BACKGROUND: Carbapenemase-producing Enterobacteriaceae (CPE) has been increasingly reported worldwide in the past 10 years, which is an important infection control concern. Since the epidemiology and characteristics of these CPEs vary according to institutes, we aimed to characterize CPEs in a university hospital during the recent 4 years. METHODS: From October 2011 to September 2015, CPE isolates from clinical specimens and hospital surveillance cultures were collected. Carbapenem resistance was confirmed by disk diffusion method and Minimal Inhibitory Concentration (MIC) was determined by agar dilution method. Carbapenemase production was tested by double disk test using aminophenylboronic acid and dipicolic acid. PCR and sequence analysis were performed to detect bla(KPC), bla(IMP-1), bla(VIM-2), bla(NDM-1)-like genes and bla(OXA-48) gene. Pulsed-field gel electrophoresis (PFGE) and Multilocus sequence typing (MLST) were conducted for KPC-producing Klebsiella pneumoniae isolates. RESULTS: Twenty-five isolates (11%) of CPE were identified among 222 carbapenem-resistant Enterobacteriacae isolates during the study period. The most prevalent CPE was KPC-producing K. pneumonia and others were IMP-1, VIM-2, NDM-1 type and OXA-48 producing CPEs. Most of these CPEs showed resistance to carbapenems with variable MICs. The sequence types (STs) of KPC-producing K. pneumoniae were ST307 and ST11. The PFGE of ST11 and ST307 showed clonality in each group suggesting the possibility of in-hospital outbreak. CONCLUSION: The prevalence of CPE has been increasing. In our institute, KPC-producing K. pneumoniae was the most frequently isolated CPE in the recent 4 years. CPE including KPC producers can easily transfer their resistance. Therefore continuous monitoring and more intensified infection control for CPE should be considered.
Academies and Institutes ; Agar ; Carbapenems ; Diffusion ; Drug Resistance, Bacterial ; Electrophoresis, Gel, Pulsed-Field ; Enterobacteriaceae* ; Epidemiology ; Infection Control ; Klebsiella pneumoniae ; Korea* ; Methods ; Molecular Epidemiology* ; Multilocus Sequence Typing ; Pneumonia ; Polymerase Chain Reaction ; Prevalence ; Sequence Analysis

BACKGROUND: Increasing numbers of Acinetobacter spp. resistant to multiple drugs, including carbapenem, has been a serious problem. The aims of this study were to determine carbapenem resistance patterns and mechanisms, as well as to study the molecular epidemiology of Acinetobacter spp. METHODS: Clinical isolates of Acinetobacter spp. were collected from May to November in 2006. Antimicrobial susceptibility testing was performed using CLSI disk diffusion and agar dilution methods. Metallo-beta-lactamase- and OXA carbapenemase-producing isolates were detected by PCR. Carbapenem resistance and hydrolytic activities were compared according to OXA type and presence of ISAba1. Pulsed-field gel electrophoresis (PFGE) was performed to determine the epidemiologic features. RESULTS: The imipenem non-susceptible rates were variable from 10% to 67%. Among 151 isolates carrying bla(OXA-51-like), 75 isolates carried both bla(OXA-51-like) and ISAba1, and 25 isolates had both bla(OXA-51-like), bla(OXA-23-like), and ISAba1. Carbapenem MICs of both bla(OXA-51-like) and ISAba1-carrying isolates were higher than those with bla(OXA-51-like) only. Carbapenem MICs of bla(OXA-23-like)-carrying isolates were higher than those with both bla(OXA-51-like) and ISAba1. Both bla(OXA-51-like) and ISAba1-carrying isolates and blaOXA-51-like, blaOXA-23-like, and ISAba1-carrying isolates demonstrated higher hydrolysis activities in oxacillin and carbapenems. Most of the tested isolates were susceptible to tigecycline, and all of them were susceptible to colistin. Pulsed-field gel electrophoresis suggested that there had been several outbreaks of bla(OXA-23-like) and bla(OXA-51-like)-positive strains. CONCLUSION: Carbapenem non-susceptible Acinetobacter isolates and OXA carbapenemase-producing isolates were prevalent. Dissemination of bla(OXA)-harboring isolates may make it difficult to treat infections due to carbapenem-resistant Acinetobacter spp. Further surveillance studies are required to prevent the spread of carbapenem resistance.
Acinetobacter ; Agar ; Carbapenems ; Colistin ; Diffusion ; Disease Outbreaks ; Electrophoresis, Gel, Pulsed-Field ; Hydrolysis ; Imipenem ; Lifting ; Minocycline ; Molecular Epidemiology ; Oxacillin ; Oxytocin ; Polymerase Chain Reaction

BACKGROUND: Avoparcin, cross-resistance with vancomycin, was added as feed-additive since 1970s and was prohibited in 1997 in Korea. After avoparcin was banned we examined prevalence and genetic relatedness of VRE in enterococci isolated from livestock and humans. MATERIALS AND METHODS: Using enrichment broth and 6 microgram/mL vancomycin-containing enterococcosel selective agar, vancomycin-resistant enterococci (VRE) were isolated from fecal sample of 255 pigs of 8 farms, 431 chickens of 9 farms, and 328 humans (Food industry employee and Institution cafeteria employee) of 5 public health centers, and 100 raw chicken meats from April to June 2003. Antimicrobial susceptibility was examined by disk diffusion and minimum inhibitory concentrations (MICs), and E-test. Species identification and genotyping were done by multiplex PCR method. Pulsed-field gel electrophoresis (PFGE) of vanA-type VRE isolates was performed by CHEF-Mapper system. RESULTS: 19 isolates from 255 pigs, 122 isolates from 431 chickens, 19 isolates from 100 raw chicken meat, and 7 isolates from 328 humans were resistant to vancomycin. Of the 167 VRE isolates, vanA gene was detected in 141 isolates; 1 isolate (0.4%) in pigs, 121 isolates (28.1%) in chickens, 18 isolates (18.0%) in raw chicken meat, and 1 isolate (0.3%) in humans. Resistant rates of streptomycin, tetracycline, and erythromycin were over 60% in vanA-type E. faecium isolated from poultry. PFGE analysis resulted in two major patterns, F and P types. Also PFGE pattern of 1 VRE from human was identical to that of 1 VRE from poultry. CONCLUSION: Despite the high prevalence of vanA-type VRE in poultry farms, VRE isolation rate in human was relatively low. This result suggests that the possibility of VRE transmission from poultry to human is low but that possibility may be not ruled out. In PFGE analysis showing 51.5% identical in 2 PFGE patterns, the dissemination of VRE isolates in poultry may be transmitted vertically and horizontally.
Agar ; Chickens ; Diffusion ; Electrophoresis, Gel, Pulsed-Field ; Enterococcus faecium ; Erythromycin ; Humans* ; Korea* ; Livestock* ; Meat ; Microbial Sensitivity Tests ; Multiplex Polymerase Chain Reaction ; Poultry ; Prevalence* ; Public Health ; Streptomycin ; Swine ; Tetracycline ; Vancomycin

BACKGROUND: Avoparcin, cross-resistance with vancomycin, was added as feed-additive since 1970s and was prohibited in 1997 in Korea. After avoparcin was banned we examined prevalence and genetic relatedness of VRE in enterococci isolated from livestock and humans. MATERIALS AND METHODS: Using enrichment broth and 6 microgram/mL vancomycin-containing enterococcosel selective agar, vancomycin-resistant enterococci (VRE) were isolated from fecal sample of 255 pigs of 8 farms, 431 chickens of 9 farms, and 328 humans (Food industry employee and Institution cafeteria employee) of 5 public health centers, and 100 raw chicken meats from April to June 2003. Antimicrobial susceptibility was examined by disk diffusion and minimum inhibitory concentrations (MICs), and E-test. Species identification and genotyping were done by multiplex PCR method. Pulsed-field gel electrophoresis (PFGE) of vanA-type VRE isolates was performed by CHEF-Mapper system. RESULTS: 19 isolates from 255 pigs, 122 isolates from 431 chickens, 19 isolates from 100 raw chicken meat, and 7 isolates from 328 humans were resistant to vancomycin. Of the 167 VRE isolates, vanA gene was detected in 141 isolates; 1 isolate (0.4%) in pigs, 121 isolates (28.1%) in chickens, 18 isolates (18.0%) in raw chicken meat, and 1 isolate (0.3%) in humans. Resistant rates of streptomycin, tetracycline, and erythromycin were over 60% in vanA-type E. faecium isolated from poultry. PFGE analysis resulted in two major patterns, F and P types. Also PFGE pattern of 1 VRE from human was identical to that of 1 VRE from poultry. CONCLUSION: Despite the high prevalence of vanA-type VRE in poultry farms, VRE isolation rate in human was relatively low. This result suggests that the possibility of VRE transmission from poultry to human is low but that possibility may be not ruled out. In PFGE analysis showing 51.5% identical in 2 PFGE patterns, the dissemination of VRE isolates in poultry may be transmitted vertically and horizontally.
Agar ; Chickens ; Diffusion ; Electrophoresis, Gel, Pulsed-Field ; Enterococcus faecium ; Erythromycin ; Humans* ; Korea* ; Livestock* ; Meat ; Microbial Sensitivity Tests ; Multiplex Polymerase Chain Reaction ; Poultry ; Prevalence* ; Public Health ; Streptomycin ; Swine ; Tetracycline ; Vancomycin

BACKGROUND: Enterococcal infections have become extremely difficult to manage because of an increase in antibiotic resistance among enterococci. In Europe, the use of avoparcin in animals was reported to be the cause of vancomycin-resistant enterococci (VRE) transmission to humans. In this study, we performed antimicrobial susceptibility testing and pulsed-field gel electrophoresis (PFGE) to characterize the genetic relatedness of VRE of human and chicken. METHODS: Ninety strains of VRE were isolated from clinical specimens in three University hospitals located in Seoul and Kyungi province in 2001-2002. Thirty isolates of VRE were collected from four chicken farms located in areas remotely distanced from each other. The isolates were identified to the species level by conventional biochemical tests and commercial kits. Antimicrobial susceptibilities were tested by the NCCLS disk diffusion and agar dilution methods. For a molecular epidemiologic analysis, PFGE was performed. RESULTS: Among the 90 clinical isolates were 73 vancomycin-resistant Enterococcus faecium (VREFM) and 17 vancomycin-resistant E. faecalis (VREFA). The resistant rates of VREFA to ampicillin, levofloxacin and tetracycline were 0%, 100%, and 100%, respectively, and for VREFM, 100%, 96%, and 26%, respectively. However, the resistant rates of VREFM isolated from chicken were 19% to ampicillin, 0% to levofloxacin, and 100% to tetracycline. The PFGE patterns of genomic DNA of the clinical isolates were very diverse, suggesting a polyclonal spread of VRE, although some isolates had an identical PFGE pattern, indicating a mini-outbreak due to a clonal spread. The PFGE patterns of genomic DNA of the chicken isolates were very different from those of the human isolates. CONCLUSIONS: VRE isolates from human and chicken showed very different antimicrobial susceptibilities and PFGE patterns. These results suggest that VRE isolated from human and chicken are not closely related genetically.
Agar ; Ampicillin ; Animals ; Chickens* ; Diffusion ; DNA ; Drug Resistance, Microbial ; Electrophoresis, Gel, Pulsed-Field ; Enterococcus faecium ; Enterococcus* ; Epidemiology ; Europe ; Hospitals, University ; Humans ; Levofloxacin ; Seoul ; Tetracycline ; Vancomycin

BACKGROUND: The importance of extended-spectrum beta-lactamases (ESBL) produced in gramnegative bacilli is now well recognized, but most clinical laboratories have problems in detecting and interpreting ESBL and implicating the findings in nosocomial infections caused by ESBL producing gram-negative bacilli. The present study aims primarily to evaluate the distributions of these enzymes among Escherichia coli and Klebsiella pneumoniae, the most frequent isolates of Enterobacteriaceae producing ESBL, to differentiate the types of enzymes in theses isolates and finally to relate the clonality of specific types within a part of Daegu city. METHODS: The clinical isolates of 1, 242 E. coli and 859 K. pneumoniae were screened for ESBL production by the disk diffusion method of the National Committee of Clinical Laboratory Standard, and it was confirmed by the double-disk synergy test (DDS). Antimicrobial susceptibility test was performed by the agar dilution method. The presence of -lactamase was tested by polymerase chain reaction (PCR) and plasmid analysis. Isoelectric focusing and nucleotide sequence analysis were performed to evaluate ESBL types. Pulsed-field gel electrophoresis (PFGE) of XbaI-digested DNA fragments was carried out to determine the extend of clonality within the hospital. RESULTS: Of 34 isolates of E. coli and 31 isolates of K. pneumoniae ramdomly selected from those isolates screened for ESBL production were further tested by DDS to confirm its production: 30 (88.2%) E. coli and 29 (93.5%) K. pneumoniae were positive. TEM-52 and SHV-12 were present both in E. coli and K. pneumoniae, but SHV-2a was distributed only in K. pneumoniae. The resistance was transferable in 66.7% of E. coli and 68.9% of K. pneumoniae. Six and 5 PFGE patterns were shown by E. coli and K. pneumoniae, respectively. Among the 5 patterns of K. pneumoniae, type B was dominant, suggesting a clonal outbreak in the hospital. CONCLUSIONS: The ESBL specific enzyme types were TEM-52, SHV-2a and SHV-12. Despite many different PFGE patterns of the ESBL producing isolates, a few outbreak and edemic clones appear to be prevalent in Dongsan Medical Center.
Agar ; Base Sequence ; beta-Lactamases* ; Clone Cells ; Cross Infection ; Daegu ; Diffusion ; DNA ; Electrophoresis, Gel, Pulsed-Field ; Enterobacteriaceae ; Escherichia coli* ; Isoelectric Focusing ; Klebsiella pneumoniae* ; Plasmids ; Pneumonia ; Polymerase Chain Reaction

BACKGROUND: ESBL-producing Klebsiella spp. are increasing worldwide, and infections with ESBL-producing organisms are usually hospital-acquired and common infections with ESBL-producing organisms include urinary tract infections, peritonitis, cholangitis, intraabdominal abscesses and nosocomial pneumonia. We studied the phenotypic and genotypic characteristics of ESBL-producing K. pneumoniae, which were isolated from the patients in St. Vincent Hospital. METHODS: Susceptibility to antibiotic was determined by standard disk diffusion and agar dilution methods for all 22 strains of K. pneumoniae. PCR amplifications were performed with primers specific for TEM. SHY, and CMY-1 genes, and the DNA of the amplified products were sequenced. Total DNA was extracted from the isolates restricted with XbaI, and fingerprinted using pulsed-field gel electrophoresis. Crude preparations of beta-lactamase were obtained by sonications and used for characterization of beta-lactamase by isoelectric focusing. RESULTS: The MIC90 values for ceftazidime and aztreonam were 128 microgram/mL and 64 microgram/mL, respectively. The MIC90 values for cefotaxime and sulperazon were 8 microgram/mL and 4microgram/mL, respectively, and that for cefoxitin was 1.0 microgram/mL, which is much lower than the value for third-generation cephalosporins. The MICs for ciprofloxacin. cefepime, and imipenem were less than 1 microgram/mL for all organisms, which is within the susceptible range. Isoelectric focusing studies demonstrated three beta-lactamases with pls of 5.. (TEM-1), 7.6 (SHV-2a), and 8.2 (SHV-12). The presence of blaSHV and blaTEM genes was confirmed by specific PCRs and DNA sequencing analysis. but blaCMY-1 genes were not found. According to DNA sequencing analysis, 21 K. pneumoniae strains produced SHV-12 ESBL and one strain produced SHV-2a ESBL. CONCLUSIONS: Our results suggest that the resistance of K. pneumoniae from clinical isolates to extended spectrum beta-lactam antibiotics may be due to the production of SHY-type ESBL. This approach in detecting and characterizing ESBL will be valuable for the control of infection and antibiotics use in medical institution.
Abscess ; Agar ; Anti-Bacterial Agents ; Aztreonam ; beta-Lactamases ; Cefotaxime ; Cefoxitin ; Ceftazidime ; Cephalosporins ; Cholangitis ; Ciprofloxacin ; Dermatoglyphics ; Diffusion ; DNA ; Electrophoresis, Gel, Pulsed-Field ; Humans ; Imipenem ; Isoelectric Focusing ; Klebsiella pneumoniae* ; Klebsiella* ; Peritonitis ; Pneumonia ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Sonication ; Urinary Tract Infections

BACKGROUND: Recently, an acquired resistance to vancomycin in enterococci has become a serious clinical problem. For the prevention of further propagation of vancomycin-resistant enterococci (VRE), epidemiological study of the infection is essential, but studies on the VRE infection are rare in Korea. We conducted an analysis of the epidemiology of a VRE outbreak in a neonatal intensive care unit (NICU) to clear up the route of propagation of the VRE. METHODS: Vancomycin-resistant Enterococcus faecium (VREFM) strains were isolated from urine specimens of seven patients, rectal swabs from seven patients, and three skin swabs from two patients in the Kosin Medical Center neonatal intensive care unit, Pusan, Korea. Antimicrobial susceptibilities were tested by a disk diffusion method and agar dilution method. Genotypes of vancomycin-resistance were determined by PCR and SmaI-digested genomic enterococcal DNAs were analyzed by pulsed-field gel electrophoresis. RESULTS: All of the 17 strains of VREFM were resistant to ampicillin, vancomycin, and teicoplanin and they showed the same genotype (vanA). SmaI-digested genomic DNAs of seven strains isolated from urine were typed as I (1), II (1), IIIb (4), and IV (1). Three strains from skin swabs were I (2) and II (1). Six strains from rectal swabs were I (2), II (1), and IIIa (3). Genomic DNA typing of one isolate from a rectal swab failed. Each genomic DNA type of VREFM strains isolated from skin swabs of two patients were the same with those from urine specimens as I and II, respectively. CONCLUSIONS: This study suggests that VRE strains colonized in the intestines can cause infections after skin colonizing and can be transmitted/propagated to other people through skin contact. In conclusion, it is important for the prevention of the dissemination of VRE that controls for patients' skin hygiene, as well as hand washing by medical persons, be put in place.
Agar ; Ampicillin ; Busan ; Colon ; Diffusion ; DNA ; DNA Fingerprinting ; Electrophoresis, Gel, Pulsed-Field ; Enterococcus faecium ; Epidemiologic Studies ; Epidemiology ; Genotype ; Hand Disinfection ; Humans ; Hygiene ; Infant, Newborn ; Intensive Care, Neonatal* ; Intestines ; Korea ; Molecular Epidemiology* ; Polymerase Chain Reaction ; Skin ; Teicoplanin ; Vancomycin

BACKGROUND: Vancomycin-resistant enterococci (VRE) with vanA gene have been reported as a significant nosocomial pathogen. The vanA gene cluster (Tn1546) located on mobile DNA elements is known to be transferable from VRE to other enterococci. The purpose of this study was to investigate the genetic relationship between the vanA VRE strains isolated from hospitalizd patients and poultry. METHODS: Total 145 isolates, including 58 E. faecium, 12 E. faecalis, 3 E. casseliflavus, and 4 E. gallinarum from humans and 68 E. faecium from poultry, were studied. Antimicrobial susceptibility tests were done by disk diffusion or agar dilution methods and molecular epidemiological analysis was performed by pulsed-field gel electrophoresis (PFGE). The internal and structural regions of vanA gene cluster were analyzed by PCR fragment length polymorphism, restriction enzyme, and sequencing of Orf2D region and vanXY intergenic region. The point mutation at Tn1546 nucleotide position 8234 (G->T) within the vanX gene was screened with DdeI restriction enzyme. RESULTS: The antibiotic resistance patterns of human isolates were different from those of poultry. PFGE patterns revealed high heterogeneity. Three PCR fragment length patterns in the vanA gene cluster were found : (I) PCR amplicon of the same size as prototype (E. faecium BM4147) in 17% of human isolates and 100% of poultry ones; (II) PCR amplicon for vanXY intergenic region due to an insertion between vanX and vanY genes in 2.5% of human isolates; (III) the insertions in vanX-vanY intergenic and Orf2 regions in 81% of human isolates. The T type in vanX gene of human and poultry isolates was not found. CONCLUSION: Despite the diverse PFGE patterns, 81% of human and all of poultry isolates belonged to vanA gene cluster type III and I, respectively. These results indicate that the horizontal spread of vanA gene is occurring among genetically diverse strains of VRE in Korea.
Agar ; Diffusion ; DNA ; DNA, Intergenic ; Drug Resistance, Microbial ; Electrophoresis, Gel, Pulsed-Field ; Humans ; Korea* ; Multigene Family ; Point Mutation ; Polymerase Chain Reaction ; Population Characteristics ; Poultry*

BACKGROUND: Pseudomonas aeruginosa is a major cause of nosocomial infections. During recent three years, the isolation rate was high in Wonkwang University Hospital, especially in intensive care unit(ICU) and wards of neurosurgery. We performed this study to investigate the isolation rate and mode of transmission of P. aeruginosa, and usefulness of antibiotic resistance, colonial characteristics and PFGE patterns in the epidemiologic survey. METHOD: From Aug. 1996 to Oct. 1998, 1,682 strains of P. aeruginosa were isolated. For the isolation of P. aeruginosa, environment and 18 nurses's hands were cultured. Antibiotic resistance were tested by NCCLS disk diffusion method for 298 selected strains. Among them 98 strains were evaluated for colonial characteristics(color and margin) on the blood agar and PFGE patterns restricted by Spe I, were evaluated. RESULTS: Overall isolation rate was 12% and was high in medical ICU (23%) and neurosurgical ICU(12%). The majority of specimens where P. aeruginosa was isolated were sputum 47%, urine 24% and wound 21% in decreasing order. The wards where isolation rate was high, had more resistant strains. Ninty eight strains could be classified into 39 different groups by PFGE patterns. But 29 strains belonged to five major patterns (P1-5) and were suspected as epidemic or cross-infected strains. Majority of these strains revealed resistant to two or more antibiotics and colonial phenotype of G2R, GIR(P1), G1I, G3I(P2), Wm(P3), G3I, G3R(P4) and G3I(P5) types. Forty four strains isolated from specimens(sputum, urine, wound, and stool) of 7 patients during hospitalization, revealed single or two PFGE patterns per patient. Conclusion: Transmission mode of P. aeruginosa was suspected to be patient's own GI-tract and cross contamination, especially via health care persons. Combined phenotypes of antibiotic resistance and colonial characteristics correlated well with PFGE patterns. So, in the early period of outbreak of P. aeruginosa, careful examination of colonial characteristics and antibiotic resistance patterns gave meaningful information.
Agar ; Anti-Bacterial Agents ; Cross Infection ; Delivery of Health Care ; Diffusion ; Drug Resistance, Microbial* ; Electrophoresis, Gel, Pulsed-Field ; Epidemiologic Studies* ; Hand ; Hospitalization ; Humans ; Critical Care ; Neurosurgery ; Phenotype ; Pseudomonas aeruginosa* ; Pseudomonas* ; Sputum ; Wounds and Injuries ; Surveys and Questionnaires