Abstract
The vancomycin, one of the family of glycopeptide antibiotics, inhibits the synthesis of bacterial cell wall peptidoglycan and has been widely used against gram-positive bacterial infections, especially for a treatment of methicillin resistant S. aureus infection. However, clinical isolate which was intermediately resistant to vancomycin (Mu50: MIC 8 $\mu\textrm{g}$/ml) was isolated in recent years. In this study we performed vancomycin susceptibility test with the increment method and population analysis with clinical isolates S. aureus. Also we did several kinds of tests with three selected isolates (s129: MIC 7 $\mu\textrm{g}$/ml, s134: MIC 7 $\mu\textrm{g}$/ml, s135: MIC 8 $\mu\textrm{g}$/ml) to find out possible mechanism of vancomycin resistance. As a result, the prevalence of vancomycin resistant S. aureus isolates among S. aureus strains resistant to methicillin was 23.3% (25/107). The vancomycin resistances of isolated strains of S. aureus were between those of Mu5O and Mu3 strains. By PCR analysis, none of the isolates with decreased vancomycin susceptibility contained known vancomycin resistant genes such as vanA, vanB, vanC1, vanC2, and vanH. Major bands of 81 kDa, 58 kDa, 33 kDa, 28 kDa were demonstrable in whole cell lysates by SDS-PAGE from all three isolates as well as reference strains. And especially,45 kDa protein was overproduced in Mu50 strains. Among them increased production of NAD$^{+}$-linked-$_{D}$-lactate dehydrogenase (dnLDH) were detected from one clinical strain (s135) and Mu5O strain. From these data, we suggest that the mechanism of vancomycin resistance in these isolates are distinct from that in enterococci.
Vancomycin은 세포벽의 합성을 억제하여 세균에 대한 항균효과를 나타내는 glycopeptide 계 항생물질로서 그람양성세균으로 인한 감염치료에 광범위하게 사용되며, 특히 methicillin 내성 포도상구균의 선택적 치료제로 쓰이고 있다. 그러나 최근 임상검체에서도 중등도의 내성을 가지는 포도상구균 (Mu50: MIC 8 $\mu\textrm{g}$/ml)이 나타나기 시작하였고 여러 가지 여건상 국내에서도 내성균주가 분리될 가능성이 높다고 사료되어 임상검체 중 methcillin 내성 포도상구균을 대상으로 vancomycin 감수성 및 내성빈도 조사를 실시하고 이에 따른 내성기전을 알아보고자 하였다. 본 실험 결과 107주 (株)의 methicillin 내성균주 중 23.3%가 vancomycin에 대하여 내성을 보였으며 vancomycin 내성을 나타내는 표준균주인 Mu50과 Mu3의 중간 정도의 내성빈도를 보였다. 중합효소 연쇄반응을 통해 장구균의 vancomycin 내성에 관여하는 vanA, vanB, vanC1, vanC2, vanH 특이 유전자는 증폭되지 않았다. SDS-PAGE를 실시하여 81 kDa, 58 kDa, 33 kDa, 28 kDa 등의 주요 단백 분획을 확인하였고, Mu50에서 45 kDa의 특징적인 단백 분획을 관찰하였다. LDH enzyme assay에서는 한 개의 검체가 Mu50과 함께 높은 LDH 활성을 보였다.
