• 대한의생명과학회지
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• 제5권1호
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• pp.95-100
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• 1999

일반적으로 임상검사실에서 vancomycin resistant enterococci (VRE)를 검출하는 일은 어렵고, 시간이 많이 들며, 검체처리 비용도 많이 든다. 따라서 본 실험은 임상검체에서 분리된 세균으로부터 VRE를 신속하게 확인하고, 진단하기 위한 방법으로서 다중 중합효소 연쇄반응을 확립하였다. 본 실험에 사용된 primer는 장구균에 특이 한 유전자인 vanA, vanB, vanC-1, vanC-2/3 각각의 염기서열을 기초로 primer를 제작하고, 다중 중합효소 연쇄 반응을 실시하여 임상검체로부터 분리된 VRE 유전자의 type및 분포율을 조사하고자 하였다. 국내에서 분리된 75주의 장구균을 대상으로 다중 중합효소 연쇄반응을 실시한 결과 36주의 분리균주에서 vancomycin에 대해 높은 저항성을 보이는 vanA 유전자를 가진 것으로 나타났다. 그리고 18주에서는 vancomycin에 낮은 저항성을 내성을 보이는 vanC-1 또는 vanC-2/3 유전자를 보유한 것으로 나타났다. 따라서 본 실험에서 확립한 다중 중합효소 연쇄 반응 기법은 신속한 VRE 진단 방법으로 이용할 수 있을 것이다.

• 대한의생명과학회지
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• 제26권3호
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• pp.149-156
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• 2020

We developed a simple and rapid method for detecting vancomycin resistance genes, such as vanA and vanB, using loop-mediated isothermal amplification (LAMP). To identify not only vancomycin resistance genes, but also the genus Enterococcus, primers were designed for vanA, vanB, and 16S rRNA. Screening for vancomycin susceptibility in Enterococcus was performed using Etest (bioMérieux Inc). The results of the LAMP assay were compared to those of real-time RT-PCR. The optimal conditions for the LAMP assay were 65℃ for 60 min. The detection limits of the LAMP assay for vanA, and vanB were 2 × 10² copies/reaction. Compared to RT-PCR, the sensitivities and specificities of LAMP for 16S rRNA, vanA, and vanB were 100/100%, 100/100%, and 100/100%, respectively. The vanA genotype-vanB phenotype accounted for 57.5% (46/80) of the vancomycin-resistant Enterococci samples collected from 2016 to 2019. In conclusion, the LAMP assay developed in this study showed high sensitivity and specificity for vancomycin-resistant genes. Moreover, due to the simplicity and rapidity of the LAMP assay, its use can be very useful in clinical microbiology laboratories.

• 대한수의학회지
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• 제39권2호
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• pp.343-352
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• 1999

Vancomycin resistant enterococci (VRE) have emerged as an important nosocomial pathogen. Since 1989 the Center for Disease Control, United States, has reported a rapid increase in the incidence of enterococcal bacteremia and endocarditis infection by VRE. It was suggested that the use of avoparcin was associated with the appearance of VRE in animal husbandry. To date, several detection methods have been used based on conventional methods of culture and gene detection. However, these methods have some limitations such as time-consuming, laborious and additional differential needs. Therefore, In this study a multiplex PCR method was established to detect and differentiate resistance types of enterococci which specifically amplify the four van genes encoding vancomycin resistance elements. Using the method, we investigated the incidence rates and types of VRE from farms using or not using avoparcin. A total of 1091 animal fecal samples were collected from 70 pig and 32 poultry farms. A total of 425 of enterococci were isolated from samples. Of the 425 isolates, 11 of the them showed a pattern of high-level vancomycin resistance (MIC : $64{\sim}256{\mu}g/ml$) which was associated with the presence of the vanA or vanB gene. Fifty-seven isolates showed a pattern of low-level vancomycin resistance (MIC : $3{\sim}8{\mu}g/ml$) associated with the vanC-1 or vanC-2 gene. Interestingly, all isolates with high-level vancomycin resistance were from farms that have never used avoparcin. Moreover, the high-level VRE isolation rate in Korea (2.58%) was much lower than that of other countries (50% in England, 7% in Belgium) where avoparcin have been used. In conclusion, the multiplex PCR method established in this study could be applied for detection of VRE.

• 대한임상검사과학회지
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• 제45권1호
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• pp.16-20
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• 2013

Outbreaks of vancomycin-resistant enterococci (VRE) are being reported more frequently in many countries. While seven glycopeptide resistance genotypes have been described in Enterococci, vanA and vanB are the most common resistance genotypes. The aim of this study was to detect antibiotic susceptibilities of 23 Enterococcus faecium strains, which caused an outbreak in a University hospital by a disk diffusion test to investigate the presence of the species specific gene, and the resistant genotypes, vanA and vanB by duplex PCR. PCR for vanA and vanB was performed on 23 enterococci. Twenty three were identified as E. faecium and were tested positive for the vanA genotype. This study will report on the validation of a simple and accurate VRE detection method that can be easily incorporated into the daily routine of a clinical laboratory. Early detection of VRE strains, including those with susceptibility to vancomycin, is of paramount clinical importance as it allows rapid initiation of strict infection control practices, as well as the therapeutic guidance for confirmed infections. The PCR method developed in the present study is simple and reliable for the rapid characterization of VRE.

• 대한임상검사과학회지
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• 제45권1호
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• pp.1-4
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• 2013

Vancomycin-resistant enterococci (VRE) have emerged as important healthcare-associated infection since last two decades. ChromID VRE agar (cIDVA) is useful for VRE rectal swab screening. We investigated all VRE were isolated on the cIDVA. A total of 363 rectal swabs of 85 patients to test VRE screening were inoculated into bile-esculin (B-E) broth with $6{\mu}g/mL$ vancomycin. After 24 hours incubation, we subcultured B-E broths were changed to black onto cIDVA. All isolates were identified by the MICROSCAN and VITEK2. The vanA gene and vancomycin minimal inhibition concentration (MIC) were detected by PCR and E-test respectively. 277 E. faecium (84.7%), 16 E. faecalis (4.9%), 25 E. avium (7.6%), 8 E. gallinarum (2.4%) and 1 E. raffinosus (0.3%) were isolated. 10.3% of VRE detected on cIDVA were other than E. faecium and E. faecalis that presented various color from colorless to pale violet. All isolates contained vanA and vancomycin MIC were > $256{\mu}g/mL$. VRE isolates other than E. faecium and E. faecalis should be objective to the contact precautions for healthcare-associated infection control if they possess vanA gene. Due to emerging enterococci carrying vanA such as E. avium, E. gallinarum, and E. raffinosus, VRE surveillance should be expanded to all isolates on chromogenic agar.

• 한국축산식품학회지
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• 제33권1호
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• pp.133-138
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• 2013

본 연구에서는 서울시내 256곳의 판매점에서 구입한 축산물 및 수산물에서 Enterococcus faecalis 와 Enterococcus facium을 분리하였으며 분리된 균주의 항생제 내성양상과 vancomycin 내성 유전자 보유여부를 검증하였다. 총 256개 시료 중 117개에서 E. faecalis(40.6%)와 E. faecium(5.1%)가 검출되어 45.7%의 분리율을 나타내었다. 축산물은 192개 중 105개 균주가 분리되어 54.7%의 분리율을 나타내었는데 닭고기에서 가장 높은 68.8%의 분리율을, 돼지고기에서 50.0%의 분리율을, 쇠고기에서 45.3%의 분리율을 나타내었다. 횟감어류에서는 18.8%의 분리율을 나타내었다. 분리된 균주에 대한 항생제 내성 양상은 10종의 항생제 디스크를 이용하여 검증하였다. Tetracycline의 내성률이 52.1%로 가장 높았으며, erythromycin의 내성률이 27.4%로 두 번째로 높게 나타났다. Ampicillin과 penicillin는 1개의 균주를 제외하고는 모두 감수성을 보였으며, amoxicillin & clavulanic acid에는 모든 균주가 감수성을 보였다. Vancomycin에는 모든 균주가 감수성을 보여 VRE는 검출되지 않았다. 분리된 균주의 vancomycin 내성유전자의 검출은 multiplex PCR을 이용하여 vanA gene과 vanB gene 보유여부를 확인하였다. vanA gene이 검출된 균주는 없었으나, vancomycin에 감수성을 나타내던 9개의 균주에서 vanB gene이 검출되어 VRE균 출현의 잠재적인 가능성을 보여주었다.

• Journal of Microbiology
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• 제39권2호
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• pp.116-120
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• 2001

The prevalence resistance genotype and antibiotic susceptibility of vancomycin-resistant enterococci (VRE) were determined. Prevalence of VRE in chickens, healthy children and intensive care unit (ICU) patients was 43.0%, 12.7% and 24.1%, respectively. Forty out of 56 isolates from chicken intestines were identified as Enterococcus faecium, and 12 were E. faecalis. All the isolates contained the vanA gene. Nine out of 13 VRE isolates from patients and two out of 21 from healthy young children were identified as E. faecium. The resistance types of E. faecium, E. gallinarium and E. casseliflavus were VanA, VanCl, and VanC2, respectively. The minimum inhibitory concentrations (MICs) of E. faecium, E. gallinarium, and E. casseliflavus to vancomycin were 512,8 and 4 g/ml, respectively. Specifically, E. faecium isolates were resistant to most of antibiotics except ampicillin and gentamicin. This is the first report of high VanA type VRE prevalence in nonhospitalized young children in Korea.

• 한국작물학회:학술대회논문집
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• 한국작물학회 2006년도 한국약용작물학회 공동춘계학술발표회
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• pp.80-81
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• 2006

• 대한수의학회지
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• 제43권1호
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• pp.103-112
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• 2003

A multiplex PCR assay, which allows simultaneous detection of vancomycin resistant genotypes and Enterococcus species-specific genes, was developed. Vancomycin resistant enterococci (VRE) from chickens and humans could be detected for vanA, vanB, vanC-1, vanC-2, $ddl_{E.faecium}$ and $ddl_{E.faecalis}$ by multiplex PCR. Eight isolates of VRE from humans (n=11) had $ddl_{E.faecium}$ and vanA, and 3 isolates of the VRE had $ddl_{E.faecium}$ and vanB. One isolate of VRE from chickens (n=6) had $ddl_{E.faecium}$ and vanA, and 5 isolates of the VRE had only vanA. E. faecium, E. faecalis, E. gallinarum and E. casseliflavus were also confirmed for the species-specific gene by multiplex PCR. This multiplex PCR could detect E. faecium, E. faecalis, E. gallinarum, E. casseliflavus, vanA, vanB, vanC-1 and vanC-2, simultaneously. The PCR assay established in the present study can be an alternative to time-consuming biochemical tests and antibiotic susceptibility tests of Enterococcus spp.

• Journal of Microbiology
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• 제45권5호
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• pp.466-472
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• 2007

A total of 98 vancomycin-resistant Enterococcus faecium (VREF) isolates (58 isolates from patients and 40 isolates from poultry) were compared based on their antimicrobial susceptibility, Tn1546 element organization, and pulsed-field gel electrophoresis (PFGE) patterns. This comparison aided in determining the relationships between the groups of isolates. All the VREF isolates harbored the vanA gene; however, 29 (29.6%) of the isolates exhibited the VanB phenotype-vanA genotype. Furthermore, the VREF isolates from humans and poultry exhibited distinct antimicrobial resistance patterns. The PCR mapping of the Tn1546 elements exhibited 12 different transposon types (A to L). The VREF isolates of poultry were classified into types A to D, whereas the human isolates were classified into types E to L. A PFGE analysis demonstrated a high degree of clonal heterogeneity in both groups of isolates; however, the distinct VREF clones appeared in each group of isolates. The deletion of the vanX-vanY genes or insertion of IS1216V in the intergenic region from the vanX-vanY genes is directly associated with the incongruence of the VanB phenotype-vanA genotype in human VREF isolates. These data suggest that the VREF isolates exhibit distinct phenotypic and genotypic traits according to their origins, which suggests that no evidence exists to substantiate the clonal spread or transfer of vancomycin resistance determinants between humans and poultry.