Journal of Microbiological Methods
Volume 75, Issue 1, September 2008, Pages 1-11
Review
Molecular diagnostic tools in mycobacteriology
Ioannis K. Neonakisa, , 1, , Zoe Gittia, 1, Elias Krambovitisb and Demetrios A. Spandidosa
aMycobacteriology Laboratory, Department of Clinical Bacteriology, Parasitology, Zoonoses and Geographical Medicine, University Hospital of Heraklion, 712 01 Heraklion, Crete, Greece bMicrobiology and Parasitology Laboratory, Department of Veterinary Medicine, School of Health Sciences, University of Thessaly, Karditsa, Greece
Received 30 January 2008; revised 14 May 2008; accepted 23 May 2008. Available online 31 May 2008.
Abstract
Although the diagnosis of mycobacteriosis and susceptibility testing are still primarily based on conventional methods (staining, culture, biochemical analysis, proportional method), a series of molecular assays are increasingly introduced and incorporated in the workflow of clinical mycobacteriology laboratories worldwide. These assays are rapid and offer high sensitivities and specificities. In the present review, we describe the molecular assays concerning the early detection of Mycobacteria in clinical specimens, the identification of mycobacterial species, the detection of drug resistance and the typing for epidemiological investigations.
Keywords: Drug resistance; Molecular diagnostics; MTBC; Mycobacteria; NTM; Tuberculosis
Article Outline
1. Introduction
2. Direct detection of mycobacteria in clinical specimens
2.1. In-house PCR for detection of mycobacteria from clinical specimens
2.2. Commercially available assays
2.2.1. Cobas Amplicor M. tuberculosis assay (Amplicor; Roche Diagnostic Systems, Branchburg, NJ )
2.2.2. Amplified M. tuberculosis direct test (AMTD; bio Merieux, Gen-probe, Inc., San Diego, Calif.)
2.2.3. DProbe Tec ET (energy transfer) M. tuberculosis Direct Detection Assay (DTB), (BDProbe Tec; Becton Dickinson Bioscience, Sparks, Md.)
2.2.4. Genotype mycobacteria direct assay for detection of M. tuberculosis complex and four atypical mycobacteria (Hain Lifescience, Nehren, Germany)
2.2.5. LCx MTBC assay (Abbott Laboratories, Diagnostic Division, Chicago, USA)
2.2.5.1. Remarks
3. Identification of mycobacterial species from culture by molecular methods
3.1. PCR-based sequencing
3.2. DNA probe technology
3.3. Line probe technology (hybridization in strips)
3.3.1. Inno LiPA Mycobacteria v2, (Innogenetics, Ghent, Belgium)
3.3.2. GenoType Mycobacterium (Hain Lifescience, Nehren, Germany)
3.4. PRA method. [polymerase chain reaction and restriction enzyme analysis for identification of mycobacteria from culture]
3.5. Pyrosequencing
3.6. DNA microarrays (DNA chips)
4. Molecular methods for detecting drug resistance in mycobacterial strains
4.1. PCR-DNA sequencing
4.2. Hybridization-based techniques
4.2.1. Line probe technology
4.2.1.1. Inno-LiPA RifTB (Innogenetics, Ghent, Belgium)
4.2.1.2. GenoType MTBDR plus (Hain Lifescience, Germany)
4.3. Hybridization on DNA chips
4.4. PCR-SSCP (single-strand-conformation-polymorphisms)
4.5. Pyrosequencing
4.6. Real-time PCR methodology
4.7. Mycobacteriophage D29-based assay
5. Molecular epidemiological methods
5.1. IS6110-RFLP method
5.2. Spoligotyping (spacer oligotyping)
5.3. MIRU-VNTR method
5.4. Repetitive sequence-based PCR method
5.5. Remarks
References
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