Evaluation of an automated extraction method for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae by Cobas Amplicor PCR from different sample materials

Abstract

A commercially available automated device (MagNA Pure LC) was adapted for nucleic acid extraction of urogenital specimen for subsequent PCR detection of Chlamydia trachomatis and Neisseria gonorrhoeae in a clinical laboratory. Results were compared to the standard manual extraction procedure and showed excellent correlation, with even slightly increased sensitivity.

First evaluation of six nucleic acid amplification tests widely used in the diagnosis of Chlamydia trachomatis in Russia

“…This is probably due to the fact that MagNA Pure LC was used for automated DNA isolation, which has previously been shown to increase the sensitivity and significantly lower the rate of inhibition. 13,14,16,17 However, in the present study, the Russian NAATs that used silica-based nucleic acid isolation techniques had significantly lower inhibition rates. Moreover, in the NAATs developed by the Central Research Institute of Epidemiology (conventional and real-time PCR, as well as realtime NASBA), the ICs are added before the nucleic acid isolation that is highly recommended, in order to examine both the inhibition of amplification and the efficiency of DNA/RNA extraction.…”

“…Nevertheless, using the reference methods, none of the samples was inhibited. This is probably due to the fact that MagNA Pure LC was used for automated DNA isolation, which has previously been shown to increase the sensitivity and significantly lower the rate of inhibition 13,14,16,17 . However, in the present study, the Russian NAATs that used silica‐based nucleic acid isolation techniques had significantly lower inhibition rates.…”

“…MagNA Pure LC (Roche Molecular Biochemicals, Mannheim, Germany), instead of manual preparation. Nevertheless, both these modifications have been effectively used and evaluated in previous studies and, especially, the latter one can certainly be an optimization that results in an increased sensitivity, substantially decreased rate of amplification inhibition, less labour intensive, etc 13–17 …”

“…Nevertheless, both these modifications have been effectively used and evaluated in previous studies and, especially, the latter one can certainly be an optimization that results in an increased sensitivity, substantially decreased rate of amplification inhibition, less labour intensive, etc. [13][14][15][16][17] The aim of the present study was to evaluate the performance characteristics of five PCRs and a recently introduced nucleic acid sequence-based amplification (NASBA) assay currently used to diagnose C. trachomatis infection in Russia. The following NAATs were evaluated: conventional and real-time PCRs developed at the DNA Technology company, Moscow, Russia (cPCR-DT and rtPCR-DT); conventional PCR developed at the Lytech company, Moscow, Russia (cPCR-Ly); conventional and real-time PCRs developed at the Central Research Institute of Epidemiology, Moscow, Russia (cPCR-Ep and rtPCR-Ep); and real-time NASBA assay based on NucliSens Technology (bioMérieux, Boxtel, the Netherlands), with the target-specific oligonucleotides developed at the Central Research Institute of Epidemiology (Table 1).…”