Tuberculosis

A public health crisis

What is Tuberculosis?
A type of bacterial infection
Mycobacterium tuberculosis
Transmitted through respiratory droplet
Infection begins in lungs & can spread to other organs








Source: Center for Biologic Counterterrorism and Emerging Diseases, 2008

2 TB-related conditions exist:
(A) Latent tuberculosis infection (LTBI)
- TB remains in dormant state without spreading or causing
any symptoms

(B) TB disease
- TB is in active state due to inability of immune system to deter
against M. tuberculosis



Source: CDC, 2012

Tuberculosis treatment
Amikacin (AMK)
Amoxiclav (AMX)
Amoxicillin/clavulanate
Bedaquiline
Capreomycin (CM)
Clarithromycin (CIP)
Clofloxacin (CFX)
Clofamizine (CLO)
Cycloserine (CS)
Ethambutol (EMB)
Ethionamide (ETH)
Imipenem/cilastatin
Isoniazid (INH)
Kanamycin (KM)
Levofloxacin (LFX)
Linezolid (LZD)
Moxifloxacin (MFX)
Ofloxacin (OFX)
Para-aminosalicylic acid (PAS)
Protionamide
Pyrazinamid (PZA)
Rifabutin (RFB)
Rifampicin/Rifampin (RIF)
Rifapentine
Streptomycin (SM)
Terizidone
Thionamide
Thioacetazone (THZ)

Issues with tuberculosis treatment
Long treatment period, usually more than 6 months

Complete the full course of TB medications

If failed to do so, TB bacteria can survive & proliferate

Overtime, TB bacteria can become resistant to treatment

Types of drug resistant tuberculosis
Multi-drug resistant tuberculosis (MDR-TB)
Resistant to isoniazid & rifampicin/rifampin

Extensively drug resistant tuberculosis (XDR-TB)
Resistant to isoniazid & rifampicin + any fluoroquinolone +
any one of the three injectable second-line drugs (amikacin,
kanamycin or capreomycin)

Total drug resistant tuberculosis (TDR-TB)
Resistance to all first and second line drugs tested
How to cure drug-resistant TB?
M. tuberculosis is a slow growing bacterium, phenotypic
conventional drug susceptibility test (DST) will take weeks
to complete.

Hence, rapid diagnostic tools to effectively test for
resistance in M. tuberculosis are urgently needed.

Since drug resistance is caused by genomic mutations in
the gene of M. tuberculosis, several rapid test kit that
detects for genomic mutations have been developed.

Detection methods
Xpert MTB/RIF (Cepheid) Genotype MTBDRplus
(hain-lifescience)
Only detects for specific
nucleotide mutations in RRDR
of rpoB gene

Only detects for specific nucleotide
mutations in rpoB, katG, inhA genes











Limitations:
- Unable to detect for resistances originating from mutations of other genes!


Source: Hain Lifescience, 2008 Source: Cepheid, 2010
Rifampicin
Reduced binding to RNA polymerase
- Cluster of mutations at Rifampicin Resistance Determining
Region, located in rpoB gene






Aim of the project
Look into existing database to identify genetic
markers or regions to assist future rapid
diagnostic development

Existing database explored includes:
(1) TBDReaMDB
(2) Genome-wide Mycobacterium tuberculosis
Variation (GMTV)
(3) Broad institute

GMTV Broad Institute TBDReaMDB
User-friendly
Lineage information
was available
27 types of drugs
resistance genomic
mutations
Validated data
Specified mutations
to drugs
GMTV Broad Institute TBDReaMDB
Isolates with at least
one drug resistant
Russian populations
Mutations are not
specified to drugs
Not user-friendly
Mutations are not
specified to drugs
Lineage information
was not available


9 types of drug
resistant genomic
mutations
Some codons
reported are only
estimated values or
not reported
Lineage information
was not available

Advantages
Limitations

Moxifloxacin
Drug resistance
Total
Yes No
Mutation at codon 94
of gyrA gene
46 10 56
No mutation at codon
94 of gyrA gene
59 285 344
Total 105 295 400
Value
Odd ratios 22.223
P-value <0.0001
Ofloxacin
Drug resistance
Total
Yes No
Mutation at codon 94
of gyrA gene
63 0 63
No mutation at codon
94 of gyrA gene
80 377 457
Total 143 377 500
Value
Odd ratios 595.6
P-value <0.0001
What can we do next?
Plot a graph with our gathered data to look for similar
regions like RRDR/QRDR
- X axis: codon positions; Y axis: absolute mutation counts








Source: Broad institute, 2014

Contribute to development of new molecular testing
methods to enhance detection for drug resistance
Mycobacterium tuberculosis
Recent works
Thank you!