INVESTIGAREA REZISTENȚEI LA ANTIBIOTICE A UNOR TULPINI BACTERIENE MULTIREZISTENTE DIN IZOLATE CLINICE PRIN SPECTROMETRIE DE MASĂ MALDI TOF

Abstract

The objectives of the research theme/2022 were the characterization of susceptibility to antibiotics of some multidrug-resistant bacterial strains from clinical isolates by phenotypic methods and the optimization of the method for determining the antibiotic resistance of the bacteria under study by implementing the STAR-BL software module (Selective Testing of Antibiotic Resistance - ß-Lactamase) for MALDI Biotyper. Materials and methods. 22 multidrug-resistant bacterial strains isolated in the hospital (received within the framework of the collaboration with the Clinical Hospital for Emergency Plastic Surgery, Repair and Burns, Bucharest) were analyzed by MALDI TOF mass spectrometry for species identification. Based on the performed antibiotic resistance profiles, the enzymatic activity of these bacterial strains resistant to beta-lactam antibiotics was investigated by mass spectrometry.

Obiective. Spectrometria de masă MALDI TOF este o tehnologie cu aplicaţii multiple în diagnosticul microbiologic, controlul microorganismelor din colecţii, analiza relaţiilor taxonomice, controlul microbiologic al alimentelor şi apei etc. Obiectivele temei de cercetare/2022 au fost caracterizarea sensibilităţii la antibiotice a unor tulpini bacteriene polifarmacorezistente din izolate clinice prin metode fenotipice şi optimizarea metodei pentru determinarea rezistenţei la antibiotice a tulpinilor studiate prin implementarea modulului software STAR-BL (Selective Testing of Antibiotic Resistance - ß-Lactamase) pentru MALDI Biotyper.

Materiale şi metode. 22 de tulpini bacteriene multirezistente (primite în cadrul colaborării cu Spitalul Clinic de Urgenţă Chirurgie plastică Reparatorie şi Arsuri, Bucureşti) au fost analizate prin metode microbiologice clasice şi spectrometrie de masă MALDI TOF pentru identificarea speciei. Rezistenţa la antibiotice a fost determinată prin antibiogramă, metode rapide imunocromatografice (NG-Test® CARBA-5) şi spectrometrie de masă cu ajutorul kiturilor MBT STAR - Cepha kit şi MBT STAR - Carba kit.

Rezultate şi discuţii. Au fost validate prin spectometrie de masă 16 tulpini de Pseudomonas aeruginosa, 4 tulpini Acinetobacter baumannii (identificate primar de laboratorul spitalului) şi identificate 2 specii bacteriene (neidentificate de laboratorul spitalului) ca fiind Providencia stuartii. Pe baza profilurilor de antibiorezistenţă realizate a fost investigată activitatea enzimatică a acestor tulpini bacteriene rezistente la antibioticele beta-lactamice prin spectrometrie de masă. Rezultatele obţinute au confirmat prezenţa beta-lactamazelor la o parte din tulpinile analizate, prin detecţia unui domeniu de masă specific cuprins între 100-1000 Da, corespunzător hidrolizării grupării funcţionale a antibioticului.

Concluzii. Sistemul MALDI Biotyper echipat cu modulul MBT STAR-BL a permis identificarea speciilor bacteriene şi a rezistenţei la antibiotice mediate de ß-lactamaze. A fost instalat software-ul MBT STAR-BL şi au fost optimizate procedurile de lucru pentru detecţia activităţii cefalosporinazelor şi carbapenemazelor la tulpinile bacteriene investigate.

Sursa de finanţare: Planul Sectorial de Cercetare Dezvoltare al MApN.

Objectives. MALDI TOF mass spectrometry is a technology with multiple applications in microbiological diagnosis, control of microorganisms in collections, analysis of taxonomic relationships, microbiological control of food and water, etc. The objectives of the research theme/2022 were the characterization of susceptibility to antibiotics of some multidrug-resistant bacterial strains from clinical isolates by phenotypic methods and the optimization of the method for determining the antibiotic resistance of the bacteria under study by implementing the STAR-BL software module (Selective Testing of Antibiotic Resistance - ß-Lactamase) for MALDI Biotyper.

Materials and methods. 22 multidrug-resistant bacterial strains isolated in the hospital (received within the framework of the collaboration with the Clinical Hospital for Emergency Plastic Surgery, Repair and Burns, Bucharest) were analyzed by MALDI TOF mass spectrometry for species identification. Antibiotic resistance was assessed by classical microbiological methods, rapid immunochromatographic methods (NG-Test® CARBA-5) and mass spectrometry, using MBT STAR - Cepha kit and MBT STAR - Carba kit.

Results and discussion. Mass spectrometry allowed validation of 16 Pseudomonas aeruginosa and 4 Acinetobacter baumannii strains (primarily identified by the hospital laboratory) and identification of 2 bacterial species as being Providencia stuartii (which the laboratory could not recognize). Based on the performed antibiotic resistance profiles, the enzymatic activity of these bacterial strains resistant to beta-lactam antibiotics was investigated by mass spectrometry. The obtained results confirmed the presence of beta-lactamases in some of the analyzed strains, by the detection of a specific mass range between 100-1000 Da, corresponding to the hydrolysis of the functional group of the antibiotic.

Conclusions. The MALDI Biotyper system equipped with the MBT STAR-BL module allows both identification of bacterial species and ß-lactamase mediated antibiotic resistance. The installed MBT STAR-BL software optimized the detection of cephalosporinases and carbapenemases activity in the studied bacterial strains.

Funding source: R&D Program funded by the Ministry of Defence.