Diagnosis of Tb

Dr.T.V.Rao

DIAGNOSIS OF DIAGNOSIS OF TUBERCULOSISTUBERCULOSIS

Emerging TrendsEmerging Trends

Dr.T.V.Rao, MD.Dr.T.V.Rao, MD.

Dr.T.V.RaoDr.T.V.Rao

Robert Koch Discovers Robert Koch Discovers MycobacteriumMycobacterium

Dr.T.V.Rao

A Global EmergencyA Global Emergency

The Tuberculosis in the The Tuberculosis in the beginning of the 21beginning of the 21stst Century Century declared as Global Emergency declared as Global Emergency

(WHO)(WHO)


Why Tuberculosis is a Important Why Tuberculosis is a Important Disease.Disease.

Tuberculosis continues to be a Tuberculosis continues to be a Important communicable disease.Important communicable disease.

A leading cause of morbidity and A leading cause of morbidity and mortality in Developing world. mortality in Developing world.

Most Important communicable Most Important communicable disease in Bangladesh, China, disease in Bangladesh, China, Indonesia, Africa, and Pakistan.Indonesia, Africa, and Pakistan.

But it is Curable DiseaseBut it is Curable Disease


Tuberculosis is a Global ProblemTuberculosis is a Global Problem


Tuberculosis - Important Tuberculosis - Important communicable disease spread by communicable disease spread by

Respiratory routeRespiratory route


Why Everybody Concerned.Why Everybody Concerned.

Tuberculosis kills young adults.Tuberculosis kills young adults. Premature death of the infected a Premature death of the infected a

prominent future.prominent future. Today many are co infected with HIV.Today many are co infected with HIV. The open cases of Tuberculosis infects a The open cases of Tuberculosis infects a

few around his/her environment.few around his/her environment. A social burden to the family, society and A social burden to the family, society and

Nations.Nations.


Tuberculosis in the era ofTuberculosis in the era ofHIV / AIDS.HIV / AIDS.

HIV / AIDS epidemic led to large HIV / AIDS epidemic led to large increase of Smear negative pulmonary increase of Smear negative pulmonary

tuberculosis which in turn has led to tuberculosis which in turn has led to poor treatment out comes, and early poor treatment out comes, and early

mortalitymortality

Frequently involves Lower lobes of Frequently involves Lower lobes of Lungs.Lungs.


Why we fail to Diagnose Why we fail to Diagnose Tuberculosis.Tuberculosis.

Lack of health infrastructure.Lack of health infrastructure. Control is plagued with lack ofControl is plagued with lack of

Accurate,Accurate,

Robust,Robust,

and Rapidand Rapid

Diagnostic methods,Diagnostic methods,

Technologies.Technologies.


Why we failed ( Cont )Why we failed ( Cont )

Diagnostic services are poor, and so we Diagnostic services are poor, and so we failed at Individual and community failed at Individual and community levels.levels.

Patients are diagnosed late.Patients are diagnosed late. Many patients are never diagnosed Many patients are never diagnosed

before death.before death.

Early deaths are burden to Social Early deaths are burden to Social Infrastructure and Economic loss. Infrastructure and Economic loss.


Importance of Clinical servicesImportance of Clinical services

Early diagnosis rests with clinicians, Early diagnosis rests with clinicians, whose contribution is immense in whose contribution is immense in

prompt treatment.prompt treatment.

A clinicians knowledge, proper A clinicians knowledge, proper documentation are immense help in documentation are immense help in

Developing countries.Developing countries.


When to suspect TuberculosisWhen to suspect Tuberculosis

Cough longer than 3 weeks.Cough longer than 3 weeks. Fever for 1 month, or both.Fever for 1 month, or both. Blood stained sputum.Blood stained sputum. Night sweats, weight lossNight sweats, weight loss Age between 14 and 70 years Age between 14 and 70 years

( ( Correlates National Tuberculosis Correlates National Tuberculosis Programme ).Programme ).

Dr.T.V.Rao

DIAGNOSTIC METHODSDIAGNOSTIC METHODS


Diagnosis.Diagnosis.

Tuberculosis is a diversified disease.Tuberculosis is a diversified disease. Any organs can be involved.Any organs can be involved. Any age group, gender no bar for Any age group, gender no bar for

Tuberculosis.Tuberculosis. Involvement of Lungs contribute to Involvement of Lungs contribute to

majority of tuberculosis.majority of tuberculosis. And involvement of Lungs is designated as And involvement of Lungs is designated as

Pulmonary tuberculosis.Pulmonary tuberculosis.


Diagnosis of Pulmonary Diagnosis of Pulmonary TuberculosisTuberculosis

Majority of Adults suffer with pulmonary Majority of Adults suffer with pulmonary tuberculosis.tuberculosis.

Microbiological examination of Sputum Microbiological examination of Sputum continues to be a Gold standard in proving continues to be a Gold standard in proving the Diagnosis.the Diagnosis.

Sputum examination in Children is not Sputum examination in Children is not sensitive in Diagnosis.sensitive in Diagnosis.

Radiological examination of Lungs, most Radiological examination of Lungs, most commonly prescribed investigation.commonly prescribed investigation.


X - ray examination of chest most X - ray examination of chest most easily available Investigation.easily available Investigation.

Dr.T.V.Rao

Microbiological Microbiological Investigations are essential Investigations are essential for definitive Diagnosis of for definitive Diagnosis of

Tuberculosis.Tuberculosis.


Importance of Optimal SpecimensImportance of Optimal Specimens

Pulmonary Tuberculosis is the Pulmonary Tuberculosis is the commonest presentation of Tuberculosiscommonest presentation of Tuberculosis

Sputum is the Most important specimen Sputum is the Most important specimen for identification and isolation of Acid for identification and isolation of Acid fast bacilli.fast bacilli.

The developing countries suffers the most The developing countries suffers the most important step in getting an ideal important step in getting an ideal sample.sample.

Dr.T.V.Rao

Sputum specimensSputum specimens

Train the staff to obtain the appropriate Train the staff to obtain the appropriate specimenspecimen

A few minutes of education to patients on A few minutes of education to patients on importance of ideal sample make a great importance of ideal sample make a great difference and improves the Diagnosis.difference and improves the Diagnosis.


Observe to identify Sputum from Observe to identify Sputum from Saliva.Saliva.

SPUTUMSPUTUM

Specimens appear mucoid Specimens appear mucoid even, blood stained. even, blood stained.

Contains many Contains many Polymorphoneutrophils.Polymorphoneutrophils.

SALIVASALIVA

Appears clear, watery, Appears clear, watery, and frothy.and frothy.

Contains many Contains many squamous epithelial squamous epithelial cellscells

Absence of Absence of Polymorphoneutrophils.Polymorphoneutrophils.


Role of Microscopy in Tuberculosis.Role of Microscopy in Tuberculosis.

Microscopy for Diagnosis of Tuberculosis is Microscopy for Diagnosis of Tuberculosis is initiated in 1880initiated in 1880

The conceptions have not changed since The conceptions have not changed since then.then.

Best efforts should be put to obtain Best efforts should be put to obtain sputum,sputum,

Processing of saliva loses all valuable Processing of saliva loses all valuable clues to diagnose.clues to diagnose.


Microscopy and TuberculosisMicroscopy and Tuberculosis

Microscopy with Microscopy with Ziehl – Neelsen’s Ziehl – Neelsen’s stainingstaining

A century old A century old

procedureprocedure


Why MicroscopyWhy Microscopy

Only we need Microscope, and few Only we need Microscope, and few stains.stains.

Most rapid, economical,Most rapid, economical, Can detect bacterial load.Can detect bacterial load. A Diagnostic, and Prognostic tool.A Diagnostic, and Prognostic tool. A little of sputum 0.2 µl is adequate.A little of sputum 0.2 µl is adequate. A prompt diagnosis after searching A prompt diagnosis after searching

as few as 100 fields.as few as 100 fields.


Limitation of Microscopy for Limitation of Microscopy for Tuberculosis.Tuberculosis.

Repeated sample examinations. load on Repeated sample examinations. load on technical staff.technical staff.

Training and dedication of Microscopist.Training and dedication of Microscopist. The load of bacilli must be more than The load of bacilli must be more than

10,000 / 1 ml of sputum.10,000 / 1 ml of sputum. Low in sensitivity < 50 %Low in sensitivity < 50 % Repeated requests for samplesRepeated requests for samples High drop out by patients, for repeated High drop out by patients, for repeated

samples.samples. Not dependable in pediatric age group.Not dependable in pediatric age group.


Smear showing Acid Fast Bacilli.Smear showing Acid Fast Bacilli.


What is Smear PositivityWhat is Smear PositivityWHOWHO

All patients who have submitted All patients who have submitted twotwo

Specimens and found to be Specimens and found to be positivepositive

for identification of AFBfor identification of AFB


Processing Direct smear negative Processing Direct smear negative specimensspecimens

Sputum Microscopy can be improved with Sputum Microscopy can be improved with Sputum liquefaction, concentration and Sputum liquefaction, concentration and gravity sedimentation.gravity sedimentation.

Popular solvents Popular solvents Sodium hypochlorite.Sodium hypochlorite. Sodium hydroxide.Sodium hydroxide. Ammonium sulphateAmmonium sulphate N-acetyl-L-cysteine –sodium N-acetyl-L-cysteine –sodium

hydroxide.hydroxide.


Benefits of Liquefaction and Benefits of Liquefaction and ConcentrationConcentration

Major studies showed processing of Major studies showed processing of sputum with chemicals and centrifugation sputum with chemicals and centrifugation improved sensitivity up to 18 %.improved sensitivity up to 18 %.

Incremental yield ( positive with bleach Incremental yield ( positive with bleach minus positives with Ziehl – Neelsen stain) minus positives with Ziehl – Neelsen stain) up to 9 %.up to 9 %.

Treating specimens with Sodium Treating specimens with Sodium hypochlorite is Mycobactericidal and also hypochlorite is Mycobactericidal and also kills HIV and improves the safety and kills HIV and improves the safety and acceptability by technical staff.acceptability by technical staff.


When Microscopy failsWhen Microscopy fails

Smear negative tuberculosis.Smear negative tuberculosis. In HIV infected patients, on many In HIV infected patients, on many

occasions prove negative. in spite of occasions prove negative. in spite of presence of bacilli, ( as few bacilli are presence of bacilli, ( as few bacilli are expectorated).expectorated).

Needs concentration and liquefaction Needs concentration and liquefaction with chemicals.with chemicals.

Time consuming, needs more Time consuming, needs more technical manpowertechnical manpower


Growing role ofGrowing role ofFluorescent MicroscopyFluorescent Microscopy

There is a growing need for screening for There is a growing need for screening for AFB by Florescent Microscopy.AFB by Florescent Microscopy.

Several studies prove, Florescent Several studies prove, Florescent Microscopy in Diagnosis of Tuberculosis is Microscopy in Diagnosis of Tuberculosis is a priority,a priority,

Developing world should opt and initiate Developing world should opt and initiate florescent microscopy.florescent microscopy.


Acid Fast Bacilli as seen under Acid Fast Bacilli as seen under Fluorescent MicroscopeFluorescent Microscope


Why we need Florescent Why we need Florescent MicroscopyMicroscopy

Useful when few bacilli are present.Useful when few bacilli are present. Increases the sensitivity in HIV patients with Increases the sensitivity in HIV patients with

tuberculosis.tuberculosis. Reduces the time needed for testing.Reduces the time needed for testing. About 15 times as many fields of view can be About 15 times as many fields of view can be

scanned by fluorescent microscopy than by Ziehl scanned by fluorescent microscopy than by Ziehl – Neelsen’method in the same period.– Neelsen’method in the same period.

Increases the sensitivity by 10 %Increases the sensitivity by 10 % Better conclusions with one or two specimens, Better conclusions with one or two specimens,

unlike Ziehl Neelsen’s method needing 3 or > 3 unlike Ziehl Neelsen’s method needing 3 or > 3 specimens.specimens.


Culturing MycobacteriumCulturing Mycobacterium

Culturing for isolation of Culturing for isolation of Mycobacterium spp continues to be a Mycobacterium spp continues to be a Gold standard, particularly in Gold standard, particularly in Developing countries.Developing countries.

Need only 10 – 100 bacilli / 1 ml of Need only 10 – 100 bacilli / 1 ml of sputum.sputum.


Culturing Culturing Most useful inMost useful in

Surveillance,Surveillance, Drug sensitivity testing patterns.Drug sensitivity testing patterns. Identify treatment failures.Identify treatment failures. Useful in Patients presenting with Useful in Patients presenting with

respiratory symptoms, X- ray’s respiratory symptoms, X- ray’s suggestive, but smear negative. Can prove suggestive, but smear negative. Can prove culture positive.culture positive.

Cultures remain suggestive and helpful in Cultures remain suggestive and helpful in early treatment periods, failed drug early treatment periods, failed drug regimes.regimes.


Methods of Culturing.Methods of Culturing.

Culturing on Lowenstein Jenson’s Culturing on Lowenstein Jenson’s culture medium remain the culture medium remain the affordable ,economical method in affordable ,economical method in developing world.developing world.


Limitation in CulturingLimitation in Culturing

Mycobacterium spp are slow Mycobacterium spp are slow growing.growing.

Need 6 – 8 weeks for growing.Need 6 – 8 weeks for growing. Specimens can be contaminated Specimens can be contaminated

while growing, needs repeated while growing, needs repeated specimens, in turn patients loose specimens, in turn patients loose confidence in Laboratories.confidence in Laboratories.


Recent facts on CulturingRecent facts on Culturing

Useful in HIV infected patients with Useful in HIV infected patients with Tuberculosis.Tuberculosis.

As even few bacilli can be grown in As even few bacilli can be grown in spite of smear negativity.spite of smear negativity.

But the specimens to be incubated But the specimens to be incubated for longer time as few bacilli are for longer time as few bacilli are present.present.


Pitfalls in CulturingPitfalls in Culturing

Specificity is lost due to Specificity is lost due to contamination.contamination.

Can yield false positive results in 1 – Can yield false positive results in 1 – 4 % of the cases.4 % of the cases.

Cultures may be negative in spite of Cultures may be negative in spite of x rays are suggestive of tuberculosis.x rays are suggestive of tuberculosis.


Growth of Acid fast bacilli on Growth of Acid fast bacilli on L J Medium.L J Medium.

Dr.T.V.Rao

ADVANCES IN ADVANCES IN CULTURING CULTURING

TECHNIQUES.TECHNIQUES.There are emerging Modern Media There are emerging Modern Media

with accurate detection, are with accurate detection, are replacing the Egg and Agar based replacing the Egg and Agar based

medium.medium.


Emerging methods in CulturingEmerging methods in Culturing

MGIT – Mycobacterium growth MGIT – Mycobacterium growth incubator tube method.incubator tube method.

Growth occurs in shorter than egg Growth occurs in shorter than egg medium.medium.

Usefulness in HIV patients established.Usefulness in HIV patients established. Contamination is lessContamination is less But expensive to people in Developing But expensive to people in Developing

world.world.


Blood culturing for MycobacteriumBlood culturing for Mycobacterium

Useful in HIV patients, and children.Useful in HIV patients, and children. Effective in isolation of Atypical Effective in isolation of Atypical

mycobacterium.mycobacterium. But not cost effective.But not cost effective. May be important tool in future for May be important tool in future for

diagnosing Tuberculosis in HIV diagnosing Tuberculosis in HIV infected.infected.

Dr.T.V.Rao

Molecular Methods in Molecular Methods in Diagnosis of TuberculosisDiagnosis of Tuberculosis

Several methods are Several methods are available, mainly used as available, mainly used as

Research toolsResearch tools


Real Time PCR replacing older Real Time PCR replacing older MethodsMethods


PCR How useful to our Patients?PCR How useful to our Patients?

PCR ( Polymerase chain reaction ) used by PCR ( Polymerase chain reaction ) used by several investigators.several investigators.

However most cases can be diagnosed However most cases can be diagnosed with simple methods if effectively used.with simple methods if effectively used.

The definite role of PCR continues to be The definite role of PCR continues to be controversial controversial

Above all not cost effective to Developing Above all not cost effective to Developing countries.countries.

Dr.T.V.Rao

Rapid Diagnostic Methods Rapid Diagnostic Methods in Tuberculosisin Tuberculosis

Past decade has seen several Past decade has seen several emerging technologiesemerging technologies

How far practicable ?How far practicable ?


Emerging Rapid Methods.Emerging Rapid Methods.

1. Fast Plaque TB uses phage amplification 1. Fast Plaque TB uses phage amplification technology.technology.

2. ELISA ( QuantiFERON – TB )2. ELISA ( QuantiFERON – TB )

3. Enzyme-Linked immunospot 3. Enzyme-Linked immunospot ( ELISPOT ) ( ELISPOT ) ELISPOT proved highly useful to detect active ELISPOT proved highly useful to detect active tuberculosis in Adults and children. tuberculosis in Adults and children.


Emerging TechnologyEmerging TechnologyMODSMODS

Microscopic observation drug susceptibility Microscopic observation drug susceptibility assay. ( MODS )assay. ( MODS )

A new method gained importance in A new method gained importance in several reviews.several reviews.

Use a tissue culture plate based assay Use a tissue culture plate based assay with use of Middle Brook 7HG.with use of Middle Brook 7HG.

Needs a inverted light microscope.Needs a inverted light microscope. Even the drug resistance can be tested Even the drug resistance can be tested

with Rifampicin,and Isoniazid.with Rifampicin,and Isoniazid. Safe to work with cultures.Safe to work with cultures.

Dr.T.V.Rao

Non Specific TestsNon Specific Tests

Tuberculin testTuberculin test

( Mantoux Test )( Mantoux Test )


Tuberculin TestTuberculin Test( Mantoux Test )( Mantoux Test )

Test to be interpreted Test to be interpreted in relation to clinical in relation to clinical evaluation.evaluation.

Even the induration of Even the induration of 5 mm to be 5 mm to be considered positive considered positive when tested on HIV when tested on HIV patients.patients.

Lacks specificity.Lacks specificity.


Serology in Tuberculosis.Serology in Tuberculosis.

Several serological methods were Several serological methods were evaluated.evaluated.

But never gained the acceptance of But never gained the acceptance of the majority of the clinicians.the majority of the clinicians.

Serological tests are low sensitivity.Serological tests are low sensitivity. Many physicians depend on serology Many physicians depend on serology

in extra pulmonary tuberculosis.in extra pulmonary tuberculosis.

Dr.T.V.Rao

Dealing with Tuberculosis Dealing with Tuberculosis in HIV / AIDS patients.in HIV / AIDS patients.

Diagnosing Tuberculosis in Diagnosing Tuberculosis in HIV infected is a priority and HIV infected is a priority and

improve quality of Lifeimprove quality of Life


HIV/AIDS HIV/AIDS - - TuberculosisTuberculosis

Consider the HIV statusConsider the HIV status Identify the severity of Tuberculosis.Identify the severity of Tuberculosis. Early use of chest radiography.Early use of chest radiography. Maximal number of sputum smear Maximal number of sputum smear

examinations.examinations. Sputum concentration methods to be Sputum concentration methods to be

encouraged even by smaller laboratories.encouraged even by smaller laboratories. Explore the use of Florescent Microscopy.Explore the use of Florescent Microscopy. All smear negative specimens should be All smear negative specimens should be

cultured. cultured.


Limitations of Rapid TestsLimitations of Rapid Tests

The testing needs advanced and The testing needs advanced and sophisticated infrastructure.sophisticated infrastructure.

These tests are known for their These tests are known for their inability to diagnose between active inability to diagnose between active disease and latent infection.disease and latent infection.

Exclusively used in Developed Exclusively used in Developed nations.nations.

Dr.T.V.Rao

Extra pulmonary Extra pulmonary TuberculosisTuberculosis

Poses several challenges, Yet Poses several challenges, Yet no optimal, specific diagnostic no optimal, specific diagnostic

methodsmethods


Extra pulmonary TuberculosisExtra pulmonary Tuberculosis

A real challenge to Clinicians and A real challenge to Clinicians and Laboratories.Laboratories.

Optimal specimen collection a priority,Optimal specimen collection a priority, Molecular Methods are growing need.Molecular Methods are growing need. Clinicians start drug regimes on empirical Clinicians start drug regimes on empirical

basis.basis. Several serological tests for antibody Several serological tests for antibody

determinations are evaluated.determinations are evaluated.

Dr.T.V.Rao

Identification of Atypical Identification of Atypical MycobacteriumMycobacterium

A growing concern on infections with A growing concern on infections with less known, uncommon Mycobacterium less known, uncommon Mycobacterium

in immunosupreesed, an emerging in immunosupreesed, an emerging infectious disease probleminfectious disease problem


Atypical MycobacteriumAtypical Mycobacterium

Needs the help of reference Needs the help of reference laboratories.laboratories.

Needs different drug regimes, unlike Needs different drug regimes, unlike typical Mycobacterium isolates.typical Mycobacterium isolates.

Now a gowning concern in the era of Now a gowning concern in the era of AIDS.AIDS.


Future perceptionsFuture perceptions It is highly essential to explore and discover It is highly essential to explore and discover

rapid, simple, and accurate tuberculosis rapid, simple, and accurate tuberculosis diagnostic tools.diagnostic tools.

A massive investment, greater scientific interest, A massive investment, greater scientific interest, political commitment a top priority,political commitment a top priority,

Man power development, Human resource Man power development, Human resource utilization a greater concern.utilization a greater concern.

Microscopy and Florescent Microscopy utilization Microscopy and Florescent Microscopy utilization should be immediate concern, and strengthening should be immediate concern, and strengthening of treatment initiation protocols.of treatment initiation protocols.

Effective methods in diagnosing smear negative Effective methods in diagnosing smear negative patients a growing priority.patients a growing priority.


Microscopy in TuberculosisMicroscopy in TuberculosisTODAYTODAY

In spite of several In spite of several scientific, and scientific, and

molecular molecular advances advances

Microscopy in Microscopy in Tuberculosis Tuberculosis

continues to be continues to be back bone in back bone in Diagnosis.Diagnosis.

Dr.T.V.Rao

Specific detection of active Specific detection of active Tuberculosis cases in Tuberculosis cases in

patients with HIV infection or patients with HIV infection or AIDS is feasible and AIDS is feasible and

improves the rate of early improves the rate of early diagnosis and successful diagnosis and successful treatment of Tuberculosis.treatment of Tuberculosis.


Created for Medical graduates and Created for Medical graduates and Health care workers in Developing worldHealth care workers in Developing world

Dr.T.V.Rao, MD.Dr.T.V.Rao, MD.

e mail; e mail; [email protected]@gmail.com

Diagnosis of Tb

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