[Molecular methods for diagnosing novel coronavirus infection: comparison of loop-mediated isothermal amplification and polymerase chain reaction].

INTRODUCTION: Currently, the basis for molecular diagnostics of most infections is the use of reverse transcription polymerase chain reaction (RT-PCR). Technologies based on reverse transcription isothermal loop amplification (RT-LAMP) can be used as an alternative to RT-PCR for diagnostic purposes. In this study, we compared the RTLAMP and RT-PCR methods in order to analyze both the advantages and disadvantages of the two approaches. MATERIAL AND METHODS: For the study, we used reagent kits based on RT-PCR and RT-LAMP. The biological material obtained by taking swabs from the mucous membrane of the oropharynx and nasopharynx in patients with symptoms of a new coronavirus infection was used. RESULTS: We tested 381 RNA samples of the SARS-CoV-2 virus (Coronaviridae: Coronavirinae: Betacoronavirus; Sarbecovirus) from various patients. The obtained values of the threshold cycle (Ct) for RT-PCR averaged 20.0 ± 3.7 s (1530 ± 300 s), and for RT-LAMP 12.8 ± 3.7 s (550 ± 160 s). Proceeding from the theoretical assumptions, a linear relationship between values obtained in two kits was proposed as a hypothesis; the correlation coefficient was approximately 0.827. At the same time, for samples with a low viral load (VL), the higher Ct values in RT-LAMP did not always correlated with those obtained in RT-PCR. DISCUSSION: We noted a significant gain in time for analysis using RT-LAMP compared to RT-PCR, which can be important in the context of testing a large number of samples. Being easy to use and boasting short turnaround time, RT-LAMP-based test systems can be used for mass screening in order to identify persons with medium and high VLs who pose the greatest threat of the spread of SARS-CoV-2, while RT-PCR-based diagnostic methods are also suitable for estimation of VL and its dynamics in patients with COVID-19.

[Detection of Treponema pallidum DNA and RNA in clinical material from patients with syphilis at different stages]. / Vyiavlenie DNK i PNK Treponema pallidum v klinicheskom materiale u patsientov s razlichnymi stadiiami sifilisa.

The results of the development and approval of methods for the detection of T. pallidum DNA and 16S rRNA in clinical material (blood plasma, serous exudates) are presented. T. pallidum DNA was detected with the use of primers to the gene coding protein with a moleculat weight of 47 kD and T. pallidum RNA, with the primers to gene 16S rRNA. The isolation, reverse transcription and amplification of DNA and RNA was carried out in the presence of inner DNA and RNA control respectively. The analytical sensitivity of the developed method was 400 DNA copies per ml. The characteristics of analytical and diagnostic specificity were 100%. The specimens of blood plasma, taken from 292 patients with syphilis at different stages before specific antibacterial therapy, were tested by the PCR. The detection rate of T. pallidum DNA and RNA in blood plasma was, respectively, 91% and 100% in primary seropositive syphilis, 68% and 79% in secondary early syphilis, 19% and 26% in latent unverified syphilis. In secondary relapsing syphilis T. pallidum DNA and RNA were detected in 92% and in latent early syphilis, in 14% of patients. T. pallidum nucleic acids were detected in 1 patient at the seronegative period of primary syphilis. No positive result was obtained in the PCR analysis in any of the patients with diagnosed seroresistance, latent late syphilis and tertiary syphilis. In the study of material taken from chancres of 11 syphilis patients the data obtained by dark-field microscopy and the PCR analysis completely coincided.