The opportunistic pathogen Candida parapsilosis is a major causative agent of candidiasis leading to death in immunocompromised individuals. Azoles are the first line of defense in their treatment. The purpose of this study was to characterize eight fluconazole-resistant and sensitive C. parapsilosis hospital isolates through a battery of phenotypic tests that target pathogenicity attributes such as virulence, biofilm formation, stress resistance, and ergosterol content. Whole genome sequencing was carried out to identify mutations in key pathogenicity and resistance genes. Phylogenetic comparison was performed to determine strain relatedness and clonality. Genomic data and phylogenetic analysis revealed that two isolates were C. orthopsilosis and C. metapsilosis misidentified as C. parapsilosis. Whole genome sequencing analysis revealed known and novel mutations in key drug resistance and pathogenicity genes such as ALS6, ALS7, SAPP3, SAP7, SAP9, CDR1, ERG6, ERG11 and UPC2. Phylogenetic analysis revealed a high degree of relatedness and clonality within our C. parapsilosis isolates. Our results showed that resistant isolates exhibited an increase in biofilm content compared to the sensitive isolates. In conclusion, our study is the first of its kind in Lebanon to describe phenotypic and genotypic characteristics of nosocomial C. parapsilosis complex isolates having a remarkable ability to form biofilms.
- MeSH
- antifungální látky * farmakologie MeSH
- biofilmy * růst a vývoj MeSH
- Candida parapsilosis * genetika izolace a purifikace klasifikace MeSH
- fenotyp * MeSH
- flukonazol farmakologie MeSH
- fungální léková rezistence * genetika MeSH
- fylogeneze * MeSH
- genotyp * MeSH
- infekce spojené se zdravotní péčí mikrobiologie MeSH
- kandidóza * mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti * MeSH
- nemocnice MeSH
- sekvenování celého genomu * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Libanon MeSH
Závěrečná zpráva o řešení grantu Agentury pro zdravotnický výzkum MZ ČR
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Narůstající incidence bakterií rezistentních k antibiotikům poslední volby představuje jeden z nejvýznamnějších medicínských problémů, který komplikuje úspěšnou léčbu život ohrožujících infekcí. V rámci navrhovaného projektu bude provedena molekulárně-epidemiologická/genomická analýza Enterobacterales produkujících karbapenemázy (CPE) izolovaných z pacientů hospitalizovaných v českých nemocnicích a srovnání s bakteriální populací citlivou ke karbapenemům. S využitím nejmodernějších postupů celogenomového sekvenování a bioinformatické analýzy budou identifikovány jedinečné rysy CPE, vysoce rizikové klony se zvýšeným významem pro zdraví populace a mobilní genetické elementy spojené s šířením rezistence ke karbapenemům v nemocnicích. Komplexní genomická analýza umožní popsat genetickou strukturu sledovaných bakteriálních populací včetně její dynamiky prostřednictvím mikroevolučních změn během šíření nemocničních nákaz. Výsledky projektu budou bezprostředně využity pro zlepšení a zpřesnění diagnostiky infekčních nemocí a návrh účinnějších postupů v prevenci a kontrole šíření patogenů.; Growing incidence of bacteria resistant to last-line antibiotics represents one of the most important medical issue, complicating the successful treatment of life-threatening infections. In this project, molecular-epidemiological and genomic analysis of carbapenemase-producing Enterobacterales (CPE) isolated from patients in Czech hospitals and its comparison with the carbapenem-susceptible population will be performed. Whole genome sequencing and bioinformatics analysis will allow to identify specific features of CPE, high-risk clones with increased importance for public health and mobile genetic elements associated with the dissemination of resistance to carbapenems in hospitals. Complex genomic analysis will reveal genetic structure of the studied bacterial populations including their dynamics through microevolutionary changes in the clinical context of a hospital outbreak. The project outcomes will be used to improve the diagnostics of infectious diseases and to outline effective interventions for the prevention and control of the transmission of bacterial pathogens.
Fosfomycin (FOS) is an effective antibiotic against multidrug-resistant Enterobacterales, but its effectiveness is reducing. Little is known on the current prevalence of FosA enzymes in low-risk pathogens, such as Citrobacter freundii. The aim of the study was the molecular characterization of a carbapenemase- and FosA-producing C. freundii collected in Italy. AK867, collected in 2023, showed an XDR profile, retaining susceptibility only to colistin. AK867 showed a FOS MIC >128 mg/L by ADM. Based on WGS, AK867 belonged to ST116 and owned a wide resistome, including fosA3, blaKPC-2, and blaVIM-1. fosA3 was carried by a conjugative pKPC-CAV1312 plasmid of 320,480 bp, on a novel composite transposon (12,907 bp). FosA3 transposon shared similarities with other fosA3-harboring pKPC-CAV1312 plasmids among Citrobacter spp. We report the first case of FosA3 production in clinical carbapenemase-producing C. freundii ST116. The incidence of FosA3 enzymes is increasing among Enterobacterales, affecting even low-virulence pathogens, as C. freundii.
- MeSH
- antibakteriální látky * farmakologie MeSH
- bakteriální proteiny * genetika metabolismus MeSH
- beta-laktamasy * genetika metabolismus MeSH
- Citrobacter freundii * genetika enzymologie účinky léků MeSH
- enterobakteriální infekce * mikrobiologie MeSH
- fosfomycin * farmakologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- mnohočetná bakteriální léková rezistence genetika MeSH
- plazmidy genetika MeSH
- sekvenování celého genomu MeSH
- transpozibilní elementy DNA MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Itálie MeSH
Fosfomycin-resistant FosA8-producing Enterobacterales are uncommon strains with extremely low incidence in Europe, based on only three reports in the literature. We detected FosA8-producing Escherichia coli ST131 in clinical isolates from two patients admitted in February 2023 to a rehabilitation unit in Italy. The occurrence of rare fosA-like genes in the high-risk clone ST131 is of clinical relevance. The dissemination of FosA-producing E. coli, although still at low levels, should be continuously monitored.
- MeSH
- antibakteriální látky * farmakologie terapeutické užití MeSH
- bakteriální léková rezistence MeSH
- beta-laktamasy genetika metabolismus MeSH
- Escherichia coli * izolace a purifikace genetika účinky léků MeSH
- fosfomycin farmakologie terapeutické užití MeSH
- infekce vyvolané Escherichia coli * mikrobiologie epidemiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- multilokusová sekvenční typizace MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- Geografické názvy
- Itálie MeSH
INTRODUCTION: In the battle against multidrug-resistant bacterial infections, ceftazidime- avibactam (CZA) stands as a pivotal defense, particularly against carbapenemresistant (CR) Gram-negative pathogens. However, the rise in resistance against this drug poses a significant threat to its effectiveness, highlighting the critical need for in-depth studies about its resistance mechanisms. METHODS: This research focuses on the genomic characterization of CR- and CZA-resistant Escherichia coli (n=26) and Klebsiella pneumoniae (n=34) strains, harboring the blaNDM and/or blaOXA-48-like genes, at a major Lebanese tertiary care medical center, using whole genome sequencing (WGS). RESULTS: Our findings revealed a notable prevalence of blaNDM in all K. pneumoniae strains isolates, with 27 of these also harboring blaOXA-48. On the other hand, E. coli strains predominantly carried the blaNDM-5 gene. Whole genome sequencing (WGS) identified a predominance of ST383 among K. pneumoniae strains, which possessed a multi-replicon IncFIB-IncHI1B plasmid harboring the blaNDM-5. Additionally, various Inc group plasmids in K. pneumoniae across multiple sequence types were found to carry the blaNDM. Similarly, diverse STs of E. coli were observed to carry blaNDM-5 on different plasmids. DISCUSSION: The study underscores NDM carbapenemases as a paramount resistance mechanism in Lebanon,jeopardizing critical last-resort treatments. It also illuminates the role of varied sequence types and mobile genetic elements in the spread of NDM resistance,stressing the urgent need for strategies to mitigate this threat, especially in nosocomial infections.
- MeSH
- antibakteriální látky * farmakologie MeSH
- azabicyklické sloučeniny * farmakologie MeSH
- bakteriální proteiny genetika metabolismus MeSH
- beta-laktamasy * genetika metabolismus MeSH
- ceftazidim * farmakologie MeSH
- centra terciární péče MeSH
- Enterobacteriaceae rezistentní na karbapenemy genetika účinky léků izolace a purifikace MeSH
- Escherichia coli * genetika účinky léků MeSH
- fixní kombinace léků * MeSH
- genom bakteriální MeSH
- karbapenemy * farmakologie MeSH
- Klebsiella pneumoniae * genetika účinky léků MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- mnohočetná bakteriální léková rezistence * genetika MeSH
- plazmidy genetika MeSH
- přenos genů horizontální MeSH
- sekvenování celého genomu * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Libanon MeSH
BACKGROUND: Aminopenicillins are recommended agents for non-invasive Haemophilus influenzae infections. One of the mechanisms of resistance to β-lactams is the alteration of the transpeptidase region of penicillin binding protein 3 (PBP3) which is caused by mutations in the ftsI gene. It was shown that exposure to beta-lactams has a stimulating effect on increase of prevalence of H. influenzae strains with the non-enzymatic mechanism of resistance. OBJECTIVES: The aim of our study was to compare the mutational potential of ampicillin and cefuroxime in H. influenzae strains, determination of minimum inhibitory concentration and the evolution of mutations over time, focusing on amino acid substitutions in PBP3. METHODS: 30 days of serial passaging of strains in liquid broth containing increasing concentrations of ampicillin or cefuroxime was followed by whole-genome sequencing. RESULTS: On average, cefuroxime increased the minimum inhibitory concentration more than ampicillin. The minimum inhibitory concentration was increased by a maximum of 32 fold. Substitutions in the PBP3 started to appear after 15 days of passaging. In PBP3, cefuroxime caused different substitutions than ampicillin. CONCLUSIONS: Our experiment observed differences in mutation selection by ampicillin and cefuroxime. Selection pressure of antibiotics in vitro generated substitutions that do not occur in clinical strains in the Czech Republic.
- MeSH
- ampicilin * farmakologie MeSH
- antibakteriální látky * farmakologie MeSH
- bakteriální proteiny genetika metabolismus MeSH
- cefuroxim * farmakologie MeSH
- Haemophilus influenzae * genetika účinky léků MeSH
- hemofilové infekce mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti * MeSH
- molekulární evoluce MeSH
- mutace * MeSH
- proteiny vázající penicilin * genetika metabolismus MeSH
- sekvenování celého genomu MeSH
- selekce (genetika) MeSH
- sériové pasážování MeSH
- substituce aminokyselin * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: The pathogenic fungus Candida albicans is a leading agent of death in immunocompromised individuals with a growing trend of antifungal resistance. METHODS: The purpose is to induce resistance to drugs in a sensitive C. albicans strain followed by whole-genome sequencing to determine mechanisms of resistance. Strains will be assayed for pathogenicity attributes such as ergosterol and chitin content, growth rate, virulence, and biofilm formation. RESULTS: We observed sequential increases in ergosterol and chitin content in fluconazole-resistant isolates by 78% and 44%. Surface thickening prevents the entry of the drug, resulting in resistance. Resistance imposed a fitness trade-off that led to reduced growth rates, biofilm formation, and virulence in our isolates. Sequencing revealed mutations in genes involved in resistance and pathogenicity such as ERG11, CHS3, GSC2, CDR2, CRZ2, and MSH2. We observed an increase in the number of mutations in key genes with a sequential increase in drug-selective pressures as the organism increased its odds of adapting to inhospitable environments. In ALS4, we observed two mutations in the susceptible strain and five mutations in the resistant strain. CONCLUSION: This is the first study to induce resistance followed by genotypic and phenotypic analysis of isolates to determine mechanisms of drug resistance.
- Publikační typ
- časopisecké články MeSH
A 2-year national genomic screening in the Czech Republic identified a notable prevalence of the New Delhi metallo-β-lactamase 5 (NDM-5)-producing Escherichia coli sequence type 38 (ST38) in the city of Brno. Forty-two ST38 E. coli isolates harbored the blaNDM-5 gene on the chromosome. Virulence factors confirmed the persistence of these isolates through biofilm formation. Single Nucleotide Polymorphisms (SNPs)-based phylogeny and CRISPR assay typing showed minimal genomic variations, implying a clonally driven outbreak. Results suggest that this high-risk clone may impose a nationwide problem.
- MeSH
- antibakteriální látky farmakologie MeSH
- beta-laktamasy * genetika MeSH
- biofilmy růst a vývoj MeSH
- epidemický výskyt choroby * MeSH
- Escherichia coli * genetika účinky léků izolace a purifikace enzymologie MeSH
- faktory virulence genetika MeSH
- fylogeneze MeSH
- genom bakteriální MeSH
- genomika metody MeSH
- infekce vyvolané Escherichia coli * mikrobiologie epidemiologie MeSH
- jednonukleotidový polymorfismus * MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
OBJECTIVE: Here we describe a novel IncFIA plasmid harbouring mcr-10 gene in a clinical Enterobacter ludwigii strain isolated at the University Hospital in Pilsen in the Czech Republic. METHODS: The strain was subjected to antibiotic susceptibility testing. Whole genome sequencing was performed using Illumina for short-read sequencing and Oxford Nanopore Technologies for long-read sequencing followed by hybrid assembly. The resulting genome was used to detect species using average nucleotide identity, resistance genes, plasmid replicon and MLST (using centre for genomic epidemiology databases; ResFinder, PlasmidFinder and MLST, respectively) and virulence genes using VFDB. RESULTS: Τhe strain showed susceptibility against tetracycline, cefuroxime and chloramphenicol, and it was susceptible to the second and third generation of cephalosporins, carbapenems and colistin. Genome analysis identified the strain as E. ludwigii sequence type ST20 and located the mcr-10 gene on an IncFIA (HI1)/IncFII (Yp) plasmid (pI9455333_MCR10; 129 863 bp). Upon blasting the nucleotide sequence of pI9455333_MCR10 against the NCBI database, no similar plasmid sequence was detected, implying a novel plasmid structure. Nevertheless, it showed a partial similarity with pRHBSTW-00123_3 and FDAARGOS 1432, which were detected in Enterobacter cloacae complex (ECC) strains in wastewater samples in 2017 in UK and in 2021 in the United States, respectively, and pEC81-mcr, which was detected in a clinical Escherichia coli strain in 2020 in China. Moreover, I9455333cz genome carried virulence genes coding for curli fibers, fimbrial adherence determinants, siderophore aerobactin, iron uptake proteins and regulators of sigma factor. CONCLUSION: In conclusion, we identified a novel IncF plasmid harbouring mcr-10 gene in a clinical Enterobacter ludwigii strain. To our knowledge, this is the first clinical report of mcr-10 in the Czech Republic.
- MeSH
- antibakteriální látky * farmakologie MeSH
- bakteriální proteiny genetika MeSH
- centra terciární péče * MeSH
- Enterobacter * genetika účinky léků izolace a purifikace MeSH
- enterobakteriální infekce * mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti * MeSH
- mnohočetná bakteriální léková rezistence genetika MeSH
- multilokusová sekvenční typizace MeSH
- plazmidy * genetika MeSH
- sekvenování celého genomu MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
