Pyrosequencing technology for resistance detection

Detection of a Range of Possible Resistance Mutations with a Single Assay

Many common resistance mutations are single nucleotide polymorphisms (SNPs) or clustered mutations in mutation “hot spots”, which makes sequencing an appropriate choice to characterize them. Sequencing has the advantage over hybridization-based approaches in that a single assay can detect a range of possible mutations in a region, as well as new and unexpected mutations that hybridization-based methods would miss (e.g. Q-PCR).

Compared to traditional biochemical and sequencing methods, Pyrosequencing technology offers significant advantages for detecting resistance: it is accurate with built-in QC, takes less than one hour to perform an analysis, and offers high throughput (96 samples in parallel) at significantly lower cost and manpower.

An Efficient Alternative to Culture-based Resistance Screening

One major advantage of applying molecular biological techniques for resistance screening is the avoidance of culturing, which is time-consuming for slow-growing organisms like Mycobacteria. Several studies have demonstrated that Pyrosequencing technology is useful for rapid screening of resistance in Mycobacterium tuberculosis. Arnold et al. (2005) performed both species identification and a rapid screen for drug resistance to rifampicin and isoniazid. Since identification and resistance screening could be performed directly from sputum, they suggest that this could function as a low-cost replacement of the existing time-consuming tests based on culture.