Molecular characterization of VIM-producing Klebsiella pneumoniae from Scandinavia reveals genetic relatedness with international clonal complexes encoding transferable multidrug resistance

VIM-producing Klebsiella pneumoniae (VPKP) has been identified as a source of hospital outbreaks and is prevalent particularly in the Mediterranean region. In this study we have characterized eight VPKP isolates identified in Scandinavia during 2005–2008. With the exception of one isolate, all were from patients with recent history of hospitalization abroad (Greece, n = 6; Turkey, n = 1). Multilocus sequence typing (MLST) resulted in five sequence types (STs), ST36 (n = 1), ST147 (n = 4), ST272 (n = 1), ST273 (n = 1) and ST383 (n = 1), which except for ST272 were part of putative international clonal complexes. All were multidrug resistant due to the presence of other resistance determinants, including extended-spectrum β-lactamases (CTX-M-3, SHV-5 and SHV-12), 16S rRNA methylases (ArmA) and plasmid-mediated quinolone resistance determinants (QnrS). One isolate harboured a novel VIM-variant (VIM-26) while VIM-1 and VIM-19 were detected in six and one isolate, respectively. Two different genetic structures surrounding the bla_VIM gene were identified in four isolates. In two isolates bla_VIM-1 and bla_VIM-26 were located in an integron similar to In-e541 (intI1;bla_VIM-1/-26;aacA7; dhfrI;aadA1;3’CS) while in the other two isolates bla_VIM-1 was located in an integron lacking 3’CS but with an IS26 element in the 3’end (intI1;bla_VIM-1;aac(6’)-Ib;IS26), as identified in the IncN plasmid pKOX105. The bla_VIM-genes were located on transferable plasmids ranging from -40 to -240 kb and associated with Tn21 in four isolates. PCR-based replicon typing indicated association of bla_VIM with IncN (n = 3) and A/C (n = 1) broad-host-range plasmids but also with unknown replicons (n = 4). In conclusion, Scandinavian VPKP is associated with importation and genetically related to international clones encoding transferable plasmid-mediated multidrug resistance.     

Related carbapenemase-producing <Emphasis Type="Italic">Klebsiella</Emphasis> isolates detected in both a hospital and associated aquatic environment in Sweden

Carbapenem antibiotics are one of the last-resort agents against multidrug-resistant (MDR) bacteria. The occurrence of carbapenemase-producing Enterobacteriaceae (CPE) in wastewater and aquatic environments is an indication of MDR bacteria in the community. This study evaluated CPE in aquatic environments and compared them to the local hospital isolates in Sweden. Phenotypic and genotypic analyses of antibiotic resistance of environmental and clinical CPE were performed. The relatedness of the isolates and possible clonal dissemination was evaluated using phylogenetic and phyloproteomic analysis. Klebsiella oxytoca carrying carbapenemase genes (bla_VIM-1, bla_IMP-29) were isolated from wastewater and the recipient river, while K. oxytoca (bla_VIM-1) and Klebsiella pneumoniae (bla_VIM-1, bla_OXA-48, bla_NDM-1, bla_KPC-3) were isolated from patients at the local clinics or hospital. The K. oxytoca classified as sequence type 172 (ST172) isolated from the river was genotypically related to two clinical isolates recovered from patients. The similarity between environmental and clinical isolates suggests the dispersion of bla_VIM-1 producing K. oxytoca ST172 from hospital to aquatic environment and the likelihood of its presence in the community. This is the first report of CPE in aquatic environments in Sweden; therefore, surveillance of aquatic and hospital environments for CPE in other urban areas is important to determine the major transfer routes in order to formulate strategies to prevent the spread of MDR bacteria.     

Dissemination of IMP-6-producing Pseudomonas aeruginosa ST244 in multiple cities in China

Pseudomonas aeruginosa is an important opportunistic pathogen responsible for nosocomial infections and is currently reported to be a worldwide nosocomial menace. The aim of this study was to investigate the epidemiological traits and the distribution of metallo-β-lactamases (MBLs)-producing P. aeruginosa clinical isolates in ten cities in China between January 2010 and May 2012. Antimicrobial susceptibility was determined by disc diffusion assay and the minimum inhibitory concentrations (MICs) of imipenem and meropenem were also determined by the Etest according to Clinical and Laboratory Standards Institute (CLSI) guidelines. In addition, polymerase chain reaction (PCR) and DNA sequencing were applied to detect bla _MBL genes, and their epidemiological relationships were investigated by multilocus sequence typing (MLST). Of 368 P. aeruginosa isolates, MLST analysis identified 138 sequence types (STs), including 122 known and 16 novel STs, and the most frequently detected clone was ST244, followed by ST235. Besides, our study revealed that 25 isolates carried the bla _IMP-6 gene and three isolates carried the bla _VIM-2 gene, and a probe specific for both genes could be hybridised to an ~1,125-kb fragment in all isolates. Interestingly, all of the bla _IMP-6-producing isolates shared an identical ST, ST244, and exhibited a higher level of resistance to several antibiotics. Overall, these observations suggest that P. aeruginosa ST244 carrying the chromosomally located bla _IMP-6 gene is widely disseminated in multiple cites in China.     

Hidden VIM-1 metallo-beta-lactamase phenotypes among Acinetobacter baumannii clinical isolates

A total of 87 Acinetobacter baumannii nonrepetitive consecutive clinical isolates were tested for the presence of metallo-β-lactamases (MBLs). Results of phenotypic assays (MBL Etest, imipenem/imipenem-EDTA combined-disk test, and imipenem/EDTA double-disk synergy test) were negative in all cases, but molecular testing revealed the presence of two bla_VIM-1-carrying isolates. One isolate had bla_VIM-1 preceded by a weak P1 promoter, and both had inactivated P2 promoters and reduced bla_VIM-1 expression, partially justifying the results revealing hidden MBL phenotypes.     

Dissemination and genetic context analysis of blaVIM-6 among Pseudomonas aeruginosa isolates in Asian-Pacific Nations

VIM-6, previously reported in two strains from Singapore recovered in 2000, was detected in 16 isolates collected in 2006 in India (12 isolates), Indonesia (two), Korea and the Philippines (one each). High genetic variability was observed among VIM-6-producing isolates (12 ribotypes and 11 pulsed-field gel electrophoresis types), but clones were observed in India and Indonesia; bla_VIM-6-carrying integrons of 3.9 kb and 5 kb were detected, and two of five Indian hospitals yielded isolates with both integrons. These two integrons, bla_VIM-6 was located in the first position, followed by bla_OXA-10 and aacA4. The 5-kb integrons also harboured aadAl and a 331-bp open reading frame encoding a putative efflux pump.     

Characterisation of VIM-2-producing <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> isolates from lower tract respiratory infections in a Spanish hospital

To analyse the antimicrobial phenotype, carbapenem mechanisms, integrons, virulence factors and molecular typing of 164 Pseudomonas aeruginosa isolates recovered from lower tract respiratory samples in a Spanish hospital (1 year) as well as the patients’ clinical data. Susceptibility testing to 12 antipseudomonal agents was determined by microdilution and metallo-beta-lactamase (MBL) phenotype by double disc method. The oprD gene was studied by PCR, sequencing and comparison with P. aeruginosa PAO1 sequence. Detection and characterisation of MBLs, class 1, 2 and 3 integrons, and virulence genes were studied by PCR and sequencing. The prevalence of carbapenem-resistant P. aeruginosa (CRPA) was 26.8%. MBL phenotype was detected in 52.3% CRPA, and all of them were disseminated throughout the intensive care unit. Most of the MBL-carrying patients presented respiratory disease, mechanical ventilation, tracheostomy, bacteraemia, ≥?30 hospitalisation days and previous treatment with carbapenems and/or ≥?3 different antimicrobial families. The bla_VIM-2 gene was the unique MBL encoding gene and was detected inside class 1 integrons. The class 1 integrons detected in 39 strains (23.8%) were associated with aminoglycosides (aadB, aadA1, aadA6, aacA4, aac(3)-I) and carbapenems resistance genes (bla_VIM-2). The aac(3)-I?+?aadA1 and bla_VIM-2 arrangements were the most prevalent ones. Thirty-one different PFGE patterns and 4 STs (ST175, ST235, ST253, ST973) were detected among the 39 intI1-positive isolates, being ST235 the most frequent. CRPA showed a great variety of alterations in oprD gene. The exoU⁺/exoS^? genotype was detected in 82.6% of bla_VIM-2-producing strains (ST235) and the exoU^?/exoS⁺ in the remaining 17.4% (ST973).     

Diversity in VIM-2-encoding class 1 integrons and occasional blaSHV2a carriage in isolates of a persistent,multidrug-resistant Pseudomonas aeruginosa clone from Tunis

From 2002 to 2006, 35 of 73 multidrug-resistant Pseudomonas aeruginosa isolates from different wards at Charles Nicolle hospital of Tunis were positive for class B carbapenemase (using the imipenem-EDTA test), owing to a bla_VIM-2 gene cassette in a class 1 integron. Twenty-three isolates additionally produced the extended-spectrum β-lactamase SHV2a. DNA sequences immediately surrounding bla_SHV2a shared extensive identity with a Klebsiella pneumoniae plasmid sequence. Despite belonging to the same chromosomal type, as shown by pulsed-field gel electrophoresis (PFGE), the VIM-2 producing P. aeruginosa isolates prevalent at Charles Nicolle hospital displayed a diversity of VIM-2-carrying integrons.     

Tn5090-like class 1 integron carrying blaVIM-2 in a Pseudomonas putida strain from Portugal

Three Pseudomonas putida strains containing bla_VIM-2 were isolated from an inanimate surface of a female ward sanitary facility in the Hospital Infante D. Pedro, Aveiro. A novel class 1 integron was found in strain Pp2 (aacA41bla_VIM-2/aac6'-IIc disrupted by an insertion sequence IS1382), and strain PpI was found to carry a class 1 integron (aacA7/bla_VIM-2/aacCl/aacA4), which is described for the first time in this species. Strain PFI carries a class 1 integron associated with a Tn5090-like transposon, constituting the first finding of this type of arrangement in a strain from Portugal. This association highlights further dissemination of bla_VIM-2 in environmental hospital isolates.     

Similar Frequencies of Pseudomonas aeruginosa Isolates Producing KPC and VIM Carbapenemases in Diverse Genetic Clones at Tertiary-Care Hospitals in Medellín,Colombia

Carbapenem-resistant Pseudomonas aeruginosa has become a serious health threat worldwide due to the limited options available for its treatment. Understanding its epidemiology contributes to the control of antibiotic resistance. The aim of this study was to describe the clinical and molecular characteristics of infections caused by carbapenem-resistant P. aeruginosa isolates in five tertiary-care hospitals in Medellín, Colombia. A cross-sectional study was conducted in five tertiary-care hospitals from June 2012 to March 2014. All hospitalized patients infected by carbapenem-resistant P. aeruginosa were included. Clinical information was obtained from medical records. Molecular analyses included PCR for detection of bla_VIM, bla_IMP, bla_NDM, bla_OXA-48, and bla_KPC genes plus pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) for molecular typing. A total of 235 patients were enrolled: 91.1% of them were adults (n = 214), 88.1% (n = 207) had prior antibiotic use, and 14.9% (n = 35) had urinary tract infections. The bla_VIM-2 and bla_KPC-2 genes were detected in 13.6% (n = 32) and 11.5% (n = 27), respectively, of all isolates. Two isolates harbored both genes simultaneously. For KPC-producing isolates, PFGE revealed closely related strains within each hospital, and sequence types (STs) ST362 and ST235 and two new STs were found by MLST. With PFGE, VIM-producing isolates appeared highly diverse, and MLST revealed ST111 in four hospitals and five new STs. These results show that KPC-producing P. aeruginosa is currently disseminating rapidly and occurring at a frequency similar to that of VIM-producing P. aeruginosa isolates (approximately 1:1 ratio) in Medellín, Colombia. Diverse genetic backgrounds among resistant strains suggest an excessive antibiotic pressure resulting in the selection of resistant strains.     

Coproduction of KPC-18 and VIM-1 Carbapenemases by Enterobacter cloacae: Implications for Newer β-Lactam–β-Lactamase Inhibitor Combinations

Enterobacter cloacae strain G6809 with reduced susceptibility to carbapenems was identified from a patient in a long-term acute care hospital in Kentucky. G6809 belonged to sequence type (ST) 88 and carried two carbapenemase genes, bla_KPC-18 and bla_VIM-1. Whole-genome sequencing localized bla_KPC-18 to the chromosome and bla_VIM-1 to a 58-kb plasmid. The strain was highly resistant to ceftazidime-avibactam. Insidious coproduction of metallo-β-lactamase with KPC-type carbapenemase has implications for the use of next-generation β-lactam–β-lactamase inhibitor combinations.     

Multiple carbapenem hydrolyzing genes in clinical isolates of Acinetobacter baumannii

Purpose: Carbapenem resistance in Acinetobacter baumannii has become highly rampant, which has been ascribed to the presence of multiple carbapenemases. The objective of the present study was to prospectively investigate the presence of multiple carbapenemase encoding genes in clinical isolates of A. baumannii. Materials and Methods: A total of 30 imipenem resistant, consecutive non-repeat clinical isolates A. baumannii from a Tertiary Care Centre of Delhi were subjected to antimicrobial susceptibility testing (AST), screening for carbapenemase production by modified Hodge test (MHT) and determination of minimum inhibitory concentration for imipenem by E-Test®. These were subjected to Real time PCR for bla_{IMP-1 and 2}, bla_{VIM-1 and 2}, bla_{OXA23, 24, 51 and 58} using SYBR green-I. These were grouped together on the basis of their genotype as each isolate harboured multiple carbapenemases and correlated with their AST profile. Detection of the novel carbapenemase bla_NDM-1 was performed by real time PCR using TaqMan probes on 14 isolates. Results: Colistin appeared to be the most effective drug in vitro, followed by tetracycline and beta lactam/beta lactamase inhibitor combinations. All, but one isolate were positive for the MHT. All 30 isolates were positive for bla_OXA-51 like gene as well as bla_IMP-1 and bla_VIM-1 genes. bla_{OXA 24 and 58} were not detected in any of the isolates. bla_IMP-2, bla_VIM-2, bla_OXA-23 were present in 15, 6 and 14 isolates respectively. Grouping based on the genotypic profile did not correlate with susceptibility pattern. Nine among the 14 isolates also harboured the novel bla_NDM-1 gene. Conclusions: This is the first study from North India, which comprehensively detected the presence of multiple carbapenemases as well the bla_NDM-1 gene. The presence of the novel gene bla_NDM-1indicated ability of A. baumannii to acquire new carbapenemase genes despite the existence of multiple carbapenemase genes. The present study confirmed the presence of multiple genetic mechanisms for carbapenemases production among the clinical isolates of A. baumannii in north India.     

Characterization of carbapenem-resistant <Emphasis Type="Italic">Escherichia coli</Emphasis> and <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> from a healthcare region in Hong Kong

Carbapenem-resistant Enterobacteriaceae represents a major public health issue. This study investigated the clonality and resistance mechanisms of 92 carbapenem-resistant E. coli (n?=?21) and K. pneumoniae (n?=?71) isolates collected consecutively from clinical specimens and patients at high risk of carriage between 2010 and 2012 in a healthcare region in Hong Kong. Combined disk tests (CDTs) and the Carba NP test were used for phenotypic detection of carbapenemases. PCR assays were used to detect carbapenemase genes. All isolates were intermediate or resistant to at least one carbapenem. Nine (9.8 %) isolates were genotypic carbapenemase producers and included six K. pneumoniae (one ST1306/bla _IMP-4, one ST889/bla _IMP-4, two ST11/bla _KPC-2, one ST258/bla _KPC-2, one ST483/bla _NDM-1) and three E. coli (one ST131/bla _IMP-4, two ST744/ bla _NDM-1) isolates. All nine isolates carrying carbapenemase genes could be detected by the CDTs and the Carba NP test. PCR identified bla _CTX-M and bla _AmpC alone or in combination in 77.8 % (7/9) and 96.4 % (80/83) of the carbapenemase-producers and non-producers, respectively. Porin loss was detected in 22.2 % (2/9) and 59.0 % (49/83) of the carbapenemase-producers and non-producers, respectively. Overall, the E. coli clones were diverse (14 different STs), but 36.6 % (26/71) of the K. pneumoniae isolates belonged to ST11. In conclusion, the prevalence of carbapenemases among carbapenem-nonsusceptible E. coli and K. pneumoniae remained low in Hong Kong. Porin loss combined with AmpC and/or CTX-M type ESBL was the major mechanism of carbapenem resistance in the study population.     

Nosocomial dissemination of VIM-2-producing ST235 <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> in Lithuania

Pseudomonas aeruginosa multidrug resistance, and particularly the production of carbapenemases linked to international high-risk clones, is of growing concern. While high levels of carbapenem resistance (>60 %) have been reported in Lithuania, so far, there is no information on the underlying mechanisms. Thus, the aim of this work was to determine the molecular epidemiology and prevalence of acquired carbapenemases among 73 carbapenem-resistant P. aeruginosa isolates recovered in a hospital from Kaunas, Lithuania in 2011–2012. The presence of acquired carbapenemases was evaluated through phenotypic (modified Hodge test, cloxacillin inhibition test, double-disc synergy test) and genetic methods [polymerase chain reaction (PCR) and sequencing]. Clonal relatedness was assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Acquired β-lactamases were detected in 19 (26 %) of the isolates, whereas resistance was exclusively chromosomal (OprD inactivation?±?AmpC hyperproduction) in the remaining 54 (74 %) isolates. The acquired β-lactamases detected included 16 VIM-2, one PER-1 and two GES enzymes. PFGE revealed that 15 of the 16 VIM-2 isolates belonged to a single clone, identified as the international high-risk clone ST235 by MLST. bla _VIM-2 was preceded by aacA7 in a class I integron, similar to epidemic ST235 isolates described in nearby countries. Additionally, sequencing of bla _GES revealed the presence of the carbapenem-hydrolysing enzyme GES-5 in one of the isolates and a novel GES variant, designated GES-27, in the other. GES-27 differed from GES-5 by a single amino acid substitution, proline 167, that was replaced by glutamine. Increasing emergence and dissemination of concerning resistance mechanisms and international clones warrants global surveillance and control strategies.     

Sporadic occurrence of CMY-2-producing multidrug-resistant Escherichia coli of ST-complexes 38 and 448, and ST131 in Norway

Clinical isolates of Escherichia coli with reduced susceptibility to oxyimino-cephalosporins and not susceptible to clavulanic acid synergy (n = 402), collected from Norwegian diagnostic laboratories in 2003–2007, were examined for the presence of plasmid-mediated AmpC β-lactamases (PABLs). Antimicrobial susceptibility testing was performed for β-lactam and non-β-lactam antibiotics using Etest and Vitek2, respectively. The AmpC phenotype was confirmed using the boronic acid test. PABL-producing isolates were detected using ampC multiplex-PCR and examined by bla_AmpC sequencing, characterization of the bla_AmpC genetic environment, phylogenetic grouping, Xbal- pulsed-field gel electrophoresis (PFGE), multi-locus sequence typed (MLST), plasmid profiling and PCR-based replicon typing. For the PABL-positive isolates (n = 38), carrying bla_CMY-2 (n = 35), bla_CMY-7 (n = 1) and bla_DHA-1 (n = 2), from out- (n = 23) and in-patients (n = 15), moderate-high MICs of β-lactams, except cefepime and carbapenems, were determined. All isolates were resistant to trimethoprim-sulphamethoxazole. Multidrug resistance was detected in 58% of the isolates. The genes bla_CMY-2 and bla_CMY-7 were linked to ISEcp1 upstream in 32 cases and in one case, respectively, and bla_DHA-1 was linked to qacEΔ1-sull upstream and downstream in one case. Twenty isolates were of phylogenetic groups B2 or D. Thirty-three XbaI-PFGE types, including three clusters, were observed. Twenty-five sequence types (ST) were identified, of which ST complexes (STC) 38 (n = 7), STC 448 (n = 5) and ST131 (n = 4) were dominant. Plasmid profiling revealed 1–4 plasmids (50–250 kb) per isolate and 11 different replicons in 37/38 isolates; bla_CMY-2 was carried on transferable multiple-replicon plasmids, predominantly of Inc groups I1 (n = 12), FII (n = 10) and A/C (n = 7). Chromosomal integration was observed for bla_CMY-2 in ten strains. CMY-2 is the dominant PABL type in Norway and is associated with ISEcp1 and transferable, multiple-replicon IncI1, IncA/C, or IncFII plasmids in nationwide strains of STC 448, STC 38 and ST131.     

The emergence and dissemination of CTX-M-producing Escherichia coli sequence type 131 causing community-onset bacteremia in Israel

Community-onset bloodstream infections caused by extended-spectrum β-lactamase-producing Escherichia coli (ESBL-EC-COBSIs) were investigated over a 7-year-period (2003–2009) in our institution. ESBL-EC-COBSI inclusion criteria were cefotaxime/ceftazidime non-susceptible blood isolates recovered during 48 h upon hospital admission. Forty-one isolates were molecularly characterized. Susceptibilities were determined (Vitek-2) and genotyping was performed [multilocus sequence typing (MLST)]. CTX-M genes were determined [polymerase chain reaction (PCR) and sequencing] and bla _CTX-M-encoding plasmids (n?=?10) were analyzed and compared. Phylogrouping and virulence genes were identified (PCR). The incidence rate of ESBL-EC-COBSIs has increased from 2.94 to 7.87 cases/10,000 admissions. All isolates were multidrug-resistant (MDR), displaying co-resistance to ciprofloxacin (93 %), trimethoprim–sulfamethoxazole (85 %), and gentamicin (51 %). MLST identified ten sequence types (STs), of which five were novel. ST131 accounted for 66 % of the cases (27/41), and dominated over the years (prevalence of 25 % in 2003 and 85 % in 2009). All isolates carried CTX-M genes with the following prevalence: bla _CTX-M-2 (6/8; 75 %) in 2003; bla _CTX-M-15 (9/13, 69 % in 2007); and bla _CTX-M-15 (11/20, 55 %) and bla _CTX-M-14 (7/20, 35 %) in 2009. bla _CTX-M-15- and bla _CTX-M-14-encoding plasmids harbored by ST131 differed. Of all isolates, 98 % belonged to virulent phylogroups B2 (28/41, 68 %) and D (12/41, 29 %), though ST131 isolates carried a higher number of virulence genes compared to other lineages (p?<?0.05). The incidence of ESBL-EC-COBSIs increased 2.7-fold during the period 2003–2009. This increase appears to be related to the emergence and clonal expansion of bla _CTX-M-15- or bla _CTX-M-14-carrying ST131. The superiority of this virulent lineage should be further explored.     

Successive emergence of extended-spectrum beta-lactamase-producing and carbapenemase-producing Enterobacter aerogenes isolates in a university hospital

Sixty-two clinical isolates of Enterobacter aerogenes resistant to expanded-spectrum cephalosporins were collected between July 2003 and May 2005. Among these isolates, 23 (37.1%) were imipenem (IPM) susceptible, and 39 (62.9%) were IPM insusceptible, of which 89.7% (35/39) were resistant and 10.3% (4/39) were intermediate. Isolate genotypes were compared by pulsed-field gel electrophoresis. Of 62 isolates, 48 belonged to epidemic pulsotype A (77.4%). This pulsotype included 37.5% and 58.4% of β-lactam phenotypes b and a, respectively. Nine isolates (14.5%) belonged to pulsotype E, which included 22.3% and 77.7% of phenotypes b and a, respectively. The β-lactamases with pIs of 5.4, 6.5, 8.2, and 8.2 corresponded to extended-spectrum β-lactamases (ESBLs) TEM-20, TEM-24, SHV-5, and SHV-12, respectively. Of 39 IPM-insusceptible E. aerogenes isolates, 26 (66.6%) were determined to be metallo-β-lactamase producers, by using a phenotypic method. Of these isolates, 24 harbored a bla_IMP-1 gene encoding a protein with a pI of >9.5, and two carried the bla_VIM-2 gene encoding a protein with a pI of 5.3, corresponding to β-lactamases IMP-1 and VIM-2, respectively. The remaining 13 (33.4%) isolates were negative for the bla_IMP-1 and bla_VIM-2 genes but showed an alteration of their outer membrane proteins (OMPs). Ten of these isolates produced the two possible OMPs (32 and 42 kDa), with IPM MICs between 8 and 32 μg/ml, and three others produced only a 32-kDa OMP with IPM MICs >32 μg/ml. This work demonstrates that, in addition to resistance to expanded-spectrum cephalosporins, IPM resistance can occur in ESBL-producing E. aerogenes isolates by carbapenemase production or by the loss of porin in the outer membrane.     

Emergence of Pseudomonas aeruginosa strains producing metallo-β-lactamases of the IMP-15 and VIM-2 types in Mexico

Eighty-six carbapenem non-susceptible Pseudomonas aeruginosa isolates collected in the National Institute of Respiratory Diseases of Mexico City were screened for the presence of metallo-β-lactamase (MBL) activity using both E-test strips and a microbiological assay with EDTA-imipenem. Genomic comparisons and sequence analyses conducted with these isolates revealed the presence of bla_VIM-2 in two clonally related isolates, and bla_IMP-15 in a clonally unrelated isolate. Both genes were found to be carried by class 1 integrons, and bla_IMP-15 was additionally present on a broad host-range plasmid. This is the first report of co-existing P. aeruginosa strains producing different MBLs in a Mexican hospital, highlighting the necessity of appropriate surveillance to prevent dissemination of carbapenem resistance.     

Characterization of Extended-Spectrum-Beta-Lactamase-Producing Escherichia coli Strains Involved in Maternal-Fetal Colonization: Prevalence of E. coli ST131

Maternal-fetal Escherichia coli infections, such as neonatal bacteremia and meningitis, are important causes of morbidity and mortality. From 2006 to 2010, we studied newborns and their mothers who were colonized with E. coli in a French hospital in order to document (i) the epidemiology and genetic characteristics of extended-spectrum-beta-lactamase (ESBL)-producing E. coli strains, (ii) the prevalence of associated virulence genes, (iii) the prevalence of clone sequence type 131 (ST131), and (iv) the genetic relationship among ESBL-producing strains. Among the 2,755 E. coli cultures recovered from vaginal or neonatal samples, 68 were ESBL producers (2.46%). We found a wide diversity of ESBL genes, with the majority being bla_CTX-M-14, bla_CTX-M-1, and bla_CTX-M-15, distributed among the 4 main phylogenetic groups. Genes encoding virulence factors were found in 90.7% of the isolates, with ≥2 virulence genes present in 76% of cases. The prevalence of ST131 among ESBL-producing E. coli isolates was 9.4% (6/64). Five of these 6 ST131 isolates possessed bla_CTX-M-15 enzymes (and also were resistant to quinolones), and one possessed bla_CTX-M-2 enzymes. Two possessed virulence genes, suggesting the presence of pathogenicity island II_J96 (PAI II_J96)-like domains. Pulsed-field gel electrophoresis (PFGE) revealed a high level of genomic diversity overall, except for 3 closely related isolates belonging to clonal group ST131. Repetitive PCR showed that the six ST131 isolates were closely related to ST131 control strains (>95% similarity). This study shows a high prevalence of ESBL-producing E. coli strains and clonal group ST131 in the French maternal-fetal population. These results suggest a widespread distribution of ESBL enzymes in the community and highlight the early transmission between mothers and neonates. These findings are worrisome, especially for this particularly vulnerable population.     

Carbapenem-resistant Enterobacter cloacae and the emergence of metallo-β-lactamase-producing strains in a third-level hospital (Santiago de Compostela, NW Spain)

The purpose of this paper was to investigate the occurrence of carbapenem-resistant Enterobacter cloacae in our institution, to detect the carbapenemase-associated resistance and to determine the genetic relatedness of the isolates. Species identification and antimicrobial susceptibility testing were performed using the Vitek 2 system and Etest. Multiplex polymerase chain reaction–enzyme linked immunosorbent assay (PCR-ELISA) was used for the detection of extended-spectrum β-lactamase (ESBL)-producers. The bla _IMP and bla _VIM genes were amplified by PCR and sequenced. The DiversiLab System was used for strain-typing. During the period 2006–2008, 12 different isolates of carbapenem-resistant E. cloacae (2.3 %) were recovered in our laboratory. Only two positive isolates for the bla _VIM gene were detected. The minimum inhibitory concentration (MIC) values were higher for all carbapenems in the group of non-metallo-β-lactamase (MBL)-producers. All isolates showed MIC values ≤2 against this tigecycline. The two bla _VIM-1-carrying isolates showed different genotypes. For non-MBL-producers, two clonally related clusters were observed. Different mechanisms can be associated with carbapenem-resistance in E. cloacae. MBL-producing strains are less prevalent than those with other mechanisms of resistance. The clonal relationship confirms the risk of spread of these organisms with the transfer of patients to different wards and the persistence of these clones over time or the ‘de novo’ acquisition of the resistance caused by the selective pressure exerted by antibiotics treatments.     

Molecular epidemiology of carbapenem-resistant <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> in an endemic area: comparison with global data

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) is an endemic problem in certain countries including Greece. CRPA and multidrug-resistant P. aeruginosa (MDRPA) firstly emerged in our region during the 80s, right after the launch of imipenem and meropenem as therapeutic agents against P. aeruginosa infections. The role of outer membrane protein (Opr) inactivation has been known to contribute to imipenem resistance since many years, while efflux overexpression systems have been mainly associated with meropenem resistance. Among carbapenemases, metallo-β-lactamases (MBL) and mostly Verona integron-mediated (VIM) MBL’s have played the most crucial role in CRPA emergence. VIM-2 and VIM-4 producing CRPA, usually belonging to clonal complexes (CC) 111 and 235 respectively, have most frequently been isolated. Bla_VIM-2 and bla_VIM-4 are usually associated with a class 1 integron. VIM-17 also has appeared in Greece. On the other hand, other VIM subtypes detected in a global level, such as VIM-3, VIM-5, VIM-6, VIM-7, VIM-11, VIM-14, VIM-15, VIM-16 and VIM-18 have not yet emerged in Greece. However, new VIM subtypes will probably emerge in the future. In addition, MBL carbapenemases other than VIM, detected worldwide have not yet appeared. A single CRPA isolate producing KPC has emerged in our region several years ago. The study of the molecular basis of Opr deficiency and efflux overexpression remains a challenge for the future. In this article, we review the molecular epidemiology of CRPA in an endemic area, compared to global data.