Influence of ethanol concentration in the phagocytic function of neutrophils against Klebsiella pneumoniae isolates in an experimental model.

BACKGROUND/PURPOSE: Although the prevalence of pneumonia or other extrapulmonary infections is higher in people with alcoholism or acute alcohol intoxication, the possible relationship of acute alcohol intoxication to phagocytic function has not been investigated. Our aim was to determine whether acute alcohol intoxication suppresses phagocytic function in human neutrophils. METHODS: Twenty healthy individuals were enrolled for isolating neutrophils to evaluate the neutrophil phagocytic function at different alcohol concentrations. Klebsiella pneumoniae was isolated from clinical specimens of liver abscesses. The rate of K. pneumonia phagocytosis (K2 and non-K1/K2 isolates) by neutrophils was determined using flow cytometry and compared among the nine groups with different alcohol concentrations. RESULTS: The rate of phagocytic uptake decreased significantly with increasing alcohol concentration in both the K2 and non-K1/K2 K. pneumonia groups (r = -0.866, p = 0.03 vs. r = -0.975, p < 0.001). Moreover, the percentage of K. pneumoniae ingested by neutrophils decreased with age. CONCLUSION: The ability of neutrophils to phagocytose virulent K2 K. pneumoniae was suppressed by ethanol at high concentrations. This finding may account for the higher prevalence of pneumonia or other extrapulmonary infection in people with acute alcohol intoxication.

Quantification and comparison of virulence and characteristics of different variants of carbapenemase-producing Klebsiella pneumoniae clinical isolates from Taiwan and the United States.

BACKGROUND/PURPOSE: The emergence of Klebsiella pneumoniae carbapenemase (KPC)-producing strains is a challenge for clinicians. The characteristics and virulence of variants of KPC-producing K. pneumoniae isolates were evaluated. METHODS: Five clinical isolates-three KPC subtypes from Taiwan (KPC2-TW, KPC3-TW, and KPC17-TW) and two clinical strains from the United States (US; KPC2-US, KPC3-US)-were included. Virulent traits and capsular serotypes were analyzed by Polymerase Chain Reaction (PCR). Serum killing, neutrophil phagocytosis, and mice lethargy studies were performed to evaluate virulence. RESULTS: Multilocus sequence typing (MLST) demonstrated that KPC2-TW and KPC17-TW belonged to sequence type (ST)11, and KPC2-US, KPC3-US, and KPC3-TW to ST258. KPC3-TW expressed capsular serotype K1, whereas the others were non-K1/K2/K5 isolates. MLST analysis indicated that ST11 strains were serum resistant, whereas ST258 isolates were serum sensitive. ST11 isolates exhibited significantly higher 15-minute phagocytic rates than ST258 isolates (70.28 ± 16.68% vs. 34.88 ± 10.52%, p < 0.001). The capsular serotype K1 strain was more resistant to neutrophil phagocytosis than non-K1/K2/K5 isolates (27.1 ± 10.23% vs. 54.46 ± 20.94%, p = 0.050). All KPC-producing strain variants from Taiwan and the US demonstrated less virulence in a mouse lethality study, where the LD50 ranged from approximately 10(6) colony-forming units (CFU) to >10(7) CFU. Immunological responses were not significantly correlated with KPC subtype; however, responses were associated with MLST and capsular serotype. CONCLUSION: Production of KPC itself was not associated with increased virulence despite different variants of KPC. The ST11 KPC-producing strain was resistant to serum killing, whereas capsular ss K1 was associated with resistance to neutrophil phagocytosis.

Regulator of the mucoid phenotype A gene increases the virulent ability of extended-spectrum beta-lactamase-producing serotype non-K1/K2 Klebsiella pneumonia.

BACKGROUND: To determine whether the presence of a capsule regulator gene [i.e., regulator of mucoid phenotype A (rmpA) gene] contributes to virulence on extended-spectrum ß-lactamase-producing Klebsiella pneumoniae (ESBL-KP) with serotype non-K1/K2 strains. METHODS: Twenty-eight ESBL-KP and non-ESBL-KP isolates were collected from the Tri-Service General Hospital (Taipei, Taiwan). The impact of the virulent rmpA gene in different capsular polysaccharide serotypes on ESBL-KP and non-ESBL-KP isolates was studied by a neutrophil phagocytosis reaction, a serum bactericidal assay, and an animal survival model. RESULTS: Resistance to broad spectrum antibiotics was more prevalent in ESBL-KP strains than in non-ESBL-KP strains (p < 0.01). The ESBL-KP strains had different molecular patterns from non-ESBL-KP strains, based on pulsed-field gel electrophoresis. The frequency of serum-resistant isolates was the highest among ESBL-KP strains with rmpA (i.e., rmpA(+)) [71.4% (5/7)] than among of non-ESBL-KP rmpA(+) strains [42.8% (6/14)], ESBL-KP strains without rmpA (rmpA(-)) [33.3% (7/21)], and non-ESBL-KP rmpA(-) strains [14.2% (2/14)]. The most significant increase in neutrophil resistance occurred in the ESBL-KP rmpA(+) strains in comparison to the non-ESBL-KP rmpA(+), ESBL-KP rmpA(-), and non-ESBL-KP rmpA(-) strains (p < 0.01). The results of the animal survival model were compatible with the neutrophil phagocytosis reaction and serum bactericidal assay. CONCLUSION: We conclude that the pathogenic potential is greater in rmpA(+) ESBL-KP strains than in rmpA(-) ESBL-KP and non-ESBL-KP strains.

Clinical features and risk factors of mortality for bacteremia due to community-onset healthcare-associated methicillin-resistant S. aureus.

Studies comparing adult community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) and community-onset healthcare-associated MRSA (COHCA MRSA bacteremia have not been available. From 1 January 2010 through 30 October 2010, a prospective observational program was conducted among all patients aged >16 years with positive Staphylococcus aureus blood cultures within 48 h after their arrival at the emergency department of our hospital. Clinical course of infection and infection foci of bacteremia were evaluated. Resistance to oxacillin was confirmed with the presence of mecA gene examined by polymerase chain reaction. Presence of TSST-1, PVL gene, SCCmec elements (I-V), mecA gene, and multilocus sequence typing were identified through methods described elsewhere. Univariate and multivariate analysis revealed that chronic renal failure was significantly more common in COHCA-MRSA than in CA-MRSA. In addition, APACHE III score was significantly higher in COHCA-MRSA than in CA-MRSA. Both the 7-day and 30-day mortality rates in COHCA-MRSA, 14.6% (7/48) and 29.2% (14/48), respectively, were higher than those in CA-MRSA without a significant difference. SCCmec II was more common in COHCA-MRSA, but SCCmecVT was more common in CA-MRSA. The majority of MRSA isolates belonged to ST59, ST239, and ST5. ST59 was significantly more common in CA-MRSA, while ST239 was nearly equally common in both CA-MRSA and COHCA-MRSA. SCCmec III and II isolates were the first and second most resistant to the antibiotics commonly used for S. aureus, whereas SCCmecVT isolates were the most susceptible to these antibiotics. We conclude that, although both CA-MRSA and COHCA-MRSA bacteremia had community onset, these 2 MRSA infections were different in underlying diseases, risk of mortality, SCCmec types, sequence types, and antimicrobial susceptibility. It is more appropriate to understand the MRSA pathogen and clinical features based on etiology and ST types than based on the location of disease onset. CA-MRSA and HCA-MRSA should be differentiated also based on etiology and ST types, in addition to location of acquisition.

Induction of alloimmune tolerance in heart transplantation through gene silencing of TLR adaptors.

Toll-like receptors (TLRs) activate biochemical pathways that evoke activation of innate immunity, which leads to dendritic cell (DC) maturation and initiation of adaptive immune responses that provoke allograft rejection. We aimed to prolong allograft survival by selectively inhibiting expression of the common adaptors of TLR signaling, namely MyD88 and TRIF, using siRNA. In vitro we demonstrated that blocking expression of MyD88 and TRIF led to reduced DC maturation. In vivo treatment of recipients with MyD88 and TRIF siRNA significantly prolonged allograft survival in the BALB/c > C57BL6 cardiac transplant model. Moreover, the combination of MyD88 and TRIF siRNA along with a low dose of rapamycin further extended the allograft survival (88.8 ± 7.1 days). Tissue histopathology demonstrated an overall reduction in lymphocyte interstitium infiltration, vascular obstruction and hemorrhage in mice treated with MyD88 and TRIF siRNA vector plus rapamycin. Furthermore, treatment was associated with an increase in the numbers of CD4(+) CD25(+) FoxP3(+) regulatory T cells and Th2 deviation. To our knowledge, this study is the first demonstration of prolonging the survival of allogeneic heart grafts through gene silencing of TLR signaling adaptors, highlighting the therapeutic potential of siRNA in clinical transplantation.

Increasing opsonizing and killing effect of serum from patients with recurrent K1 Klebsiella pneumoniae liver abscess.

BACKGROUND: Klebsiella pneumoniae liver abscess (KLA) is an emerging infectious disease caused by the virulent K pneumoniae strains of capsular serotype K1 and commonly associated with diabetes mellitus. Recurrent KLA is rarely reported and the mechanism of recurrence is uncertain. In this study we evaluated both phagocytosis by neutrophils and serum killing ability of serum from recurrent K1 KLA patients compared to normal healthy subjects (NHS). METHODS: This prospective study included six cases of recurrent K1 KLA consisting of three male and three female patients with a mean age of 67.2 years (range, 56-88 years). The different serotypes of K pneumoniae were reacted with serum from patients with recurrent KLA and NHS. Subsequent phagocytosis by neutrophils was determined using flow cytometry and serum killing assays were performed. RESULTS: The most common underlying disease in patients with recurrent KLA was diabetes mellitus, occurring in about 83.3% (5/6) of patients. The antibiogram of the strains associated with recurrent KLA remained uniquely resistant to ampicillin. The average percentage derived from the serum killing assays showed serotype K1 and K2 resistance to serum from NHS (1281% and 621%, respectively); however, serum susceptibly was observed in the serum of patients with recurrent K1 KLA (0.3% and 1.1%, respectively). A significant increase in neutrophil phagocytosis of serotype K1 was observed following opsonisation with serum from patients with recurrent KLA compared with serum from NHS (p = 0.008). No significant difference in the phagocytic rate of non-K1/K2 or K2 serotypes was observed between NHS and patients with recurrent KLA (p = 0.76 and p = 0.132, respectively). CONCLUSION: These preliminary results showed possible immunologic protection in patients with recurrent KLA due to increasing opsonization and serum killing.

Eradication of multidrug-resistant Acinetobacter baumannii from the respiratory tract with inhaled colistin methanesulfonate: a matched case-control study.

Repeated isolation of multidrug-resistant Acinetobacter baumannii (MDRAB) from respiratory secretions poses a great challenge for infection control. We conducted a retrospective case-control study to evaluate the efficacy and adverse effect of inhaled colistin methanesulfonate (CMS) in the eradication of MDRAB from the respiratory tract. Patients who were admitted to Taipei Veterans General Hospital between February 2009 and June 2010, had at least two sets of monomicrobial culture of MDRAB from respiratory secretions, and remained in hospital for at least 14 days after the first isolation of MDRAB (index day) were included. Patients who received intravenous CMS were excluded. Patients who received CMS inhalation for ≥ 3 days were selected as cases whereas the controls were matched for age and Acute Physiology and Chronic Health Evaluation II score. Thirty-nine cases and controls were identified. The duration of CMS inhalation was 10.9 ± 3.6 days. The use of inhaled CMS was the only independent factor associated with the eradication of MDRAB within 14 days after the index day (OR 266.33; 95% CI 11.26-6302.18, p <0.001), and shortened the duration of MDRAB recovery from the respiratory tract by 13.3 ± 1.45 days. The adverse effects were similar for both groups. The increase of colistin minimal inhibitory concentrations in the last isolate compared with the index isolate from the same patient did not differ between the two groups. In conclusion, our study demonstrated that inhaled CMS enhanced the eradication of MDRAB from the respiratory tract without significant clinical adverse effect or impact on colistin resistance.

Antimicrobial resistance of Moraxella catarrhalis isolates in Taiwan.

BACKGROUND/PURPOSE: The prevalence of ampicillin-resistant Moraxella catarrhalis has been higher in Taiwan than in other countries, with reports of 97.7% in the 1990s. The aims of this study were to assess resistance trends for M. catarrhalis, which causes respiratory tract infections, against several classes of oral antibiotics and to compare the minimum inhibitory concentration (MIC) of antimicrobial agents against M. catarrhalis isolates between 1993-1994 and 2001-2004. METHODS: Clinical isolates of M. catarrhalis (n = 314) were collected from 11 large medical centers in Taiwan between 2001 and 2004. ß-Lactamase production tests were performed. The MICs for 13 different oral antibiotics were calculated using the agar dilution method. Pulsed-field gel electrophoresis (PFGE) was performed for 18 randomly selected high-level ampicillin-resistant (BRO-1 ß-lactamase-positive, MIC ≥ 32 µg/mL) isolates to investigate their genetic relatedness. RESULTS: The overall rate of ß-lactamase-producing isolates was 97.8% (307/314). All isolates were susceptible to amoxicillin + clavulanate, chloramphenicol, cefixime, ciprofloxacin, erythromycin, levofloxacin, moxifloxacin, and roxithromycin. The rate of resistance to cefaclor and cefuroxime was 8.3% and 1.3%, respectively, while no resistance was found in 1993-1994. Resistance to trimethoprim-sulfamethoxazole (SXT) and tetracycline was 18.5% and 19.8%, respectively. Comparison of 1993-1994 and 2001-2004 isolates revealed that the zone diameter for amoxicillin + clavulanate disks decreased from 43 mm in 1993-1994 to 32 mm in 2001-2004 (p < 0.001). However, MIC(50) (0.25 µg/mL in both 1993-1994 and 2001-2004) and MIC(90) (0.5 µg/mL in both 1993-1994 and 2001-2004) for amoxicillin + clavulanate did not differ between the study periods. The PFGE typing results demonstrate that at least two closely related BRO-1 clones are spreading in Taiwan. CONCLUSION: The rates of resistance to cefaclor, cefuroxime, tetracycline and SXT are now increasing in Taiwan. Molecular typing showed that at least two closely related BRO-1 clones are circulating. Although amoxicillin + clavulanate remains the antimicrobial therapy of choice for M. catarrhalis infections, continued surveillance of antimicrobial susceptibility and application of control measures against further transmission are required to inhibit the emergence of the resistant strains.

Targeted gene silencing of TLR4 using liposomal nanoparticles for preventing liver ischemia reperfusion injury.

RNAi-based therapy is a promising strategy for the prevention of ischemia-reperfusion injury (IRI). However, systemic administration of small interfering RNA (siRNA) may cause globally nonspecific targeting of all tissues, which impedes clinical use. Here we report a hepatocyte-specific delivery system for the treatment of liver IRI, using galactose-conjugated liposome nanoparticles (Gal-LipoNP). Heptocyte-specific targeting was validated by selective in vivo delivery as observed by increased Gal-LipoNP accumulation and gene silencing in the liver. Gal-LipoNP TLR4 siRNA treatment resulted in a significant decrease of serum alanine transferase (ALT) and aspartate transaminase (AST) in a hepatic IRI model. Histopathology displayed an overall reduction of the injury area in the Gal-LipoNP TLR4 siRNA treated mice. Additionally, neutrophil accumulation and lipid peroxidase-mediated tissue injury, detected by MPO, MDA and ROS respectively, were attenuated after Gal-LipoNP TLR4 siRNA treatment. Moreover, therapeutic effects of Gal-LipoNP TLR4 siRNA were associated with suppression of the inflammatory cytokines IL-1 and TNF-α. Taken together, this study is the first demonstration of liver IRI treatment using liver-specific siRNA delivery.

Different culture medium formulations induce variant protein expression patterns of outer membrane porins in Klebsiella pneumoniae.

Outer membrane porin (OMP) expression has been shown to play an important role in antimicrobial resistance. In this study, we observed that OmpK35 of Klebsiella pneumoniae had varied expression profiles in different nutrient broths. the potential factors that could influence protein expression were assessed. K. pneumoniae (ATCC 13883) was cultured into two commercial available nutrient broths and also into solutions of the individual ingredients. To ensure that OmpK35 was detected, an OmpK35 deficient mutant was generated as control. When OmpK protein expression profiles were analyzed by SDS-PAGE, OmpK35 exhibited two different isoforms. Expression of an additional isoform-like OmpK35 protein was identified in one of the broths. No OmpK35 isoforms were observed when the individual ingredients of beef extract, casein or gelatin were used as culture medium. OmpK35 isoform expression could be repressed by adding more beef extract. In summary, OmpK can exhibit varied protein expression profiles when growing in different nutrient broths. The isoform-like protein expression of OmpK35 may lead to confusion in OmpK protein analysis.

Clinical and microbiological characteristics of community-acquired thoracic empyema or complicated parapneumonic effusion caused by Klebsiella pneumoniae in Taiwan.

Klebsiella pneumoniae is the major cause of community-acquired pyogenic infections in Taiwan and is becoming an increasing problem in acute thoracic empyema. This study evaluated the clinical and microbiological characteristics of community-acquired thoracic empyema or complicated parapneumonic effusion caused by K. pneumoniae in Taiwanese adults treated during the period 2001-2008 at a tertiary medical center. All clinical isolates were examined for capsular serotypes K1/K2, and pulsed-field gel electrophoresis (PFGE) was performed on strains of the same serotype. K. pneumoniae was the most frequent cause of community-acquired thoracic empyema or complicated parapneumonic effusion. It was associated with high mortality (32.4%) and was an independent risk factor for fatal outcome. Diabetes mellitus, liver cirrhosis, and bronchogenic carcinoma were independent risk factors for K. pneumoniae infection. Serotypes K1 (9/37, 24.3%) and K2 (13/37, 35.1%) were the prevalent strains but did not predispose patients to poor outcome compared with other non-K1/K2 serotypes. There was no major cluster of isolates found among serotype K1/K2 strains. In summary, physicians should be aware of the risk factors for thoracic empyema or complicated parapneumonic effusion caused by K. pneumoniae and the associated high mortality, and monitor these patients more closely.

Molecular characterization of beta-lactamase genes and their genetic structures in Acinetobacter genospecies 3 isolates in Taiwan.

The genetic structure of beta-lactamases in Acinetobacter genospecies 3 (AG3) isolates in Taiwan was studied to analyze their high rates of resistance to beta-lactams, including carbapenems (57.9%). bla(IMP-1) and bla(IMP-8) were located in a class 1 integron. bla(OXA-58) was bracketed by ISAba3. A novel TnpF-like integrase gene was identified upstream of bla(VEB-3). Adjacent to the 5' sequence of the bla(ADC) gene, folE was identified. Four new Acinetobacter-derived cephalosporinase (ADC) enzymes were found, which clustered phylogenetically with published AG3 ADC proteins.

Contribution of outer membrane protein K36 to antimicrobial resistance and virulence in Klebsiella pneumoniae.

OBJECTIVES: Loss of outer membrane protein (Omp) is commonly encountered in multidrug-resistant Klebsiella pneumoniae. However, little is known about the association between Omp loss and virulence. In the present study, this association was investigated in K. pneumoniae. METHODS: An OmpK36-deficient mutant (DeltaOmpK36) was derived from a virulent clinical isolate by targeted gene insertion. Antimicrobial susceptibility was tested by microbroth dilution and disc diffusion. Virulence was assessed by serum resistance, phagocytosis, clearance of viable bacteria in the liver and lethality in mice following inoculation with bacteria. RESULTS: Susceptibility tests showed that DeltaOmpK36 contributed to the resistance to cefazolin and cefoxitin but not to resistance to late-generation cephalosporins. In vitro assays demonstrated that loss of OmpK36 decreased the resistance to neutrophil phagocytosis and increased the resistance to serum killing during the first hour of the assay, but did not influence the growth rate when compared with the parental strain. Intraperitoneal injection of similar doses ( approximately 4 x 10(4) cfu) of the parental strain and DeltaOmpK36 led to significantly fewer viable bacteria in the liver 24 h post-inoculation in DeltaOmpK36-inoculated mice. In the mice LD(50) (the bacterial dose that caused 50% death) assay, the parental strain was approximately 100-fold more lethal ( approximately 10(3) cfu) than the DeltaOmpK36 mutant ( approximately 10(5) cfu). CONCLUSIONS: Loss of OmpK36 in K. pneumoniae resulted in increased antimicrobial resistance, increased susceptibility to neutrophil phagocytosis, increased resistance to serum killing and reduced virulence.

First identification of blaOXA-51-like in non-baumannii Acinetobacter spp.

Bla(OXA-51-like), the intrinsic carbapenemase gene in Acinetobacter baumannii previously found only in this species, was detected in a clinical isolate of Acinetobacter genomic species 13tU. this study aimed to characterize this gene in the isolate. Genomic species identification was confirmed by amplified ribosomal DNA restriction analysis and sequence analysis of 16S-23S ribosomal DNA intergenic spacer, rpoB and recA. The bla(OXA-51-like) gene, with an upstream ISAba1 insertion, was plasmid-encoded and the surrounding sequences suggested that its origin was from A. baumannii. Transformation of Acinetobacter genomic species 13TU AtCC 17903 with recombinant plasmid bearing ISAba1-bla(OXA-51-like) from the isolate increased the minimum inhibitory concentrations (MICs) of meropenem and imipenem 256-fold. This is the first report of bla(OXA-51-like) in an organism other than A. baumannii. This plasmid-borne bla(OXA-51-like) gene with an upstream ISAba1 insertion confers a high level of carbapenem resistance to Acinetobacter genomic species 13TU.

Methicillin-resistant Staphylococcus aureus and Acinetobacter baumannii on computer interface surfaces of hospital wards and association with clinical isolates.

BACKGROUND: Computer keyboards and mice are potential reservoirs of nosocomial pathogens, but routine disinfection for non-water-proof computer devices is a problem. With better hand hygiene compliance of health-care workers (HCWs), the impact of these potential sources of contamination on clinical infection needs to be clarified. METHODS: This study was conducted in a 1600-bed medical center of southern Taiwan with 47 wards and 282 computers. With education and monitoring program of hand hygiene for HCWs, the average compliance rate was 74% before our surveillance. We investigated the association of methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa and Acinetobacter baumannii, three leading hospital-acquired pathogens, from ward computer keyboards, mice and from clinical isolates in non-outbreak period by pulsed field gel electrophoresis and antibiogram. RESULTS: Our results revealed a 17.4% (49/282) contamination rate of these computer devices by S. aureus, Acinetobacter spp. or Pseudomonas spp. The contamination rates of MRSA and A. baumannii in the ward computers were 1.1% and 4.3%, respectively. No P. aeruginosa was isolated. All isolates from computers and clinical specimens at the same ward showed different pulsotypes. However, A. baumannii isolates on two ward computers had the same pulsotype. CONCLUSION: With good hand hygiene compliance, we found relatively low contamination rates of MRSA, P. aeruginosa and A. baumannii on ward computer interface, and without further contribution to nosocomial infection. Our results suggested no necessity of routine culture surveillance in non-outbreak situation.

Recurrent Klebsiella pneumoniae liver abscess: clinical and microbiological characteristics.

Recurrent Klebsiella pneumoniae liver abscesses (KLAs) are rarely reported. Six cases of recurrent KLAs are characterized. Most of the patients had diabetes and K1 serotype KLAs. All of the isolates were uniformly susceptible to cefazolin. Distinct molecular fingerprints were found for the strains isolated from both primary and recurrent KLAs.

Emergence of carbapenem-resistant Escherichia coli in Taiwan: resistance due to combined CMY-2 production and porin deficiency.

Eight pairs of Escherichia coli isolates with various carbapenem susceptibilities from 8 patients were prospectively collected to study the development of resistance. All carbapenem-resistant E. coli isolates were resistant to all tested ss-lactams antibiotics except tigecycline. Identical pulsed-field gel electrophoresis (PFGE) patterns were found in carbapenem-susceptible and -resistant isolates but different PFGE patterns occurred among patients. A CMY-2 ss-lactamase was found in all E. coli isolates. No previously reported carbapenemase genes were detected. Examination of outer membrane protein (OMP) profiles revealed that OmpA was not found in all isolates, while OmpC and OmpF were lost in carbapenem-resistant isolates. Loss of both OmpC and OmpF represents the major mechanism of the development of carbapenem resistance in those patients with CMY-2-producing E. coli infections.

Widespread dissemination of aminoglycoside resistance genes armA and rmtB in Klebsiella pneumoniae isolates in Taiwan producing CTX-M-type extended-spectrum beta-lactamases.

Among 235 extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL) isolates collected from a nationwide surveillance performed in Taiwan, 102 (43.4%) were resistant to amikacin. Ninety-two of these 102 (90.2%) isolates were carrying CTX-M-type beta-lactamases individually or concomitantly with SHV-type or CMY-2 beta-lactamases. The armA and rmtB alleles were individually detected in 44 and 37 of these 92 isolates, respectively. One isolate contained both armA and rmtB. The coexistence of the aac(6')-Il and rmtB genes was detected in three isolates. CTX-M-type beta-lactamase genes belonging to either group 1 (CTX-M-3 and CTX-M-15) or group 9 (CTX-M-14) were found in all armA- or rmtB-bearing ESBL-producing K. pneumoniae isolates, and all were conjugatively transferable. All except one of the isolates bearing armA produced CTX-M enzymes of group 1, and the remaining isolate bearing armA produced a group 9 CTX-M-type beta-lactamase. On the contrary, in the majority of rmtB carriers, the CTX-M-type beta-lactamase belonged to group 9 (62.2%). Molecular typing revealed that the amikacin-resistant ESBL-producing K. pneumoniae isolates were epidemiologically unrelated, indicating that the acquisition of resistance was not through the spread of a resistant clone or a resistance plasmid. A tandem repeat or multiple copies of bla(CTX-M-3) were found in some armA-bearing isolates. An ISEcp1 insert was found in all CTX-M ESBL-producing K. pneumoniae isolates carrying armA or rmtB. In conclusion, the concomitant presence of a 16S rRNA methylase gene (armA or rmtB) and bla(CTX-M) among amikacin-resistant ESBL-producing K. pneumoniae isolates is widespread in Taiwan.

Dissemination of multidrug-resistant, class 1 integron-carrying Acinetobacter baumannii isolates in Taiwan.

In this study, 283 multidrug-resistant Acinetobacter baumannii (MDR-AB) bloodstream isolates were collected between 1996 and 2004, from three teaching hospitals located in different regions of Taiwan. Susceptibility data showed that strains carrying class 1 integrons were significantly more resistant (p <0.01) to all tested antibiotics (except aztreonam and chloramphenicol) than strains lacking integrons, Seven types of gene cassette were identified among these strains, including two that have not been previously reported. The vast majority of the cassettes encoded aminoglycoside resistance genes, including aacA4, aacC1, aac(6')-II, aadA1, aadA2, aadA4 and aadDA1. Sixteen distinct ribotypes were identified in MDR-AB isolates carrying class 1 integrons. Only one strain was found to produce an extended-spectrum beta-lactamase, i.e. VEB-3. In the 18 imipenem-resistant strains, two carbapenenmase genes, bla(VIM-11) and bla(OXA-58), were found concomitantly in one isolate. An island-wide epidemic clone and an endemic clone from a hospital located in the northern region were identified by ribotyping. On the basis of the susceptibility data among the different ribogroups, the epidemic clone was associated more significantly with resistance to cefepime and ampicillin-sulbactam than was the endemic clone. In conclusion, the presence of class 1 integrons was significantly associated with resistance in MDR-AB, and the epidemic, class 1 integron-carrying MDR-AB clone was found to be widespread in Taiwan.