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
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
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
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
OBJECTIVE: Helicobacter pylori is a Gram-negative organism. Its outer membrane protein Q(Hop Q) mediates host-pathogen interactions; Hop Q genotypes 1 and 2 are found associating with gastroduodenal pathologies. Th...OBJECTIVE: Helicobacter pylori is a Gram-negative organism. Its outer membrane protein Q(Hop Q) mediates host-pathogen interactions; Hop Q genotypes 1 and 2 are found associating with gastroduodenal pathologies. The authors measured the anti-adhesion effects of the extracts of Abelmoschus esculentus, Zingiber officinale, Trachyspermum ammi, Glycyrrhiza glabra, Curcuma longa and Capsicum annum against Hop Q genotypes and H. pylori cytotoxin-associated gene A(Cag A).METHODS: DNA was extracted by polymerase chain reaction of the Hop Q genotypes(i.e., type 1, type 2 and Cag A) from 115 H. pylori strains. The effect of the extracts from selected dietary ingredients was determined using a gastric adenocarcinoma cell line and a quantitative DNA fragmentation assay. The anti-adhesive effect of these extracts on H. pylori was tested using an anti-adhesion analysis.RESULTS: C. annum, C. longa and A. esculentus showed prominent anti-adhesion effects with resultant values of 17.3% ± 2.9%, 14.6% ± 3.7%, 13.8% ± 3.6%, respectively, against Hop Q type 1 and 13.1% ± 1.7%, 12.1% ± 2%, 11.1% ± 1.6%, respectively, against Hop Q type 2. C. longa(93%), C. annum(89%) and A. esculentus(75%) had better anti-adhesive activity against H. pylori with Hop Q type 1 compared to Hop Q type 2 with respective values of 70%, 64% and 51%. Extracts of C. annum(14.7% ± 4.1%), A. esculentus(12.3% ± 4.1%) and Z. officinale(8.4% ± 2.8%) had an anti-adhesion effect against Cag A-positive H. pylori strains compared to Cag A-negative strains.展开更多
SurA is the major chaperone of outer membrane proteins(OMPs)in the periplasm.The molecular mechanism when SurA performs its chaperoning function is still unclear.Here,a purification-after-crosslinking(PAC)procedure wa...SurA is the major chaperone of outer membrane proteins(OMPs)in the periplasm.The molecular mechanism when SurA performs its chaperoning function is still unclear.Here,a purification-after-crosslinking(PAC)procedure was combined with single-molecule fluorescence resonance energy transfer(smFRET)to probe the conformations of SurA and OmpC in their complex.We found that SurA in the free state rearranges itself based on the crystal structure,except that the P2 domain moves towards the core domain with two major positions,forming a clamp-like conformation to accommodate OmpC.The obvious rearrangement of the P2 domain of SurA helps SurA to hold OmpC.OmpC attaches to SurA randomly and has the propensity to be near the middle part of the crevice.The noncollapsed and disordered conformations of OMPs provided by the OMPs?SurA complex are important to the subsequent delivery and folding process.展开更多
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
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
Objective:To assess the efficacy of various types of vaccines developed for leptospirosis.Methods:A comprehensive search was conducted in three databases:PubMed,Scopus,and Cochrane Library.Two authors(YS and MN)select...Objective:To assess the efficacy of various types of vaccines developed for leptospirosis.Methods:A comprehensive search was conducted in three databases:PubMed,Scopus,and Cochrane Library.Two authors(YS and MN)selected the articles based on manual screening.The study eligibility criteria are all Leptospira species regardless of any cluster(pathogenic,intermediate and non-pathogenic).This study recorded articles with positive and negative results and showed a comparison among various membrane proteins as vaccine candidates.The studies on the effectiveness of outer membrane protein as vaccine candidates were also included.The articles obtained in the databases were imported into the WPS spreadsheet,and duplicate documents were removed manually.Results:A total of 24 studies were included in the review,which evaluated various types of leptospirosis vaccines.Multiple vaccines were developed and tested;however,the heterogeneity of Leptospira species pose a challenge.As an effective approach,an epitope based vaccine shows quite a promising result.However,sufficient validation,testing and clinical trials are required.Conclusions:Developing an effective vaccine for leptospirosis remains a global health priority.While significant progress has been made in recent years,there is a need for further research to optimize vaccine development and to ensure that vaccines are accessible and effective for high-risk populations.展开更多
Helicobacter pylori(H.pylori)is one of the most important human pathogens,infecting approximately half of the global population.Despite its high prevalence,only a subset of H.pylori infected individuals develop seriou...Helicobacter pylori(H.pylori)is one of the most important human pathogens,infecting approximately half of the global population.Despite its high prevalence,only a subset of H.pylori infected individuals develop serious gastroduodenal pathology.The pathogenesis of H.pylori infection and disease outcome is thus thought to be mediated by an intricate interplay between host,environmental and bacterial virulence factors.H.pylori has adapted to the harsh milieu of the human stomach through possession of various virulence genes that enable survival of the bacteria in the acidic environment,movement towards the gastric epithelium,and attachment to gastric epithelial cells.These virulence factors enable successful colonization of the gastric mucosa and sustain persistent H.pylori infection,causing chronic inflammation and tissue damage,which may eventually lead to the development of peptic ulcers and gastric cancer.Numerous studies have focused on the prevalence and role of putative H.pylori virulence genes in disease pathogenesis.While several virulence factors with various functions have been identified,disease associations appear to be less evident,especially among different study populations.This review presents key findings on the most important H.pylori virulence genes,including several bacterial adhesins and toxins,in children and adults,and focuses on their prevalence,clinical significance and potential relationships.展开更多
[Objective]This study aimed to investigate the mutations of OmpF from an isolated antibiotic resistant Escherichia coli strain.[Methods]The mutant OmpF(mOmpF)from antibiotic resistant E.coli was amplified by PCR wit...[Objective]This study aimed to investigate the mutations of OmpF from an isolated antibiotic resistant Escherichia coli strain.[Methods]The mutant OmpF(mOmpF)from antibiotic resistant E.coli was amplified by PCR with Pfu DNA polymerase and ligated into the expression vector pET28a.Subsequently,the expression vector pET28-mOmpF was sequenced and analyzed by DNAMAN software and Swiss-Model online.[Result]Sequence analysis revealed that the open reading fragment of mOmpF was 903 bp long,which was mutated dramatically compared to that of the 1 020 bp long model OmpF.The DNA sequence shared only54.5%homology with OmpF.mOmpF was 44.6%identical to that of OmpF.Protein structure predication and analysis through Swiss-Model online suggested that the structure of mOmpF changed dramatically compared to OmpF.[Conclusion]The present study provided basis for further analyzing the relationships between the structure and functions of mOmpF from antibiotic resistant E.coli.展开更多
Chemokines are cytokines that can promote the activation and migration of immune cells,and increase the recognition of antigen by antigen-presenting cells(APC).Previous studies showed that a DNA vaccine can induce hum...Chemokines are cytokines that can promote the activation and migration of immune cells,and increase the recognition of antigen by antigen-presenting cells(APC).Previous studies showed that a DNA vaccine can induce humoral and cellular immune responses of flounder after immunization.To explore the improvement of chemokines on the efficiency of OmpK vaccine,two bicistronic DNA candidate vaccines were constructed and the immune responses they induced in the flounder were investigated by reverse transcription polymerase chain reaction(RT-PCR),indirect immunofl uorescent assay(IFA),H&E staining,fl ow cytometry(FCM),and quantifi cational real-time polymerase chain reaction(qRT-PCR).pBudCE4.1 plasmid as an expression vector,bicistronic DNA vaccines encoding OmpK gene and CC-motif ligand 4 gene(p-OmpK-CCL4),or Ompk gene and CC-motif ligand 19 gene(p-OmpK-CCL19)were successfully constructed.The results showed that two bicistronic DNA vaccines expressed Ompk protein of Vibrio anguillarum and CCL4/CCL19 proteins of fl ounder both in vitro and in vivo.After immunization,a large number of leucocytes in muscle were recruited at the injection site in treatment groups.The constructed vaccines induced signifi cant increases in CD4-1^(+) and CD4-2^(+) T lymphocytes,and sIgM^(+) B lymphocytes in peripheral blood,spleen,and head kidney.The percentage of T lymphocytes peaked on the 14^(th) post-vaccination day whereas that of B lymphocytes peaked in the 6^(th) post-vaccination week.Moreover,the expression profi les of 10 immune-related genes increased in muscles around the injection site,spleen,and head kidney.After the challenge,p-OmpK-CCL4 and p-OmpK-CCL19 conferred a relative percentage survival(RPS)of 74.1%and 63.3%,respectively,higher than p-OmpK alone(40.8%).In conclusion,both CCL4 and CCL19 can improve the protection of p-OmpK via evoking local immune response and then humoral and cellular immunity.CCL4 and CCL19 will be potential molecular adjuvants for use in DNA vaccines.展开更多
Neisseria meningitidis (N. meningitidis) is classified into 13 serogroups based on the immunological reactivity of the capsular polysaccharide.Serogourp-s A,B and C are responsible for over 90% of meningococcal dise...Neisseria meningitidis (N. meningitidis) is classified into 13 serogroups based on the immunological reactivity of the capsular polysaccharide.Serogourp-s A,B and C are responsible for over 90% of meningococcal disease.2 In developed countries, endemic disease is generally caused by serogroups B and C.展开更多
基金supported by grants from the National Natural Science Foundation of China (Grant No. 30901352)Innovative Research Team in University of Hunan Province (Number: [2008] 51)Hunan Provincial Innovation Foundation for Postgraduate and Hunan Provincial Training and Innovation Base for Post-graduate
文摘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.
基金the National Natural Science Foundation of China(31072151)the Specialized Research Fund for the Doctoral Program of Higher Education,China(20090097110007)the Priority Academic Program Development of Jiangsu Higher Education Institutions,China(PAPD)
文摘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.
基金supported by the China Agricultural Research System(nycytx-44-3-2)the Zhejiang Key Project on Agricultural Development through Science and Technology(2011C12028)+1 种基金the National Natural Science Foundation of China(31302068)the Zhejiang Provincial Natural Science Foundation of China(LQ13C180002)
文摘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.
文摘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.
文摘OBJECTIVE: Helicobacter pylori is a Gram-negative organism. Its outer membrane protein Q(Hop Q) mediates host-pathogen interactions; Hop Q genotypes 1 and 2 are found associating with gastroduodenal pathologies. The authors measured the anti-adhesion effects of the extracts of Abelmoschus esculentus, Zingiber officinale, Trachyspermum ammi, Glycyrrhiza glabra, Curcuma longa and Capsicum annum against Hop Q genotypes and H. pylori cytotoxin-associated gene A(Cag A).METHODS: DNA was extracted by polymerase chain reaction of the Hop Q genotypes(i.e., type 1, type 2 and Cag A) from 115 H. pylori strains. The effect of the extracts from selected dietary ingredients was determined using a gastric adenocarcinoma cell line and a quantitative DNA fragmentation assay. The anti-adhesive effect of these extracts on H. pylori was tested using an anti-adhesion analysis.RESULTS: C. annum, C. longa and A. esculentus showed prominent anti-adhesion effects with resultant values of 17.3% ± 2.9%, 14.6% ± 3.7%, 13.8% ± 3.6%, respectively, against Hop Q type 1 and 13.1% ± 1.7%, 12.1% ± 2%, 11.1% ± 1.6%, respectively, against Hop Q type 2. C. longa(93%), C. annum(89%) and A. esculentus(75%) had better anti-adhesive activity against H. pylori with Hop Q type 1 compared to Hop Q type 2 with respective values of 70%, 64% and 51%. Extracts of C. annum(14.7% ± 4.1%), A. esculentus(12.3% ± 4.1%) and Z. officinale(8.4% ± 2.8%) had an anti-adhesion effect against Cag A-positive H. pylori strains compared to Cag A-negative strains.
基金supported by the National Natural Science Foundation of China(21233002,21521003)the National Key Basic Research Special Foundation of China(2012CB917304)。
文摘SurA is the major chaperone of outer membrane proteins(OMPs)in the periplasm.The molecular mechanism when SurA performs its chaperoning function is still unclear.Here,a purification-after-crosslinking(PAC)procedure was combined with single-molecule fluorescence resonance energy transfer(smFRET)to probe the conformations of SurA and OmpC in their complex.We found that SurA in the free state rearranges itself based on the crystal structure,except that the P2 domain moves towards the core domain with two major positions,forming a clamp-like conformation to accommodate OmpC.The obvious rearrangement of the P2 domain of SurA helps SurA to hold OmpC.OmpC attaches to SurA randomly and has the propensity to be near the middle part of the crevice.The noncollapsed and disordered conformations of OMPs provided by the OMPs?SurA complex are important to the subsequent delivery and folding process.
基金supported by the National Science and Technology Key Program for Infectious Diseases of China (Grant No. 2008ZX10004‐015)the Natural Science Foundation of Zhejiang Medical College in China (No. 2007XZA02)
文摘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.
基金supported by National Natural Science Foundation of China (General Project,No. 30970094)National Sci-Tech Key Project (2009ZX10004-201,2009ZX10004-203)
文摘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
基金funded by Grant Putra Inisiatif Putra Muda,Universiti Putra Malaysia.Grant number GP-IPM/2020/9690700.
文摘Objective:To assess the efficacy of various types of vaccines developed for leptospirosis.Methods:A comprehensive search was conducted in three databases:PubMed,Scopus,and Cochrane Library.Two authors(YS and MN)selected the articles based on manual screening.The study eligibility criteria are all Leptospira species regardless of any cluster(pathogenic,intermediate and non-pathogenic).This study recorded articles with positive and negative results and showed a comparison among various membrane proteins as vaccine candidates.The studies on the effectiveness of outer membrane protein as vaccine candidates were also included.The articles obtained in the databases were imported into the WPS spreadsheet,and duplicate documents were removed manually.Results:A total of 24 studies were included in the review,which evaluated various types of leptospirosis vaccines.Multiple vaccines were developed and tested;however,the heterogeneity of Leptospira species pose a challenge.As an effective approach,an epitope based vaccine shows quite a promising result.However,sufficient validation,testing and clinical trials are required.Conclusions:Developing an effective vaccine for leptospirosis remains a global health priority.While significant progress has been made in recent years,there is a need for further research to optimize vaccine development and to ensure that vaccines are accessible and effective for high-risk populations.
文摘Helicobacter pylori(H.pylori)is one of the most important human pathogens,infecting approximately half of the global population.Despite its high prevalence,only a subset of H.pylori infected individuals develop serious gastroduodenal pathology.The pathogenesis of H.pylori infection and disease outcome is thus thought to be mediated by an intricate interplay between host,environmental and bacterial virulence factors.H.pylori has adapted to the harsh milieu of the human stomach through possession of various virulence genes that enable survival of the bacteria in the acidic environment,movement towards the gastric epithelium,and attachment to gastric epithelial cells.These virulence factors enable successful colonization of the gastric mucosa and sustain persistent H.pylori infection,causing chronic inflammation and tissue damage,which may eventually lead to the development of peptic ulcers and gastric cancer.Numerous studies have focused on the prevalence and role of putative H.pylori virulence genes in disease pathogenesis.While several virulence factors with various functions have been identified,disease associations appear to be less evident,especially among different study populations.This review presents key findings on the most important H.pylori virulence genes,including several bacterial adhesins and toxins,in children and adults,and focuses on their prevalence,clinical significance and potential relationships.
基金Supported by the National Natural Science Foundation of China(No.31100089)Scientific Research Foundation of the Education Department of Sichuan Province(No.11ZB102)the Talent Project of Sichuan University of Science&Engineering(No.2011RC12)
文摘[Objective]This study aimed to investigate the mutations of OmpF from an isolated antibiotic resistant Escherichia coli strain.[Methods]The mutant OmpF(mOmpF)from antibiotic resistant E.coli was amplified by PCR with Pfu DNA polymerase and ligated into the expression vector pET28a.Subsequently,the expression vector pET28-mOmpF was sequenced and analyzed by DNAMAN software and Swiss-Model online.[Result]Sequence analysis revealed that the open reading fragment of mOmpF was 903 bp long,which was mutated dramatically compared to that of the 1 020 bp long model OmpF.The DNA sequence shared only54.5%homology with OmpF.mOmpF was 44.6%identical to that of OmpF.Protein structure predication and analysis through Swiss-Model online suggested that the structure of mOmpF changed dramatically compared to OmpF.[Conclusion]The present study provided basis for further analyzing the relationships between the structure and functions of mOmpF from antibiotic resistant E.coli.
基金Supported by the National Natural Science Foundation of China(Nos.32173005,31730101,31672684)the National Key Research and Development Program of China(No.2018YFD0900503)+5 种基金the Shandong Provincial Natural Science Foundation(No.ZR2020KC025)the Fundamental Research Funds for the Central Universities(No.201822015)the Director Foundation of Functional Laboratory for Marine Fisheries Science and Food Production Processes,Qingdao National Laboratory for Marine Science and Technology(No.2018MFSD-01)the NBRPC(No.2012CB114406)the Key Research and Development Program of Shandong Province(No.2016GNC115001)the Taishan Scholar Program of Shandong Province。
文摘Chemokines are cytokines that can promote the activation and migration of immune cells,and increase the recognition of antigen by antigen-presenting cells(APC).Previous studies showed that a DNA vaccine can induce humoral and cellular immune responses of flounder after immunization.To explore the improvement of chemokines on the efficiency of OmpK vaccine,two bicistronic DNA candidate vaccines were constructed and the immune responses they induced in the flounder were investigated by reverse transcription polymerase chain reaction(RT-PCR),indirect immunofl uorescent assay(IFA),H&E staining,fl ow cytometry(FCM),and quantifi cational real-time polymerase chain reaction(qRT-PCR).pBudCE4.1 plasmid as an expression vector,bicistronic DNA vaccines encoding OmpK gene and CC-motif ligand 4 gene(p-OmpK-CCL4),or Ompk gene and CC-motif ligand 19 gene(p-OmpK-CCL19)were successfully constructed.The results showed that two bicistronic DNA vaccines expressed Ompk protein of Vibrio anguillarum and CCL4/CCL19 proteins of fl ounder both in vitro and in vivo.After immunization,a large number of leucocytes in muscle were recruited at the injection site in treatment groups.The constructed vaccines induced signifi cant increases in CD4-1^(+) and CD4-2^(+) T lymphocytes,and sIgM^(+) B lymphocytes in peripheral blood,spleen,and head kidney.The percentage of T lymphocytes peaked on the 14^(th) post-vaccination day whereas that of B lymphocytes peaked in the 6^(th) post-vaccination week.Moreover,the expression profi les of 10 immune-related genes increased in muscles around the injection site,spleen,and head kidney.After the challenge,p-OmpK-CCL4 and p-OmpK-CCL19 conferred a relative percentage survival(RPS)of 74.1%and 63.3%,respectively,higher than p-OmpK alone(40.8%).In conclusion,both CCL4 and CCL19 can improve the protection of p-OmpK via evoking local immune response and then humoral and cellular immunity.CCL4 and CCL19 will be potential molecular adjuvants for use in DNA vaccines.
文摘Neisseria meningitidis (N. meningitidis) is classified into 13 serogroups based on the immunological reactivity of the capsular polysaccharide.Serogourp-s A,B and C are responsible for over 90% of meningococcal disease.2 In developed countries, endemic disease is generally caused by serogroups B and C.