Molecular Diagnostics in Tuberculosis

Abstract

The processing of clinical specimens in the mycobacterial diagnostic laboratory has undergone remarkable improvements during the last decade. While microscopy and culture are still the major backbone for laboratory diagnosis of tuberculosis on a worldwide basis, new methods including molecular diagnostic tests have evolved over the last two decades. The majority of molecular tests have been focused on (i) detection of nucleic acids, both DNA and RNA, that are specific to Mycobacterium tuberculosis, by amplification techniques such as polymerase chain reaction (PCR); and (ii) detection of mutations in the genes that are associated with resistance to antituberculosis drugs by sequencing or nucleic acid hybridization. Recent developments in direct and rapid detection of mycobacteria, with emphasis on M. tuberculosis species identification by 16S rRNA gene sequence analysis or oligohybridization and strain typing, as well as detection of drug susceptibility patterns, all contribute to these advances. Generally, the balance between genome instability and genome maintenance as the basis for evolutionary development, strain diversification and resistance development is important, because it cradles the resulting M. tuberculosis phenotype. At the same time, semi-automated culture systems have contributed greatly to the increased sensitivity and reduced turnaround time in the mycobacterial analysis of clinical specimens. Collectively, these advances are particularly important for establishing the diagnosis of tuberculosis in children. More basic and operational research to appraise the impact and cost effectiveness of new diagnostic technologies must, however, be carried out. Furthermore, the design and quality of clinical trials evaluating new diagnostics must be improved to allow clinical and laboratory services that would provide rapid response to test results. Thus, important work remains before the new diagnostic tools can be meaningfully integrated into national tuberculosis control programs of high-burden countries.

Genotypic Susceptibility Testing of Mycobacterium tuberculosis Isolates for Amikacin and Kanamycin Resistance by Use of a Rapid Sloppy Molecular Beacon-Based Assay Identifies More Cases of Low-Level Drug Resistance than Phenotypic Lowenstein-Jensen Testing

“…Molecular tests offer the promise of more rapid drug resistance detection. A number of tests are available to detect resistance to many of the first-line anti-TB drugs (6)(7)(8). However, there are fewer rapid tests available to test for resistance to the injectable second-line drugs amikacin (AMK), kanamycin (KAN), and capreomycin (CAP).…”

Operating Cooperatively (OC) Sensor for Highly Specific Recognition of Nucleic Acids

“…These analytes were DNA olignonucleotide mimics of actual RNA sequence. Analysis of 16S rRNA is important for classification of bacteria species and for molecular diagnostics of infectious diseases [41]–[44]. On the other hand, 16S rRNAs, as well as their DNA amplicons can fail to hybridize to oligonucleotide probes [45], [46].…”

Nucleic acid testing for tuberculosis at the point-of-care in high-burden countries

“…Immunoassays have so far shown limited performance [11], and serological assays are not recommended for diagnosis of active TB [102]. Nucleic acid amplification testing (NAAT) enables sensitive and specific TB diagnosis [12,13]. NAAT systems with rapid turn-around time facilitate testing and treatment initiation in the same visit, which avoids loss to follow-up.…”