Mycobacterium tuberculosis 6-kDa early secretory antigenic target （ESAT-6） is a dominant target antigen for cell-mediated immunity in the early phase of tuberculosis. The fms-like tyrosine kinase 3 ligand （FL） tha...Mycobacterium tuberculosis 6-kDa early secretory antigenic target （ESAT-6） is a dominant target antigen for cell-mediated immunity in the early phase of tuberculosis. The fms-like tyrosine kinase 3 ligand （FL） that induces potent immune response has been used as an adjuvant in vaccine development. In this study, a new recombinant plasmid （plRES-epitope-peptides-FL） encoding three T cell epitopes of ESAT-6 and FL was constructed, and the immunogenicity of the DNA vaccine was assessed in C57BL/6 mice immunized with the plasmid DNA vaccine. Additionally, a strategy of intramuscular injection with the DNA vaccine （prime） and intranasal administration of the epitope peptides （boost） was employed to induce higher immune reaction of the mice. The results showed that mice vaccinated with the recombinant plasmid DNA vaccine and boosted with the peptides not only increased the levels of Thl cytokines （IFN-γ and IL-12）, the number of IFN-γ＋ T cells and activities of cytotoxic T lymphocytes as well as IgG, but also enhanced protection against Mycobacterium tuberculosis challenge. In conclusion, these data indicate that the novel recombinant plRES-epitope-peptides-FL plasmid is a useful DNA vaccine for pre- venting Mycobacterium tuberculosis infection.展开更多
Background Prompt diagnosis of Mycobacterium tuberculosis （MTB） infection is an essential step in tuberculosis control and elimination. However, it is often difficult to accurately diagnose pediatric tuberculosis ...Background Prompt diagnosis of Mycobacterium tuberculosis （MTB） infection is an essential step in tuberculosis control and elimination. However, it is often difficult to accurately diagnose pediatric tuberculosis （TB）. The tuberculin test （TST） may have a low specificity because of cross-reactivity with antigens present in Mycobacterium bovis bacillus Calmette-Guerin （BCG） and other mycobacteria, especially in China with a predominantly BCG-vaccinated population.Early-secreted antigenic target 6-kDa protein （ESAT-6） and culture filtrate protein 10 （CFP-10）, stand out as suitable antigens that induce an interferon-gamma （IFN-γ） secreting, T-cell-mediated immune response to infection. While,considered the higher costs and complexity of the IFN-γ release assay （TSPOT）, we aimed to evaluate the TSPOT and TST test in the clinical diagnosis of pediatric tuberculosis and to establish a diagnostic process suitable for China.Methods The sensitivity and specificity of the assay were evaluated in total seventy four children with active tuberculosis and fifty one nontuberculous children with other disease, and then the results were compared with TST.Logistic regression models were used to identify variables that were associated with positive results for each assay. The independent variables included sex, age, birth place, vaccination history, close contract with an active TB patient.Results The sensitivity of TSPOT was higher than TST in active TB children with or without BCG vaccination, as well as in children with culture-confirmed TB. But the difference was not significant statistically. Combining results of the TSPOT and TST improved the sensitivity to 94.6%. Agreement of the TST and TSPOT was low （77.0%, k=0.203） in active TB patients. The difference in specificity between TSPOT and TST test was statistically significant （94.1% vs.70.6%, P=0.006）. Specificity of the two tests in patients without prior BCG vaccination history was similar （80.0% vs.60.0%）. The concordance between the two tests results in BCG vaccinated subjects was low （71.7%, k=0.063）. For TSPOT, none of the included risk factors was significantly associated with positive results. For TST, BCG vaccination （OR：1.78; 95% CI： 1.30-2.00） was significantly associated with positive results.Conclusions Although IFN-γ release assay had relatively high sensitivity and specificity, we also should consider the higher costs and complexity of this test. Therefore, TSPOT could be used as the complementary tool of TST in circumstances when a suspected patient with negative TST results, or to exclude a positive TST result caused by BCG vaccination.展开更多
基金supported by the Natural Scientific Fund (09KJA310002DG216D50162011NJMU263 and 11JC005) of China
文摘Mycobacterium tuberculosis 6-kDa early secretory antigenic target （ESAT-6） is a dominant target antigen for cell-mediated immunity in the early phase of tuberculosis. The fms-like tyrosine kinase 3 ligand （FL） that induces potent immune response has been used as an adjuvant in vaccine development. In this study, a new recombinant plasmid （plRES-epitope-peptides-FL） encoding three T cell epitopes of ESAT-6 and FL was constructed, and the immunogenicity of the DNA vaccine was assessed in C57BL/6 mice immunized with the plasmid DNA vaccine. Additionally, a strategy of intramuscular injection with the DNA vaccine （prime） and intranasal administration of the epitope peptides （boost） was employed to induce higher immune reaction of the mice. The results showed that mice vaccinated with the recombinant plasmid DNA vaccine and boosted with the peptides not only increased the levels of Thl cytokines （IFN-γ and IL-12）, the number of IFN-γ＋ T cells and activities of cytotoxic T lymphocytes as well as IgG, but also enhanced protection against Mycobacterium tuberculosis challenge. In conclusion, these data indicate that the novel recombinant plRES-epitope-peptides-FL plasmid is a useful DNA vaccine for pre- venting Mycobacterium tuberculosis infection.
基金This study was supported by the grants from the National Natural Science Foundation of China （No. 30872788） and from the Beijing Municipal Science and Technology Commission （No.Z09050700940903）.
文摘Background Prompt diagnosis of Mycobacterium tuberculosis （MTB） infection is an essential step in tuberculosis control and elimination. However, it is often difficult to accurately diagnose pediatric tuberculosis （TB）. The tuberculin test （TST） may have a low specificity because of cross-reactivity with antigens present in Mycobacterium bovis bacillus Calmette-Guerin （BCG） and other mycobacteria, especially in China with a predominantly BCG-vaccinated population.Early-secreted antigenic target 6-kDa protein （ESAT-6） and culture filtrate protein 10 （CFP-10）, stand out as suitable antigens that induce an interferon-gamma （IFN-γ） secreting, T-cell-mediated immune response to infection. While,considered the higher costs and complexity of the IFN-γ release assay （TSPOT）, we aimed to evaluate the TSPOT and TST test in the clinical diagnosis of pediatric tuberculosis and to establish a diagnostic process suitable for China.Methods The sensitivity and specificity of the assay were evaluated in total seventy four children with active tuberculosis and fifty one nontuberculous children with other disease, and then the results were compared with TST.Logistic regression models were used to identify variables that were associated with positive results for each assay. The independent variables included sex, age, birth place, vaccination history, close contract with an active TB patient.Results The sensitivity of TSPOT was higher than TST in active TB children with or without BCG vaccination, as well as in children with culture-confirmed TB. But the difference was not significant statistically. Combining results of the TSPOT and TST improved the sensitivity to 94.6%. Agreement of the TST and TSPOT was low （77.0%, k=0.203） in active TB patients. The difference in specificity between TSPOT and TST test was statistically significant （94.1% vs.70.6%, P=0.006）. Specificity of the two tests in patients without prior BCG vaccination history was similar （80.0% vs.60.0%）. The concordance between the two tests results in BCG vaccinated subjects was low （71.7%, k=0.063）. For TSPOT, none of the included risk factors was significantly associated with positive results. For TST, BCG vaccination （OR：1.78; 95% CI： 1.30-2.00） was significantly associated with positive results.Conclusions Although IFN-γ release assay had relatively high sensitivity and specificity, we also should consider the higher costs and complexity of this test. Therefore, TSPOT could be used as the complementary tool of TST in circumstances when a suspected patient with negative TST results, or to exclude a positive TST result caused by BCG vaccination.
