Dataset Information

ORFeome-based identification of biomarkers for serodiagnosis of Mycobacterium tuberculosis latent infection.

ABSTRACT:

Background

The challenges posed by Mycobacterium tuberculosis infection require the gradual removal of the pool of latent tuberculosis infection (LTBI). The current cell-immune-based diagnostic tests used to identify LTBI individuals have several irreversible drawbacks. In the present study, we attempted to identify novel diagnostic antigens for LTBI.

Methods

A high-throughput glutathione S-transferase (GST)-fusion technology was used to express over 409 TB proteins and sera from LTBI and healthy individuals was used to interrogate these GST-TB fusion proteins.

Results

Of 409 TB proteins, sixty-three reacted seropositive and defined the immuno-ORFeome of latent M. tuberculosis. Within the immuno-ORFeome, the rare targets were predominantly latency-associated proteins and secreted proteins, while the preferentially recognized antigens tended to be transmembrane proteins. Six of novel highly-reactive antigens had the potential to distinguish LTBI from active TB and healthy individuals. A multiple-antigen combination set was selected through analysis of various combinations. A panel of 94 archived serum samples was used to validate the diagnostic performance of the multiple-antigen combination set, which had sensitivity of 66.1% (95% CI 52.9, 77.4) and specificity of 87.5% (95% CI 70.1, 95.1).

Conclusion

These results provide experimental evidence of the immunogenicity of novel TB proteins that are suitable for the development of serodiagnostic tools for LTBI.

SUBMITTER: Zhou F

PROVIDER: S-EPMC5745629 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Publications

ORFeome-based identification of biomarkers for serodiagnosis of Mycobacterium tuberculosis latent infection.

Zhou Fangbin F Xu Xindong X Wu Sijia S Cui Xiaobing X Pan Weiqing W

BMC infectious diseases 20171228 1

<h4>Background</h4>The challenges posed by Mycobacterium tuberculosis infection require the gradual removal of the pool of latent tuberculosis infection (LTBI). The current cell-immune-based diagnostic tests used to identify LTBI individuals have several irreversible drawbacks. In the present study, we attempted to identify novel diagnostic antigens for LTBI.<h4>Methods</h4>A high-throughput glutathione S-transferase (GST)-fusion technology was used to express over 409 TB proteins and sera from ...[more]

PMID: 29281987

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Project description:Increasing evidence demonstrates that antigen-specific cellular and humoral immunity plays an indispensable role in protection against Mycobacterium tuberculosis infection. Antigen is a key element in the development of a successful diagnostic method and vaccine. However, few antigens are available, and a systemic study on M. tuberculosis ORFeome-based antigen screening is still lacking. In the current study, a genome-wide examination was conducted on high-throughput M. tuberculosis encoding proteins and novel antigens were identified via a comprehensive investigation of serological and antigen-specific cellular responses. The serological immunoglobulin G level of each protein was detected in pooled sera from 200 pulmonary tuberculosis patients by means of semi-quantitative Western blot. Of the 1,250 detected proteins, 29 were present at a higher level relative to the commercialized 38-kDa protein. Furthermore, the top 12 of the 29 proteins had not been previously reported, and their antigenicity was validated in serum from each individual patient. Results confirmed that the 12 proteins displayed nearly identical immunoglobulin G antibody levels in patients with pulmonary and extrapulmonary tuberculosis. Antigen-specific cellular interferon-γ secretion was also evaluated using a cell-based ELISPOT assay. Thirty-four of the proteins were able to induce positive interferon-γ production by peripheral blood mononuclear cells from pulmonary tuberculosis patients as judged by positive (commercial ESAT-6 antigen) and negative controls. The top 4 candidates out of the 34 proteins displayed good accuracy ranging from 50% to 80% compared with the commercial ESAT-6 antigen. Subsequent epitope examination confirmed that a pool of peptides, including a 25aa peptide from Rv1198, demonstrated significant tuberculosis-specific cellular interferon-γ production. Overall, the current study draws significant attention to novel M. tuberculosis antigens, many of which have not been previously reported. This discovery provides a large amount of useful information for the diagnosis of tuberculosis and the development of vaccines to provide protection against tuberculosis.

Project description:BackgroundLatent tuberculosis infection (LTBI) relies on a homeostasis of macrophages and Mycobacterium tuberculosis (Mtb). The small heat shock protein, Mtb Hsp16.3 (also known as latency-associated antigen), plays an important role in Mtb persistence within macrophages. However, the mechanism of LTBI remains elusive. The aim of this study was to delineate LTBI-related miRNA expression in U937 macrophages expressing Mtb Hsp16.3 protein. U937 macrophages were infected with an integrase-deficient Lentivirus vector to transiently express Mtb Hsp16.3, and green fluorescent protein (GFP) as a control. We used a microRNA (miRNA) microarray chip containing more than 1000 probes to identify the significant differentially expressed miRNAs in the infected U937 cells, and employed real-time quantitative polymerase chain reaction (qRT-PCR) for validation. Furthermore, we confirmed these candidate LTBI-related miRNAs in peripheral blood mononuclear cells from subjects with LTBI and in healthy control individuals. Functional annotation prediction of miRNA target genes and pathway enrichment analyses were used to explore the putative links between these miRNAs and LTBI.ResultsAnalysis of the miRNA expression profile identified 149 miRNAs that were differentially expressed in U937 macrophages expressing Mtb Hsp16.3 compared with the control expressing GFP. The expression level of seven miRNAs (miR-424-5p, miR-493-5p, miR-296-5p, miR-27b-3p, miR-377-5p, miR-3680-5p, miR-191-5p) were validated by qRT-PCR. The expression level of four miRNAs (miR-424-5p, miR-27b-3p, miR-377-5p, miR-3680-5p) in the peripheral blood mononuclear cells samples from LTBI and healthy participants reflected the altered patterns observed in the microarray profile. The bioinformatic analyses suggest that the miRNAs may regulate Mtb latent infection by affecting the development of macrophage cells.ConclusionsThe results suggest that miRNA expression may play a considerable role in the pathogenesis of LTBI, and this would increase our understanding of the molecular basis of Hsp16.3-facilitated Mtb survival in macrophages.

Dataset Information

ORFeome-based identification of biomarkers for serodiagnosis of Mycobacterium tuberculosis latent infection.

Background

Methods

Results

Conclusion

Publications

ORFeome-based identification of biomarkers for serodiagnosis of Mycobacterium tuberculosis latent infection.

Similar Datasets

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