Molecular Genetic Characterization of Staphylococcus aureus Clinical Isolates from Urinary Tract Infection Xuan, Xianglan Major in Diagnostic Laboratory Science Catholic University of Pusan : Genotyping and Antibiotic Resistance Analysis of Staphylococcus aureus Clinical Isolates from Urinary Tract Infection ,Virulence Factor Gene Profiling Based on Genotype of Staphylococcus aureus Clinical Isolates from Urinary Tract Infection = 요로 감염에서 분리된 황색 포도알균 임상 분리 균주의 분자 유전학적 특성 규명

다국어 초록 (Multilingual Abstract)

Genotyping and Antibiotic Resistance Analysis of Staphylococcus aureus Clinical Isolates from Urinary Tract Infection Xuan, Xianglan Department of Clinical Laboratory Science, the Graduate School, Catholic University of Pusan Advisor : Professor Kim, Sunghyun Ph.D. Staphylococcus aureus (S. aureus) is a common bacteria that causes mild skin infections as well as potentially deadly infections. T he main worry with S. aureus infections is the growing antimicrobial resistance (AMR) caused by methicillin-resistant S. aureus (MRSA), which is resistant to β-lactam antibiotics and multidrugs. The mecA gene is found within the Staphylococcal Cassette Chromosome mec (SCCmec), and its classification as a specific type of SCCmec is dete rmined by the size of the DNA fragment. Defining SCCmec type is c ritical since this mobile genetic element contains a variety of unique gene complexes, resulting in varying methods and levels of resistanc e. A urinary tract infection (UTI) is described as the presence of bac teria in urine, which causes infections in the urinary system, includin g the ureter, bladder, or kidney. Urine catheterization is contributes to an increase in the incidence of UTIs caused by S. aureus. The prese nt study aims to elucidate the particular patterns of MRSA clinical is olates associated with urinary tract infections, as well as describe the entire SCCmec sequence patterns and conduct an intensive whole gen ome sequencing data analysis. Epidemiology of MRSA clones is dete rmined by the unique SCCmec element found inside them. Additionall y, to supplement the previous SCCmec typing approach with whole g enome sequencing (WGS) data in order to generate comprehensive da ta, including insights on AMR for further clinical research. Therefore, compared to antimicrobial susceptibility test (AST), WGS offered a m ore comprehensive detection of AMR gene data. Clinically and epidem iologically, the isolates' pathogenicity would have been underestimated if AST had been the only method used. Consequently, in situations w here there is genetic variety or mobile element variability, WGS offer s a more thorough and accurate analysis of virulence gene transmissi on. Furthermore, pathogens like S. aureus, especially MRSA, which d eveloped into MDR, are still responsible for the global problem of anti biotic resistance. We contrasted the conventional AST approach with WGS because it has been proposed that MRSA might be linked at th e molecular level. While certain SCCmec elements in this study could not be typed using previously designed PCR primers, next generation sequencing (NGS) analysis verified that they were SCCmec IVc. Sequ ence variability within the SCCmec cassette, specifically in the joining (J) region, is assumed to be the cause of this difference. The study c onfirms that using the same primer set consistently is not feasible du e to the ongoing evolution of clinical pathogens. Consequently, it mig ht be more accurate to do WGS across time in order to gather evolut ionary information. Keywords: Staphylococcus aureus, MRSA, SCCmec, Whole genome sequencing, Antimicrobial resistance

다국어 초록 (Multilingual Abstract)

Virulence Factor Gene Profiling Based on Genotype of Staphylococcus aureus
Clinical Isolates from Urinary Tract Infection Xuan, Xianglan Department of Clinical Laboratory Science, the Graduate School, Catholic University of Pusan Advisor : Professor Kim, Sunghyun Ph.D In contrast to infections caused by other bacterial pathogens, S. aureus infections depend on a variety of virulence factors. In order to invade the host, each pathogen detects a variety of environmental signals and responds through altering the production of virulence factors, such as extracellular enzymes, poisons, and cell surface adhesions. The majority of the virulence genes of S. aureus are mobile genetic elements. Consequently, the identification of virulence factors in staphylococci has resulted in the identification of mobile genetic elements. Therefore, the aim of the present study was to identify factors linked to virulence factor genes that were expressed in S. aureus clinical isolates from urine samples. To assess whether there are any differences between the virulence characteristics of MRSA and MSSA, as well as between the virulence characteristics of the UTIs and the sepsis-obtained S. aureus from the previous study, a total of 21 virulence factor genes were detected using the qPCR SYBR Green assay. Consequently, it was established that there was not a significant distinction between the virulence factor patterns of MRSA and MSSA. In contrast, the isolates from the UTI in the study and the sepsis isolates in a prior study had different adhesion and toxin characteristics. The isolates of urinary tract infections displayed a pattern of decreased expression of systemic pathogenic markers, which is consistent with a local infection phenotype that does not require systemic virulence factors linked to significant superantigen-mediated immunity but does require strong adhesion and cytolytic capacity to survive in the urinary tract. Depending on the organ and environment from which it originates, S. aureus probably displays distinct virulence factor patterns even within the same species. This implies that virulence factor expression may be more directly impacted by environmental factors than by the mere existence or lack of antibiotic resistance.

Keywords: Virulence factors, urinary tract infection, Sepsis, S. aureus

목차 (Table of Contents)

• Ⅰ Chapter 1
• 1. Introduction 2
• 2. Materials and Methods 7
• 3. Results 26
• 4. Discussion 47
• Ⅱ Chapter II 59
• 1. Introduction 59
• 2. Materials and Methods 62
• 3. Results 66
• 4. Discussion 82
• References 84
• Abstract in Korean 95