Capillary Electrophoresis–Single-Strand Conformation Polymorphism Analysis for Rapid Identification of Pseudomonas aeruginosa and Other Gram-Negative Nonfermenting Bacilli Recovered from Patients with Cystic Fibrosis

We used capillary electrophoresis-single-strand conformation polymorphism (CE-SSCP) analysis of PCR-amplified 16S rRNA gene fragments for rapid identification of Pseudomonas aeruginosa and other gram-negative nonfermenting bacilli isolated from patients with cystic fibrosis (CF). Target sequences were amplified by using forward and reverse primers labeled with various fluorescent dyes. The labeled PCR products were denatured by heating and separated by capillary gel electrophoresis with an automated DNA sequencer. Data were analyzed with GeneScan 672 software. This program made it possible to control lane-to-lane variability by standardizing the peak positions relative to internal DNA size markers. Thirty-four reference strains belonging to the genera Pseudomonas, Brevundimonas, Burkholderia, Comamonas, Ralstonia, Stenotrophomonas, and Alcaligenes were tested with primer sets spanning 16S rRNA gene regions with various degrees of polymorphism. The best results were obtained with the primer set P11P-P13P, which spans a moderately polymorphic region (Escherichia coli 16S rRNA positions 1173 to 1389 [M. N. Widjojoatmodjo, A. C. Fluit, and J. Verhoef, J. Clin. Microbiol. 32:3002-3007, 1994]). This primer set differentiated the main CF pathogens from closely related species but did not distinguish P. aeruginosa from Pseudomonas alcaligenes-Pseudomonas pseudoalcaligenes and Alcaligenes xylosoxidans from Alcaligenes denitrificans. Two hundred seven CF clinical isolates (153 of P. aeruginosa, 26 of Stenotrophomonas maltophilia, 15 of Burkholderia spp., and 13 of A. xylosoxidans) were tested with P11P-P13P. The CE-SSCP patterns obtained were identical to those for the corresponding reference strains. Fluorescence-based CE-SSCP analysis is simple to use, gives highly reproducible results, and makes it possible to analyze a large number of strains. This approach is suited for the rapid identification of the main gram-negative nonfermenting bacilli encountered in CF.     

In vitro antimicrobial activity of "last-resort" antibiotics against unusual nonfermenting Gram-negative bacilli clinical isolates

In this prospective multicentric study, we assessed the in vitro antimicrobial activity of carbapenems (imipenem, meropenem, and doripenem), tigecycline, and colistin against 166 unusual nonfermenting Gram-negative bacilli (NF-GNB) clinical isolates collected from nine French hospitals during a 6-month period (from December 1, 2008, to May 31, 2009). All NF-GNB isolates were included, except those phenotypically identified as Pseudomonas aeruginosa or Acinetobacter baumannii. Minimal inhibitory concentrations (MICs) of antimicrobial agents were determined by using the E-test technique. The following microorganisms were identified: Stenotrophomonas maltophilia (n=72), Pseudomonas spp. (n=30), Achromobacter xylosoxidans (n=25), Acinetobacter spp. (n=18), Burkholderia cepacia complex (n=9), Alcaligenes faecalis (n=7), and Delftia spp. (n=5). All isolates of Acinetobacter spp., A. faecalis, and Delftia spp. were susceptible to the three carbapenems. Imipenem exhibited the lowest MICs against Pseudomonas spp., and meropenem, as compared with imipenem and doripenem, displayed an interesting antimicrobial activity against A. xylosoxidans and B. cepacia complex isolates. Conversely, no carbapenem exhibited any activity against S. maltophilia. Except for S. maltophilia isolates, tigecycline and colistin exhibited higher MICs than carbapenems, but covered most of the microorganisms tested in this study. To our knowledge, no prior study has compared antimicrobial activity of these five antibiotics, often considered as "last-resort" treatment options for resistant Gram-negative infections, against unusual NF-GNB clinical isolates. Further studies should be carried out to assess the potential clinical use of these antibiotics for the treatment of infections due to these microorganisms.     

Ribosomal DNA-directed PCR for identification of Achromobacter (Alcaligenes) xylosoxidans recovered from sputum samples from cystic fibrosis patients

The opportunistic human pathogen Achromobacter (Alcaligenes) xylosoxidans has been recovered with increasing frequency from respiratory tract culture of persons with cystic fibrosis (CF). However, confusion of this species with other closely related respiratory pathogens has limited studies to better elucidate its epidemiology, natural history, and pathogenic role in CF. Misidentification of A. xylosoxidans as Burkholderia cepacia complex is especially problematic and presents a challenge to effective infection control in CF. To address the problem of accurate identification of A. xylosoxidans, we developed a PCR assay based on a 16S ribosomal DNA sequence. In an analysis of 149 isolates that included 47 A. xylosoxidans and several related glucose-nonfermenting species recovered from CF sputum, the sensitivity and specificity of this PCR assay were determined to be 100 and 97%, respectively. The availability of this assay will enhance identification of A. xylosoxidans, thereby facilitating study of the pathogenic role of this species and improving infection control efforts in CF.     

P594 Intracellular killing of drug-resistant Staphylococcus aureus by different antibiotics

Objectives:  The aim of this study was to assess the carbapenem susceptibility of four nosocomial pathogens and to evaluate the reliability of the susceptibility results determined by E-test and disc diffusion (DD) methods.
Methods:  Escherichia coli ( n  = 73), Klebsiella pneumoniae ( n  = 60), Pseudomonas aeruginosa ( n  = 70) and Acinetobacter spp. ( n  = 70) isolated from nosocomial infections in 2002–2003 were included in the study. Thirty-five per cent of the strains were isolated from intensive care units. After determining antimicrobial susceptibility against imipenem and meropenem by DD (10  μ g; Oxoid, UK) and Etest (AB Biodisk, Solna, Sweden) methods, the results were categorised as susceptible (S), intermediate (I) and resistant (R) according to the NCCLS criteria. For statistical analyses, the intermediate group was included in the resistant category because of the low numbers of bacteria in the former group.
Results:  None of E. coli or K. pneumoniae strains were resistant to carbapenems, whereas, resistance reached up to 59.0% in Acinetobacter spp. and P. aeruginosa isolates. By either method, the pattern of the susceptibility of the four bacteria was not statistically significantly different for imipenem vs. meropenem. Total agreement of DD and E-test methods for susceptibility to imipenem was 95.7%, and 90.0% in Acinetobacter spp. and P. aeruginosa , respectively; and susceptibility to meropenem was 90.0% for both bacteria. However, the difference of the results obtained by either method was statistically significant for Acinetobacter spp.
Conclusion:  Study results suggest a high resistance rate for Acinetobacter spp. and P. aeruginosa strains against carbapenem antibiotics in our hospital. Further studies are needed to clarify whether E-test should be used to confirm meropenem resistance of Acinetobacter spp. determined by DD method.     

Antimicrobial susceptibility testing of carbapenems: multicenter validity testing and accuracy levels of five antimicrobial test methods for detecting resistance in Enterobacteriaceae and Pseudomonas aeruginosa isolates

From January 1996 to May 1999, Project ICARE (Intensive Care Antimicrobial Resistance Epidemiology) received 448 nonduplicate clinical isolates of Enterobacteriaceae and Pseudomonas aeruginosa that were reported to be imipenem intermediate or resistant. However, broth microdilution (BMD) confirmatory testing at the Project ICARE central laboratory confirmed this result in only 11 of 123 (8.9%) Enterobacteriaceae isolates and 241 of 325 (74.2%) P. aeruginosa isolates. To investigate this overdetection of imipenem resistance, we tested 204 selected isolates from the Project ICARE collection plus five imipenem-resistant challenge strains at the Centers for Disease Control and Prevention against imipenem and meropenem by agar dilution, disk diffusion, Etest (AB BIODISK North America, Inc., Piscataway, N.J.), two MicroScan WalkAway conventional panels (Neg MIC Plus 3 and Neg Urine Combo 3) (Dade MicroScan, Inc., West Sacramento, Calif.), and two Vitek cards (GNS-116 containing meropenem and GNS-F7 containing imipenem) (bioMérieux Vitek, Inc., Durham, N.C.). The results of each test method were compared to the results of BMD testing using in-house-prepared panels. Seven imipenem-resistant and five meropenem-resistant isolates of Enterobacteriaceae and 43 imipenem-resistant and 21 meropenem-resistant isolates of P. aeruginosa were identified by BMD. For Enterobacteriaceae, the imipenem and meropenem test methods produced low numbers of very major and major errors. All test systems in the study produced low numbers of very major and major errors when P. aeruginosa was tested against imipenem and meropenem, except for Vitek testing (major error rate for imipenem, 20%). Further testing conducted in 11 of the participating ICARE hospital laboratories failed to pinpoint the factors responsible for the initial overdetection of imipenem resistance. However, this study demonstrated that carbapenem testing difficulties do exist and that laboratories should consider using a second, independent antimicrobial susceptibility testing method to validate carbapenem-intermediate and -resistant results.     

Antimicrobial activity of doripenem and other carbapenems against gram-negative pathogens from Korea

A total of 950 gram-negative bacterial isolates from patients with bacteremia and urinary tract infections were collected from tertiary-care hospitals in Korea. In vitro antimicrobial susceptibility testing was performed using broth microdilution test according to Clinical and Laboratory Standards Institute protocol. In general, carbapenems such as doripenem, imipenem, and meropenem were very active against Enterobacteriaceae, Moraxella catarrhalis, Pseudomonas aeruginosa, and Acinetobacter sp. isolates. Doripenem was more potent than imipenem against most Enterobacteriaceae species except Proteus spp. based on minimum inhibitory concentration (MIC)(50) and MIC(90). In addition, doripenem displayed similar activity to meropenem but was superior to imipenem against ceftazidime-resistant Enterobacteriaceae isolates. For P. aeruginosa and Acinetobacter spp. isolates, MIC(50)s of doripenem were 1 and 0.5 mg/L, respectively, which were the same with those of meropenem but two- to fourfold lower than those of imipenem (both 2 mg/L). On the basis of the in vitro data, we conclude that doripenem has equivalent or more activity than other carbapenems such as imipenem and meropenem against most gram-negative pathogens from Korea. Thus, doripenem may be a promising new antimicrobial agent for the treatment of infections caused by gram-negative pathogens in Korea.     

The activity of meropenem and comparators against Acinetobacter strains isolated from European hospitals, 1997–2000

In vitro susceptibilities to meropenem and comparators of Acinetobacter strains isolated from serious infections in 37 European hospital centers participating in the Meropenem Yearly Susceptibility Test Information Collection (MYSTIC) Program (1997–2000) were tested. There were 635 Acinetobacter strains collected: 490 A. baumannii ; 51 A. calcoaceticus var . lwoffii ; and 94 other Acinetobacter strains. Overall, meropenem and imipenem were the most effective agents tested. Resistance to the antimicrobials was: 14%, meropenem; 16%, imipenem; 39%, piperacillin–tazobactam; 41%, tobramycin; 45%, ceftazidime; and 53%, ciprofloxacin. Thus, the carbapenems have useful activity against Acinetobacter spp. and represent a viable choice for treating infections caused by these organisms.     

Identification of Pandoraea Species by 16S Ribosomal DNA-Based PCR Assays

The recently described genus Pandoraea contains five named species (Pandoraea apista, Pandoraea pulmonicola, Pandoraea pnomenusa, Pandoraea sputorum, and Pandoraea norimbergensis) and four unnamed genomospecies. Pandoraea spp. have mainly been recovered from the respiratory tracts of cystic fibrosis (CF) patients. Accurate genus- and species-level identification by routine clinical microbiology methods is difficult, and differentiation from Burkholderia cepacia complex organisms may be especially problematic. This can have important consequences for the management of CF patients. On the basis of 16S ribosomal DNA sequences, PCR assays for the identification of Pandoraea spp. were developed. A first PCR assay was developed for the identification of Pandoraea isolates to the genus level. PCR assays for the identification of P. apista and P. pulmonicola as a group, P. pnomenusa, P. sputorum, and P. norimbergensis were also developed. All five assays were evaluated with a panel of 123 bacterial isolates that included 69 Pandoraea sp. strains, 24 B. cepacia complex strains, 6 Burkholderia gladioli strains, 9 Ralstonia sp. strains, 5 Alcaligenes xylosoxidans strains, 5 Stenotrophomonas maltophilia strains, and 5 Pseudomonas aeruginosa strains. The use of these PCR assays facilitates the identification of Pandoraea spp. and avoids the misidentification of a Pandoraea sp. as a B. cepacia complex isolate.     

Evaluation of the 4-hour RapID NF Plus method for identification of 345 gram-negative nonfermentative rods.

The ability of the RapID NF Plus system (Innovative Diagnostic Systems, Inc., Atlanta, Ga.) to identify 345 nonfermentative gram-negative rods was evaluated. Kits were inoculated with no. 1 McFarland suspensions, and reactions were interpreted after a 4-h incubation at 35 degrees C. Overall, the method correctly identified 311 strains (90.1%) without additional tests and 21 strains (6.1%) with additional tests, and 13 strains (3.8%) were misidentified. Five of 13 misidentified strains were Alcaligenes faecalis-Alcaligenes odorans misidentified as Alcaligenes xylosoxidans; however, all strains were xylose negative but nitrate positive and could have been A. faecalis group I-Alcaligenes piechaudii. The system does not differentiate between Pseudomonas fluorescens and Pseudomonas putida, and all Acinetobacter species are identified as Acetinobacter calcoaceticus. Additionally, no subspecies differentiation is made between A. xylosoxidans subsp. xylosoxidans and A. xylosoxidans subsp. denitrificans. All strains of the former Flavobacterium group IIb are identified as Flavobacterium indologenes-Flavobacterium gleum, and no species identification of the genus Methylobacterium is attempted. The system is easy to set up and interpret and provides an accurate commercial nonautomated method for same-day identification of gram-negative nonfermenters.     

Use of random amplified polymorphic DNA PCR to examine epidemiology of Stenotrophomonas maltophilia and Achromobacter (Alcaligenes) xylosoxidans from patients with cystic fibrosis

Stenotrophomonas maltophilia and Achromobacter (Alcaligenes) xylosoxidans have been increasingly recognized as a cause of respiratory tract colonization in cystic fibrosis (CF). Although both organisms have been associated with progressive deterioration of pulmonary function, demonstration of causality is lacking. To examine the molecular epidemiology of S. maltophilia and A. xylosoxidans in CF, isolates from patients monitored for up to 2 years were fingerprinted using a PCR-based randomly amplified polymorphic DNA (RAPD-PCR) method. Sixty-one of 69 CF centers screened had 183 S. maltophilia culture-positive patients, and 46 centers had 92 A. xylosoxidans-positive patients. At least one isolate from each patient was genotyped, and patients with > or =10 positive cultures (12 S. maltophilia cultures, 15 A. xylosoxidans cultures) had serial isolates genotyped. In addition, centers with multiple culture-positive patients were examined for evidence of shared clones. There were no instances of shared genotypes among different CF centers. Some patients demonstrated isolates with a single genotype throughout the observation period, and others had intervening or sequential genotypes. At the six centers with multiple S. maltophilia culture-positive patients and the seven centers with multiple A. xylosoxidans-positive patients, there were three and five instances of shared genotypes, respectively. The majority of shared isolates were from pairs who were siblings or otherwise epidemiologically linked. These findings suggest RAPD-PCR typing can distinguish unique CF isolates of S. maltophilia and A. xylosoxidans, person-to-person transmission may occur, there are not a small number of clones infecting CF airways, and patients with long-term colonization may either have a persistent organism or may acquire additional organisms over time.     

Fluorescence in situ hybridization for rapid identification of Achromobacter xylosoxidans and Alcaligenes faecalis recovered from cystic fibrosis patients

Achromobacter xylosoxidans is frequently isolated from the respiratory secretions of cystic fibrosis (CF) patients, but identification with biochemical tests is unreliable. We describe fluorescence in situ hybridization assays for the rapid identification of Achromobacter xylosoxidans and Alcaligenes faecalis. Both assays showed high sensitivities and high specificities with a collection of 155 nonfermenters from CF patients.     

Evaluation of the VITEK 2 system for the identification and susceptibility testing of three species of nonfermenting gram-negative rods frequently isolated from clinical samples

VITEK 2 is a new automatic system for the identification and susceptibility testing of the most clinically important bacteria. In the present study 198 clinical isolates, including Pseudomonas aeruginosa (n = 146), Acinetobacter baumannii (n = 25), and Stenotrophomonas maltophilia (n = 27) were evaluated. Reference susceptibility testing of cefepime, cefotaxime, ceftazidime, ciprofloxacin, gentamicin, imipenem, meropenem, piperacillin, tobramycin, levofloxacin (only for P. aeruginosa), co-trimoxazole (only for S. maltophilia), and ampicillin-sulbactam and tetracycline (only for A. baumannii) was performed by microdilution (NCCLS guidelines). The VITEK 2 system correctly identified 91.6, 100, and 76% of P. aeruginosa, S. maltophilia, and A. baumannii isolates, respectively, within 3 h. The respective percentages of essential agreement (to within 1 twofold dilution) for P. aeruginosa and A. baumannii were 89.0 and 88.0% (cefepime), 91.1 and 100% (cefotaxime), 95.2 and 96.0% (ceftazidime), 98.6 and 100% (ciprofloxacin), 88.4 and 100% (gentamicin), 87.0 and 92.0% (imipenem), 85.0 and 88.0% (meropenem), 84.2 and 96.0% (piperacillin), and 97.3 and 80% (tobramycin). The essential agreement for levofloxacin against P. aeruginosa was 86.3%. The percentages of essential agreement for ampicillin-sulbactam and tetracycline against A. baumannii were 88.0 and 100%, respectively. Very major errors for P. aeruginosa (resistant by the reference method, susceptible with the VITEK 2 system [resistant to susceptible]) were noted for cefepime (0.7%), cefotaxime (0.7%), gentamicin (0.7%), imipenem (1.4%), levofloxacin (2.7%), and piperacillin (2.7%) and, for one strain of A. baumannii, for imipenem. Major errors (susceptible to resistant) were noted only for P. aeruginosa and cefepime (2.0%), ceftazidime (0.7%), and piperacillin (3.4%). Minor errors ranged from 0.0% for piperacillin to 22.6% for cefotaxime against P. aeruginosa and from 0.0% for piperacillin and ciprofloxacin to 20.0% for cefepime against A. baumannii. The VITEK 2 system provided co-trimoxazole MICs only for S. maltophilia; no very major or major errors were obtained for co-trimoxazole against this species. It is concluded that the VITEK 2 system allows the rapid identification of S. maltophilia and most P. aeruginosa and A. baumannii isolates. The VITEK 2 system can perform reliable susceptibility testing of many of the antimicrobial agents used against P. aeruginosa and A. baumannii. It would be desirable if new versions of the VITEK 2 software were able to determine MICs and the corresponding clinical categories of agents active against S. maltophilia.     

Evaluation of MicroScan Autoscan for identification of Pseudomonas aeruginosa isolates from cystic fibrosis patients

Accurate identification of gram-negative bacilli from cystic fibrosis (CF) patients is essential. Only 57% (108 of 189) of nonmucoid strains and 40% (24 of 60) of mucoid strains were definitively identified as Pseudomonas aeruginosa with MicroScan Autoscan. Most common misidentifications were Pseudomonas fluorescens-Pseudomonas putida (i.e., the strain was either P. fluorescens or P. putida, but the system did not make the distinction and yielded the result P. fluorescens/putida) and Alcaligenes spp. Extending the incubation to 48 h improved identification, but 15% of isolates remained misidentified. The MicroScan Autoscan system cannot be recommended for the identification of P. aeruginosa isolates from CF patients.     

Comparison of three molecular techniques for typing Pseudomonas aeruginosa isolates in sputum samples from patients with cystic fibrosis

Monitoring the emergence and transmission of Pseudomonas aeruginosa strains among cystic fibrosis (CF) patients is important for infection control in CF centers internationally. A recently developed multilocus sequence typing (MLST) scheme is used for epidemiologic analyses of P. aeruginosa outbreaks; however, little is known about its suitability for isolates from CF patients compared with that of pulsed-field gel electrophoresis (PFGE) and enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR). As part of a prevalence study of P. aeruginosa strains in Australian CF clinics, we compared the discriminatory power and concordance of ERIC-PCR, PFGE, and MLST among 93 CF sputum and 11 control P. aeruginosa isolates. PFGE and MLST analyses were also performed on 30 paired isolates collected 85 to 354 days apart from 30 patients attending two CF centers separated by 3,600 kilometers in order to detect within-host evolution. Each of the three methods displayed high levels of concordance and discrimination; however, overall lower discrimination was seen with ERIC-PCR than with MLST and PFGE. Analysis of the 50 ERIC-PCR types yielded 54 PFGE types, which were related by ≤ 6 band differences, and 59 sequence types, which were classified into 7 BURST groups and 42 singletons. MLST also proved useful for detecting novel and known strains and for inferring relatedness among unique PFGE types. However, 47% of the paired isolates produced PFGE patterns that within 1 year differed by one to five bands, whereas with MLST all paired isolates remained identical. MLST thus represents a categorical analysis tool with resolving power similar to that of PFGE for typing P. aeruginosa. Its focus on highly conserved housekeeping genes is particularly suited for long-term clinical monitoring and detecting novel strains.     

Molecular characterization of a salt-tolerant bacterial community in the rice rhizosphere

The diversity of salt-tolerant bacteria present in the rhizosphere of Oryza sativa was investigated. Fourteen bacterial strains, isolated after enrichment in nitrogen-free, semi-solid medium and showing tolerance to 3% NaCl, were analyzed by restriction patterns produced by amplified DNA coding for 16S rDNA (ARDRA) with enzymes Sau3AI, AluI and RsaI which showed that they were represented by 4 ARDRA types. Biodiversity among the 14 strains was also analyzed by the random amplified polymorphic DNA (RAPD) technique with a 10-mer primer. Partial nucleotide sequence of 16S rDNA assigned these clusters to Serratia marcescens, Pseudomonas aeruginosa, Alcaligenes xylosoxidans and Ochrobactrum anthropi. Notably, all four bacterial species are potential human pathogens that infect immunocompromised patients.     

PCR typing of genetic determinants for metallo-beta-lactamases and integrases carried by gram-negative bacteria isolated in Japan, with focus on the class 3 integron

From January 2001 to December 2002, 587 strains of gram-negative bacterial isolates demonstrating resistance to ceftazidime and a combination of sulbactam and cefoperazone were subjected to a disk diffusion screening test using sodium mercaptoacetic acid; 431 strains (73.4%) appeared to produce metallo-beta-lactamase (MBL). Of these 431 strains, 357 were found by PCR to carry genes for IMP-1 type MBL (bla(IMP-1)), while only 7 and 67 strains carried the IMP-2 gene (bla(IMP-2)) and the VIM-2 gene (bla(VIM-2)), respectively. Neither VIM-1 nor SPM-1 type MBL genes were found among the strains tested. Of 431 strains, 427 carried the intI1 gene, and 4 strains carrying both the intI1 and intI3 genes were reidentified as Pseudomonas putida harboring bla(IMP-1). Of these four P. putida strains, three strains and one strain, respectively, were separately isolated from two hospitals located in the same prefecture, and the three strains showed very similar pulsed-field gel electrophoresis patterns. Of 357 bla(IMP-1) carriers, 116, 53, 51, 47, and 30 strains were identified as Pseudomonas aeruginosa, Alcaligenes xylosoxidans, P. putida/fluorescens, Serratia marcescens, and Acinetobacter baumannii, respectively. Four strains carrying bla(IMP-2) were reidentified as P. putida. Sixty-three P. aeruginosa strains and four P. putida strains carried bla(VIM-2). Of 427 intI1-positive strains, 180, 53, 51, 47, and 35 were identified as P. aeruginosa, A. xylosoxidans, P. putida/fluorescens, S. marcescens, and A. baumannii, respectively. In the present study, it was confirmed that strains carrying bla(IMP-1) with a class 1 integron are the most prevalent type in Japan, although several intI3 carriers have also been identified sporadically in this country.     

南京市江宁医院2015年~2017年血培养阳性病原菌分布及耐药性分析

目的 研究血培养阳性检出的病原菌分布及耐药情况,指导临床合理用药。方法 采用法国梅里埃公司Bact/Alert 3D全自动血培养仪对我院2015年1月~2017年12月送检的血培养标本进行培养,阳性标本用VITEK 2 Compact全自动细菌鉴定及药敏分析系统进行细菌鉴定和药敏试验,观察病原菌分布及耐药情况。结果 共分离出病原菌459株,检出率为9.57%,其中革兰阴性杆菌246株（53.59%）,革兰阳性球菌202株（44.01%）,真菌11株（2.40%）;前3位病原菌依次为大肠埃希菌、肺炎克雷伯菌、金黄色葡萄球菌。大肠埃希菌对美罗培南、阿米卡星、厄他培南、亚胺培南均较敏感,耐药率低于3%。肺炎克雷伯菌对美罗培南、阿米卡星、厄他培南、亚胺培南、头孢替坦较敏感,耐药率低于6%。未检出对万古霉素、利奈唑胺、喹努普汀/达福普汀、替加环素耐药的金黄色葡萄球菌。结论 本院血培养的病原菌种类多,以革兰阴性杆菌为主,且耐药情况复杂。应加强对病原菌的鉴定和药敏监测,指导临床合理用药,防止广谱抗生素的滥用,减少耐药菌株的产生。     

Alcaligenes xylosoxidans bacteremia: clinical features and microbiological characteristics of isolates.

Bacteremia caused by Alcaligenes xylosoxidans is rare. Between 1999 and 2002, 12 cases of bacteremia caused by A. xylosoxidans were diagnosed at a tertiary referral center in central Taiwan. The clinical features of these patients and the antimicrobial susceptibilities and pulsed-field gel electrophoresis (PFGE) pattern of their blood isolates were studied. All infections were acquired nosocomially. All of the adult patients had underlying diseases, and 10 (83%) had undergone an invasive procedure. The clinical syndrome included primary bacteremia in 7 patients (58%), and catheter-associated bacteremia, surgical wound infection, pneumonia, urinary tract infection, and empyema in 1 each. Polymicrobial bacteremia was found in 1 patient. The case-fatality rate was 17% (2/12). All isolates were susceptible to piperacillin and ceftazidime and resistant to aminoglycoside, ciprofloxacin and cefepime. Susceptibility to imipenem (67%), ampicillin-sulbactam (75%) and trimethoprim-sulfamethoxazole (92%) was variable. Genetic fingerprints obtained by PFGE showed identical pattern in the isolates from 2 neonates, indicating the epidemiologic relatedness of these infections. We conclude that A. xylosoxidans isolates are multi-resistant and A. xylosoxidans bacteremia should be considered as a possible etiology of infection after invasive procedures in patients with underlying diseases. Strict infection control is needed to prevent this infection.     

Oligonucleotide array-based identification of species in the Acinetobacter calcoaceticus-A. baumannii complex in isolates from blood cultures and antimicrobial susceptibility testing of the isolates

Acinetobacter calcoaceticus, A. baumannii, Acinetobacter genomic species (gen. sp.) 3, and Acinetobacter gen. sp. 13TU, which are included in the A. calcoaceticus-A. baumannii complex, are difficult to distinguish by phenotypic methods. An array with six oligonucleotide probes based on the 16S-23S rRNA gene intergenic spacer (ITS) region was developed to differentiate species in the A. calcoaceticus-A. baumannii complex. Validation of the array with a reference collection of 52 strains of the A. calcoaceticus-A. baumannii complex and 137 strains of other species resulted in an identification sensitivity and specificity of 100%. By using the array, the species distribution of 291 isolates of the A. calcoaceticus-A. baumannii complex from patients with bacteremia were determined to be A. baumannii (221 strains [75.9%]), Acinetobacter gen. sp. 3 (67 strains [23.0%]), Acinetobacter gen. sp. 13TU (2 strains [0.7%]), and unidentified Acinetobacter sp. (1 strain [0.3%]). The identification accuracy of the array for 12 randomly selected isolates from patients with bacteremia was further confirmed by sequence analyses of the ITS region and the 16S rRNA gene. Antimicrobial susceptibility testing of the 291 isolates from patients with bacteremia revealed that A. baumannii strains were less susceptible to antimicrobial agents than Acinetobacter gen. sp. 3. All Acinetobacter gen. sp. 3 strains were susceptible to ampicillin-sulbactam, imipenem, and meropenem; but only 67.4%, 90%, and 86% of the A. baumannii strains were susceptible to ampicillin-sulbactam, imipenem, and meropenem, respectively. The observed significant variations in antimicrobial susceptibility among different species in the A. calcoaceticus-A. baumannii complex emphasize that the differentiation of species within the complex is relevant from a clinical-epidemiological point of view.     

Imipenem resistance in nonfermenters causing nosocomial urinary tract infections

Nonfermenting gram-negative bacilli (nonfermenters) have emerged as important nosocomial pathogens causing opportunistic infections in immunocompromised hosts. These organisms show high level of resistance to b-lactam agents, fluoroquinolones and aminoglycosides. Imipenem is a carbapenem antibiotic, which can be very useful for treatment of infections caused by nonfermenters. Eighty-five nonfermenters causing nosocomial UTI were tested for MIC to imipenem by agar dilution method. Resistance to other antimicrobial agents was compared between imipenem sensitive (S) and resistance (R) groups. Overall 36.4% of nonfermenters were resistant to imipenem. Forty two percent of P. aeruginosa and 18.5% of Acinetobacter baumanii were imipenem resistant. Other nonfermenters showed variable resistance, resistance in Alcaligenes spp. being very high. More than 70% of the nonfermenters were resistant to ceftazidime, gentamicin and ciprofloxacin. Piperacillin and amikacin had the best in vitro susceptibility. No significant difference was found in the antibiotic susceptibility profile among imipenem sensitive (S) or resistant (R) strains.