Evaluation of Carbapenem Resistance Enterobacteriaceae(CRE) detection by Matrix Assisted Laser Desorption / Ionization-Time of Flight mass spectrometry (MALDI-TOF)

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Evaluation of Carbapenem Resistance Enterobacteriaceae(CRE) detection by Matrix Assisted Laser Desorption / Ionization-Time of Flight mass spectrometry (MALDI-TOF)


Author: Cheung, Kwok Chi
Title: Evaluation of Carbapenem Resistance Enterobacteriaceae(CRE) detection by Matrix Assisted Laser Desorption / Ionization-Time of Flight mass spectrometry (MALDI-TOF)
Degree: M.Sc.
Year: 2015
Subject: Anti-infective agents.
Communicable diseases -- Diagnosis.
Communicable diseases -- Treatment.
Hong Kong Polytechnic University -- Dissertations
Department: Dept. of Health Technology and Informatics
Pages: xiv, 93 pages : illustrations
Language: English
InnoPac Record: http://library.polyu.edu.hk/record=b2811878
URI: http://theses.lib.polyu.edu.hk/handle/200/7999
Abstract: Introduction: Enterobacteriaceae represents a group of gram negative bacilli that is common in infection. It infect various sites that including severe infections such as sepsis and Central Nervous System(CNS) infection. Enterobacteriaceae has been developing different mechanisms to survive from antibiotics, and finally able to survive from one of the most advanced antibiotic, carbapenem. Carbapenem is cidal both gram negative and positive organisms effectively. Carbapenem resistance makes therapy of Enterobacteriaceae more challenging. Furthermore, carbapenemase production can be transmitted within Enterobacteriaceae through plasmid exchange. Therefore, rapid detection and infection control of Carbapenem Resistance Enterobacteriaceae(CRE) is essential for therapy management. Sensitivity test based detection is commonly performed in routine laboratory, but turnaround time is its limitation. Polymerase Chain Reaction(PCR) based method is a rapid detection of CRE now, but it also have its weakness Thus, a novel detection of CRE is needed. Matrix Assisted Laser Desorption / Ionization-Time of Flight mass spectrometry (MALDI-TOF) has been implanting into diagnostic laboratory for bacterial identification for several years. Now it takes a step forward to provide rapid detection in antibiotic resistance including carbapenem. In the study, MALDI-TOF BL-STAR detection was evaluated as a rapid CRE detection method, in compare with PCR and disc diffusion test. Aims: In the study, a new version of MALDI-TOF firmware developed by Bruker database, MALDI-TOF BL-STAR, was used for detecting CRE. This firmware has manufacture provided setting for CRE detection. Optimization and evaluation of CRE detection were performed and finally compare results with PCR and disc diffusion based CRE detection. The comparison can estimate can BL-STAR serve as a rapid CRE detection in routine laboratory.
Methods: 33 CRE positive clinical strains, and 12 carbapenem susceptible clinical strains were involved in the study. All were subcultured on blood agar for 24 hours incubation in 35℃, 0.5% CO2. CRE confirmation of those strains were done by combine disc method; disc diffusion method; PCR detection of blaVIM, blaKPC, blaIMP, blaNDM and blaOXA-48-like genes. Followed by MALDI-TOF BL-STAR detection using 4mg/mL Meropenem in H2O for 2 hours incubation for comparison. Results: All CRE stains were not susceptible to Ertapenem by disc diffusion. 30 CRE strains were positive for combined disc test, and 23 out of these 30 CRE strains were bla positive strains. All susceptible strains were negative for both tests. MALDI-TOF BL-STAR detection detected 21 out of 23 carbapenemase producing CRE successfully. The sensitivity was 91.3% and specificity was 100% in compare with PCR detection. 2 undetected CRE were OXA-48-like carbapenemase, and MALDI-TOF BL-STAR detection interpreted the result as unconcluded. Extended incubation time solved the problem. In compare with CDT, sensitivity was 70% and specificity was 100%. 9 CDT positive strains were not detected by BL-STAR detection. Conclusion: MALDI-TOF BL-STAR detection showed good performance in detecting carbapenemase producing CRE in technical mean, with high sensitivity and specificity. Also the analysis time was around 3 hours in current setting; easy handling of test and affordable running cost will also benefit to routine diagnostic laboratory. However, improvement in OXA-48-like carbapenemase detection and stability of software are suggested in the further study.

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