Browsing by Author "Wiker, Harald G."
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- ItemEffect of non-tuberculous mycobacteria on host biomarkers potentially relevant for tuberculosis management(PLoS, 2014-10-16) Dhanasekaran, S.; Jenum, Synne; Stavrum, Ruth; Wiker, Harald G.; Kenneth, John; Vaz, Mario; Doherty, T. Mark; Grewal, Harleen M. S.; Hesseling, A. C.Background: Non-tuberculous mycobacteria (NTM) are different from Mycobacterium tuberculosis (MTB) both in their ubiquitous environmental distribution and in their reduced capacity to cause disease. While often neglected in favour of other infectious diseases, NTM may interfere with important aspects of TB control and management, namely the efficacy of new anti-tuberculosis (TB) vaccines; the immuno-diagnostic Tuberculin skin test (TST) and QuantiFERON TB Gold In Tube assay (QFTGIT); and immune biomarkers explored for their diagnostic and/or predictive potential. Our objective was therefore to explore host immune biomarkers in children who had NTM isolated from respiratory and/or gastric specimens. Methodology and Principle Findings: The present study was nested within a prospective cohort study of BCG-vaccinated neonates in Southern India. In this setting, immune biomarkers from peripheral blood were analyzed in 210 children aged , 3 years evaluated for TB using dual-colour-Reverse-Transcriptase-Multiple-Ligation-dependent-Probe-Amplification (dcRTMLPA) and Bio-Plex assays. The children were classified based on clinical examination, chest X-rays and mycobacterial culture reports as either: 1) TB disease, 2) NTM present and 3) controls. The study shows a down-regulation of RAB33A (p, 0.001) and up-regulation of TGFb1, IL-2 and IL-6 (all p,0.05) in children with TB disease, and that RAB33A, TGFBR2 and IL-10 (all p,0.05) were differentially expressed in children with NTM present when compared to children that were culture negative for MTB and NTM (controls). Conclusions and Significance: Carriage of NTM may reduce the specificity of future diagnostic and predictive immune biomarkers relevant to TB management.
- ItemOn the impact of the pangenome and annotation discrepancies while building protein sequence databases for bacteria proteogenomics(Frontiers Media, 2019) Machado, Karla C. T.; Fortuin, Suereta; Tomazella, Gisele Guicardi; Fonseca, Andre F.; Warren, Robin Mark; Wiker, Harald G.; De Souza, Sandro Jose; De Souza, Gustavo AntonioENGLISH ABSTRACT: In proteomics, peptide information within mass spectrometry (MS) data from a specific organism sample is routinely matched against a protein sequence database that best represent such organism. However, if the species/strain in the sample is unknown or genetically poorly characterized, it becomes challenging to determine a database which can represent such sample. Building customized protein sequence databases merging multiple strains for a given species has become a strategy to overcome such restrictions. However, as more genetic information is publicly available and interesting genetic features such as the existence of pan- and core genes within a species are revealed, we questioned how efficient such merging strategies are to report relevant information. To test this assumption, we constructed databases containing conserved and unique sequences for 10 different species. Features that are relevant for probabilistic-based protein identification by proteomics were then monitored. As expected, increase in database complexity correlates with pangenomic complexity. However, Mycobacterium tuberculosis and Bordetella pertussis generated very complex databases even having low pangenomic complexity. We further tested database performance by using MS data from eight clinical strains from M. tuberculosis, and from two published datasets from Staphylococcus aureus. We show that by using an approach where database size is controlled by removing repeated identical tryptic sequences across strains/species, computational time can be reduced drastically as database complexity increases.
- ItemPhosphoproteomics analysis of a clinical mycobacterium tuberculosis Beijing isolate : expanding the mycobacterial phosphoproteome catalog(Frontiers Media, 2015) Fortuin, Suereta; Tomazella, Gisele G.; Nagaraj, Nagarjuna; Sampson, Samantha L.; Gey Van Pittius, Nicolaas C.; Soares, Nelson C.; Wiker, Harald G.; De Souza, Gustavo A.; Warren, Robin M.Reversible protein phosphorylation, regulated by protein kinases and phosphatases, mediates a switch between protein activity and cellular pathways that contribute to a large number of cellular processes. The Mycobacterium tuberculosis genome encodes 11 Serine/Threonine kinases (STPKs) which show close homology to eukaryotic kinases. This study aimed to elucidate the phosphoproteomic landscape of a clinical isolate of M. tuberculosis. We performed a high throughput mass spectrometric analysis of proteins extracted from an early-logarithmic phase culture. Whole cell lysate proteins were processed using the filter-aided sample preparation method, followed by phosphopeptide enrichment of tryptic peptides by strong cation exchange (SCX) and Titanium dioxide (TiO2) chromatography. The MaxQuant quantitative proteomics software package was used for protein identification. Our analysis identified 414 serine/threonine/tyrosine phosphorylated sites, with a distribution of S/T/Y sites; 38% on serine, 59% on threonine and 3% on tyrosine; present on 303 unique peptides mapping to 214 M. tuberculosis proteins. Only 45 of the S/T/Y phosphorylated proteins identified in our study had been previously described in the laboratory strain H37Rv, confirming previous reports. The remaining 169 phosphorylated proteins were newly identified in this clinical M. tuberculosis Beijing strain. We identified 5 novel tyrosine phosphorylated proteins. These findings not only expand upon our current understanding of the protein phosphorylation network in clinical M. tuberculosis but the data set also further extends and complements previous knowledge regarding phosphorylated peptides and phosphorylation sites in M. tuberculosis.