[1]张艳英,张召兴,郭家媚,等.鸡源致病性大肠杆菌地方流行株外膜蛋白的制备及其免疫原性研究[J].中国预防兽医学报,2019,(11):1141-1146.[doi:0.3969/j.issn.1008-0589.201904002]
 ZHANG Yan-ying,ZHANG Zhao-xing,GUO Jia-mei,et al.A study on preparation and immunogenicity of OMPs from endemic strains of Chicken pathogenic Escherichia coli[J].Chinese journal of preventive veterinary medicine,2019,(11):1141-1146.[doi:0.3969/j.issn.1008-0589.201904002]
点击复制

鸡源致病性大肠杆菌地方流行株外膜蛋白的制备及其免疫原性研究()
分享到:

《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2019年11
页码:
1141-1146
栏目:
免疫学
出版日期:
2019-12-25

文章信息/Info

Title:
A study on preparation and immunogenicity of OMPs from endemic strains of Chicken pathogenic Escherichia coli
文章编号:
1008-0589(2019)11-1141-06
作者:
 

张艳英12张召兴23郭家媚2李蕴玉2李佩国2*索 勋1*

 (1. 中国农业大学 动物医学学院,北京 100094;2. 河北科技师范学院 河北省预防兽医学重点实验室,河北 秦皇岛 066004;
3. 河北旅游职业学院 畜牧兽医系,河北 承德 067000)
Author(s):
 

ZHANG Yan-ying12 ZHANG Zhao-xing23 GUO Jia-mei2 LI Yun-yu2 LI Pei-guo2* SUO Xun1*

 (1. Agricultural University of China, Beijing 10094, China; 2. Hebei Key Laboratory of Preventive Veterinary Medicine,
Hebei Normal University of Science &Technology, Qinhuangdao 066004, China; 3. Department of Animal Husbandry and Veterinary Medicine, Hebei Vocational College of Tourism, Chengde 067000, China)
关键词:
致病性大肠杆菌外膜蛋白制备免疫原性
Keywords:
chicken  pathogenic Escherichia coli  Outer membrane protein preparation  Immunogenicity
分类号:
S852.61
DOI:
0.3969/j.issn.1008-0589.201904002
文献标志码:
A
摘要:
为了分析鸡源致病性大肠杆菌地方流行株外膜蛋白(OMPs)免疫保护性,本研究采用Sarkosyl法提取10株鸡源致病性大肠杆菌地方流行株的OMPs,进行浓度测定和超微结构观察,经SDS-PAGE和western blot分析其免疫原性。结果显示,制备的OMPs浓度为1026.632 μg/mL~1715.589 μg/mL;电镜结果显示其超微结构呈双层泡状的C形或O形;SDS-PAGE分析显示其大小集中在25 ku~55 ku;动物免疫结果显示其具有一定的免疫原性。用提取的OMPs免疫小鼠和雏鸡后,以鸡源致病性大肠杆菌强毒株EC-9攻毒,结果显示:OMPs对小鼠和雏鸡均具有较好保护力,对小鼠保护率为70 %,对鸡的保护率为80 %;间接EILSA法检测免疫小鼠后的抗体效价最高为1:400,免疫雏鸡后抗体效价最高为1∶800;MTT法检测免疫小鼠后的T淋巴细胞转化率最高为1.582,免疫雏鸡后的T淋巴细胞转化率最高为2.215;常规法分析免疫小鼠脾脏指数为5.334,雏鸡脾脏指数为3.981,表明OMPs可以诱导小鼠和雏鸡产生良好的体液免疫应答和细胞免疫应答。本研究提取的鸡源致病性大肠杆菌地方流行株Omps对小鼠和雏鸡均具有一定的免疫保护性,为致病性大肠杆菌的亚单位疫苗的研制奠定基础。
Abstract:
In order to investigate the immunoprotection of outer membrane proteins (OMPs) of endemic strains of chicken-derived E.coli, this study used Sarkosyl method to extract OMPs of 10 local strains of chicken-borne pathogenic coliforms for concentration determination and ultrastructural observation, followed by analyzing their immunogenicity with SDS-PAGE and western blot. The results showed that the concentration of OMPs was between 1,026.632 μg/mL and 1,715.589 μg/mL, and the ultrastructure was C-shaped or O-shaped bilayer bubbles. The size of immunoreactiveprotein was between 25ku and 55ku, and they are immunogenic. After immunizing mice and chicks with OMPs extracted, they were challenged with EC-9. The results showed that OMPs had good protection against mice and chicks. The protection rateswere 70% on mice, and 80% on chickens, respectively. The OMPs can induce good humoral and cellular immune responses in mice and chicks. Indirect EILSA method was used to detect specific antibody titers. The highest antibody titers was 1∶400 in immunized mice and 1:800 in immunized chickens. MTT method was used to detect T lymphocyte transformation rate in peripheral blood. The highest T lymphocyte transformation rate was 1.582 in immunized mice and 2.215 in immunized chickens. The spleen index of immune organs was 5.334 in mice and 3.981 in chickens. In conclusion, the OMPs prepared in this study hadimmunoprotective effects on mice and chicken, and this study laid a foundation for the development of subunit vaccine of pathogenic E.coli.

参考文献/References:

[1]张春丽,张宇. 大肠埃希菌235株药敏状况分析[J]. 临床合理用药杂志,2011,4(1A):127.
[2]陈爱平,吴旭辰,熊美琴,等. 实时荧光PCR检测肠致泻性大肠杆菌在腹泻人群中的分布[J]. 海峡预防医学杂志,2012,18(2):1-3.
[3]DrRienzo J M, KInouye M. The outer membrane proteins of Gramnegative bacteria: biosynthesis, assembly and functions [J]. Annu Rev Biochem, 1978, 47: 481-532.
[4]Fairman J W, Noinaj N, Buchanan S K. The structural biology of β-barrel membrane proteins: a summary of recent reports [J]. Curr Opin Struct Biol , 2011, 21(4): 523-531.
[5]Confer A W, Ayalew S. The OmpA family of proteins: roles in bacterial pathogenesis and immunity [J]. Vet Microbiol, 2013, 163(3-4): 207-222.
[6]Zheng W Y, Wang, Y, Zhang Z C, et al. Immunological characteristics of outer membrane protein omp31 of goat Brucella and its monoclonal antibody [J]. Genet Mol Res, 2015, 14(4): 11965-11974.
[7]Simborio H L, Reyes A W, Hop H T, et al. Immune modulation of recombinant OmpA against Brucellaabortus 544 infection in mice [J]. J Microbiol Biotechnol, 2016, 26(3): 603-609.
[8]徐晶,陈立志,刘晓颖,等. 坏死梭杆菌OMPs提取及免疫原性分析[J]. 动物医学进展,2016,37([]6):26-29.
[9]强付,王洪伦,周昌范,等. 考马斯亮蓝G-250染色法测定柠条锦鸡儿种子中可溶性蛋白含量[J]. 氨基酸和生物资源,2011,33(3):74-76.
[10]徐全圆,石明,杨夷平,等. 马链球菌马亚种 FNEB 蛋白的截短表达及其免疫保护性研究[J]. 中国预防兽医学报,2015,37(02):132-135.
[11]张志强,杨楠,李永慧,等. 迟缓爱德华菌OMPsOmpA原核表达及其免疫原性研究[J]. 中国预防兽医学报,2018,40(6):534-537.
[12]潘廷云. 鸡大肠杆菌病脂多糖缺失多价灭活苗安全性、最适抗原含量、免疫持续期及交叉免疫保护研究[D]. 扬州:扬州大学,2018.
[13]李晓霞,邱玉玉,王海荣. 肠致病性大肠杆菌OMPs免疫保护性研究[J]. 中国免疫学杂志,2007,23(5):394-397.
[14]上官明君,王芳,张红岗,等. 菊粉对蛋雏鸡生长性能、免疫器官指数和血清免疫球蛋白的影响[J]. 动物营养学报,2009,21(1):118-122.
[15]张召兴,张香斋,李蕴玉,等. 鸡致病性E.coli地方株血清型的鉴定、毒力基因及致病性检测[J]. 中国兽医学报,2017,37(12):2260-2265.
[16]程安春,汪铭书,刘兆宇,等. 血清 2型鸭疫里默氏杆菌OMPs多肽的组成及免疫原性[J]. 中国兽医学报,2004,24(4):329-331.
[17]苏敬良,郭玉璞,吕艳丽. 鸭疫里氏杆菌OMPs免疫原性研究[J]. 畜牧兽医学报,2009,30(5):444-448.
[18]Franklin M W, Nepomnyachyi S, Feehan R, et al. Evolutionary pathways of repeat protein topology in bacterial outer membrane proteins [J]. Elife, 2018(7): 1-19.
[19]张斌,汤承,岳华. 副猪嗜血杆菌主要OMPs基因研究进展[J]. 动物医学进展,2013,34(01):91-96.
[20]Udhayakumar V, Muthukkaruppan V R. Protective immunity induced by outer membrane proteins of salmonellatyphimurium in mice [J]. Infec Immun, 1987, 55(3): 816-821.
[21]Phan G, Picard M, Broutin I. Focus on the outer membrane factor oprM, the forgotten player from efflux pumps assemblies [J]. antibiotics (Basel), 2015, 4(4): 544-566.
[22]张彦明,王晶钰,鱼艳荣. 禽大肠埃希氏菌融合菌株疫苗的研制及应用[J]. 中国兽医科技,2003,(11):42-45.
[23]高崧,彭大新,文其乙,等. 禽大肠杆菌病免疫保护机理的研究[J]. 畜牧兽医学报,2001,32(4):345-353.
[24]高崧,吴晓东,张扬,等. 禽大肠杆菌OMPs、脂多糖疫苗的免疫保护试验[J]. 中国兽医学报,2002,22(5):457-459.
[25]王文豪,张丹琳,张显浩,等. 猪致病性大肠埃希菌OMPs亚单位疫苗免疫效果观察[J]. 动物医学进展,2013,34(09):33-36.
(本文编辑:彭永刚;英文编辑:蔡文通)

相似文献/References:

[1]闫鸿斌,贾万忠,田广孚,等.四价鸡球虫苗的免疫效果研究[J].中国预防兽医学报,2009,(03):236.
 YAN Hong-bin,JIA Wan-zhong,TIAN Guang-fu,et al.The protective efficacy of the live vaccine against coccidiosis [J].Chinese journal of preventive veterinary medicine,2009,(11):236.
[2]谭复善,李文超,崔雷,等.鸡Mx 基因的表达及其抗血清的制备[J].中国预防兽医学报,2009,(05):388.
 TAN Fu-shan,LI Wen-chao,CUI Lei,et al.Preparation of the rabbit antiserum against chicken Mx[J].Chinese journal of preventive veterinary medicine,2009,(11):388.
[3]闫鸿斌,田广孚,贾万忠,等.6种鸡艾美耳球虫兰州株的致病性研究[J].中国预防兽医学报,2009,(04):266.
 YAN Hong-bin,TIAN Guang-fu,JIA Wan-zhong,et al.Evaluation of the virulence of 6 Eimeria spp. Lanzhou strains[J].Chinese journal of preventive veterinary medicine,2009,(11):266.
[4]赵绪永*,马 辉,赵 丽,等.鸡β-防御素-3 在毕赤酵母中的分泌表达及其生物活性测定 [J].中国预防兽医学报,2009,(01):24.
 ZHAO Xu-yong*,MA Hui,ZHAO Li,et al.Secreted expression of chicken β-gallinacin-3 inPichia pastoris and biological activities detection [J].Chinese journal of preventive veterinary medicine,2009,(11):24.
[5]邱 妍,崔保安,董发明,等.4种中药多糖对免疫雏鸡黏膜免疫功能的影响[J].中国预防兽医学报,2009,(12):974.
 QIU Yan,CUI Bao-an,DONG Fa-ming,et al.Effects of four polysaccharides on mucosal immunity in vaccinated chickens[J].Chinese journal of preventive veterinary medicine,2009,(11):974.
[6]叶秀华,蔡建平*,吴志光,等.rChIFN-γ 对E.tenella卵囊免疫雏鸡肠道分泌型IgA+细胞数量的影响[J].中国预防兽医学报,2008,(12):959.
 YE Xiu-hua,CAI Jian-ping*,WU Zhi-guang,et al.Immuno-modulatory effects of rChIFN-γ on sIgA+ plasma cell numbers in the intestine of chickens vaccinated with Eimeria tenella oocysts[J].Chinese journal of preventive veterinary medicine,2008,(11):959.
[7]周作勇,聂 奎*,王芝英,等.鸡MyD88、TRIF和TIRAP基因荧光定量PCR检测方法的建立[J].中国预防兽医学报,2011,(01):66.[doi:10.3969/j.issn.1008-0589.2011.01.16]
 ZHOU Zuo-yong,NIE kui*,WANG Zhi-ying,et al.Development of real-time PCR for the detection of chicken MyD88, TRIF and TIRAP genes[J].Chinese journal of preventive veterinary medicine,2011,(11):66.[doi:10.3969/j.issn.1008-0589.2011.01.16]
[8]张小燕,潘志明,胡青海,等.鸡白细胞介素2分子单克隆抗体的研制[J].中国预防兽医学报,2008,(01):39.
 ZHANG Xiao-yan,PAN Zhi-ming,HU Qing-hai,et al.Development of monoclonal antibodies against chicken IL-2[J].Chinese journal of preventive veterinary medicine,2008,(11):39.
[9]闫鸿斌,田广孚,贾万忠,等.鸡球虫苗(LCV-2号)安全性和免疫效果研究[J].中国预防兽医学报,2007,(05):368.
 YAN Hong-bin,TIAN Guang-fu,JIA Wan-zhong,et al.Efficacy of the vaccine (LCV-2) against chicken coccidiosis[J].Chinese journal of preventive veterinary medicine,2007,(11):368.
[10]肖根辉,孙 浩,何后军,等.鹅致病性大肠杆菌耐药性与整合子研究[J].中国预防兽医学报,2012,(10):790.[doi:doi: 10.3969/j.issn.1008-0589.2012.10.08]
 XIAO Gen-hui,SUN Hao,HE Hou-jun,et al.The research on integrons and drug resistance ofthe pathogenic E. coli from goose[J].Chinese journal of preventive veterinary medicine,2012,(11):790.[doi:doi: 10.3969/j.issn.1008-0589.2012.10.08]
[11]贾青辉?,张召兴?,李蕴玉,等.河北地区鸡源致病性大肠杆菌毒力基因检测与耐药性分析[J].中国预防兽医学报,2018,(05):401.[doi:0.3969/j.issn.1008-0589.201706016]
 JIA Qing-hui,ZHANG Zhao-xing,LI Yun-yu,et al.Analysis of virulence genes and drug resistance of the pathogenic Escherichia coli isolates from chicken in Hebei province[J].Chinese journal of preventive veterinary medicine,2018,(11):401.[doi:0.3969/j.issn.1008-0589.201706016]

更新日期/Last Update: 2019-12-09