Multi Drug Resistant Tuberculosis

Basics, Concerns and Diagnosis
Dr.T.V.Rao MD

A Tribute to Robert Koch
Discoverer of Mycobacterium Tuberculosis

Dr.T.V.Rao MD


Nobody is absolutely Immune to Tuberculosis

Dr.T.V.Rao MD


Basic concepts – Keep facts
Primary (Initial) resistance
TB patient’s initial Mycobacterium tuberculosis population resistant to drugs

Secondary (Acquired) resistance
Drug-resistant M. tuberculosis in initial population selected by inappropriate drug use (inadequate treatment or non-adherence)

Dr.T.V.Rao MD


Changing Definition of MDR TB
M. tb resistant to INH, streptomycin and/or PAS 1980s-current: M. tb resistant to at least INH and Rifampin

Definition of MDR - TB
MDR-TB caused by strains of Mycobacterium Tuberculosis resistant both Rifampicin and Isoniazid with or without resistance to other drugs. Single Isoniazid or Rifampicin resistance is not MDR - TB

MDR TB is a laboratory diagnosis, Not a Clinical assumption
Dr.T.V.Rao MD

Why INH and Rifampin
Most potent and bactericidal drugs for Tuberculosis
Tb can be treated effectively with INH+Rif alone Mono-resistance to one of them can be treated effectively with a regimen containing the other agent with very low failure rate (2.55%) Failure rate when INH+Rif resistant is 44% in non-HIV and 70% in HIV patients

Duration required for cure doubles to triples.
Dr.T.V.Rao MD


% of MDR-TB among new TB cases 1994-2007

Dr.T.V.Rao MD


Dr.T.V.Rao MD


Epidemiology of MDR TB

Genesis of MDR TB
Resistance is a man-made amplification of a natural phenomenon.

Inadequate drug delivery is main cause of secondary drug resistance. Secondary drug resistance is the main cause of primary drug resistance due to transmission of resistant strains. MDR due to spontaneous mutations is not possible as the genes encoding resistance for anti TB are unlinked. 14 Dr.T.V.Rao MD

When to suspect MDR TB Re-treatment patients who’s sputum smear
remains positive after three months’ of intensive therapy

Treatment failure and interruption cases
Close contacts of MDR tuberculosis cases

Positive diagnoses with;
TB culture and susceptibility testing
Dr.T.V.Rao MD

Factors Contributing to Development and Spread of MDR and XDR TB
Weak TB programs (DOTS)

Low completion/cure rates Lack of treatment follow up and patient support

Unreliable drug supply
Diagnostic delay
Absent or inadequate infection control measures Uncontrolled use of 2nd line drugs

Dr.T.V.Rao MD


Mechanism of resistance
Chromosomally mediated
Loss of catalase/peroxidase Mutation in mycolic acid synthesis

Regulators of peroxide response
Dr.T.V.Rao MD 18

Mechanism of resistance
Reduced binding to RNA polymerase Clusters of mutations at “Rifampin Resistance Determining Region” (RRDR)

Reduced Cell wall permeability
Dr.T.V.Rao MD 19

The laboratory is an essential component in TB control programs, and broader access to DST is a priority for most countries. Early choice of appropriate treatment is an essential determinant of favourable outcome, and rapid determination of drug resistance can allow a customized approach to treatment early in the course of the disease and can potentially reduce morbidity, mortality and infectiousness
Dr.T.V.Rao MD

Using standardized DST procedures with conventional methods, eight to 12 weeks are required to identify drug-resistant microorganisms on solid media (ie, Lowenstein-Jensen medium). In general, such methods assess inhibition of M tuberculosis growth in the presence of antibiotics to distinguish between susceptible and resistant strains
Dr.T.V.Rao MD

Proportion method
The proportion method allows precise determination of the proportion of resistant mutants to a certain drug; the resistance ratio method compares the resistance of an unknown strain with that of a standard laboratory strain. While relatively inexpensive and undemanding of sophisticated equipment, results usually take weeks and this is challenging; inappropriate choice of treatment regimen may result in death within weeks of initiation, such as in the case of XDR-TB (especially in HIV-infected patients)
Dr.T.V.Rao MD

Conventional Methods are Outdated
In addition, delayed identification of drug resistance results in inadequate treatment, which may generate additional drug resistance and continued transmission in the community.

The BACTEC 460 TB radiometric system (Becton Dickinson, USA) was considered to be a major advancement when it was introduced, but has been replaced by the Mycobacteria Growth Indicator Tube system (Becton Dickinson, USA). Several published studies have shown the excellent performance of the Mycobacteria Growth Indicator Tube system for the rapid detection of resistance to first- and second-line anti-TB drugs . Detection of drug resistance can be accomplished in days rather than weeks, although still constrained by high cost (equipment and consumables).

Diagnosis of MDR-TB and XDR-TB
The diagnosis of MDR-TB and XDR-TB is hampered by the absence of effective and affordable rapid diagnostic techniques for drug sensitivity. Several approaches, phenotypic and molecular, have been explored to develop rapid, reliable and accurate methods for the rapid detection of drug resistance in M tuberculosis. These methods should also be evaluated and applied in high-incidence areas
Dr.T.V.Rao MD

Susceptibility Testing
Direct and indirect testing Primary Drugs testing


Ethambutol (*)
Pyrazinamide (*)

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Drug susceptibility testing (DST)
DST is recommended for all new cases for all first line drugs with specimens taken before initiating treatment.?

Accuracy is more important than speed
DST results should come from a small number of wellequipped, experienced laboratories who participate and perform well in an international DST quality control scheme.

The WHO Supranational Laboratory Quality Control Network offers the greatest global coverage.
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Drug susceptibility Testing
Assessment of growth inhibition on solid media containing various dilutions of the drug, in comparison with the test strains.
As the method depend observation of growth Results are not available until several weeks after isolation of the organism.
Dr.T.V.Rao MD 28

Microscopic Observation of Drug Susceptibility Testing

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MODS affordable Technically Feasible
MODS arose during experiments conducted by Luz Caviedes under the guidance of Professor Robert Gilman at Universidad Peruana Cayetano Heredia in Lima, Peru in the late 1990s in which a colorimetric test for TB growth was being investigated. The observation that micro colonies could be seen under the microscope long before a colour change occurred prompted the development of MODS.
Dr.T.V.Rao MD

Observation of Growth in liquid Media
MODS depends upon three key principles (which have been known for decades): (1) Mycobacterium tuberculosis grows faster in liquid (broth) than on solid media, (2) in liquid cultures M. tuberculosis grows in a visually characteristic manner (tangles, cording) which can be observed under the microscope long before the naked eye could visualize colonies on solid agar
Dr.T.V.Rao MD

Least time required for detection of MDR

Incorporation of anti-TB drugs into broth cultures at the outset enables direct susceptibility testing from sputum samples

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Advantages of MODS methodology in MDR detection
• All the chemical ingredients are available locally, except few which can be acquired easily. • Existing infrastructure in District and Teaching hospital can be adopted for implementation of MODS • Risk to technician handling the specimens is minimal, there is no absolute need to obtain grade III safety cabinets, Technology transfer is easier all the new technical manpower can be trained easily.
Dr.T.V.Rao MD 33

Other accredited Methods
Radiometric and Non radiometric methods Nucleic acid technology – effective up to 95% in mutations to rifampicin resistance to gene rpoB gene

Drug susceptibility testing (DST)
As a minimum, laboratories supplying DST data, should correctly identify resistance to isoniazid and rifampicin in over 90% of quality control samples in two out of the last three quality control rounds.
Dr.T.V.Rao MD 35

Detection of Rifampicin Drug susceptibility testing (DST) is more important.
Early identification of mycobacterial growth as M. tuberculosis complex and the identification of rifampicin resistance should be the first priority as rifampicin resistance invalidates standard 6 month short-course chemotherapy and is a useful marker in most countries for MDR-TB. Laboratories should aim to identify isolates as M. tuberculosis complex and perform rifampicin resistance in 90% of isolates within 1-2 working days. This is technologically feasible.
Dr.T.V.Rao MD

Drug susceptibility testing
For DST laboratories, modern molecular techniques permit the successful identification of isoniazid resistance in at least 75% of mycobacterial cultures within 1-2 working days and are useful preliminary screens for isoniazid resistance.

Secondary Drugs testing: [lack of standardized methods!]
Ofloxacin, quinolones Ethionamide Kanamycin

! Ensure quality control and quality assurance ?

Other WHO-Endorsed Tools
Liquid culture (e.g. MGIT, BacT/ALERT) Capilia TB Rapid strip test that detects a TB-specific antigen from culture

Molecular line probe assays (e.g. Genotype MTBDRplus, INNO-LiPA Rif.TB)
Strip test for detection of TB and drug-resistance conferring mutations

Dr.T.V.Rao MD


The identification of specific mutations responsible for drug resistance has facilitated the development of novel, rapid molecular tools for DST. The detection of RIF resistance is traditionally used as a predictor of MDR-TB – its positive predictive value is a function of the sensitivity and specificity of RIF resistance testing and the prevalence of MDR and non-MDR RIF resistance, which is highest among previously treated cases in settings with high MDR prevalence and low non-MDR RIF resistance.

Dr.T.V.Rao MD



The Xpert MTB/RIF
The Xpert MTB/RIF is a cartridge-based, automated diagnostic test that can identify Mycobacterium tuberculosis (MTB)DNA and resistance to rifampicin (RIF)by nucleic acid amplification technique(NAAT
Dr.T.V.Rao MD 42

WHO Endorses Xpert MTB/RIF
In December 2010, the World Health Organization (WHO) endorsed the Xpert MTB/RIF for use in TB endemic countries[2] and declared it a major milestone for global TB diagnosis.
Dr.T.V.Rao MD 44

Xpert MTB/RIF detects DNA sequences
The Xpert MTB/RIF detects DNA sequences specific for Mycobacterium tuberculosis and rifampicin resistance by polymerase chain reaction It is based on the Cepheid GeneXpert system, a platform for rapid and simple-to-use nucleic acid amplification tests (NAAT).

Dr.T.V.Rao MD


How Xpert® MTB/RIF Works
The Xpert® MTB/RIF purifies and concentrates Mycobacterium tuberculosis bacilli from sputum samples, isolates genomic material from the captured bacteria by sonication and subsequently amplifies the genomic DNA by PCR. Dr.T.V.Rao MD 46

Xpert® MTB/RIF Helps in Faster Diagnosis of Resistance to Rifampicin
The process identifies all the clinically relevant Rifampicin resistance inducing mutations in the RNA polymerase beta (rpoB) gene in the Mycobacterium tuberculosis genome in a real time format using fluorescent probes called molecular beacons. Results are obtained from unprocessed sputum samples in 90 minutes, with minimal biohazard and very little technical training required to operate.This test was developed as an ondemand near patient technology which could be performed even in a doctor's office if necessary.

Summary Drug resistant TB
Drug-resistant TB poses a grave public health threat especially in high HIV prevalence settings XDR-TB strains have been found in all regions of the world XDR-TB occurs as a result inadequate TB control programmes XDR-TB, if identified early, can be treated and cured but experience limited to low HIV prevalence settings Infection control measures must be strengthened XDR-TB underlines the need for investment in basic TB control plus development of new TB diagnostics, treatments and vaccines

Dr.T.V.Rao MD


No testing method replaces clinical assessment for Tuberculosis

Dr.T.V.Rao MD


Progress in diagnosing multidrugresistant tuberculosis
20 March 2014 | Geneva - Almost half a million people fell ill with multidrugresistant tuberculosis (MDR-TB) in 2012, yet less than one in 4 of these people was diagnosed, mainly due to a lack of access to quality diagnostic services.
Dr.T.V.Rao MD 51

Koch failed to conquer tuberculosis, which still causes enormous health problems worldwide 100 years after his Nobel award. The scientific academies noted that the triumphant discovery of 1882 was followed by a succession of failures: first of all, the failed attempt to present tuberculin as a remedy against tuberculosis in 1890-91, which severely damaged Koch's reputation
Medical History, 2001, 45: 1-32 CHRISTOPH GRADMANN*

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Tuberculosis is a Concern for Everyone

Dr.T.V.Rao MD


MDR Tuberculosis is Global Emergency

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Visit me for more Articles of Interest on Infectious Diseases

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Programme Created by Dr.T.V.Rao MD for Medical and Health Professionals in the Developing World

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