First Isolation of Vancomycin-Resistant Enterococcus faecium Carrying Plasmid-Borne vanD1.

Vancomycin-resistant Enterococcus faecium carrying the vanD1 gene on plasmid pEF-D was isolated from a fecal sample of a hospitalized patient in Japan. The strain JH5687 showed moderate resistance to vancomycin (MIC, 16 µg/mL) but remained susceptible to teicoplanin (MIC, 1 µg/mL). The backbone gene organization of pEF-D was highly homologous to that of conjugative plasmid pMG1 or pHTß. The calculated conjugation frequency of JH5687 was 10-4 to 10-5 per donor cell.

Is elevation of the serum ß-d-glucan level a paradoxical sign for trichosporon fungemia in patients with hematologic disorders?

The detection of serum 1,3-ß-d-glucan (BDG) has been reported to be useful for the diagnosis and therapeutic monitoring of various invasive fungal infections. Although Trichosporon fungemia is increasingly recognized as a fatal mycosis in immunocompromised patients, the utility of this assay for Trichosporon fungemia is still unknown. In our experience (28 cases), the level of BDG rose in about half of the patients with hematologic disorders who developed Trichosporon fungemia. Among them, early death from this infection was more frequently seen in BDG-negative patients than in BDG-positive patients. In addition, overall survival was also significantly worse in BDG-negative patients than in BDG-positive patients. There were no significant differences between these two patient groups in terms of clinical background. Unlike for other invasive fungal infections, elevation of BDG level may indicate a paradoxical sign for Trichosporon fungemia in patients with hematologic disorders.

Fatal Trichosporon fungemia in patients with hematologic malignancies.

OBJECTIVE: Invasive Trichosporon infection has been increasingly recognized in patients with hematologic malignancies. Our study aims to clarify the clinical characteristics of this disease and factors influencing patient prognosis. PATIENTS AND METHODS: We retrospectively analyzed 33 cases of Trichosporon fungemia (TF) in patients with hematologic malignancies treated at our collaborating five hospitals in Japan between 1992 and 2007. RESULTS: The majority of these patients had acute leukemia (82%), neutropenia (85%), and a history of intensive chemotherapy (91%). TF occurred as a breakthrough infection during antifungal therapy in 30 patients (91%), 18 of whom were receiving micafungin. The surveillance cultures of most patients were negative for Trichosporon. Only a few patients exhibited elevated levels of 1,3-beta-d-glucan before positive blood culture. Twenty-five patients (76%) died of this infection. The resolution of infection was associated with neutrophil recovery (P = 0.0001), absence of hyperglycemia (P = 0.023), and azole inclusive therapy (P = 0.031). Survival was significantly longer in patients receiving antifungal therapies containing azole than in those who did not receive azole (P = 0.0034). CONCLUSIONS: At present, the diagnosis of invasive trichosporonosis depends on blood culture studies, and the mortality of this disease is high; however, azole therapy and control of blood glucose level, together with hematopoietic recovery could help in improving the clinical outcome. When we use antifungals lacking anti-Trichosporon activity, sufficient care should be taken to prevent the development of breakthrough trichosporonosis.

Increased prevalence and clonal dissemination of multidrug-resistant Pseudomonas aeruginosa with the blaIMP-1 gene cassette in Hiroshima.

OBJECTIVES: The aim of this study was to evaluate the dissemination of metallo-beta-lactamase (MBL)-encoding genes among multidrug-resistant (MDR) Pseudomonas aeruginosa isolates recovered from major hospitals in the Hiroshima region. METHODS: During July to December from 2004 to 2006, a surveillance of eight major hospitals in the Hiroshima region identified 387 non-duplicate isolates resistant to imipenem (MIC >or= 16 mg/L). They were screened for resistance to amikacin (MIC >or= 64 mg/L) and ciprofloxacin (MIC >or= 4 mg/L) and MBL-encoding genes. The structure of the variable regions of the integrons was determined using PCR mapping. Clonality was assessed using PFGE and multilocus sequence typing (MLST). RESULTS: The frequency of MBL-positive isolates in MDR P. aeruginosa isolates significantly increased from 42.3% in 2004 to 81.4% in 2006. Most of the MBL-positive isolates produced IMP-1 followed by VIM-2. The bla(IMP-1) and bla(VIM-2) genes were present in class 1 integrons. Characterization of the variable regions of the integron showed the presence of six different gene cassette arrays in bla(IMP-1) cassettes and a single array in bla(VIM-2) cassettes. The IMP-1 producers belonged to two clonal lineages using PFGE and MLST analyses and the integron variations correlated well with the clonal complexes. Among them, strains positive for a newly identified In113-derived bla(IMP-1) gene cassette array were most widely distributed in Hiroshima. CONCLUSIONS: This study shows a dramatic increase in MBL genes, primarily bla(IMP-1), in MDR P. aeruginosa isolates in Hiroshima during these 3 years. In addition, MDR P. aeruginosa with the newly discovered In113-derived bla(IMP-1) gene cassette array appears to be clonally expanding.

Molecular characterization of imipenem-resistant Pseudomonas aeruginosa in Hiroshima, Japan.

Pseudomonas aeruginosa showing resistance to imipenem were found in 100 of 1,058 strains (9.5%) from six hospitals (a-f) in Hiroshima City, Japan. Of the 100 strains, 14 (14%) were double disk synergy test positive using sodium mercaptoacetic acid disks, and 18 (18%) were bla(IMP-1) or bla(VIM-2) allele positive by polymerase chain reaction (PCR). Among 100 imipenem-resistant strains, 32 were categorized into multi-drug resistant strains, in which 13 were positive for the metallo-beta-lactamase gene. Fifty-one strains (51%) among the 100 imipenem-resistant strains had elevated RND efflux pump activity against levofloxacin. But only 6 of 51 strains were classified as multi-drug resistant strains. The pulsed field gel electrophoresis analysis of the Spe I-digested DNA from the 100 isolates suggested not only clonal spread but spread of heterogeneous clones started to contribute to the prevalence of metallo-beta-lactamase producing P. aeruginosa strains in Japanese hospitals.

Nocardia araoensis sp. nov. and Nocardia pneumoniae sp. nov., isolated from patients in Japan.

Two actinomycete strains isolated from two patients with lung nocardiosis between 1995 and 1997 in Japan were assigned to novel species of the genus Nocardia based on morphological and chemical criteria. Comparative 16S rRNA gene sequence analysis of the two strains revealed that they belong to the genus Nocardia and are most closely related to the species Nocardia beijingensis. Determination of DNA-DNA relatedness indicated that these strains could be assigned to two novel species. Based on their phenotypic and phylogenetic characters, two novel species of the genus Nocardia are proposed: Nocardia araoensis sp. nov. for IFM 0575(T) (=NBRC 100135(T)=JCM 12118(T)=DSM 44729(T)) and Nocardia pneumoniae sp. nov. for IFM 0784(T) (=NBRC 100136(T)=JCM 12119(T)=DSM 44730(T)).

Legionella micdadei pneumonia diagnosed by culture isolation and DNA-dNA hybridization from bronchial lavage fluid.

An 80-year-old man was admitted because of dyspnea on effort. We suspected an acute exacerbation of chronic heart failure and idiopathic interstitial pneumonia caused by right-sided pneumonia. A nodular shadow in right upper lobe spread and consolidated into the airspace, and it failed to improve despite administration of meropenem trihydrate, vancomycin hydrochloride and clindamycin. A definitive diagnosis of Legionella micdadei pneumonia was made on the basis of this organism being isolated in culture from bronchial lavage fluid and subsequent identification of Legionella micdadei using DNA-DNA hybridization. The airspace consolidation gradually improved following treatment with intravenous erythromycin and minocycline hydrochloride.

[Detection of MRSA with antimicrobial susceptibility by using chemiluminescent assay].

Chemiluminescent assay can give the result of the detection of MRSA about 13 hours more rapidly than conventional broth microdilution method. In order to apply chemiluminescent assay to detection of MRSA, we compared MIC and antimicrobial susceptibility to MPIPC in using chemiluminescent assay with these in using broth microdilution method. In MSSA, rate of concordance of MIC and antimicrobial susceptibility to MPIPC obtained by both methods was 87%, but all MICs come to be agreed by modifying the concentration of bacterial liquid. In MRSA, all MICs and susceptibility to MPIPC are agreed in both methods. Although we have used chemiluminescent assay to detect MRSA for one year, no trouble has been reported by clinical side. The chemiluminescent assay is evaluated to be good in detecting MRSA.

Molecular epidemiology of invasive stomatitis due to Aspergillus flavus in patients with acute leukemia.

BACKGROUND: Invasive oral aspergillosis is a rare complication and only little information on the epidemiology of Aspergillus flavus infection is available. We present here the molecular analysis of the epidemiology of invasive stomatitis due to Aspergillus flavus in patients with acute leukemia. METHODS: During a 5-year period (1992-1996), six isolates of A. flavus were obtained from leukemic patients with invasive Aspergillus stomatitis. Random amplification of polymorphic DNA (RAPD) with three different PCR primers was carried out to investigate the DNA typing of the isolates. RESULTS: The molecular analysis using RAPD revealed that three isolates of A. flavus obtained in 1992 from three patients were of the same type, whereas each of the isolates from the other three patients had a distinct unique band, resulting in four groups of A. flavus. CONCLUSION: As the three patients with invasive oral aspergillosis detected in 1992 were infected by a single strain of A. flavus, the strain was suspected to have caused a nosocomial outbreak of invasive oral aspergillosis in the hematology unit.

[Evaluation of the Rapid Lumi 'Eiken' using chemiluminescent assay for rapid antimicrobial susceptibility testing].

One hundred ninety eight clinical isolates, including Enterobacteriaceae (70 strains), Pseudomonas aeruginosa (20 strains), Acinetobacter baumannii (10 strains), staphylococci (50 strains), enterococci (20 strains), Streptococcus pneumoniae (15 strains) and Haemophilus influenzae (13 strains) were tested for their antimicrobial susceptibilities by using the Rapid Lumi 'Eiken' (RL). As a reference method, broth microdilution method according to the National Committee for Clinical Laboratory Standards was used. Then, each MIC obtained by both of these methods was compared. In order to improve the discrepancy between MICs obtained by both methods, modification of the RL method was studied. All MICs using the RL method were obtained with an incubation period of 4 hours. The essential agreement (to within one twofold dilution) between MICs obtained by both methods was 82%. The false susceptible in the RL method test results (very major error) and the false resistant in the RL tests (major error) were 0.9% and 2.3%, respectively. The agreement of interpretive category (that is, when the categories obtained by both methods are in perfect agreement) was 89%. Proteus spp. and A. baumannii showed low essential agreements, 59% and 46% respectively. The differences were resulted from the RL method's MICs being higher than the reference method. In order to improve the difference between both methods, the RL method's procedure was modified in the inoculum size (10(6) CFU/mL to 10(5) CFU/mL), the menadione concentration (5 mg/L to 25 mg/L) and the interpretive criteria for Enterobacteriaceae and A. baumannii. As the results of the modification, the essential agreement in Proteus spp. and A. baumannii increased to 82% and 81%, respectively, and there was no significant change in the other species of Enterobacteriaceae. In the case of the modified RL method to Enterobacteriaceae and A. baumannii, the essential agreement, the very major error, the major error and the agreement of interpretive category of all 198 strains were 87%, 1.4%, 1.5% and 90%, respectively. In conclusion, with only 4-hour incubation period, the RL method based on chemiluminescent assay gave reliable susceptibility testing among the most clinically important bacteria. Although several tests showed low essential agreement, it was possible to improve by use of the modified RL method. The Rapid Lumi 'Eiken' will provide useful information for choosing the most effective antibiotic for primary treatment to bacterial infections.