Whole Genome Sequencing For Tuberculosis
Posted By HealthcareOnTime Team
Posted on 2022-02-02
Tuberculosis (TB) has been highlighted in medical history as
The Leading Killer. Some common terms for TB worldwide include
'The Captain of all these Men of Death', 'Phthisis', 'Scrofula', 'The
White Plague'. The emergence and spread of Drug Resistant (DR)
Tuberculosis is a major concern that threatens global health.
India has the highest estimated deaths among countries
reporting TB statistics to WHO, with 4,36,000 deaths in
HIVnegative adults and 9,500 in HIV-positive adults. Rapid
diagnosis and identification of DR strains are prerequisites
for eradicating TB. Accurate diagnosis aids in customizing
individualized treatment, thus reducing infectiousness,
morbidity and mortality. Without treatment, about 5 to
10% of infected people develop TB disease at some point
in their lives.
Although, there exists a number of technologies for early
detection of threats, constant improvisation has elevated
the degree of usage for a wider population. Let's look at
some interesting detection techniques, with a focus on
sequencing, a technological advancement in the field of
Sites of TB Infection
Based on site of infection, TB is classified as,
Pulmonary TB - It is a contagious bacterial infection that affects the lungs.
This disease is indicated by symptoms like cough and abnormal chest
-Extrapulmonary TB - It affects organs other than the lungs, including
larynx, lymph nodes, pleura, brain, kidneys,bones and joints. Infected
individuals usually are not infectious, unless they have pulmonary along
with extrapulmonary disease or have an infected oral cavity or larynx,
or an infected open abscess or lession.
Miliary TB-A potentially fatal form of active TB, occurs when bacteria
enters the bloodstream and disseminates to all parts of the body, causing
infection in multiple sites.
TB genome sequencing/WGS-TB
After tissue invasion, the concern that adds to severity of TB infection
is the property of Mycobacterium tuberculosis (MTB) to develop
resistance against the antimicrobial drugs used for treatment. It
limits the treatment options thereby putting the patient at higher
risk of incurable TB.
Types of drug resistant TB
-Multi drug resistant TB (MDR-TB)
MDR-TB is caused by TB bacteria resistant to two of the most important
anti-TB drugs- Isoniazid and Rifampicin. It occurs either when a person
is infected with a resistant strain or when improper treatment leads
to drug resistance in a particular infecting strain. In 2018, India had
58,347 (44%) MDR-TB cases out of 1,30,000 drug-resistant TB cases.
-Extensively drug resistant TB (XDR-TB)
XDR-TB is caused by rare type of MDR-TB that is resistant to Isoniazid,
Rifampin, as well as Fluoroquinolones and second-line injectable drugs
like Amikacin, Kanamycin, and Capreomycin. Among the MDR-TB patients,
XDR-TB rate was found to be 1.3%
MTB spreads through infectious droplet nuclei (1-5 microns in diameter)
via air, when a person with pulmonary or laryngeal TB disease coughs,
sneezes, shouts, or sings.
Types of TB diagnosis
It is necessary to choose appropriate screening or diagnostic testing for
detecting mycobacteria, as accurate diagnosis will help in customizing
a treatment, thus reducing infectiousness, morbidity and mortality,
WHO recommended TB diagnostics tools
-TB Screening Test - Mantoux tuberculin skin test (TST) and
interferon-gamma release assays (IGRAs) are two widely
used tests for detecting Latent TB infection. The IGRAs
include tests like QuantiFERON-TB Gold Plus (QFT-Plus)
and T-SPOT.TB test (T-Spot)
-Smear Microscopy - Sputum microscopic examination is the
most conventional diagnostic test for pulmonary TB. It is widely
used in low-income countries due to its rapidity, low cost and
high positive predictive value for TB.
-Chest Radiograph - A posterior anterior radiograph aids in detecting
chest abnormalities. These abnormalities may suggest pulmonary TB,
but cannot be used to make a definitive diagnosis.
-Lipoarabinomannan Urine Strip Test - Lateral flow urine Lipoarabinomannan
strip test (LF-LAM) developed for TB diagnosis is based on detection of
mycobacterial lipoarabinomannan antigen in urine. It is used to detect
active TB disease in HIV patients.
-Culture (MGIT) and Drug Susceptibility Testing (DST) - Culture is World Health
Organization (WHO) recommended gold standard technique for TB diagnosis.
Organism isolated on solid and liquid culture medium i only important for
definitive diagnosis but also for determining phenotypic DST. Identifying
drug resistance at the earliest ensures effective treatment.
-TB-PCR - Nucleic acid amplification tests (NAAT) and alternative methods
like ligase chain reaction, strain displacement amplification, loop-mediated
isothermal amplification (LAMP), transcription mediated amplification
and real time PCR are used for detection and identification of MTB.
-CB-NAAT/ GeneXpert MTB/RIF Test - Cartridgebased nucleic
test (CB-NAAT) or Xpert MTB/RIF assay, is an automated semi-quantitative
nested real-time PCR that uses molecular beacon technology to detect M.
tuberculosis complex (MTBC) and rifampin resistance. It is a rapid, highly
sensitive and specific assay that generate results in less than 2 hours.
-Line Probe Assay (LPA) - LPA is novel technique designed to identify MTBC
and drug sensitivity to Rifampicin and Isoniazid. It uses PCR and reverse
hybridization methods for detection of mutations associated with drug
resistance. Resistance of second line of drugs can be detected by second
-Sequencing Based Technique - DNA sequencing of variable genomic regions
allows for the rapid and accurate identification of drug resistant TB. It is
based on determination of species-specific nucleotide sequences, which
are then compared to known quality-controlled sequences from in-house
or commercially available databases.
Advances in diagnosis and drug susceptibility testing methods for
MTBC are required to control transmission as well as provide better
individualized treatment against MDR/XDR-TB. One of the important
criteria for successfully treating TB patients is to administer the right
treatment at an early stage.
This state-of-art sequencing approach overcomes many challenges
associated with conventional phenotypic testing as well as the
limitations of other less comprehensive molecular tests by providing
rapid, detailed sequence information for multiple gene regions or
whole genomes of interest. Whole genome sequencing (WGS) is a
WHO-approved probe-based method for TB diagnosis and detection
of drug resistance, genetic diversity as well as transmission dynamics
of MTBC. Additional information obtained by Whole genome sequencing on transmission
would help in controlling the outbreak.
WGS-TB can identify mutations in genes that predict phenotypic
resistance against anti-TB drugs within the entire region of microbial
genome. It also allows for more robust MTBC classification into lineages
and sublineages, via the identification of single-nucleotide polymorphisms
(SNPs). This emphasizes the importance of Whole genome sequencing in making informed decisions
prior to antituberculosis therapy.
Benefits of WGS-TB
-Single genome-wide test would replace multiple panel tests
-Detect drug resistance mutations for a broad range of antibiotics
-Reliable and highly sensitive diagnosis
-Speed up TB diagnosis from months to days
-Rapid diagnosis allows immediate action, avoiding wastage of resources
Testing @ Healthcare Ontime
TB genome sequencing offered by Thyrocare is a WGS based test that combines genome sequencing
and analysis for a faster and more sensitive detection of tuberculosis. It is a comprehensive single
test solution for DST profiling against 18 anti-TB drugs.