Peptide microarray-based identification of dormancy-associated Mycobacterium tuberculosis antigens inducing immune responses among latent tuberculosis infection individuals in Thailand.

Multi-stage tuberculosis (TB) vaccines composed of active- and dormancy-associated antigens are promising to trigger the immune protection against all TB stages. However, scientists are still in quest of the suitable vaccine candidates. In this study, we identified the potential targets for this vaccine in a high TB burden country, Thailand. Peptide microarray was applied to gauge IgA and IgG antibodies specific to 16,730 linear epitopes of 52 dormancy-associated Mycobacterium tuberculosis (M. tb) proteins in three study groups: active tuberculosis (ATB), latent tuberculosis infection (LTBI) and endemic healthy control (EHC). Preferential IgA recognition against epitopes of dormancy-associated proteins was identified in LTBI group. Validation of these findings revealed that LTBI subjects exhibited the greater levels of Rv2659c- and Rv1738-specific IgA than those of household contacts, but less than did ATB subjects. Frequencies of IFNγ-producing CD4+ and CD8+ T cells induced by proteins Rv2659c and Rv1738 were higher in LTBI than ATB individuals. The results indicated that LTBI group in a high TB burden country demonstrated cell-mediated immune response to proteins Rv2659c and Rv1738 stronger than those of ATB. These immune responses likely contribute to natural protection against dormant M. tb and might be potential targets for a multi-stage TB vaccine.

Toll-like receptor-mediated innate immune responses by recognition of the recombinant dormancy-associated Mycobacterium tuberculosis proteins Rv2659c and Rv1738.

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) poses a major threat to the global public health. Importantly, latent tuberculosis infection (LTBI) still impedes the elimination of TB incidence since it has a substantial risk to develop active disease. A multi-stage subunit vaccine comprising active and latency antigens of Mtb has been raised as the promising vaccine to trigger immune protection against all stages of TB. Therefore, the discovery of new antigens that could trigger broad immune response is essential. While current development of TB vaccine mainly focuses on protective immunity mediated by adaptive immune response, the knowledge on triggering the innate immune response by antigens is still limited. We showed that recombinant dormancy-associated Mtb proteins Rv2659c and Rv1738 were recognized by human innate immune recognition molecules, Toll-like receptors (TLRs) 2 and 4 by using HEK-Blue™ hTLR2/hTLR4 systems. We further demonstrated that these two proteins activated phosphorylated NF-κB p65 (Ser536) in the human CD14+ blood cells. We also investigated that these two proteins significantly induced level of pro- and anti-inflammatory cytokines (IL-1ß, IL-6, IL-8, IL-10 and TNF-α) which were mediated through TLR2 and TLR4 pathways in human peripheral blood mononuclear cells (hPBMCs). These findings suggest that proteins Rv2659c and Rv1738 stimulated innate immune response targeting TLR2 and TLR4 to produce inflammatory cytokines, and their benefits would be valuable for the development of an effective prophylactic tuberculosis vaccine.

Circulating IgA/IgG memory B cells against Mycobacterium tuberculosis dormancy-associated antigens Rv2659c and Rv3128c in active and latent tuberculosis.

OBJECTIVE: To elucidate the antigenic potential of dormancy-associated antigens Rv2659c and Rv3128c of Mycobacterium tuberculosis by examining the persistence of specific IgG and IgA memory B cells (MBCs) among patients with active tuberculosis (TB), household contacts with latent tuberculosis (LTBI), and an endemic healthy control group. METHODS: Fresh peripheral blood mononuclear cells from the three study groups were used to enumerate the numbers of IgG and IgA MBCs specific to recombinant protein Rv2659c and Rv3128c by ELISpot assay. The composition of MBC subsets IgA+ and IgG + was analyzed by flow cytometry. RESULTS: The number of IgA MBCs specific to antigen Rv2659c was significantly higher in the LTBI group than the TB group. In contrast, no significant difference was found in IgA or IgG MBCs against antigen Rv3128c. The number of IgA+ MBCs was significantly higher than that of IgG+ MBCs in the classical MBC subset of the LTBI group. CONCLUSION: The results indicated that the dormancy-associated antigen Rv2659c induced an IgA MBCs response in individuals with latent TB, and IgA+ classical MBCs formed a major portion of the MBCs subset. This new knowledge will be beneficial for the development of novel TB vaccines and their control of latent TB.

Immunogenicity of the Plasmodium vivax merozoite surface protein 1 paralog in the induction of naturally acquired antibody and memory B cell responses.

BACKGROUND: The Plasmodium vivax merozoite surface protein 1 paralog (PvMSP1P-19) is a glycosylphosphatidylinositol (GPI)-anchored blood-stage protein that is expressed on the merozoite surface. It is proposed as a blood-stage vaccine candidate against P. vivax because of its ability to induce immune responses upon natural P. vivax exposure and in immunized animals. This study aimed to demonstrate the presence of inhibitory antibodies and memory B cell responses to the PvMSP1P-19 antigen during acute P. vivax infection and after recovery from infection. METHODS: To evaluate the antibody responses to PvMSP1P-19 during and after recovery from P. vivax infection, heparinized blood was collected from P. vivax-infected patients and recovered subjects to detect the total IgG response. The seropositive samples were defined into high and low responders, according to their optical density (OD) values obtained from ELISA. High responders were the subjects who had OD values above the OD of antisera from non-exposed controls plus 4× standard deviations, whereas low responders were the subjects who had OD values less than OD of antisera from non-exposed controls plus 4× standard deviations. The plasma from high and low responders were taken for testing the inhibitory activity against PvMSP1P-19-erythrocyte binding by in vitro EBIA. The sustainability of PvMSP1P-19-specific memory B cell responses after recovery from infection was analysed by ELISPOT. RESULTS: The anti-PvMSP1P-19 antibody levels were significantly higher in acutely infected P. vivax patients compared to healthy controls (P <  0.0001). Monitoring of the anti-PvMSP1P-19 antibody titre showed that the antibody was maintained for up to 9 months after recovery. Almost all high-responder groups strongly inhibited PvMSP1P-19 binding to erythrocytes, whereas no inhibition was shown in most low-responder samples. Interestingly, the inhibitory activity of the antibodies in some individuals from high-responder samples were stable for at least 12 months. The longevity of the antibody response was associated with the presence of PvMSP1P-19-specific memory B cells at 9 months after recovery from infection. CONCLUSIONS: The PvMSP1P-19 antigen has immunogenicity during the induction of the antibody response, in which both the levels and inhibitory activity are maintained after the patient recovered from P. vivax infection. The maintenance of the antibody response was associated with the response of PvMSP1P-19-specific memory B cells. Therefore, the PvMSP1P-19 antigen should also be considered as a reliable vaccine candidate to develop a blood-stage vaccine against P. vivax.