A novel and more sensitive loop-mediated isothermal amplification assay targeting IS6110 for detection of Mycobacterium tuberculosis complex.
Developing improved tuberculosis (TB) diagnostics is one of the international research priorities, as TB remains globally a major health threat. Loop-mediated isothermal amplification (LAMP) is a new nucleic acid detection method that can be used in low-resource settings, because it does not require expensive or complex instruments. Using the repetitive insertion sequence IS6110 as a target gene, we developed an efficient LAMP assay, which specifically detects members of the Mycobacterium tuberculosis complex (MTBC). This assay proved 20 times more sensitive than IS6110-based conventional PCR. Moreover, its sensitivity was, respectively, 50 and 20 times higher than the one obtained with the two previously described LAMP assays for M. tuberculosis, based on gyrB and rrs, respectively. Identical sensitivities were obtained for LAMP and nested PCR, but the LAMP assay was more rapid and cost-effective than the latter. Although, our LAMP assay can successfully be performed using a non-denatured template, this results in a 200-fold reduction in the sensitivity of the assay. Moreover, by performing our LAMP assay on 15 clinical sputum samples from TB patients we were able to detect MTB. Taken together, our preliminary results indicate that IS6110-based MTBC-LAMP assay is a promising new TB-diagnostic test, with high sensitivity and that could easily be applied for the diagnosis of TB in a low-resource setting.
Aryan
Ehsan
E
Department of Medical Microbiology, Faculty of Medicine, Ahvaz Jondishapour University of Medical Sciences, 61357-15794 Ahvaz, Iran. ehsanaryan@hotmail.com <ehsanaryan@hotmail.com>
Makvandi
Manoochehr
M
Farajzadeh
Ahmad
A
Huygen
Kris
K
Bifani
Pablo
P
Mousavi
Seyed-Latif
SL
Fateh
Abolfazl
A
Jelodar
Abbass
A
Gouya
Mohammad-Mehdi
MM
Romano
Marta
M
D001431:Bacteriological Techniques / Q000191:economics / Q000379:methods*; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors; D014376:Tuberculosis / Q000175:diagnosis* / Q000382:microbiology
D004251:DNA Transposable Elements; D004269:DNA, Bacterial