Evaluation of host biomarkers for early diagnosis of tuberculosis disease in children

Manyelo, Masilo Charles (2018-12)

Thesis (MSc)--Stellenbosch University, 2018.

Thesis

Background: The diagnosis of tuberculosis (TB) remains a challenge in children. There is an urgent need for new tools for early diagnosis of TB disease in children Objectives: To evaluate the usefulness of a previously described 3-marker cerebrospinal fluid (CSF) biosignature (VEGF, IL-13 and cathelicidin LL-37) and other CSF biomarkers for diagnosis of tuberculous meningitis (TBM), and evaluate the utility of a previously identified adult 7-marker serum protein biosignature (CRP, IFN-γ, IP-10, CFH, Apo-AI, SAA and transthyretin) and other blood biomarkers for diagnosis of pulmonary TB (PTB) and TBM in children. Methods: CSF and serum samples were collected from children with suspected meningitis, whereas serum samples were collected from children with suspected PTB for investigation of biomarkers for the diagnosis of childhood TBM and PTB, respectively. Children in the TBM project were enrolled at the Tygerberg Academic Hospital, whereas those in the PTB study were enrolled at the Red Cross War Memorial Children’s Hospital in Cape Town, South Africa. Children were classified as TBM or no-TBM and PTB or no-PTB, using combination of clinical, radiological and laboratory findings. Using a multiplex platform, the concentrations of 69 host biomarkers were evaluated in CSF and serum samples from children in the TBM study whereas 40 host markers were evaluated in serum samples from children in the PTB study. The diagnostic accuracies of individual biomarkers were assessed by receiver operator characteristics (ROC) curve, whereas the General Discriminant Analysis (GDA) was used to assess the accuracies of combinations between different host biomarkers. Results: Of the 69 host biomarkers evaluated in CSF and serum samples from children in the TBM study, multiple individual host biomarkers showed potential as diagnostic candidates for TBM as ascertained by area under the ROC curve (AUC). The previously described 3-marker CSF biosignature was validated in the project. However, refinement of the biosignature by substitution of IL-13 and cathelicidin LL-37 with two new proteins (MPO and IFN-γ) resulted in a new biosignature with improved accuracy (AUC of 0.97). Furthermore, we identified a 4- marker CSF biosignature (sICAM-1, MPO, CXCL8 and IFN-γ), which also diagnosed TBM with AUC of 0.97. The adult 7-marker serum biosignature, modified by the replacement of transthyretin with NCAM1, diagnosed TBM with AUC of 0.80. However, a childhood TBM-specific serum biosignature (adipsin, Aβ42 and IL-10) diagnosed TBM with AUC of 0.84. The adult signature performed with an AUC of 0.79 in children with PTB, showing no significant difference in the diagnosis of childhood PTB or TBM. However, novel childhood PTB-specific biosignatures performed better than the adult 7-marker signature. Conclusion: The adult 7-marker signature showed potential in the diagnosis of both PTB and TBM in children recruited from a high TB incidence area. We validated a previously established 3- marker CSF biosignature, but a refined signature showed much improved accuracy. The biosignatures identified in this thesis hold potential for development of new diagnostic tools for PTB and TBM in children for possible use at the point-of-care. Our findings require further validation in larger and multi-site studies.

Agtergrond: Die diagnose van tuberkulose (TB) by kinders bly ‘n uitdaginig. Daar is ‘n dringende behoefte aan nuwe toestelle vir die vroeë diagnose van TB-siekte by kinders. Doelwitte: Om die doeltreffendheid van ‘n voorheen beskryfde 3-merker serebrospinale vloeistof (CSF) bioprofiel (VEGF, IL-13 and cathelicidin LL-37) en ander CSF biomerkers vir die diagnose van tuberkulose meningitis (TBM) te evalueer, asook die bruikbaarheid van ‘n voorheen geïndetifiseerde volwasse 7-merker serumproteïn bioprofiel (CRP, IFN-γ, IP-10, CFH, ApoAI, SAA en transthyretin) en ander bloedbiomerkers te evalueer vir die diagnose van pulmonale TB (PTB) en TBM by kinders. Metodes: CSF- en serummonsters is verkry van kinders wat vermoedelik meningitis het, terwyl serummonsters van kinders met vermeende PTB verkry is om die biomerkers se vermoëns om kindertyd TBM and PTB, onderskeidelik, te diagnoseer te ondersoek. Die kinders in die TBM-projek is ingeskryf by die Tygerberg Akademiese Hospitaal, terwyl dié wat aan die PTB studie deelgeneem het by die Rooikruis Oorlogsgedenkhospitaal in Kaapstad ingeskryf is. Die kinders is geklassifiseer as TBM of nie-TBM, en PTB of nie-PTB deur gebruik te maak van ‘n kombinasie van kliniese-, radiologiese-, en laboratoriumbevindings. ‘n Multipleks-platform is benut om die konsentrasies van 69 gasheerbiomerkers in CSF- en serummonsters van kinders in die TBM-studie en 40 gasheerbiomerkers in serummonsters van die kinders in die PTB-studie, onderskeidelik, te evalueer. Die diagnostiese akkuraatheid van individuele biomerkers is met die ontvanger-operateur-eienskappe (ROC) kurwe geassesseer, terwyl die Algemene Diskriminant Analise (GDA) gebruik is om die akkuraatheid van kombinasies tussen verskillende gasheerbiomerkers te bepaal. Resultate: Van die 69 gasheerbiomerkers wat in die CSF- en serummonsters van kinders in die TBMstudie geëvalueer is, het verskeie individuele gasheerbiomerkers potensiaal getoon as diagnostiese kandidate vir TBM, soos vasgestel deur die area onder die ROC-kurwe (AUC). Validasie van die voorheen beskryfde 3-merker CSF bioprofiel is in die projek uitgevoer. Verfyning van die bioprofiel deur die vervanging van IL-13 en cathekicidin LL-37 met twee nuwe proteïene (MPO en IFN-γ), het gelei tot ‘n nuwe bioprofiel met verbeterde akkuraatheid (AUC van 0.97). Daarbenewens, is daar ‘n 4-merker CSF-bioprofiel geïdentifiseer (sICAM-1, MPO, CXCL8 en IFN-γ) wat ook TBM met ‘n AUC van 0.97 gediagnoseer het. Die volwasse 7-merker serum bioprofiel, gewysig deur die vervanging van transthyretin met NCAM1, het TBM met ‘n AUC van 0.80 gediagnoseer. ‘n TBM-spesifieke serum bioprofiel (adipsin, Aβ42 en IL-10) het egter TBM met ‘n AUC van 0.84 gediagnoseer. Die volwasse teken het ‘n AUC van 0.79 in kinders met PTB opgelewer en dus geen beduidende verskil tussen die diagnose van kindertyd PTB of TBM getoon nie. Daarteenoor, het nuwe kindertyd PTB-spesifieke bioprofiele beter gevaar as die volwasse 7-merker profiel. Afsluiting: Die volwasse 7-merker profiel het potensiaal getoon om beide PTB en TBM by kinders afkomkstig van hoë TB-voorkomsgebiede te diagnoseer. ‘n Voorheen beskryfde 3-merkerCSF-bioprofiel is bevestig, maar ‘n verfynde profiel het heelwat verbeterde akkuraatheid getoon. Die bioprofiel wat in hierdie proefskrif geïdentifiseer is, het die potensiaal om gebruik te word om nuwe diagnostiese intrumente te ontwikkel vir punt-van-sorg gebruik by PTB en TBM in kinders. Die bevindings verg egter verdere validasie in groter en multi-setel studies.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/105172
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