Abstract
This study was conducted to compare the prevalences of antimicrobial resistance profiles of clinical isolates in the Acinetobacter calcoaceticus-Acinetobacter baumannii complex from sterile and nonsterile sites and to further study the relationship of antimicrobial resistance profiles and genospecies by amplified rRNA gene restriction analysis (ARDRA). A total of 1,381 isolates were tested with 12 different antibiotics to show their antimicrobial susceptibility profiles. A total of 205 clinical isolates were further analyzed by ARDRA of the intergenic spacer (ITS) region of the 16S-23S rRNA gene. It was found that the overall percentage of isolates from nonsterile sites (urine, sputum, pus, or catheter tip) that were resistant to the 12 antibiotics tested was significantly higher than that of isolates from sterile sites (cerebrospinal fluid [CSF], ascites fluid, and bloodstream) (46% versus 22%; P < 0.05). After ARDRA, it was found that 97% of the 62 isolates resistant to all antibiotics tested were the A. baumannii genospecies, which was identified in only 31% of the isolates susceptible to all antibiotics tested. More genospecies diversity was identified in the isolates susceptible to all antibiotics tested, including genospecies of 13TU (34%), genotype 3 (29%), and A. calcoaceticus (5%). Furthermore, as 91% (10/11) of the isolates from CSF were susceptible to all antibiotics tested, the A. calcoaceticus-A. baumannii complex isolates with multidrug resistance could be less invasive than the more susceptible isolates. This study also indicated current emergence of carbapenem-, fluoroquinolone-, aminoglycoside-, and cephalosporin-resistant A. calcoaceticus-A. baumannii complex isolates in Taiwan.
INTRODUCTION
Members of the genus Acinetobacter have been recognized to be widespread and persistent on a variety of surfaces and inanimate environments (4, 42). The bacteria have recently become one of the emerging nosocomial pathogens (1, 28). Higher mortality could be observed in patients with a long period of hospitalization, on mechanical ventilation, on chemotherapy, and with underlying diseases (8, 29, 33).
Acinetobacter spp. are glucose-nonfermentative, nonfastidious, catalase-positive, oxidase-negative, strictly aerobic Gram-negative coccobacilli and commonly occur in diploid formation or in chains of variable length (32). However, different genospecies cannot be easily identified using traditional methods. Members of the genus have been classified in various ways, and therefore it is difficult to understand the true status of the epidemiology and clinical importance of these organisms (41). Since 1986, the taxonomy of the genus Acinetobacter has undergone extensive revision (2). The original single species named Acinetobacter calcoaceticus has been abandoned, and at least 32 genospecies have now been proposed, 17 of which have been correlated with species names. Identification of the members of the genus Acinetobacter to the species level by traditional methods is problematic (5, 10). Among the different genospecies, A. baumannii and its close relatives (genospecies 3 and 13TU), together forming the “A. calcoaceticus-A. baumannii complex,” belong to the molecular group of greatest clinical importance, which accounts for the vast majority of human infections and hospital outbreaks (13, 31, 39).
In recent years, clinical cases due to A. calcoaceticus-A. baumannii complex infection have raised major concerns worldwide. The infection can be fatal, and clinical A. calcoaceticus-A. baumannii complex isolates were frequently resistant to the last-line antibiotics for clinical treatment, including cephalosporins, aminoglycosides, fluoroquinolones, and carbapenems (30). In Taiwan, several studies further indicated the emergence of the infection, especially with regard to multidrug resistance and nosocomial infections (14, 15, 17). Since 1998, several studies on isolates in the A. calcoaceticus-A. baumannii complex have been conducted in Taiwan, and several genospecies associated with antibiotic resistance have been identified (11, 24, 26). The genotyping methods included restriction analysis of the amplified rRNA gene (amplified rRNA gene restriction analysis [ARDRA]) (3, 40), the 16S-23S rRNA gene intergenic spacer (ITS) region (6), and the whole ribosomal operon (9). ARDRA is easy to conduct and can be considered a useful tool for clinical identification. Through molecular epidemiology, trends in the prevalence of antibiotic resistance in different genospecies have also been documented (24, 26).
Clinically, it is important to differentiate the different genospecies of isolates in the A. calcoaceticus-A. baumannii complex. The main reason is that different genospecies have different biological characteristics and pathogenicity, which could be relevant to treatment efficacy (11). The previous study indicated that genospecies 3 and 13TU had different antibiotic susceptibilities (2, 14). Therefore, there is an urgent need to conduct molecular epidemiological investigations to understand the relationship of genospecies and their antimicrobial resistance profiles. The results will be helpful for better clinical treatment and control of the infection (7, 11, 18).
The objectives of this study were to compare the prevalences of antimicrobial resistance profiles of isolates in the A. calcoaceticus-A. baumannii complex from sterile and nonsterile sites in patients and to further study the relationship of antimicrobial resistance profiles and genospecies by amplified rRNA gene restriction analysis (ARDRA) of the intergenic spacer (ITS) region of the 16S-23S rRNA gene.
MATERIALS AND METHODS
Collection of isolates in the A. calcoaceticus-A. baumannii complex and antibiotic susceptibility testing.
A total of 1,381 isolates in the A. calcoaceticus-A. baumannii complex from clinical specimens from various sites in patients were collected at Teaching Hospital of China Medical University, Taiwan, in 2008. The hospital is a medical center in central Taiwan with more than 2,000 beds. The BD Phoenix NMIC/ID-2 commercial kit (Becton, Dickinson Diagnostic Systems, Sparks, MD) was used for antibiotic susceptibility testing. The antibiotics tested included amikacin (AN) (MIC, 8 to 32 μg/ml), ceftazidime (CAZ) (0.5 to 16 μg/ml), ciprofloxacin (CIP) (0.5 to 2 μg/ml), colistin (CL) (1 to 4 μg/ml), cefepime (FEP) (2 to 16 μg/ml), gentamicin (GM) (2 to 8 μg/ml), imipenem (IPM) (1 to 8 μg/ml), levofloxacin (LVX) (0.5 to 2 μg/ml), meropenem (MEM) (1 to 8 μg/ml), ampicillin-sulbactam (SAM) (4/2 to 16/8 μg/ml), trimethoprim-sulfamethoxazole (SXT) (0.95/9.5 to 2/38 μg/ml), and piperacillin-tazobactam (TZP) (4/4 to 64/4 μg/ml).
DNA extraction and ARDRA for the ITS region of the 16S-23S rRNA gene of isolates in the A. calcoaceticus-A. baumannii complex.
A total of 205 clinical isolates (8 from ascites fluid, 11 from cerebrospinal fluid [CSF], and 186 from bloodstream) collected from 2007 to 2009 were further analyzed by amplified rRNA gene restriction analysis (ARDRA) of the intergenic spacer (ITS) region of the 16S-23S rRNA gene (6). All the isolates and reference strains used in this study were cultured and identified to be in the A. calcoaceticus-A. baumannii complex according to a previous study (32). Briefly, isolates were grown on BAP agar plates (Becton, Dickinson and Company, Sparks, MD), and then pure bacterial colonies were suspended in 600 μl of TE buffer (10 mM Tris, 1 mM EDTA, pH 8.0) and the pellet was obtained after brief centrifugation using a microcentrifuge. DNA was extracted from the pellet with a Genomic DNA Mini Kit (Geneaid, New Taipei City, Taiwan) following the manufacturer's recommendations. The DNA extracts were stored at −20°C for further molecular analysis. For genospecies identification of the isolates, ARDRA for the ITS region of the 16S-23S rRNA gene was applied as previously described by Vaneechoutte et al. (40). Briefly, the specific segment was amplified with primer set 5′-TGGCTCAGATTGAACGCTGGCGGC-3′ and 5′-TACCTTGTTACGACTTCACCCCA-3′. Amplification was performed using 5 μl of DNA extract in a 50-μl PCR mixture containing 1 U of Taq polymerase (Fermentas, Vilnius, Lithuania), 200 mM each deoxynucleoside triphosphate (Viogen, New Taipei City, Taiwan), and 0.2 mM each primer in reaction buffer (1.5 mM MgCl2 and 50 mM KCl in 10 mM Tris-HCl, pH 8.3). The PCR conditions were established as follows: first denaturation at 94°C for 5 min, 35 cycles of denaturation at 95°C for 45 s, annealing at 50°C for 45 s, and extension at 72°C for 1 min, and final extension at 72°C for 7 min. The expected amplified product was approximately 1,500 bp. Three restriction endonucleases, AluI, HhaI, and MboI (New England BioLabs, Beverly, MA), were used for DNA digestion following the manufacturer's instructions. Using 5 μl of a digested PCR product, the restriction fragment pattern of each sample was visualized by 2% agarose (Amresco Inc., Solon, OH) gel electrophoresis at 100 V for 30 min in Tris-borate-EDTA (TBE) buffer. Genospecies were determined by the digestion patterns as described by Vaneechoutte et al. (40).
RESULTS
Analysis of antibiotic susceptibility results for isolates from different clinical specimens.
After using antibiotic susceptibility testing to analyze a total of 1,381 isolates from 2008, the percentages of resistance were found to be 68% for amikacin, 75% for ceftazidime, 73% for ciprofloxacin, 0% for colistin, 75% for cefepime, 75% for gentamicin, 65% for imipenem, 71% for levofloxacin, 68% for meropenem, 67% for ampicillin-sulbactam, 74% for trimethoprim-sulfamethoxazole, and 78% for piperacillin-tazobactam. As shown in Table 1, among the 983 isolates from nonsterile sites, the highest percentage of resistance to all antibiotics tested was in sputum, followed by pus and central venous pressure (CVP) catheter tips. The highest percentage of resistance of all antibiotics tested for the 398 isolates from sterile sites was in ascites fluid, followed by pleural fluid and cerebrospinal fluid (CSF). The overall percentage of isolates from nonsterile sites that were resistant to all antibiotics was significantly higher than that of isolates from sterile sites (46% versus 22%; P < 0.05).
Table 1.
Comparison of percentages of antibiotic-susceptible A. calcoaceticus-A. baumannii complex isolates from different specimens
| Susceptibility to all 12 antibiotics testeda | No. (%) of isolates from: |
|||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Nonsterile areas |
Sterile areas |
|||||||||||
| Bile | Tip | Urine | Pus | Sputum | Total | Body Fluid | Ascites | Pleural fluid | CSF | Bloodstream | Total | |
| Susceptible | 0 (0) | 18 (11) | 15 (7) | 24 (11) | 13 (3) | 70 (7) | 7 (58) | 4 (22) | 3 (16) | 11 (44) | 116 (36) | 141 (35) |
| Resistant | 4 (36) | 78 (46) | 68 (32) | 103 (47) | 204 (55) | 457 (46) | 2 (17) | 6 (33) | 5 (26) | 6 (24) | 69 (21) | 88 (22) |
| Total | 11 | 170 | 210 | 219 | 373 | 983 | 12 | 18 | 19 | 25 | 324 | 398 |
The antibiotics tested were amikacin, ceftazidime, ciprofloxacin, colistin, cefepime, gentamicin, imipenem, levofloxacin, meropenem, ampicillin-sulbactam, trimethoprim-sulfamethoxazole, and piperacillin-tazobactam.
Molecular typing of isolates by ARDRA.
After analyzing 205 isolates (including 62 isolates resistant to the 12 antibiotics tested and 143 isolates susceptible to the 12 antibiotics tested) for genospecies identification by ARDRA, except for one isolate with an undetermined genospecies, 97% of the all resistant isolates were the unique A. baumannii genospecies (Table 2). However, isolates susceptible to all antibiotics tested had more genetic diversity; the major genospecies determined by ARDRA was 13TU (34%), followed by A. baumannii (31%), genotype 3 (29%), and A. calcoaceticus (5%). 13TU, A. baumannii, genotype 3, and A. calcoaceticus could all be identified in blood isolates. However, isolates from ascites fluid or CSF were only A. baumannii and genotype 3.
Table 2.
Comparison of genospecies of A. calcoaceticus-A. baumannii complex isolates in resistant and susceptible groups from different clinical sources
| Susceptibility to all 12 antibiotics testeda | Source of specimens | Genospecies (no. of isolates) |
|---|---|---|
| Resistant (n = 62) | CSF | Undetermined (1) |
| Bloodstream | A. baumannii (56), untypeable (1) | |
| Ascites fluid | A. baumannii (4) | |
| Susceptible (n = 143) | CSF | A. baumannii (5), genotype 3 (5) |
| Bloodstream | 13TU (49), A. baumannii (37), genotype 3 (35), A. calcoaceticus (7), undetermined (1) | |
| Ascites fluid | A. baumannii (2), genotype 3 (2) |
The antibiotics tested were amikacin, ceftazidime, ciprofloxacin, colistin, cefepime, gentamicin, imipenem, levofloxacin, meropenem, ampicillin-sulbactam, trimethoprim-sulfamethoxazole, and piperacillin-tazobactam.
Further analysis of the results by year (Table 3) showed that the A. baumannii genospecies with resistance to all 12 antibiotics tested was dominant in 2007 and 2008. However, in 2009, isolates were still mainly of the A. baumannii genospecies but were susceptible to all 12 antibiotics tested.
Table 3.
Genospecies distribution of A. calcoaceticus-A. baumannii complex isolates, 2007 to 2009
| Genospecies | No. (%) of isolates resistant or susceptiblea in: |
|||||
|---|---|---|---|---|---|---|
| 2007 (n = 53) |
2008 (n = 82) |
2009 (n = 70) |
||||
| Resistant | Susceptible | Resistant | Susceptible | Resistant | Susceptible | |
| A. baumannii | 22 (42) | 5 (9) | 30 (37) | 9 (11) | 9 (13) | 38 (54) |
| A. calcoaceticus | 0 (0) | 0 (0) | 0 (0) | 7 (9) | 0 (0) | 0 (0) |
| Genotype 3 | 1 (0) | 13 (25) | 0 (0) | 14 (17) | 0 (0) | 15 (21) |
| 13TU | 2 (0) | 9 (17) | 0 (0) | 22 (27) | 0 (0) | 18 (26) |
| Undetermined | 1 (2) | 0 (0) | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
Isolates resistant or susceptible to the 12 antibiotics tested in this study.
DISCUSSION
In Taiwan, the emergence of infections due to Acinetobacter bacteria has recently been emphasized with respect to the aspects of multidrug resistance and nosocomial infections (12, 14, 15, 20). Nevertheless, most clinical microbiological laboratories using traditional methods or commercial identification systems could not easily distinguish various genospecies of the isolates in the A. calcoaceticus-A. baumannii complex (32). Therefore, further genotyping of isolates in the A. calcoaceticus-A. baumannii complex may offer more clinical information. This study first identified that the overall percentage of resistance to all antibiotics tested was significantly different between the isolates from sterile sites and nonsterile sites (sterile sites, 22%; nonsterile sites, 46% [P < 0.05]). Furthermore, it was found that isolates resistant to all antibiotics tested were mostly of the unique A. baumannii genospecies, but more genospecies could be identified in isolates susceptible to all antibiotics tested.
In this study, very high percentages of resistance to imipenem, meropenem, amikacin, and ceftazidime (65%, 68%, 68%, and 75%, respectively) were identified in isolates from 2008. It is important to note that these are the last-line agents for clinical treatments. Previous studies conducted in intensive care units (ICUs) of several teaching hospitals in Taiwan had already indicated that the percentage of resistance to imipenem in A. baumannii was increasing, from 22% in 2000 to 25% in 2005 (16, 21). In other countries, such as Estonia, 5% of A. baumannii strains were resistant to meropenem and imipenem (27). Furthermore, in the MYSTIC report from 48 European hospitals for isolates from the period of 2002 to 2004 (38), 27% and 30% of isolates were resistant to meropenem and imipenem, respectively. Therefore, such high percentages of resistance in the current study, compared to the previous findings in Taiwan as well as in other countries (16, 21, 27, 38), may suggest that the prevalence of the isolates in the A. calcoaceticus-A. baumannii complex that are resistant to carbapenems has recently been increasing in Taiwan. In our hospital, the prevalence of isolates in the A. calcoaceticus-A. baumannii complex that were resistant to imipenem was below 25% before 2004 but abruptly increased to 57% in 2006 and approached 71% in 2011 (unpublished data). In addition, most of the isolates in the A. calcoaceticus-A. baumannii complex were resistant to fluoroquinolones (76%), aminoglycosides (75%), and cephalosporins (77%). The situation is very similar to what has been found in Europe, where 68%, 66%, and 52% of the isolates were resistant to ceftazidime, ciprofloxacin, and gentamicin (38).
A previous study (22) applied the oligonucleotide array system to characterize 52 blood isolates in the A. calcoaceticus-A. baumannii complex and found that antimicrobial susceptibility profiles were different among different genospecies of the A. calcoaceticus-A. baumannii complex. Nevertheless, limited information is currently available to further explore whether A. calcoaceticus-A. baumannii complex isolates from different collection sites can be associated with different antimicrobial resistance characteristics. In this study, it was found that the overall percentage of isolates from nonsterile sites that were resistant to all antibiotics was significantly higher than that of isolates from sterile sites (46% versus 22%; P < 0.05). However, the overall percentage of isolates from sterile sites that were susceptible to all antibiotics tested was significantly higher than that of isolates from nonsterile sites (35% versus 7%; P < 0.05). After ARDRA, the Acinetobacter isolates from sterile sites (CSF, bloodstream, and ascites fluid) showed genetic diversity. Furthermore, compared to all resistant strains, more genetic diversity was identified in all susceptible strains. The results may imply a close relationship of genospecies and antimicrobial resistance profiles. Several Acinetobacter ribotypes have been reported to be associated with pan resistance (35). Lim et al. (25) also reported that the majority of MBL-producing Acinetobacter isolates were Acinetobacter phenon 6/ct 13TU, genospecies, 3 and 13TU. The study also documented that 40 strains of A. baumannii showed resistance to most antimicrobial agents, except carbapenems, but isolates of other genospecies, such as Acinetobacter phenon 6/ct 13TU, genospecies 3, and 13TU, were usually susceptible to ciprofloxacin and ampicillin-sulbactam. Similarly, we also found that isolates resistant to all 12 antibiotics tested were mainly of the A. baumannii genospecies (97%), but only 31% of the isolates susceptible to all antimicrobials tested were A. baumannii genospecies.
In this study, 10 isolates from CSF were all susceptible to the 12 antimicrobials tested. This may raise the concern that the A. calcoaceticus-A. baumannii complex isolates that are susceptible to more antibiotics could be more invasive than the isolates with multidrug resistance. The phenomenon has never been discussed for isolates in the A. calcoaceticus-A. baumannii complex, but it has clinical importance. In human uropathogenic Escherichia coli (UPEC), chloramphenicol-, tetracycline-, and streptomycin-resistant isolates were associated with reduced virulence potential, and their virulence-related genes were found to be less frequent (37). Similarly, a study by Piatti et al. (34) also indicated that quinolone-resistant E. coli strains harbored fewer virulence genes than the susceptible strains. Future research should be conducted to further elucidate whether the same is the case for Acinetobacter bacteria.
Previous studies applied multilocus sequence typing (MLST) (43), randomly amplified polymorphic DNA (RAPD) (36), amplified fragment length polymorphism (AFLP) (19), and rpoB gene sequence (23) analysis for the identification of clinically important and emerging A. calcoaceticus-A. baumannii complex isolates. These methods are time-consuming and are not easy to apply in clinical settings. MLST analysis even needs to include analysis of sequences of several housekeeping genes (gltA, gyrB, gdhB, recA, cpn60, gpi, and rpoD). The ARDRA using 5 different restriction enzymes offers an efficient way to differentiate genospecies of the A. calcoaceticus-A. baumannii complex (3, 40). The results of our study not only support the application of ARDRA for genospecies determination of A. calcoaceticus-A. baumannii complex isolates but also suggest that most of the genospecies could be identified for clinical consideration using only three restriction enzymes. There were still two isolates for which the genospecies could not be determined in this study. According to molecular pattern comparison, one isolate could be genospecies 13BJ or 14TU A. haemolyticus/A. johnsonii, and the other one could be genospecies 5 (A. junii), 15BJ, 16BJ, or 17BJ; these genospecies currently have less clinical importance. In a clinical microbiology laboratory, one could always consider the use of more restriction enzymes in ARDRA for better identification.
Conclusion.
This study indicated the current emergence of carbapenem-, fluoroquinolone-, aminoglycoside-, and cephalosporin-resistant A. calcoaceticus-A. baumannii complex isolates in Taiwan. Moreover, ARDRA was shown to be a useful tool to study the genetic diversity of A. calcoaceticus-A. baumannii complex isolates for clinical consideration. Clinicians need to aware that not only does the so-called “A. baumannii” have clinical importance but other genospecies also may have different antimicrobial resistance and virulence potential characteristics. Beyond these major observations, after further analyzing our results by year, it was shown that A. baumannii genospecies isolates resistant to all 12 antibiotics tested were dominant in 2007 and 2008. However, in 2009, isolates were still mainly of the A. baumannii genospecies, but most of them were susceptible to all 12 antibiotics tested. The reason for such a change remains unknown and needs further investigation.
ACKNOWLEDGMENTS
We appreciate the collaborating work with the staff in the Department of Laboratory Medicine, China Medical University Hospital, Taichung, Taiwan.
This work was supported by grants DMR-98-098 and DMR-99-123 from China Medical University Hospital, Taichung, Taiwan.
We have no conflicts to declare.
Footnotes
Published ahead of print 24 September 2012
REFERENCES
- 1. Bergogne-Bérézin E, Towner KJ. 1996. Acinetobacter spp. as nosocomial pathogens: microbiological, clinical, and epidemiological features. Clin. Microbiol. Rev. 9:148–165 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2. Bouvet PJM, Grimont PAD. 1986. Taxonomy of the genus Acinetobacter with the recognition of Acinetobacter baumannii sp. nov., Acinetobacter haemolyticus sp. nov., Acinetobacter johnsonii sp. nov., and Acinetobacter junii sp. nov., and emended descriptions of Acinetobacter calcoaceticus and Acinetobacter lwoffii. Int. J. Syst. Bacteriol. 36:228–240 [Google Scholar]
- 3. Chang HC, et al. 2005. Species-level identification of isolates of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex by sequence analysis of the 16S-23S rRNA gene spacer region. J. Clin. Microbiol. 43:1632–1639 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4. Chang HL, et al. 2009. Nosocomial outbreak of infection with multidrug-resistant Acinetobacter baumannii in a medical center in Taiwan. Infect. Control. Hosp. Epidemiol. 30:34–38 [DOI] [PubMed] [Google Scholar]
- 5. Dijkshoorn L, et al. 1993. Correlation of typing methods for Acinetobacter isolates from hospital outbreaks. J. Clin. Microbiol. 31:702–705 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6. Dolzani L, Tonin E, Lagatolla C, Prandin L, Monti-Bragadin C. 1995. Identification of Acinetobacter isolates in the A. calcoaceticus-A. baumannii complex by restriction analysis of the 16S-23S rRNA intergenic-spacer sequences. J. Clin. Microbiol. 33:1108–1113 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7. Dueñas Díez AI, et al. 2004. Susceptibility of the Acinetobacter calcoaceticus-A. baumannii complex to imipenem, meropenem, sulbactam and colistin. Int. J. Antimicrob. Agents 23:487–493 [DOI] [PubMed] [Google Scholar]
- 8. Fournier PE, Richet H. 2006. The epidemiology and control of Acinetobacter baumannii in health care facilities. Clin. Infect. Dis. 42:692–699 [DOI] [PubMed] [Google Scholar]
- 9. GarcíA-Arata MI, Gerner-Smidt P, Ibrahim A. 1997. PCR-amplified 16S and 23S rDNA restriction analysis for the identification of Acinetobacter strains at the DNA group level. Res. Microbiol. 148:777–784 [DOI] [PubMed] [Google Scholar]
- 10. Gerner-Smidt P, Tjernberg I, Ursing J. 1991. Reliability of phenotypic tests for identification of Acinetobacter species. J. Clin. Microbiol. 29:277–282 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11. Harald S, Rainer B, Andrea S, Gerhard P. 1993. Antimicrobial susceptibility of Acinetobacter species. Antimicrob. Agent Chemother. 37:750–753 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12. Ho M, et al. 1999. 1998 Surveillance of antibiotic resistance in Taiwan. J. Microbiol. Immunol. Infect. 32:239–249 [PubMed] [Google Scholar]
- 13. Horrevorts A, et al. 1995. Clinical and epidemiological investigations of Acinetobacter genomospecies 3 in a neonatal intensive care unit. J. Clin. Microbiol. 33:1567–1572 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14. Houang ET, et al. 2003. Significance of genomic DNA group delineation in comparative studies of antimicrobial susceptibility of Acinetobacter spp. Antimicrob. Agents Chemother. 47:1472–1475 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15. Hsueh PR, Liu CY, Luh KT. 2002. Current status of antimicrobial resistance in Taiwan. Emerg. Infect. Dis. 8:132–137 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16. Hsueh PR, et al. 2001. Multicenter surveillance of antimicrobial resistance of major bacterial pathogens in intensive care units in 2000 in Taiwan. Microb. Drug Resist. 7:373–382 [DOI] [PubMed] [Google Scholar]
- 17. Huang YT, Hsueh PR. 2008. Antimicrobial drug resistance in Taiwan. Int. J. Antimicrob. Agents. 32:S174–S178 [DOI] [PubMed] [Google Scholar]
- 18. Idzenga D, Schouten MA, van Zanten AR. 2006. Outbreak of Acinetobacter genomic species 3 in a Dutch intensive care unit. J. Hosp. Infect. 63:485–487 [DOI] [PubMed] [Google Scholar]
- 19. Janssen P, et al. 1997. Discrimination of Acinetobacter genomic species by AFLP fingerprinting. Int. J. Syst. Bacteriol. 47:1179–1187 [DOI] [PubMed] [Google Scholar]
- 20. Jean SS, Hsueh PR. 2011. Antimicrobial drug resistance in Taiwan. J. Formos. Med. Assoc. 110:4–13 [DOI] [PubMed] [Google Scholar]
- 21. Jean SS, et al. 2009. Nationwide surveillance of antimicrobial resistance among non-fermentative gram negative bacteria in intensive care units in Taiwan: SMART programmed data 2005. Int. J. Antimicrob. Agents. 33:266–271 [DOI] [PubMed] [Google Scholar]
- 22. Ko WC, et al. 2008. 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. J. Clin. Microbiol. 46:2052–2059 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 23. Lee Y, et al. 2011. Carbapenem-non-susceptible Acinetobacter baumannii of sequence type 92 or its single-locus variants with a G428T substitution in zone 2 of the rpoB gene. J. Antimicrob. Chemother. 66:66–72 [DOI] [PubMed] [Google Scholar]
- 24. Lee YC, et al. 2011. Acinetobacter baumannii and Acinetobacter genospecies 13TU and 3 bacteraemia: comparison of clinical features, prognostic factors and outcomes. J. Antimicrob. Chemother. 66:1839–1846 [DOI] [PubMed] [Google Scholar]
- 25. Lim YM, Shin KS, Kim J. 2007. Distinct antimicrobial resistance patterns and antimicrobial resistance-harboring genes according to genomic species of Acinetobacter isolates. J. Clin. Microbiol. 45:902–905 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 26. Lin MF, et al. 2009. Clinical features and molecular epidemiology of multidrug-resistant Acinetobacter calcoaceticus-A baumannii complex in a regional teaching hospital in Taiwan. Am. J. Infect. Control. 37:e1–e3 [DOI] [PubMed] [Google Scholar]
- 27. Lõivukene K, Sepp E, Adamson V, Mitt Kallandi PU, Otter K, Naaber P. 2006. Prevalence and antibiotic susceptibility of Acinetobacter baumannii, Pseudomonas aeruginosa and Klebsiella pneumoniae in Estonian intensive care unit on comparison with European data. Scand. J. Infect. Dis. 38:1001–1008 [DOI] [PubMed] [Google Scholar]
- 28. Maragakis LL, Perl TM. 2008. Acinetobacter baumannii: epidemiology, antimicrobial resistance, and treatment options. Clin. Infect. Dis. 46:1254–1263 [DOI] [PubMed] [Google Scholar]
- 29. Montefour K, et al. 2008. Acinetobacter baumannii: an emerging multidrug-resistant pathogen in critical care. Crit. Care. Nurse 28:15–25 [PubMed] [Google Scholar]
- 30. Murray CK, Hospenthal DR. 2005. Treatment of multidrug resistant Acinetobacter. Curr. Opin. Infect. Dis. 18:502–506 [DOI] [PubMed] [Google Scholar]
- 31. Nemec A, Janda L, Melter O, Dijkshoorn L. 1999. Genotypic and phenotypic similarity of multiresistant Acinetobacter baumannii isolates in the Czech Republic. J. Med. Microbiol. 48:287–296 [DOI] [PubMed] [Google Scholar]
- 32. Paul CS, Maryan ID, Dannie GH. 2007. Acinetobacter, Alcaligenes, Chryseobacterium, Moraxella, and other nonfermentative gram-negative rods, p 770–802 In Murray PR, Baron EJ, Jorgensen JH, Landry ML, Pfaller MA. (ed), Manual of clinical microbiology, 9th ed ASM Press, Washington, DC [Google Scholar]
- 33. Peleg AY, Seifert H, Paterson DL. 2008. Acinetobacter baumannii: emergence of a successful pathogen. Clin. Microbiol. Rev. 21:538–582 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 34. Piatti G, Mannini A, Balistreri M, Schito AM. 2008. Virulence factors in urinary Escherichia coli strains: phylogenetic background and quinolone and fluoroquinolone resistance. J. Clin. Microbiol. 46:480–487 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 35. Quale J, Bratu S, Landman D, Heddurshetti R. 2003. Molecular epidemiology and mechanisms of carbapenem resistance in Acinetobacter baumannii endemic in New York City. Clin. Infect. Dis. 37:214–220 [DOI] [PubMed] [Google Scholar]
- 36. Spence RP, van der Reijden TJ, Dijkshoorn L, Towner KJ. 2004. Comparison of Acinetobacter baumannii isolates from United Kingdom hospitals with predominant Northern European genotypes by amplified-fragment length polymorphism analysis. J. Clin. Microbiol. 42:832–834 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 37. Starcic Erjavec M, Rijavec M, Krizan-Hergouth V, Fruth A, Zgur-Bertok D. 2007. Chloramphenicol- and tetracycline-resistant uropathogenic Escherichia coli (UPEC) exhibit reduced virulence potential. Int. J. Antimicrob. Agents 30:436–442 [DOI] [PubMed] [Google Scholar]
- 38. Unal S, Garcia-Rodriguez JA. 2005. Activity of meropenem and comparators against Pseudomonas aeruginosa and Acinetobacter spp. isolated in the MYSTIC Program, 2002-2004. Diagn. Microbiol. Infect. Dis. 53:265–271 [DOI] [PubMed] [Google Scholar]
- 39. van Dessel H, et al. 2002. Outbreak of a susceptible strain of Acinetobacter species 13 (sensu Tjernberg and Ursing) in an adult neurosurgical intensive care unit. J. Hosp. Infect. 51:89–95 [DOI] [PubMed] [Google Scholar]
- 40. Vaneechoutte M, et al. 1995. Identification of Acinetobacter genomic species by amplified ribosomal DNA restriction analysis. J. Clin. Microbiol. 33:11–15 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 41. Van Looveren M, Goossens H, ARPAC Steering Group 2004. Antimicrobial resistance of Acinetobacter spp. in Europe. Clin. Microbiol. Infect. 10:684–704 [DOI] [PubMed] [Google Scholar]
- 42. Villers D, et al. 1998. Nosocomial Acinetobacter baumannii infections: microbiological and clinical epidemiology. Ann. Intern. Med. 129:182–189 [DOI] [PubMed] [Google Scholar]
- 43. Wisplinghoff H, et al. 2008. Molecular epidemiology of clinical Acinetobacter baumannii and Acinetobacter genomic species 13TU isolates using a multilocus sequencing typing scheme. Clin. Microbiol. Infect. 214:708–715 [DOI] [PubMed] [Google Scholar]