Peran Tes Cepat Molekular

dalam Penegakkan Diagnosis TB dan TB RO

Seminar Ilmiah – Maret 2019

RSPI Prof Dr Sulianti Saroso
Alur Prosedur Laboratorium Mikrobiologi Klinik
 Patient with TB symptoms Cascade of Opportunities
presents to clinic for Poor Outcomes with
Physician suspects TB? TB Diagnostic Tests
No
Physician orders diagnostic
test? No

Diagnostic test performed No

properly?

Test result positive? No Diagnosis Delayed.

Test sensitivity Patient returns to clinic?
No
Results made available to
clinician and patient? Patient suffers from
No
untreated TB
Patient returns and is offered
treatment? No

Patient initiates and

completes treatment?

TB morbidity averted, after Adapted from: Dowdy et al., PLoS Medicine

diagnostic delay 2011:8(7): e1001063
 The spectrum of TB
from Mycobacterium tuberculosis infection to active
(pulmonary) TB disease

NATURE REVIEWS | DISEASE PRIMERS VOLUME 2 | 2016

Tuberculosis diagnostic

Non
Molecular
molecular
base
base
Culture
base

Microscopy

Lancet Infect Dis 2018 Published Online March 23, 2018

Tuberculosis
diagnostic
pipeline
 International Standards for Tuberculosis Care
(ISTC)
ISTC 2009
Standard 3
For all patients (adults, adolescents, and
children) suspected of having
extrapulmonary tuberculosis, appropriate
specimens from the suspected sites of
involvement should be obtained for
microscopy, culture, and histopathological
examination.
ISTC 2014
• All patients, including children, who are
suspected of having pulmonary
tuberculosis and are capable of
producing sputum should have at least
two sputum specimens submitted for
smear microscopy or a single sputum
specimen for Xpert® MTB/RIF* testing in
a quality-assured laboratory.
• Patients at risk for drug resistance, who
have HIV risks, or who are seriously ill,
should have Xpert MTB/RIF performed as
the initial diagnostic test.
• Blood-based serologic tests and
interferon-gamma release assays should
not be used for diagnosis of active
tuberculosis.
 Key differences between the 2009 and 2014
Standard 4 • The importance of microbiological diagnosis of
extrapulmonary tuberculosis is emphasized.
• WHO recommendations for the use of rapid molecular testing
for samples from extrapulmonary sites are included.

Standard 5 The WHO recommendations for use of rapid molecular testing for
diagnosis of tuberculosis among persons who are suspected of
having the disease but have negative sputum smear microscopy
are presented.

Standart 6 The WHO recommendations for the use of rapid molecular

testing for the diagnosis of tuberculosis in children are presented.

Standart 10 The role of microscopy in monitoring response in patients who

had the diagnosis established by a rapid molecular test is
described.

Standart 11 Describes the use of Xpert® MTB/RIF in assessing for rifampicin

resistance and line probe assay for detecting resistance to both
isoniazid and rifampicin.
 Pemeriksaan
Tes Cepat Molekular
(Xpert MTB/Rif)
Penggunaan

Xpert MTB/RIF berfungsi :

• mendeteksi keberadaan dari kompleks DNA bakteri Mycobacterium
tuberculosis (MTB)
• resistensi (kekebalan) bakteri tersebut terhadap pengobatan dengan
Rifampisin.

Secara lebih spesifik lagi, Xpert MTB/RIF mendeteksi gen

rpoB (dan mutasi yang terjadi, apabila ditemukan) yang
berhubungan dengan resistensi bakteri MTB terhadap
Rifampisin.
• Xpert MTB/RIF merupakan tes in vitro Diagnostik
dengan hasil semi kuantitatif

• Xpert MTB/RIF hanya digunakan untuk diagnosa

pasien terduga TB (bukan untuk monitoring pasien)
 Perbandingan kinerja
Deteksi kuman dalam sputum

Xpert

•Limit deteksi pada ZN: 10000 kuman /ml sputum

•Limit deteksi pada MGIT: 10-100 kuman/ml sputum
•Limit deteksi pada GeneXpert: 100-1000 kuman /ml sputum

 Lebih sensitif dibandingkan ZN, lebih sederhana

pengerjaannya
 Dasar Biologi Molekuler : adanya DNA Mtb pd sputum

DNA  mRNA  Protein

Mutasi DNA  protein mutanresistensi obat

M. Tuberculosis GeneXpert
rifampicin terikat pada rpoB (RNA
Deteksi DNA mutan yg
polymerase) yg disandi gen rpoβ
menyebabkan resistensi
Mutasi gen rpoB RNA polymerase mutan
------- resistensi RIF terhadap RIF
Mekanisme Deteksi Bakteri TB

81 bp daerah inti gen rpoB

 Sistem otomatis yang mengintegrasikan proses purifikasi
spesimen, amplifikasi asam nukleat, dan deteksi sekuen target

Xpert®

Pencucian Ekstraksi Pemurnian

DNA DNA

Sampel
pasien asterM
Mix

ume
er re real-time lainnya
Instrumen

Positif
Analisis Deteksi Amplifikasi
Negati
f

11
Meta-analysis of the
sensitivity and specificity
of Xpert MTB/RIF in
diagnosing
extrapulmonary TB and
rifampicin resistance in
adults and children
(WHO, 2013)
TCM – TB RO

Smith SE; Int J Tuberc Lung Dis. 2012 February ; 16(2): 203–205
Hasil Pelaporan

Xpert® MTB/RIF Interpretasi

MTB DETECTED; MTB terdeteksi, Rifampisin

RIF Resistance resisten
DETECTED

MTB DETECTED; MTB terdeteksi, Rifampicin

RIF Resistance resisten tidak terdeteksi
NOT DETECTED (sensitif)

MTB DETECTED; MTB terdeteksi, Rifampicin

RIF Resistance resisten belum dapat
INDETERMINATE ditentukan
Interpreting results from Xpert MTB/RIF tests

WHO, 2014
 Sampel (sputum)
• Jumlah minimum sampel yang digunakan adalah 1 mL

• Sampel harus murni dahak (sputum). Dilarang

menggunakan sampel yang mengandung partikel
makanan dan/atau partikel padat lain

• penyimpanan spesimen dahak dilakukan pada:

• Suhu 2-8o C. Spesimen dahak harus diolah dalam 7
hari, ATAU
• Suhu ruang. Spesimen dahak harus segera diolah
dalam 3 hari.

• Untuk sampel kiriman; selama proses transportasi

sampel harus disimpan pada suhu 2 – 8 °C
Prosedur kerja