Use of the cyanobacterial lectin, microvirin-N, for specific detection of the tuberculosis biomarker mannose-capped lipoarabinomannan

Abstract

Novel methods for TB diagnosis are urgently needed to control the transmission and unacceptably high number of deaths due to this otherwise curable disease. The goal of this project is to utilize the lectin microvirin (MVN) as a novel molecular recognition element for mannose-capped lipoarabinomannan (ManLAM). It has previously been established that MVN binds to Man-alpha-(1,2)-Man linkages. These linkages are present in the mannose caps of ManLAM providing the rationale for this work. First, the binding of MVN to ManLAM and other variants of LAM was investigated using bio-layer interferometry (BLI). The results showed that MVN is specific for ManLAM versus variants of LAM capped with non-mannose motifs. In contrast, anti-LAM antibodies were not able to distinguish between the variants of LAM. Then, a MVN-based on-bead enzyme-linked immunosorbent assay (OB-ELISA) was developed that allows in vitro ManLAM to be detected in urine with a limit of detection of 1.1 ng/mL. Importantly, no signal was observed when the OB-ELISA was tested with PILAM, confirming the BLI results. Lastly, clinical urine samples were tested with the MVN-based OB-ELISA to show that MVN is capable of binding to mannose caps on in vivo ManLAM. Specifically, a significant difference in the OB-ELISA signal of TB-positive versus TB-negative samples was identified in a subset of samples from Peru, opening up avenues for future exploration of geographic differences in ManLAM structure and/or concentration. As a result, this work represents a critical advance in TB diagnostic technology.