Genexpert For Global Health Problem: TB

 

Introduction:

Tuberculosis (TB) remains a major global health problem. The TB epidemic is larger than previously estimated with an incidence of 10.4 million new TB cases worldwide in 2015, of which 5.9 million (56%) were among men, 3.5 million (34%) among women and 1.0 million (10%) among children. India accounts for one fourth of the global TB burden. In 2015, an estimated 2,800,000 cases occurred and 480,000 people died due to TB. India has the highest burden of both TB and MDR-TB, based on estimates reported in Global TB Report 2016 (0.13 million of 0.48 million of MDR cases).

 

Due to the paucibacillary nature of pediatric TB, the yield of identifying TB bacilli using the traditional method of sputum microscopy is very low. Because of the problem of AFB-smear-negative cases, clinical diagnosis of TB is the norm. According to the latest Revised National TB Control Program (RNTCP) and Indian Academy of Pediatrics (IAP) guidelines, all attempts should be made to obtain a bacteriological diagnosis. Early detection is the key to successful treatment and reduction of the disease transmission.

 

Xpert MTB/RIF:

 

The Xpert MTB/RIF is a cartridge-based nucleic acid amplification test (NAAT) for simultaneous rapid tuberculosis diagnosis and rapid antibiotic sensitivity test. It is an automated diagnostic test that can identify Mycobacterium tuberculosis (MTB) DNA and resistance to rifampicin (RIF). It was co-developed by the laboratory of Professor David Alland at the University of Medicine and Dentistry of New Jersey (UMDNJ), Cepheid Inc. and Foundation for Innovative New Diagnostics, with additional financial support from the US National Institutes of Health (NIH).

In December 2010, the World Health Organization (WHO) endorsed the Xpert MTB/RIF for use in tuberculosis (TB) endemic countries. This followed 18 months of assessment of its field effectiveness in TB, MDR-TB and TB/HIV co-infection. The test may enable the diagnosis of TB in patients likely to be missed by traditional tests.

 

According to the Centers for Disease Control and Prevention (CDC) in 2015, the Xpert MTB/RIF test was "revolutionizing TB control by contributing to the rapid diagnosis of TB disease and drug resistance. The test simultaneously detects Mycobacterium tuberculosis complex (MTBC) and resistance to rifampin (RIF) in less than 2 hours. In comparison, standard cultures can take 2 to 6 weeks for MTBC to grow and conventional drug resistance tests can add 3 more weeks."

 

Sample required for testing:

 

Specimens that can be sent for testing include respiratory specimens such as sputum, bronchial or tracheal aspirates, broncho-alveolar lavage and gastric lavage as well as extra pulmonary specimens like tissue biopsy including lymph node, pus from abscess, CSF, ascitic and pericardial fluid, pleural fluid. CB-NAAT testing for TB on other samples such as stool, urine and blood is not recommended. According to WHO, a positive yield of 60–70% of culture positive cases, availability of results in 2 hours, detection threshold of 130–150 cfu/ml, and increasing number of other specimens that can be included along with respiratory specimens.

Though Xpert MTB/RIF test is a useful tool for rapid identification of rifampicin resistant M. tuberculosis, results must always be confirmed by culture and drug susceptibility testing (DST) to detect additional drug resistance. Culture-based methods currently remain the reference standard for drug susceptibility testing. Conventional microscopy and culture are essential for monitoring therapy and for performing DST for anti-TB agents other than rifampicin (including for isoniazid and second-line anti-TB drugs). Specimens of any type can be used for culture; the threshold for detection is low (10–100 cfu/ml compared to 130–150 cfu/ml for CB-NAAT). Patients with a negative GeneXpert result can still have TB with MTB or mycobacteria other than tuberculosis (MOTT) species. Additionally, the PCR test amplifies any DNA, whether of live or dead bacilli. Obtaining a clear history of treatment with ATT is required to avoid false positive results. 

 

Working:

The Xpert MTB/RIF detects DNA sequences specific for Mycobacterium tuberculosis and rifampicin resistance by polymerase chain reaction. It is based on the Cepheid GeneXpert system, a rapid, simple-to-use nucleic acid amplification test (NAAT). The Xpert® MTB/RIF purifies and concentrates Mycobacterium tuberculosis bacilli from sputum samples, isolates genomic material from the captured bacteria by sonication and subsequently amplifies the genomic DNA by PCR. The process identifies most of the clinically relevant Rifampicin resistance inducing mutations in the RNA polymerase beta (rpoB) gene in the Mycobacterium tuberculosis genome in a real time format using fluorescent probes called molecular beacons. Results are obtained from unprocessed sputum samples in 90 minutes, with minimal biohazard and very little technical training required to operate. This test was developed as an on-demand near patient technology which could be performed even in a doctor's office if necessary.

A review to assess the diagnostic accuracy of Xpert TB found that when used as an initial test to replace smear microscopy it had pooled sensitivity of 89% and specificity of 99% . However, when Xpert TB was used as an add-on for cases of negative smear microscopy the sensitivity was only 67% and specificity 99%. In a clinical study conducted the sensitivity of the MTB/RIF test on just 1 sputum sample was 92.2% for culture-positive TB; 98.2% for smear+ and culture-positive cases; and 72.5% for smear-negative, culture-positive cases, with a specificity of 99.2%. Sensitivity and higher specificity were slightly higher when 3 samples were tested.

 

Advantages:

 

The main advantages of the test are, for diagnosis, reliability when compared to sputum microscopy and the speed of getting the result when compared with the culture test. For diagnosis of TB, although sputum microscopy is both quick and cheap, it is often unreliable. It is particularly unreliable when people are HIV positive. Although culture gives a definitive diagnosis, to get the result usually takes weeks rather than the hours of the Genexpert test.

The main advantage in respect of identifying rifampicin resistance, is again the matter of speed. Normally to get any drug resistance result takes weeks rather than hours.

 

Disadvantages:

The disadvantages include:

• The shelf life of the cartridges is only 18 months;
• A very stable electricity supply is required;
• The instrument needs to be recalibrated annually;
• The cost of the test;
• The temperature ceiling is critical.