ABSTRACT
Tuberculosis (TB), caused by Mycobacterium tuberculosis (M.tb), is a leading infectious disease and cause of
death worldwide. The growing burden of drug-resistant (DR)-TB is complicating TB treatment. Early diagnosis of
TB with drug susceptibility testing (DST) is critical for successful treatment and is the first pillar of the World
Health Organization’s (WHO) End TB Strategy. DST is achieved via phenotypic or genotypic methods.
Traditionally, phenotypic DST is performed on solid (Löwenstein Jensen) or liquid media (MGIT) in a two-step
process: first a culture to identify M.tb growth, and then re-culture of the isolate with the drugs to be tested. In
addition to requiring biosafety level II-plus labs, the DST process, if available in low-middle income settings, can
take 42 to ~6 months from sample collection to notification of results to the clinical provider resulting in treatment
delays, continued transmission, and higher mortality. Conversely, genotypic DST has many advantages,
including a reduced time to result (< 2h for GeneXpert) and the possibility of deployment to at or near point of
care (POC). However, its widespread use in high TB burden resource-limited settings is hindered by the need
for regular power supply and importantly cost. Thus, NIH/NIAID is redirecting attention to innovative and simple
phenotypic DST solutions to be deployed at or near POC.
The goal of this application is to develop the 1G test into the 2G test, providing higher flexibility to perform DST
for 1st and 2nd frontline drugs, including drugs prescribed for DS- and DR-TB regimens such RIPE (DS-TB oral
regimen composed of RIF/INH/PZA/Ethambutol), HPMZ (DS-TB 4-month short course oral drug regimen
composed of INH/Rifapentine/MFX/PZA) and BPaL [MDR- and pre-XDR oral drug regimen composed of
bedaquiline (BDQ), pretomanid (PMD) and linezolid (LNZ)], as well as clofazimine (CFZ) and delamanid (DLM),
other WHO recommended oral agents for DR-TB. Because the 2G test is non-proprietary, its cost is expected to
be extremely low (< $8) and mainly driven by the cost of drugs. Further, for the 1G test we tested a simple step
to digest/decontaminate sputa that does not require equipment, meeting the near to POC test definition. We will
optimize this sputum-processing protocol for use with the 2G test. We propose to: Aim 1) Develop and validate
the 2G test by defining the stability and critical concentration (CC) for new drugs against known DR-M.tb strains,
and optimize appropriate sputum digestion and decontamination protocols for this test; Aim 2) Determine the
agreement of the 2G test with current gold standard methods for phenotypic DST for each of the 11 drugs, and
Aim 3) Determine the accuracy of the 2G test against reference phenotypic DST protocols using freshly collected
sputa in field settings and assess its usability, acceptability, and feasibility.
We expect that the novel, simple, affordable and sustainable 2G test will provide a significant improvement when
compared to current phenotypic DST reference methods, allowing rapid and tailored treatment for DS-/DR-TB
in low- and middle-income countries with high TB burden.
