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Phenotypic and genotypic detection of metallo-beta-lactamases in Carbapenem resistant Acinetobacter baumannii.

ABSTRACT: Background:Carbapenem resistance in Acinetobacter baumannii has become a major concern for treating physicians. The aim of this study was to investigate the prevalence of metallo ?-lactamase (MBL) genes (bla VIM , and bla IMP) among isolated multidrug-resistant A. baumannii . Methods:Fifty non-repetitive carbapenem-resistant A. baumannii isolates were collected. Antibiotic susceptibility was performed by disk diffusion method. MICs were determined by E test method. The resistant strains were tested for the production of carbapenemases by the Modified Hodge Test (MHT) followed by EDTA-disk synergy test was performed for metallo-?-lactamases (MBL) phenotypic detection. Detection of bla VIM , and bla IMP was performed by PCR followed by sequencing. Results:All isolates had a multidrug resistant profile, and were all resistant to all antibiotics including the carbapenems but remained susceptible to colistin. Among these isolates, Carbapenemase production was confirmed by the Modified Hodge test for 42 (84%) isolates. Phenotypic method showed the production of MBL in 15 (30%) isolates. PCR techniques revealed that out of 50 isolates, 13 (26%) were positive for bla VIM and all were negative for bla IMP . Conclusion:Our study concludes that the high prevalence of carbapenem resistant Acinetobacter species with MBL production is one of the main concerns in our country and this situation needs strict infection control measures.

SUBMITTER: Moulana Z

PROVIDER: S-EPMC7265521 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Phenotypic and genotypic detection of metallo-beta-lactamases in Carbapenem resistant Acinetobacter baumannii.

Moulana Zahra Z Babazadeh Arefeh A Eslamdost Zohreh Z Shokri Mehran M Ebrahimpour Soheil S

Caspian journal of internal medicine 20200101 2

<h4>Background</h4>Carbapenem resistance in <i>Acinetobacter baumannii</i> has become a major concern for treating physicians. The aim of this study was to investigate the prevalence of metallo β-lactamase (MBL) genes (<i>bla</i> <sub>VIM</sub> <i>, and bla</i> <sub>IMP</sub>) among isolated multidrug-resistant <i>A. baumannii</i> .<h4>Methods</h4>Fifty non-repetitive carbapenem-resistant <i>A. baumannii</i> isolates were collected. Antibiotic susceptibility was performed by disk diffusion metho ...[more]

PMID: 32509245

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Project description:Background:Antibiotic use is largely under-regulated in Egypt leading to the emergence of resistant isolates. Carbapenems are last resort agents to treat Acinetobacter baumannii infections resistant to other classes of antibiotics. However, carbapenem-resistant isolates are emerging at an alarming rate. This study aimed at phenotypically and molecularly characterizing seventy four carbapenem-unsusceptible A. baumannii isolates from Egypt to detect the different enzymes responsible for carbapenem resistance. Methods:Carbapenemase production was assessed by a number of phenotypic methods: modified Hodge test (MHT), carbapenem inactivation method (CIM), combined disc test (CDT), CarbAcineto NP test and boronic acid disc test. Polymerase chain reaction (PCR) was used to screen the isolates for the presence of some genes responsible for resistance to carbapenems, as well as some insertion sequences. Results:PCR amplification of class D carbapenemases revealed the prevalence of bla OXA-51 and bla OXA-23 in 100% of the isolates and of bla OXA-58 in only one isolate (1.4%). bla VIM and bla NDM-1 belonging to class B metallo-?-lactamases were present in 100 and 12.1% of the isolates, respectively. The prevalence of ISAba1, ISAba2 and ISAba3 was 100, 2.7 and 4.1%, respectively. None of the tested isolates carried bla OXA-40 , bla IMP , bla SIM , bla SPM , bla GIM or the class A bla KPC . Taking PCR as the gold standard method for the detection of different carbapenemases, the sensitivities of the MHT, CIM, CDT, CarbAcineto NP test and boronic acid disc/imipenem or meropenem test for this particular collection of isolates were 78.4, 68.9, 79.7, 95.9, and 56.8% or 70.3%, respectively. Conclusions:The widespread detection of carbapenem-resistant A. baumannii (CR-AB) has become a real threat to the efficacy of treatment regimens. Among the studied cohort of CR-AB clinical isolates, bla OXA-51 , bla OXA-23 and bla VIM were the most prevalent, followed by bla NDM-1 and bla OXA-58 . The genotypic detection of carbapenemases among CR-AB clinical isolates using PCR was most conclusive, followed closely by the phenotypic testing using CarbAcineto NP test.

Project description:(1) Background: Acinetobacter baumannii is well known as a causative agent of severe hospital-acquired infections, especially in intensive care units. The present study characterised the genetic traits of biofilm-forming carbapenem-resistant A. baumannii (CRAB) clinical isolates. Additionally, this study determined the prevalence of biofilm-producing A. baumannii isolates from a tertiary care hospital and investigated the association of biofilms with the distribution of biofilm-related and antibiotic resistance-associated genotypes. (2) Methods: The 995 non-duplicate A. baumannii isolates were identified, and their susceptibilities to different antibiotics were determined using the disk diffusion method. Using the modified microtiter plate assay, the CRAB isolates were investigated for their biofilm formation ability. Hemolysin and protease activities were determined. CRABs were subjected to polymerase chain reaction (PCR) assays targeting blaVIM, blaNDM, blaIMP, blaOXA-23-like, blaOXA-24-like, blaOXA-51-like, csuE and pgaB genes. Individual CRAB isolates were identified for their DNA fingerprint by repetitive element sequence-based (REP)-PCR. (3) Results: Among all A. baumannii isolates, 172 CRABs were identified. The major antibiotic resistance gene among the CRAB isolates was blaOXA-51-like (100%). Ninety-nine isolates (57.56%) were biofilm producers. The most prevalent biofilm gene was pgaB (79.65%), followed by csuE (76.74%). Evidence of virulence phenotypes revealed that all CRAB exhibited proteolytic activity; however, only four isolates (2.33%) were positive for the hemolytic-producing phenotype. REP-PCR showed that 172 CRAB isolates can be divided into 36-DNA fingerprint patterns. (4) Conclusions: The predominance of biofilm-producing CRAB isolates identified in this study is concerning. The characterisation of risk factors could aid in controlling the continual selection and spreading of the A. baumannii phenotype in hospitals, thereby improving patient care quality.

Project description:BackgroundThe rapid emergence of carbapenem resistant Acinetobacter baumannii (CRAB) has resulted in an alarming situation worldwide. Realizing the dearth of literature on susceptibility of CRAB in genetic context in the developing region, this study was performed to determine the susceptibility profile against standard drugs/combinations and the association of in-vitro drug synergy with the prevalent molecular determinants.Methods and findingsA total of 356 clinical isolates of A. baumannii were studied. Confirmation of the isolates was done by amplifying recA and ITS region genes. Susceptibility against standard drugs was tested by Kirby Bauer disc diffusion. Minimum inhibitory concentration (MIC), MIC50 and MIC90 values against imipenem, meropenem, doripenem, ampicillin/sulbactam, minocycline, amikacin, polymyxin B, colistin and tigecycline was tested as per guidelines. Genes encoding enzymes classes A (bla GES, bla IMI/NMC-A, bla SME, bla KPC), B (bla IMP, bla VIM, bla NDM) and D (bla OXA-51, bla OXA-23 and bla OXA-58) were detected by multiplex polymerase chain reaction. Synergy against meropenem-sulbactam and meropenem-colistin combinations was done by checkerboard MIC method. Correlation of drug synergy and carbapenemase encoding genes was statistically analyzed.ResultsOf the total, resistance above 90% was noted against gentamicin, ciprofloxacin, levofloxacin, ceftazidime, cefepime, ceftriaxone, cotrimoxazole and piperacillin/tazobactam. By MIC, resistance rates from highest to lowest was seen against imipenem 89.04% (n=317), amikacin 80.33% (n=286), meropenem 79.49% (n=283), doripenem 77.80% (n=277), ampicillin/sulbactam 71.62% (n=255), tigecycline 55.61% (n=198), minocycline 14.04% (n=50), polymyxin B 10.11% (n=36), and colistin 2.52% (n=9). CRAB was 317 (89.04%), 81.46% (n=290) were multidrug resistant and 13.48% (n=48) were extensively drug resistant. All the CRAB isolates harboured bla OXA-51 gene (100%) and 94% (n=298) bla OXA-23 gene. The bla IMP gene was most prevalent 70.03% (n=222) followed by bla NDM, 59.62% (n=189). Majority (87.69%, 278) were co-producers of classes D and B carbapenemases, bla OXA-23 with bla IMP and bla NDM being the commonest. Synergy with meropenem-sulbactam and meropenem-colistin was 47% and 57% respectively. Reduced synergy (p= <0.0001) was noted for those harbouring bla OXA-51+blaOXA-23with bla NDM gene alone or co-producers.ConclusionPresence of bla NDM gene was a significant cause of synergy loss in meropenem-sulbactam and meropenem-colistin. In bla NDM endemic regions, tigecycline, minocycline and polymyxins could be viable options against CRAB isolates with more than one carbapenemase encoding genes.

Project description:BackgroundWith an increase in the use of colistin methansulfonate (CMS) to treat carbapenem-resistant Acinetobacter baumannii infections, colistin resistance is emerging.MethodsPatients with infection or colonization due to colistin-resistant A. baumannii were identified at a hospital system in Pennsylvania. Clinical data were collected from electronic medical records. Susceptibility testing, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST) were performed. To investigate the mechanism of colistin resistance, lipid A was subjected to matrix-assisted laser desorption/ionization mass spectrometry.ResultsTwenty patients with colistin-resistant A. baumannii were identified. Ventilator-associated pneumonia was the most common type of infection. Nineteen patients had received intravenous and/or inhaled CMS for treatment of carbapenem-resistant, colistin-susceptible A. baumannii infection prior to identification of colistin-resistant isolates. The 30-day all-cause mortality rate was 30%. The treatment regimen for colistin-resistant A. baumannii infection associated with the lowest mortality rate was a combination of CMS, a carbapenem, and ampicillin-sulbactam. The colistin-susceptible and -resistant isolates from the same patients were highly related by PFGE, but isolates from different patients were not, suggesting evolution of resistance during CMS therapy. By MLST, all isolates belonged to the international clone II, the lineage that is epidemic worldwide. Phosphoethanolamine modification of lipid A was present in all colistin-resistant A. baumannii isolates.ConclusionsColistin-resistant A. baumannii occurred almost exclusively among patients who had received CMS for treatment of carbapenem-resistant, colistin-susceptible A. baumannii infection. Lipid A modification by the addition of phosphoethanolamine accounted for colistin resistance. Susceptibility testing for colistin should be considered for A. baumannii identified from CMS-experienced patients.

Dataset Information

Phenotypic and genotypic detection of metallo-beta-lactamases in Carbapenem resistant Acinetobacter baumannii.

Publications

Phenotypic and genotypic detection of metallo-beta-lactamases in Carbapenem resistant Acinetobacter baumannii.

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