et al. Alternative splicing, muscle calcium sensitivity and the modulation of dragonfly flight performance[J]. Proc Natl Acad Sci USA, 1999, 96(26):15304–15309.
94. Miyoshi T, Tsuji N, Islam MK, et al. A set of serine proteinase paralogs are required for blood-digestion in the ixodid tick Haemaphysalis longicornis[J]. Parasitol Int, 2008, 57(4): 499–505.
95. Miyoshi T, Tsuji N, Islam MK, et al. A Kunitz-type proteinase inhibitor from the midgut of the ixodid tick, Haemaphysalis longicornis, and its endogenous target serine proteinase[J]. Mol Biochem Parasitol, 2010, 170(2):112–115.
96. Minami and Tshihara T. Babesia ovata sp. isolate fromcattle in Japan[J]. Natl Inst Anim Health, 1980, 20(3):101-110.
97. Motobu M, Tsuji N, T Miyoshi, et al. Molecular characterization of a blood-induced serine carboxypeptidase from the ixodid tick Haemaphysalis longicornis[J]. FEBS J, 2007, 274: 3299–3312.
98. Mulenga C, Sugimoto M, Onuma. Issues in tick vaccine development: identification and characterization of potential candidate vaccine antigens[J]. Microbes Infect. 2000, 2: 1353–1361.
99. Mulenga A, Sugimoto C, Sako Y, et al. Molecular characterization of a Haemaphysalis longicornis tick salivary gland associated 29-kilodalton protein and its effectas a vaccine against tick infestation in rabbits[J]. Infec Immun, 1999, 67 (4): 1652-1658.
100.Mulenga A, Sugimoto C, Ingram G, et al. Characterization of two cDNAs encoding serineproteinases from the hard tick Haemaphysalis longicornis[J]. Insect Biochemd Mol Biol, 2001, 31(8):817-825.
101.Nuttall PA, Trimnell AR, Kazimirova M, et al. Exposed and concealed antigens as vaccine targets for controlling ticks and tick-borne diseases[J]. Parasite Immunol, 2006, 28(4),155–163.
102.Nakajima Y, Van der Goes Van Naters-Yasui A, Taylor D, et al. Antibacterial peptide defensin is involved in midgut immunity of the soft tick Ornithodoros moubata (Acari :Argasidae)[J]. Insect Mol Biol, 2002, 11 (6): 611-618.
103.Nakajima Y, Van der Goes Van Naters-Yasui A, Taylor D, et al. Two isoforms of a member of the arthropod defensin family from the soft tick Ornithodoros moubata (Acari :Argasidae)[J]. Insect Biochem Mol Biol, 2001, 31 (8): 747-751.
104.Ohnishi A, Hull JJ, Matsumoto S. Targeted disruption of genes in the Bombyx mori sex pheromone biosynthetic pathway[J]. Proc Natl Acad Sci U S A, 2006, 103(12): 4398–4403.
105.Perry SV. Troponin T: genetics, properties and function[J]. J Muscle Res Cell Motil, 1998, 19(6): 575–602.
106.Parola P and Raoult D. Tick-borne bacterial diseases emerging in Europe[J]. Clin Microbiol Infec, 2001, 7: 80–83.
107.Rossi V, Banfield DK, Vacca M, et al. Longins and their longin domains: regulated SNAREs and multifunctional SNARE regulators[J]. Trends Biochem Sci, 2004, 29(12): 682–688.
108.Rothel JS, Wood PR, Seowf HF, et al. Urea/DTT solubiliiation of a recombinant Taenia ovis antigen, 45W, expressed as a GST fusion protein results in enhanced protective immune response to the 45W moiety[J]. Vaccine, 1997, 15(5): 469-472.
109.Rudenko N, Golovchenko M, Edwards MJ, et al. Differential expression of Ixodes ricinus tick genes induced by blood feeding or Borrelia burgdorferi infection[J]. J Med Entomol, 2005, 42(1): 36–41.
110.Sauer JR, M cswain JL, Bowman AS, et al. Tick salivary glandphysiology[J]. Annu Rev Entomol, 1995, 40: 245-267.
111.Selsted ME and Ouellette AJ. Mammalian defensins in the antimicrobial immune response[J]. Nat Immunol, 2005, 6(6):551–557.
112.Shahein YE. Molecular cloning and expression of a larval immunogenic protein from the cattle tick Boophilus annulatus[J]. Vet Immunol Immunopathol, 2008, 121(3-4): 281-289.
113.Silverman GA, Bird Pl, Carrell RW,et al. The serpins are an expanding superfamily of structurally similar but functionally diverse proteins: evolution, mechanism of inhibition, novel functions, and a revised nomenclature[J]. J Biol Chem, 2001, 276(36): 33293–33296.
114.Sojka D, Hajdusek O, Dvorák J et al. IrAE: an asparaginyl endopeptidase (legumain) in the gut of the hard tick Ixodes ricinus[J]. Int J Parasitol, 2007, 37(7): 713-724.
115.Sojka D, Franta Z, Horn M, et al. Profiling of proteolytic enzymes in the gut of the tick Ixodes ricinus reveals an evolutionarily conserved network of aspartic and cysteine peptidases[J]. Parasit Vectors, 2008, 1(1):7.
116.Solis CF, Vázquez-Talavera J, Laclette JP. Toward development of a Taenia solium paramyosin based vaccine against porcine cysticercosis[J]. Gac Med Mex, 2004, 140(2): 129-38.
117.Sonenshine DE. Biology of Ticks, vol. 1(M). New York: Oxford University Press, 1991.
118.Sonenshine DE, CeraulSM, Hynes WE, et al. Expression of defensin-like peptides in tick hemolymph and midgut in response to challenge with Borrelia burgdorferi, Escherichia coli and Bacillus subtilis[J]. Exp Appl Acarol, 2002, 28(1-4): 127–134.
119.Sugino M, Imamura S, Mulenga A, et al. A serine proteinase inhibitor (serpin) from ixodid tick Haemaphysalis longicornis cloning and preliminary assessment of its suitability as a candidate for a tick vaccine[J]. Vaccine, 2003, 21(21-22): 2844–2851.
120.SuginoM, Imamura S, Mulenga A, et al. A serine proteinase inhibitor (serpin ) from ixodid tick Haemaphysalis longicornis: cloning and preliminary assessment of its suitability as a candidatefor a tick vaccine[J]. Vaccine, 2003, 21: 2844-2851.
121.Taylor MG, Huggins MC, Shi F, et al. Production and testing of Schistosoma japonicum candidate vaccine antigens in the natural ovine host. Vaccine, 1998, 16(13): 1290-8.
122.Temeyer KB, Davey RB, Chen AC. Identification of a third Boophilus microplus (Acari: Ixodidae) cDNA presumptively encoding anacetyl cholinesterase[J]. J Med Entomol, 2004, 41 (3): 259-268.
123.Tsuji N and Fujisaki K. Longcin plays a crucial role inhibiting the transmission of babesia parasites in the vector tick Hamaphysalis longicornis[J]. Future Microbiol, 2007, 2(6): 575-578.
124.Tsuji N, Battsetseg B, Boldbaatar D, et al. Babesial Vector Tick Defensin against Babesia sp. Parasites[J]. Infect Immun, 2007, 75(7): 3633–3640.
125.Tsuda A, Mulenga A, Sugimoto C, et al. cDNA cloning, characterization and vaccine effect analysis of Haemaphysalis longicornis tick saliva proteins[J]. Vaccine, 2001, (19) : 4287～4296.
126.Tsuji N, Kamio T, Isobe T, et al. Molecular characterization of a peroxiredoxin from the hard tick Haemaphysalis longicornis[J]. Insect Mol Biol, 2001,10(2): 121-129.
127.Tsuji N, Miyoshi T, Battsetseg B, et al. a Cysteine Protease is Critical for Babesia spp. Transmission in Haemaphysalis ticks[J]. PLoS Pathog, 2008, 4(5): 1-14.
128.Valenzuela JG. Exploring tick saliva: from biochemistry to‘sialome’and functional genomics[J]. Parasitology, 2004, 129: S83–S94.
129.Vázquez-Talavera J, Solís CF, Terrazas LI, et a1. Characterization and protective potential of the immune response to Taenia solium paramyosin in a murine model of cysticercosis[J]. Infect Immun, 2001, 69(9): 5412-6.
130.Willadsen P. Anti-tick vaccines[J]. Parasitology, 2004, 129: S367–387.
131.Willadsen P. Tick control: Thoughts on a research agenda[J]. Vet Parasitol, 2006, 138(1-2): 161–168.
132.Xu G, Fang QQ, Keirans JE. Cloning and sequencing of putative calreticulin complementary DNAs from four hard tick species[J]. J Parasitol, 2004, 90 (1): 73-78.
133.Yamaji K, Tsuji N, Miyoshi T, et al. A salivary cystatin, HlSC-1, from the ixodid tick Haemaphysalis longicornis play roles in the blood feeding processes[J]. Parasitol Res, 2009,106(1):61-68.
134.Yamaji K, Tsuji N, Miyoshi T, et al. Hemoglobinase activity of a cysteine protease from the ixodid tick Haemaphysalis longicornis[J]. Parasitol Int, 2009, 58:232-237.
135.Yamaji K, Tsuji N, Miyoshi T, et al. Hlcyst-1 and Hlcyst-2 are potential inhibitors of HlCPL-A in the midgut of the ixodid tick Haemaphysalis longicornis[J]. J Vet Med Sci, 2010, 72(5):599-604.
136.You M and Fujisaki K. Vaccination Effects of Recombinant Chitinase Protein from the Hard Tick Haemaphysalis longicornis (Acari: Ixodidae)[J]. J Vet Med Sci, 2009, 71(6): 709–712.
137.You M, Xuan X, Tsuji N, et al. Molecular characterization of a troponin I-like protein from the hard tick Haemaphysalis longicornis[J]. Insect Biochem Mol Biol, 2001, 32(1): 67-73.
138.You M, Xuan X, Tsuji N, et al. Identification and molecular characterization of a chitinase from the hard tick Haemaphysalis longicornis[J]. J Biol Chem, 2003, 278(10): 8556–8563.
139.Zhou J, Ueda M, Umemiya R, et al. A secreted cystatin from the tick Haemaphysalis longicornis and its distinct expression patterns in relation to innate immunity[J]. Insect Biochem Mol Biol, 2006, 36:527–535.
140.Zhou J, Liao M, Hatta T, et al. Identification of a follistatin-related protein from the tick Haemaphysalis longicornis and its effect on tick oviposition[J]. Gene, 2006, 372, 191–198.
141.Zhou J, Liao M, Ueda M, et al. Sequence characterization and expression patterns of two defensin-like antimicrobial peptides from the tick Haemaphysalis longicornis[J]. Peptides, 2007, 28 (6) :304-310.
142.Zhou J, Liao M, Ueda M, et al. Characterization of an intracellular cystatin homolog from the tick Haemaphysalis longicornis. Vet Parasitol, 2009, 160: 180–183.
143.Zhou J, Liao M, Gong H, et al. Characterization of Hlcyst-3 as a member of cystatins from the tick Haemaphysalis longicornis[J]. Exp Appl Acarol, 2010, Jan 27.
2.白启,刘光远,韩根凤.卵形巴贝斯虫在长角血血蜱饱血雌虫及其卵内的发育形态观察[J].中国农业科学,1996,29 (1):89-92.
3.白启,刘光远,张林.牛血孢子虫单一种的分离技术及其保藏方法的研究[J].卵形巴贝斯虫的传播实验.畜牧兽医学报,1992 ,23 (2):166-171.
4.柴慧萍,刘光远,张林,等.长角血蜱饥饿雌蜱cDNA表达文库的构建及免疫学筛选[J].中国寄生虫与寄生虫病学杂志,2009,27(1):6-10.
5.程天印,周金林,周勇志,等.亚洲璃眼蜱唾液腺差异表达基因文库的构建与分析[J].中国预防兽医学报,2006,3(28):261-266.
6.程远国,吴厚永,李德昌,等.长角血蜱唾液腺中腺苷三磷酸双磷酸酶的纯化及其酶切机制[J].昆虫学报,2002, 45 (3) :307-311.
7.程远国,吴厚永,李德昌,等.长角血蜱唾液腺Apyrase的生物学特性和功能的研究[J].寄生虫与医学昆虫学报,2000,7(3):175-182.
8.程远国,吴厚永,李德昌.长角血蜱唾液腺中胶原特异性血小板聚集抑制因子的纯化及机制的研究[J].中国科学(C辑),1999,29(6):587-591.
9.邓国藩,姜在阶.中国经济昆虫志(第39册) :蜱螨亚纲硬蜱科[M].北京:科学出版社,1991:
2.
10.刘光远,谢俊仁,田占成,等.长角血蜱雌蜱唾液腺cDNA表达文库的免疫筛选与初步分析,甘肃农业大学学报,2008,43(1):18～24.
11.刘建刚,刘光远,田占成,等.长角血蜱雄蜱抑制消减杂交cDNA文库的构建与分析[J].中国兽医科学,2009,39(08):659-664.
12.李有全.我国羊泰勒虫的生物学特性研究[李有全博士学位论文].北京:中国农业科学院,2007.
13.李金杰.青海血蜱cDNA表达文库的免疫筛选及基因克隆的表达[李金杰硕士学位论文] .北京:中国农业科学院,2008.
14.李晓明,张志平,杨小龙,等.长角血蜱卵黄蛋白单克隆抗体的制备及鉴定[J].昆虫学报,2008,51(10):1028-1032.
15.马腾.镰形扇头蜱肌钙蛋白Ⅰ基因的克隆及表达[马腾硕士学位论文].北京:中国农业科学院,2006.
16.邱咏梅,夏庆友.EST规模测序在家蚕功能基因组中的应用[J].蚕学通讯,2002 ,22 (4) :16-19.
17.宋广忠,石君帆.寄生蠕虫副肌球蛋白研究进展[J].中国人兽共患病学报,2006, 22 (10):989-991.
18.萨姆布鲁克J.,拉塞尔D W.分子克隆实验指南,3版(,黄培堂,王嘉玺,朱厚础,等)[M]北京:科学出版社,2002 :1193-1194.
19.谢俊仁,刘光远,田占成,等.长角血蜱雌蜱唾液腺cDNA表达文库的构建[J].甘肃农业大学学报,2007,42(6):13-17.
20.王玉霞,周金林,沈杰.蜱功能基因研究进展[J].动物医学进,2005,26(7):31-35.
21.王光雷.蜱害引起家畜大批死亡的调查报告.草食家畜,1997,2:43
22.温进坤,韩梅.Western印记,医学分子生物学理论与研究技术[M].北京:科学出版社,2002,228-238.
23.吴乃虎.基因工程原理[M].北京:科学出版社,第二版,1998,208.
24.向飞宇,周金林,周勇志,等.镰形扇头蜱抑制消减杂交文库的构建与分析[J].中国农业科学,2006,39(11):2347-2353.
25.杨成君,王军.cDNA文库的构建策略及其应用[J].生物技术通报,2007,1:5-9.
26.闫晓菲,韩红玉,岳城,等.cDNA文库及其在原虫研究中的应用[J].生物技术通报,2008,1:52-55.
27.杨小龙,高志华,胡永红,等.长角血蜱卵黄蛋白的纯化及其性质[J].昆虫学报,2004,47(3):316-319.
28.殷宏,吕文顺,张其才,等.甘肃省牛羊部分蜱传性血液原虫的传播实验[J].中国兽医寄生虫病杂志,2000,8 (1) :17-19.
29.赵忠芳,孔繁瑶,等.兔非特异性免疫力抗长角血蜱幼虫感染能力的研究[J].中国农业大学学报,1998,(增刊):79-81.
30.赵海平.青海血蜱幼蜱cDNA表达文库的构建及其免疫学筛选[赵海平硕士学位论文] .北京:中国农业科学院,2007.
31.张宝.吕氏泰勒虫疫苗候选抗原的免疫筛选及其免疫功能的研究[张宝硕士学位论文].北京:中国农业科学院,2009.
32.张维谊.镰形扇头蜱唾液腺cDNA文库的构建及其表达序列标签的研究[张维谊硕士学位论文].南宁:广西大学, 2004.
33.张弗盈,王文杰.膜融合中的SNAREs及其相关蛋白研究进展[J].医学综述2004,10(12).
34. Alekseev AN and Chunikhin SP. The fractions, functions, methods of isolation and the significance for transmission of causative agents of the saliva of ixodid ticks[J]. Med Parazitol (Mosk) , 1989, (4) : 3～9.
35. Aljamali MN, Sauer JR, Essenberg RC. RNA interference: applicability in tick research[J]. Exp Appl Acarol, 2002, 28(124): 89-96.
36. Alim MA, Tsujia N, Miyoshi T, et al. Characterization of asparaginyl endopeptidase, legumain induced byblood feeding in the ixodid tick Haemaphysalis longicornis[J]. Insect Biochem Mol Biol, 2007, 37: 911–922.
37. Alim MA, Tsuji N, Miyoshi T, et al. HlLgm2, a member of asparaginyl endopeptidases/legumains in the midgut of the ixodid tick Haemaphysalis longicornis, is involved in blood-meal digestion[J]. J Insect Physiol, 2008, 54(3):573–585.
38. Alim MA, Tsuji N, Miyoshi T, et al. Developmental Stage- and Organ-Specific Expression Profilesof Asparaginyl Endopeptidass / Legumains in the Ixodid Tick Haemaphysalis longicornis[J]. J Vet Med Sci, 2008, 70(12): 1363–1366.
39. Alim MA, Tsuji N, Miyoshi T et al. Legumains from the hard tick Haemaphysalis longicornis play modulatory roles in blood feeding and gut cellular remodelling and impact on embryogenesis[J]. Int J Parasitol, 2009, 39(1):97–107.
40. Alim MA, Tsuji N, Miyoshi T, et al. Legumains from the hard tick Haemaphysalis longicornis play modulatory roles in blood feeding and gut cellular remodelling and impact on embryogenesis[J]. Int J Parasitol. 2009, 39: 97-107.
41. Attardo GM, Higgs S, Klingler KA, et al. RNA interference-mediated knockdown of a GATA factor reveals a link to anautogeny in the mosquito Aedes aegypti[J]. Proc Natl Acad Sci USA, 2003, 100, 13374–13379.
42. Bates SM and Weitz JI. The status of new anticoagulants[J]. Br.J. Haematol. 2006, 134: 3–19.
43. Boldbaatar D, Sikalizyo Sikasunge C, Battsetseg B, et al. Molecular cloning and functional characterization of an aspartic protease from the hard tick Haemaphysalis longicornis[J].Insect Biochem. Mol. Biol. 2006, 36:25-36.
44. Boisson B, Jacques J C, Choumet V, et al. Gene silencing in mosquito salivary glands by RNAi[J]. FEBS Lett. 2006, 580, 1988–1992.
45. Boutros M, Kiger AA, Armknecht S, et al. Heidelberg Fly Array Consortium, Genome-wide RNAi analysis of growth and viability in Drosophila cells[J]. Science. 2004, 303, 832–835.
46. Bullard B, Leonard K, Larkins A, et al. Troponin of asynchronous flight muscle[J]. J Mol Biol. 1988, 204, 621–637.
47. Chen HG, Nara T, Zeng XJ, et al. Hirayama vaccination of domestic pig with recombinant paramyosin against Schistosoma japonicum in China[J]. Vaccine, 2000, 18 : 214-26.
48. Ceraul SM, Sonenshine DE, Ratzlaff RE, et al. An art hropod defensin expressed by the hemocytes of the American dog tick, Dermacentor variabilis ( Acari: Ixodidae )[J]. Insect Biochem Mol Biol, 2003, 33 (11): 1099-1103.
49. Chen A and Holmes S P. Molecular cloning and functional expression of a serotonin receptor from the Southern cattle tick, Boophilus microplus (Acari: Ixodidae) [J]. Insect Mol Biol, 2004, 13(1): 45-54.
50. Ceraul S M, SM, Dreher-Lesnick SM, Gillespie JJ, et al. New tick defensin isoform and antimicrobial gene expression in response to Rickettsia montanensis challenge[J]. Infect Immun. 2007, 75:1973–1983.
51. de la Fuente J and Kocan KM. Advances in the identification and characterization of protective antigens for development of recombinant vaccines against tick infestations[J]. Expert Rev Vaccines.2003, 2: 583–593.
52. de la Fuente J and Kocan KM. Strategies for development of vaccines for control of ixodid tick species[J]. Parasite Immunol, 2006, 28(7): 275–283.
53. de la Fuente J, Kocan KM, Blouin EF. Tick Vaccines and the Transmission of Tick-BornePathogens[J]. Vet Res Commun, 2007, 31(1): 85–90.
54. da Silva Vazl Jr, Torino Lermen T, Michelon A, er al.Effect of acaricides on the activity of a Boophilus microplus glutathione S-transferase[J]. Vet Parasitol, 2004, 119 (223): 237-245.
55. Emini EA, Hughes JV, Perlow DS, et al. Induction of hepatitis A virus-neutralizing antibody by a virus specific synthetic peptide[J]. J Virol, 1985, 55(3): 836-839.
56. Estrada-Pe?a A and Jongejan F. Ticks feeding on humans: a review of records on humanbiting Ixodoidea with special reference to pathogen transmission[J]. Exp Appl Acarol, 1999, 23(9): 685–715.
57. Ferreira CA, Barbosa MC, Silveira TC, et al. cDNA cloning, expression and characterization of a Boophilus microplus paramyosin[J]. Parasitology, 2002, 125(3): 265-274.
58. Fogaca AC, Lorenzini DM, Kaku LM, et al. Cysteine-rich antimicrobial peptides of the cattle tick Boophilus microplus: isolation, structural characterization and tissue expression profile[J]. Dev Comp Immunol, 2004, 28(3): 191–200.
59. Francischetti IM, Mather TN , Ribeiro JM. Tick saliva is a potent inhibitor of endothelial cell proliferation and angiogenesis[J]. Thromb Haemost, 2005, 94(1): 167–174.
60. Fyrberg E, Fyrberg CC, Beall C, et al. Drosophila melanogaster troponin-T mutations engender three distinct syndromes of myofibrillar abnormalities. J Mol Biol., 1990, 216(3), 657–675.
61. Gao J, Luo J, Fan R, et al. Molecular characterization of a myosin alkali light chain-like protein, a“concealed”antigen from the hard tick Haemaphysalis qinghaiensis[J]. Vet Parasitol, 2007, 147(1-2):140–149.
62. Gao J, Luo J, Fan R, et al. Characterization of a concealed antigen Hq05 from the hard tick Haemaphysalis qinghaiensis and its effect as a vaccine against tick infestation in sheep[J]. Vaccine, 2009, 27(3):483-490.
63. Gong H, LiaoM, Zhou J, et al. Gene silencing of ribosomal protein P0 is lethal to the tick Haemaphysalis longicornis[J]. Veterinary Parasitology, 2007, 151: (2008) 268–278.
64. Gong H, Umemiya R, Zhou J, et al. Blocking the secretion of saliva by silencing the HlYkt6 gene in the tick Haemaphysalis longicorni[J]s. Insect Biochem Mol Biol, 2009, 39(5-6): 372–381.
65. Graf JF, Gogolewski R, Leach-Bing N, et al. Tick control: an industry point of view[J]. Parasitology, 2004, 129: S427–442.
66. GrunclováL, Horn M, VancováM, et al. Two secreted cystatins of the soft tick Ornithodoros moubata: differential expression pattern and inhibitory specificity[J]. Biol Chem, 2006, 387(12): 1635–1644.
67. Guilfoile PG and Packila M. Identification of four genes expressed by feeding female Ixodes scapularis, including three with sequence similarity to previously recognized genes[J]. Exp Appl Acarol, 2004, 32 (122): 103-110.
68. Hatta T, Kazama K, Miyoshi T, et al. Identification and characterisation of a leucine aminopeptidase from the hard tick Haemaphysalis longicornis[J]. Int J Parasitol, 2006, 36(10-11): 1123–1132.
69. Hatta T, Tsuji N, Miyoshi T, et al. Leucine Aminopeptidase in the Ixodid Tick Haemaphysalis longicornis: Endogenous Expression Profiles in Midgut[J]. J Vet Med Sci, 2009, 71(5): 589–594.
70. Hatta T, Umemiya R, Liao M, et al. RNA interference of cytosolic leucine aminopeptidase reduces fecundity in the hard tick, Haemaphysalis longicornis[J]. Parasitol Res, 2007, 100(4):847–854.
71. Horn M, NussbaumerováM, Sanda M, et al. Hemoglobin digestion in blood-feeding ticks: mapping a multipeptidase pathway by functional proteomics[J]. Chem Biol, 2009, 16(10):1053-1063.
72. Hynes WL, Ceraul SM, Todd SM, et al. A defensin-like gene expressed in the black-legged tick, Ixodes scapularis[J]. Med Vet Entomol, 2005, 19(4): 339–344.
73. Imamura S, da Silva Vaz Junior l, Sugino M, et al. A serine protease inhibitor (serpin) from Haemaphysalis longicornis as an anti-tick vaccine[J]. Vaccine, 2005, 23(10): 1301–1311.
74. Imamura S, da Silva Vaz Jr Itabajara, Konnai S, et al. Effect of vaccination with a recombinant metalloprotease from Haemaphysalis longicornis[J]. Exp Appl Acarol, 2009, 48(4): 345–358.
75. Islam MK, Tsuji N, Miyoshi T, et al. The Kunitz-Like Modulatory Protein Haemangin Is Vital for Hard Tick Blood-Feeding Success[J]. PLoS Pathog. 2009, 5(7):1-13.
76. Jameson BA, Wolf H. The antigenic index: a novel algorithm for prediction antigenic determinants[J]. Comput Appl Biosci, 1988, 4(1):181-186.
77. Jaworski DC. Tick‘‘talk’’: protein release by tick salivary cells[J]. Trends Parasitol. 2003, 19(10): 427–429.
78. Jaworski DC, J asinskas A, Metz CN. Identification and characterization of a homologue of the proinflammatory cytokine macrophage migration inhibitory factor in the tick amblyomma americanum[J]. Insect Mol Biol, 2001, 10 (4): 323-331.
79. Karim S, Miller NJ, Valenzuela J, et al. RNAi-mediated gene silencing to assess the role of synaptobrevin and cystatin in tick blood feeding[J]. Biochem Biophys Res Commu, 2005, 334(4) :1336– 1342.
80. Konnai S, Nakajima C, Imamura S, et al. Suppression of cell proliferation and cytokine expression by HL-p36, a tick salivary gland-derived protein of Haemaphysalis longicornis[J]. Immunology, 2009, 126(2): 209–21.
81. Kotsyfakis M, Sá-Nunes A, Francischetti IM, et al. Anti-inflammatory and immunosuppressive activity of sialostatin L, a salivary cystatin from the tick Ixodes scapularis[J]. J Biol Chem, 2006, 281:26298–26307.
82. Kotsyfakis M, Karim S, Andersen JF, et al. Selective cysteine protease inhibition contributes to blood-feeding success of the tick Ixodes scapularis[J]. J Biol Chem, 2007, 282(40):29256–29263.
83. Kyte J and Doolittle RF. A simple method for displaying the hydropathic character of a protein[J]. J Mol Biol, 1982, 157(1):105-132.
84. Lai R, Lomas LO, Jonczy J, et al. Two novel non-cationic defensin-like antimicrobial peptides from haemolymph of the female tick, Amblyomma hebraeum[J]. Biochem J, 2004, 379(Pt3): 681-685.
85. Levy O. Antimicrobial proteins and peptides of blood: templates for novel antimicrobial agents[J]. Blood, 2000, 96(8):2664–2672.
86. Liao M, Zhou J, Hatta T, et al. Molecular characterization of Rhipicephalus (Boophilus) microplus Bm86 homologue from Haemaphysalis longicornis ticks[J]. Vet Parasitol, 2007, 146 (1-2):148-157.
87. Liao M, Hatta T, Umemiya R, et al. Identification of three protein disulfide isomerase members from Haemaphysalis longicornis tick Insect[J]. Biochem Mol Biol, 2007, 37: 641–654.
88. Liao M, Boldbaatar D, Gong H, et al. Functional analysis of protein disulfide isomerases in blood feeding, viability and oocyte development in Haemaphysalis longicornis ticks[J]. Insect Biochem Mol Bioly, 2008, 38(3): 285–295.
89. Lima CA, Sasaki SD, Tanaka AS. Bmcystatin, a cysteine proteinase inhibitor characterized from the tick Boophilus microplus[J]. Biochem Biophys Res Commun, 2006, 347(1): 44–50.
90. Meiringer CT, Auffarth K, Hou H, et al. Depalmitoylation of Ykt6 prevents its entry into the multivesicular body pathway[J]. Traffic, 2008, 9(9): 1510–1521.
91. Martín Hernández R, Cuellar del Hoyo C, Olmeda Garcia AS, et al. Analysis of stage-specific and shared antigens derived from Rhipicephalus sanguineus by electrophoresis and western blotting[J]. Med Vet Entomol, 1995, 9: 358~364.
92. Marden JH, Fitzhugh GH, Girgenrath M, et al. Alternative splicing, muscle contraction and intraspecific variation: associations between troponin T transcripts, Ca(2+) sensitivity and the force and power output of dragonfly flight muscles during oscillatory contraction[J]. J Exp Biol, 2001, 204(20), 3457–3470.
93. Marden JH, Fitzhugh GH, Wolf MR, et al. Alternative splicing, muscle calcium sensitivity and the modulation of dragonfly flight performance[J]. Proc Natl Acad Sci USA, 1999, 96(26):15304–15309.
94. Miyoshi T, Tsuji N, Islam MK, et al. A set of serine proteinase paralogs are required for blood-digestion in the ixodid tick Haemaphysalis longicornis[J]. Parasitol Int, 2008, 57(4): 499–505.
95. Miyoshi T, Tsuji N, Islam MK, et al. A Kunitz-type proteinase inhibitor from the midgut of the ixodid tick, Haemaphysalis longicornis, and its endogenous target serine proteinase[J]. Mol Biochem Parasitol, 2010, 170(2):112–115.
96. Minami and Tshihara T. Babesia ovata sp. isolate fromcattle in Japan[J]. Natl Inst Anim Health, 1980, 20(3):101-110.
97. Motobu M, Tsuji N, T Miyoshi, et al. Molecular characterization of a blood-induced serine carboxypeptidase from the ixodid tick Haemaphysalis longicornis[J]. FEBS J, 2007, 274: 3299–3312.
98. Mulenga C, Sugimoto M, Onuma. Issues in tick vaccine development: identification and characterization of potential candidate vaccine antigens[J]. Microbes Infect. 2000, 2: 1353–1361.
99. Mulenga A, Sugimoto C, Sako Y, et al. Molecular characterization of a Haemaphysalis longicornis tick salivary gland associated 29-kilodalton protein and its effectas a vaccine against tick infestation in rabbits[J]. Infec Immun, 1999, 67 (4): 1652-1658.
100.Mulenga A, Sugimoto C, Ingram G, et al. Characterization of two cDNAs encoding serineproteinases from the hard tick Haemaphysalis longicornis[J]. Insect Biochemd Mol Biol, 2001, 31(8):817-825.
101.Nuttall PA, Trimnell AR, Kazimirova M, et al. Exposed and concealed antigens as vaccine targets for controlling ticks and tick-borne diseases[J]. Parasite Immunol, 2006, 28(4),155–163.
102.Nakajima Y, Van der Goes Van Naters-Yasui A, Taylor D, et al. Antibacterial peptide defensin is involved in midgut immunity of the soft tick Ornithodoros moubata (Acari :Argasidae)[J]. Insect Mol Biol, 2002, 11 (6): 611-618.
103.Nakajima Y, Van der Goes Van Naters-Yasui A, Taylor D, et al. Two isoforms of a member of the arthropod defensin family from the soft tick Ornithodoros moubata (Acari :Argasidae)[J]. Insect Biochem Mol Biol, 2001, 31 (8): 747-751.
104.Ohnishi A, Hull JJ, Matsumoto S. Targeted disruption of genes in the Bombyx mori sex pheromone biosynthetic pathway[J]. Proc Natl Acad Sci U S A, 2006, 103(12): 4398–4403.
105.Perry SV. Troponin T: genetics, properties and function[J]. J Muscle Res Cell Motil, 1998, 19(6): 575–602.
106.Parola P and Raoult D. Tick-borne bacterial diseases emerging in Europe[J]. Clin Microbiol Infec, 2001, 7: 80–83.
107.Rossi V, Banfield DK, Vacca M, et al. Longins and their longin domains: regulated SNAREs and multifunctional SNARE regulators[J]. Trends Biochem Sci, 2004, 29(12): 682–688.
108.Rothel JS, Wood PR, Seowf HF, et al. Urea/DTT solubiliiation of a recombinant Taenia ovis antigen, 45W, expressed as a GST fusion protein results in enhanced protective immune response to the 45W moiety[J]. Vaccine, 1997, 15(5): 469-472.
109.Rudenko N, Golovchenko M, Edwards MJ, et al. Differential expression of Ixodes ricinus tick genes induced by blood feeding or Borrelia burgdorferi infection[J]. J Med Entomol, 2005, 42(1): 36–41.
110.Sauer JR, M cswain JL, Bowman AS, et al. Tick salivary glandphysiology[J]. Annu Rev Entomol, 1995, 40: 245-267.
111.Selsted ME and Ouellette AJ. Mammalian defensins in the antimicrobial immune response[J]. Nat Immunol, 2005, 6(6):551–557.
112.Shahein YE. Molecular cloning and expression of a larval immunogenic protein from the cattle tick Boophilus annulatus[J]. Vet Immunol Immunopathol, 2008, 121(3-4): 281-289.
113.Silverman GA, Bird Pl, Carrell RW,et al. The serpins are an expanding superfamily of structurally similar but functionally diverse proteins: evolution, mechanism of inhibition, novel functions, and a revised nomenclature[J]. J Biol Chem, 2001, 276(36): 33293–33296.
114.Sojka D, Hajdusek O, Dvorák J et al. IrAE: an asparaginyl endopeptidase (legumain) in the gut of the hard tick Ixodes ricinus[J]. Int J Parasitol, 2007, 37(7): 713-724.
115.Sojka D, Franta Z, Horn M, et al. Profiling of proteolytic enzymes in the gut of the tick Ixodes ricinus reveals an evolutionarily conserved network of aspartic and cysteine peptidases[J]. Parasit Vectors, 2008, 1(1):7.
116.Solis CF, Vázquez-Talavera J, Laclette JP. Toward development of a Taenia solium paramyosin based vaccine against porcine cysticercosis[J]. Gac Med Mex, 2004, 140(2): 129-38.
117.Sonenshine DE. Biology of Ticks, vol. 1(M). New York: Oxford University Press, 1991.
118.Sonenshine DE, CeraulSM, Hynes WE, et al. Expression of defensin-like peptides in tick hemolymph and midgut in response to challenge with Borrelia burgdorferi, Escherichia coli and Bacillus subtilis[J]. Exp Appl Acarol, 2002, 28(1-4): 127–134.
119.Sugino M, Imamura S, Mulenga A, et al. A serine proteinase inhibitor (serpin) from ixodid tick Haemaphysalis longicornis cloning and preliminary assessment of its suitability as a candidate for a tick vaccine[J]. Vaccine, 2003, 21(21-22): 2844–2851.
120.SuginoM, Imamura S, Mulenga A, et al. A serine proteinase inhibitor (serpin ) from ixodid tick Haemaphysalis longicornis: cloning and preliminary assessment of its suitability as a candidatefor a tick vaccine[J]. Vaccine, 2003, 21: 2844-2851.
121.Taylor MG, Huggins MC, Shi F, et al. Production and testing of Schistosoma japonicum candidate vaccine antigens in the natural ovine host. Vaccine, 1998, 16(13): 1290-8.
122.Temeyer KB, Davey RB, Chen AC. Identification of a third Boophilus microplus (Acari: Ixodidae) cDNA presumptively encoding anacetyl cholinesterase[J]. J Med Entomol, 2004, 41 (3): 259-268.
123.Tsuji N and Fujisaki K. Longcin plays a crucial role inhibiting the transmission of babesia parasites in the vector tick Hamaphysalis longicornis[J]. Future Microbiol, 2007, 2(6): 575-578.
124.Tsuji N, Battsetseg B, Boldbaatar D, et al. Babesial Vector Tick Defensin against Babesia sp. Parasites[J]. Infect Immun, 2007, 75(7): 3633–3640.
125.Tsuda A, Mulenga A, Sugimoto C, et al. cDNA cloning, characterization and vaccine effect analysis of Haemaphysalis longicornis tick saliva proteins[J]. Vaccine, 2001, (19) : 4287～4296.
126.Tsuji N, Kamio T, Isobe T, et al. Molecular characterization of a peroxiredoxin from the hard tick Haemaphysalis longicornis[J]. Insect Mol Biol, 2001,10(2): 121-129.
127.Tsuji N, Miyoshi T, Battsetseg B, et al. a Cysteine Protease is Critical for Babesia spp. Transmission in Haemaphysalis ticks[J]. PLoS Pathog, 2008, 4(5): 1-14.
128.Valenzuela JG. Exploring tick saliva: from biochemistry to‘sialome’and functional genomics[J]. Parasitology, 2004, 129: S83–S94.
129.Vázquez-Talavera J, Solís CF, Terrazas LI, et a1. Characterization and protective potential of the immune response to Taenia solium paramyosin in a murine model of cysticercosis[J]. Infect Immun, 2001, 69(9): 5412-6.
130.Willadsen P. Anti-tick vaccines[J]. Parasitology, 2004, 129: S367–387.
131.Willadsen P. Tick control: Thoughts on a research agenda[J]. Vet Parasitol, 2006, 138(1-2): 161–168.
132.Xu G, Fang QQ, Keirans JE. Cloning and sequencing of putative calreticulin complementary DNAs from four hard tick species[J]. J Parasitol, 2004, 90 (1): 73-78.
133.Yamaji K, Tsuji N, Miyoshi T, et al. A salivary cystatin, HlSC-1, from the ixodid tick Haemaphysalis longicornis play roles in the blood feeding processes[J]. Parasitol Res, 2009,106(1):61-68.
134.Yamaji K, Tsuji N, Miyoshi T, et al. Hemoglobinase activity of a cysteine protease from the ixodid tick Haemaphysalis longicornis[J]. Parasitol Int, 2009, 58:232-237.
135.Yamaji K, Tsuji N, Miyoshi T, et al. Hlcyst-1 and Hlcyst-2 are potential inhibitors of HlCPL-A in the midgut of the ixodid tick Haemaphysalis longicornis[J]. J Vet Med Sci, 2010, 72(5):599-604.
136.You M and Fujisaki K. Vaccination Effects of Recombinant Chitinase Protein from the Hard Tick Haemaphysalis longicornis (Acari: Ixodidae)[J]. J Vet Med Sci, 2009, 71(6): 709–712.
137.You M, Xuan X, Tsuji N, et al. Molecular characterization of a troponin I-like protein from the hard tick Haemaphysalis longicornis[J]. Insect Biochem Mol Biol, 2001, 32(1): 67-73.
138.You M, Xuan X, Tsuji N, et al. Identification and molecular characterization of a chitinase from the hard tick Haemaphysalis longicornis[J]. J Biol Chem, 2003, 278(10): 8556–8563.
139.Zhou J, Ueda M, Umemiya R, et al. A secreted cystatin from the tick Haemaphysalis longicornis and its distinct expression patterns in relation to innate immunity[J]. Insect Biochem Mol Biol, 2006, 36:527–535.
140.Zhou J, Liao M, Hatta T, et al. Identification of a follistatin-related protein from the tick Haemaphysalis longicornis and its effect on tick oviposition[J]. Gene, 2006, 372, 191–198.
141.Zhou J, Liao M, Ueda M, et al. Sequence characterization and expression patterns of two defensin-like antimicrobial peptides from the tick Haemaphysalis longicornis[J]. Peptides, 2007, 28 (6) :304-310.
142.Zhou J, Liao M, Ueda M, et al. Characterization of an intracellular cystatin homolog from the tick Haemaphysalis longicornis. Vet Parasitol, 2009, 160: 180–183.
143.Zhou J, Liao M, Gong H, et al. Characterization of Hlcyst-3 as a member of cystatins from the tick Haemaphysalis longicornis[J]. Exp Appl Acarol, 2010, Jan 27.