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Antimicrobial Susceptibility and Characterization of Outer Membrane Proteins of Aeromonas hydrophila Isolated in China 被引量:2
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作者 GUO Peng WANG Na +1 位作者 LIU Yong-jie LU Cheng-ping 《Journal of Integrative Agriculture》 SCIE CAS CSCD 2014年第4期911-917,共7页
Aeromonas hydrophila isolates from clinical cases (n=43) were tested against 8 antimicrobial agents and typed by outer membrane protein (OMP) pattern by using sodium dodecyl sulfate gel electrophoresis. All isolat... Aeromonas hydrophila isolates from clinical cases (n=43) were tested against 8 antimicrobial agents and typed by outer membrane protein (OMP) pattern by using sodium dodecyl sulfate gel electrophoresis. All isolates were resistant to ampicillin (MICs, ≥16 μg mL-1) and sulfamonomethoxine (MICs≥64 μg mLl), but susceptible to norfloxacin (MICs,≤0.5 μg mL-1). There was a high incidence of resistance to erythromycin (90.70%) and tylosin (93.02%), while a low incidences of resistance to ciprofloxacin (2.33%), enrofloxacin (2.33%) and florfenicol (4.65%). Six different outer membrane protein patterns were found among 34 isolates by analyzing proteins in the range of 22 to 50 kDa, other than 9 isolates with their respective profiles. The strains with the similar OMP profiles had similar resistances. Compared with the other strains from the same OMP patterns, NB-1, A.Pun and MR-1 had lacked the proteins in the range of 30 to 45 kDa and their resistance to florfenicol substantially increased. It is speculated that the outer membrane protein changes might correlate with decreased susceptibility to florfenicol in the three strains. Some strains which showed completely identical OMP types had a little difference in their resistance to fluoroquinolones, indicating that there might be other factors that were involved in the antimicrobial resistance of A. hydrophila. 展开更多
关键词 Aeromonas hydrophila antimicrobial resistance outer membrane proteins
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Determination of the genus-specific antigens in outer membrane proteins from the strains of Leptospira interrogans and Leptospira biflexa with different virulence 被引量:2
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作者 罗依惠 严杰 +1 位作者 毛亚飞 李淑萍 《Journal of Zhejiang University Science》 CSCD 2004年第4期462-466,共5页
Objective:To determine the existence of genus-specific antigens in outer membrane proteins (OMPs) of leptospira with different virulence. Methods: Microscope agglutination test (MAT) was applied to detect the agglutin... Objective:To determine the existence of genus-specific antigens in outer membrane proteins (OMPs) of leptospira with different virulence. Methods: Microscope agglutination test (MAT) was applied to detect the agglutination between commercial rabbit antiserum against leptospiral genus-specific TR/Patoc I antigen and 17 strains of Leptospira interrongans belonging to 15 serogroups and 2 strains of Leptospira biflexa belonging to 2 serogroups.The outer envelopes (OEs) of L.interrogans serogroup Icterohaemorrhagiae serovar lai strain lai (56601) with strong virulence and serogroup Pomona serovar pomona strain Luo (56608) with low virulence,and L.biflexa serogroup Semaranga serovar patoc strain Patoc I without virulence were prepared by using the method reported in Auran et al.(1972).OMPs in the OEs were obtained by treatment with sodium deoxycholate. SDS-PAGE and western blot were used for analyzing the features of the OMPs on electrophoretic pattern and the immunoreactivity to the antiserum against TR/Patoc I antigen, respectively. Results:All the tested strains belonging to different leptospiral serogroups agglutinated to the antiserum against leptospiral genus-specific TR/Patoc I antigen with agglutination titers ranging from 1:256-1:512. A similar SDS-PAGE pattern of the OMPs from the three strains of leptospira with different virulence was shown and the molecular weight of a major protein fragment in the OMPs was found to be approximately 60 KDa.A positive protein fragment with approximately 32 KDa confirmed by Western blot,was able to react with the antiserum against leptospiral genus-specific TR/Patoc I antigen, and was found in each the OMPs of the three stains of leptospira.Conclusion: There are genus-specific antigens on the surface of L.interrogans and L.biflexa. The OMP with molecular weight of 32 KDa may be one of the genus-specific protein antigens of leptospira. 展开更多
关键词 LEPTOSPIRA outer membrane protein Genus-specific antigen
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Immunogenicity and protective role of antigenic regions from five outer membrane proteins of Flavobacterium columnare in grass carp Ctenopharyngodon idella 被引量:2
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作者 罗璋 刘志新 +3 位作者 付建平 张秋胜 黄贝 聂品 《Chinese Journal of Oceanology and Limnology》 SCIE CAS CSCD 2016年第6期1247-1257,共11页
Flavobacterium columnare causes columnaris disease in freshwater fi sh. In the present study, the antigenic regions of fi ve outer membrane proteins(OMPs), including zinc metalloprotease, prolyl oligopeptidase, thermo... Flavobacterium columnare causes columnaris disease in freshwater fi sh. In the present study, the antigenic regions of fi ve outer membrane proteins(OMPs), including zinc metalloprotease, prolyl oligopeptidase, thermolysin, collagenase and chondroitin AC lyase, were bioinformatically analyzed, fused together, and then expressed as a recombinant fusion protein in Escherichia coli. The expressed protein of 95.6 k Da, as estimated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was consistent with the molecular weight deduced from the amino acid sequence. The purifi ed recombinant protein was used to vaccinate the grass carp, C tenopharyngodon idella. Following vaccination of the fi sh their Ig M antibody levels were examined, as was the expression of I g M, Ig D and Ig Z immunoglobulin genes and other genes such as MHC Iα and MHC I I β, which are also involved in adaptive immunity. Interleukin genes( IL), including I L- 1β, IL- 8 and I L- 10, and type I and type II interferon(I FN) genes were also examined. At 3 and 4 weeks post-vaccination(wpv), signifi cant increases in Ig M antibody levels were observed in the fi sh vaccinated with the recombinant fusion protein, and an increase in the expression levels of I g M, Ig D and Ig Z genes was also detected following the vaccinations, thus indicating that an adaptive immune response was induced by the vaccinations. Early increases in the expression levels of IL and IFN genes were also observed in the vaccinated fi sh. At four wpv, the fi sh were challenged with F. column a re, and the vaccinated fi sh showed a good level of protection against this pathogen, with 39% relative percent survival(RPS) compared with the control group. It can be concluded, therefore, that the fi ve OMPs, in the form of a recombinant fusion protein vaccine, induced an immune response in fi sh and protection against F. columnare. 展开更多
关键词 Flavobacterium columnare outer membrane protein antigen immunogenicity vaccine immune response grass carp
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Immunoproteomic Analysis of Bordetella bronchiseptica Outer Membrane Proteins and Identification of New Immunogenic Proteins
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作者 LIU Yan QIN Feng-yan +4 位作者 BAO Guo-lian CHEN Hui XIAO Chen-wen WEI Qiang JI Quan-an 《Journal of Integrative Agriculture》 SCIE CAS CSCD 2014年第9期2010-2018,共9页
Bordetella bronchiseptica is a Gram-negative pathogen that causes acute and chronic respiratory infection in a variety of animals. To identify useful antigen candidates for diagnosis and subunit vaccine of B. bronchis... Bordetella bronchiseptica is a Gram-negative pathogen that causes acute and chronic respiratory infection in a variety of animals. To identify useful antigen candidates for diagnosis and subunit vaccine of B. bronchiseptica, immunoproteomic analysis was adopted to analyse outer membrane proteins of it. The outer membrane proteins extracted from B. bronchiseptica were separated by two-dimensional gel electrophoresis and analyzed by Western blotting for their reactivity with the convalescent serum against two strains. Immunogenic proteins were identified by matrix-assisted laser desorption/ionization time of flight-mass spectrometry(MALDI-TOF-MS), a total of 14 proteins are common immunoreactive proteins, of which 1 was known antigen and 13 were novel immunogenic proteins for B. bronchiseptica. Putative lipoprotein gene was cloned and recombinantly expressed. The recombinant protein induced high titer antibody, but showed low protective indices against challenges with HB(B. bronchiseptica strain isolated from a infected rabbit). The mortality of mice was 80% compared to 100% of positive controls. The identification of these novel antigenic proteins is an important resource for further development of a new diagnostic test and vaccine for B. bronchiseptica. 展开更多
关键词 Bordetella bronchiseptica outer membrane proteins immunoproteomic analysis
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Study on Outer Membrane Protein Patterns of Escherichia coli O38,O53 and O75 Isolated from Chickens
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作者 张艳英 高桂生 +5 位作者 高光平 史秋梅 刘玉芹 张海莹 房海 陈翠珍 《Agricultural Science & Technology》 CAS 2012年第10期2070-2072,2122,共4页
[Objective] This study aimed to investigate the outer membrane protein (OMP) patterns of Escherichia coli 038, 053 and 075 isolates from chickens. [Method] Eight pathogenic E. coil isolates with various serotypes we... [Objective] This study aimed to investigate the outer membrane protein (OMP) patterns of Escherichia coli 038, 053 and 075 isolates from chickens. [Method] Eight pathogenic E. coil isolates with various serotypes were used as experimental materials to extract OMP by using supersonic schizolysis method and Sarcosyl. After SDS-PAGE electrophoresis, OMP patterns of the extracted products were determined based on the OMP model diagram. [Result] OMP of eight E. coil isolates with three serotypes were divided into three patterns, to be specific, 2 075 isolates respectively belonged to OMP-I and OMP-II pattern, 1 053 isolate belonged to OMP-II pattern, and 5 038 isolates belonged to OMP-I and OMP-III pattern. [Conclusion] Experimental results showed that E. coli isolates with the same serotype may belong to completely different OMP patterns, while serologically unrelated isolates may belong to the same OMP pattern. OMP of E. coil isolates with the same serotype may generate genetic differentiation; in addition, OMP of E. coli isolates with different serotypes may have different genetic correlation. 展开更多
关键词 Escherichia coli from chickens outer membrane protein pattern SDS- PAGE
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30 and 32 kDa outer membrane proteins of Bordetella pertussis as a modulator on promoting degranulation of mast cells
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作者 YONG LIANG LOU JIE YAN +1 位作者 YI HUI LUO YA FEI MAO 《Journal of Microbiology and Immunology》 2005年第2期111-119,共9页
The correlation between the activities of the outer menbrane proteins (OMPs) of Bordetella pertussis and the lgE-mediated asthma was investigated in the present study, in which the OMPs of B. pertussis and their com... The correlation between the activities of the outer menbrane proteins (OMPs) of Bordetella pertussis and the lgE-mediated asthma was investigated in the present study, in which the OMPs of B. pertussis and their components were prepared by detergent treatment and chromatography, and the molecular weights of the OMPs components were determined by SDS-PAGE. The amounts of total as well as the ovalbumin (OVA)-specific IgE induced by dead B. pertussis whole bacterial vaccine on guinea pigs were detected by ELISA. Meanwhile, the effect of the OMPs and their components to promote the degranulation of guinea pig mast cells was observed by using the mast cell degranulation test, and ELISA assay was used to measure the histatmine levels in the supematants from the mast cell cultures. Histamine sensitive test was used to demonstrate the effects of the OMPs and their components to increase the histamine lethal sensitivity in mice. It was found that four components with molecular weights of 30, 32, 38 and 69 kDa could be obtained from the OMPs of B. pertussts, and the dead whole bacteria vaccine of B. pertussis had the ability to increase the levels of the total as well as the OVA-specific IgE in sera of guinea pigs. The OMPs and their 30 and 32 kDa components demonstrated significantly enhancing effect on the degranulation of guinea pig mast cells, and the histamine levels in the supematants from the mast cell culture treated with OMPs and their 30 and 32 kDa components were also significantly increased. It is evident that the strong adjuvant activity and the enhancing effect to degranulation of mast cells and the release of histamine of certain outer membrane components of B. pertussis could be demonstrated as revealed by the results of the present study, suggesting the possibility of a close relationship between the infection of vaccination with B. pertussis and the IgE-mediated asthma. 展开更多
关键词 Bordetella pertussis Bacterial outer membrane protein Immunoglobulin E Mast cells Cell degranulation Histamine Asthma
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Prokaryotic Expression and Identification of Outer Membrane Protein 2 of Chlamydia trachomatis
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作者 陈超群 吴移谋 +2 位作者 李忠玉 朱翠明 尹卫国 《Chinese Journal of Sexually Transmitted Infections》 2004年第2期67-71,i001,共6页
Objective: To construct a recombinant plasmid containing the outer membrane protein 2 (Omp2) gene of Chlamydia trachomatis and express Omp2 in E.coli. Methods: The omp2 gene of C. trachomatis serovar D was cloned into... Objective: To construct a recombinant plasmid containing the outer membrane protein 2 (Omp2) gene of Chlamydia trachomatis and express Omp2 in E.coli. Methods: The omp2 gene of C. trachomatis serovar D was cloned into pQE30 vector following PCR amplification from genomic DNA. E. coli M15 transformants were induced to express the fusion protein by IPTG and the product was identified by SDS-PAGE and Western blot. Results: Confirmed by enzyme cleavage analysis and DNA sequencing, a correct recombinant plasmid pQE30/omp2 was constructed. The fusion protein from the transformants was approximately 60 kDa in size in SDS-PAGE analysis, which could specially react with anti-6 X His mouse monoclonal IgG antibodies. Conclusion: We successfully expressed Omp2 in E. coli M15, providing an efficient and simple system for assaying the immunological properties of Omp2. 展开更多
关键词 Chlamydia trachomatis outer membrane protein 2(omp2) expression.
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Development and Evaluation of a MAb-Based ELISA for Detection of Chlamy- dophila pneumoniae Infection with Variable Domain 2 and 3 of the Major Outer Membrane protein 被引量:2
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作者 ZHOU Zhou WU Yi Mou CHEN Li Li LIU Guang Chao LIU Liang Zhuan ZHOU An Wen ZHANG Jun Hua 《Biomedical and Environmental Sciences》 SCIE CAS CSCD 2012年第6期690-696,共7页
Objective This paper aims to develop a monoclonal antibodies (MAbs)- based ELISA for detecting Chlamydophila pneumoniae (C. pneumonioe) antigens in humans with the variable domains (VD) 2 and 3 of the major oute... Objective This paper aims to develop a monoclonal antibodies (MAbs)- based ELISA for detecting Chlamydophila pneumoniae (C. pneumonioe) antigens in humans with the variable domains (VD) 2 and 3 of the major outer membrane protein (MOMPvD2-VD~) and to assess its sensitivity and specificity by comparing with a widely used MAb that is able to recognize the elementary bodies of C. pneumoniae. Methods MOMPvo2-vo3were overexpressed in Escherichia coil and purified by affinity chromatography. Mice were immunized with the recombinant antigen, and hybridomas secreting MAbs were screened. Three stable hybridomas clones were selected and named 5D6, 7G3, and 8C9. The MAbs-based ELISA was scrutinized for species-specific recognition with a number of human throat swab samples from Group I (156 patients with typical respiratory illness clinically confirmed before) and Group II (57 healthy donors). Results In Group I, 55 positive cases were detected by anti-EB MAb-based ELISA, 51 cases were positive by MAbs 5D6-based ELISA, and 33 and 38 cases were positive by MAb 8C9 and 7G3-based ELISA respectively. Of the 57 samples from Group II "healthy donors", 5 were positive and 52 were negative with both anti-EB and 5D6-based tests, while 2 and 3 positive cases were identified by the other two MAb-based ELISAs respectively. Conclusion The novel MOMPvD2.VD3 MAb-based assay may have higher specificity than the anti-EB MAb, which may possibly be used as an alternative tool for the diagnosis of C. pneumoniae infection. 展开更多
关键词 Chlamydophila pneumoniae Major outer membrane protein Monoclonal antibody ELISA
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Gene cloning and prokaryotic expression of recombinant outer membrane protein from Vibrio parahaemolyticus
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作者 袁野 王秀利 +1 位作者 郭设平 仇雪梅 《Chinese Journal of Oceanology and Limnology》 SCIE CAS CSCD 2011年第5期952-957,共6页
Gram-negative Vibrio parahaemolyticus is a common pathogen in humans and marine animals, The outer membrane protein of bacteria plays an important role in the infection and pathogenicity to the host. Thus, the outer m... Gram-negative Vibrio parahaemolyticus is a common pathogen in humans and marine animals, The outer membrane protein of bacteria plays an important role in the infection and pathogenicity to the host. Thus, the outer membrane proteins are an ideal target for vaccines. We amplified a complete outer membrane protein gene (ompW) from E parahaemolyticus ATCC 17802. We then cloned and expressed the gene into Escherichia coli BL21 (DE3) cells. The gene coded for a protein that was 42.78 kDa. We purified the protein using Ni-NTA affinity chromatography and Anti-His antibody Western blotting, respectively. Our results provide a basis for future application of the OmpW protein as a vaccine candidate against infection by E parahaemolyticus. In addition, the purified OmpW protein can be used for further functional and structural studies. 展开更多
关键词 vibrio parahaemolyticus outer membrane protein ompW) CLONING prokaryotic expression protein characterization recombinant proteins
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Research progress on Helicobacter pyloriouter membrane protein
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作者 Shi-HeShao HuaWang +1 位作者 Shun-GenChai Li-MeiLiu 《World Journal of Gastroenterology》 SCIE CAS CSCD 2005年第20期3011-3013,共3页
Helicobacter pylori (H pylori), one of the most common bacterial pathogens on human beings, colonizes the gastric mucosa. In its 95 paralogous gene families, there is a large outer membrane protein (OMP) family. It in... Helicobacter pylori (H pylori), one of the most common bacterial pathogens on human beings, colonizes the gastric mucosa. In its 95 paralogous gene families, there is a large outer membrane protein (OMP) family. It includes 32 members. These OMP are important for the diagnosis, protective immunity, pathogenicity of H pylori and so on. They are significantly associated with high H pylori density,the damage of gastric mucosa, high mucosal IL-8 levels and severe neutrophil infiltration. We introduce their research progress on pathogenicity. 展开更多
关键词 H pylori outer membrane protein OIPA Gastric carcinoma
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Recombinant outer membrane protein F-B subunit of LT protein as a prophylactic measure against Pseudomonas aeruginosa burn infection in mice
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作者 Hassan Heydari Farsani Iraj Rasooli +2 位作者 Seyed Latif Mousavi Gargari Shahram Nazarian Shakiba Darwish Alipour Astaneh 《World Journal of Methodology》 2015年第4期230-237,共8页
AIM: To study immunogenicity of outer membrane protein F(Opr F) fused with B subunit of LT(LTB), against Pseudomonas aeruginosa(P. aeruginosa). METHODS: The Opr F, a major surface exposed outer membrane protein that i... AIM: To study immunogenicity of outer membrane protein F(Opr F) fused with B subunit of LT(LTB), against Pseudomonas aeruginosa(P. aeruginosa). METHODS: The Opr F, a major surface exposed outer membrane protein that is antigenically conserved in various strains of P. aeruginosa, is a promising immunogen against P. aeruginosa. In the present study recombinant Opr F and Opr F-LTB fusion gene was cloned, expressed and purified. BALB/c mice and rabbits were immunized using recombinant Opr F and Opr F-LTB and challenged at the burn site with P. aeruginosa lethal dose of 104 CFU. The protective efficacy of rabbit anti Opr F Ig G against P. aeruginosa burn infection was investigated by passive immunization. RESULTS: It has been well established that the LTB is a powerful immunomodulator with strong adjuvant activity. LTB as a bacterial adjuvant enhanced immunogenicity of Opr F and anti Opr F Ig G titer in serum was increased. Experimental findings showed significantly higher average survival rate in burned mice immunized with Opr F-LTB than immunized with Opr F or the control group. Rabbits anti Opr F Ig G brought about 75% survival of mice following challenge with P. aeruginosa. Post challenge hepatic and splenic tissues of mice group immunized with Opr F-LTB had significantly lower bacterial load than those immunized with Opr F or the control groups. CONCLUSION: These results demonstrate that LTBfused Opr F might be a potential candidate protein for a prophylactic measure against P. aeruginosa in burn infection. 展开更多
关键词 Pseudomonas AERUGINOSA outer membrane protein F B SUBUNIT of LT IMMUNIZATION Burn
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鲍曼不动杆菌OmpA蛋白的生物学特性分析
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作者 赵丹 李尼克 +3 位作者 李雯 彭春红 赵德刚 张湘燕 《贵州医药》 CAS 2024年第3期339-344,共6页
目的提取临床分离的鲍曼不动杆菌外膜蛋白A(OmpA)的基因,并进行生物学预测和分析。方法查询鲍曼不动杆菌OmpA基因序列,设计特异性PCR引物,以提取的鲍曼不动杆菌基因组DNA为模板,PCR扩增OmpA片段。回收目的片段,采用生物信息学软件分析O... 目的提取临床分离的鲍曼不动杆菌外膜蛋白A(OmpA)的基因,并进行生物学预测和分析。方法查询鲍曼不动杆菌OmpA基因序列,设计特异性PCR引物,以提取的鲍曼不动杆菌基因组DNA为模板,PCR扩增OmpA片段。回收目的片段,采用生物信息学软件分析OmpA蛋白的生物学特性。结果12株鲍曼不动杆菌多位点序列分型(MLST)序列分析有6个ST分型,分别是ST 208、ST 229、ST 191、ST 195、ST540、ST 1145,鲍曼不动杆菌OmpA基因序列全长为1070 bp的,单核苷酸多态性(SNP)位点较少,预测蛋白为跨膜蛋白,具有6种三级结构,均由β-桶状结构组成的保守结构域。结论临床分离的不同鲍曼不动杆菌株OmpA蛋白为结构相对保守的跨膜蛋白,具有维持细胞的形状和稳定性,参与细菌耐药机制及免疫原性作用。 展开更多
关键词 鲍曼不动杆菌 外膜蛋白A 空间结构 跨膜结构
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钩端螺旋体OmpR家族相关TCS 鉴定及功能初探
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作者 戴欣珏 夏文浩 +2 位作者 任雨轩 车名花 胡玮琳 《中国人兽共患病学报》 CAS CSCD 北大核心 2024年第3期197-202,共6页
目的建立钩端螺旋体(钩体)OmpR家族双组分系统(TCS)的鉴定方法,初步探讨OmpR家族TCS在钩体感染人巨噬细胞过程中对钩体主要外膜蛋白(OMPs)表达水平的调控作用。方法采用生物信息学技术预测致病性钩体赖株TCS的组氨酸激酶(HK)和应答调节... 目的建立钩端螺旋体(钩体)OmpR家族双组分系统(TCS)的鉴定方法,初步探讨OmpR家族TCS在钩体感染人巨噬细胞过程中对钩体主要外膜蛋白(OMPs)表达水平的调控作用。方法采用生物信息学技术预测致病性钩体赖株TCS的组氨酸激酶(HK)和应答调节蛋白(RR)OmpR功能结构域。采用激酶活性检测试剂盒和细菌双杂交技术进一步验证各HK激酶活性以及各HK与RR的相互作用。采用qRT-PCR法检测HK抗血清封闭前后钩体主要OMPs基因mRNA水平的变化。结果钩体56601株中LA2828、LA1710含有HATPase_c激酶催化结构域,LA2827、LA1709含有Response_reg与OmpR功能结构域,提示其分别可构成HK/OmpR-TCS。体外重组HK-2828和HK-1710蛋白均具有激酶活性,且分别与OmpR-2827和OmpR-1709存在直接的相互作用。HK-2828抗体封闭处理可使感染过程中显著下降的外膜蛋白编码基因lipL32、lipL41和ompL1的mRNA水平明显回升。结论成功构建钩体OmpR家族-TCS鉴定及功能探究体系,为阐明感染过程中问号钩体OMPs表达改变调控机制,以及制定基因工程疫苗OMPs抗原筛选新策略提供依据。 展开更多
关键词 钩端螺旋体 双组分系统 外膜蛋白
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外膜蛋白OmpA在蛙源米尔伊丽莎白菌致病性中的功能
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作者 刘芳园 胡瑞雪 +3 位作者 余芳 侯家昊 于子润 顾泽茂 《华中农业大学学报》 CAS CSCD 北大核心 2024年第1期203-209,共7页
为探究外膜蛋白A(outer membrane protein A,OmpA)对米尔伊丽莎白菌致病作用的影响,以蛙源米尔伊丽莎白菌FL160902为研究对象,通过同源重组法构建OmpA缺失株△ompA,比较缺失株和野生株的生长特性、生物膜形成能力、抗血清杀伤能力、对... 为探究外膜蛋白A(outer membrane protein A,OmpA)对米尔伊丽莎白菌致病作用的影响,以蛙源米尔伊丽莎白菌FL160902为研究对象,通过同源重组法构建OmpA缺失株△ompA,比较缺失株和野生株的生长特性、生物膜形成能力、抗血清杀伤能力、对细胞的黏附能力以及对蛙的致病性差异。结果显示:△ompA的生长能力和抗血清杀伤能力与野生株无显著差异;但与野生株相比,△ompA的生物膜形成能力增加了66%,△ompA对bEnd.3细胞的黏附能力降低了61%;黑斑蛙感染试验显示,△ompA在黑斑蛙血液、脾和脑组织中的载菌量分别为(3.15×10^(8)±0.09×10^(8))、(2.11×10^(8)±0.07×10^(8))和(6.61×10^(8)±0.16×10^(8))copies/g,均显著低于野生株,且△ompA对黑斑蛙的致死率为37%,显著低于野生株的致死率(75%)。上述结果表明,ompA基因缺失不改变米尔伊丽莎白菌的抗血清杀伤能力,但增加了菌株的生物膜形成能力,减弱了菌株的黏附能力,从而降低了该菌对蛙的致病性。 展开更多
关键词 米尔伊丽莎白菌 基因缺失 外膜蛋白A 致病性
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Immunogenic proteins and their vaccine development potential evaluation in outer membrane proteins(OMPs)of Flavobacterium columnare 被引量:1
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作者 Zhang Luo Jianping Fu +4 位作者 Nan Li Zhixing Liu Ting Qin Xiaolin Zhang Pin Nie 《Aquaculture and Fisheries》 2016年第1期1-8,共8页
Flavobacterium columnare is a Gram-negative bacterium that causes columnaris disease in freshwater fish worldwide.Many studies have focused on the identification of protective antigens to aid in the development of nov... Flavobacterium columnare is a Gram-negative bacterium that causes columnaris disease in freshwater fish worldwide.Many studies have focused on the identification of protective antigens to aid in the development of novel vaccines against the disease.In this study,an immunoblotting approach was employed to identify immunogenic outer membrane proteins(OMPs)from F.columnare in two-dimensional electrophoresis(2-DE)map gels using antibacterial sera obtained from grass carp(Ctenopharyngodon idella),and anti-grass carp-recombinant Ig(rIg)monoclonal antibodies.Five unique immunogenic proteins,including the gliding motility lipoprotein GldJ(GldJ),hypothetical protein FCOL_13420(Fco1),lipoprotein(Lip),F0F1 ATP synthase subunit beta(F0f1)and outer membrane efflux protein precursor(Omep),were characterized.Over-expression of these proteins in Escherichia coli DE3,and their immunogenicity and protective efficacy were evaluated in grass carp.The relative percent survival(RPS)of the groups immunized separately with recombinant GldJ,Lip and Omep was 72%,64%and 68%,respectively when compared to control fish.Up-regulation of immuno-related genes and specific antibodies were detected in immunized fish and sera of immunized fish inhibited the growth of F.columnare.The results suggest that GldJ,Lip and Omep are major protective antigens and may be considered as novel candidates in the development of vaccines against columnaris disease in fish. 展开更多
关键词 Flavobacterium columnare outer membrane protein Immunogenic protein VACCINE Grass carp
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军团菌MOMPS基因的克隆并在原核系统中表达
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作者 张雷 陈建平 +2 位作者 王涛 张莉 田玉 《生物医学工程学杂志》 EI CAS CSCD 北大核心 2006年第2期379-382,共4页
以军团菌DNA为模板,PCR扩增获得军团菌主要外膜蛋白基因(M a jor ou ter m em brane prote in gene,om pS),与原核表达质粒pUC 18定向重组,构建重组质粒,转化大肠杆菌BL 21,并用限制性酶酶切分析、聚合酶链式反应、核酸序列分析、十二... 以军团菌DNA为模板,PCR扩增获得军团菌主要外膜蛋白基因(M a jor ou ter m em brane prote in gene,om pS),与原核表达质粒pUC 18定向重组,构建重组质粒,转化大肠杆菌BL 21,并用限制性酶酶切分析、聚合酶链式反应、核酸序列分析、十二烷基磺酸钠-聚丙烯酰胺凝胶电泳、W estern印迹进行鉴定。实验结果表明我们扩增出了军团菌914 bp的om pS基因,成功构建了重组质粒pLPom pS,并在原核系统中得到了表达。 展开更多
关键词 军团菌 主要外膜蛋白基因(omps) 克隆 表达
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A Sensitive and Specific IgM-ELISA for the Serological Diagnosis of Human Leptospirosis Using a rLipL32/1-LipL21-OmpL1/2 Fusion Protein 被引量:6
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作者 SUN AiHua WANG Yuan +2 位作者 DU Peng WU ShengLing YAN Jie 《Biomedical and Environmental Sciences》 SCIE CAS CSCD 2011年第3期291-299,共9页
Objective To construct a lipL32//1-1ipL21-OmpL1//2 fusion gene and its prokaryotic expression system, and to establish an enzyme-linked immunosorbent assay (ELISA) using the rLipL32/1-LipL21-OmpL1/2 fusion antigen o... Objective To construct a lipL32//1-1ipL21-OmpL1//2 fusion gene and its prokaryotic expression system, and to establish an enzyme-linked immunosorbent assay (ELISA) using the rLipL32/1-LipL21-OmpL1/2 fusion antigen of Leptospira interrogans for sensitive and specific detection of IgM in the serum of patients with leptospirosis. Methods lipL32/1-1ipL21-OmpL1/2 fusion genes were constructed using a primer-linking PCFI. The target recombinant protein antigens, rLipL32/1, rLipL21, rOmpL1/2 and rLipL32/1-LipL21-OmpL1/2, were expressed and the purified antigens were then immobilized to the surface of microplate wells for ELISA-based detection of IgM in the sera of leptospirosis patients; Results Of 493 acute leptospirosis patients, 95.7% and 97.8% were positive by rLipL32/1-LipL21- OmpL1/2-1gM-ELISA using different serum dilutions, which was higher than the rLipL32/1-1gM-ELISA (93.1% and 90.3%), rLipL21-1gM-ELISA (90.3% and 87.0%), and rOmpLI-lgM-ELISA (85.6% and 81.1%) (P〈0.01). All IgM-ELISAs tested negative against 56 non-leptospirosis patients with typhoid fever, hemorrhagic fever or dengue fever. Conclusion Trigeminal fusion antigen increases ELISA sensitivity and the rLipL32/1-LipL21-OmpL1/2- IgM-ELISA is a sensitive and specific serological diagnostic method for clinical leptospirosis. 展开更多
关键词 LEPTOSPIRA outer membrane protein Fusion antigen Recombinant expression IgM-ELISA
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布鲁氏菌外膜蛋白OMP10间接ELISA检测方法的建立及免疫原性评估 被引量:2
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作者 邓肖玉 徐锦凤 +4 位作者 何金科 谢珊珊 王勇 王震 陈创夫 《中国人兽共患病学报》 CAS CSCD 北大核心 2023年第2期107-114,共8页
目的建立布鲁氏菌外膜蛋白OMP10间接ELISA检测方法,评价OMP10在小鼠体内的免疫效果。方法本研究表达和纯化了布鲁氏菌外膜蛋白OMP10,通过Western Blot进行验证,建立了OMP10间接ELISA检测方法,然后将OMP10与弗氏佐剂和LDH佐剂配伍成2种... 目的建立布鲁氏菌外膜蛋白OMP10间接ELISA检测方法,评价OMP10在小鼠体内的免疫效果。方法本研究表达和纯化了布鲁氏菌外膜蛋白OMP10,通过Western Blot进行验证,建立了OMP10间接ELISA检测方法,然后将OMP10与弗氏佐剂和LDH佐剂配伍成2种亚单位疫苗,免疫小鼠后检测抗体水平、脾脏淋巴细胞的增殖水平、CD4^(+)和CD8^(+)T细胞比值以及细胞因子分泌,最后通过攻毒试验评估OMP10的免疫保护效果。结果OMP10作为诊断抗原的特异性和符合率均高于70%;免疫小鼠后2种亚单位疫苗均能产生高滴度的IgG抗体,CD4^(+)/CD8^(+)T的比值均高于PBS对照组,IFN-γ高于PBS对照组,差异具有统计学意义(P<0.01),但IL-4未发生明显变化,表明OMP10能够诱导很好的体液免疫及Th1型免疫应答。免疫保护结果显示,2种亚单位疫苗免疫组小鼠脾脏指数和脾脏载菌量低于布鲁氏菌M5感染组,差异具有统计学意义(P<0.01),表明OMP10蛋白配伍的亚单位疫苗对布鲁氏菌M5感染小鼠具有高度的保护作用。结论外膜蛋白OMP10在布鲁氏菌诊断和疫苗方面具有一定的应用价值。 展开更多
关键词 布鲁氏菌 外膜蛋白omp10 间接ELISA 抗体水平 细胞因子 免疫保护
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SDS-PAGE analysis of whole cell protein and outer memrbane protein patterns of clinical isolates of Burkholderia pseudomallei 被引量:1
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作者 Aoichart Nontprasert Cheeraratana Cheeramakara +5 位作者 Sasithon Pukrittayakamee David AB Dance Ty L Pitt Michael D Smith Sirivan Vanijanonta Nicholas J White 《Asian Pacific Journal of Tropical Medicine》 SCIE CAS 2009年第5期14-19,共6页
Objective:To investigate the banding patterns of whole cell protein(WCP) and outer membrane protein (OMP) of Burkholderia pseudomallei(B.pseudomallei) in clinical isolates from patients with melioidosis. Methods:WCP a... Objective:To investigate the banding patterns of whole cell protein(WCP) and outer membrane protein (OMP) of Burkholderia pseudomallei(B.pseudomallei) in clinical isolates from patients with melioidosis. Methods:WCP and OMP of of B.pseudomallei in 50 clinical isolates,from 47 patients with melioidosis were prepared and separated by polyacrylamide gel electrophoresis(SDS-PAGE) using 10%gels and stained with Coomassie brilliant blue.The banding patterns were compared by using a laser densitometer and dendrogram. Results:There were 6 different banding patterns of WCP and 2 types of OMP.Type 1 -5 WCP had 8 common protein bands at 19.0 - 45.0 kDa with identical OMP pattern.The banding patterns of WCP in type 6 were distinct from the others and also its OMP profile.The majority of clinical isolates(37/50,74%) were in type 1 WCP.Of the remaining isolates,8 were in type 2,2 in type 3,and one each was in type 4 to 6.There was no significant association between the WCP typing and the demographic or clinical features of the investigated patients.Conclusion:Despite the wide variation of clinical features of melioidosis,the results of this study show that B.pseudomallei had a few differences in the WCP and OMP profiles.Therefore typing of WCP and OMP,using SDS-PAGE analysis,could be an alternative method for phenotypic differentiation in clinical isolates of B.pseudomallei. 展开更多
关键词 SDS-PAGE BURKHOLDERIA pseudomallei WHOLE cell protein outer membrane protein MELIOIDOSIS
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Comparison of Lipopolysaccharide and Protein Immunogens from Pathogenic Yersinia enterocolitica Bio-serotype 1B/O:8 and 2/O:9 using SDS-PAGE
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作者 GU Wen Peng WANG Xin +6 位作者 QIU Hai Yan LUO Xia XIAO Yu Chun TANG Liu Ying KAN Biao XU Jian Guo JING Huai Qi 《Biomedical and Environmental Sciences》 SCIE CAS CSCD 2012年第3期282-290,共9页
Objective Yersinia enterocolitica is an extracellular pathogen and its related antigens interact with the host immune system. We investigated the difference in immunological characteristics between a highly pathogenic... Objective Yersinia enterocolitica is an extracellular pathogen and its related antigens interact with the host immune system. We investigated the difference in immunological characteristics between a highly pathogenic and poorly pathogenic strain of Y. enterocolitico. Methods We used SDS-PAGE and western blotting to characterize lipopolysaccharide (LPS), Yersinio outer membrane proteins (Yops), membrane proteins, and whole-cell proteins from poorly pathogenic Y. enterocolitico bio-serotype 2/0:9, isolated from China, and highly pathogenic bio-serotype 1B/O:8, isolated from Japan. Results These two strains of Y. enterocolitica had different LPS immune response patterns. Comparison of their Yops also showed differences that could have accounted for their differences in pathogenicity. The membrane and whole-cell proteins of both strains were similar; immunoblottting showed that the 35 kD and perhaps the 10 kD proteins were immunogens in both strains. Conclusion The major antigens of the two strains e and membrane proteins, as shown by comparing preparations. citing the host immune protein samples with response were the LPS reference and purified 展开更多
关键词 Immunogens LIPOPOLYSACCHARIDES outer membrane proteins Whole-cell proteins Pathogenic Yersinia enterocolitica
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