학술논문
A Comparative Study on Visual Detection of Mycobacterium tuberculosis by Closed Tube Loop-Mediated Isothermal Amplification: Shedding Light on the Use of Eriochrome Black T.
Document Type
Academic Journal
Author
Neshani A; Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad 91967-73113, Iran.; Zare H; Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad 91967-73113, Iran.; Sadeghian H; Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad 91967-73113, Iran.; Department of Laboratory Sciences, School of Paramedical Sciences, Mashhad University of Medical Sciences, Mashhad 91779-48964, Iran.; Safdari H; Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad 91967-73113, Iran.; Department of Laboratory Sciences, School of Paramedical Sciences, Mashhad University of Medical Sciences, Mashhad 91779-48964, Iran.; Riahi-Zanjani B; Medical Toxicology Research Center, Mashhad University of Medical Sciences, Mashhad 91779-48564, Iran.; Aryan E; Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad 91967-73113, Iran.; Department of Medical Microbiology, Ghaem University Hospital, Mashhad University of Medical Sciences, Mashhad 91766-99199, Iran.
Source
Publisher: MDPI AG Country of Publication: Switzerland NLM ID: 101658402 Publication Model: Electronic Cited Medium: Print ISSN: 2075-4418 (Print) Linking ISSN: 20754418 NLM ISO Abbreviation: Diagnostics (Basel) Subsets: PubMed not MEDLINE
Subject
Language
English
ISSN
2075-4418
Abstract
Loop-mediated isothermal amplification is a promising candidate for the rapid detection of Mycobacterium tuberculosis . However, the high potential for carry-over contamination is the main obstacle to its routine use. Here, a closed tube LAMP was intended for the visual detection of Mtb to compare turbidimetric and two more favorable colorimetric methods using calcein and hydroxy naphthol blue (HNB). Additionally, a less studied dye (i.e., eriochrome black T (EBT)) was optimized in detail in the reaction for the first time. Mtb purified DNA and 30 clinical specimens were used to respectively determine the analytical and diagnostic sensitivities of each method. The turbidimetric method resulted in the best analytical sensitivity (100 fg DNA/reaction), diagnostic sensitivity and specificity (100%), and time-to-positivity of the test (15 min). However, this method is highly prone to subjective error in reading the results. Moreover, HNB-, calcein-, and EBT-LAMP could respectively detect 100 fg, 1 pg, and 1 pg DNA/reaction (the analytical sensitivities) in 30, 15, and 30 min, while the diagnostic sensitivity and specificity were respectively 93.3% and 100% for them all. Interestingly, EBT-LAMP showed the lowest potential for subjective error in reading the results. This report helps judiciously choose the most appropriate visual method, taking a step forward toward the field applicability of LAMP for the detection of Mtb, particularly in resource-limited settings.