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Optimization of the MALDIxin test for the rapid identification of colistin resistance in Klebsiella pneumoniae using MALDI-TOF MS.

ABSTRACT: BACKGROUND:With the dissemination of carbapenemase producers, a revival of colistin was observed for the treatment of infections caused by MDR Gram-negatives. Unfortunately, the increasing usage of colistin led to the emergence of resistance. In Klebsiella pneumoniae, colistin resistance arises through addition of 4-amino-l-arabinose (l-Ara4N) or phosphoethanolamine (pEtN) to the native lipid A. The underlying mechanisms involve numerous chromosome-encoded genes or the plasmid-encoded pEtN transferase MCR. Currently, detection of colistin resistance is time-consuming since it still relies on MIC determination by broth microdilution. Recently, a rapid diagnostic test based on MALDI-TOF MS detection of modified lipid A was developed (the MALDIxin test) and tested on Escherichia coli and Acinetobacter baumannii. OBJECTIVES:Optimize the MALDIxin test for the rapid detection of colistin resistance in K. pneumoniae. METHODS:This optimization consists of an additional mild-acid hydrolysis of 15?min in 1% acetic acid. The optimized method was tested on a collection of 81 clinical K. pneumoniae isolates, including 49 colistin-resistant isolates (45 with chromosome-encoded resistance, 3 with MCR-related resistance and 1 with both mechanisms). RESULTS:The optimized method allowed the rapid (<30?min) identification of l-Ara4N- and pEtN-modified lipid A of K. pneumoniae, which are known to be the real triggers of polymyxin resistance. At the same time, it discriminates between chromosome-encoded and MCR-related polymyxin resistance. CONCLUSIONS:The MALDIxin test has the potential to become an accurate tool for the rapid determination of colistin resistance in clinically relevant Gram-negative bacteria.

SUBMITTER: Dortet L

PROVIDER: S-EPMC6910190 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Optimization of the MALDIxin test for the rapid identification of colistin resistance in Klebsiella pneumoniae using MALDI-TOF MS.

Dortet Laurent L Broda Agnieszka A Bernabeu Sandrine S Glupczynski Youri Y Bogaerts Pierre P Bonnin Rémy R Naas Thierry T Filloux Alain A Larrouy-Maumus Gerald G

The Journal of antimicrobial chemotherapy 20200101 1

<h4>Background</h4>With the dissemination of carbapenemase producers, a revival of colistin was observed for the treatment of infections caused by MDR Gram-negatives. Unfortunately, the increasing usage of colistin led to the emergence of resistance. In Klebsiella pneumoniae, colistin resistance arises through addition of 4-amino-l-arabinose (l-Ara4N) or phosphoethanolamine (pEtN) to the native lipid A. The underlying mechanisms involve numerous chromosome-encoded genes or the plasmid-encoded pE ...[more]

PMID: 31580426

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Project description:BackgroundThis study aimed to evaluate antibiotic resistance genes and virulence genes and the clonal relationship of the carbapenem-nonsusceptible Klebsiella pneumoniae strains by molecular methods which are isolated from various clinical specimens from patients treated in tertiary care hospital in Turkey.MethodsIdentification of 32 carbapenem non-susceptible K. pneumoniae were determined by VITEK-2 (BioMérieux, France) automated system. Thirteen colistin-resistant strains were tested with the broth microdilution method. Various antibiotic resistance genes and virulence genes frequently seen in carbapenem-resistant strains were screened by PCR. Immunochromatographic tests used in the rapid diagnosis of carbapenemases were compared with PCR results. In addition, PFGE, MLST and MALDI-TOF MS methods were used to determine the clonal relationship among these strains.ResultsPCR demonstrated that 31 of the strains carried at least one of the carbapenemase genes. In one strain, the coexistence of blaOXA-48+NDM was shown. The most common resistance genes were determined as blaSHV (84.3%), blaCTX-M-1 (46.8%), blaOXA-48 (40.6%), blaKPC (40.6%), blaTEM (31.2%), blaNDM (18.8%) respectively. Among the virulence genes; magA (68.7%) was the most common, followed by kpn (59.3%) and K2 (9.3%). Immunochromatographic tests were found to be 100% compatible with PCR results. All colistin-resistant isolates were also found to be resistant by colistin broth microdilution. In PFGE analysis, 25 different genotypes were determined and clustering isolates were collected in 5 different clusters and the clustering rate was 35.4%. In MLST analysis, ST101 type was determined as the most common ST type with a rate of 29%. ST101 is followed by ST16, ST307, ST14, ST147, ST309, ST377, ST395 and ST2096, respectively. The compatibility rate between MALDI-TOF MS and VITEK-2 was found 94.3%, in bacterial identification. In MALDI-TOF MS typing, the maximum similarity between the strains was less than 70% and clustering not shown.ConclusionIn addition to OXA-48, which is endemic in our country, it has been determined that KPC, which is more common in the world, is becoming increasingly common in our region. ST101 type was determined as the most common type between the strains. To the best of our knowledge, this is the first study that compares these three methods in our country. There may be differences between bacterial identifications made with VITEK-2 and MALDI-TOF MS. In this study, it was observed that MALDI-TOF MS analyses were not compatible with the typing of strains according to PFGE and MLST analysis results.

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Optimization of the MALDIxin test for the rapid identification of colistin resistance in Klebsiella pneumoniae using MALDI-TOF MS.

Publications

Optimization of the MALDIxin test for the rapid identification of colistin resistance in Klebsiella pneumoniae using MALDI-TOF MS.

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