Drugresistanttuberculosis 100227020029 Phpapp01

Drug ResistantDrug Resistant TuberculosisTuberculosis

A Great Human ConcernA Great Human Concern

DrTVRao MDDrTVRao MD

MDR TB ndashGreat Human ConcernMDR TB ndashGreat Human Concern

HISTORY ofHISTORY ofTuberculosisTuberculosis

Tuberculosis Is an Tuberculosis Is an Ancient Disease Ancient Disease Identified as Spinal Identified as Spinal Tuberculosis in Tuberculosis in Egyptian Mummies Egyptian Mummies History dates to History dates to 1550 ndash 1080 BC 1550 ndash 1080 BC Identified Identified by PCRby PCR

A Tribute to Robert Koch A Tribute to Robert Koch Discoverer of MycobacteriumDiscoverer of Mycobacterium

TuberculosisTuberculosis

Global StatusGlobal Status

Nine million people suffer from Nine million people suffer from tuberculosis tuberculosis

Two million people die each Two million people die each yearyear

Tuberculosis accounts for one-Tuberculosis accounts for one-third of third of AIDSAIDS deaths world deaths world wide every yearwide every year

USAIDUSAID Report on Tuberculosis Report on Tuberculosis in Indiain India

India has more new tuberculosis (TB) cases India has more new tuberculosis (TB) cases annually than any other country ranking first annually than any other country ranking first among the 22 high-burden TB countries among the 22 high-burden TB countries worldwide according to the World Health worldwide according to the World Health Organizationrsquos (WHOrsquos) Global TB Report 2009 Organizationrsquos (WHOrsquos) Global TB Report 2009 TB remains one of the leading infectious causes TB remains one of the leading infectious causes of mortality in India causing more than 331000 of mortality in India causing more than 331000 deaths in 2007 There were approximately 196 deaths in 2007 There were approximately 196 million new TB cases in India in 2007 million new TB cases in India in 2007 representing more than 21 percent of all TB representing more than 21 percent of all TB cases worldwide cases worldwide

Nobody is absolutely Immune to Nobody is absolutely Immune to TuberculosisTuberculosis

1908-1920 (Calmette and Guerin)1908-1920 (Calmette and Guerin) Vaccine (BCG)Vaccine (BCG)

Attenuated strain Mycobacterium Bovis Attenuated strain Mycobacterium Bovis

19431943 Streptomycin developedStreptomycin developed

20th November 1944 20th November 1944 Critically ill TB patient injected dramatically Critically ill TB patient injected dramatically

recoveredrecovered

Pharmacological discoveriesPharmacological discoveries

Selman Abraham WaksmanSelman Abraham Waksman Nobel Nobel Prize for his discovery in 1952Prize for his discovery in 1952


1956-1960 1956-1960 Combination therapy of INH and PZA cures TBCombination therapy of INH and PZA cures TB

1955 Cycloserine1955 Cycloserine 1962 Ethambutol 1962 Ethambutol 1963 Rifampicin1963 Rifampicin 1970-19771970-1977

Combination of Rifampicin and Combination of Rifampicin and IIsoniazid adopted as soniazid adopted as International regime for treatment of TBInternational regime for treatment of TB

IntroductionIntroduction

Tuberculosis is an ancient disease amp it Tuberculosis is an ancient disease amp it remains the leading cause of death of remains the leading cause of death of human being human being

It is mainly caused by It is mainly caused by Mycobacterium Mycobacterium tuberculosistuberculosis

Typical tubercle bacilliTypical tubercle bacilli

Human type Human type MtuberculosisMtuberculosis

Bovine type Bovine type MbovisMbovis

Vole type Vole type MmicrotiMmicroti

Human type Human type MafricanumMafricanum

Multi Drug Resistant Multi Drug Resistant TuberculosisTuberculosis

MDR-TBMDR-TB

DefinitionDefinition

MDR-TB caused by strains of MDR-TB caused by strains of Mycobacterium Tuberculosis resistant Mycobacterium Tuberculosis resistant both both Rifampicin and IsoniazidRifampicin and Isoniazid with or with or without resistance to other drugswithout resistance to other drugs

Single Isoniazid or Rifampicin Single Isoniazid or Rifampicin resistance is not resistance is not MDR - TBMDR - TB

MDR TB is a laboratory diagnosisMDR TB is a laboratory diagnosis

MDR-TB amp XDR-TBMDR-TB amp XDR-TBTHE 2008 REPORTTHE 2008 REPORT

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

Classification of DrugsClassification of Drugs 3 Groups depending upon the degree of 3 Groups depending upon the degree of

effectiveness and potential side effectseffectiveness and potential side effects First Line (Primary agents)First Line (Primary agents)

are the most effective and have lowest toxicity are the most effective and have lowest toxicity IsoniazidIsoniazid RifampinRifampin

Second LineSecond Line Less effective and more toxic effectsLess effective and more toxic effects include (in no particular order) p-amino include (in no particular order) p-amino

salicylic acid Streptomycin Ethambutolsalicylic acid Streptomycin Ethambutol Third LineThird Line

are least effective and most toxicare least effective and most toxic Amikacin Amikacin Kanamycin Capreomycin Viomycin Kanamycin Capreomycin Viomycin Kanamycin CycloserineKanamycin Cycloserine

Several Drugs becoming Several Drugs becoming resistantresistant

Basic concepts ndash Keep factsBasic concepts ndash Keep facts

Primary (Initial) resistancePrimary (Initial) resistance TB patientrsquos initial TB patientrsquos initial Mycobacterium Mycobacterium

tuberculosis tuberculosis population resistant population resistant to drugsto drugs

Secondary (Acquired) resistanceSecondary (Acquired) resistance Drug-resistant Drug-resistant M tuberculosis M tuberculosis in initial in initial

population selected by inappropriate drug use population selected by inappropriate drug use (inadequate treatment or non-adherence(inadequate treatment or non-adherence))

When to suspect MDR TBWhen to suspect MDR TB

Re-treatment patients whorsquos sputum Re-treatment patients whorsquos sputum smear remains positive after three monthsrsquo smear remains positive after three monthsrsquo of intensive therapyof intensive therapy

Treatment failure and interruption casesTreatment failure and interruption cases Close contacts of MDR tuberculosis casesClose contacts of MDR tuberculosis cases Positive diagnoses with Positive diagnoses with

TB culture and susceptibility testingTB culture and susceptibility testing

What is extensively drug What is extensively drug resistant tuberculosis (XDR TB)resistant tuberculosis (XDR TB)

Extensively drug resistant TB (XDR TB) is Extensively drug resistant TB (XDR TB) is a relatively rare type of MDR TB XDR TB a relatively rare type of MDR TB XDR TB is defined as TB which is resistant to is defined as TB which is resistant to isoniazid and rifampin plus resistant to isoniazid and rifampin plus resistant to any any fluoroquinolonefluoroquinolone and at least one of and at least one of three injectable second-line drugs three injectable second-line drugs (ie (ie amikacin kanamycin or capreomycin)amikacin kanamycin or capreomycin)

Why XDR - TB a grave concernWhy XDR - TB a grave concern

Because XDR TB is resistant to first-line and Because XDR TB is resistant to first-line and secondline drugs patients are left with treatment secondline drugs patients are left with treatment options that are much less effective options that are much less effective

XDR TB is of special concern for persons with XDR TB is of special concern for persons with HIV infection or other conditions that can HIV infection or other conditions that can weaken the immune system These persons are weaken the immune system These persons are more likely to develop TB disease once they are more likely to develop TB disease once they are infected and also have a higher risk of death infected and also have a higher risk of death once they develop TBonce they develop TB

Global Estimates

ClassificatClassificationion

Estimated Number Estimated Number of Casesof Cases

Estimated Estimated Number of Number of

DeathsDeaths

All forms All forms TBTB

88 million88 million 16 million16 million

MDR TBMDR TB 424000424000 116000116000

XDR TBXDR TB 2700027000 1600016000

Extensively Drug-Resistant Extensively Drug-Resistant Mycobacterium tuberculosisMycobacterium tuberculosis India India

The first XDR TB cases in India The first XDR TB cases in India and the emergence of XDR TB is and the emergence of XDR TB is reported by reported by Rajesh Mondal and Rajesh Mondal and Amita JainAmita Jain King Georges Medical King Georges Medical University Lucknow IndiaUniversity Lucknow India Volume 13 Volume 13 Number 9ndashSeptember 2007 in Number 9ndashSeptember 2007 in Emerging Infectious Emerging Infectious DiseasesDiseases

Global incidence of tuberculosis Still rising as a result of the growing epidemic in

Africa

0

100

200

300

400

500

600

1990 1995 2000 2005 2010 2015

Inci

den

ce p

er 1

000

00 p

er y

ear

World

Cent Euro

East Europe

Est Market

East Medit

Lat America

West Pacific

Sth East Asia

AFR high HIV

AFR low HIV

Drug susceptible TBsect

MDR-TB

1990sect

XDR-TB

2006sect

Total DR

Resistance to HampR ndash

Treatable with 2nd line drugs

Resistance to 2nd line drugs ndashTreatment options seriously restricted

Resistance to all available drugs ndash

No treatment options

or limited resistance manageable with 4 drug regimen - DOTS

Are we Returning to a Pre-Are we Returning to a Pre-antibiotic Eraantibiotic Era

WHO Surveillance and Incidence of WHO Surveillance and Incidence of MDR TBMDR TB

CountryCountry MDR TB of all new cases MDR TB of all new cases

EstoniaEstonia 141141

LatviaLatvia 9090

China (non-DOTS)China (non-DOTS) 7777

China (DOTS)China (DOTS) 2828

RussiaRussia 6060

IndiaIndia 3434

IranIran 5858

DominicanDominican 6666

Ivory CostIvory Cost 5353

Dye et al Global Burden of Multidrug-Resistant TB JID 185(8) 2002Dye et al Global Burden of Multidrug-Resistant TB JID 185(8) 2002

Genesis of MDR TBGenesis of MDR TB

Resistance is a Resistance is a man-made amplificationman-made amplification of a of a natural phenomenonnatural phenomenon

Inadequate drug delivery is main cause of Inadequate drug delivery is main cause of secondary drug resistancesecondary drug resistance

Secondary drug resistance is the main cause Secondary drug resistance is the main cause of primary drug resistance due to transmission of primary drug resistance due to transmission of resistant strainsof resistant strains

MDR due to spontaneous mutations is not possible MDR due to spontaneous mutations is not possible as the genes encoding resistance for anti TB are as the genes encoding resistance for anti TB are unlinkedunlinked

Strains with genetic drug resistance

Wild M TB strain

Acquired drug resistance

Primary drug resistance

Spontaneous mutationSpontaneous mutation

Selection inadequate treatmentSelection inadequate treatment

TransmissionTransmission

Development of anti-tuberculosis drug resistanceDevelopment of anti-tuberculosis drug resistance

Pablos-Mendez et al WHO 1997Pablos-Mendez et al WHO 1997

Factors Contributing to Development Factors Contributing to Development

and Spread of MDR and XDR TBand Spread of MDR and XDR TB Weak TB programs (DOTS) Weak TB programs (DOTS)

Low completioncure ratesLow completioncure rates Lack of treatment follow up and Lack of treatment follow up and

patient support patient support Unreliable drug supply Unreliable drug supply Diagnostic delay Diagnostic delay

Absent or inadequate infection control Absent or inadequate infection control measuresmeasures

Uncontrolled use of 2Uncontrolled use of 2ndnd line drugs line drugs

INHINH Chromosomally mediatedChromosomally mediated Loss of catalaseperoxidaseLoss of catalaseperoxidase Mutation in mycolic acid synthesisMutation in mycolic acid synthesis Regulators of peroxide responseRegulators of peroxide response

Mechanism of resistanceMechanism of resistance

RifampinRifampin Reduced binding to RNA polymeraseReduced binding to RNA polymerase

Clusters of mutations at ldquoRifampin Resistance Clusters of mutations at ldquoRifampin Resistance Determining Regionrdquo (RRDR)Determining Regionrdquo (RRDR)

Reduced Cell wall permeabilityReduced Cell wall permeability


Gene location associated Gene location associated Drug-Resistant Mtuberculosis Drug-Resistant Mtuberculosis

DrugDrug Gene Gene Isoniazid Isoniazid Kat G Inh A Kas A Kat G Inh A Kas A RifampicinRifampicin rpo B rpo B EthambutolEthambutol emb B emb B StreptomycinStreptomycin rps L rps L PyrazinamidePyrazinamide pnc A pnc A FluoroquinolonesFluoroquinolones gyr A gyr A

Dubaniewicz A et al Molecular sub-type of the HLA-DR antigens Dubaniewicz A et al Molecular sub-type of the HLA-DR antigens in pulmonary tuberculosis Int J Infect Dis20004129-33in pulmonary tuberculosis Int J Infect Dis20004129-33

Drug Susceptibility Drug Susceptibility TestingTesting

Susceptibility TestingSusceptibility Testing

1048708 1048708 Direct and indirect testingDirect and indirect testing 1048708 1048708 Primary Drugs testingPrimary Drugs testing 1048708 1048708 IsoniazidIsoniazid 1048708 1048708 RifampicinRifampicin 1048708 1048708 Ethambutol ()Ethambutol () 1048708 1048708 Pyrizinamide ()Pyrizinamide ()

Drug susceptibility testing (DST)Drug susceptibility testing (DST)

DST is recommended for all new cases for all DST is recommended for all new cases for all first line drugs with specimens taken before first line drugs with specimens taken before initiating treatmentinitiating treatment

Accuracy Accuracy is more important than is more important than speedspeed DST results should come from a small number DST results should come from a small number

of well-equipped experienced laboratories who of well-equipped experienced laboratories who participate and perform well in an participate and perform well in an international international DSTDST quality control scheme quality control scheme

The WHO Supranational Laboratory Quality The WHO Supranational Laboratory Quality Control Network offers the greatest global Control Network offers the greatest global coverage for thiscoverage for this

Drug susceptibility TestingDrug susceptibility Testing

Assessment of grwoth inhibition on solid Assessment of grwoth inhibition on solid media containing various dilutions of the media containing various dilutions of the drug in comparison with the test strainsdrug in comparison with the test strains

As the method depend observation of As the method depend observation of grwoth grwoth Results are not available until several Results are not available until several weeks after isolation of the organismweeks after isolation of the organism

Other accredited MethodsOther accredited Methods

Radiometric and Non radiometric methodsRadiometric and Non radiometric methods Nucleicacid technology ndash effective upto Nucleicacid technology ndash effective upto

95 in mutations to rifampicin resistance 95 in mutations to rifampicin resistance to gene rpoB geneto gene rpoB gene

Drug susceptibility testing Drug susceptibility testing (DST(DST))

As a minimum laboratories As a minimum laboratories supplying DST data should supplying DST data should correctly identify resistance to correctly identify resistance to isoniazid and rifampicin in over isoniazid and rifampicin in over 90 of quality control samples 90 of quality control samples in two out of the last three in two out of the last three quality control roundsquality control rounds

Detection of Rifampicin Drug Detection of Rifampicin Drug susceptibility testing (DST) is more susceptibility testing (DST) is more

importantimportant Early identification of mycobacterial growth as Early identification of mycobacterial growth as

M tuberculosis M tuberculosis complex and the identification of complex and the identification of rifampicin resistance should be the rifampicin resistance should be the first priority first priority as rifampicin resistance invalidates standard as rifampicin resistance invalidates standard 6 month short-course chemotherapy and is a 6 month short-course chemotherapy and is a useful marker in most countries for MDR-TBuseful marker in most countries for MDR-TB

Laboratories should aim to identify isolates as Laboratories should aim to identify isolates as M tuberculosis M tuberculosis complex and perform rifampicin complex and perform rifampicin resistance in resistance in 9090 of isolates within 1-2 working of isolates within 1-2 working days This is technologically feasible days This is technologically feasible

Drug susceptibility testingDrug susceptibility testing

For DST laboratories modern molecular For DST laboratories modern molecular techniques permit the successful techniques permit the successful identification of isoniazid resistance in at identification of isoniazid resistance in at least least 7575 of mycobacterial cultures within of mycobacterial cultures within 1-2 working days and are useful 1-2 working days and are useful preliminary screens for isoniazid preliminary screens for isoniazid resistance resistance

Secondary Drugs testing[lack of Secondary Drugs testing[lack of standardized methods]standardized methods]

Ofloxacin quinolonesOfloxacin quinolones EthionamideEthionamide

KanamycinKanamycin CapreomycinCapreomycin Ensure quality control and quality Ensure quality control and quality

assurance assurance

MODSMODS MMicroscopicicroscopic OObservation ofbservation of DDrug Susceptibility rug Susceptibility TTestingesting

MODS affordable Technically MODS affordable Technically Feasible Feasible

MODS arose during experiments conducted by MODS arose during experiments conducted by Luz Caviedes under the guidance of Professor Luz Caviedes under the guidance of Professor Robert GilmanRobert Gilman at Universidad Peruana at Universidad Peruana Cayetano Heredia in Lima Peru in the late Cayetano Heredia in Lima Peru in the late 1990s in which a colorimetric test for TB growth 1990s in which a colorimetric test for TB growth was being investigated The observation that was being investigated The observation that microcolonies could be seen under the microcolonies could be seen under the microscope long before a colour change microscope long before a colour change occurred prompted the development of MODS occurred prompted the development of MODS

Review Article in Indian Journal Review Article in Indian Journal of Medical Microbiologyof Medical Microbiology

Caviedes L Moore Caviedes L Moore DA Introducing DA Introducing mods A low-cost mods A low-cost low-tech tool for high-low-tech tool for high-performance performance detection of detection of tuberculosis and tuberculosis and multidrug resistant multidrug resistant tuberculosis Indian J tuberculosis Indian J Med Microbial Med Microbial 20072587-8 20072587-8

Observation of Grwoth in liquid Observation of Grwoth in liquid MediaMedia

MODS depends upon three key principles MODS depends upon three key principles (which have been known for decades) (1) (which have been known for decades) (1) Mycobacterium tuberculosisMycobacterium tuberculosis grows faster grows faster in liquid (broth) than on solid media (2) in in liquid (broth) than on solid media (2) in liquid cultures liquid cultures M tuberculosisM tuberculosis grows in a grows in a visually characteristic manner (tangles visually characteristic manner (tangles cording) which can be observed under the cording) which can be observed under the microscope long before the naked eye microscope long before the naked eye could visualize colonies on solid agarcould visualize colonies on solid agar

Least time required for detection Least time required for detection of MDRof MDR

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

MODS more streamlinedMODS more streamlined

Recently completed operational field Recently completed operational field studies have served to refine and studies have served to refine and streamline the methodology further and streamline the methodology further and importantly validate MODS as a test for TB importantly validate MODS as a test for TB detection and MDRTB detection directly detection and MDRTB detection directly from sputum from sputum

Inverted Microscope a minimal Inverted Microscope a minimal needneed

Characteristic ldquo Characteristic ldquo tangles ldquo of tangles ldquo of Mtuberculosis can Mtuberculosis can be visualised under be visualised under microscope long microscope long before colonies to before colonies to the naked eyethe naked eye

MODS for detection of MODS for detection of MDR - TBMDR - TB

The scientific observations have proved The scientific observations have proved that a single MODS culture of sputum that a single MODS culture of sputum

sample offers more rapid and sensitive sample offers more rapid and sensitive detection of tuberculosis and Multidrug-detection of tuberculosis and Multidrug-resistant tuberculosis than the existing resistant tuberculosis than the existing

gold standard methods usedgold standard methods used

Advantages of MODS methodology in MDR detection

bull All the chemical ingredients are available locally except few which can be acquired easily

bull Existing infrastructure in District and Teaching hospital can be adopted for implementation of MODS

bull Risk to technician handling the specimens is minimal there is no absolute need to obtain grade III safety cabinets

bull Technology transfer is easier all the new technical manpower can be trained easily

Performing MODS AssayPerforming MODS Assay

The MODS assay was performed as The MODS assay was performed as described in standard protocolsdescribed in standard protocols

Broth cultures were prepared in 24 well Broth cultures were prepared in 24 well tissue culture plates ( Becton Dickinson) tissue culture plates ( Becton Dickinson) each containing decontaminant 7H9 broth each containing decontaminant 7H9 broth (Becton Dickinson) oxalic acid albumin (Becton Dickinson) oxalic acid albumin dextrose and catlase (OADC) (Becton dextrose and catlase (OADC) (Becton Dickinson) and Dickinson) and Polymyxin Amphotericin B Polymyxin Amphotericin B Nalidixic acidtrimethoprim and azlocillinNalidixic acidtrimethoprim and azlocillin (PANTA) (PANTA)

MODS Assay ( Contd)MODS Assay ( Contd)

For each sample 12 wells were usedFor each sample 12 wells were used Four in control wells no drug was used Four in control wells no drug was used

and each of the remaining 8 wells and each of the remaining 8 wells contained one of the four drugs at one of contained one of the four drugs at one of the two concentrations testedthe two concentrations tested

The cultures were examined under an inverted The cultures were examined under an inverted light microscope at magnification of 40x every light microscope at magnification of 40x every day ( except weekends ) from 4 to day 15 on day ( except weekends ) from 4 to day 15 on alternative days from 16 today 25 and twice alternative days from 16 today 25 and twice weekly from 26 to 40day weekly from 26 to 40day

Sample layout on MODS plate Sample layout on MODS plate (2 samples per plate)(2 samples per plate)

No plate contained 2 samples from the same No plate contained 2 samples from the same patientpatient

Drug susceptibility TestingDrug susceptibility TestingIn MODSIn MODS

Drug susceptibility testing was performed Drug susceptibility testing was performed with the use of MODS assay with the use of MODS assay

Growth in the drug free control wells but Growth in the drug free control wells but not in drug containing wells indicates not in drug containing wells indicates susceptibilitysusceptibility

The drug concentration were as follows The drug concentration were as follows Isoniazid 01 and 04 microgmilliliterIsoniazid 01 and 04 microgmilliliter Rifampicin 1 and 2 microg per millilitreRifampicin 1 and 2 microg per millilitre

Differentiation from Typical and Differentiation from Typical and Atypical MycobacteriumAtypical Mycobacterium

Nontuberculous Nontuberculous mycobacteria (NTM) mycobacteria (NTM) were recognized by were recognized by their lack of cording or their lack of cording or (in the case of (in the case of Mycobacterium Mycobacterium chelonae that uniquely chelonae that uniquely among NTM does among NTM does form cords) rapid form cords) rapid overgrowth of wells by overgrowth of wells by day 5day 5

In MODS growth is In MODS growth is identified by cording on identified by cording on

MicroscopyMicroscopy

MODS assay ( Contd)MODS assay ( Contd) To minimize cross To minimize cross

contamination and contamination and occupational occupational exposure plates exposure plates were permanently were permanently sealed inside plastic sealed inside plastic zip lock bags after zip lock bags after inoculation and were inoculation and were subsequently subsequently examined with in the examined with in the bagbag

Observation of growth in Observation of growth in MODSMODS

Positive cultures Positive cultures were identified by were identified by cord formation cord formation characteristic of characteristic of Mtuberculosis Mtuberculosis grwothin liquid grwothin liquid medium in drug medium in drug free control wellsfree control wells

MODS in Atypical MODS in Atypical MycobacteriumMycobacterium

Non tuberculous Non tuberculous mycobacterium mycobacterium were recognised by were recognised by their lack of cording their lack of cording or for Mchelonae ( or for Mchelonae ( which forms cords) which forms cords) by rapid by rapid overgrwoth by day overgrwoth by day 55

Contamination in MODS Contamination in MODS AssayAssay

Fungal or bacterial Fungal or bacterial contamination was contamination was recognised by rapid recognised by rapid overgrowth and overgrowth and clouding in wellsclouding in wells

If contamination was If contamination was detected the original detected the original samples was cultured samples was cultured again after being again after being decontaminated once decontaminated once moremore

Honduras study comparing the Honduras study comparing the LJ mediumLJ medium

Per specimen there was concordance Per specimen there was concordance between MODS and LJ culture in 942 between MODS and LJ culture in 942 MODS tests were also less prone to MODS tests were also less prone to contamination than LJ cultures 62 [38] contamination than LJ cultures 62 [38] vs (95 [58] of 1639 samples vs (95 [58] of 1639 samples respectively (P le001) respectively (P le001)

PCR Molecular susceptibility PCR Molecular susceptibility testingtesting

RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay

Confirming Confirming MODSMODS results results

Spacer Spacer Oligonucleotide typing Oligonucleotide typing SpoligotypingSpoligotyping polymerase chain polymerase chain reaction with multiple reaction with multiple primers or both were primers or both were applied to all isolates applied to all isolates from each of the three from each of the three types of cultures in types of cultures in order to confirm the order to confirm the presence of presence of MtuberculosisMtuberculosis

MODS and MDR detection

bull The drug sensitivity for Rifampicin and Isoniazid can be tested and established the presence of MDR

bull In view of being chronic disease it is highly essential to establish MDR Tuberculosis at centers serving DOTS under WHO guidelines

bull Starting and establishing centers to identify MDR at every district and Teaching Medical centers leads to better control of Tuberculosis

Why MODS is a better Why MODS is a better method for MDR TB method for MDR TB

detectiondetection If a MODS culture was negative on day If a MODS culture was negative on day

15there is 997 chance that the 15there is 997 chance that the sample is truly culture negativesample is truly culture negative

The negative MODS cultures can be The negative MODS cultures can be discarded after 3 weeksdiscarded after 3 weeks

Biosafety concerns in MODS Biosafety concerns in MODS technologytechnology

Legitimate concerns about biosafety with Legitimate concerns about biosafety with other liquid culture systems do not really other liquid culture systems do not really apply to MODS indeed the converse is apply to MODS indeed the converse is the case After inoculation with the case After inoculation with decontaminated sample the MODS plates decontaminated sample the MODS plates are permanently sealed in ziplock are permanently sealed in ziplock polythene bags through which the polythene bags through which the microscopic examination is made thus microscopic examination is made thus spillage of the mycobacterial soup spillage of the mycobacterial soup cannot occur cannot occur

Lower Grade Biosafety is Lower Grade Biosafety is adequateadequate

N 95 mask protects from BiohazardN 95 mask protects from Biohazard

No transfer of Materials needed No transfer of Materials needed in MODSin MODS

As no secondary sub-culture is needed As no secondary sub-culture is needed (because this is direct and not indirect (because this is direct and not indirect susceptibility testing) no further susceptibility testing) no further manipulation is required - this zero manipulation is required - this zero potential for aerosolisation or accident potential for aerosolisation or accident compares favourably with the hazard compares favourably with the hazard associated with preparation of a associated with preparation of a standardized inoculum for indirect DST standardized inoculum for indirect DST

Computer pattern Computer pattern recognitionrecognition

of of Mycobacterium Mycobacterium tuberculosistuberculosis

in MODS culturein MODS culture

Automation in MODSAutomation in MODS

M tuberculosis in MODS x10 objective (sputum sample inoculation)

Day 6

Day 16 Day 17

Day 7 Day 8 Day 9 Day 10 Day 11 Day 12 Day 13 Day 14 Day 15

MODS can be used in Extra pulmonary MODS can be used in Extra pulmonary TuberculosisTuberculosis

Draw backs of MODSDraw backs of MODS

One possible drawback however could be One possible drawback however could be the inability of the laboratory technicians to the inability of the laboratory technicians to distinguish between TB and some NTM distinguish between TB and some NTM This could potentially have clinical impact This could potentially have clinical impact in settings where NTM prevalence is high in settings where NTM prevalence is high and not all mycobacteria respond to anti-and not all mycobacteria respond to anti-TB treatment TB treatment

Other WHO-Endorsed ToolsOther WHO-Endorsed Tools

Liquid culture (eg MGIT BacTALERT)Liquid culture (eg MGIT BacTALERT)

Capilia TBCapilia TB Rapid strip test that detects a TB-specific Rapid strip test that detects a TB-specific

antigen from cultureantigen from culture

Molecular line probe assays (eg Molecular line probe assays (eg GenoType MTBDRplus INNO-LiPA RifTB)GenoType MTBDRplus INNO-LiPA RifTB) Strip test for detection of TB and drug-Strip test for detection of TB and drug-

resistance conferring mutationsresistance conferring mutations

WHO Controls the Tuberculosis WHO Controls the Tuberculosis related workrelated work

The laboratory methods for anti-The laboratory methods for anti-tuberculosis drug susceptibility testing tuberculosis drug susceptibility testing should be selected from among those that should be selected from among those that are WHO-recommended and all are WHO-recommended and all laboratory processes should be quality-laboratory processes should be quality-assured in cooperation with a partner assured in cooperation with a partner Supranational Reference Laboratory Supranational Reference Laboratory (SRL)(SRL)

XDR-TB in South AfricaXDR-TB in South AfricaAugust 2006August 2006

53 of 544 patients defined as XDR-TB 53 of 544 patients defined as XDR-TB casescases

bull bull 52 of the 53 patients died on average 52 of the 53 patients died on average within 25within 25

days including those on antiretroviral days including those on antiretroviral therapytherapy

bull bull Further investigations being carried outFurther investigations being carried out bull bull XDR-TB likely in bordering African XDR-TB likely in bordering African

countriescountries

Molecular FingerprintingMolecular Fingerprinting

26 of 30 (87) XDR TB isolates found to 26 of 30 (87) XDR TB isolates found to be genetically similarbe genetically similar

Majority of patients had no previous history Majority of patients had no previous history of TB treatment Suggestive of recent of TB treatment Suggestive of recent infection with drug-resistant straininfection with drug-resistant strain

Additional cases identified in 28 of the 68 Additional cases identified in 28 of the 68 hospitals in KZN KwaZulu Natalhospitals in KZN KwaZulu Natal

CDC Updates Guidelines for Nucleic AcidCDC Updates Guidelines for Nucleic AcidAmplification Techniques to Diagnose Amplification Techniques to Diagnose

TuberculosisTuberculosis NAAT results should be interpreted in NAAT results should be interpreted in

conjunction with the AFB smear conjunction with the AFB smear resultsresults

NAAT and smear positive start Rx NAAT and smear positive start Rx despite pending culture results PPV despite pending culture results PPV 9595

Smear negative NAAT positive use Smear negative NAAT positive use clinical judgment to either treat or await clinical judgment to either treat or await cultureculture

Selection from automated systems for Selection from automated systems for molecular andmolecular and

bacteriological rapid diagnosticsbacteriological rapid diagnostics

PCRPCR RocheCOBASreg Amplicorreg RocheCOBASreg Amplicorreg

amplification kitsamplification kits RocheCOBASreg LightCyclerreg (real-RocheCOBASreg LightCyclerreg (real-

time-PCR)time-PCR) RocheCOBASreg TaqMan 48regRocheCOBASreg TaqMan 48reg (increases the specificity of real-time-(increases the specificity of real-time-

PCR)PCR)

Microscopy and Culturing still a Microscopy and Culturing still a top prioritytop priority

Is PCR methods a solution Is PCR methods a solution

PCR cant yet PCR cant yet replace neither replace neither microscopy microscopy culturing and culturing and competent clinical competent clinical examinationexamination

No testing method replaces No testing method replaces clinical assessmentclinical assessment

Extreme Drug resistant Extreme Drug resistant Tuberculosis (XDR-TB)Tuberculosis (XDR-TB)

Resistant to all first line drugs namely Resistant to all first line drugs namely Isoniazid Isoniazid and Rifampin and Rifampin and and

Three or more Three or more second line drugs (SLDrsquoS) that are second line drugs (SLDrsquoS) that are used to treat MDR-TBused to treat MDR-TB Thequinalones like OfloaxinThequinalones like Ofloaxin

OrOr Aminoglygocides like Capreomycin amp Kanamycin Aminoglygocides like Capreomycin amp Kanamycin

No third-line drugs available to treat XDR-TB No third-line drugs available to treat XDR-TB since none since none has been developed in the last 40 has been developed in the last 40 yearsyears


XDR-TB 82306XDR-TB 82306

ldquo ldquoRapidly Fatal in South Africardquo Rapidly Fatal in South Africardquo Tugela Tugela Ferry KwaZulu-NatalFerry KwaZulu-Natal

10 isolates resistant to ALL 110 isolates resistant to ALL 1stst and and 22ndnd line agents line agents

5152 XDR dead in median 16 days 5152 XDR dead in median 16 days after first positive sputumafter first positive sputum

67 AIDS deaths w MDR TB67 AIDS deaths w MDR TB

Czech Republic

The b

oundarie

s and n

am

es sh

ow

n a

nd th

e d

esig

natio

ns u

sed o

n th

is map d

o n

ot im

ply

the e

xpre

ssion o

f any o

pin

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whatso

ever o

n th

e p

art o

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rnin

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gal sta

tus o

f any co

untry

territo

ry city

or a

rea o

r of its a

uth

oritie

s or co

nce

rnin

g th

e d

elim

itatio

n o

f its frontie

rs or b

oundarie

s Dotte

d lin

es o

n m

aps re

pre

sent a

ppro

xim

ate

bord

er lin

es fo

r w

hich

there

may n

ot y

et b

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ll agre

em

ent

WH

O 2

005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile

Based on information provided to WHO Stop TB Department 13 September 2007

SpainIslamic Republic of Iran

China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam

Countries with confirmed XDR-TB cases as of September 2007

Summary Summary Drug resistant TBDrug resistant TB

bull Drug-resistant TB poses a grave public health threat Drug-resistant TB poses a grave public health threat especially in high HIV prevalence settingsespecially in high HIV prevalence settings

bull XDR-TB strains have been found in all regions of the world XDR-TB strains have been found in all regions of the world

bull XDR-TB occurs as a result inadequate TB control XDR-TB occurs as a result inadequate TB control programmesprogrammes

bull XDR-TB if identified early canXDR-TB if identified early can be treated and cured but be treated and cured but experience limited to low HIV prevalence settings experience limited to low HIV prevalence settings

bull Infection control measures must be strengthened Infection control measures must be strengthened

bull XDR-TB underlines the need for investment in basic TB XDR-TB underlines the need for investment in basic TB control plus development of new TB diagnostics control plus development of new TB diagnostics treatments and vaccinestreatments and vaccines

Health Care Workers and MDR Health Care Workers and MDR TBTB

Recognised risk for health care workersRecognised risk for health care workers Risk assessmentRisk assessment High risk ndash Prolonged closed contact with High risk ndash Prolonged closed contact with

infectious patientsinfectious patients Smear positive MDR TB patientsSmear positive MDR TB patients Medium risk ndashPrimary health care centres Medium risk ndashPrimary health care centres

involvedinvolved Sputum collection on TB suspectsSputum collection on TB suspects Low risk ndashHealth care support staff eg Low risk ndashHealth care support staff eg

cleanerscleaners Porters and admin staffPorters and admin staff


Koch failed to conquer tuberculosis which still Koch failed to conquer tuberculosis which still causes enormous health problems worldwide causes enormous health problems worldwide

100 years after his Nobel award100 years after his Nobel award The scientific academies The scientific academies

noted that the triumphant noted that the triumphant discovery of 1882 was discovery of 1882 was followed by a succession followed by a succession of failures first of all the of failures first of all the failed attempt to present failed attempt to present tuberculin as a remedy tuberculin as a remedy against tuberculosis in against tuberculosis in 1890-91 which severely 1890-91 which severely damaged Kochs damaged Kochs reputationreputation

Medical History 2001 45 1-32 Medical History 2001 45 1-32 CHRISTOPH GRADMANNCHRISTOPH GRADMANN

Are there any solutions for Are there any solutions for effective Diagnosis in TB effective Diagnosis in TB

Many more powerful hands needed Many more powerful hands needed to Control Tuberculosisto Control Tuberculosis

Contribute your Knowledge Wisdom Contribute your Knowledge Wisdom to prevent spread and control of to prevent spread and control of


Created by DrTVRao MD for Created by DrTVRao MD for lsquoersquo learninglsquoersquo learning Programme Programme

EmailEmail

doctortvraogmailcomdoctortvraogmailcom

MDR TB ndashGreat Human ConcernMDR TB ndashGreat Human Concern
















recoveredrecovered
















MDR-TBMDR-TB




























Global Estimates




DeathsDeaths



MDR TBMDR TB 424000424000 116000116000

XDR TBXDR TB 2700027000 1600016000




Africa

0

100

200

300

400

500

600

1990 1995 2000 2005 2010 2015

Inci

den

ce p

er 1

000

00 p

er y

ear

World

Cent Euro

East Europe

Est Market

East Medit

Lat America

West Pacific

Sth East Asia

AFR high HIV

AFR low HIV


MDR-TB

1990sect

XDR-TB

2006sect

Total DR











LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















PCR)PCR)
















Czech Republic

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ent

WH

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005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam
























EmailEmail

















recoveredrecovered
















MDR-TBMDR-TB




























Global Estimates




DeathsDeaths



MDR TBMDR TB 424000424000 116000116000

XDR TBXDR TB 2700027000 1600016000




Africa

0

100

200

300

400

500

600

1990 1995 2000 2005 2010 2015

Inci

den

ce p

er 1

000

00 p

er y

ear

World

Cent Euro

East Europe

Est Market

East Medit

Lat America

West Pacific

Sth East Asia

AFR high HIV

AFR low HIV


MDR-TB

1990sect

XDR-TB

2006sect

Total DR











LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















PCR)PCR)
















Czech Republic

The b

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WH

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005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam
























EmailEmail
















MDR-TBMDR-TB




























Global Estimates




DeathsDeaths



MDR TBMDR TB 424000424000 116000116000

XDR TBXDR TB 2700027000 1600016000




Africa

0

100

200

300

400

500

600

1990 1995 2000 2005 2010 2015

Inci

den

ce p

er 1

000

00 p

er y

ear

World

Cent Euro

East Europe

Est Market

East Medit

Lat America

West Pacific

Sth East Asia

AFR high HIV

AFR low HIV


MDR-TB

1990sect

XDR-TB

2006sect

Total DR











LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















PCR)PCR)
















Czech Republic

The b

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005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam
























EmailEmail





























Global Estimates




DeathsDeaths



MDR TBMDR TB 424000424000 116000116000

XDR TBXDR TB 2700027000 1600016000




Africa

0

100

200

300

400

500

600

1990 1995 2000 2005 2010 2015

Inci

den

ce p

er 1

000

00 p

er y

ear

World

Cent Euro

East Europe

Est Market

East Medit

Lat America

West Pacific

Sth East Asia

AFR high HIV

AFR low HIV


MDR-TB

1990sect

XDR-TB

2006sect

Total DR











LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















PCR)PCR)
















Czech Republic

The b

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WH

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005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

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Israel

Azerbaijan

Poland

Slovenia

India

Australia

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Vietnam
























EmailEmail





Africa

0

100

200

300

400

500

600

1990 1995 2000 2005 2010 2015

Inci

den

ce p

er 1

000

00 p

er y

ear

World

Cent Euro

East Europe

Est Market

East Medit

Lat America

West Pacific

Sth East Asia

AFR high HIV

AFR low HIV


MDR-TB

1990sect

XDR-TB

2006sect

Total DR











LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















PCR)PCR)
















Czech Republic

The b

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005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam
























EmailEmail



MDR-TB

1990sect

XDR-TB

2006sect

Total DR











LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















PCR)PCR)
















Czech Republic

The b

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005 A

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Ecuador

Georgia

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Germany

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Armenia

Russian Federation

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Mexico

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USA

Brazil

UK

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China Hong Kong SAR

France

Japan

NorwayCanada

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Lithuania

Ireland

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Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam
























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LatviaLatvia 9090



RussiaRussia 6060

IndiaIndia 3434

IranIran 5858










Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















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Czech Republic

The b

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Ecuador

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Germany

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Mexico

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USA

Brazil

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Chile



China Hong Kong SAR

France

Japan

NorwayCanada

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Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

Azerbaijan

Poland

Slovenia

India

Australia

Mozambique

Vietnam
























EmailEmail








Wild M TB strain
















































assurance assurance




















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















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Czech Republic

The b

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Ecuador

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Germany

Republic of Korea

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Russian Federation

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Mexico

Peru

USA

Brazil

UK

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Chile



China Hong Kong SAR

France

Japan

NorwayCanada

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Ireland

Romania

Israel

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Poland

Slovenia

India

Australia

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Vietnam
























EmailEmail










































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RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















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Czech Republic

The b

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005 A

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Ecuador

Georgia

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Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

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Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

Romania

Israel

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Poland

Slovenia

India

Australia

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Vietnam
























EmailEmail





















































RMP resistance

INH resistance

HainGenotypeMTBDR

INNO-LiPARifTBassay






















Day 6

Day 16 Day 17


















countriescountries















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Czech Republic

The b

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005 A

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Ecuador

Georgia

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Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

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Portugal

Latvia

Mexico

Peru

USA

Brazil

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Sweden

Thailand

Chile



China Hong Kong SAR

France

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Lithuania

Ireland

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Poland

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India

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Day 6

Day 16 Day 17


















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Czech Republic

The b

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serv

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Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

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Netherlands

Estonia

Lithuania

Ireland

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Israel

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Poland

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serv

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Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

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China Hong Kong SAR

France

Japan

NorwayCanada

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Czech Republic

The b

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Argentina

Bangladesh

Germany

Republic of Korea

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Russian Federation

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Mexico

Peru

USA

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UK

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Czech Republic

The b

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ll agre

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ent

WH

O 2

005 A

ll rights re

serv

ed

Ecuador

Georgia

Argentina

Bangladesh

Germany

Republic of Korea

Armenia

Russian Federation

South Africa

Portugal

Latvia

Mexico

Peru

USA

Brazil

UK

Sweden

Thailand

Chile



China Hong Kong SAR

France

Japan

NorwayCanada

Italy

Netherlands

Estonia

Lithuania

Ireland

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Israel

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Poland

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Drugresistanttuberculosis 100227020029 Phpapp01

Health & Medicine

Transcript of Drugresistanttuberculosis 100227020029 Phpapp01