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

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S283-S284
Author(s):  
Wenming Zhu ◽  
Gillian A McAllister ◽  
Maria Jose Machado ◽  
Davina Campbell ◽  
Maria Karlsson ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Illumina Miseq ◽  
Multidrug Resistant ◽  
Urine Specimen ◽  
Emerging Infections ◽  
E Coli ◽  
Long Read ◽  
Protein Sequence Identity ◽  
Topo Ii ◽  
Lactamase Gene

Abstract Background Carbapenemases, 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. Methods We 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. Results The 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. Conclusion A 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.

scholarly journals Presence of β-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals

2021 ◽  
Vol 22 (11) ◽  
pp. 5905
Author(s):  
Olivia M. Grünzweil ◽  
Lauren Palmer ◽  
Adriana Cabal ◽  
Michael P. Szostak ◽  
Werner Ruppitsch ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Marine Mammals ◽  
Virulence Genes ◽  
Multidrug Resistant ◽  
Healthcare Sector ◽  
Whole Genome ◽  
E Coli ◽  
High Prevalence ◽  
Lactamase Gene ◽  
Sequence Types

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum β-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant β-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-β-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.

scholarly journals Post-antibiotic era in hemodialysis? Two case reports of simultaneous colonization and bacteremia by multidrug-resistant bacteria

2020 ◽  
Author(s):  
Johanna M. Vanegas ◽  
Lorena Salazar-Ospina ◽  
Gustavo A. Roncancio ◽  
Julián Builes ◽  
Judy Natalia Jiménez
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Infections ◽  
Case Reports ◽  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Control Measures ◽  
Resistant Bacteria ◽  
Multidrug Resistant Bacteria ◽  
E Coli ◽  
Carbapenem Resistant

ABSTRACT The emergence of resistance mechanisms not only limits the therapeutic options for common bacterial infections but also worsens the prognosis in patients who have conditions that increase the risk of bacterial infections. Thus, the effectiveness of important medical advances that seek to improve the quality of life of patients with chronic diseases is threatened. We report the simultaneous colonization and bacteremia by multidrug-resistant bacteria in two hemodialysis patients. The first patient was colonized by carbapenem- and colistin-resistant Klebsiella pneumoniae, carbapenem-resistant Pseudomonas aeruginosa, and methicillin-resistant Staphylococcus aureus (MRSA). The patient had a bacteremia by MRSA, and molecular typing methods confirmed the colonizing isolate was the same strain that caused infection. The second case is of a patient colonized by extended-spectrum beta-lactamases (ESBL)-producing Escherichia coli and carbapenem-resistant Pseudomonas aeruginosa. During the follow-up period, the patient presented three episodes of bacteremia, one of these caused by ESBL-producing E. coli. Molecular methods confirmed colonization by the same clone of ESBL-producing E. coli at two time points, but with a different genetic pattern to the strain isolated from the blood culture. Colonization by multidrug-resistant bacteria allows not only the spread of these microorganisms, but also increases the subsequent risk of infections with limited treatments options. In addition to infection control measures, it is important to establish policies for the prudent use of antibiotics in dialysis units.

scholarly journals Acquired fluoroquinolone resistance genes in corneal isolates of Pseudomonas aeruginosa

2020 ◽  
Author(s):  
Mahjabeen Khan ◽  
Stephen Summers ◽  
Scott A Rice ◽  
Fiona Stapleton ◽  
Mark D P Willcox ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Resistance Genes ◽  
Empirical Therapy ◽  
Illumina Miseq ◽  
Whole Genome Sequence ◽  
Fluoroquinolone Resistance ◽  
Beta Lactam ◽  
Long Reads ◽  
Long Read ◽  
Class I Integrons

AbstractFluroquinolones are widely used as an empirical therapy for pseudomonal ocular infections. Based on increasing reports on acquired fluroquinolone resistance genes in clinical isolates of Pseudomonas aeruginosa, we investigated 33 strains of P. aeruginosa isolated from the cornea of microbial keratitis patients in India and Australia between 1992 and 2018 to understand the prevalence of acquired fluroquinolone resistance genes in ocular isolates and to assess whether the possession of those genes was associated with fluoroquinolone susceptibility. We obtained the whole genome sequence of 33 isolates using Illumina MiSeq platform and investigated the prevalence of two fluoroquinolone resistance genes crpP and qnrVC1. To examine the associated mobile genetic elements of qnrVC1 positive strains, we obtained long read sequences using Oxford Nanopore MinION and performed hybrid assembly to combine long reads with Illumina short sequence reads. We further assessed mutations in QRDRs and antibiotic susceptibilities to ciprofloxacin, levofloxacin and moxifloxacin to examine the association between resistance genes and phenotype. Twenty strains possessed crpP in genetic islands characterised by possession of integrative conjugative elements. The qnrVC1 gene was carried by four isolates on class I integrons and Tn3 transposons along with aminoglycoside and beta-lactam resistance genes. We did not observe any evidence of plasmids carrying fluroquinolone resistance genes. Resistance to fluroquinolones was observed in those strains which possessed crpP, qnrVC1 and that had QRDRs mutations. The presence of crpP was not a sole cause of fluroquinolone resistance.

scholarly journals Carbapenem and colistin-resistant bacteria in North Lebanon: Coexistence of mcr-1 and NDM-4 genes in Escherichia coli

2021 ◽  
Vol 15 (07) ◽  
pp. 934-342
Author(s):  
Charbel Al-Bayssari ◽  
Tania Nawfal Dagher ◽  
Samar El Hamoui ◽  
Fadi Fenianos ◽  
Nehman Makdissy ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Resistance Genes ◽  
Multidrug Resistant ◽  
Resistant Bacteria ◽  
Ionization Time ◽  
E Coli ◽  
Flight Mass Spectrometry ◽  
Carbapenem Resistant ◽  
The Matrix

Introduction: The increasing incidence of infections caused by multidrug-resistant bacteria is considered a global health problem. This study aimed to investigate this resistance in Gram-negative bacteria isolated from patients hospitalized in North-Lebanon. Methodology: All isolates were identified using the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Antibiotic susceptibility testing was achieved using disk diffusion, E-test and Broth microdilution methods. Phenotypic detection of carbapenemase was carried out using the CarbaNP test. RT-PCR, standard-PCR and sequencing were performed to detect resistance genes and oprD gene. Conjugal transfer was carried out between our isolates and Escherichia coli J53 to detect the genetic localization of resistance genes. MLST was conducted to determine the genotype of each isolate. Results: Twenty-three carbapenem-resistant Enterobacterales of which eight colistin-resistant Escherichia coli, and Twenty carbapenem-resistant Pseudomonas aeruginosa were isolated. All isolates showed an imipenem MIC greater than 32 mg/mL with MICs for colistin greater than 2 mg/L for E. coli isolates. All the Enterobacterales isolates had at least one carbapenemase-encoding gene, with E. coli isolates coharboring blaNDM-4 and mcr-1 genes. Moreover, 16/20 Pseudomonas aeruginosa harbored the blaVIM-2 gene and 18/20 had mutations in the oprD gene. MLST revealed that the isolates belonged to several clones. Conclusions: We report here the first description in the world of clinical E. coli isolates coharboring blaNDM-4 and mcr-1 genes, and K. pneumoniae isolates producing NDM-6 and OXA-48 carbapenemases. Also, we describe the emergence of NDM-1-producing E. cloacae in Lebanon. Screening for these isolates is necessary to limit the spread of resistant microorganisms in hospitals.

scholarly journals Multidrug-Resistant Sequence Type 235 Pseudomonas aeruginosa Clinical Isolates Producing IMP-26 with Increased Carbapenem-Hydrolyzing Activities in Vietnam

2016 ◽  
Vol 60 (11) ◽  
pp. 6853-6858 ◽  
Author(s):  
Tatsuya Tada ◽  
Pham Hong Nhung ◽  
Tohru Miyoshi-Akiyama ◽  
Kayo Shimada ◽  
Mitsuhiro Tsuchiya ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Multidrug Resistant ◽  
Sequence Type ◽  
Clinical Isolates ◽  
Medical Setting ◽  
Single Nucleotide ◽  
Content Type ◽  
E Coli ◽  
Whole Genomes ◽  
Encoding Genes

ABSTRACTForty clinical isolates of multidrug-resistantPseudomonas aeruginosawere obtained in a medical setting in Hanoi, Vietnam. Whole genomes of all 40 isolates were sequenced by MiSeq (Illumina), and phylogenic trees were constructed from the single nucleotide polymorphism concatemers. Of these 40 isolates, 24 (60.0%) harbored metallo-β-lactamase-encoding genes, includingblaIMP-15,blaIMP-26,blaIMP-51, and/orblaNDM-1. Of these 24 isolates, 12 harboredblaIMP-26and belonged to sequence type 235 (ST235).Escherichia coliexpressingblaIMP-26was significantly more resistant to doripenem and meropenem thanE. coliexpressingblaIMP-1andblaIMP-15. IMP-26 showed higher catalytic activity against doripenem and meropenem than IMP-1 and against all carbapenems tested, including doripenem, imipenem, meropenem, and panipenem, than did IMP-15. These data suggest that clinical isolates of multidrug-resistant ST235P. aeruginosaproducing IMP-26 with increased carbapenem-hydrolyzing activities are spreading in medical settings in Vietnam.

scholarly journals IMP-43 and IMP-44 Metallo-β-Lactamases with Increased Carbapenemase Activities in Multidrug-Resistant Pseudomonas aeruginosa

2013 ◽  
Vol 57 (9) ◽  
pp. 4427-4432 ◽  
Author(s):  
Tatsuya Tada ◽  
Tohru Miyoshi-Akiyama ◽  
Kayo Shimada ◽  
Masahiro Shimojima ◽  
Teruo Kirikae
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Amino Acid ◽  
Amino Acid Residue ◽  
Active Site ◽  
Multidrug Resistant ◽  
Amino Acid Sequences ◽  
Medical Settings ◽  
Content Type ◽  
E Coli ◽  
Catalytic Activities

ABSTRACTTwo novel IMP-type metallo-β-lactamase variants, IMP-43 and IMP-44, were identified in multidrug-resistantPseudomonas aeruginosaisolates obtained in medical settings in Japan. Analysis of their predicted amino acid sequences revealed that IMP-43 had an amino acid substitution (Val67Phe) compared with IMP-7 and that IMP-44 had two substitutions (Val67Phe and Phe87Ser) compared with IMP-11. The amino acid residue at position 67 is located at the end of a loop close to the active site, consisting of residues 60 to 66 in IMP-1, and the amino acid residue at position 87 forms a hydrophobic patch close to the active site with other amino acids. AnEscherichia colistrain expressingblaIMP-43was more resistant to doripenem and meropenem but not to imipenem than one expressingblaIMP-7. AnE. colistrain expressingblaIMP-44was more resistant to doripenem, imipenem and meropenem than one expressingblaIMP-11. IMP-43 had more efficient catalytic activities against all three carbapenems than IMP-7, indicating that the Val67Phe substitution contributed to increased catalytic activities against carbapenems. IMP-44 had more efficient catalytic activities against all carbapenems tested than IMP-11, as well as increased activities compared with IMP-43, indicating that both the Val67Phe and Phe87Ser substitutions contributed to increased catalytic activities against carbapenems.

scholarly journals Emergence of Metallo-β-Lactamase NDM-1-Producing Multidrug-Resistant Escherichia coli in Australia

2010 ◽  
Vol 54 (11) ◽  
pp. 4914-4916 ◽  
Author(s):  
Laurent Poirel ◽  
Emilie Lagrutta ◽  
Peter Taylor ◽  
Jeanette Pham ◽  
Patrice Nordmann
Keyword(s):  
Escherichia Coli ◽  
16S Rrna ◽  
Multidrug Resistant ◽  
E Coli ◽  
Extended Spectrum ◽  
High Level ◽  
Lactamase Gene

ABSTRACT A multidrug-resistant Escherichia coli isolate recovered in Australia produced a carbapenem-hydrolyzing β-lactamase. Molecular investigations revealed the first identification of the bla NDM-1 metallo-β-lactamase gene in that country. In addition, this E. coli isolate expressed the extended-spectrum β-lactamase CTX-M-15, together with two 16S rRNA methylases, namely, ArmA and RmtB, conferring a high level of resistance to aminoglycosides.

scholarly journals Recovering Escherichia coli Plasmids in the Absence of Long-Read Sequencing Data

2021 ◽  
Vol 9 (8) ◽  
pp. 1613
Author(s):  
Julian A. Paganini ◽  
Nienke L. Plantinga ◽  
Sergio Arredondo-Alonso ◽  
Rob J. L. Willems ◽  
Anita C. Schürch
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Multidrug Resistant ◽  
Special Focus ◽  
Valuable Insight ◽  
Sequencing Data ◽  
Extended Spectrum Beta Lactamase ◽  
E Coli ◽  
Long Read

The incidence of infections caused by multidrug-resistant E. coli strains has risen in the past years. Antibiotic resistance in E. coli is often mediated by acquisition and maintenance of plasmids. The study of E. coli plasmid epidemiology and genomics often requires long-read sequencing information, but recently a number of tools that allow plasmid prediction from short-read data have been developed. Here, we reviewed 25 available plasmid prediction tools and categorized them into binary plasmid/chromosome classification tools and plasmid reconstruction tools. We benchmarked six tools (MOB-suite, plasmidSPAdes, gplas, FishingForPlasmids, HyAsP and SCAPP) that aim to reliably reconstruct distinct plasmids, with a special focus on plasmids carrying antibiotic resistance genes (ARGs) such as extended-spectrum beta-lactamase genes. We found that two thirds (n = 425, 66.3%) of all plasmids were correctly reconstructed by at least one of the six tools, with a range of 92 (14.58%) to 317 (50.23%) correctly predicted plasmids. However, the majority of plasmids that carried antibiotic resistance genes (n = 85, 57.8%) could not be completely recovered as distinct plasmids by any of the tools. MOB-suite was the only tool that was able to correctly reconstruct the majority of plasmids (n = 317, 50.23%), and performed best at reconstructing large plasmids (n = 166, 46.37%) and ARG-plasmids (n = 41, 27.9%), but predictions frequently contained chromosome contamination (40%). In contrast, plasmidSPAdes reconstructed the highest fraction of plasmids smaller than 18 kbp (n = 168, 61.54%). Large ARG-plasmids, however, were frequently merged with sequences derived from distinct replicons. Available bioinformatic tools can provide valuable insight into E. coli plasmids, but also have important limitations. This work will serve as a guideline for selecting the most appropriate plasmid reconstruction tool for studies focusing on E. coli plasmids in the absence of long-read sequencing data.

scholarly journals Wild Boars Carry Extended-Spectrum β-Lactamase- and AmpC-Producing Escherichia coli

2021 ◽  
Vol 9 (2) ◽  
pp. 367 ◽  
Author(s):  
Anna R. Holtmann ◽  
Diana Meemken ◽  
Anja Müller ◽  
Diana Seinige ◽  
Kathrin Büttner ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multidrug Resistant ◽  
Phylogenetic Groups ◽  
Fecal Samples ◽  
Wild Boars ◽  
E Coli ◽  
Extended Spectrum ◽  
Nasal Swabs ◽  
Typing Methods ◽  
Lactamase Gene

Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA) represent major healthcare concerns. The role of wildlife in the epidemiology of these bacteria is unclear. The purpose of this study was to determine their prevalence in wild boars in Germany and to characterize individual isolates. A total of 375 fecal samples and 439 nasal swabs were screened for the presence of ESBL-/AmpC-E. coli and MRSA, respectively. The associations of seven demographic and anthropogenic variables with the occurrence of ESBL-/AmpC-E. coli were statistically evaluated. Collected isolates were subjected to antimicrobial susceptibility testing, molecular typing methods, and gene detection by PCR and genome sequencing. ESBL-/AmpC-E. coli were detected in 22 fecal samples (5.9%) whereas no MRSA were detected. The occurrence of ESBL-/AmpC-E. coli in wild boars was significantly and positively associated with human population density. Of the 22 E. coli, 19 were confirmed as ESBL-producers and carried genes belonging to blaCTX-M group 1 or blaSHV-12. The remaining three isolates carried the AmpC-β-lactamase gene blaCMY-2. Several isolates showed additional antimicrobial resistances. All four major phylogenetic groups were represented with group B1 being the most common. This study demonstrates that wild boars can serve as a reservoir for ESBL-/AmpC-producing and multidrug-resistant E. coli.

scholarly journals Epidemiology of Multidrug-resistant Bacteria Isolated from Hospitalized Patients in a Regional Central Hospital in China

2020 ◽  
Author(s):  
Jixun Zhang ◽  
Rui Li ◽  
Zhenzhong Liu ◽  
Chao Wang
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Acinetobacter Baumannii ◽  
Multidrug Resistant ◽  
Hospitalized Patients ◽  
Resistant Bacteria ◽  
Central Hospital ◽  
Gram Negative ◽  
E Coli ◽  
Significant Difference ◽  
The Impact

Abstract Objectives: Considering the dynamic changes of MDR, we did an up-to-date study and analyzed the impact of MDR on the outcome of patients. Design: Collected MDR isolated from hospitalized patients between June 2018 and May 2020 and performed retrospective analysis. Setting: This study was conducted in a public regional central hospital in China.Patients: 1156 patients with MDR infections.Results: Total 1291 MDRS were isolated, intensive care unit (ICU) accounted for 32.3% as the most. The main samples were sputum (75.1%) and 89.6% MDR were Gram-negative. The most common MDR were Acinetobacter baumannii, carbapenemase-producing K. pneumoniae, Pseudomonas aeruginosa, ESBL-producing E. coli. Methicillin-resistant Staphylococcus aureus (MRSA) and ESBL-producing K.pneumoniae. 35.6% were nosocomial infections and 64.4% were community-acquired infections. There was a statistically significant difference in mortality between patients infected with MDR and those with non-MDR (7.4% [32/432] vs 2.6% [17/655]; P = 0.001). The Acinetobacter baumannii and Klebsiella pneumoniae were mainly sensitive to tigecycline. The Pseudomonas aeruginosa was mainly sensitive to amikacin and levofloxacin. More than 80% of the Escherichia coli were sensitive to tigecycline and carbapenems. More than 90% of MRSA were sensitive to vancomycin, linezolid, and quinoprptin / daptoptin.Conclusions: The MDRS are mainly gram-negative bacteria. ICU contributes most MDR and pulmonary infection is the main origin of MDR. MDR infection is an independent risk factor for death. ESBL-producing Enterobacteriaceae, especially carbapenemase producing Enterobacteriaceae, should be paid more attention. This study is helpful to understand the distribution of MDR in hospital and the extent of antibiotic resistance.

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