Jing-Jou Yan

Change of Major Genotype of Enterovirus 71 in Outbreaks of Hand-Foot-and-Mouth Disease in Taiwan between 1998 and 2000

ABSTRACT Two outbreaks of hand-foot-and-mouth disease (HFMD) occurred in Taiwan between 1998 and 2000. Enteroviruses were isolated from a total of 1,892 patients in this laboratory during this period. Of the virus isolates, enterovirus 71 (EV71) was diagnosed in 44.4% of the patients (132 of 297) in 1998, 2% (13 of 646) in 1999, and 20.5% (195 of 949) in 2000. Genetic analyses of the 5′-untranslated and VP1 regions of EV71 isolates by reverse transcription-PCR and sequencing were performed to understand the diversity of EV71 in these outbreaks of HFMD. Most EV71 isolates from the 1998 epidemic belonged to genotype C, while only one-tenth of the isolates were genotype B. Interestingly, all EV71 isolates tested from 1999 to 2000 belonged to genotype B. This study indicated that two genogroups of EV71 capable of inducing severe clinical illness have been circulating in Taiwan. Furthermore, the predominant EV71 genotypes responsible for each of the two major HFMD outbreaks within the 3-year period in Taiwan were different.

Prevalence of Plasmid-Mediated Quinolone Resistance Determinants QnrA, QnrB, and QnrS among Clinical Isolates of Enterobacter cloacae in a Taiwanese Hospital

ABSTRACT The prevalence of three plasmid-mediated quinolone resistance determinants, QnrA, QnrB, and QnrS, among 526 nonreplicate clinical isolates of Enterobacter cloacae collected at a Taiwanese university hospital in 2004 was determined by PCR and colony hybridization, and the association of Qnr with the IMP-8 metallo-β-lactamase was investigated. Eighty-six (16.3%) of all isolates were qnr positive, and the qnrA1-like, qnrB2-like, and qnrS1-like genes were detected alone or in combination in 3 (0.6%), 53 (10.1%), and 34 (6.5%) isolates, respectively. Among 149 putative extended-spectrum-β-lactamase-producing isolates, 59 (39.6%) isolates, all of which were SHV-12 producers, harbored qnrA (0.7%; 1 isolate), qnrB (28.9%; 43 isolates), or qnrS (12.1%; 18 isolates). Forty-four (78.6%) of 56 IMP-8 producers carried qnrB (58.9%; 33 isolates), qnrS (25.0%; 14 isolates), or both. PCR and sequence analysis revealed that qnrA1 was located in a complex sul1-type integron that contains dhr15, aadA2, qacEΔ1, sul1, orf513, qnrA1, ampR, and qacEΔ1. Conjugation experiments revealed the coexistence of qnrB and blaIMP-8 on the transferred plasmids and the absence of β-lactamase content on the transferred qnrS-positive plasmids. The transferred blaIMP-8-positive plasmids with and without qnrB had very similar restriction patterns, suggesting the horizontal mobility of qnrB. Pulsed-field gel electrophoresis showed six major patterns among the 44 qnr-positive IMP-8-producing isolates. Thus, the extremely high prevalence of qnr among the metallo-β-lactamase-producing E. cloacae isolates in the hospital may be due mainly to the intrahospital spread of a few clones and the dissemination of plasmids containing both qnrB and blaIMP-8.

Prevalence of SHV-12 among Clinical Isolates of Klebsiella pneumoniae Producing Extended-Spectrum β-Lactamases and Identification of a Novel AmpC Enzyme (CMY-8) in Southern Taiwan

ABSTRACT Twenty (8.5%) of 234 nonrepetitive clinical isolates ofKlebsiella pneumoniae from southern Taiwan were found to produce extended-spectrum β-lactamases (ESBLs): 10 strains produced SHV-12, 4 produced SHV-5, 2 produced a non-TEM non-SHV ESBL with a pI of 8.3, 3 produced a novel AmpC β-lactamase designated CMY-8 with a pI of 8.25, and 1 produced SHV-12 and an unidentified AmpC enzyme with a pI of 8.2. The CMY-8 enzyme confers a resistance phenotype similar to CMY-1 and MOX-1, and sequence comparisons showed high homologies (>95%) of nucleotide and amino acid sequences among these three enzymes. Plasmid and pulse-field gel electrophoresis analyses revealed that all isolates harboring an SHV-derived ESBL were genetically unrelated, indicating that dissemination of resistance plasmids is responsible for the spread of SHV ESBLs among K. pneumoniaein this area. All three isolates carrying CMY-8 had identical genotypic patterns, suggesting the presence of an epidemic strain.

Outbreak of Infection with Multidrug-Resistant Klebsiella pneumoniae Carrying bla IMP-8 in a University Medical Center in Taiwan

ABSTRACT Klebsiella pneumoniae strains with the transferable carbapenem-hydrolyzing metallo-β-lactamases, which include IMP- and VIM-type enzymes, remain extremely rare. To investigate whether IMP- or VIM-producing K. pneumoniaeisolates had spread at a university medical center in Taiwan, a total of 3,458 clinical isolates of K. pneumoniaeconsecutively collected in 1999 and 2000 were tested by the agar diffusion method, colony hybridization, PCR, and nucleotide sequencing. A total of 40 isolates (1.2%), or 17 nonrepetitive isolates, from 16 patients were found to carry blaIMP-8, a metallo-β-lactamase gene recently identified from a K. pneumoniae strain in Taiwan. Carriage ofblaVIM or otherblaIMP genes was detected in none of the remaining isolates. Of the 17 nonrepetitiveblaIMP-8-positive isolates, 15 isolates (88.2%) appeared susceptible to imipenem (MICs, ≤4 μg/ml) and meropenem (MICs, ≤1 μg/ml), indicating the difficulty in detecting blaIMP-8 in K. pneumoniae by routine susceptibility tests; 14 isolates (82.4%) produced SHV-12 as well; and 14 isolates (82.4%) were also resistant to fluoroquinolones. The organisms caused wound infections in eight patients and bloodstream infections in three patients. They were not directly associated with the death of nine patients. Before the recovery of the blaIMP-8-positive isolates, all 16 patients had undergone various surgical procedures, and 15 patients had been admitted to the surgical intensive care unit, suggesting a nosocomial outbreak. Two major patterns were observed by pulsed-field gel electrophoresis for 14 of the 17 nonrepetitive isolates, indicating that the clonal spread was mainly responsible for the outbreak.

Complete Nucleotide Sequence of pK245, a 98-Kilobase Plasmid Conferring Quinolone Resistance and Extended-Spectrum-beta-Lactamase Activity in a Clinical Klebsiella pneumoniae Isolate

ABSTRACT A plasmid containing the qnrS quinolone resistance determinant and the gene encoding the SHV-2 β-lactamase has been discovered from a clinical Klebsiella pneumoniae strain isolated in Taiwan. The complete 98-kb sequence of this plasmid, designated pK245, was determined by using a whole-genome shotgun approach. Transfer of pK245 conferred low-level resistance to fluoroquinolones in electroporant Escherichia coli epi300. The sequence of the immediate region surrounding qnrS in pK245 is nearly identical (>99% identity) to those of pAH0376 from Shigella flexneri and pINF5 from Salmonella enterica serovar Infantis, the two other qnrS-carrying plasmids reported to date, indicating a potential common origin. Other genes conferring resistance to aminoglycosides (aacC2, strA, and strB), chloramphenicol (catA2), sulfonamides (sul2), tetracycline (tetD), and trimethoprim (dfrA14) were also detected in pK245. The dfrA14 gene is carried on a class I integron. Several features of this plasmid, including three separate regions containing putative replicons, a partitioning-control system, and a type II restriction modification system, suggest that it may be able to replicate and adapt in a variety of hosts. Although no critical conjugative genes were detected, multiple insertion sequence elements were found scattered throughout pK245, and these may facilitate the dissemination of the antimicrobial resistance determinants. We conclude that pK245 is a chimera which acquired its multiple antimicrobial resistance determinants horizontally from different sources. The identification of pK245 plasmid expands the repertoire of the coexistence of quinolone and extended-spectrum-β-lactam resistance determinants in plasmids carried by various species of the family Enterobacteriaceae in different countries.

Identification of a Plasmid Encoding SHV-12, TEM-1, and a Variant of IMP-2 Metallo-β-Lactamase, IMP-8, from a Clinical Isolate of Klebsiella pneumoniae

ABSTRACT A multidrug-resistant plasmid encoding TEM-1, SHV-12, and a variant of IMP-2 metallo-β-lactamase, designated IMP-8, was identified from a clinical isolate of Klebsiella pneumoniae. There are four nucleotide differences between blaIMP-2 andblaIMP-8, resulting in two amino acid differences. blaIMP-8 was also found to be carried by an integron-borne gene cassette similar to theblaIMP-2 cassette.

Multicenter surveillance of antimicrobial resistance of major bacterial pathogens in intensive care units in 2000 in Taiwan

A susceptibility surveillance study of 1,274 bacterial isolates recovered from various clinical specimens from patients in intensive care units (ICUs) of five major teaching hospitals was carried out from March, 2000, to June, 2000, in Taiwan. This study demonstrated a high rate (66%) of oxacillin resistance in Staphylococcus aureus (ORSA), a high rate of nonsusceptibility to penicillin (intermediate, 50% and highly resistant, 8%), and high rates of cefotaxime nonsusceptibility for S. pneumoniae (intermediate, 29% and resistant, 4%), Enterobacter cloacae (57%), Serratia marcescens (34%), and Citrobacter freundii (60%). High rate of ceftazidime nonsusceptibility for Pseudomonas aeruginosa (22%), and high rates of imipenem nonsusceptibility for P. aeruginosa (15%) and Acinetobacter baumannii (22%) were also found. The percentage (11.9%) of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli was greater than that (11.3%) for Klebsiella pneumoniae. Rates of quinupristin-dalfopristin nonsusceptibility for S. pneumoniae (42%), Enterococcus faecium (71%), and ORSA (39%) were high, but no vancomycin-resistant enterococci were found in this study. The resistance rates of some pathogen varied by institution or type of ICUs. Surveillance for antimicrobial resistance among bacterial pathogens in hospitals, particularly in ICU settings with a preexisting higher resistance burden, is mandatory in establishing and/or modifying guidelines for empirical treatment of severe infections in ICU patients caused by these antimicrobial-resistant pathogens.

Emergence of Klebsiella pneumoniae Isolates Producing Inducible DHA-1 β-Lactamase in a University Hospital in Taiwan

ABSTRACT Ten nonrepetitive clinical isolates of Klebsiella pneumoniae exhibiting an unusual inducible β-lactam resistance phenotype were identified between January 1999 and September 2001 in a university hospital in Taiwan. In the presence of 2 μg of clavulanic acid, the isolates showed a one to four twofold concentration increase in the MICs of ceftazidime, cefotaxime, and aztreonam but remained susceptible to cefepime (MICs, ≤0.5 μg/ml) and imipenem (MICs, ≤0.5 μg/ml). PCR, sequence analysis, and isoelectric focusing revealed production by these isolates of TEM-1, SHV-11, and DHA-1, a plasmid-encoded inducible AmpC β-lactamase originally found in a Salmonella enterica serovar Enteritidis strain. Transfer of the resistance by conjugation experiments was not successful, but Southern hybridization showed that blaDHA-1 was located on 70-kb plasmids, suggesting that the blaDHA-1-containing plasmids in the K. pneumoniae isolates were non-self-transmissible. Five isolates were recovered from patients in two surgery wards and two intensive care units. Acquisition of the DHA-1 producers could be traced back to previous hospitalizations 1 to 5 months earlier for the other five patients. Six and seven patterns among the isolates were demonstrated by plasmid analysis and ribotyping, respectively, indicating that the spread of the DHA-1 producers was due to both horizontal transfer of blaDHA-1 and dissemination of endemic clones.

Dissemination of blaCMY-2 among Escherichia coli Isolates from Food Animals, Retail Ground Meats, and Humans in Southern Taiwan

ABSTRACT Twenty-six Escherichia coli isolates recovered from food animal feces and retail ground meats and 14 urinary E. coli isolates from outpatients were shown to carry blaCMY-2. Similar CMY-2-encoding plasmids were found among seven human and three ground-pork isolates. These data indicate the community spread of blaCMY-2 in southern Taiwan.

Complexity of Klebsiella pneumoniae Isolates Resistant to Both Cephamycins and Extended-Spectrum Cephalosporins at a Teaching Hospital in Taiwan

ABSTRACT Among 99 clinical Klebsiella pneumoniae isolates resistant to cefoxitin and extended-spectrum cephalosporins, coexistence of AmpC (DHA-1, CMY-2, or CMY-8) and extended-spectrum β-lactamases (CTX-M and/or SHV) was detected in a total of 35. The remainder produced AmpC (n = 42), extended-spectrum β-lactamases (n = 9), metallo-β-lactamases (n = 2), or none of these enzymes (n = 11). Phenotypic characteristics of these isolates were demonstrated.