Variability in carbapenemase activity of intrinsic OxaAb (OXA-51-like) β-lactamase enzymes in Acinetobacter baumannii

Abstract Objectives To measure the variability in carbapenem susceptibility conferred by different OxaAb variants, characterize the molecular evolution of oxaAb and elucidate the contribution of OxaAb and other possible carbapenem resistance factors in the clinical isolates using WGS and LC–MS/MS. Methods Antimicrobial susceptibility tests were performed on 10 clinical Acinetobacter baumannii isolates. Carbapenem MICs were evaluated for all oxaAb variants cloned into A. baumannii CIP70.10 and BM4547, with and without their natural promoters. Molecular evolution analysis of the oxaAb variants was performed using FastTree and SplitsTree4. Resistance determinants were studied in the clinical isolates using WGS and LC–MS/MS. Results Only the OxaAb variants with I129L and L167V substitutions, OxaAb(82), OxaAb(83), OxaAb(107) and OxaAb(110) increased carbapenem MICs when expressed in susceptible A. baumannii backgrounds without an upstream IS element. Carbapenem resistance was conferred with the addition of their natural upstream ISAba1 promoter. LC–MS/MS analysis on the original clinical isolates confirmed overexpression of the four I129L and L167V variants. No other differences in expression levels of proteins commonly associated with carbapenem resistance were detected. Conclusions Elevated carbapenem MICs were observed by expression of OxaAb variants carrying clinically prevalent substitutions I129L and L167V. To drive carbapenem resistance, these variants required overexpression by their upstream ISAba1 promoter. This study clearly demonstrates that a combination of IS-driven overexpression of oxaAb and the presence of particular amino acid substitutions in the active site to improve carbapenem capture is key in conferring carbapenem resistance in A. baumannii and other mechanisms are not required.

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Variability in carbapenemase activity of intrinsic OxaAb (OXA-51-like) beta-lactamase enzymes in Acinetobacter baumannii

10.1101/2020.07.02.183822 ◽

2020 ◽

Cited By ~ 2

Author(s):

Yuiko Takebayashi ◽

Jacqueline Findlay ◽

Kate J. Heesom ◽

Philip J. Warburton ◽

Matthew B. Avison ◽

...

Keyword(s):

Molecular Evolution ◽

Efflux Pump ◽

Carbapenem Resistance ◽

Resistance Mechanisms ◽

Clinical Isolates ◽

Beta Lactamase ◽

Resistance Factors ◽

Evolution Analysis ◽

Ms Analysis ◽

Additional Resistance

ABSTRACTObjectivesThis study aimed to measure the variability in carbapenem susceptibility conferred by different OxaAb variants, characterise the molecular evolution of oxaAb and elucidate the contribution of OxaAb and other possible carbapenem resistance factors in the clinical isolates using WGS and LC-MS/MS.MethodsDisc susceptibility and MIC broth microdilution tests were performed on ten clinical A. baumannii isolates and interpreted according to CLSI guidelines. Imipenem and meropenem MICs were evaluated for all oxaAb variants cloned into susceptible A. baumannii CIP70.10 and increased adeABC efflux pump gene expression BM4547 backgrounds, with and without their natural promoters. Molecular evolution analysis of the oxaAb variants was performed using FastTree and SplitsTree4. Resistance determinants were studied in the clinical isolates using WGS and LC-MS/MS analysis.ResultsOxaAb(82), OxaAb(83), OxaAb(107), and OxaAb(110) carrying I129L and L167V substitutions increased carbapenem MICs when transferred into susceptible A. baumannii backgrounds without an upstream IS element. Carbapenem resistance was conferred with the addition of their natural upstream ISAba1 promoter. LC-MS/MS analysis on the original clinical isolates showed no differences in expression levels of proteins commonly associated with carbapenem resistance.ConclusionsISAba1-driven overexpression of OxaAb variants with substitutions I129L and L167V confers carbapenem resistance with no need for additional resistance mechanisms.

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DETECTION OF AMINOGLYCOSIDE AND QUINOLONE RESISTANCE GENES AND EVALUATION OF POLYMYXIN B SUSCEPTIBILITY PROFILE IN ACINETOBACTER BAUMANNII CLINICAL ISOLATES IN TEHRAN, IRAN DURING 2015-2016

Mediterranean Journal of Hematology and Infectious Diseases ◽

10.4084/mjhid.2018.044 ◽

2018 ◽

Vol 10 (1) ◽

pp. e2018044 ◽

Cited By ~ 5

Author(s):

Mohsen Heidary

Keyword(s):

Acinetobacter Baumannii ◽

Resistance Genes ◽

Antimicrobial Susceptibility ◽

Polymyxin B ◽

Quinolone Resistance ◽

Clinical Isolates ◽

Diffusion Method ◽

Clinical Samples ◽

Antimicrobial Susceptibility Tests ◽

Susceptibility Tests

ABSTRACTAcinetobacter baumannii is an important opportunistic pathogen, responsible for approximately 10% of all gram-negative nosocomial infection. The main aims of this study were to detect aminoglycoside and quinolone resistance genes among clinical isolates of A. baumannii and determine the antimicrobial susceptibility profiles. Current study was performed from February 2015 to April 2016, at two teaching hospitals. One-hundred A. baumannii isolates were collected from different clinical samples. Antimicrobial susceptibility tests were done by disk diffusion method according to CLSI guidelines. Detection of the qnrA, anrB, qnrS, aac(3)-IIa, and aac(6′)-Ib genes was done by PCR assay. The results of antibiotic susceptibility tests indicated that polymyxin B was the most effective drug against isolates of A. baumannii and the isolates were most resistant to cefepime (97%), ceftriaxone (95%), and amikacin (82%). The aac(3)-IIa, aac(6′)-Ib, and qnrA genes were found in 45%, 50%, and 50% of isolates, respectively. However, qnrB and qnrS genes could not be detected in any A. baumannii isolate.This study showed that there is a high level of resistance genes among clinical isolates of A. baumannii circulating in hospitals in Iran. This high prevalence rate highlights the necessity for establishing rapid diagnostic assays, more antimicrobial susceptibility tests, continuous antibiotic resistance monitoring.Keywords: Acinetobacter baumannii, Aminoglycoside, Quinolone, Iran

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Molecular Characterization of Carbapenem-Resistant Acinetobacter baumannii Clinical Isolates from Egyptian Patients

10.1101/2020.08.24.264911 ◽

2020 ◽

Author(s):

Reem M Hassan ◽

Sherifa T Salem ◽

Saly Ismail Mostafa Hassan ◽

Asmaa Sayed Hegab ◽

Yasmine S Elkholy

Keyword(s):

Acinetobacter Baumannii ◽

Molecular Characterization ◽

Antimicrobial Agents ◽

Treatment Options ◽

Carbapenem Resistance ◽

Clinical Isolates ◽

Common Mechanism ◽

Carbapenem Resistant ◽

Egyptian Patients ◽

Global Threat

AbstractAcinetobacter baumannii (A. baumannii) represents a global threat owing to its ability to resist most of the currently available antimicrobial agents. Moreover, emergence of carbapenem resistant A. baumannii (CR-AB) isolates limits the available treatment options. Enzymatic degradation by variety of ß-lactamases, have been identified as the most common mechanism of carbapenem resistance in A. baumannii. The alarming increase in the prevalence of CR-AB necessitates continuous screening and molecular characterization to appreciate the problem. The present study was performed to assess the prevalence and characterize carbapenemases among 206 CR-AB isolated from various clinical specimens collected from different intensive care units at Kasr Al-Aini Hospital.All isolates were confirmed to be A. baumannii by detection of the blaOXA-51-like gene. Molecular screening of 13 common Ambler class bla carbapenemases genes in addition to insertion sequence (IS-1) upstream OXA-23 was performed by using four sets of multiplex PCR, followed by identification using gene sequencing technology. Among the investigated genes, the prevalence of blaOXA-23, and blaOXA-58 were 77.7%, and 1.9%, respectively. The ISAba1 was detected in 10% of the blaOXA-23 positive isolates. The prevalence of metallo-β-lactamases (MBLs) studied; blaNDM-1, blaSPM, blaVIM, blaSIM-1 were 11.7%, 6.3%, 0.5%, and 0.5% respectively. One of class A; bla KPC was detected in 10.7% of the investigated isolates. blaOXA-24/40, blaIMP, blaGES, blaVEB and blaGIM were not detected in any of the studied isolates. Moreover, 18.4% of the isolates have shown to harbor two or more of the screened bla genes. We concluded that the most prevalent type of ß-lactamases genes among CR-AB isolates collected from Egyptian patients were blaOXA-23 followed by blaNDM-1 and blaKPC.Author summaryCarbapenem-resistant A. baumannii has become a real global health threat. The aim of the present study was to characterize and to assess the prevalence of carbapenemases among 206 CR-AB clinical isolates from Egyptian patients. We concluded that the most prevalent type of ß-lactamases genes among CR-AB isolates collected from Egyptian patients were blaOXA-23 followed by blaNDM-1 and blaKPC. In this study, ISAba1 was detected upstream 10% of blaOXA-23 positive isolates only which indicates that the spread of resistance among Acinetobacter isolates could be either chromosomal or plamid-mediated.

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In vitro activity of colistin against multidrug-resistant Acinetobacter baumannii isolates harboring blaOXA-23-like and blaOXA-24-like genes: A multicenter based study

Acta Microbiologica et Immunologica Hungarica ◽

10.1556/030.2020.01031 ◽

2020 ◽

Vol 67 (3) ◽

pp. 182-186

Author(s):

Susan Khanjani ◽

Hadi Sedigh Ebrahim-Saraie ◽

Mohammad Shenagari ◽

Ali Ashraf ◽

Ali Mojtahedi ◽

...

Keyword(s):

Acinetobacter Baumannii ◽

Carbapenem Resistance ◽

Multidrug Resistant ◽

Resistance Rate ◽

Clinical Isolates ◽

Clinical Samples ◽

Cross Sectional ◽

The North ◽

North Of Iran

AbstractThis study was aimed to evaluate occurrence of antibiotic resistance and the presence of resistance determinants among clinical isolates of Acinetobacter baumannii. This cross-sectional study from January to September 2018 was performed on 59 A. baumannii strains isolated from clinical samples in the north of Iran. Isolates were identified by standard microbiologic tests and molecular method. Antimicrobial susceptibility testing was carried out by disk diffusion and broth microdilution methods. The presence of carbapenem resistance genes was detected by PCR method. All isolates were resistant to cefepime, meropenem, imipenem and ceftazidime. The lowest resistance rate was observed against doxycycline with 33.9%. Minimum inhibitory concentration (MIC) results showed that all carbapenem-resistant A. baumannii (CRAB) isolates were susceptible to colistin with MIC50 and MIC90 values of 1/2 µg/mL. Among 59 CRAB, blaOXA-23-like was the most prevalent gene (86.4%) followed by blaOXA-24-like (69.5%). Meanwhile, none of the clinical isolates harbored blaOXA-58-like gene. We found a high prevalence of CRAB strains harboring OXA-type carbapenemases in the north of Iran. Our results suggests that the presence of OXA-type genes was not directly correlated with the increase of imipenem MIC level, but can be clinically important as they contribute to the selection of CRAB strains.

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Genomic Analysis of Carbapenem-Resistant Acinetobacter baumannii Isolates Belonging to Major Endemic Clones in South America

Frontiers in Microbiology ◽

10.3389/fmicb.2020.584603 ◽

2020 ◽

Vol 11 ◽

Author(s):

Carolina Silva Nodari ◽

Rodrigo Cayô ◽

Ana Paula Streling ◽

Felipe Lei ◽

Julia Wille ◽

...

Keyword(s):

South America ◽

Acinetobacter Baumannii ◽

Outer Membrane Proteins ◽

Acquired Resistance ◽

Clinical Isolates ◽

Efflux System ◽

Illumina Platform ◽

Synonymous Mutations ◽

Resistance Determinants ◽

Carbapenem Resistant

Carbapenem-resistant Acinetobacter baumannii (CRAB) are emerging worldwide. In South America, clinical isolates presenting such a phenotype usually do not belong to the globally distributed international clone 2 (IC2). The majority of these isolates are also resistant to multiple other antimicrobials and are often designated extremely drug-resistant (XDR). The aim of this study was to characterize the resistance mechanisms presented by 18 carbapenem-resistant A. baumannii isolates from five different Brazilian hospitals. Species identification was determined by rpoB sequencing, and antimicrobial susceptibility was determined by broth microdilution. Isolates were submitted to whole genome sequencing using Illumina platform and genetic similarity was determined by PFGE, MLST, and cgMLST. Genome analysis was used to identify intrinsic and acquired resistance determinants, including mutations in the AdeRSABC efflux system and in outer membrane proteins (OMPs). All isolates were identified as A. baumannii and grouped into 4 pulsotypes by PFGE, which belonged to clonal complexes (CC) 15Pas/103Ox (n = 4) and 79Pas/113Ox (n = 14), corresponding to IC4 and IC5, respectively. High MIC values to carbapenems, broad-spectrum cephalosporins, amikacin, and ciprofloxacin were observed in all isolates, while MICs of ampicillin/sulbactam, gentamicin, and tigecycline varied among the isolates. Minocycline was the most active antimicrobial agent tested. Moreover, 12 isolates (66.7%) were considered resistant to polymyxins. Besides intrinsic OXA-51 and ADC variants, all isolates harbored an acquired carbapenem-hydrolyzing class D β-lactamase (CHDL) encoding gene, either blaOXA–23 or blaOXA–72. A diversity of aminoglycoside modifying enzymes and resistance determinants to other antimicrobial classes were found, as well as mutations in gyrA and parC. Non-synonymous mutations have also been identified in the AdeRSABC efflux system and in most OMPs, but they were considered natural polymorphisms. Moreover, resistance to polymyxins among isolates belonging to IC5 were associated to non-synonymous mutations in pmrB, but no known polymyxin resistance mechanism was identified in isolates belonging to IC4. In conclusion, A. baumannii clinical isolates belonging to South America’s major clones present a myriad of antimicrobial resistance determinants. Special attention should be paid to natural polymorphisms observed in each clonal lineage, especially regarding non-synonymous mutations in constitutive genes associated with distinct resistance phenotypes.

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Novel Acquired Metallo-β-Lactamase Gene, blaSIM-1, in a Class 1 Integron from Acinetobacter baumannii Clinical Isolates from Korea

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.11.4485-4491.2005 ◽

2005 ◽

Vol 49 (11) ◽

pp. 4485-4491 ◽

Cited By ~ 218

Author(s):

Kyungwon Lee ◽

Jong Hwa Yum ◽

Dongeun Yong ◽

Hyuk Min Lee ◽

Heung Dong Kim ◽

...

Keyword(s):

Acinetobacter Baumannii ◽

Tertiary Care ◽

Carbapenem Resistance ◽

Gene Cassette ◽

Clinical Isolates ◽

Care Hospital ◽

Class 1 Integron ◽

Class 1 ◽

Broad Array ◽

Tract Infections

ABSTRACT Carbapenem resistance mediated by acquired carbapenemase genes has been increasingly reported, particularly for clinical isolates of Pseudomonas aeruginosa and Acinetobacter spp. Of 1,234 nonduplicate isolates of carbapenem-resistant Pseudomonas spp. and Acinetobacter spp. isolated at a tertiary-care hospital in Seoul, Korea, 211 (17%) were positive for metallo-β-lactamase (MBL). Of these, 204 (96%) had either the bla IMP-1 or bla VIM-2 allele. In addition, seven Acinetobacter baumannii isolates were found to have a novel MBL gene, which was designated bla SIM-1. The SIM-1 protein has a pI of 7.2, is a new member of subclass B1, and exhibits 64 to 69% identity with the IMP-type MBLs, which are its closest relatives. All SIM-1-producing isolates exhibited relatively low imipenem and meropenem MICs (8 to 16 μg/ml) and had a multidrug resistance phenotype. Expression of the cloned bla SIM-1 gene in Escherichia coli revealed that the encoded enzyme is capable of hydrolyzing a broad array of β-lactams, including penicillins, narrow- to expanded-spectrum cephalosporins, and carbapenems. The bla SIM-1 gene was carried on a gene cassette inserted into a class 1 integron, which included three additional cassettes (arr-3, catB3, and aadA1). The strains were isolated from sputum and urine specimens from patients with pneumonia and urinary tract infections, respectively. All patients had various underlying diseases. Pulsed-field gel electrophoresis of SmaI-digested genomic DNAs showed that the strains belonged to two different clonal lineages, indicating that horizontal transfer of this gene had occurred and suggesting the possibility of further spread of resistance in the future.

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