DEVELOPMENT OF PRIMER SYSTEM AND PROBE FOR IDENTIFICATION OF STAPHYLOCOCCUS AUREUS BY PCR-RV

The article presents the results of the development and testing of original system of primers and a probe for the identification of bacteria of the Staphylococcus aureus species by polymerase chain reaction in «real time» mode. The extension of domestic poultry industry opens up opportunities for the incidence rate of zoonoses. Staphylococci are one of the leading pathogens of bacterial infections in birds, and Staphylococcus aureus causes a wide range of diseases in chickens, including septic arthritis, subcutaneous abscesses and gangrenous dermatitis. Therefore, the development of methods for the timely detection of pathogenic variants of Staphylococcus aureus is an important task of food safety. For the development of a molecular genetic identification system, a unique DNA region of Staphylococcus aureus K39 NODE_3_length_262400_cov_12.378: 113,355..114,623 bp encoding transcription regulator U32 family peptidase was selected. With the help of the BLAST-primer NCBI resource, the UGENA program and the Oligoevaluator resource, the selection and design of oligonucleotide primers and a fluorescent probe for PCR were carried out. Optimization of primer systems showed that the optimal concentration is 3 pM of each primer per reaction, and an increase in the concentration of primers does not affect the effectiveness of the reaction. It has been experimentally established that the optimal concentration of the fluorescent probe in the reaction is 0.4 pM. Protocols have been selected for PCR-RV both in the presence of intercalating dye SYBR Green and using a fluorescent probe.The species specificity of selected primer systems was confirmed experimentally on 9 strains of heterologous bacterial genera and amounted to 100%. The sensitivity of primer system and probe was 102 copies.