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The cyanobacterial lectin, microvirin-N, enhances the specificity and sensitivity of lipoarabinomannan-based TB diagnostic tests.

ABSTRACT: Tuberculosis (TB) is one of the top ten causes of death globally, despite being treatable. The eradication of TB disease requires, amongst others, diagnostic tests with high specificity and sensitivity that will work at the point of care (POC) in low-resource settings. The TB surface glycolipid antigen, mannose-capped lipoarabinomannan (ManLAM) currently serves as the only POC molecular diagnostic biomarker suitable for use in low cost immunoassays. Here, we demonstrate the high affinity and exceptional specificity of microvirin-N (MVN), a 14.3 kDa cyanobacterial lectin, toward H37Rv TB ManLAM and utilize it to develop a novel on-bead ELISA. MVN binds to ManLAM with sub-picomolar binding affinity, but does not bind to other variants of LAM expressed by non-pathogenic mycobacteria - a level of binding specificity and affinity that current commercially available anti-LAM antibodies cannot achieve. An on-bead ELISA was subsequently developed using MVN-functionalized magnetic beads which allows for the specific capture of ManLAM from human urine with a limit of detection (LOD) of 1.14 ng mL^-1 and no cross-reactivity when tested with PILAM, a variant of LAM found on non-pathogenic mycobacteria.

SUBMITTER: van der Horst M

PROVIDER: S-EPMC8374243 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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The cyanobacterial lectin, microvirin-N, enhances the specificity and sensitivity of lipoarabinomannan-based TB diagnostic tests.

van der Horst Megan M Karamchand Leshern L Bauer Westley S WS Nel Andrew J M AJM Blackburn Jonathan M JM Wright David W DW

The Analyst 20201224 4

Tuberculosis (TB) is one of the top ten causes of death globally, despite being treatable. The eradication of TB disease requires, amongst others, diagnostic tests with high specificity and sensitivity that will work at the point of care (POC) in low-resource settings. The TB surface glycolipid antigen, mannose-capped lipoarabinomannan (ManLAM) currently serves as the only POC molecular diagnostic biomarker suitable for use in low cost immunoassays. Here, we demonstrate the high affinity and exc ...[more]

PMID: 33367346

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Project description:BackgroundLassa fever virus has been enlisted as a priority pathogen of epidemic potential by the World Health organization Research and Development (WHO R & D) Blueprint. Diagnostics play a crucial role in epidemic preparedness. This systematic review was conducted to determine the sensitivity and specificity of Lassa fever diagnostic tests for humans.MethodsWe searched OVID Medline, OVID Embase, Scopus and Web of Science for laboratory based and field studies that reported the performance of diagnostic tests for Lassa fever in humans from 1 January 1990 to 25 January 2019. Two reviewers independently screened all the studies and included only studies that involved the evaluation of a Lassa fever diagnostic test and provided data on the sensitivity and specificity. The quality of the studies was assessed using the QUADAS-2 criteria. Data on the study location, study design, type of sample, index test, reference tests and diagnostic performance were extracted from the studies.ResultsOut of a total of 1947 records identified, 1245 non-duplicate citations were obtained. Twenty-six (26) full-text articles examined which identified 08 studies meeting pre-defined criteria. Only one study was a field evaluation study. The sensitivity and specificity of the point of care (RDT) against the Nikisins qPCR were 91.2%(95% CI:75.2-97.7) and 86%(95% CI: 71.4-94.2) at temperatures 18-30 °C, while the sensitivity and specificity of the single IgM ELISA assay against standard RT-PCR were 31.1%(95%CI: 25.6-37) and 95.7%(95%CI:92.8-97.7). The sensitivity of the combined ELISA Antigen/IgM assay(against virus isolation), the recombinant IgM/IgG ELISA(against standard RT-PCR), and the IgM/IgG immunoblot(against IFA) were 88%(95%CI:77-95), 25.9%(95%CI:20.8-31.6), and 90.7%(95%CI:84.13-97.27) respectively. The specificity of the combined ELISA Antigen/IgM assay(against virus isolation), the recombinant IgM/IgG ELISA(against standard RT-PCR), and the IgM/IgG immunoblot(against IFA) were 90%(95%CI:88-91), 100%(95%CI:98.2-100), and 96.3%(95%CI:92.2-100) respectively.ConclusionLassa fever has assays for antigenaemia, IgM, IgG and PCR detection. The RDT reportedly performed well but more data are needed from other countries and at temperatures above 30 °C. Most combined immunoassays perform better than the single IgM. Multiplex and pan-Lassa assays are needed. More well conducted field studies are needed.Trial registrationProspero registration number: CRD42018091585 .

Project description:Lipoarabinomannan (LAM), a component of the Mycobacterium tuberculosis (MTB) cell wall, is detectable in the urine of MTB infected patients with active tuberculosis (TB). LAM-specific antibodies (Igs) have been developed by a variety of traditional and recombinant methods for potential use in a rapid diagnostic test (RDT). We evaluated the analytical performance of the TB LAM Igs to identify pairs that offer superior performance over existing urine LAM tests. We assessed 25 new and 4 existing Igs in a matrixed format using a multiplex electrochemiluminescence-based liquid immunoassay. A total of 841 paired Ig combinations were challenged with in vitro cultured LAM (cLAM) derived from MTB strains representing diverse phylogenetic lineages, alongside urinary LAM (uLAM) from the urine of adults with active pulmonary TB. Analytical sensitivity of down-selected Ig pairs was determined using MTB Aoyama-B cLAM, while diagnostic accuracy was determined using clinical samples. When testing cLAM, the reactivity of Ig pairs was similar across MTB lineages 1-4 but lineage 5:6 had significantly more reactivity among Ig pairs. Overall, 41 Ig pairs had a strong binding affinity to cLAM, as compared to the reference pair of S4-20/A194-01, and 28 Ig pairs therein exhibited a strong affinity for both cLAM and uLAM. Retrospective testing on clinical urine specimens demonstrated varying sensitivities (12-80%) and specificities (14-100%). The five top pairs had a similar analytical limit of detection to the reference pair but in four instances, the sensitivity and specificity with clinical uLAM samples was poor. Overall, epitopes presented by uLAM are different from cLAM, which may affect antibody performance when testing uLAM in patient samples. Several new Ig pairs had similar ranges of high sensitivity to cLAM but overall, there were no new candidate Ig pairs identified in this round of screening with increased performance with uLAM as compared to an existing optimal pair.

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Dataset Information

The cyanobacterial lectin, microvirin-N, enhances the specificity and sensitivity of lipoarabinomannan-based TB diagnostic tests.

Publications

The cyanobacterial lectin, microvirin-N, enhances the specificity and sensitivity of lipoarabinomannan-based TB diagnostic tests.

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