24 October 2018, New York - WHO and partners released a series of documents today to improve drug susceptibility testing for TB in laboratories worldwide.
The technical documents put together the latest knowledge on molecular mechanisms of drug resistance in Mycobacterium tuberculosis - the cause of human TB - and describe state-of-the art testing methods for determining drug resistance in the laboratory in order to design the most appropriate regimens for patients requiring treatment for drug-resistant TB.
These documents include a Technical Guide on the use of next-generation sequencing technologies for the detection of mutations associated with drug resistance in M. tuberculosis; a Technical Guide on interpreting results from molecular line probe assays for the detection of drug-resistant TB; and a Technical Manual for drug susceptibility testing of medicines used in the treatment of TB.
"Innovations in basic science and new platforms for testing are elevating our understanding of TB drug resistance’, said Dr Tereza Kasaeva, Director of the WHO Global TB Programme. "We know more about TB drug resistance than ever before, thanks to molecular science and the ability of innovators to transform such findings into modern technologies for rapid diagnosis."
Drug resistance in TB occurs mainly through specific changes (mutations) in the DNA of TB bacteria. For some anti-TB drugs these mutations are clustered together in specific segments (or genes) in the DNA, while for others the mutations are spread throughout the DNA (or genome) of the bacterium.
These mutations can be detected by advanced and sophisticated molecular techniques called ‘sequencing’, which extracts and amplifies the bacterial DNA and identifies specific mutations that may confer drug resistance against individual anti-TB medications (such as rifampicin or isoniazid) or groups of anti-TB medicines (such as fluoroquinolones or injectables).
Current technologies are able to generate sequence data at specific DNA segments involved in TB drug resistance (also called ‘targeted sequencing’) or the whole genome of M. tuberculosis (also called ‘whole genome sequencing’). Sequencing platforms generate a huge amount of data that is highly complex to interpret, as not all mutations necessarily predict the development of drug resistance clinically in patients. In addition, the genetic mutations that may confer resistance to newer anti-TB drugs such as bedaquiline and delamanid are not yet fully known.
FIND and WHO, in collaboration with technical partners, have developed a standardized procedure that objectively evaluated the frequency of different mutations in M. tuberculosis and whether they can be used to accurately ‘predict’ resistance against the medicines used in treatment. The Technical Guide released today includes a consensus list of graded mutations along with the level of confidence that these mutations are associated with clinical drug resistance. Evidence on sequencing is rapidly increasing and the Technical Guide will be updated regularly to reflect the most recent changes.
The joint development of these guides reflects the critical role of diagnostics in fighting the spread of TB drug resistance. Innovative rapid tools, such as sequencing, improve patient care while also protecting the efficacy of anti-TB drugs,” said Catharina Boehme, CEO, FIND. “This work complements our collaborative efforts to establish the ReSeqTB knowledgebase at WHO to support sequencing for surveillance, and ultimately clinical diagnosis, of drug-resistant TB.”
Improved understanding of the molecular basis of resistance has greatly facilitated the development of genotypic tests for the rapid detection of resistance to specific anti-TB medicines drugs. These include commercial line probe assays that screen specific genes of TB bacteria for mutations conferring resistance to key medicines such as rifampicin, isoniazid and the fluoroquinolones.
As with whole genome sequencing technologies, line probe assay results require careful interpretation of the mutations detected in order to guide treatment. WHO and the Global Laboratory Initiative (GLI) of the Stop TB Partnership has therefore developed a technical guide for interpretation of LPA results and the clinical/treatment implications for specific mutations detected.
Testing for drug resistance classically requires growing TB bacteria in the laboratory and exposing them to different levels of individual medicines to determine which ‘critical concentration’ best defines resistance or drug susceptibility. This approach, called ‘phenotypic testing’ has been the reference standard for TB drug susceptibility testing for many years and remains essential for several medicines used in TB treatment.
Following a Technical Report released by WHO and FIND in March 2018, that included internationally agreed critical concentrations for drug-susceptibility testing, a Technical Manual has now been developed that describes the standardized laboratory protocols for performing these tests for both first- and second-line anti-TB medicines.
"Together, this series of technical documents provide the current global norms and standards for TB drug susceptibility testing", said Dr Karin Weyer, Coordinator of TB Diagnostics, Laboratories & Drug Resistance at WHO. "Increasingly, precision-medicine is expected to guide the diagnosis of drug-resistant TB and ensure appropriate treatment for all patients".