Evaluation of multiplex polymerase chain reaction utilising multiple targets in Mycobacterium tuberculosis direct test negative but culture positive cases: a potential method for enhancing the diagnosis of tuberculosis.
OBJECTIVE
To evaluate multiplex Polymerase Chain Reaction (MPCR) utilising multiple targets (IS6110, Protein b [Pab] and MPB64 genes) in Mycobacterium tuberculosis Direct Test (MTD) negative but culture positive cases and comparison of MPCR with Real-Time polymerase chain reaction (RT-PCR) for diagnosis of tuberculosis.


METHODS
MPCR was carried out on 28 culture positive sputum samples. Out of 28 culture positive samples, 17 were originally reported, as MTD test negative and 11 were MTD test positive, respectively. The results of MPCR were compared with RT-PCR. To check the specificity of the tests, MPCR and RT-PCR were also evaluated with 16 non-tuberculous mycobacterial (NTM) isolates.


RESULTS
Out of 28 culture positive sputum samples, MPCR was positive in all 28/28 samples, whereas RT-PCR was positive in 27/28 samples and MTD test was originally tested positive in six sputum samples and on repeating MTD testing, five more sputum samples were positive and thus total number of MTD positive were 11/28 sputum samples, respectively. All the tests were negative on evaluation with all the 16 NTMs, thus giving specificity of 100% to all the tests; sensitivity of MPCR, RT-PCR and MTD tests were 100%, 96.42% and 39.28%, respectively, in these specifically selected samples.


CONCLUSIONS
MPCR may be an important tool in the rapid diagnosis of tuberculosis especially in disease endemic, resource limited countries.
Department of Medical Microbiology, Post Graduate Institute of Medical Education and Research, Chandigarh, India.
K
K
Sharma
D
D
Ashkin
P
P
Fiorella
D
D
Willis
S
S
Dean
A
A
Sharma
K K
KK
Singh
Y
Y
Lee
M
M
Pedrosa
G
G
Singh
M
M
Sharma
S
S
Laal
D000942:Antigens, Bacterial / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D004251:DNA Transposable Elements; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D013183:Sputum / Q000382:microbiology; D013997:Time Factors; D014376:Tuberculosis / Q000175:diagnosis*
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; C080865:MPB64 protein, Mycobacterium