scholarly journals Carbapenemase Producing Klebsiella pneumoniae (KPC): What Is the Best MALDI-TOF MS Detection Method

Antibiotics ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1549
Author(s):  
Lukáš Hleba ◽  
Miroslava Hlebová ◽  
Anton Kováčik ◽  
Juraj Čuboň ◽  
Juraj Medo
Keyword(s):  
Mass Spectrum ◽  
Klebsiella Pneumoniae ◽  
Direct Detection ◽  
Degradation Products ◽  
Hospital Setting ◽  
Morbidity And Mortality ◽  
Klebsiella Pneumoniae Carbapenemase ◽  
The Third ◽  
Maldi Tof ◽  
Positive Mode

Klebsiella pneumoniae carbapenemase (KPC)-producing bacteria is a group of highly dangerous antibiotic resistant Gram-negative Enterobacteriaceae. They cause infections associated with significant morbidity and mortality. Therefore, the rapid detection of KPC-producing bacteria plays a key role in clinical microbiology. Matrix assisted laser desorption/ionization time-of- flight (MALDI-TOF) is a rapidly evolving technology that finds application in various clinical, scientific, and industrial disciplines. In the present study, we demonstrated three different procedures of carbapenemase-producing K. pneumoniae (KPC) detection. The most basic model of MALDI-TOF instrument MS Microflex LT was used, operating in the linear ion-positive mode, commonly used in modern clinical laboratories. The first procedure was based on indirect monitoring of carbapenemase production with direct detection of hydrolyzed carbapenem antibiotic degradation products in the mass spectrum. The second procedure was based on direct detection of blaKPC accompanying peak with an 11,109 Da in the mass spectrum of carbapenemase-producing K. pneumoniae (KPC), which represents the cleaved protein (pKpQIL_p019) expressed by pKpQIL plasmid. In addition, several unique peaks were detected in the carbapenemase-producing K. pneumoniae (KPC) mass spectrum. The third procedure was the identification of carbapenemase-producing K. pneumoniae (KPC) based on the protein fingerprint using local database created from the whole mass spectra. By comparing detection procedures, we determined that the third procedure was very fast and relatively easy. However, it requires previous verification of carbapenemase-producing K. pneumoniae (KPC) using other methods as genetic blaKPC identification, detection of carbapenem degradation products, and accompanying peak with 11,109 Da, which represents cleaved pKpQIL_p019 protein expressed by pKpQIL plasmid. Detection of carbapenemase-producing K. pneumoniae using MALDI-TOF provides fast and accurate results that may help to reduce morbidity and mortality in hospital setting when applied in diagnostic situations.

scholarly journals Occurrence of the p019 gene in the blaKPC-harbouring plasmids: adverse clinical impact for direct tracking of KPC-producing Klebsiella pneumoniae by MALDI-TOF MS

2021 ◽  
Author(s):  
Eva Gato ◽  
Ignacio Pedro Constanso ◽  
Bruno Kotska Rodiño-Janeiro ◽  
Paula Guijarro-Sánchez ◽  
Tyler Alioto ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Computational Analysis ◽  
Direct Detection ◽  
Clinical Impact ◽  
Mass Peak ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Direct Tracking ◽  
Geographical Locations ◽  
Tof Ms

MALDI-TOF MS has recently been used for the direct detection of KPC-producing isolates by analysis of the 11,109 Da mass peak representing the P019 protein. In this study we evaluate the presence of the 11,109 Da mass peak in a collection of 435 unduplicated K. pneumoniae clinical isolates. The prevalence of the P019 peak in the blaKPC K. pneumoniae isolates was 49.2% (32/65). The 11,109 Da mass peak was not observed in any of the other carbapenemase (319) or non carbapenemase producers (116). Computational analysis of the presence of the p019 gene was performed in the aforementioned carbapenemase-producing K. pneumoniae isolates fully characterized by WGS and in a further collection of 1,649 K. pneumoniae genomes included in EuSCAPE. Herein, we have demonstrated that the p019 gene is not exclusively linked to the pKpQil plasmid, but it is present in the following plasmids: IncFIB(K)/IncFII(K)/ColRNAI, IncFIB(pQil), IncFIB(pQil)/ColRNAI, IncFIB(pQil)/IncFII(K), IncFIB(K)/IncFII(K) and IncX3. Besides, we have proven the independent movement of the Tn4401 and the ISKpn31, of which the p019 gene is a component. The absence of the p019 gene was obvious in Col440I, Col(pHAD28), IncFIB(K)/IncX3/IncFII(K), IncFIB(K)/IncFII(K) plasmids. In addition, we also observed another plasmid in which neither Tn4401 nor ISKpn31 was found, IncP6. In the EuSCAPE, the occurrence of p019 varied from 0% to 100% among the different geographical locations. The adverse clinical impact of the diminished prevalence of the p019 gene within the plasmid encoding KPC-producing Klebsiella pneumoniae puts forward the need for reconsideration when applying this technique in a clinical setting.

scholarly journals 536. Challenges in Using MALDI-TOF Technology to Assess for KPC Resistance in Klebsiella pneumoniae isolates in America

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S256-S257
Author(s):  
Michael Christopher Thompson ◽  
David Banach ◽  
Christina Nishimura ◽  
Anthony Muyombwe
Keyword(s):  
Klebsiella Pneumoniae ◽  
Protein Extraction ◽  
High Sensitivity ◽  
Rapid Identification ◽  
Health Concern ◽  
Ionization Time ◽  
Klebsiella Pneumoniae Carbapenemase ◽  
Maldi Tof ◽  
Control And Prevention ◽  
Low Sensitivity

Abstract Background The emergence of Klebsiella pneumoniae Carbapenemase-producing Enterobacteriaceae (KPC-E) has created a major public health concern. In clinical practice, rapid identification of KPC-KP has important implications for clinical management and infection control. In some settings matrix-assisted laser desorption ionization time of flight (MALDI-TOF) software has been used for rapid detection of KPC producing K. pneumoniae with high sensitivity and specificity. Genomic sequencing has determined that the 11.09 m/z peak is related to protein expression from the P109 gene found mostly in Tna4401a isoform among KPC-E. In our study, we evaluated the use of MALDI-TOF automated detection software to evaluate for KPC detection among a diverse group of KPC-E isolates. Methods We tested 52 KPC-E isolates from various hospitals in Connecticut and the Centers for Disease Control and Prevention (CDC) Antibiotic Resistance (AR) Bank. All specimens were verified as KPC-producing strains by detection of the blaKPCgene through polymerase chain reaction. Protein extraction using the standard extraction method was performed on sub-cultured isolates. Each isolate was tested three times on MALDI-TOF MS with the incorporated bio-subtype KPC module. An organism confidence or log score value of 2 or higher was considered valid. Results Among 52 tested KPC-K. pneumoniae isolates, 44 (85%) were from various hospitals in Connecticut, eight (15%) came from the AR Bank. Only 15 (25.1%) of the isolates were detected as KPC-producing using the MALDI-TOF KPC module. Further investigation by peak analysis confirmed all 15 isolates detected positive demonstrated a peak at 11.09 m/z. The 11.09 m/z peak was not found in the 37 specimens that were not detected. Conclusion The results from our study suggest low sensitivity using this software and contradicts results seen in previous European studies. The Tna4401a isoform is often seen in KPC-2 strains, which may be less prevalent in our sample of isolates, explaining the poor sensitivity of MALDI-TOF. Further study is needed to explore this finding and potential opportunities for MALDI-TOF for rapid identification of KPC-KP. Disclosures All authors: No reported disclosures.

scholarly journals Direct detection of intact Klebsiella pneumoniae carbapenemase variants from cell lysates: Identification, characterization and clinical implications

2020 ◽  
Vol 17 ◽  
pp. 12-21
Author(s):  
William M. McGee ◽  
Matthew L. Faron ◽  
Jason R. Neil ◽  
Scott R. Kronewitter ◽  
Blake W. Buchan ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Direct Detection ◽  
Clinical Implications ◽  
Cell Lysates ◽  
Klebsiella Pneumoniae Carbapenemase

scholarly journals 12. Evaluation of Rapid Phenotypic Testing for KPC Carbapenemase Producing klebsiella Pneumoniae Directly from Positive Blood Cultures by Use of “Hot Chocolate” Plates

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S7-S7
Author(s):  
Alexander Lawandi ◽  
Gleice C Leite ◽  
Brigitte Lefebvre ◽  
Jean Longtin ◽  
Todd C Lee
Keyword(s):  
Klebsiella Pneumoniae ◽  
Carbapenem Resistance ◽  
Empiric Therapy ◽  
Turnaround Time ◽  
Rapid Identification ◽  
Blood Cultures ◽  
Minimal Growth ◽  
Maldi Tof ◽  
Considerable Morbidity ◽  
Hot Chocolate

Abstract Background Invasive infections with Carbapenemase Producing Enterobacterales are associated with considerable morbidity and mortality, in part due to the risk of inappropriate empiric therapy. Consequently, the rapid identification of carbapenem resistance is crucial to the management of these infections. We sought to evaluate possible reductions in turnaround time to identification of this resistance in blood cultures growing these organisms by applying rapid phenotypic test kits to growth from “hot chocolate” plates. Methods 30 blood cultures, spiked with carbapenem resistant Klebsiella pneumoniae isolates or susceptible controls, were inoculated onto chocolate agars that had pre-warmed at 37°C. These plates were incubated at 37ºC for 3.5 hours. The resulting minimal growth was then identified using MALDI-TOF and underwent rapid phenotypic testing using three commercially available products (β-lacta and β-carba, from Bio-Rad, Marnes-la-Coquette, France, and Carba-NP, from bioMérieux, Durham, NC). The time to identification of carbapenem resistance using this method was then compared to that of the conventional laboratory workup. Results The identification was 100% accurate to the species level using MALDI-TOF paired to the 3.5 hour growth on the “hot choocolate” plates. The β-lacta kit identified resistance to 3rd generation cephalosporins for all ESBL and carbapenemase producing Klebsiella pneumoniae isolates, while the β-carba and Carba-NP kits identified carbapenem resistance only in the carbapenemase producers. The sensitivity of all assays was 100% (95% CI 0.87–1.0) and the specificity of carbapenemase detection was 100% (97.5% one-sided CI 0.4–1.0). The corresponding sensitivities and specificities of direct disc diffusion for ertapenem resistance detection were 88.5% (95% CI 0.70–0.98) and 100% (95%CI 0.40–1.0) respectively. The turnaround time for the rapid kits coupled to the “hot chocolate” plates was 4.25 to 5.1 hours as compared to 16 hours for the conventional workup. Conclusion Rapid phenotypic tests performed after inoculation of “hot chocolate” plates are highly sensitive for the presence of carbapenemase production and can be incorporated into the laboratory workflow for Klebisella pneumoniae with important reductions in turnaround time. Disclosures All Authors: No reported disclosures

scholarly journals Meropenem-vaborbactam as salvage therapy for ceftazidime-avibactam-, cefiderocol-resistant ST-512 Klebsiella pneumoniae producing KPC-31, a D179Y variant of KPC-3

2021 ◽  
Author(s):  
Giusy Tiseo ◽  
Marco Falcone ◽  
Alessandro Leonildi ◽  
Cesira Giordano ◽  
Simona Barnini ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Salvage Therapy ◽  
Target Region ◽  
Klebsiella Pneumoniae Carbapenemase ◽  
Variant Enzyme

Abstract A 68-year-old man had recurrent bacteremia by Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae resistant to ceftazidime-avibactam and cefiderocol. The sequencing of a target region showed that it harbored a KPC-3 variant enzyme (D179Y; KPC-31), which confers resistance to ceftazidime-avibactam and restores meropenem susceptibility. The patient was successfully treated with meropenem-vaborbactam.

A multi-institutional outbreak of New Delhi metallo-β-lactamase–producing Escherichia coli with subsequent acquisition of the Klebsiella pneumoniae carbapenemase gene

2020 ◽  
pp. 1-4
Author(s):  
Ester Solter ◽  
Jason C. Kwong ◽  
Aaron Walton ◽  
Norelle Sherry ◽  
Benjamin P. Howden ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Common Ancestor ◽  
Sequence Type ◽  
Second Phase ◽  
New Delhi ◽  
Genetic Sequencing ◽  
Klebsiella Pneumoniae Carbapenemase ◽  
Carbapenemase Gene

Abstract We characterized 57 isolates from a 2-phase clonal outbreak of New Delhi metallo-β-lactamase–producing Eschericha coli, involving 9 Israeli hospitals; all but 1 isolate belonged to sequence-type (ST) 410. Most isolates in the second phase harbored blaKPC-2 in addition to blaNDM-5. Genetic sequencing revealed most dual-carbapenemase–producing isolates to be monophyletically derived from a common ancestor.

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