Common Carbapenemase Research Articles

Tigecycline in combination with other antibiotics against clinical isolates of carbapenem-resistant Klebsiella pneumoniae in vitro.

To investigate the activity of 5 antibiotic monotherapies, including colistin (COL), meropenem (MEM), amikacin (AMK), levofloxacin (LEV), and tigecycline (TGC), when combined with 4 other antibiotics against clinical isolates of carbapenem-resistant Klebsiella pneumoniae (CRKP) in vitro. The minimum inhibitory concentrations (MICs) of 5 antibiotics against 40 CRKP isolates were determined by micro-broth dilution method. There were synergistic effects between TGC combinations in the 10 CRKP isolates detected with checkerboard microdilution method. Time-kill assay was used to assess the monotherapies and the TGC combinations against 4 distinct sequence typing (STs) CRKP isolates. Polymerase chain reaction (PCR) tests were used to detect the carbapenemase genes, extended-spectrum beta lactamase (ESBL) genes, colistin resistance gene, and quinolone resistance genes, while multilocus sequence typing (MLST) was performed for 10 CRKP isolates. The MICs of TGC, COL, MEM, AMK, and LEV were 0.5-2, 2-32, 4-256, 1-16,384, and 0.5-64 µg/mL, respectively. The combinations exerted a significant synergism or additive effect via the checkerboard technique for most tested CRKP isolates, but a portion of the CRKP isolates had an indifferent effect except for the TGC-AMK combination. In addition, time-kill assays revealed that TGC enhanced the bactericidal activity of the 4 other antibiotics. Among 10 CRKP isolates, blaKPC-2 (90%), blaSHV (100%), and blaacc(6')-Ib (100%) were the most common carbapenemase genes, ESBL genes, and quinolone resistance genes, respectively. ST76 (70%) was the most predominant clone, followed by ST11 (10%), ST375 (10%), and ST530 (10%). In contrast to the currently recommended TGC therapy, our in vitro data suggest that TGC combinations may be a valid therapeutic option against CRKP, even in the presence of 1 antibiotic resistant isolate in TGC combination therapy. TGC-AMK combination is a cost-effective option for treating CRKP in the eastern region of Heilongjiang Province. In addition, TGC combinations might circumvent the overuse of carbapenems during the era of multi-drug resistance in Klebsiella pneumoniae (KP).

Transmission and evolution of OXA-48-producing Klebsiella pneumoniae ST11 in a single hospital in Taiwan.

Epidemic spread of OXA-48-producing Klebsiella pneumoniae, mainly mediated by the transmission of a blaOXA-48-carrying plasmid, has threatened global health during the last decade. Since its introduction to Taiwan in 2013, OXA-48 has become the second most common carbapenemase. We described the transmission and evolution of an OXA-producing K. pneumoniae clone in a single hospital. Twenty-two OXA-48 K. pneumoniae were isolated between October 2013 and December 2015. Comparative genomic analysis was performed based on the WGS data generated with Illumina and MinION techniques. Seventeen of the 22 OXA-48 K. pneumoniae that belonged to ST11, with the same capsular genotype, KL64, and differed from each other by seven or fewer SNPs, were considered outbreak strains. Eight of the 17 outbreak strains harboured a 65499 bp blaOXA-48-carrying IncL plasmid (called pOXA48). pOXA48 was absent from the remaining nine strains. Instead, a 24.9 kb blaOXA-48-carrying plasmid fragment was integrated into a prophage region of their chromosomes. Transmission routes of the ST11_KL64 K. pneumoniae sublineages, which carried either pOXA48 or chromosomally integrated blaOXA-48, were reconstructed. Clonal expansion of ST11_KL64 sublineages contributed to the nosocomial outbreak of OXA-48 K. pneumoniae. The chromosome-borne blaOXA-48 lineage emerged during a 2 year period in a single hospital. Dissemination of OXA-48, which is vertically transmitted in K. pneumoniae even in the absence of selective pressure from antimicrobials, deserves public health attention.

625. Genomic Epidemiology of Carbapenem-Resistant Enterobacteriaceae from Colombia: A Prospective Multicenter Study

BackgroundCarbapenem-resistant Enterobacteriaceae (CRE) is a serious public health threat. A major epidemic of carbapenemase-producing Klebsiella pneumoniae has occurred in Colombia through complex mechanisms of blaKPC dissemination. In the framework of a prospective, observational cohort study (CRACKLE-2), we aimed to characterize the genomic epidemiology of CRE circulating in Colombia.MethodsWe performed whole-genome sequencing of 52 isolates collected from the same number of patients (July 2017–April 2018) in 5 Colombian hospitals. Species confirmation and sequence type were determined using Strain Seeker and MLST database. Resistance genes were detected using ResFinder and CARD databases. Phylogenetic reconstruction included additional 108 isolates of carbapenem-resistant K. pneumoniae from a previous study.Results K. pneumoniae (36%), Escherichia coli (17%), and Enterobacter cloacae complex(17%) were the most frequent species. Genes conferring resistance to carbapenems were detected in 93% of isolates. blaKPC-2/3,blaNDM-1 andblaVIM-2/24 were identified in 81%, 15%, and 5% of isolates, respectively. Phylogenetic reconstructions of K. pneumoniae showed that clonal group 258 (CG258) were the predominant genetic lineage (Figure 1). Among CG258, ST11 was the most common comprising ca. 26% of isolates. Of note, ST11 had been extremely rare in previous surveillance studies in Colombia. The non-CG258 were from 9 different STs and exhibited high genomic diversity. Among E. coli isolates 33% belonged to the high-risk clone ST131 harboring blaKPC-2 and we detected both blaKPC-2 and blaVIM-24 in 1 E. coli ST131 isolate. ST510 E. cloacae complex harboring blaKPC-2 was the most common (44%) lineage.Conclusion K. pneumoniae and E. coli are the most frequent CRE isolated from patients in Colombian hospitals. Dissemination of blaKPC through horizontal gene transfer to several species of Enterobacteriaceae continues to be a common mechanism of spread. Although KPC continues to be the most common carbapenemase, a rise in high-risk clonal lineages harboring metallo-carbapenemases, in particular NDM-1 is worrisome. Our results indicate emergence of virulent genetic lineages of K. pneumoniae ST11 and E. coli ST131 carrying carbapenemases in Colombia. Disclosures All authors: No reported disclosures.

606. Identification and Characterization of HMB-2, a Novel Metallo-β-Lactamase in a Pseudomonas aeruginosa Isolate

BackgroundCarbapenemases, a global health threat, are a diverse group of β-lactamases active against cephalosporins and carbapenems, which are often last resort treatments for multidrug-resistant gram-negative infections. The most common carbapenemases reported among Pseudomonas aeruginosa are metallo-β-lactamase (MBLs). We describe a novel MBL (designated HMB-2) identified in a P. aeruginosa isolate from a urine specimen collected in 2015 as part of CDC’s Emerging Infections Program.MethodsWe performed antimicrobial susceptibility testing (AST) by broth microdilution, real-time PCR to screen for common carbapenemases (IMP, KPC, NDM, VIM, and OXA-48), and modified carbapenem inactivation method (mCIM) to test for carbapenemase production. The isolate underwent whole-genome sequencing (WGS) using Illumina MiSeq and PacBio RS II (Pacific Biosciences) platforms. Long read sequences were polished using Quiver and corrected by Pilon utilizing Illumina reads. We further characterized a putative novel MBL identified in WGS data by amplifying and cloning the gene into the pCR2.1-TOPO II vector (Invitrogen), which was then sub-cloned into a pET21 expression vector (Sigma–Aldrich). The resulting hmb2+ pET21 plasmid was transformed into a susceptible Escherichia coli for AST, including the imipenem-EDTA method to confirm MBL activity.ResultsThe isolate displayed resistance to carbapenems and demonstrated phenotypic carbapenemase activity (mCIM positive), but was negative for carbapenemase genes by PCR. WGS analyses identified a putative MBL gene located on the chromosome. The gene shared 98% DNA and protein sequence identity with an MBL reported in 2016 in a P. aeruginosa isolate from Germany (HMB-1) and thus was named hmb-2. The cloned hmb-2 gene conferred resistance to carbapenems (meropenem and ertapenem) and third-generation cephalosporins (cefotaxime and ceftazidime) in transformed E. coli. The Minimum Inhibitory Concentrationratio for the imipenem-EDTA method was ≥4.ConclusionA putative, novel β-lactamase gene, blaHMB-2, was identified and cloned. The imipenem-EDTA results indicated that HMB-2 is an MBL. This discovery underscores the important role WGS plays in identifying new mechanisms of antimicrobial resistance.Disclosures All authors: No reported disclosures.

518. Comparing the Mortality of Carbapenemase-Producing and Non-Carbapenemase-Producing Carbapenem-Resistant Enterobacteriaceae Bacteremia

BackgroundCarbapenem-resistant Enterobacteriaceae (CRE) infection is an emerging clinical issue. One of the mechanisms of carbapenem-resistance is carbapenemase production. This study aimed to identify whether clinical outcomes differ by CRE resistance mechanism and to evaluate risk factors for mortality in patients with CRE bacteremia.MethodsWe conducted a retrospective cohort study comparing 14-day mortality between patients with carbapenemase-producing (CP)-CRE and non-CP-CRE bacteremia during January 2011 to October 2018. Only monomicrobial Escherichia coli or Klebsiella pneumoniae bacteremia were included in the study. A modified carbapenem inactivation method was used for phenotypic detection of carbapenemase production. The presence of a variety of carbapenemase genes was evaluated by PCR with specific primers.ResultsOf 134 patients with monomicrobial CRE bacteremia, 48 (35.8%) were infected with CP-CRE, and 86 (64.1%) were infected with non-CP-CRE. The most common carbapenemase in CP-CRE isolates was KPC (66.7%), followed by NDM-1 (18.8%), OXA-48-like (10.4%), and VIM (4.1%). Baseline characteristics were similar between the two groups (Table 1). However, the CP-CRE group was significantly more likely to undergo removal of eradicable foci and to have meropenem MIC >8 µg/mL. A total of 33 (24.6%) patients died within 14 days, including 9 (18.8%) in the CP-CRE group and 24 (27.9%) in the non-CP-CRE group. Deceased patients were more likely to have a higher Pitt bacteremia score, nosocomial acquisition, ineradicable or not-eradicated foci, immunosuppressant use, inappropriate definitive treatment (Table 2). Combination therapy for definitive treatment was associated with decreased mortality. In a multivariate analysis including carbapenemase production, a higher Pitt bacteremia score (aOR, 5.15), ineradicable or not-eradicated foci (aOR, 4.05) and combination therapy for definitive treatment (aOR, 0.35) were independent risk factors for mortality.ConclusionOur study suggests that carbapenemase production is not a mortality risk factor in CRE bacteremia and provides additional evidence for early source control and combination therapy. Disclosures All authors: No reported disclosures.

2217. Frequency and Antimicrobial Susceptibility of Bacteria Isolated from Patients Hospitalized with Pneumonia in US Medical Centers During 2018

BackgroundRapidly introducing appropriate antimicrobial therapy is crucial to reduce morbidity and mortality of patients hospitalized with pneumonia (PHP), and therapy is determined mostly by understanding causative pathogens. Ceftazidime–avibactam (CAZ-AVI) was recently approved and ceftolozane–tazobactam (C-T) is in late-stage clinical development for treating nosocomial pneumonia, including ventilator-associated.MethodsBacterial isolates were consecutively collected from PHP (1/patient) in 67 US medical centers in 2018 and the Gram-negative bacilli (GNB) were tested by reference broth microdilution methods for susceptibility (S) to CAZ-AVI, C-T, and many comparators at a central laboratory.ResultsThe most common organisms isolated from PHP were S. aureus (27.0%), P. aeruginosa (PSA) (24.6%), K. pneumoniae (KPN; 7.6%), E. coli (6.8%), S. marcescens (5.4%), and S. maltophilia (XM; 4.5%). Colistin (99.7%S), CAZ-AVI (95.7%S), and C-T (94.9%S) were the most active compounds against PSA; CAZ-AVI (99.9%S), amikacin (AMK; 98.8%S), and meropenem (MEM; 97.6%S) were the most active compounds against Enterobacterales (ENT). CAZ-AVI and C-T retained activity against PSA isolates non-S (NS) to piperacillin–tazobactam (PIP-TAZ), MEM, and cefepime (FEP), whereas PSA isolates NS to PIP-TAZ, MEM, or FEP exhibited low S rates to PIP-TAZ (≤ 39.2%), MEM (≤ 37.8%), and FEP (≤ 38.0%; Table). CAZ-AVI and tigecycline were the only compounds with good activity against carbapenem-resistant ENT (CRE), both with 96.6%S. Among ENT, the most common ESBL and carbapenemase were CTX-M-15 (73%) and KPC-2/3 (76%), respectively. CAZ-AVI was active against all ESBL producers (100.0%S), whereas the S rate to C-T was 82.4%. The most active compounds against multidrug-resistant (MDR) ENT were CAZ-AVI (98.9%S), AMK (91.5%S), and MEM (80.8%S). XM and A. baumannii exhibited low S rates to most antimicrobials tested.ConclusionGram-negative bacteria were isolated from 70% of PHP, and PSA and ENT represented >80% of these organisms. CAZ-AVI and C-T showed similar coverage (%S) against PSA (95.7–94.9%S). In contrast, C-T was less active than CAZ-AVI against ENT in general and exhibited limited activity against ENT-resistant subsets. Disclosures All authors: No reported disclosures.

721. In Vitro Activity of Cefiderocol Against Gram-Negative Clinical Isolates From New York City

BackgroundMultidrug-resistant Gram-negative bacteria have become a serious problem in hospitals worldwide. Cefiderocol (CFDC) is a novel siderophore cephalosporin with activity against a wide range of carbapenemase- and ESBL-producing bacteria. We tested the activity of CFDC against (1) a recent collection of clinical isolates and (2) a separate collection of carbapenem-resistant isolates gathered from NYC hospitals.MethodsSusceptibility testing was performed on isolates of E. coli, K. pneumoniae, Enterobacter spp., P. aeruginosa, and A. baumannii gathered in 2017 from 7 hospitals in Brooklyn, NY. Consecutive unique patient clinical isolates from all sources were collected for a three month period. Testing was also done on a collection of carbapenem-resistant isolates from a similar surveillance study conducted in 2013–2014. MICs were performed with iron-depleted cation-adjusted Mueller–Hinton broth for CFDC and agar dilution for other antibiotics according to CLSI methodology. The provisional CLSI breakpoint (≤4 μg/mL susceptible) was used for CFDC. Cephalosporin-resistant isolates were tested for common carbapenemases by PCR.ResultsThe susceptibility results for CFDC and meropenem for the isolates gathered in 2017 are listed in the Table. All of the Enterobacteriacae were susceptible to CFDC including KPC-possessing E. coli (n = 4), K. pneumoniae (n = 20), and Enterobacter spp (n = 3). 99.6% of P. aeruginosa and 100% of A. baumannii (including 8 with blaOXA-23, 2 with blaOXA-24, and 1 with blaKPC) were susceptible to CFDC. For the collection of carbapenem-resistant isolates gathered in 2013–14, 100% of K. pneumoniae (n = 111), 100% of P. aeruginosa (n = 130), and 90% of A. baumannii (n = 78) were susceptible to CFDC.ConclusionCFDC has excellent in vitro activity against Gram-negative clinical isolates from NYC, including a large collection of carbapenem-resistant Enterobacteriaceae, P. aeruginosa, and A. baumannii. Disclosures All authors: No reported disclosures.

695. Activity of Imipenem–Relebactam and Ceftolozane–Tazobactam Against a Contemporary Collection of Gram-Negative Bacteria from New York City

BackgroundCarbapenem-resistant Gram-negative bacteria are important nosocomial pathogens, and therapeutic options are often limited.MethodsClinical isolates were gathered during a surveillance study in 2017 involving 7 hospitals in Brooklyn, NY. Isolates underwent susceptibility testing using the agar dilution method; for the combination of imipenem-relebactam and ceftolozane-tazobactam, the concentrations of relebactam and tazobactam were fixed at 4 µg/mL. Breakpoints were defined according to CLSI criteria; for imipenem-relebactam, the breakpoint of imipenem was utilized. Isolates were screened by PCR for common carbapenemases.ResultsOverall susceptibility patterns are given in the Table. Of 1805 isolates of E. coli (including 4 with blaKPC), 100% were susceptible to imipenem and imipenem-relebactam. Of 503 isolates of K. pneumoniae (including 19 isolates with blaKPC), all were susceptible to imipenem-relebactam. Of 171 isolates of Enterobacter spp. (including 3 with blaKPC), 100% were susceptible to imipenem-relebactam. Of 260 isolates of P. aeruginosa, 96% were susceptible to imipenem-relebactam and nearly all to ceftolozane-tazobactam. Against A. baumannii, the activity of imipenem-relebactam was the same as imipenem and the ceftolozane-tazobactam MIC was ≤ 4 µg/mL in 65% of isolates.ConclusionImipenem-relebactam possesses promising activity against multidrug-resistant Enterobacteriaceae endemic to New York City. Ceftolozane-tazobactam demonstrated excellent activity against P. aeruginosa, including isolates resistant to carbapenems. Disclosures All authors: No reported disclosures.

728. Activity of Eravacycline Against Contemporary Gram-Negative Clinical Isolates From New York City Hospitals

BackgroundAntibiotic-resistant Gram-negative bacteria, including KPC-producing Enterobacteriaceae and carbapenem-resistant A. baumannii, have been problematic hospital pathogens in NYC and other areas. Eravacycline (ERV), a fluorocycline antibiotic released in the USA in 2018, has demonstrated in vitro activity against many of these strains. We tested the activity of ERV against a recent collection of clinical isolates from NYC hospitals.MethodsFor a 3-month period in 2017, all unique patient isolates of E. coli, K. pneumoniae, Enterobacter spp., and A. baumannii were collected from 7 hospitals in Brooklyn, NY. MICs were performed by broth microdilution for ERV and Tigecycline (TGC) and agar dilution for other antibiotics according to CLSI methodology. Cephalosporin-resistant isolates were screened by PCR for common carbapenemases.ResultsThe susceptibility results for tetracycline and ERV are listed in the Table. Overall, 95% of the Enterobacteriaceae were inhibited by ≤ 0.5 μg/mL of ERV, the FDA-suggested breakpoint. Of 1,876 isolates of E. coli, 4 possessed KPC. ERV MICs for these 4 isolates were 0.125–0.25 μg/mL. Of 518 isolates of K. pneumoniae, 20 possessed KPC. The ERV MIC50 and MIC90 for these isolates were 1 and 1 μg/mL, respectively. Of 172 isolates of Enterobacter spp., 3 possessed KPC. ERV MICs for these 3 isolates were 0.5–1 μg/mL. Of 45 isolates of A. baumannii, 11 isolates possessed a carbapenemase (OXA23 in 8, OXA24 in 2, and KPC in 1). The ERV MIC50 and MIC90 for these isolates were 1 and 2 μg/mL, respectively. Overall, ERV MICs were two-fold lower than TGC MICs for A. baumannii.ConclusionERV possesses significant in vitro activity against contemporary clinical isolates of Enterobacteriaceae and A. baumannii from NYC, including many carbapenemase producing strains. Disclosures All authors: No reported disclosures.

Characterization of phenotypic and genotypic traits of carbapenem-resistant Acinetobacter baumannii clinical isolates recovered from a tertiary care hospital in Taif, Saudi Arabia.

Background and objectiveAcinetobacter baumannii (A. baumannii) is a common nosocomial pathogen, which developed multi-drug-resistance to different classes of antibiotics including carbapenems. This study examined ten common carbapenemase genes among 32 carbapenem-resistant A. baumannii clinical isolates recovered from Taif, Saudi Arabia.MethodsIsolates were phenotypically identified to the genus level by Vitek®2 and API 20NE®. The species level was confirmed by the amplification of blaOXA-51. The susceptibility for 21 different antibiotics was performed by Vitek 2 and modified Kirby-Bauer method. Isolates were genetically screened for 10 carbapenemases. Phylogenetic relatedness between isolates was determined by ERIC-PCR.ResultsGenotypically identified A. baumannii represented 100% of the total phenotypically identified Acinetobacter spp. All the carbapenem-resistant isolates were sensitive to polymyxin B and colistin. Among the other antibiotics, ampicillin/sulbactam and tigecycline were the most effective agents. 90.8% of the isolates were resistant to all ten investigated β–lactams. blaOXA-51, blaIPM, blaNDM and blaOXA-23 were detected in 100%, 87.5%, 62.5% and 59.4% of isolates, respectively. Also, blaVIM and blaOXA-40 were less prevalent and were detected in 9.3% and 3.1% of the isolates, respectively. In addition, blaKPC, blaOXA-48, blaOXA-58, blaOXA-181 were not detected in any isolate. The A. baumannii isolates were categorised into ten genotypes on the basis of the detected carbapenemase genes and ERIC-PCR revealed a remarkable clonal diversity among these isolates.ConclusionClass A and class D carbapenemase genes were the most commonly detected among carbapenem resistant A. baumannii (CRAB) clinical isolates.

Carbapenemase Genes and Multidrug Resistance of Acinetobacter Baumannii: A Cross Sectional Study of Patients with Pneumonia in Southern Vietnam.

Background: Acinetobacter baumannii (Ab) is an opportunistic bacterial pathogen found in hospital-acquired infections including nosocomial pneumonia, especially multidrug-resistant Ab. This study aims to survey the drug resistance profiles of Ab isolated from patients in Thong Nhat Dong Nai General Hospital and assess the relationship between genotypes and antibiotic resistance; Methods: Ninety-seven Ab strains isolated from 340 lower respiratory tract specimens among pneumonia patients were used to screen the most common local carbapenemase genes. Antimicrobial susceptibility testing results and demographic data were collected and minimum inhibitory concentrations (MIC) of colistin were also determined; Results: Over 80% and 90% of Ab strains were determined as carbapenem-resistant and multidrug-resistant (MDR), respectively. Most of the strains carried carbapenemase genes, including blaOXA-51, blaOXA-23-like, blaOXA-58-like, and blaNDM-1, with proportions of 97 (100%), 76 (78.4%), 10 (10.3%), 6 (6.2%), respectively. Amongst these genes, blaOXA-23-like was the only gene which significantly influenced the resistance (p < 0.0001); and Conclusions: The severity of Ab antibiotic resistance is urgent and specifically related to carbapenemase encoding genes. Therefore, screening of MDR Ab and carbapenemase for better treatment options is necessary.

Isolation and characterisation of carbapenem-resistant Xanthomonas citri pv. mangiferaeindicae-like strain gir from the faecal material of giraffes.

The purpose of this study was to determine if giraffes (Giraffa camelopardalis) living in captivity at the Jacksonville Zoo and Gardens, Jacksonville, FL were colonised with carbapenem-resistant bacteria and, if found, to identify underlying genetic mechanisms contributing to a carbapenem-resistant phenotype. Faecal samples from seven giraffes were examined for carbapenem-resistant bacteria. Only one isolate (a Xanthomondaceae) was found to be carbapenem-resistant by antibiotic susceptibility testing. This isolate was selected for additional characterization, including whole genome sequencing (WGS). Based on average nucleotide identity, the bacterium was identified as Xanthomonas citri pv. mangiferaeindicae-like strain gir. Phenotypic carbapenemase tests and PCR for the most common carbapenemase genes produced negative results, suggesting that carbapenem resistance was mediated by another mechanism. Resistance gene profile analysis of WGS results confirmed these results. Among identified resistance genes, a chromosomal class A beta-lactamase with 71% identity to the penP beta-lactamase gene from Xanthomonas citri ssp. citri was identified, which could contribute to a carbapenem-resistant phenotype.

Susceptibility of carbapenemase-producing Enterobacterales (CPE) to nitroxoline.

Infections caused by carbapenemase-producing Enterobacterales (CPE) constitute a major global health concern and are associated with increased morbidity and mortality. Nitroxoline is an old antibiotic, which has recently been re-launched for the treatment of uncomplicated urinary tract infection. Because of low resistance rates it could be an interesting option for treatment of MDR isolates, yet data on CPE susceptibility are scarce. To analyse the in vitro activity of nitroxoline against CPE. MICs of nitroxoline were determined by agar dilution for a collection of well-characterized carbapenemase producers (n = 105), producing OXA-48-like (n = 36), VIM (n = 21), IMI (n = 9), IMP (n = 6), NDM (n = 22), KPC (n = 11), OXA-58 (n = 2) and GES (n = 2). For comparison, MICs of ertapenem, imipenem and meropenem were determined by agar gradient diffusion. For all 105 isolates, the MIC50/90 of nitroxoline was 8/16 mg/L. All Escherichia coli isolates (30/30, 100%) showed low MICs of 2-8 mg/L and were susceptible to nitroxoline. MICs of 32 mg/L were recorded for five isolates of VIM- and IMI-producing Enterobacter cloacae (n = 3) and OXA- and VIM-producing Klebsiella pneumoniae (n = 2). Nitroxoline exhibited excellent in vitro activity against most isolates producing common and rare carbapenemases. If the current EUCAST susceptibility breakpoint of ≤16 mg/L for E. coli in uncomplicated urinary tract infections was applied, 95.2% (100/105) of isolates would be classified as susceptible. Nitroxoline could therefore be an alternative oral option for treatment of uncomplicated urinary tract infections caused by CPE.

Resistance mechanisms and molecular epidemiology of carbapenem-nonsusceptible Escherichia coli in Taiwan, 2012-2015.

PurposeThis study aimed to investigate the resistance mechanisms and molecular epidemiology of carbapenem-nonsusceptible Escherichia coli (CnsEC) in Taiwan.Patients and methodsFrom 2012 to 2015, 237 E. coli isolates with minimum inhibitory concentrations of imipenem or meropenem >1 μg/mL were collected in a nationwide surveillance and subjected to polymerase chain reaction (PCR) for carbapenemase, AmpC-type β-lactamase, and extended spectrum β-lactamase (ESBL) genes. We evaluated outer membrane proteins (OmpF and OmpC) loss and conducted multilocus sequence typing and pulsed-field gel electrophoresis (PFGE). Isolates that were resistant to all carbapenems were designated as pan-carbapenem-resistant E. coli (pCREC) in this study.ResultsThe predominant resistance mechanism of CnsEC in Taiwan was the CMY-2 β-lactamase in combination with OmpF and OmpC loss. Sequence type 131 was the most prevalent type (29.2%). Among 237 CnsEC isolates, 106 (44.7%) isolates were pCREC and 18 (7.59%) produced carbapenemase. The prevalence of carbapenemases increased from 6% in 2012 to 11.36% in 2015. Various carbapenemases including KPC-2, IMP-8, NDM-1, NDM-5, VIM-1, OXA-48, and OXA-181 were identified, with NDM-1 being the most common (38.9%) carbapenemase. Comparison between pCREC and non-pCREC among the non-carbapenemase-producing CnsEC isolates revealed SHV, CMY, co-carriage of SHV and CTX-M and concurrent loss of both OmpF and OmpC were more commonly detected in the pCREC group. PFGE revealed no nationwide clonal spread of carbapenemase-producing E. coli.ConclusionNDM-1 was the most common carbapenemase and combination of CMY-2 and concurrent OmpF and OmpC porin loss was the most prevalent resistance mechanism in CnsEC in Taiwan.

Arising Prevalence of OXA-48 producer Escherichia coli and OXA-48 with NDM co-producer Klebsiella pneumoniae Strains

Abstract Background/aim: This prospective study aimed to determine the presence of the most common carbapenemase genes, blaOXA-48, blaKPC, blaIMP, blaVIM and blaNDM on carbapenem resistant clinical K.pneumoniae and E.coli isolates. Materials and methods: Isolates were selected according to EUCAST guideline; gradient test and disc diffusion with both meropenem and ertapenem discs. Resistance rates of these isolates to other antimicrobial agents were also examined by disc diffusion method. Carbapenem resistance gene were investigated by using Real-Time PCR. Results: A total of 3845 E. coli and 1689 K.pneumoniae isolates from clinical samples between January 2015 and April 2017 were evaluated. The 419 isolates were found as carbapenem resistant but only the first resistant isolate (n=155; 126 K.pneumoniae and 29 E.coli) of each patient were included. Carbapenem resistant isolates were most frequently isolated from intensive care units (48.8%). Colistin was the most effective antibiotic (91.0%). The 121 (78.1%) of the tested isolates were positive for OXA-48 (103 K.pneumoniae and 18 E.coli) and 9 K. pneumoniae carrying blaNDM were also positive for blaOXA-48. VIM, IMP and KPC type carbapenemases were not detected in any isolates. Conclusion: Carbapenem-resistant pathogens have been shown to be able to develop resistance mechanisms with more than one carbapenemase encoding gene.

Sewage effluent from an Indian hospital harbors novel carbapenemases and integron-borne antibiotic resistance genes

BackgroundHospital wastewaters contain fecal material from a large number of individuals, of which many are undergoing antibiotic therapy. It is, thus, plausible that hospital wastewaters could provide opportunities to find novel carbapenemases and other resistance genes not yet described in clinical strains. Our aim was therefore to investigate the microbiota and antibiotic resistome of hospital effluent collected from the city of Mumbai, India, with a special focus on identifying novel carbapenemases.ResultsShotgun metagenomics revealed a total of 112 different mobile antibiotic resistance gene types, conferring resistance against almost all classes of antibiotics. Beta-lactamase genes, including encoding clinically important carbapenemases, such as NDM, VIM, IMP, KPC, and OXA-48, were abundant. NDM (0.9% relative abundance to 16S rRNA genes) was the most common carbapenemase gene, followed by OXA-58 (0.84% relative abundance to 16S rRNA genes). Among the investigated mobile genetic elements, class 1 integrons (11% relative abundance to 16S rRNA genes) were the most abundant. The genus Acinetobacter accounted for as many as 30% of the total 16S rRNA reads, with A. baumannii accounting for an estimated 2.5%. High throughput sequencing of amplified integron gene cassettes identified a novel functional variant of an IMP-type (proposed IMP-81) carbapenemase gene (eight aa substitutions) along with recently described novel resistance genes like sul4 and blaRSA1. Using a computational hidden Markov model, we detected 27 unique metallo-beta-lactamase (MBL) genes in the shotgun data, of which nine were novel subclass B1 genes, one novel subclass B2, and 10 novel subclass B3 genes. Six of the seven novel MBL genes were functional when expressed in Escherichia coli.ConclusionBy exploring hospital wastewater from India, our understanding of the diversity of carbapenemases has been extended. The study also demonstrates that the microbiota of hospital wastewater can serve as a reservoir of novel resistance genes, including previously uncharacterized carbapenemases with the potential to spread further.

Research progress in epidemiological characteristics of carbapenem-resistant Enterobacteriaceae

Carbapenem-resistant Enterobacteriaceae (CRE) are defined as bacteria of the Enterobacteriaceae family exhibiting resistance to any carbapenem antibiotics. As some of the most common pathogens, Enterobacteriaceae spread worldwide and often cause large-scale outbreaks and transmission in medical institutions or regions. Increasing resistance of these bacteria to clinically common-used anti-infective drugs results in more and more treatment failure and brings enormous challenges to nosocomial infection control and anti-infection treatment. Understanding the distribution and transmission of CRE is of great significance for controlling the spread of such pathogens. This article reviews the progress in the distribution and epidemic characteristics of CRE. Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae (K.pneumoniae) was first isolated in the United States and has become the predominant species of CRE worldwide. In recent years, KPC-2- and KPC-3-producing K. pneumoniae are widely spread in many regions. NDM-1 has been detected worldwide since it was isolated from Escherichia coli (E.coli) and K. pneumoniae in India and Britain in 2010 and become the second most common carbapenemase produced by Enterobacteriaceae other than KPC. OXA-48, which was first detected from a carbapenem-resistant K. pneumoniae in 2004, has weak hydrolytic activity against carbapenems. Large-scale outbreaks caused by OXA-48-producing Enterobacteriaceae have occurred in many countries such as Turkey, Spain, Morocco and Tunisia. In China, the predominant species of CRE is KPC-producing K. pneumoniae mainly belonging to the sequence type 11 (ST11). In addition, KPC-producing Klebsiella oxytoca (K.oxytoca), E. coli, Enterobacter cloacae (E.cloacae), Citrobacter freundii (C.freundii), Serratia marcescens, Proteus mirabilis (P.mirabilis), Enterobacter fluvialis (E.fluvialis) and Morgan morganii have also been reported. The prevalent NDM-producing species isolated in China are E. coli and K. pneumoniae. Moreover, blaNDM is also carried by E. cloacae, K. oxytoca, C. freundii, P. mirabilis and Enterobacter aerogenes (E.aerogenes). Small-scale outbreaks caused by OXA-48-producing K. pneumoniae have only been reported in Beijing and Taiwan in China. Key words: Carbapenemase; Drug resistance; Enterobacteriaceae; Carbapenem-resistant Enterobacteriaceae

Comparison of five methods for detection of carbapenemases in Enterobacterales with proposal of a new algorithm

ObjectivesThe aim of this study was to evaluate the performance of five different carbapenemase tests and to develop an algorithm which will permit the detection of most common and rare carbapenemases in routine microbiology laboratories. MethodsThe immunochromatographic tests CARBA-5 (NG), RESIST-4 O.K.N.V. (Coris), the colorimetric β-CARBA (BioRad), a newly developed carbapenem-inactivation method (CIM) supplemented with zinc (zCIM), and the Xpert Carba-R (Cepheid) were challenged with a collection of 189 molecularly characterized Enterobacterales isolates, including 146 carbapenemase producers (CPE): VIM (n = 48), OXA-48-like (n = 40), NDM (n = 29), KPC (n = 13), IMI (n = 9), IMP (n = 9), OXA-58 (n = 2), and GES (n = 2). ResultsThe overall sensitivity/specificity values for the five carbapenemase detection tests were 84.2% (CI 77.6–89.2%)/100% (CI 91.8–100%) for RESIST-4, 88.2% (CI 82.1–92.4%)/100% (CI 91.8–100%) for CARBA-5, 88.2% (CI 82.1–92.4%)/100% (CI 91.8–100%) for Xpert Carba-R, 73.7% (CI 66.2–80.0%)/100% (CI 93.4–99.0%) for β-CARBA, and 97.4% (CI 87.9–99.6%)/97.7% (CI 87.9–99.6%) for zCIM. The four common carbapenemases (KPC, OXA-48-like, NDM, and VIM) were detected with ≥97.6% sensitivity by all tests except for β-CARBA (76.6% (CI 68.4–83.2%)). IMI and GES were only detected by zCIM (sensitivity 90.9% (CI 62.3–98.4%)). Based on these results a new algorithm was developed, consisting of an immunochromatographic assay as the first test followed by zCIM, which allows detection of 99.3% of all carbapenemases assessed. ConclusionsExcept for β-CARBA, all methods showed excellent sensitivity/specificity for the detection of the four most frequent carbapenemases. With the new algorithm, rare variants can also be detected. It is rapid, simple, and inexpensive and can be performed in any microbiology laboratory, as no PCR equipment is required.

Epidemiological, clinical, and microbiological characteristics of carbapenemase-producing Enterobacteriaceae bloodstream infection in the Republic of Korea

BackgroundCarbapenemase-producing Enterobacteriaceae (CPE) is an important pathogen in nosocomial infections; nevertheless, only a few studies regarding CPE infection and its epidemiological factors have been conducted in the Republic of Korea (ROK). We aimed to analyze the clinical, microbiological, and epidemiological characteristics of CPE bloodstream infections (BSIs) in the ROK.MethodsThis retrospective cohort study included data collected from the National Surveillance System from January 2015 to December 2016 based on the epidemiologic survey performed by an epidemiologist from the Korea Centers for Disease Control and Prevention. We selected patients with CPE BSI from the Korea National Institute of Health based on carbapenemase genotyping.ResultsIn this study, 131 CPE BSIs were identified, and the proportion of CPE BSI among total CPE isolates was 7%. Klebsiella pneumoniae accounted for 69% of all CPE BSIs, and 66% of these produced K. pneumoniae carbapenemase. Among nine provinces in ROK, one province had NDM as the most common carbapenemase. CPE was susceptible to amikacin, tigecycline, and gentamicin (76, 41, and 39%, respectively). Of 29 patients tested for colistin sensitivity, one patient showed colistin resistance. The most common CPE BSI sources were pneumonia, primary bacteremia, and biliary tract infection. Multivariable analysis showed that adequate antibiotic use at CPE detection was significantly associated with decreased 30-day mortality.ConclusionsCPE BSIs are prevalent in the ROK. Moreover, most CPE BSIs originated from hospital-acquired infection, demonstrating the need to improve hospital infection control strategies.

Performance evaluation of the PANA RealTyper™ CRE Kit for detecting carbapenemase genes in Gram-negative bacilli.

The spread of carbapenemase-producing organisms has been continuously reported over the past decade. Rapid and accurate detection of carbapenemase production is essential for adequate infection control and appropriate antimicrobial treatment in the clinical setting. In this study, the performance of the newly developed PANA RealTyper™ CRE Kit (PANAGENE, Daejeon, South Korea) for the detection of six common carbapenemase genes (blaKPC, blaGES, blaIMP, blaNDM, blaVIM and blaOXA-48-like) was evaluated. A total of 479 non-duplicate clinical isolates of Gram-negative bacilli, including 391 carbapenemase-producers and 88 non-producers, were tested. Conventional PCR and sequencing were performed as reference methods for performance evaluation of the PANA RealTyper™ CRE Kit. The PANA RealTyper™ CRE Kit showed a reliable performance for the detection of carbapenemase-producing organisms compared with the monoplex PCR system, with sensitivity and specificity values both >95%, with only a few discrepant results for six types of carbapenemase genes. This kit is rapid and accurate for simultaneously detecting various carbapenemase genes in a single reaction and could contribute to early decisions for appropriate antimicrobial treatment in the clinical setting.