卫氏并殖吸虫TPx基因的克隆、表达和免疫学诊断价值

doi:10.12140/j.issn.1000-7423.2021.01.003

中国寄生虫学与寄生虫病杂志 ›› 2021, Vol. 39 ›› Issue (1): 20-26.doi: 10.12140/j.issn.1000-7423.2021.01.003

卫氏并殖吸虫TPx基因的克隆、表达和免疫学诊断价值

周岩¹(), 陈韶红¹, 李浩¹, 程娜¹, 洪加林², 许学年¹^,^*()

1 中国疾病预防控制中心寄生虫病预防控制所,国家热带病研究中心,世界卫生组织热带病合作中心,科技部国家级热带病国际联合研究中心,卫生部寄生虫病原与媒介生物学重点实验室,上海 200025
2 永嘉县人民医院,永嘉 325100

收稿日期:2020-07-27 修回日期:2020-10-10 出版日期:2021-02-28 发布日期:2021-03-10
通讯作者: 许学年
作者简介:周岩（1982-）女,硕士,助理研究员,从事寄生虫病防控研究。E-mail: zhouyan@nipd.chinacdc.cn
基金资助:
国家公益性卫生行业科研专项(201502021);国家寄生虫种质资源共享服务平台(20170804)

Cloning, expression of the thioredoxin peroxidase gene of Paragonimus westermani and its immunodiagnostic potential

ZHOU Yan¹(), CHEN Shao-hong¹, LI Hao¹, CHENG Na¹, HONG Jia-lin², XU Xue-nian¹^,^*()

1 National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention; Chinese Center for Tropical Diseases Research; WHO Collaborating Center for Tropical Diseases; National Center for International Research on Tropical Diseases, Ministry of Science and Technology; Key Laboratory of Parasite and Vector Biology, Ministry of Health, Shanghai 200025, China
2 People’s Hospital of Yongjia County, Yongjia 325100, China

Received:2020-07-27 Revised:2020-10-10 Online:2021-02-28 Published:2021-03-10
Contact: XU Xue-nian
Supported by:
Chinese Special Program for Scientific Research of Public Health(201502021);National Sharing Service Platform for Parasite Resources(20170804)

摘要/Abstract

摘要：

目的免疫筛选卫氏并殖吸虫成虫cDNA表达文库,克隆并表达卫氏并殖吸虫硫氧还蛋白过氧化物酶（TPx）,初步评价其免疫诊断价值。方法用卫氏并殖吸虫病患者的混合血清筛选卫氏并殖吸虫成虫λ ZAP cDNA表达文库,取阳性噬菌体进行克隆、测序和序列比对分析。将TPx基因全长片段和前端截短片段分别克隆至原核表达质粒pET28a(+)中,构建rPwTPx（全长型）和rPwTPx_1（截短型）表达载体。构建的重组蛋白经1 mmol/L的异丙基-β-D-半乳糖苷（IPTG）诱导表达。使用蛋白抽提剂、溶菌酶和核酸酶裂解表达细菌,用组氨酸标签亲和纯化柱（Ni-NTA树脂）纯化rPwTPx和rPwTPx_1,SDS-PAGE检测重组蛋白的表达情况。以rPwTPx、rPwTPx_1和粗抗原作为检测抗原,间接ELISA法检测36份卫氏并殖吸虫病、15份华支睾吸虫病、15份日本血吸虫病、15份片形吸虫病和15份猪囊尾蚴病的患者血清,以及36份健康人血清,评价该重组蛋白的免疫诊断价值。以健康人血清对照组的平均值加上4个标准差作为阳性判断值。所有数据均采用SAS 9.2软件进行统计学分析。结果克隆的卫氏并殖吸虫TPx基因,经原核表达、纯化,获得其可溶性重组蛋白rPwTPx和rPwTPx_1,相对分子质量（M_r）分别为25 000和22 000。以rPwTPx作为检测抗原的ELISA检测结果显示,卫氏并殖吸虫病患者、健康者、华支睾吸虫病患者、日本血吸虫病患者、片形吸虫病患者和猪囊尾蚴病患者血清组的吸光度（A₄₅₀值）分别为0.150 ± 0.092、0.036 ± 0.014、0.043 ± 0.019、0.047 ± 0.013、0.060 ± 0.022和0.048 ± 0.021。卫氏并殖吸虫病患者血清阳性率为58.3%（21/36）,36份健康人血清、15份华支睾吸虫病患者血清、15份日本血吸虫病患者血清、15份猪囊尾蚴病患者血清均未检出阳性,15份片形吸虫病患者血清假阳性为2/15。以rPwTPx_1作为检测抗原ELISA检测结果则为0.144 ± 0.092、0.022 ± 0.009、0.027 ± 0.015、0.033 ± 0.022、0.036 ± 0.015和0.032 ± 0.018。卫氏并殖吸虫病患者血清阳性率为88.9%（32/36）,健康人血清未检出阳性,15份华支睾吸虫病患者血清仅1份存在交叉反应,15份日本血吸虫病患者血清假阳性为3份,15份片形吸虫病患者和15份猪囊尾蚴病患者血清假阳性均为2份。重组蛋白rPwTPx的ELISA检测敏感性为58.3%,特异性为97.9%,总符合率为87.1%;rPwTPx_1则分别为88.9%、91.7%、90.9%。rPwTPx_1敏感性高于rPwTPx（P < 0.05）。以成虫粗抗原为检测抗原的ELISA检测结果显示,36份健康者血清A₄₅₀值为0.012,标准差为0.006。敏感性为100%,特异性为83.3%,总符合率为87.9%,与rPwTPx-ELISA和rPwTPx_1-ELISA检测的总符合率差异无统计学意义（P > 0.05）。结论筛选并克隆了卫氏并殖吸虫TPx基因,表达并纯化全长型和截短型两种重组TPx抗原,构建的全长型和截短型重组TPx抗原作为人体卫氏并殖吸虫病的免疫诊断抗原具有一定的诊断意义。

关键词: 卫氏并殖吸虫, 硫氧还蛋白过氧化物酶, 免疫学筛选, 原核表达, ELISA

Abstract:

Objective To immunologically screen the cDNA library of adult Paragonimus westermani for the gene encoding thioredoxin peroxidase (TPx), clone and express the gene, and evaluate the immunodiagnostic value of TPx recombinant protein. Methods The λ ZAP cDNA library was immunologically screened with the pooled serum of P. westermani patients. The obtained positive clones were sequenced and analyzed by multiple sequence alignment. The full length (rPwTPx) and N-terminal truncated (rPwTPx_1) sequences of PwTPX were subcloned into the prokaryotic plasmid pET28a (+), respectively, to construct the expression vectors rPwTPX and rPwTPX_1. The expression of the two constructed recombinant plasmids was induced by 1 mmol/L IPTG. Protein extraction reagents, lysozyme and nuclease were used to lyse and express the bacterial fluid. The recombinant proteins of rPwTPx and rPwTPx_1 were purified using the His-tagged affinity column (Ni-NTA). SDS-PAGE was used to analyzed the expression of the recombinant proteins. rPwTPx and rPwTPx_1 were used to test the sera of 36 paragonimiasis westermani patients, 15 patients with clonorchiasis sinensis, 15 schistosomiasis japonica patients, 15 fascioliasis gigantica patients, 15 cysticercosis patients, and 36 healthy donors, using the indirect ELISA method. Data were analyzed with the SAS 9.2 software. Results The TPx recombinant proteins rPwTPx and rPwTPx_1 were obtained through expression and purification, with relative molecular mass M_r of 25 000 and 22 000, respectively. The rPwTPx ELISA results showed that the A₄₅₀ values for the sera from patients of paragonimiasis westermani, healthy persons, clonorchiasis sinensis, schistosomiasis japonica, fascioliasis gigantic, and cysticercosis patients were 0.150 ± 0.092, 0.036 ± 0.014, 0.043 ± 0.019, 0.047 ± 0.013, 0.060 ± 0.022 and 0.048 ± 0.021, respectively, The positive rate in serum of paragonimiasis westermani patients was 58.3% (21/36). There was no cross-reaction with sera of healthy donors, clonorchiasis sinensis, schistosomiasis japonica and cysticercosis patients. The cross-reaction with sera of fascioliasis gigantic patients was 2 samples. The ELISA using rPwTPx_1 as the antigen presented the A₄₅₀ values to the above testing sera was 0.144 ± 0.092, 0.022 ± 0.009, 0.027 ± 0.015, 0.033 ± 0.022, 0.036 ± 0.015 and 0.032 ± 0.018, respectively. The positive rate of paragonimiasis westermani patients sera was 88.9% (32/36), while the sera of healthy donors showed no cross-reaction. Only 1 serum sample of 15 clonorchiasis sinensis patients had cross-reaction. The cross-reaction with sera of schistosomiasis japonica, fascioliasis gigantic and cysticercariasis patients were 3, 2 and 2 samples. The total diagnostic coincidence rate, sensitivity and specificity of ELLSA using rPwTPx and rPwTPx_1 as antigen were 87.1%, 58.3%, 97.9%, and 90.9%, 88.9%, 91.7%, respectively. The sensitivity of the rPwTPx_1 ELISA was significantly higher than that of the rPwTPx ELISA (P < 0.05). Crude antigen-ELISA showed that the A₄₅₀ value of serum samples from the 36 healthy donors was 0.012, and S was 0.006. The sensitivity, specificity and total diagnostic coincidence rate of crude antigen were 100%, 83.3% and 87.9%, which did not significantly differ from those of rPwTPx and rPwTPx_1 (P > 0.05). Conclusion The gene encoding TPx was screened and expressed, and two structural types of recombinant protein were obtained, including full length and truncated types, which recombinant protein shows proper sensitivity and high specificity for the serodiagnosis of P. westermani infection.

Key words: Paragonimus westermani, Thioredoxin peroxidase, Immunoscreen, Expression, ELISA

中图分类号:

R532.221

周岩, 陈韶红, 李浩, 程娜, 洪加林, 许学年. 卫氏并殖吸虫TPx基因的克隆、表达和免疫学诊断价值[J]. 中国寄生虫学与寄生虫病杂志, 2021, 39(1): 20-26.

ZHOU Yan, CHEN Shao-hong, LI Hao, CHENG Na, HONG Jia-lin, XU Xue-nian. Cloning, expression of the thioredoxin peroxidase gene of Paragonimus westermani and its immunodiagnostic potential[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2021, 39(1): 20-26.

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参考文献 24

[1]	Ayako Y, Kayoko M, Junji M, et al. Venison, another source of Paragonimus westermani infection[J]. Parasitology, 2016,65(6Pt A):607-612.
[2]	Itoh N, Tsukahara M, Yamasaki H, et al. Paragonimus westermani infection mimicking recurrent lung cancer: a case report[J]. J Infect Chemother, 2016,22(12):815-818. doi: 10.1016/j.jiac.2016.07.002 pmid: 27498617
[3]	Kalhan S, Sharma P, Sharma S, et al. Paragonimus westermani infection in lung: a confounding diagnostic entity[J]. Lung India, 2015,32(3):265. doi: 10.4103/0970-2113.156248 pmid: 25983414
[4]	Shim SS, Kim Y, Lee JK, et al. Pleuropulmonary and abdominal paragonimiasis: CT and ultrasound findings[J]. Br J Radiol, 2012,85(1012):403-410. doi: 10.1259/bjr/30366021 pmid: 22457403
[5]	Singh TS, Mutum SS, Razaque MA. Pulmonary paragonimiasis: clinical features, diagnosis and treatment of 39 cases in Manipur[J]. Trans Royal Soc Trop Med Hyg, 1986,80(6):967-971. doi: 10.1016/0035-9203(86)90275-0
[6]	Keiser J, Utzinger J. Emerging foodborne trematodiasis[J]. Emerg Infect Dis, 2005,11(10):1507-1514. doi: 10.3201/eid1110.050614 pmid: 16318688
[7]	Wu XP, Gao LF, Zhang L, et al. Construction and screening of a cDNA expresssion library of Clonorchis sinensis adult worm[J]. Chin J Vet Sci, 2009,29(7):868-872. (in Chinese)
	( 吴秀萍, 高丽芳, 张玲, 等. 华支睾吸虫成虫cDNA表达文库的构建与筛选[J]. 中国兽医学报, 2009,29(7):868-872.)
[8]	McGonigle S, Curley GP, Dalton JP. Cloning of peroxiredoxin, a novel antioxidant enzyme, from the helminth parasite Fasciola hepatica[J]. Parasitology, 1997,115(Pt 1):101-104.
[9]	Kumagai T, Osada Y, Kanazawa T. 2-Cys peroxiredoxins from Schistosoma japonicum: the expression profile and localization in the life cycle[J]. Mol Biochem Parasitol, 2006,149(2):135-143. doi: 10.1016/j.molbiopara.2006.05.004 pmid: 16806527
[10]	Shu B, Cheng XJ. Peroxiredoxin and parasite[J]. Chin J Vet parasitol, 2002,20(2):115-118. (in Chinese)
	( 舒勃, 程训佳. Peroxiredoxin与寄生虫[J]. 中国寄生虫学与寄生虫病杂志, 2002,20(2):115-118.)
[11]	Suttiprapa S, Loukas A, Laha T, et al. Characterization of the antioxidant enzyme, thioredoxin peroxidase, from the carcinogenic human liver fluke, Opisthorchis viverrini[J]. Mol Biochem Parasitol, 2008,160(2):116-122. doi: 10.1016/j.molbiopara.2008.04.010 pmid: 18538872
[12]	Wang YQ, Zhou Y, Cheng N, et al. Cloning, expression and immunodiagnostic evaluation of the Fasciola gigantica thioredoxin peroxidase[J]. Chin J Parasitol Parasit Dis, 2015,33(2):81-85. (in Chinese)
	( 王月祺, 周岩, 程娜, 等. 巨片形吸虫硫氧还蛋白过氧化物酶的克隆、表达和免疫学诊断价值评价[J]. 中国寄生虫学与寄生虫病杂志, 2015,33(2):81-85.)
[13]	Macalanda AMC, Angeles JMM, Moendeg KJ, et al. Evaluation of Schistosoma japonicum thioredoxin peroxidase-1 as a potential circulating antigen target for the diagnosis of Asian schistosomiasis[J]. J Vet Med Sci, 2018,80(1):156-163. doi: 10.1292/jvms.17-0579 pmid: 29187698
[14]	Hu KM, Chen SH, Ai L, et al. Sequence analysis of ribosomal ITS2 gene and mitochondrial CO1 gene of Paragonimus metacercariae from freshwater crabs in Henan, Anhui, Fujian and Zhejiang Provinces, China[J]. Chin J Parasitol Parasit Dis, 2020,38(1):87-94. (in Chinese)
	( 胡坤敏, 陈韶红, 艾琳, 等. 豫皖闽浙4省溪蟹并殖吸虫囊蚴核糖体ITS2和线粒体CO1基因序列分析[J]. 中国寄生虫学与寄生虫病杂志, 2020,38(1):87-94.)
[15]	Chen SH, Zhou XN, Zhang YN, et al. Dynamics of specific antibody and circulating antigen in sera from the dogs infected with Paragonimus westermani[J]. Chin J Vet Parasitol, 2007,15(5):11-14. (in Chinese)
	( 陈韶红, 周晓农, 张永年, 等. 卫氏并殖吸虫感染犬循环抗原和特异性抗体的动态观察[J]. 中国兽医寄生虫病, 2007,15(5):11-14.)
[16]	Yang YP, Zhu XP, Yang J, et al. Immunoscreening and sequence analysis of a cdna library of adult Trichinella spiralis[J]. Chin J Parasitol Parasiti Dis, 2002,20(5):270-273. (in Chinese)
	( 杨雅平, 诸欣平, 杨静, 等. 旋毛虫成虫cDNA文库免疫筛选及序列分析[J]. 中国寄生虫学与寄生虫病杂志, 2002,20(5):270-273.)
[17]	Zhu YM, Li WG. Advanced on the study of molecular biology of Echinococcus granulosus[J]. Chin J Parasit Dis Can, 2005,18(3):217-220. (in Chinese)
	( 朱佑明, 李文桂. 细粒棘球绦虫分子生物学研究进展[J]. 中国寄生虫病防治杂志, 2005,18(3):217-220.)
[18]	Yin C, Luo XN, Wang S, et al. Prokaryotic expression and biological properties of thioredoxin peroxidase from Taenia solium[J]. Chin J Animal Vet Sci, 2014,45(9):1512-1517.(in Chinese)
	( 尹才, 骆学农, 王帅, 等. 猪带绦虫硫氧还蛋白过氧化物酶的原核表达及其生物学特性分析[J]. 畜牧兽医学报, 2014,45(9):1512-1517.) doi: 10.11843/j.issn.0366-6964.2014.09.018
[19]	Chae HZ, Robison K, Poole LB, et al. Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes[J]. Proc Natl Acad Sci USA, 1994,91(15):7017-7021. doi: 10.1073/pnas.91.15.7017 pmid: 8041738
[20]	Liu JY, Li WH, Qu ZG, et al. Cloning and bioinformatics analysis of thioredoxin peroxidase genes from Trichinella spiralis[J]. Chin Vet Sci, 2014,44(4):331-339. (in Chinese)
	( 刘静宜, 李文卉, 曲自刚, 等. 旋毛虫硫氧还蛋白过氧化物酶基因的克隆与生物信息学分析[J]. 中国兽医科学, 2014,44(4):331-339.)
[21]	Chen Y, Qiao J, Meng QL, et al. Molecular characteristics and reactionogenicity of Echinococcus granulosus TP_x protein[J]. Guizhou Agric Sci, 2018,46(1):68-73. (in Chinese)
	( 陈英, 乔军, 孟庆玲, 等. 细粒棘球蚴TP_x蛋白的分子特征与反应原性[J]. 贵州农业科学, 2018,46(1):68-73.)
[22]	Robinson MW, Hutchinson AT, Dalton JP, et al. Peroxiredoxin: a central player in immune modulation[J]. Parasite Immunol, 2010,32(5):305-313. doi: 10.1111/j.1365-3024.2010.01201.x pmid: 20500659
[23]	Sun L, Lin RY, Fang BB, et al. Cloning, expression, and determination of the immunodiagnostic value of thioredoxin peroxidase from Echinococcus multilocularis[J]. Chin J Pathogen Biol, 2019,14(6):634-638, 642. (in Chinese)
	( 孙立, 林仁勇, 房彬彬, 等. 多房棘球绦虫硫氧还蛋白过氧化物酶基因的克隆、表达及其免疫诊断价值初步评价[J]. 中国病原生物学杂志, 2019,14(6):634-638, 642.)
[24]	Sangpairoj K, Changklungmoa N, Vanichviriyakit R, et al. Analysis of the expression and antioxidant of 2-Cys peroxiredoxin protein in fasciola gigantic[J]. Exp Parasitol, 2014,140:24-32. doi: 10.1016/j.exppara.2014.02.017

卫氏并殖吸虫TPx基因的克隆、表达和免疫学诊断价值

Cloning, expression of the thioredoxin peroxidase gene of Paragonimus westermani and its immunodiagnostic potential

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血清Serum	检测份数 No. detected	血清抗体阳性数No. of positive
血清Serum	检测份数 No. detected	rPwTPx	rPwTPx_1	成虫粗抗原 Crude antigen
卫氏并殖吸虫病患者Paragonimiasis westermani	36	21	32	36
华支睾吸虫病患者Clonorchiasis sinensis	15	0	1	5
日本血吸虫病患者Schistosomiasis japonica	15	0	3	1
片形吸虫病患者Fascioliasis gigantica	15	2	2	10
猪囊尾蚴病患者Cysticercosis	15	0	2	0
健康者 Healthy donors	36	0	0	0
合计Total	132	23	40	52

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[13]	余琴1*，朱艳红2，关飞3，杨海2. 日本血吸虫重组抗原rSj26的磁分离酶联免疫分析的建立及其在低感染度血清抗体检测中的应用[J]. 中国寄生虫学与寄生虫病杂志, 2016, 34(4): 2-303-307.
[14]	李瑾，崔勇，尹昆，刘功振，肖婷，徐超，魏庆宽，黄炳成，孙慧*. 刚地弓形虫微线蛋白16基因片段原核表达、纯化及免疫反应性分析[J]. 中国寄生虫学与寄生虫病杂志, 2016, 34(3): 3-198-202.
[15]	赵殷奇1,2,3，李子华1,2,3，王浩2,4，朱明星1,2,3，牛楠2,3,5，王娅娜6，赵嘉庆2,4，李娜2,3,5，佟雪琪2,3,5，宋佳卉1,2,3，赵. 细粒棘球绦虫原头节抗原分子Eg-01883的克隆、表达及免疫原性分析[J]. 中国寄生虫学与寄生虫病杂志, 2016, 34(3): 5-208-214.