Module 1

Understanding Tuberculosis, the Global Emergency

Learning Objectives
At the end of this module, the participant will be able to:

1. Explain the TB epidemic and the annual global TB burden 2. Describe the forms of TB and how TB is transmitted 3. Discuss the ISTC Standards for Diagnosis 4. Define and compare various methods of TB diagnosis,

Learning Objectives
5. Describe NTP and its purposes 6. Describe the role of the laboratory in NTP 7. Describe the DOTS component of STOP TB strategy 8. Explain the importance of AFB microscopy in the DOTS program 9. Describe levels of TB laboratory services.

 What is tuberculosis? The TB epidemic and the annual global TB burden Transmission and forms of TB Risk of Disease ISTC Standards for Diagnosis TB Diagnosis

Content Overview

Content Overview
National TB Program and its purposes The role of the laboratory in NTP The DOTS component of STOP TB strategy The importance of AFB-microscopy in DOTS
programs Levels of laboratory services

Global Emergency
Tuberculosis kills 5,000 people a day !

2.3 million die each year !

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

Disturbing Statistics
1/3 of worlds population is infected with TB 8 Million people develop active TB every year TB kills more young women than any other
disease

More than 100,000 children will die from TB

this year

Hundreds of thousands of children will become

TB orphans
Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

PHILIPPINE STATUS

9th amongst the 22 high-burdened countries

worldwide (WHO watchlist)

4th amongst the countries in the Western Pacific

Region of WHO (2nd among the HBCs in WPRO)

8th amongst the countries with high burden for

MDRTB TB is 6th in mortality and morbidity
TB morbidity & mortality Natl Objectives for Health 2005-2010 DOH

What is TB?
TB is an infectious disease that affects mainly the lungs (pulmonary TB or PTB) but can also attack any part of the body (extra-pulmonary TB or EPTB)

A person with PTB is infectious to others!

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

The Causes of TB
Mycobacterium tuberculosis complex

Mycobacterium tuberculosis Mycobacterium bovis

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

10

Staining Characteristics

Mycobacteria are called Acid-Fast Bacilli (AFB) due to their microscopic appearance after decolorization. Organisms appear red on a blue background

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

11

TB Transmission (infection)
Person to person via Airborne transmission in Confined environment
12

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

Risk Factors for Infection

Exposure to TB bacilli Duration of exposure to a person with PTB Intensity of exposure

Untreated AFB smear positive PTB cases are the most infectious

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

13

Risk Factors for Disease

Development of disease depends on
individual susceptibility

HIV increases the risk of getting TB disease

10% Life time risk of TB in HIV negative
10% Annual risk of TB in HIV positive

14
Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

What is MDR-TB?
Multidrug-resistant TB (MDR TB) is TB that is
resistant to at least two of the best anti-TB drugs, isoniazid and rifampicin. These drugs are considered first-line drugs and are used to treat all persons with TB disease.

ISTC TB Training Modules 2009

15

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

ISTC TB Training Modules 2009

16

Purpose ISTC Purpose of ISTC

17

ISTC TB Training Modules 2009

18

THE ISTC
Intended to facilitate the effective engagement of all care providers in delivering high quality care for patients of all ages. intended to complement, not replace, national and local recommendations. The ISTC should be viewed as a living document that will be revised as technology, resources, and circumstances change.
19

Introduction

20

ISTC TB Training Modules 2009

Standards for Diagnosis

21

ISTC TB Training Modules 2009

Standards for Diagnosis

22
ISTC TB Training Modules 2009

Microbiologic Diagnosis of TB
Significance of microbiologic testing for public health goals and patient care:
WHO global target of 70% case detection of new smearpositive cases Rapid and accurate case detection coupled with effective treatment is essential to reduce the incidence of TB Failure to perform a proper diagnostic evaluation before initiating treatment potentially:
Exposes the patient to the risks of unnecessary or wrong treatment May delay accurate diagnosis and proper treatment
23
ISTC TB Training Modules 2009

ISTC Standard 1
All persons with otherwise unexplained productive cough lasting two-three weeks or more should be evaluated for tuberculosis
24

ISTC Standard 2
All patients (adults, adolescents, and children who are capable of producing sputum) suspected of having pulmonary TB should have at least two sputum specimens obtained for microscopic examination in a quality-assured laboratory. When possible, at least one early morning specimen should be obtained.
25

ISTC Standard 3
For all patients (adults, adolescents, and children) suspected of having extrapulmonary TB, appropriate specimens from the suspected sites of involvement should be obtained for microscopy, culture, and histopathological examination.

26

NTP
Extra-Pulmonary TB (EP)
A patient with at least one mycobacterial smear/culture positive from an extra-pulmonary site (organs other than the lungs: pleura, lymph nodes, genito-urinary tract, skin, joints and bones, meninges, intestines, peritoneum, and pericardium, among others) A patient with histological and/or clinical evidence consistent with active extra pulmonary TB and there is a decision by a physician to treat the patient with anti-TB drugs All EP cases shall undergo DSSM prior to treatment.
NTP MOP 2005
27

ISTC Standard 4
All persons with chest radiographic findings suggestive of tuberculosis should have sputum specimens submitted for microbiological examination.
28

ISTC Standard 5
The diagnosis of sputum smear-negative pulmonary tuberculosis should be based on the following criteria: At least two negative sputum smears (including at least one early morning specimen) Chest radiography findings consistent with tuberculosis Lack of response to a trial of broad-spectrum antimicrobial agents (Note: Because the fluoroquinolones are active against M. tuberculosis complex, and thus may cause transient improvement in persons with tuberculosis, they should be avoided.)

29

ISTC Standard 5
For such patients, sputum cultures should be obtained. In persons who are seriously ill or have known or suspected HIV infection, the diagnostic evaluation should be expedited and if clinical evidence strongly suggests TB, a course of antituberculosis treatment should be initiated.
ISTC TB Training Modules 2009
30

ISTC Standard 6
In all children suspected of having intrathoracic (i.e., pulmonary, pleural, and mediastinal or hilar lymph node) TB, bacteriological confirmation should be sought through examination of sputum (by expectoration, gastric washings, or induced sputum) for smear microscopy and culture. In the event of negative bacteriological results, a diagnosis of TB should be based on: The presence of abnormalities consistent with TB on chest radiography A history of exposure to an infectious case, evidence of TB infection (positive tuberculin skin test or interferon gammarelease assay), and Clinical findings suggestive of TB
31

ISTC Standard 6
For children suspected of having EPTB, appropriate specimens from the suspected sites of involvement should be obtained for microscopy and for culture and histopathological examination.
32

Diagnosis of TB
Direct demonstration of AFB in sample Growth of TB bacilli in culture Skin Test Nucleic Acid Amplification tests

(NAATs) T-cell-based interferon-gamma release assay (IGRAs) X-Ray
33

ISTC TB Training Modules 2009

Sputum Smear Microscopy

Sputum smear microscopy is the most important test for the diagnosis of pulmonary TB in many areas of the world Direct smears (unconcentrated specimen) are most common Fluorescence microscopy and chemical processing can increase sensitivity Assessment of laboratory quality is essential
34

ISTC TB Training Modules 2009

Sputum Microscopy: Direct Smears

Direct smears of unconcentrated sputum:
Fast, simple, inexpensive, widely applicable Extremely specific for M. tuberculosis in high-incidence areas Ziehl-Neelsen staining (carbol fuchsin type) most common
ISTC TB Training Modules 2009
35

Sputum Smear Microscopy

Carbolfuchsin-based stains Utilize a regular light microscope Must be read at a higher magnification Two types: Ziehl-Neelsen and Kinyoun. Both use

carbolfuchsin/phenol as the primary dye Smear is then decolorized with acid (HCI) alcohol and counter-stained with methylene blue

ISTC TB Training Modules 2009

36

Ziehl-Neelsen (ZN) Stain

ISTC TB Training Modules 2009

37

Sputum Microscopy: Fluorochrome Stain

Fluorochrome stain

Fluorochrome stained smears require a fluorescent microscope Generally read at 250X-450X magnification which allows rapid scanning of the smear Auramine-rhodamine is an example of such a stain where the AFB appear yellow against a black background

ISTC TB Training Modules 2009

38

Auramine-rhodamine Stain

ISTC TB Training Modules 2009

39

Advantages of AFB Smear Microscopy

Microscopy is a simple convenient test Requires minimal infrastructure and

equipment Highly accurate, inexpensive and fast Accessible to the majority of patients Prioritizes infectious cases

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

40

Limitations of Microscopy
Can not distinguish between dead or live
bacteria

High bacterial load >30005000 AFB /mL is

required for detection

Can not do species identification Can not perform Drug Susceptibility Testing

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

41

Fluorescence Microscopy
Advantages:

More accurate: 10% more sensitive than light microscopy, with specificity comparable to ZN staining Faster to examine = less technician time Disadvantages:

Higher cost and technical complexity, less feasible in many areas

Steingart KR, et al. Lancet Infect. Dis. 2006; 6 (9):570-81

42

Culture and Drug Susceptibility Testing

Although sputum microscopy is the first bacteriologic diagnostic test of choice, both culture and drug susceptibility testing (DST) can offer significant advantages in the diagnosis and management of TB.
ISTC TB Training Modules 2009
43

Culture: Advantages
Higher sensitivity than smear microscopy (culture
can make diagnosis despite fewer bacilli in specimen) If TB suspected and sputum smears are negative, culture may provide diagnosis Allows for identification of mycobacterial species Allows for drug susceptibility testing

ISTC TB Training Modules 2009

44

Culture: Disadvantages

Cost Technical complexity May take weeks to get results Requires ongoing quality assurance

Therefore, more likely to be found in major

referral centers. Avoid delaying appropriate TB treatment in suspicious cases while awaiting results.
ISTC TB Training Modules 2009
45

Culture: Solid Media

Solid media have the advantage that organisms (colonies) can be seen on the surface of the medium Types most commonly used are:
Lowenstein-Jensen: egg-based Middlebrook 7H 10 or 7H11: agar-based Ogawa
ISTC TB Training Modules 2009
46

Culture: Liquid Media

More sophisticated equipment Faster detection of growth Higher sensitivity than solid media Can also be used for drugsusceptibility testing Two examples: BACTEC MGIT
MGIT

BACTEC MGIT Incubator

ISTC TB Training Modules 2009

47

Culture: Identification of Mycobacteria

Growth characteristics (preliminary ID) Preliminary indication of M.tb can be determined from colony characteristics

Rate of growth Colonial morphology Pigmentation Biochemical tests There is a battery of 8 12 biochemical tests used to differentiate M.tb within the genus Nitrate reduction and niacin production are definitive for M.tb

ISTC TB Training Modules 2009

48

Culture: Identification of Mycobacteria

Visual assessment of colony morphology:

Smooth, buff-colored colonies suggestive of Mycobacterium avium complex

ISTC TB Training Modules 2009

Rough, buff-colored colonies suggestive of Mycobacterium tuberculosis

49

Culture: Cross-Contamination

Be aware that faulty technique can lead to laboratory crosscontamination of specimens (difficult to verify without access to more technical testing). Adequate quality control is an essential component of any mycobacteriology laboratory.
ISTC TB Training Modules 2009
50

Rapid Diagnostic Testing

Nucleic acid probe tests (non-amplified) to identify organisms grown in culture: DNA probe tests are species or complex specific
Commercial probes are available for M.tb complex, MAC, M. kansasii and M. gordonae

Nucleic acid amplification tests (NAAT): These tests are designed to amplify and detect DNA specific to M.tb Enables direct detection of M.tb in clinical specimens
ISTC TB Training Modules 2009
51

Other Rapid Diagnostic Tests

Loop-mediated isothermal amplification (LAMP)
Rapid, simplified NAAT still under investigation May be more feasible in lower resource settings

Immunological tests
Serologic tests for antibody, antigens, and immune complexes; not currently accurate enough to replace microscopy and culture.

ISTC TB Training Modules 2009

52

Other Rapid Diagnostic Tests

High performance liquid chromatography
(HPLC)
HPLC uses a liquid chromatography method to identify mycobacteria based on their mycolic acid profiles (cell wall composition) The equipment is expensive and is usually reserved for larger, specialized, reference laboratories

ISTC TB Training Modules 2009

53

Rapid Drug Susceptibility Tests

Line-probe assays
Identifies M.tb and genetic mutations associated with INH and RIF resistance

Can be used directly on sputum specimens, results within 1-2 days

Molecular beacons

*GenoType MTDBRplus strips (Hain Lifescience)

Bacteriophage-based assays
*Barnard et al. Am. J. Respir. Crit. Care Med 2008; 177: 787-792
54

National Tuberculosis Control Program (NTP)

Objectives Reduce mortality, morbidity and disease
transmission and avoid the development of drug resistance

In the long term, to eliminate suffering due to

TB

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

55

Goals of the NTP

Detect at least 70% of the infectious
cases

Cure at least 85% of newly detected

cases of smear-positive TB

Reduce prevalence of and deaths due to

TB

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

56

NTP

Case finding, which is the identification and diagnosis of TB cases among individuals with suspected signs and symptoms of TB, is a basic step in TB control. Fundamental to case finding is the detection of infectious cases through direct sputum smear microscopy (DSSM).
DSSM results serve as basis for categorizing TB symptomatics according to standard case definition, monitor progress of patients with sputum smear-positive TB while they are receiving antiTB treatment These are also used to confirm cure at the end of treatment.
NTP MOP 2005
57

NTP
National TB Reference Laboratory

Quality assurance of sputum microscopy smear are done quarterly

Objectives of QA program:
ensure that the reported results are accurate identify practices that are potential sources of error ensure that appropriate corrective actions are initiated

NTP MOP 2005

58

What is STOP TB Strategy

1. Pursuing quality DOTS expansion and enhancement 2. Addressing TB/HIV and MDR-TB 3. Contributing to health system strengthening

4. Engaging all care providers 5. Empowering patients and communities 6. Enabling and promoting research

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

59

Who has Tuberculosis? TB Pathogenesis

Exposure

Infection

Active Disease

STOP TB AT THE SOURCE!

Inactive Disease

60

DOTS Component of STOP TB Strategy

Political commitment to TB control Case detection by quality assured

bacteriology

Regular, uninterrupted supply of high quality

anti-TB drugs

Standardized treatment with supervision

and patient support

Standardized recording and reporting

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

61

Benefits of DOTS
Produces cure rates of up to 95 % Prevents new infections
Prevents the development of MDR-TB

Cost effective
Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

62

IMPORTANCE OF ACID-FAST BACILLI (AFB) MICROSCOPY IN DOTS PROGRAMS

Diagnosis

Treatment Follow-up

63

Role of Laboratory

Detection of infectious cases Monitoring of treatment progress

Documentation of cure

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

64

Detection and treatment of infectious cases reduces the spread of Tuberculosis!

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

65

 AFB smear-positive patients are usually

sick and seek treatment.

Pulmonary Positive Patients

AFB smear-positive patients are much

more likely to die if untreated.

Untreated, an AFB smear-positive patient

may infect 1015 persons/year.

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

66

NTP RESPONSE TO MDR-TB PROBLEM

Policy and technical support: AO 2008-0018: Guidelines for the Implementation of PMDT Implementing guidelines and training modules for PMDT Mainstreaming of MDR-TB services to the NTP Public-private collaboration from diagnosis to management of cases
67

NTP MOP 2005

ISTC Standard 12

Patients with or highly likely to have tuberculosis caused by drug-resistant (especially MDR/XDR) organisms should be treated with specialized regimens containing 2nd-line antituberculosis drugs The regimen chosen may be standardized or based on suspected or confirmed drug susceptibility patterns At least four drugs to which the organisms are known or presumed to be susceptible, including an injectable agent, should be used, and treatment should be given for at least 1824 months beyond culture conversion

68

Laboratory Network
Central Laboratory

Intermediate laboratories

Peripheral Laboratories

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

69

Peripheral Laboratory
Located within a general dispensary,
clinic or hospital

Limited services for TB diagnosis

Sputum specimen collection AFB sputum smear microscopy

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

70

Intermediate Laboratory
Regional/provincial or large hospital Services for TB diagnosis
Sputum specimen collection Sputum smear microscopy Culture and identification of MTB

Support for peripheral laboratories

Supply of reagents and materials Training, supervision, EQA of sputum smear

microscopy
Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

71

Central Laboratory

Country/regional level Services for TB diagnosis Sputum smear microscopy Culture and identification of MTB Drug susceptibility testing of TB Support for the laboratory network Advice on procurement Organization and participation in training, supervision, EQA of sputum smear microscopy Other activities Participation in operational research Drug resistance surveillance
72

Source: http://wwwn.cdc.gov/dls/ila/acidfasttraining/section4.aspx

Laboratory is the key Component in TB Control

NO LABS
NO DIAGNOSIS

NO TREATMENT NO DOTS

NO TB CONTROL
73

Summary
What is TB and how it is transmitted? What are the ISTP Standards for Diagnosis? What are the goals of NTP? Why is microscopy an effective diagnostic
technique?

What is DOTS? What is the role of the laboratory in TB

control?
74

References
1.
2. 3. 4. 5.
PTSI,DOH,RITM,NTRL, Training Manual (Training course on Direct Sputum Microscopy http://wwwn.cdc.gov/dls/ila/acidfasttrainin g/section1.aspx#training International Standards for Tuberculosis Care, 2009 http://www.who.int/tb/strategy/en/ NTP MOP 2005
75