• Biomedical Science Letters
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• v.26 no.3
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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.

• The Korean Journal of Physiology and Pharmacology
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• v.25 no.4
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• pp.321-331
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• 2021

Vancomycin, an antibiotic used occasionally as a last line of treatment for methicillin-resistant Staphylococcus aureus, is reportedly associated with nephrotoxicity. This study aimed at evaluating the protective effects of lutein against vancomycin-induced acute renal injury. Peroxisome proliferator-activated receptor gamma (PPARγ) and its associated role in renoprotection by lutein was also examined. Male BALB/c mice were divided into six treatment groups: control with normal saline, lutein (200 mg/kg), vancomycin (250 mg/kg), vancomycin (500 mg/kg), vancomycin (250 mg/kg) with lutein, and vancomycin (500 mg/kg) with lutein groups; they were euthanized after 7 days of treatment. Thereafter, samples of blood, urine, and kidney tissue of the mice were analyzed, followed by the determination of levels of N-acetyl-β-D-glucosaminidase (NAG) in the urine, renal creatine kinase; protein carbonyl, malondialdehyde, and caspase-3 in the kidney; and the expression of PPARγ, nuclear factor erythroid 2-related factor 2 (Nrf2), and nuclear factor-kappaB (NF-κB) in renal tissue. Results showed that the levels of protein carbonyl and malondialdehyde, and the activity of NAG, creatine kinase and caspase-3, were significantly increased in the vancomycin-treatment groups. Moreover, the levels of Nrf2 significantly decreased, while NF-κB expression increased. Lutein ameliorated these effects, and significantly increased PPARγ expression. Furthermore, it attenuated vancomycin-induced histological alterations such as, tissue necrosis and hypertrophy. Therefore, we conclude that lutein protects against vancomycin-induced renal injury by potentially upregulating PPARγ/Nrf2 expression in the renal tissues, and consequently downregulating the pathways: inflammation by NF-κB and apoptosis by caspase-3.

• Korean Journal of Clinical Pharmacy
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• v.9 no.1
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• pp.1-14
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• 1999

Vancomycin-resistant Enterococci (VRE) have recently emerged in Korean hospitals, as well as in those of other countries. VRE have been partially attributed to the overuse and misuse of vancomycin. The mecbanisms of VRE resistance are related to VanA, VanB, and VanC. Both VanA and VanB produce abnormal ligase enzymes to form D-ala-D-lactate termini in E. faecium and E. faecalis, instead of D-ala-D-ala termini. Meanwhile, Van C produces D-ser-D-ala termini in E. gallinarum and E. casseliflavus. These abnormal termini have a low affinity to vancomycin. As a result, VRE avoid the activity of vancomycin by these mechanisms. Unfortunately, there is no approved therapy for the treatment of VRE. Thus, available but uncommonly prescribed antibiotics (due to their toxicity or unproven efficacy) may become possible options. They include chloramphenicol, novobiocin, fosfomycin, and bacitracin. The combination therapy of available agents may also be the other options. They include high doses of a penicillin- or ampicillin-aminoglycoside combination, high doses of an ampicillin/sulbactam and aminoglyoosidcs combination, an ampicillin and vancomycin combination, and a ciprofloxacin, aminoglycosides, and rifampin combination. With respect to the near future, many types of investigational agents will most likely expand their treatment options for VRE. Teicoplanin, a glycopeptide, can be used for VanB- and VanC-related VRE. LY333328, a new generation of glycopeptide, is effective in treating VanA as well as VanB and VanC. RP59500 (quinupristin/dalfopristin), a streptogramin, is effective in treating vancomycin-resistant E. faecium. New generation quinolones (especially clinatloxacin) are potential options for the treatment of VRE, even though they cannot work as effectively against VRE as they can against Staphylococci. Both glycylcyclines (a new generation of tetracyclines) and ketolides (a new generation of macrolides) show good activity against Enterococci, regardless of vancomycin susceptibility. Oxazolidinones (i. e. eperezolid and 1inezolid) and everninomicins (i. e. SCH27899) are new groups of antibiotics, which also demonstrate good activity against VRE. It is imperative that clinical pharmacists take the responsibility of investigating new treatment options for VRE in order to combat this growing problem throughout the world.

• Biomedical Science Letters
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• v.6 no.3
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• pp.201-208
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• 2000

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.

• Korean Journal of Clinical Laboratory Science
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• v.45 no.1
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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.

• Korean Journal of Clinical Laboratory Science
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• v.45 no.1
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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.

• Food Science and Biotechnology
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• v.15 no.4
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• pp.511-515
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• 2006

Antibiotic resistance of foodborne pathogens adapted to garlic (Allium sativum Linn.) was determined in order to understand the relationship between antibiotic resistance and garlic. The Gram (-) strains of Escherichia coli and Salmonella typhimurium and the Gram (+) strains of Bacillus cereus and Staphylococcus aureus were subcultured consecutively in a garlic broth, and the surviving colonies on the agar were selected as the adapted strains. Minimal inhibitory concentrations (MIC) for 15 antibiotics on the adapted strains were determined on Muller-Hinton Infusion agar. Adaptation to 1.3%(v/v) garlic juice increased MIC for vancomycin, aminoglycoside, and erythromycin on B. cereus, and for ampicillin and erythromycin on E. coli O157:H7. MIC of aminoglycosides, chloramphenicol, and vancomycin on the adapted S. aureus increased. The adapted S. typhimurium was more resistant to penicillin and vancomycin than the non-treated strain. The adapted S. typhimurium and S. aureus lost their antibiotic resistance in non-garlic stress conditions. However, the adapted B. cereus was still resistant to erythromycin and vancomycin, and the adapted E. coli was also resistant to erythromycin. Antibacterial garlic might increase the antibiotic resistance of E. coli, B. cereus, S. aureus, and S. typhimurium and this resistance can continue even without the stress of garlic. Therefore, garlic as a food seasoning could influence the resistance of such pathogens to these antibiotics temporarily or permanently.

• Biomedical Science Letters
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• v.5 no.1
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• pp.95-100
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• 1999

It is generally difficult, time-consuming, and expensive for the clinical laboratory to detect vancomycin resistant enterococci (VRE). The aim of this study was to develop and evaluate the multiplex polymerase chain reaction (PCR) assay system as a diagnostic tool for the rapid detection of VRE from clinical samples and/or for the identification of VRE from the bacterial strains isolated from clinical specimens. Specific primers, designed from the nucleotide sequences respectively encoding the vanA, vanB, vanC-1, vanC-2/3 genes in enterococci, were coupled in a multiplex PCR assay system. With this multiplex PCR assay system, we investigated the incidence rates and types of VRE isolated from clinical samples. A total of 75 strains of enterococci were isolated in 3 general hospitals in Korea. Of these isolates, 36 strains showed a pattern of high-level vancomycin resistance which associated with vanA gene, whereas 18 strains showed low-level vancomycin resistance associated with vanC-1 or vanC-2/3 gene. Thus, multiplex PCR assay method established in this study could be applied for the rapid detection of VRE.

• Korean Journal of Clinical Laboratory Science
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• v.46 no.2
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• pp.64-67
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• 2014

Vancomycin-resistant Enterococci (VRE) are a leading cause of a nosocomial infection. While seven glycopeptide resistance genotypes have been found in Enterococci, vanA and vanB are the most common resistance genotypes. Aims of this study were to detect antibiotic susceptibilities of 23 Enterococcus spp, which broke out in a university hospital by the disk diffusion test, to investigate specific genes of vanA and vanB by conventional and real-time PCR. PCR for vanA and vanB was performed on 23 Enterococci, all 23 were positive for vanA type. This study reports the validation of a simple and rapid 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 a rapid initiation of strict infection control practices as well as a therapeutic guidance for a confirmed infection. The real-time PCR method is a rapid technique to detect vanA in Enterococci. It is simple and reliable for the rapid characterization of VRE.

• Korean Journal of Clinical Laboratory Science
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• v.49 no.4
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• pp.401-406
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• 2017

In this study, using the VITEK 2 system, 74 samples (22.6%) out of 327 specimens were identified by the growth of Enterococcosel media (EV6 agar) supplemented with $6{\mu}g/mL$ of vancomycin. Enterococcus faecium was identified as 55 strains (74.3%), Enterococcus casseliflavus as 2 strains (2.7%), Enterococcus avium as 1 strain (1.4%), and Enterococcus gallinarum as 16 strains (21.6%). Among the 55 phenotypes of Enterococcus faecium, 42 (76.4%), 9 (16.4%), and 4 strains (7.3%) showed the vanA, vanB, and vanC phenotype, respectively. The 16 strains of Enterococcus gallinarum and 2 strains of Enterococcus casseliflavus showed the vanC phenotype and the 1 strain of Enterococcus avium had the vanB phenotype. The one strain of Enterococcus faecium propagated only in EV4 and was susceptible to both vancomycin and teicoplanin according to the antimicrobial susceptibility test using the VITEK 2 system. The vancomycin resistance phenotype gene was not detected by PCR. A total of 327 specimens were cultured in Enterococcosel broth supplemented with $6{\mu}g/mL$ of vancomycin (EV6 broth), and 120 strains (36.7%) were isolated. These 120 strains were subjected to vancomycin resistant genotyping by a multiplex real-time polymerase chain reaction and 51 strains (42.5%) showed vanA; 5 strains (4.2%) showed vanA and vanC; and 18 strains (15%) showed vanC. Vancomycin resistance genotypes were not detected in the remaining 46 strains (38.3%).