SBIR-STTR Award

TB is an infection caused by species of the Mycobacterium tuberculosis complex (MTBC). In 2014, 1.5 million people died from the disease, which is unacceptable given that TB is curable. The lack of a technology for accurate TB detection in limited-resource settings is a major obstacle in the fight against the disease. However, available detection methods have poor performance and are not suited for diagnosis in limited resource settings. The most commonly used method to detect TB is smear microscopy, a procedure developed more than 100 years ago that only detects 50% of the cases. Here, we propose to develop a next generation molecular diagnostic test that can dramatically improve TB diagnosis. The new platform will be based on a novel technology capable of detecting single molecules using simple equipment. In the single molecule technique, probe hybridization to a target nucleic acid is directly detected without PCR amplification. The technique is capable of detecting 50 CFU/ml of Mycobacterium tuberculosis cells which is below the limit of detection of the leader PCR-based TB detection assay. In addition, the technique applies a disruptive force to the probe-target duplexes which makes detection highly specific even at room temperature. Thanks to its non- enzymatic detection approach, reagents and equipment are simple and robust, ideal for settings with limited resources. Our overall goal is to develop a molecular platform for TB testing that will be easy-to-use without training, accurate, rapid, inexpensive, portable and battery-operated. In this proposal, we will demonstrate all the properties of the final product by developing a non-automated TB assay and testing it using sputum samples spiked with known quantities of the bacillus. The focus of Aim 1 is to develop an efficient method to capture MTBC RNA targets from sputum samples containing MTBC cells. Aim 2 focuses on developing a multiplex SM-Scanning test and determining the limit of detection, cross-reactivity, sensitivity and specificity of the test. If successful, the new platfom could support other tests that are urgently needed in settings with limited resources such as TB drug susceptibility, Ebola and sexually transmitted diseases.

Public Health Relevance Statement:


Public Health Relevance:
We propose the development of a rapid, sensitive and affordable molecular diagnostic test for the detection of active tuberculosis in limited resource settings.

Project Terms:
Bacillus (bacterium); base; Biological Assay; Cells; Complex; cross reactivity; Cytolysis; Detection; Developed Countries; Development; Diagnosis; Diagnostic; Digestion; Disease; Ebola virus; Equipment; fight against; Filtration; Genus Mycobacterium; Goals; improved; Infection; instrument; internal control; Killings; Legal patent; Marketing; Methods; Microscopy; Molecular; Molecular Diagnostic Testing; Motor; Mycobacterium tuberculosis; new technology; next generation; novel; Nucleic Acids; Oligonucleotides; Patient Care; Patients; Performance; Phase; point of care; Predisposition; Preparation; Primary Health Care; Procedures; Process; Property; public health relevance; Reader; Reagent; Reproduction spores; Resources; RNA; Sampling; Scanning; Sensitivity and Specificity; Sexually Transmitted Diseases; single molecule; Sputum; Techniques; Technology; Temperature; Testing; tool; Training; Tuberculosis; tuberculosis drugs

Tuberculosis (TB) is a major global burden that kills 1.5 million people every year. TB mostly affects developing countries, but the rise of multi-drug and extensively drug resistant strains is a global threat. In order to control and eradicate this disease, it is imperative to have adequate tools to detect active TB in limited-resource settings and point-of-care (POC) settings, which is where most TB patients are seen. However, available detection methods have poor performance and are not suited for diagnosis in limited-resource or POC settings. The most commonly used method to detect TB is smear microscopy, a procedure developed more than 100 years ago that only detects 50% of the cases. In Phase I, we developed a sample preparation and detection procedure that is rapid, robust, easy to automate, and highly sensitive and specific. The new assay is based on Single Molecule Scanning (SMS), a novel technology capable of detecting single molecules using simple equipment. Here, we will develop a fully automated next generation molecular test based on the feasibility of SMS stablished in Phase I. The goal is to replace smear-microscopy for sputum- based diagnosis of TB in high TB-burden countries and to replace PCR-based TB diagnosis in developed countries. If successful, the new assay will be more accurate than the current leader molecular assay (>95% sensitivity, >99% specificity) and its price will be affordable for limited- resource settings ($6/test, $1,200/instrument). In addition, the new assay will satisfy all the characteristics that according to a WHO report are required in a product to replace smear microscopy. The overall goal of Phase II, is to develop and to validate a disposable cartridge and a fully automated instrument for TB testing. Aim 1 focuses on the development of the disposable cartridge, Aim 2 on the development of the portable instrument and Aim 3 on the verification and clinical validation of the final instrument and cartridge design. This evaluation together with the design documentation accumulated during the project will be combined with additional clinical evaluations to obtain CE mark, FDA approval, approval in high TB-burden countries, and the endorsement of the World Health Organization.

Public Health Relevance Statement:
Narrative We propose the development of a novel technology for rapid detection of active tuberculosis without enzymatic reactions. We will develop and validate a disposable cartridge and a fully automated instrument.

Project Terms:
Affect; Automobile Driving; Biochemical Reaction; Biological Assay; Cells; Characteristics; Clinical; Country; Cytolysis; design; Detection; Developed Countries; Developing Countries; Development; Devices; Diagnosis; Disease; Docking; Documentation; Ensure; Epidemic; Equipment; Evaluation; extensive drug resistance; Goals; Healthcare Systems; Image; improved; instrument; Killings; Methods; Microscopy; Molecular; Molecular Diagnostic Testing; Mycobacterium tuberculosis; Mycobacterium tuberculosis H37Rv; new technology; next generation; operation; Patients; Performance; Pharmaceutical Preparations; Phase; point of care; portability; Predisposition; Preparation; Price; Procedures; Production; Protocols documentation; prototype; rapid detection; Reagent; Reporting; research clinical testing; resistant strain; Resources; Ribosomal RNA; Sampling; Scanning; Sexually Transmitted Diseases; single molecule; Specificity; Sputum; System; Temperature; Testing; tool; Tuberculosis; tuberculosis drugs; usability; Validation; World Health Organization