Faster and more portable tuberculosis diagnosis will be reality in a few years

Infectious diseases physician Ana Luíza Gibertoni Cruz alerts that the current delay diagnosing and treating the disease, besides affecting the patient’

Imagine a device that, within minutes, sequences the whole genome of tuberculosis (TB), thus providing a fast and effective answer as to which combination of anti-TB drugs is the ideal one to treat each patient, in a very personalised fashion. Please stop imaging now, for this is already a reality with CRyPTIC, Comprehensive Resistance Prediction for Tuberculosis: an International Consortium. The processes involved in this scientific enterprise are still expensive and do require specialised workforce. But this is bound to change quite soon.

According to the Infectious Disease physician Ana Luíza Gibertoni Cruz, in a few years this diagnostic solution will be embedded in routine laboratory workflows, leaving the realms of research to become mainstream microbiology. She is a clinical research fellow with CRyPTIC and a member of the team at the University of Oxford responsible for getting the project out of the ground. This consortium unites more than 10 research institutions all over the world, a truly global effort in the fight against TB.

This fast diagnostic solution is achieved by means of whole genome sequencing (WGS) of the TB mycobacteria that infect each patient. Bespoke software analyse the longer than 4-million base-pair sequence that forms TB genome, identifying the mutations associated with antimicrobial resistance. The end-product of this analysis is a laboratory report that indicates which anti-TB drugs will likely be more effective against infection; the report is issued in a straightforward language so that clinicians are able to come up with the ideal combination of drugs for their patients, even without any prior knowledge of genomics.

This is how it works: DNA is initially extracted from clinical TB specimens, and subjected to chemical processes that chop it into small fragments—this is known as library prep. These fragments are subsequently amplified and assembled, within the sequencing machine, in a nucleotide sequence. Electronic files storing this information go through analytical pipelines, pieces of software that, after confirming the identity of the infectious agent, map the obtained DNA sequence against a “wild-type”—or pan-susceptible— TB reference-genome. Different nucleotides occupying same genome positions, what commonly constitutes mutations, are then matched to a genotypic database and, voila: a list with the susceptibility profiles to each drug is emitted.

“We also hope to discover new mutations using sophisticated in vitro selection of mutants techniques and also machine learning approaches”, says the Brazilian researcher.

The ID doctor ponders that difficulties underlying TB diagnosis and treatment not only affect the lives of the patients, but also those of their families and acquaintances, all of whom are posed at high-risk for getting infected with TB as it is transmitted through the air.

And what is it that makes TB diagnosis such a time-consuming process? According to Ana Luíza Cruz, mycobacteriology still relies heavily on culture-dependent methods; ones needs to wait for the mycobacteria to grow on culture media containing the tested antibiotic so that drug-resistance can be detected. “Unlikely most of other common bacteria, TB has a very slow growth rate, hence the delays”, she explains.

In 2015, more than 9 million people fell ill with TB, out of whom 1.5 million are estimated to have died. TB is the top infectious disease killer in the world, and too heavy a burden on those affected, who are left too poorly to lead productive, worthy-living lives.

An expensive solution

Equipment and consumables used in CRyPTIC are not portable yet. Sample preparation is done in molecular microbiology labs, and the sequencer is still large and expensive. For the time being, sequencing is performed by reference centres that have the expertise in sample preparation, the financial resources to purchase equipment, and the support of a bioinformatics team to analyse results.

Nonetheless, she affirms that portable technologies are being developed and are likely to be available in the coming 5 years. This will provide enough time for the development of automated techniques to deal with samples and for the compilation of a complete genotypic database that does not let any mutation go unrecognised.

When this happens, this technology will have become even simpler, allowing for a diagnostic revolution to take place at the bedside. Ana Luíza Cruz believes that “it is a matter of time until WGS is a reality wherever it is needed the most”.

For more information:

Click here to ead the project’s release, published on the Oxford University website.

Read here the article published in the British scientific journal “New Scientist”.

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