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 Antibiotic Resistance and Molecular Epidemiological Characteristics of Streptococcus agalactiae Isolated from Pregnant Women in Guangzhou, South China

2020 ◽  
Vol 2020 ◽  
pp. 1-11
Author(s):  
Zhaomin Cheng ◽  
Pinghua Qu ◽  
Peifeng Ke ◽  
Xiaohan Yang ◽  
Qiang Zhou ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Pregnant Women ◽  
Resistance Genes ◽  
Streptococcus Agalactiae ◽  
Virulence Genes ◽  
Mass Peak ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Epidemiological Characteristics ◽  
Tof Ms

Streptococcus agalactiae colonization in pregnant women can cause postpartum intrauterine infections and life-threatening neonatal infections. To formulate strategies for the prevention and treatment of S. agalactiae infections, we performed a comprehensive analysis of antibiotic resistance and a molecular-based epidemiological investigation of S. agalactiae in this study. Seventy-two S. agalactiae strains, collected from pregnant women, were subjected to antibiotic susceptibility tests; then, the screened erythromycin and clindamycin nonsusceptible isolates were used for macrolides and clindamycin resistance genes detection, respectively. Detection of resistance genes, serotyping, and determination of virulence genes were performed by polymerase chain reaction. The clonal relationships among the colonized strains were evaluated by multilocus sequence typing. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) mass peak analysis was performed to discriminate the specific sequence types (STs). In our study, 69.4% and 47.2% of the strains were nonsusceptible to erythromycin and clindamycin, respectively; the multidrug resistance rate was 66.7%. All erythromycin nonsusceptible strains harbored resistance genes, whereas only 52.9% of the clindamycin nonsusceptible strains possessed the linB gene. Erythromycin resistance was mainly mediated by the ermB or mefA/E genes. Four serotypes were identified, and the most common serotype was serotype III (52.8%), followed by Ib (22.2%), Ia (18.0%), and II (4.2%). All the strains were divided into 18 STs that were assigned to nine clonal complexes. Most of the major STs were distributed into specific serotypes, including ST19/serotype III, ST17/serotype III, ST485/serotype Ia, ST862/serotype III, and ST651/serotype III. Analysis of virulence genes yielded seven clusters, of which bca-cfb-scpB-lmb (61.6%) was the predominant virulence gene cluster. Among all ST strains distributed in this region, only the ST17 strains had a mass peak at 7620 Da. The outcomes of this study are beneficial for the epidemiological comparison of colonized S. agalactiae in different regions and may be helpful for developing the strategies for the prevention of S. agalactiae infection in Guangzhou. Furthermore, our results show that MALDI-TOF MS can be used for the rapid identification of the ST17 strains.

scholarly journals Direct detection of humoral marker corelates of COVID-19, glycated HSA and hyperglycosylated IgG3, by MALDI-ToF mass spectrometry.

2021 ◽  
Author(s):  
Raymond Kruse Iles ◽  
Jason Kruse Iles ◽  
Raminta Zmuidinaite ◽  
Anna Gardiner ◽  
Jonathan Lacey ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Respiratory Distress ◽  
Direct Detection ◽  
Plasma Samples ◽  
Total Blood ◽  
Laser Desorption Mass Spectrometry ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Immune Correlates ◽  
Tof Ms

The prefusion Spike protein of SARS-CoV2 binds advanced glycation end product (AGE) glycated human serum albumin (HSA) and a higher mass, hyperglycosylated/glycated, IgG3, as determined by matrix assisted laser desorption mass spectrometry (MALDI-ToF MS). We set out to investigate if the total blood plasma of patients who had recovered from acute respiratory distress as a result of COVID-19, contained more glycated HSA and higher mass (glycosylated/glycated) IgG3 than those with only clinically mild or asymptomatic infections. A direct dilution and disulphide bond reduction method was development and applied to plasma samples from SARS-CoV2 seronegative (N = 30) and seropositive (N = 31) healthcare workers and 38 convalescent plasma samples from patients who had been admitted with acute respiratory distress syndrome (ARDS) associated with COVID-19. Patients recovering from COVID-19 ARDS had significantly higher mass, AGE-glycated HSA and higher mass IgG3 levels. This would indicate that increased levels and/or ratios of hyper-glycosylation (probably terminal sialic acid) IgG3 and AGE glycated HSA may be predisposition markers for development of ARDS as a result of COVID-19 infection. Furthermore, rapid direct analysis of plasma samples by MALDI-ToF MS for such humoral immune correlates of COVID-19 presents a feasible screening technology for the most at risk; regardless of age or known health conditions.

scholarly journals Direct Detection and Identification of Bacterial Pathogens from Urine with Optimized Specimen Processing and Enhanced Testing Algorithm

2017 ◽  
Vol 55 (5) ◽  
pp. 1488-1495 ◽  
Author(s):  
Bin Huang ◽  
Lei Zhang ◽  
Weizheng Zhang ◽  
Kang Liao ◽  
Shihong Zhang ◽  
...  
Keyword(s):  
Predictive Value ◽  
Urine Culture ◽  
Direct Detection ◽  
Bacterial Pathogens ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Detection And Identification ◽  
Specimen Processing ◽  
Tract Infections ◽  
Tof Ms

ABSTRACT Rapid and accurate detection and identification of microbial pathogens causing urinary tract infections allow prompt and specific treatment. We optimized specimen processing to maximize the limit of detection (LOD) by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) and evaluated the capacity of combination of MALDI-TOF MS and urine analysis (UA) for direct detection and identification of bacterial pathogens from urine samples. The optimal volumes of processed urine, formic acid/acetonitrile, and supernatant spotted onto the target plate were 15 ml, 3 μl, and 3 μl, respectively, yielding a LOD of 1.0 × 10 5 CFU/ml. Among a total of 1,167 urine specimens collected from three hospital centers, 612 (52.4%) and 351 (30.1%) were, respectively, positive by UA and urine culture. Compared with a reference method comprised of urine culture and 16S rRNA gene sequencing, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of MALDI-TOF MS alone and MALDI-TOF MS coupled with UA were 86.6% versus 93.4% (χ 2 = 8.93; P < 0.01), 91.5% versus 96.3% (χ 2 = 7.06; P < 0.01), 81.5% versus 96.4% (χ 2 = 37.32; P < 0.01), and 94.1% versus 93.1% (χ 2 = 0.40; P > 0.05), respectively. No significant performance differences were revealed among the three sites, while specificity and NPV of MALDI-TOF MS for males were significantly higher than those for females (specificity, 94.3% versus 77.3%, χ 2 = 44.90, P < 0.01; NPV, 95.5% versus 86.1%, χ 2 = 18.85, P < 0.01). Our results indicated that the optimization of specimen processing significantly enhanced analytical sensitivity and that the combination of UA and MALDI-TOF MS provided an accurate and rapid detection and identification of bacterial pathogens directly from urine.

scholarly journals Conserved mass peaks in MALDI-TOF mass spectra of bacterial species at the genus and species levels

2018 ◽  
Author(s):  
Wenfa Ng
Keyword(s):  
Mass Spectrum ◽  
Mass Spectra ◽  
Mass Peak ◽  
Biological Basis ◽  
Maldi Tof Ms ◽  
Microbial Identification ◽  
Genus Level ◽  
Maldi Tof ◽  
Maldi Tof Mass Spectra ◽  
Tof Ms

Microbes are identified based on their distinguishing characteristics such as gene sequence or metabolic profile. Nucleic acid approaches such as 16S rRNA gene sequencing provide the gold standard method for microbial identification in the contemporary era. However, mass spectrometry-based microbial identification is gaining credence through ease of use, speed, and reliability. Specifically, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used in identifying bacteria, fungus, molds and archaea to the species level with high accuracy. The approach relies on the existence of unique mass spectrum fingerprint for individual microbial species. By comparing the mass spectrum of an unknown microbe with that catalogued in a reference database of known microorganisms, microbes could be identified through mass spectrum fingerprinting. However, the approach lacks fundamental biological basis given the relative difficulty in assigning specific protein to particular mass peak in the profiled mass spectrum, which hampers a deeper understanding of the mass spectrum obtained. This study seeks to examine the existence of conserved mass peaks in MALDI-TOF mass spectra of bacteria at the species and genus levels using open access data from SpectraBank. Results revealed that conserved mass peaks existed for all bacterial species examined. Large number of conserved mass peaks such as that of Escherichia coli and Morganella morganii suggested more closely-related strains of a species though functional annotation of the mass peaks is required to provide a deeper understanding of the mechanisms underlying the conservation of specific proteins. On the other hand, strains of Staphylococcus aureus and Pseudomonas putida had the least number of conserved mass peaks. Presence of conserved mass peaks in many genus provided further evidence that MALDI-TOF MS microbial identification had a biological basis in identification of microbial species to the genus level. In addition, it also highlighted that a subset of proteins could define the taxonomical boundary between the species and genus level. Finally, existence of only one conserved mass peak in Bacillus genus corroborated the difficulty of discriminating Bacillus species based on MALDI-TOF mass spectra. Similarly, no conserved mass peak at the genus level could be found for the Staphylococcus genus. Overall, existence of conserved mass peaks of bacteria at the species and genus levels provided evidence of a firm biological basis in the mass spectrum fingerprinting approach of MALDI-TOF MS microbial identification. This could help identify specific species in mass spectrum of single or multiple microbial species. Further functional annotation of the conserved mass peaks could illuminate in greater detail the biological mysteries of why certain proteins are conserved in specific genus and species.

scholarly journals An improved MALDI-TOF MS data analysis pipeline for the identification of carbapenemase-producing Klebsiella pneumoniae

2021 ◽  
Author(s):  
Eva Gato ◽  
Ignacio Pedro Constanso ◽  
Ana Candela ◽  
Fátima Galán ◽  
Bruno Kotska Rodiño-Janeiro ◽  
...  
Keyword(s):  
Data Analysis ◽  
Klebsiella Pneumoniae ◽  
Operating Procedure ◽  
Standard Operating Procedure ◽  
Resistant Bacteria ◽  
Analysis Pipeline ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Data Analysis Pipeline ◽  
Tof Ms

The increasing emergence of carbapenemase-producing Klebsiella pneumoniae (CPK) is a global health alarm. Rapid methods that require minimum sample preparation and rapid data analysis are urgently required. MALDI-TOF MS has recently been used by clinical laboratories for identification of antibiotic resistant bacteria; however, discrepancies have arisen regarding biological and technical issues. The aim of this study was to standardize an operating procedure and data analysis for identification of CPK by MALDI-TOF MS. To evaluate this approach, a series of 162 K. pneumoniae (112 CPK and 50 non CPK), were processed in the MALDI BioTyper system (Bruker Daltonik, Germany) following a standard operating procedure. The study was conducted in two stages, the first denominated the “Reproducibility stage” and the second, “CPK identification”. The first stage was designed to evaluate the biological and technical variation associated with the entire analysis of CPK and the second stage, to assess the final accuracy of MALDI-TOF for the identification of CPK. Therefore, we present an improved MALDI-TOF MS data analysis pipeline using neural network analysis implemented in Clover MS data analysis software (Clover Biosoft, Spain), that is designed to reduce variability, guarantee inter-laboratory reproducibility and maximize the information selected from the bacterial proteome. Using the Random Forest (RF) algorithm, 100% of CPK producing isolates were correctly identified when all the peaks in the spectra were selected as input features and TIC normalization was applied. Thus, we have demonstrated that real-time direct tracking of CPK is possible using MALDI-TOF MS.

scholarly journals Clinical Impact of MALDI-TOF MS Identification and Rapid Susceptibility Testing on Adequate Antimicrobial Treatment in Sepsis with Positive Blood Cultures

PLoS ONE ◽  
2016 ◽  
Vol 11 (5) ◽  
pp. e0156299 ◽  
Author(s):  
Alexia Verroken ◽  
Lydwine Defourny ◽  
Olivier le Polain de Waroux ◽  
Leïla Belkhir ◽  
Pierre-François Laterre ◽  
...  
Keyword(s):  
Susceptibility Testing ◽  
Antimicrobial Treatment ◽  
Blood Cultures ◽  
Clinical Impact ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Tof Ms
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