Title : Genetic and biochemical equivalent characterization for Staphylococcus and Micrococcus NCCP resources as reference strains
Bacterial pathogens have been used during quality control while developing medicine, cosmetics, and feed in healthcare and industrial fields worldwide. Some countries including America, China, and Japan, as well as a few from Europe, have their own pharmacopoeias and they approve medical supplies that have undergone quality assurance based on sterility testing and antimicrobial susceptibility testing using the ATCC reference strains reported in the pharmacopoeias. Moreover, Korean pharmaceutical companies have been performing quality control tests specified in the Korean Pharmacopoeia (KP). They spend large periods of time and cost on the quality control of and quality certification for their candidate medicines because they use imported ATCC reference strains.
The National Culture Collection for Pathogens (NCCP), Korea collects and analyzes bacterial pathogen resources isolated from Korea and distributes them to researchers from several fields of study. We have conducted an equivalent analysis to replace ATCC reference strains with the Korean isolates in NCCP. In order to find alternative strains for Staphylococcus aureus ATCC 6538P, Staphylococcus epidermidis ATCC 12228, and Micrococcus luteus ATCC 10240, we conducted equivalent experiments with these three strains and 45 Korean strains.
First, we analyzed their molecular, biological, and biochemical properties using MALDI-TOF, VITEK-2, and antibiotic disc diffusion method. Additionally, 16S rRNA sequencing, MLST analysis, and NGS analyses were performed to investigate their molecular genetic characteristics. Korean candidates selected were Staphylococcus aureus NCCP 16830, Staphylococcus epidermidis NCCP 16828, and Micrococcus luteus NCCP 16831. They were evaluated for their usefulness by conducting an experiment applying the Microbial Assay for Antibiotics from the general test methods of the KP. The test was carried out with the Cylindrical-Plate Method. Ten replicates were performed, and the effective range was determined by obtaining the average titer of ATCC strains. NCCP strains were also tested ten times. The mean titers of the three selected NCCP strains were considered useful because they were within the effective range. In addition, whole genome sequencing was used to compare genetic homology between the ATCC and NCCP strains. S. aureus, S. epidermidis, and luteus showed 99.00%, 99.60%, and 98.10% homology between the two strains, respectively.
Through this study, the possibility of using NCCP strains to replace ATCC strains was confirmed. In addition, the characteristics of the ATCC and informative NCCP strains can be obtained and provided to the biomedical researchers.