Panel of primers for evaluation of antibiotic resistance genes with real time detection of results

Abstract

Introduction. The problem of the spread of antibiotic resistance among pathogens of nosocomial infections is becoming increasingly important. In order to improve microbiological monitoring, it is advisable to use methods that allow you to quickly determine the largest number of antibiotic resistance determinants. In this regard, it seems relevant to develop PCR test systems for detecting antibiotic resistance genes, in particular, for screening hospitalized patients and identifying cases of nosocomial infection.Purpose. Develop a panel of primers for detection of antibiotic resistance genes using PCR with real-time visualization of results. Materials and methods. For the design of specific primers and probes, reference sequences from the databases of the National Center for Biotechnology Information (NCBI) were used. Primer selection and specificity assessment were performed using the NCBI PrimerBlast and Primer3 programs. Primers for PCR amplification of a number of antibiotic resistance genes were tested on strains of Klebsiella pneumoniae. These strains were isolated from the clinical material of intensive care patients with COVID-19 and whole genome sequencing of the strains was carried out with a detailed assessment of the resistome, virulome. The results of whole genome sequencing and multiplex real-time PCR were compared.Results. We have developed a set of primers for the detection of antibiotic resistance genes, including carbapenemase, using real-time multiplex PCR.Conclusion. The developed panel of primers can be used to screen Klebsiella pneumoniae isolates for the presence of resistance genes; further expansion of the spectrum of detected genes and testing of the panel on clinical material is required.