Identification of VIM-2-Producing Pseudomonas aeruginosa from Tanzania Is Associated with Sequence Types 244 and 640 and the Location of blaVIM-2 in a TniC Integron

Epidemiological data on carbapenemase-producing Gram-negative bacteria on the African continent are limited. Here, we report the identification of VIM-2-producing Pseudomonas aeruginosa isolates in Tanzania. Eight out of 90 clinical isolates of P. aeruginosa from a tertiary care hospital in Dar es Salaam were shown to harbor bla_VIM-2. The bla_VIM-2-positive isolates belonged to two different sequence types (ST), ST244 and ST640, with bla_VIM-2 located in an unusual integron structure lacking the 3′ conserved region of qacΔE1-sul1.     

Carbapenemase -producing Pseudomonas aeruginosa isolates from Turkey: first report of P. aeruginosa high-risk clones with VIM-5– and IMP-7–type carbapenemases in a tertiary hospital

We investigated the presence of carbapenemases in carbapenem-resistant Pseudomonas aeruginosa isolates, which were collected over a 14-month period in a Turkish hospital, with in-depth molecular characterization of carbapenemase-producing isolates. Among 45 study isolates, 2 isolates were identified as carbapenemase producers by both Carba NP and Carbapenem Inactivation Method tests, and only 1 of them gave a positive result in polymerase chain reaction tests for a carbapenemase gene (bla_VIM). Whole genome sequencing of the 2 isolates revealed the presence of bla_VIM-5 gene in an ST308 isolate, while the other one expressed IMP-7 in an ST357 isolate; both STs are considered high-risk clones. The 2 carbapenemase-producing isolates were multidrug resistant, as they harbored other resistance determinants, including a variant of the recently described plasmid-encoded fluoroquinolone resistance determinant crpP gene, crpP-2. We report for the first time P. aeruginosa high-risk clones carrying VIM-5– and IMP-7–type carbapenemases with multiple resistance determinants in Turkey.     

Evaluation of Clonality and Carbapenem Resistance Mechanisms among Acinetobacter baumannii-Acinetobacter calcoaceticus Complex and Enterobacteriaceae Isolates Collected in European and Mediterranean Countries and Detection of Two Novel β-Lactamases,GES-22 and VIM-35

We evaluated doripenem-resistant Acinetobacter baumannii-Acinetobacter calcoaceticus complex (ACB; n = 411) and Enterobacteriaceae (n = 92) isolates collected from patients from 14 European and Mediterranean countries during 2009 to 2011 for the presence of carbapenemase-encoding genes and clonality. Following susceptibility testing, carbapenem-resistant (doripenem MIC, >2 μg/ml) isolates were screened for carbapenemases. New β-lactamase genes were expressed in a common background and susceptibility was tested. Class 1 integrons were sequenced. Clonality was evaluated by pulsed-field gel electrophoresis and multilocus sequence typing (Pasteur scheme). Relative expression of β-lactam intrinsic resistance mechanisms was determined for carbapenemase-negative Enterobacteriaceae. ACB and Enterobacteriaceae displayed 58.9 and 0.9% doripenem resistance, respectively. bla_OXA-23, bla_OXA-58, and bla_{OXA-24/OXA-40} were detected among 277, 77, and 29 ACB, respectively (in 8, 6, and 5 countries). Ten Turkish isolates carried bla_GES-11 or bla_GES-22. GES-22 (G243A and M169L mutations in GES-1) had an extended-spectrum β-lactamase profile. A total of 33 clusters of ≥2 ACB isolates were observed, and 227 isolates belonged to sequence type 2/international clone II. Other international clones were limited to Turkey and Israel. Doripenem-resistant Enterobacteriaceae increased significantly (0.7 to 1.6%), and 15 bla_KPC-2- and 22 bla_KPC-3-carrying isolates, mostly belonging to clonal complexes 11 and 258, were observed. Enterobacteriaceae isolates producing OXA-48 (n = 16; in Turkey and Italy), VIM-1 (n = 10; in Greece, Poland, and Spain), VIM-26 (n = 1; in Greece), and IMP-19, VIM-4, and the novel VIM-35 (n = 1 each from Poland) were detected. VIM-35 had one substitution compared to VIM-1 (A235T) and a similar susceptibility profile. One or more resistance mechanisms were identified in 4/6 carbapenemase-negative Enterobacteriaceae. This broad evaluation confirms results from country-specific surveys and shows a highly diverse population of carbapenemase-producing ACB and Enterobacteriaceae in Europe and Mediterranean countries.     

Prospective Multicenter Study of Carbapenemase-Producing Enterobacteriaceae from 83 Hospitals in Spain Reveals High In Vitro Susceptibility to Colistin and Meropenem

The aim of this study was to determine the impact of carbapenemase-producing Enterobacteriaceae (CPE) in Spain in 2013 by describing the prevalence, dissemination, and geographic distribution of CPE clones, and their population structure and antibiotic susceptibility. From February 2013 to May 2013, 83 hospitals (about 40,000 hospital beds) prospectively collected nonduplicate Enterobacteriaceae using the screening cutoff recommended by EUCAST. Carbapenemase characterization was performed by phenotypic methods and confirmed by PCR and sequencing. Multilocus sequencing types (MLST) were determined for Klebsiella pneumoniae and Escherichia coli. A total of 702 Enterobacteriaceae isolates met the inclusion criteria; 379 (54%) were CPE. OXA-48 (71.5%) and VIM-1 (25.3%) were the most frequent carbapenemases, and K. pneumoniae (74.4%), Enterobacter cloacae (10.3%), and E. coli (8.4%) were the species most affected. Susceptibility to colistin, amikacin, and meropenem was 95.5%, 81.3%, and 74.7%, respectively. The most prevalent sequence types (STs) were ST11 and ST405 for K. pneumoniae and ST131 for E. coli. Forty-five (54.1%) of the hospitals had at least one CPE case. For K. pneumoniae, ST11/OXA-48, ST15/OXA-48, ST405/OXA-48, and ST11/VIM-1 were detected in two or more Spanish provinces. ST11 isolates carried four carbapenemases (VIM-1, OXA-48, KPC-2, and OXA-245), but ST405 isolates carried OXA-48 only. A wide interregional spread of CPE in Spain was observed, mainly due to a few successful clones of OXA-48-producing K. pneumoniae (e.g., ST11 and ST405). The dissemination of OXA-48-producing E. coli is a new finding of public health concern. According to the susceptibilities determined in vitro, most of the CPE (94.5%) had three or more options for antibiotic treatment.     

Dispersal of Carbapenemase blaVIM-1 Gene Associated with Different Tn402 Variants,Mercury Transposons,and Conjugative Plasmids in Enterobacteriaceae and Pseudomonas aeruginosa

The emergence of bla_VIM-1 within four different genetic platforms from distinct Enterobacteriaceae and Pseudomonas aeruginosa isolates in an area with a low prevalence of metallo-β-lactamase producers is reported. Forty-three VIM-1-producing isolates (including 19 Enterobacter cloacae, 2 Escherichia coli, and 2 P. aeruginosa isolates, 18 Klebsiella pneumoniae isolate, and 2 Klebsiella oxytoca isolate) recovered from 2005 to 2007 and corresponding to 15 pulsed-field gel electrophoresis types were studied. The Enterobacteriaceae isolates corresponded to a hospital outbreak, and the P. aeruginosa isolates were sporadically recovered. The genetic context of the integrons carrying bla_VIM-1 (arbitrarily designated types A, B, C, and D) was characterized by PCR mapping based on known Tn402 and mercury transposons and further sequencing. Among Enterobacteriaceae isolates, bla_VIM-1 was part of integrons located either in an In2-Tn402 element linked to Tn21 (type A; In110-bla_VIM-1-aacA4-aadA1) or in a Tn402 transposon lacking the whole tni module [type B; In113-bla_VIM-1-aacA4-dhfrII (also called dfrB1)-aadA1-catB2] and the transposon was associated with an IncHI2 or IncI1 plasmid, respectively. Among P. aeruginosa isolates, bla_VIM-1 was part of a new gene cassette array located in a defective Tn402 transposon carrying either tniBΔ3 and tniA (type C; bla_VIM-1-aadA1) or tniC and ΔtniQ (type D; bla_VIM-1-aadB), and both Tn402 variants were associated with conjugative plasmids of 30 kb. The dissemination of bla_VIM-1 was associated with different genetic structures and bacterial hosts, depicting a complex emergence and evolutionary network scenario in our facility, Ramón y Cajal University Hospital, Madrid, Spain. Knowledge of the complex epidemiology of bla_VIM-1 is necessary to control this emerging threat.The emergence of acquired metallo-β-lactamases (MBLs) is of clinical concern since they confer resistance to all available β-lactams except aztreonam and they are not inhibited by class A β-lactamase inhibitors (42). MBLs have been categorized into different groups; IMP and VIM are the most commonly identified types, and they are predominant in Asia and Europe, respectively. VIM enzymes have been grouped into three main clusters designated VIM-1, VIM-2, and VIM-7 (42). Although they are more prevalent among nonfermenting gram-negative bacteria, they are increasingly recognized among members of the family Enterobacteriaceae (42). To date, VIM-2 is more widely spread among Pseudomonas aeruginosa isolates, whereas VIM-1 is normally confined to Enterobacteriaceae isolates (41, 42). MBL-producing isolates have been identified rarely in Spain, in contrast to other European countries such as Greece and Italy (28, 34, 41). Nevertheless, the prevalence of MBL producers in Spain may have been underestimated since Enterobacteriaceae and P. aeruginosa isolates with VIM-1 and VIM-2 enzymes have been identified in different Spanish cities (11, 25, 26, 32, 37).The bla_VIM allelic variants are part of class 1 integrons which are derivatives of Tn402 (also called Tn5090), a transposon characterized by the presence of a transposition module comprising a suite of four genes (tniR/tniC, tniQ, tniB, and tniA) (12, 21). Although diverse gene cassette arrays have been identified among class 1 integrons containing bla_VIM alleles, a detailed characterization of the whole Tn402-harboring structure has hardly been achieved. Recently, MBL genes have been identified as part of sul1 integrons linked to a Tn402 element lacking tniR/tniC and tniQ, as reported for VIM-1- and VIM-2-producing P. aeruginosa isolates from Italy, Poland, and Australia (5, 29, 34, 35). Integrons lacking the 3′ conserved sequence (3′-CS) and associated with either a complete Tn402 transposon, as described previously for an IMP-4-producing Citrobacter youngae isolate (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"AF288045","term_id":"827747829","term_text":"AF288045"}}AF288045), or a Tn402 element containing tniC, like the integrons in P. aeruginosa VIM-2 producers from Italy, Norway, Russia, Ghana, India, Taiwan, and the United States, have also been reported previously (13, 16, 31, 36, 44). Some of these Tn402 elements were identified within different Tn3-like transposons (5, 35, 40).The objective of this work was to characterize the genetic platforms linked to the bla_VIM-1 gene from clinical isolates of different bacterial species recovered at Ramón y Cajal University Hospital over a period of 3 years (2005 to 2007). These isolates included the VIM-1-producing Enterobacteriaceae isolates involved in a recent outbreak (32) and two sporadic VIM-1-producing P. aeruginosa isolates contemporarily identified in the same hospital. In addition, these genetic platforms were compared with those already described.     

Carbapenemase-Producing Enterobacteriaceae in Spain in 2012

We report the epidemiological impact of carbapenemase-producing Enterobacteriaceae (CPE) in Spain in 2012. Of the 237 carbapenemases detected, 163 were from the OXA-48 group, 60 were from VIM-1, 8 were from KPC-2, 5 were from IMP, and 1 was from NDM-1. Interhospital spread of carbapenemase-producing Klebsiella pneumoniae was due to a limited number of multilocus sequence types (MLST) and carbapenemase types, including ST15–VIM-1, ST11–OXA-48, ST405–OXA-48, ST101–KPC-2, and ST11–VIM-1. The number of CPE cases in Spain has increased sharply in recent years, due mainly to the emergence of OXA-48.     

In Vitro Susceptibility of Characterized β-Lactamase-Producing Strains Tested with Avibactam Combinations

Avibactam, a broad-spectrum β-lactamase inhibitor, was tested with ceftazidime, ceftaroline, or aztreonam against 57 well-characterized Gram-negative strains producing β-lactamases from all molecular classes. Most strains were nonsusceptible to the β-lactams alone. Against AmpC-, extended-spectrum β-lactamase (ESBL)-, and KPC-producing Enterobacteriaceae or Pseudomonas aeruginosa, avibactam lowered ceftazidime, ceftaroline, or aztreonam MICs up to 2,048-fold, to ≤4 μg/ml. Aztreonam-avibactam MICs against a VIM-1 metallo-β-lactamase-producing Enterobacter cloacae and a VIM-1/KPC-3-producing Escherichia coli isolate were 0.12 and 8 μg/ml, respectively.     

Survey of Carbapenemase-Producing Enterobacteriaceae in Companion Dogs in Madrid,Spain

We found a low prevalence (0.6%) of carbapenemase-producing Enterobacteriaceae (CPE) in fecal microbiota of companion dogs. A single VIM-1-producing Klebsiella pneumoniae isolate belonging to sequence type (ST) 2090 was detected. bla_VIM-1 was carried on a class 1 integron and an untypeable ∼48-kb plasmid. Emergence and spread of CPE in this group of animals may be a threat to public health in human and veterinary medicine. This finding supports the need for active surveillance studies in companion animals that live close to humans, as interspecies transmission may occur within the same household.     

Differences between meropenem and imipenem disk to detect carbapenemase in gram-negative bacilli using simplified carbapenem inactivation method

The spread of carbapenemase-producing Enterobacteriaceae (CPE) is a major threat to public health. In the present study, we compared the difference between meropenem and imipenem disk for detecting carbapenemase-producing gram-negative bacilli using simplified carbapenem inactivation method (sCIM). 106 Enterobacteriaceae, including 74 CPE, 17 Pseudomonas aeruginosa including 10 carbapenemase-producing isolates and 36 Acinetobacter baumannii including 20 carbapenem-resistant isolates preserved in our laboratory were tested. Based on sCIM method, the test bacteria were tested with both meropenem and imipenem disk, respectively. In Enterobacteriaceae, the usage of both meropenem and imipenem disk showed high concordance (99.1%). Meropenem disk cannot identify positive isolates among the 10 P. aeruginosa and 20 A. baumannii isolates due to low carbapenem hydrolytic ability of the carbapenemase produced by these strains. Thus, meropenem disk was found to be similar to imipenem disk, presenting high specificity and sensitivity in the detection of carbapenemase in Enterobacteriaceae, but it cannot be used for the detection of carbapenemase in P. aeruginosa and A. baumannii.     

Metallo-β-lactamase–producing Pseudomonas aeruginosa isolates in Tunisia

This study was conducted to identify metallo-β-lactamase (MBL) producers among a collection of 75 nonrepetitive carbapenem-resistant Pseudomonas aeruginosa isolates recovered between November 2003 and May 2007 at the University Hospital Sahloul, Sousse, Tunisia. Five isolates produced the MBL VIM-2. Those bla_VIM-2-positive isolates were clonally related according to pulsed-field gel electrophoresis analysis. This carbapenemase was very likely chromosomally located and as a form of a gene cassette in a class 1 integron. This is the 1st report of spread of VIM-2 producers in Tunisia.     

Emergence of Imipenem-Resistant Gram-Negative Bacilli in Intestinal Flora of Intensive Care Patients

Intestinal flora contains a reservoir of Gram-negative bacilli (GNB) resistant to cephalosporins, which are potentially pathogenic for intensive care unit (ICU) patients; this has led to increasing use of carbapenems. The emergence of carbapenem resistance is a major concern for ICUs. Therefore, in this study, we aimed to assess the intestinal carriage of imipenem-resistant GNB (IR-GNB) in intensive care patients. For 6 months, 523 consecutive ICU patients were screened for rectal IR-GNB colonization upon admission and weekly thereafter. The phenotypes and genotypes of all isolates were determined, and a case control study was performed to identify risk factors for colonization. The IR-GNB colonization rate increased regularly from 5.6% after 1 week to 58.6% after 6 weeks in the ICU. In all, 56 IR-GNB strains were collected from 50 patients: 36 Pseudomonas aeruginosa strains, 12 Stenotrophomonas maltophilia strains, 6 Enterobacteriaceae strains, and 2 Acinetobacter baumannii strains. In P. aeruginosa, imipenem resistance was due to chromosomally encoded resistance (32 strains) or carbapenemase production (4 strains). In the Enterobacteriaceae strains, resistance was due to AmpC cephalosporinase and/or extended-spectrum β-lactamase production with porin loss. Genomic comparison showed that the strains were highly diverse, with 8 exceptions (4 VIM-2 carbapenemase-producing P. aeruginosa strains, 2 Klebsiella pneumoniae strains, and 2 S. maltophilia strains). The main risk factor for IR-GNB colonization was prior imipenem exposure. The odds ratio for colonization was already as high as 5.9 (95% confidence interval [95% CI], 1.5 to 25.7) after 1 to 3 days of exposure and increased to 7.8 (95% CI, 2.4 to 29.8) thereafter. In conclusion, even brief exposure to imipenem is a major risk factor for IR-GNB carriage.     

Novel VIM Metallo-β-Lactamase Variant from Clinical Isolates of Enterobacteriaceae from Algeria

Five different strains of bacteria belonging to the family Enterobacteriaceae were isolated from two patients hospitalized in the intensive care unit of the Central Military Hospital of Algiers, Algeria. All five strains, one Providencia stuartii strain, two Escherichia coli strains, and two Klebsiella pneumoniae strains, were intermediate or resistant to all β-lactams, including carbapenems. Synergy between imipenem and EDTA was observed for all five strains. The results of the PCR experiment confirmed the presence of a bla_VIM gene in all five strains. The bla_VIM genes were located as part of a class 1 integron on a 180-kb conjugative plasmid. They encoded a novel metallo-β-lactamase designated VIM-19, which differed from the parental enzyme VIM-1 by only two substitutions: Ser228Arg, previously observed in the closely related enzyme VIM-4, and Asn215Lys, not previously observed in other VIM-type carbapenemases. VIM-19 was further characterized after purification through determination of its kinetic constants. This enzyme was inhibited by EDTA and hydrolyzed penicillins, cephalosporins, and carbapenems, as observed for other VIM-type carbapenemases but with greater catalytic efficiency against penicillins than VIM-1. VIM-19 is the first carbapenemase enzyme identified from an isolate from Algeria. These results confirm the emergence of VIM-4-like enzymes in members of the family Enterobacteriaceae from Mediterranean countries.Carbapenems are the most active molecule in β-lactams with 98% susceptibility worldwide among bacteria belonging to the family Enterobacteriaceae (24). However, a growing number of carbapenemase-producing strains have been identified since the beginning of the 1990s. They displayed variable in vitro levels of resistance to carbapenems, including susceptibility (24), but were clinically resistant to these molecules (35). The possible low level of resistance makes reliable detection of such strains difficult (35).The geographical distribution of these strains differs: those producing class A KPC-type carbapenemases have been more frequently observed in the United States and in Israel, whereas those producing VIM- and IMP-type class B β-lactamases were generally encountered in Asia and in the northern part of the Mediterranean basin (24). Among these enzymes, metallocarbapenemases are especially worrying because they virtually hydrolyze all classes of β-lactams, except aztreonam (35). Initially observed in Pseudomonas aeruginosa and Acinetobacter spp., they spread to members of the family Enterobacteriaceae during the late 1990s and the 2000s (24). The most frequently acquired metallo-β-lactamases (MBLs) are the IMP and VIM types (24). Four other types of acquired MBLs in P. aeruginosa isolates from Brazil (SPM-1) (34) and Germany (GIM-1) (5), in Acinetobacter baumannii isolates from Korea (SIM-1) (17), and in a Citrobacter freundii isolate from Japan (KHM-1) (30) have recently been described.VIM-producing members of the family Enterobacteriaceae have been involved in different outbreaks in particular in Italy and Greece during the 2000s (4, 23, 33). The southern part of the Mediterranean basin seemed to be spared by such strains until 2006, when VIM-4-producing strains of Klebsiella pneumoniae were identified during a nosocomial outbreak in a Tunisian hospital (14), suggesting a possible dissemination in northwestern Africa.Between January and May 2008, five imipenem-resistant strains belonging to bacteria in the family Enterobacteriaceae were recovered from two patients in the intensive care unit of the Central Military Hospital of Algiers, Algeria. We identified a novel metallocarbapenemase, VIM-19, in these strains. This is the first report of VIM-producing strains in Algeria.     

In Vitro Activity of Aztreonam-Avibactam against a Global Collection of Gram-Negative Pathogens from 2012 and 2013

The combination of aztreonam plus avibactam is being developed for use in infections caused by metallo-β-lactamase-producing Enterobacteriaceae strains that also produce serine β-lactamases. The in vitro activities of aztreonam-avibactam and comparator antimicrobials were determined against year 2012 and 2013 clinical isolates of Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumannii using the broth microdilution methodology recommended by the Clinical and Laboratory Standards Institute (CLSI). A total of 28,501 unique clinical isolates were obtained from patients in 190 medical centers within 39 countries. MIC₉₀ values of aztreonam and aztreonam-avibactam against all collected isolates of Enterobacteriaceae (n = 23,516) were 64 and 0.12 μg/ml, respectively, with 76.2% of the isolates inhibited by ≤4 μg/ml of aztreonam (the CLSI breakpoint) and 99.9% of the isolates inhibited by ≤4 μg/ml of aztreonam-avibactam using a fixed concentration of 4 μg/ml of avibactam. The MIC₉₀ was 32 μg/ml for both aztreonam and aztreonam-avibactam against P. aeruginosa (n = 3,766). Aztreonam alone or in combination with avibactam had no in vitro activity against isolates of A. baumannii. PCR and sequencing were used to characterize 5,076 isolates for β-lactamase genes. Aztreonam was not active against most Enterobacteriaceae isolates producing class A or class C enzymes alone or in combination with class B metallo-β-lactamases. In contrast, >99% of Enterobacteriaceae isolates producing all observed Ambler classes of β-lactamase enzymes were inhibited by ≤4 μg/ml aztreonam in combination with avibactam, including isolates that produced IMP-, VIM-, and NDM-type metallo-β-lactamases in combination with multiple serine β-lactamases.     

Antimicrobial Activity of Ceftolozane-Tazobactam Tested against Enterobacteriaceae and Pseudomonas aeruginosa with Various Resistance Patterns Isolated in U.S. Hospitals (2011-2012)

Ceftolozane/tazobactam, a novel antimicrobial agent with activity against Pseudomonas aeruginosa (including drug-resistant strains) and other common Gram-negative pathogens (including most extended-spectrum-β-lactamase [ESBL]-producing Enterobacteriaceae strains), and comparator agents were susceptibility tested by a reference broth microdilution method against 7,071 Enterobacteriaceae and 1,971 P. aeruginosa isolates. Isolates were collected consecutively from patients in 32 medical centers across the United States during 2011 to 2012. Overall, 15.7% and 8.9% of P. aeruginosa isolates were classified as multidrug resistant (MDR) and extensively drug resistant (XDR), and 8.4% and 1.2% of Enterobacteriaceae were classified as MDR and XDR. No pandrug-resistant (PDR) Enterobacteriaceae isolates and only one PDR P. aeruginosa isolate were detected. Ceftolozane/tazobactam was the most potent (MIC_50/90, 0.5/2 μg/ml) agent tested against P. aeruginosa and demonstrated good activity against 310 MDR strains (MIC_50/90, 2/8 μg/ml) and 175 XDR strains (MIC_50/90, 4/16 μg/ml). Ceftolozane/tazobactam exhibited high overall activity (MIC_50/90, 0.25/1 μg/ml) against Enterobacteriaceae and retained activity (MIC_50/90, 4/>32 μg/ml) against many 601 MDR strains but not against the 86 XDR strains (MIC₅₀, >32 μg/ml). Ceftolozane/tazobactam was highly potent (MIC_50/90, 0.25/0.5 μg/ml) against 2,691 Escherichia coli isolates and retained good activity against most ESBL-phenotype E. coli isolates (MIC_50/90, 0.5/4 μg/ml), but activity was low against ESBL-phenotype Klebsiella pneumoniae isolates (MIC_50/90, 32/>32 μg/ml), explained by the high rate (39.8%) of meropenem coresistance observed in this species phenotype. In summary, ceftolozane/tazobactam demonstrated high potency and broad-spectrum activity against many contemporary Enterobacteriaceae and P. aeruginosa isolates collected in U.S. medical centers. Importantly, ceftolozane/tazobactam retained potency against many MDR and XDR strains.     

Characteristics of carbapenem-resistant Enterobacteriaceae isolates from Korea

In this study, the characteristics of carbapenem-resistant Enterobacteriaceae (CRE) isolates from Korea was investigated. A total of 22 CRE isolates were investigated, and most were identified as Klebsiella pneumoniae (16 isolates). In vitro antimicrobial susceptibility testing, multilocus sequence typing, and pulsed-field gel electrophoresis were performed. Extended-spectrum β-lactamase (ESBL) and carbapenemase genes were detected using gene amplification and sequencing. Efflux pump activity and inactivating mutations in OmpK35/36 were also investigated. Among 22 CRE isolates, only 5 produced metallo-β-lactamases (3 NDM-1, one VIM-2 and one IMP-1). Four and 2 K. pneumoniae and Serratia marcescens isolates showed resistance to polymyxins, respectively, and 2 CRE isolates (1 K. pneumoniae and C. freundii) were resistant to tigecycline. The prevalent carbapenem resistance mechanism found in K. pneumoniae might be porin defects. The most prevalent clone of carbapenem-resistant K. pneumoniae was ST11 (56.3%), which is the most frequently identified clone among ESBL-producing K. pneumoniae isolates from Korea. Three NDM-1-producing K. pneumoniae isolates belonged to a single clone (ST340) despite their different antimicrobial susceptibilities. In the present study, the clonal dissemination of carbapenem-resistant K. pneumoniae isolates (ST11) and NDM-1-producing K. pneumoniae isolates (ST340) was determined. Polymyxin- and tigecycline-resistant CRE isolates were also identified, which limits treatment options for infections causes by these organisms.     

Isolation of VIM-2-Producing Pseudomonas monteilii Clinical Strains Disseminated in a Tertiary Hospital in Northern Spain

We describe here the occurrence of bla_VIM-2 in 10 carbapenem-resistant Pseudomonas monteilii strains isolated from different clinical samples from patients at the University Hospital Marqués de Valdecilla in northern Spain. All the bla_VIM-2-harboring P. monteilii isolates possessed a class 1 integron, with the cassette array [intI1_bla_VIM-2_aac(6′)-Ib_qacEΔ1_sul1]. Our results show the emergence of VIM-2-producing multidrug-resistant species other than Pseudomonas aeruginosa or Pseudomonas putida in a Spanish hospital. P. monteilii, although sporadically isolated, should also be considered an important metallo-β-lactamase (MBL) reservoir.     

Rapid Identification of International Multidrug-Resistant Pseudomonas aeruginosa Clones by Multiple-Locus Variable Number of Tandem Repeats Analysis and Investigation of Their Susceptibility to Lytic Bacteriophages

The objective of this study was to determine the genetic diversity of multidrug-resistant (MDR) Pseudomonas aeruginosa strains isolated over a period of 12 months in two French hospitals and to test their susceptibility to bacteriophages. A total of 47 MDR isolates recovered from hospitalized patients were genotyped using multiple-locus variable number of tandem repeats analysis. The genotypes were distributed into five clones (including 19, 5, 5, 3, and 3 isolates, respectively) and 12 singletons. Comparison to 77 MDR strains from three other countries, and MLST analysis of selected isolates showed the predominance of international MDR clones. The larger clone, CC235, contained 59 isolates displaying different antibiotic resistance mechanisms, including the presence of the GES1, VIM-2, VIM-4, and IMP-1 β-lactamases. Three newly isolated P. aeruginosa bacteriophages were found to lyse 42 of the 44 analyzed strains, distributed into the different clonal complexes. This pilot study suggests that systematic genotyping of P. aeruginosa MDR strains could improve our epidemiological understanding of transmission at both the local (hospital) and the national level and that phage therapy could be an alternative or a complementary treatment to antibiotics for treating MDR-infected patients.     

Tn6249, a New Tn6162 Transposon Derivative Carrying a Double-Integron Platform and Involved with Acquisition of the blaVIM-1 Metallo-β-Lactamase Gene in Pseudomonas aeruginosa

The In70.2 integron platform appears to be a conserved structure involved in the dissemination of the bla_VIM-1 metallo-β-lactamase gene in Pseudomonas aeruginosa. The genetic context of the In70.2 integron platform from P. aeruginosa VR-143/97, the VIM-1-producing index strain isolated in Italy in 1997, was fully characterized by a next-generation sequencing approach refined by conventional sequencing. The In70.2 integron platform from VR-143/97 was found to be associated with a defective Tn402-like transposon inserted into the urf2 gene of a Tn3 family transposon of an original structure, named Tn6249, which also carried a partially deleted mer operon and an In90 integron platform in a tail-to-tail orientation. Tn6249 was inserted into a PACS171b-like genomic island, which was in turn inserted into the endA gene of the Pseudomonas chromosomal backbone. Tn6249 showed a similar structure and a conserved location with respect to that of Tn6060, a Tn3 family transposon associated with In70.2 and carrying a double-integron platform, which was detected in a VIM-1-producing P. aeruginosa strain isolated in Australia in 2008. Both Tn6249 and Tn6060 are apparently derived from Tn6162, a mercury resistance transposon carrying an integron platform, which was found in P. aeruginosa isolates from different geographic locations. The conservation of the genetic context of Tn6249 and Tn6060 suggests an in situ evolution of these elements after the insertion of a Tn6162-like ancestor into the PACS171b-like genomic island (GI) present in the genome of a successful widespread P. aeruginosa clonal lineage.     

Emergence of VIM-2 and IMP-15 Carbapenemases and Inactivation of oprD Gene in Carbapenem-Resistant Pseudomonas aeruginosa Clinical Isolates from Lebanon

We report here the emergence of VIM-2 and IMP-15 carbapenemases in a series of clinical isolates of carbapenem-resistant Pseudomonas aeruginosa in Lebanon. We also describe the disruption of the oprD gene by either mutations or insertion sequence (IS) elements ISPa1328 and ISPre2 isoform. Our study reemphasizes a rapid dissemination of the VIM-2 carbapenemase-encoding gene in clinical isolates of P. aeruginosa in the Mediterranean basin.