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| Image created by Dr. Michael J. Miller |
Researchers have used laboratory-on-a-chip technology to create a low cost, sensitive tuberculosis (TB) test that could improve disease detection in high-endemic, under-resourced areas.
Writing in The Journal of Molecular Diagnostics, collaborators at St George’s University of London and QuantuMDx describe the limitations of existing TB tests. Smear microscopy, in which sputum samples are smeared onto microscope slides for examination, is cheap and requires minimal facilities but suffers from poor sensitivity, and the quality of the results can vary between sites and operators.
Growing the bacteria in sputum samples improves sensitivity; however, TB grows slowly, delaying diagnosis, and biosafety level 3 facilities are required to perform the work. Those factors are limitations in parts of the world where resources are restricted.
The St George’s and QuantuMDx collaborators identified dielectrophoresis as a way to improve testing for TB. Dielectrophoresis enables the selective attraction or repulsion of specific particles or cells based on their properties. Using the technique, the researchers isolated the bacteria that cause TB from the rest of the content of sputum samples.
“This chip-based technology exploits the physiological property of the TB bacteria to be specifically collected onto the device so that small numbers can be visualized on the chip electrodes and act as a visual readout to replace the lab-based sputum smear/microscopic methods, which typically have low detection rates and require training laboratory staff, at molecular-like sensitivities and at a fraction of the price,” QuantuMDx CEO Jonathan O’Halloran, PhD, said in a statement (see below).
Purification of the bacterial population serves two purposes. First, visual analysis of the purified sample may enable the diagnosis of TB infection, serving as a substitute for smear microscopy. Second, the sample is then ready for further testing such as quantitative PCR and genotypic drug-susceptibility analysis that can confirm TB infection and show whether the pathogen is resistant to certain treatments.
The researchers used a panel of 50 characterized sputum samples to optimize the prototype, and then compared its performance to culture diagnosis. A blinded screening of 100 characterized sputum samples found the prototype reported the same result as culture diagnosis for all of the smear-negative samples and for 87% of the smear-positive samples. Limiting the analysis to samples with high bacterial burdens increased the smear-positive concordance to 100%.
Work is underway to improve the prototype. Increasing sample throughput could improve sensitivity and accelerate testing, and refinements to biological processing and device design are expected to further enhance performance.
Statement from Elsevier Press Release:
Lead investigator Philip D. Butcher, PhD, St George’s, University of London, explained, “The global burden of TB is not improving. Although it is infectious, it is highly treatable. However, affordable diagnostics suitable for implementation at the point of care are needed to reach the ‘missing millions.’ Our TB research group at St George’s has a longstanding appreciation of the global imperative for improved diagnostics for TB, and we realized that novel technologies may provide an answer. We saw an opportunity by collaborating on a new chip-based technology using dielectrophoresis to selectively isolate Mtb bacilli from sputum samples.”
The investigators describe a prototype microfluidic lab-on-a-chip system called CAPTURE-XT® from QuantuMDx that can process solubilized sputum from suspected TB patients, capture Mtb bacilli for visual analysis (as a substitute for smear microscopy), and provide a purified sample for molecular confirmation by quantitative PCR (qPCR) and ultimately for genotypic drug-susceptibility analysis. CAPTURE-XT® technology relies on the principle of dielectrophoresis – a little-utilized technique that can be tuned to selectively attract or repel specific particles or cells based upon their dielectric properties. In this case, it is the Mtb bacteria that cause TB that are specifically captured and concentrated, while the other sputum contents are washed away.
After optimization using a panel of 50 characterized sputum samples, the performance of the prototype was assessed by a blinded screening of 100 characterized and bio-banked sputum samples provided by the Foundation for Innovative New Diagnostics (FIND).
Concordance with culture diagnosis was 100% for smear negative samples and 87% for smear positive samples. Of the smear positive samples, the high burden sample concordance was 100%. These results demonstrate the potential of the technology to provide a powerful sample preparation tool that could function as a front-end platform for enhanced molecular detection. This versatile tool could equally be applied as a visual detection diagnostic, potentially associated with bacterial identification for low-cost screening.
Jonathan O’Halloran, PhD, founder and Chief Executive Officer of QuantuMDx Group Ltd said, “The CAPTURE-XT® technology is truly revolutionary and will have an impact in many different diseases from sepsis to oncology (circulating tumor cells). This application in Mtb is truly exceptional as its ultra-low cost and ultra-high sensitivity will profoundly improve equitable access to quality diagnoses for hundreds of millions of people. This chip-based technology exploits the physiological property of the TB bacteria to be specifically collected onto the device so that small numbers can be visualized on the chip electrodes and act as a visual readout to replace the lab-based sputum smear/microscopic methods, which typically have low detection rates and require training laboratory staff, at molecular-like sensitivities and at a fraction of the price. Moreover, when used as a front-end to downstream cellular, protein, and molecular devices, the possibilities are almost limitless.”
Professor Butcher concluded, “Collaboration between University-based academic researchers and biotechnology industry scientists presents a way forward to develop new approaches for some of the world’s greatest healthcare challenges, such as TB. This new chip-based technology could bring diagnostics to the patients that need it and also, by more accessible case-finding, prevent the further spread of this disease.”
Co-author Heather Murton, PhD, LEX Diagnostics, Melbourn, UK, and formerly of QuantuMDx Group Ltd, said, “Tuberculosis is one of the oldest challenges faced in human healthcare. This technology has the potential to meet the expectations for a mobile TB diagnostic, and it is exciting to see a seemingly abstract physics principle successfully applied to a neglected disease area.”
TB is the 13th leading cause of death worldwide, and until COVID-19 was the leading cause of death from a single infectious disease – more than malaria and HIV. Globally it infects 10 million individuals and kills 1.4 million individuals every year, of whom 230,000 are children.
Diagnostics with increased sensitivity and expanded drug susceptibility testing are also needed to address drug resistance and diagnose low-bacterial burden cases.