Objectives: To evaluate the immunogenicity of Methods: Protein extracts from Results: Immunization with Conclusion This is the advanced study to investigate the immunogenicity of
Animals ; Antibodies, Bacterial/immunology* ; Antigens, Bacterial/immunology* ; Bacterial Proteins/immunology* ; Cross Reactions ; Cytokines/immunology* ; Female ; Genome, Bacterial ; Immunoglobulin G/immunology* ; Immunoglobulin M/immunology* ; Macrophages/immunology* ; Mice, Inbred BALB C ; Mycobacterium avium Complex/immunology* ; Mycobacterium tuberculosis/immunology* ; Tuberculosis Vaccines/administration & dosage* ; Whole Genome Sequencing

OBJECTIVE: This study aimed to characterize the diagnostic and vaccine potential of a novel Mycobacterium tuberculosis antigen Rv0674. METHODS: To evaluate the diagnostic potential and antigenicity of Rv0674, IgG was evaluated using ELISA and interferon (IFN)-γ was done by using ELISpot assay among TB patients and healthy donors. For immunogenicity evaluation, BALB/c mice were immunized with Rv0674. Cytokine production was determined by cytokine release assay using an ELISA kit, and the antibodies were tested using ELISA. RESULTS: The results of serum Elisa tests showed that Rv0674 specific immunoglobulin G (IgG) response was higher in TB patients than negative controls. And Rv0674 had good performance in serological test with sensitivity and specificity of 77.1% and 81.1%, respectively. While it shows poor sensitivity and specificity of 26.23% and 79.69% for IFN-γ tests. In BALB/c mice, Rv0674 adjuvant by DDA/Poly I:C could also induce a high level of IFN-γ, interleukin-2 and interleukin-6 as well as a high IgG titer in both high- and low-dose groups indicating that Rv0674 is essential in humoral and cellular immunity. Moreover, the cytokine profile and IgG isotype characterized Rv0674 as a Th1/Th2-mixed-type protective immunity with the predominance of Th1 cytokines. CONCLUSION Rv0674 may be a good potential candidate for the development of TB serological diagnosis and a new TB vaccine.
Adult ; Aged ; Animals ; Antigens, Bacterial ; immunology ; Female ; Humans ; Immunity, Cellular ; Immunity, Humoral ; Male ; Mice ; Mice, Inbred BALB C ; Middle Aged ; Tuberculosis ; diagnosis ; immunology ; Young Adult

Adult ; Alternaria ; immunology ; Alveolitis, Extrinsic Allergic ; immunology ; Animals ; Antibodies ; immunology ; Antibodies, Bacterial ; immunology ; Antibodies, Fungal ; immunology ; Antigens ; immunology ; Antigens, Bacterial ; immunology ; Antigens, Fungal ; immunology ; Aspergillus fumigatus ; immunology ; Asymptomatic Diseases ; Candida albicans ; immunology ; Cladosporium ; immunology ; Columbidae ; immunology ; Female ; Healthy Volunteers ; Humans ; Immunoglobulin G ; immunology ; Male ; Melopsittacus ; immunology ; Middle Aged ; Mucor ; immunology ; Nocardia ; immunology ; Parrots ; immunology ; Penicillium chrysogenum ; immunology ; Stachybotrys ; immunology ; Thermoactinomyces ; immunology

The levels of Streptococcus (S.) mutans infections in saliva were evaluated and a comparison for specific antibody levels among children with different levels of S. mutans infection was made. The promising epitopic regions of antigen AgI/II (PAc) and glucosyltransferase (GTF) for potential vaccine targets related to S. mutans adherence were screened. A total of 94 children aged 3-4 years were randomly selected, including 53 caries-negative and 41 caries-positive children. The values of S. mutans and those of salivary total secretory immunoglobulin A (sIgA), anti-PAc and anti-Glucan binding domain (anti-GLU) were compared to determine the correlation among them. It was found the level of s-IgA against specific antigens did not increase with increasing severity of S. mutans infection, and the complete amino acid sequence of PAc and GTFB was analyzed using the DNAStar Protean system for developing specific anti-caries vaccines related to S. mutans adherence. A significantly positive correlation between the amount of S. mutans and children decayed, missing, and filled teeth index was observed. No significant difference was detected in specific sIgA against PAc or GLU between any two groups. No significant correlation was found between such specific sIgA and caries index. A total of 16 peptides from PAc as well as 13 peptides from GTFB were chosen for further investigation. S. mutans colonization contributed to early children caries as an important etiological factor. The level of sIgA against specific antigens did not increase with increasing severity of S. mutans infection in children. The epitopes of PAc and GTF have been screened to develop the peptide-based or protein-based anti-caries vaccines.
Antibodies, Bacterial ; biosynthesis ; Antigens, Bacterial ; chemistry ; immunology ; Bacterial Proteins ; chemistry ; immunology ; Case-Control Studies ; Child, Preschool ; Dental Caries ; immunology ; pathology ; prevention & control ; Epitopes ; chemistry ; immunology ; Female ; Glucosyltransferases ; chemistry ; immunology ; Humans ; Immunoglobulin A, Secretory ; biosynthesis ; Male ; Peptides ; chemistry ; immunology ; Saliva ; chemistry ; microbiology ; Severity of Illness Index ; Streptococcal Vaccines ; biosynthesis ; chemistry ; immunology ; Streptococcus mutans ; chemistry ; immunology ; pathogenicity ; Vaccines, Subunit ; Virulence Factors ; chemistry ; immunology

Lipooligosacharide (LOS) of Neisseria gonorrhoeae (gonococci, GC) is involved in the interaction of GC with host cells. Deletion of the alpha-oligosaccharide (alpha-OS) moiety of LOS (lgtF mutant) significantly impairs invasion of GC into epithelial cell lines. GC opacity (Opa) proteins, such as OpaI, mediate phagocytosis and stimulate chemiluminescence responses in neutrophils in part through interaction with members of the carcinoembryonic antigen (CEA) family, which includes CEACAM3 (CD66d), a human neutrophil specific receptor for phagocytosis of bacteria. In the present work, we examined the effects of OpaI-expressing lgtF mutant on phagocytosis by HeLa-CEACAM3 cells and chemiluminescence responses in neutrophils. The results showed that lgtF mutant even expressing OpaI completely lost the ability to promote either phagocytosis mediated by CEACAM3 interaction in HeLa cells or chemiluminescence responses in neutrophils. These data indicated that Opa proteins in the lgtF mutant, which might result from the conformational change, cannot be functional.
Antigens, Bacterial ; chemistry ; genetics ; immunology ; metabolism ; Carbohydrate Sequence ; Carcinoembryonic Antigen ; genetics ; immunology ; Gene Expression Regulation ; HeLa Cells ; Host-Pathogen Interactions ; Humans ; Lipopolysaccharides ; chemistry ; immunology ; Luminescent Measurements ; Mutation ; Neisseria gonorrhoeae ; genetics ; metabolism ; pathogenicity ; Neutrophils ; immunology ; microbiology ; Phagocytosis

Recombinant Fl-V (rFl-V) fusion protein is the main ingredient of the current candidate vaccine against Yersinia pestis infection, which has been under investigation in clinical trial in USA. We investigated the soluble expression conditions of rF1-V in Escherichia coli BL21 (DE3) that we constructed before. After scale-up and optimization of fermentation processes, we got the optimized fermentation process parameters: the culture was induced at the middle exponential phase with 50 µmol/L of IPTG at 25 °C for 5 h. Soluble rFl-V protein was isolated to 99% purity by ammonium sulfate precipitation, ion exchange chromatography, hydrophobic chromatography and gel filter chromatography. The protein recovery was above 20%. Protein identity and primary structure were verified by mass spectrometry and Edman sequencing. Results of purity, quality and western blotting analysis indicated that the target protein is a consistent and properly folded product. Furthermore, the immunogenicity of various antigens formulated with aluminum hydroxide adjuvant was evaluated in mice. Serum antibody titers of 4 groups including 20 µg rFl, rV and rFl-V and 10 µg rFl+10 µg rV, were assayed by ELISA after 2 doses. The antibody titers of anti-Fl with 20 µg rFl-V were obviously higher than titers with other groups; meanwhile there were no significant difference of anti-V antibody titers among them. These findings confirm that rFl-V would be the active pharmaceutical ingredient of the plague subunit vaccine.
Adjuvants, Immunologic ; Animals ; Antibodies, Bacterial ; blood ; Antibody Formation ; Antigens, Bacterial ; immunology ; Blotting, Western ; Chromatography, Ion Exchange ; Enzyme-Linked Immunosorbent Assay ; Mice ; Plague ; prevention & control ; Plague Vaccine ; immunology ; Recombinant Fusion Proteins ; immunology ; Vaccines, Subunit ; immunology ; Yersinia pestis

To analyze the immunogenicity and protective ability of recombinant IgG-binding protein (EAG) of Streptococcus equi subspecies equi and to evaluate its value when used as equine vaccine antigen, EAG gene was amplified by PCR and inserted into pET-28a vector. The EAG recombinant proteins were expressed and purified to immune mice. The serum antibody and challenge protection were tested. The purified recombinant protein of EAG was 26 kDa, and the protein reacted specifically with positive serum of Streptococcus equi subspecies equi. The mice antibody level for EAG immunization group was 1∶8 100. The immunological protection result showed that the protection rate of the EAG recombinant protein was 90%. The results suggested that the EAG protein has good immunogenicity and immunological protection, and it can effectively increase the humoral immune response and immunological protection of mice.
Animals ; Antibodies, Bacterial ; blood ; Antigens, Bacterial ; immunology ; Bacterial Proteins ; immunology ; Bacterial Vaccines ; immunology ; Immunity, Humoral ; Immunoglobulin G ; blood ; Mice ; Polymerase Chain Reaction ; Protein Binding ; Recombinant Proteins ; immunology ; Streptococcal Infections ; prevention & control ; Streptococcus equi ; Vaccination

We tested correlations between anti-Helicobacter pylori IgG and IgA levels and the urease test, anti-CagA protein antibody, degree of gastritis, and age. In total, 509 children (0-15 years) were enrolled. Subjects were stratified as 0-4 years (n = 132), 5-9 years (n = 274), and 10-15 years (n = 103) and subjected to the urease test, histopathology, ELISA, and western blot using whole-cell lysates of H. pylori strain 51. The positivity rate in the urease test (P = 0.003), the degree of chronic gastritis (P = 0.021), and H. pylori infiltration (P < 0.001) increased with age. The median titer for anti-H. pylori IgG was 732.5 IU/mL at 0-4 years, 689.0 IU/mL at 5-9 years, and 966.0 IU/mL at 10-15 years (P < 0.001); the median titer for anti-H. pylori IgA was 61.0 IU/mL at 0-4 years, 63.5 IU/mL at 5-9 years, and 75.0 IU/mL at 10-15 years (P < 0.001). The CagA-positivity rate was 26.5% at 0-4 years, 36.5% at 5-9 years, and 46.6% at 10-15 years for IgG (P = 0.036), and 11.3% at 0-4 years, 18.6% at 5-9 years, and 23.3% at 10-15 years for IgA (P < 0.001). Anti-H. pylori IgG and IgA titers increased with the urease test grade, chronic gastritis degree, active gastritis, and H. pylori infiltration. Presence of CagA-positivity is well correlated with a high urease test grade and high anti-H. pylori IgG/IgA levels.
Adolescent ; Antibodies, Bacterial/*blood ; Antigens, Bacterial/*analysis/immunology ; Bacterial Proteins/*analysis/immunology/metabolism ; Blotting, Western ; Child ; Child, Preschool ; Chronic Disease ; Enzyme-Linked Immunosorbent Assay ; Female ; Gastritis/pathology ; Helicobacter Infections/blood/microbiology/*pathology ; Helicobacter pylori/isolation & purification/*metabolism ; Humans ; Immunoglobulin A/*blood ; Immunoglobulin G/*blood ; Infant ; Infant, Newborn ; Male ; Severity of Illness Index ; Urease/metabolism

Leptospirosis, a zoonotic disease that is caused by many serovars which are more than 200 in the world, is an emerging worldwide disease. Accurate and rapid diagnostic tests for leptospirosis are a critical step to diagnose the disease. There are some commercial kits available for diagnosis of leptospirosis, but the obscurity of a species- or genus-specific antigen of pathogenic Leptospira interrogans causes the reduced sensitivity and specificity. In this study, the polysaccharide derived from lipopolysaccharide (LPS) of nonpathogenic Leptospira biflexa serovar patoc was prepared, and the antigenicity was confirmed by immunoblot and enzyme linked immunosorbent assay (ELISA). The performance of the rapid diagnostic test (RDT) kit using the polysaccharide as a diagnostic antigen was evaluated in Korea, Bulgaria and Argentina. The sensitivity was 93.9%, 100%, and 81.0% and the specificity was 97.9%, 100%, and 95.4% in Korea (which is a rare region occurring with 2 serovars mostly), Bulgaria (epidemic region with 3 serovars chiefly) and Argentina (endemic region with 19 serovars mainly) respectively. These results indicate that this RDT is applicable for global diagnosis of leptospirosis. This rapid and effective diagnosis will be helpful for diagnosis and manage of leptospirosis to use and the polysaccharide of Leptospira may be called as genus specific antigen for diagnosis.
Antigens, Bacterial/*immunology ; Argentina ; Bulgaria ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Leptospira/isolation & purification/metabolism ; Leptospira interrogans/isolation & purification/metabolism ; Leptospirosis/*diagnosis/microbiology ; Male ; Polysaccharides/*immunology ; Reagent Kits, Diagnostic/*standards ; Republic of Korea ; Sensitivity and Specificity

Leptospirosis, a zoonotic disease that is caused by many serovars which are more than 200 in the world, is an emerging worldwide disease. Accurate and rapid diagnostic tests for leptospirosis are a critical step to diagnose the disease. There are some commercial kits available for diagnosis of leptospirosis, but the obscurity of a species- or genus-specific antigen of pathogenic Leptospira interrogans causes the reduced sensitivity and specificity. In this study, the polysaccharide derived from lipopolysaccharide (LPS) of nonpathogenic Leptospira biflexa serovar patoc was prepared, and the antigenicity was confirmed by immunoblot and enzyme linked immunosorbent assay (ELISA). The performance of the rapid diagnostic test (RDT) kit using the polysaccharide as a diagnostic antigen was evaluated in Korea, Bulgaria and Argentina. The sensitivity was 93.9%, 100%, and 81.0% and the specificity was 97.9%, 100%, and 95.4% in Korea (which is a rare region occurring with 2 serovars mostly), Bulgaria (epidemic region with 3 serovars chiefly) and Argentina (endemic region with 19 serovars mainly) respectively. These results indicate that this RDT is applicable for global diagnosis of leptospirosis. This rapid and effective diagnosis will be helpful for diagnosis and manage of leptospirosis to use and the polysaccharide of Leptospira may be called as genus specific antigen for diagnosis.
Antigens, Bacterial/*immunology ; Argentina ; Bulgaria ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Leptospira/isolation & purification/metabolism ; Leptospira interrogans/isolation & purification/metabolism ; Leptospirosis/*diagnosis/microbiology ; Male ; Polysaccharides/*immunology ; Reagent Kits, Diagnostic/*standards ; Republic of Korea ; Sensitivity and Specificity