ABSTRACT
The discovery of antibiotics more than 80 years ago has led to considerable improvements in human and animal health. Although antibiotic resistance in environmental bacteria is ancient, resistance in human pathogens is thought to be a modern phenomenon that is driven by the clinical use of antibiotics1. Here we show that particular lineages of methicillin-resistant Staphylococcus aureus-a notorious human pathogen-appeared in European hedgehogs in the pre-antibiotic era. Subsequently, these lineages spread within the local hedgehog populations and between hedgehogs and secondary hosts, including livestock and humans. We also demonstrate that the hedgehog dermatophyte Trichophyton erinacei produces two ß-lactam antibiotics that provide a natural selective environment in which methicillin-resistant S. aureus isolates have an advantage over susceptible isolates. Together, these results suggest that methicillin resistance emerged in the pre-antibiotic era as a co-evolutionary adaptation of S. aureus to the colonization of dermatophyte-infected hedgehogs. The evolution of clinically relevant antibiotic-resistance genes in wild animals and the connectivity of natural, agricultural and human ecosystems demonstrate that the use of a One Health approach is critical for our understanding and management of antibiotic resistance, which is one of the biggest threats to global health, food security and development.
Subject(s)
Anti-Bacterial Agents/history , Arthrodermataceae/metabolism , Hedgehogs/metabolism , Hedgehogs/microbiology , Methicillin Resistance/genetics , Methicillin-Resistant Staphylococcus aureus/genetics , Selection, Genetic/genetics , Animals , Anti-Bacterial Agents/metabolism , Arthrodermataceae/genetics , Denmark , Europe , Evolution, Molecular , Geographic Mapping , History, 20th Century , Humans , Methicillin-Resistant Staphylococcus aureus/metabolism , New Zealand , One Health , Penicillins/biosynthesis , Phylogeny , beta-Lactams/metabolismABSTRACT
Whole genome sequencing data of 874 Escherichia coli isolates carrying bla NDM-5 from 13 European Union/European Economic Area countries between 2012 and June 2022 showed the predominance of sequence types ST167, ST405, ST410, ST361 and ST648, and an increasing frequency of detection. Nearly a third (30.6%) of these isolates were associated with infections and more than half (58.2%) were predicted to be multidrug-resistant. Further spread of E. coli carrying bla NDM-5 would leave limited treatment options for serious E. coli infections.
Subject(s)
Escherichia coli Infections , Escherichia coli , Humans , beta-Lactamases/genetics , Drug Resistance, Multiple, Bacterial/genetics , Escherichia coli/genetics , Escherichia coli/isolation & purification , Escherichia coli Infections/drug therapy , Escherichia coli Infections/epidemiology , European Union , Microbial Sensitivity Tests , Europe/epidemiologyABSTRACT
Objectives: To identify and characterize a novel MBL gene conferring carbapenem resistance to an isolate of Enterobacter cloacae from Austria. Methods: The novel MBL gene was heterologously expressed in Escherichia coli TOP10 to conduct comparative MIC studies and biochemical assays. Furthermore, WGS was performed using Illumina MiSeq and Oxford Nanopore MinION instruments to analyse the genetic environment of the novel MBL gene. Results: The novel MBL showed highest sequence homology to a predicted MBL precursor from the marine bacterium Rheinheimera pacifica and hence belongs to Ambler subgroup B3. The comparative MIC studies and biochemical assays showed activity of the novel enzyme against penicillins, cephalosporins and carbapenems, but not against aztreonam. It was named Linz MBL (LMB-1). The blaLMB-1 gene was shown to be located on a 108 kb plasmid of Inc type IncFIB(K). Of note, a gene adjacent to blaLMB-1 coded for a glycerophosphoryl diester phosphodiesterase that was also previously detected in R. pacifica. Conclusions: Homologies of the MBL gene itself and another gene located on the same plasmid to genes detected in marine bacterial species strongly suggest that this novel MBL was transferred to E. cloacae from a marine bacterium. This underlines the importance of natural reservoirs supplying hitherto unknown resistance genes to clinically relevant bacterial species and the importance of ongoing surveillance and research.
Subject(s)
Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Enterobacter cloacae/enzymology , Enterobacteriaceae Infections/microbiology , Escherichia coli/drug effects , beta-Lactamases/genetics , beta-Lactams/pharmacology , Austria , Bacterial Proteins/biosynthesis , Enterobacter cloacae/genetics , Enterobacter cloacae/isolation & purification , Escherichia coli/genetics , Gene Expression , Humans , Microbial Sensitivity Tests , Plasmids/analysis , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics , Sequence Homology , Whole Genome Sequencing , beta-Lactamases/biosynthesisABSTRACT
Since colistin resistance based on the plasmid-encoded mcr-1 gene was first described, this resistance gene in Enterobacteriaceae has been found worldwide. These organisms are typically of heterogeneous genetic background and show exceptional clonal diversity. We describe the first confirmation of mcr-1 in a human Escherichia coli strain cultured from a surveillance stool sample of an Austrian oncology patient.
Subject(s)
Escherichia coli/metabolism , Anti-Bacterial Agents/pharmacology , Austria , Colistin/pharmacology , Drug Resistance, Bacterial/genetics , Escherichia coli/drug effects , Escherichia coli/genetics , Escherichia coli Infections/microbiology , Feces/microbiology , Humans , Microbial Sensitivity TestsABSTRACT
BACKGROUND: Morganella spp are opportunistic pathogens involved in various infections. Intrinsic resistance to multiple antibiotics (including colistin) combined with the emergence of carbapenemase producers reduces the number of active antimicrobials. The aim of this study was to characterise genetic features related to the spread of carbapenem-resistant Morganella spp. METHODS: This comparative genomic study included extensively drug-resistant Morganella spp isolates collected between Jan 1, 2013, and March 1, 2021, by the French National Reference Center (NRC; n=68) and European antimicrobial resistance reference centres in seven European countries (n=104), as well as one isolate from Canada, two reference strains from the Pasteur Institute collection (Paris, France), and two colistin-susceptible isolates from Bicêtre Hospital (Kremlin-Bicêtre, France). The isolates were characterised by whole-genome sequencing, antimicrobial susceptibility testing, and biochemical tests. Complete genomes from GenBank (n=103) were also included for genomic analysis, including phylogeny and determination of core genomes and resistomes. Genetic distance between different species or subspecies was performed using average nucleotide identity (ANI). Intrinsic resistance mechanisms to polymyxins were investigated by combining genetic analysis with mass spectrometry on lipid A. FINDINGS: Distance analysis by ANI of 275 isolates identified three groups: Morganella psychrotolerans, Morganella morganii subspecies sibonii, and M morganii subspecies morganii, and a core genome maximum likelihood phylogenetic tree showed that the M morganii isolates can be separated into four subpopulations. On the basis of these findings and of phenotypic divergences between isolates, we propose a modified taxonomy for the Morganella genus including four species, Morganella psychrotolerans, Morganella sibonii, Morganella morganii, and a new species represented by a unique environmental isolate. We propose that M morganii include two subspecies: M morganii subspecies morganii (the most prevalent) and M morganii subspecies intermedius. This modified taxonomy was supported by a difference in intrinsic resistance to tetracycline and conservation of metabolic pathways such as trehalose assimilation, both only present in M sibonii. Carbapenemase producers were mostly identified among five high-risk clones of M morganii subspecies morganii. The most prevalent carbapenemase corresponded to NDM-1, followed by KPC-2, and OXA-48. A cefepime-zidebactam combination was the most potent antimicrobial against the 172 extensively drug-resistant Morganella spp isolates in our collection from different European countries, which includes metallo-ß-lactamase producers. Lipid A analysis showed that the intrinsic resistance to colistin was associated with the presence of L-ARA4N on lipid A. INTERPRETATION: This global characterisation of, to our knowledge, the widest collection of extensively drug-resistant Morganella spp highlights the need to clarify the taxonomy and decipher intrinsic resistance mechanisms, and paves the way for further genomic comparisons. FUNDING: None.
Subject(s)
Anti-Bacterial Agents , Bacterial Proteins , Enterobacteriaceae Infections , Genome, Bacterial , Microbial Sensitivity Tests , Morganella , Phylogeny , beta-Lactamases , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , beta-Lactamases/genetics , beta-Lactamases/metabolism , Anti-Bacterial Agents/pharmacology , Genome, Bacterial/genetics , Humans , Enterobacteriaceae Infections/microbiology , Enterobacteriaceae Infections/epidemiology , Morganella/genetics , Genomics , Whole Genome Sequencing , Europe/epidemiology , Carbapenems/pharmacology , Drug Resistance, Multiple, Bacterial/genetics , Colistin/pharmacologyABSTRACT
The introduction of multidimensional NMR spectroscopy was a breakthrough in biological NMR methodology because it allowed the unequivocal correlation of different spin states of the system. The introduction of large pressure perturbations in the corresponding radio frequency (RF) pulse sequences adds an extra structural dimension into these experiments. We have developed a microprocessor-controlled pressure jump unit that is able to introduce fast, strong pressure changes at any point in the pulse sequences. Repetitive pressure changes of 80 MPa in the sample tube are thus feasible in less than 30 ms. Two general forms of these experiments are proposed here, the pressure perturbation transient state spectroscopy (PPTSS) and the pressure perturbation state correlation spectroscopy (PPSCS). PPTSS can be used to measure the rate constants and the activation energies and activation volumes for the transition between different conformational states including the folded and unfolded state of proteins, for polymerization-depolymerization processes, and for ligand binding at atomic resolution. PPSCS spectroscopy correlates the NMR parameters of different pressure-induced states of the system, thus allowing the measurement of properties of a given pressure induced state such as a folding intermediate in a different state, for example, the folded state. Selected examples for PPTSS and PPSCS spectroscopy are presented in this Article.
Subject(s)
Bacterial Proteins/chemistry , Nuclear Magnetic Resonance, Biomolecular/instrumentation , Staphylococcus/chemistry , Equipment Design , Pressure , Protein Conformation , Protein FoldingABSTRACT
The genetic mechanisms associated with acquisition of linezolid (LZD) resistance are diverse, including point mutations in the V domain of the 23S rRNA and the 50S ribosomal proteins as well as cfr, optrA, and/or poxtA genes, which may be plasmid- or chromosomally encoded. The aim of this study was to investigate through Whole Genome Sequencing (WGS)-based typing the presence and location of genes and point mutations associated with LZD resistance in two Enterococcus faecalis isolates from Upper Austrian patients. The isolates were retrieved during screening by LZD disk diffusion test of a total of 911 clinical E. faecalis isolates in 2017. The two E. faecalis isolates had LZD minimum inhibitory concentrations of 8 and 32 mg/L and were optrA-positive (ST476 and ST585). Bioinformatic analysis revealed the presence of optrA located in the chromosome of both isolates. One isolate carried the optrA gene in the transposon 6674, previously reported as chromosomally encoded, and the second isolate in fragments originating from the integrative plasmid pEF10748. Additional mechanisms of LZD resistance on the 23S rRNA and the 50S ribosomal proteins were detected. None of the patients reported travels to geographical areas with high LZD resistance or previous LZD treatments. This is the first report of optrA carrying E. faecalis, including characterization by WGS from Austria. LZD resistance in a low-prevalence setting is of concern and should be further monitored.
Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Resistance, Bacterial/genetics , Enterococcus faecalis/drug effects , Enterococcus faecalis/genetics , Linezolid/pharmacology , Adult , Austria/epidemiology , Female , Genes, Bacterial , Humans , Male , Microbial Sensitivity Tests , Middle Aged , Oxazolidinones/pharmacology , Plasmids , RNA, Ribosomal, 23S/genetics , Retrospective Studies , Whole Genome SequencingABSTRACT
Background: Enterococcus faecium is part of the human gastrointestinal flora but may act as opportunistic pathogen. Environmental persistence, high colonization capability and diverse intrinsic and acquired resistance mechanisms make it especially successful in nosocomial high-risk settings. In March 2014, an outbreak of Linezolid resistant Enterococcus faecium (LREfm) was observed at the hematooncology department of a tertiary care center in Upper Austria. Methods: We report on the outbreak investigation together with the whole genome sequencing (WGS)-based typing results including also non-outbreak LREfm and susceptible isolates. Results: The 54 investigated isolates could be divided in six clusters based on cgMLST. Cluster one comprised LREfm isolates of genotype ST117 and CT24, which was identified as the causative clone of the outbreak. In addition, the detection of four other clusters comprising isolates originating from hematooncology patients but also at other hospitals, pointed to LREfm transmission between local healthcare facilities. LREfm patients (n = 36) were typically at risk for acquisition of nosocomial pathogens because of immunosuppression, frequent hospitalization and antibiotic therapies. Seven of these 36 patients developed LREfm infection but were successfully treated. After termination of the initial outbreak, sporadic cases occurred despite a bundle of applied outbreak control interventions. Conclusions: WGS proved to be an effective tool to differentiate several LREfm clusters in an outbreak. Active screening for LREfm is important in a high-risk setting such as hematooncology, where multiple introductions are possible and occur despite intensified infection control measures.
Subject(s)
Cross Infection/epidemiology , Disease Outbreaks , Drug Resistance, Bacterial , Enterococcus faecium/isolation & purification , Gram-Positive Bacterial Infections/epidemiology , Adult , Aged , Aged, 80 and over , Austria , Cross Infection/microbiology , Cross Infection/transmission , Enterococcus faecium/classification , Enterococcus faecium/drug effects , Female , Genome, Bacterial , Gram-Positive Bacterial Infections/microbiology , Gram-Positive Bacterial Infections/transmission , Humans , Linezolid , Male , Middle Aged , Molecular Typing , Phylogeny , Tertiary Care Centers , Whole Genome Sequencing , Young AdultABSTRACT
Enterobacterales belonging to the genus Enterobacter are well-known human pathogens and blaIMI carrying strains have been described in several countries. From Austria, we report on the first human Enterobacter mori isolate, typically a plant pathogen, which showed an undescribed multilocus sequence type and carried a blaIMI-2 carbapenemase.
Subject(s)
Bacterial Proteins/genetics , Enterobacter/genetics , Enterobacter/isolation & purification , Genome, Bacterial/genetics , beta-Lactamases/genetics , Anti-Bacterial Agents/therapeutic use , Austria , Carbapenems/therapeutic use , Enterobacter/drug effects , Enterobacteriaceae Infections/drug therapy , Enterobacteriaceae Infections/microbiology , Humans , Microbial Sensitivity Tests/methods , Middle Aged , Multilocus Sequence Typing/methods , Otitis Externa/drug therapy , Otitis Externa/microbiologyABSTRACT
In 2016, the Austrian Agency for Health and Food Safety started a pilot project to investigate antimicrobial resistance in surface water. Here we report on the characterisation of carbapenem resistant and ESBL-producing K. pneumoniae isolates from Austrian river water samples compared to 95 clinical isolates recently obtained in Austrian hospitals. Ten water samples were taken from four main rivers, collected upstream and downstream of major cities in 2016. For subtyping and comparison, public core genome multi locus sequence typing (cgMLST) schemes were used. The presence of AMR genes, virulence genes and plasmids was extracted from whole genome sequence (WGS) data. In total three ESBL-producing strains and two carbapenem resistant strains were isolated. WGS based comparison of these five water isolates to 95 clinical isolates identified three clusters. Cluster 1 (ST11) and cluster 2 (ST985) consisted of doublets of carbapenem resistant strains (one water and one clinical isolate each). Cluster 3 (ST405) consisted of three ESBL-producing strains isolated from one water sample and two clinical specimens. The cities, in which patient isolates of cluster 2 and 3 were collected, were in concordance with the water sampling locations downstream from these cities. The genetic concordance between isolates from river water samples and patient isolates raises concerns regarding the release of wastewater treatment plant effluents into surface water. From a public health perspective these findings demand attention and strategies are required to minimize the spread of multiresistant strains to the environment.
Subject(s)
Bacteria/genetics , Bacterial Proteins/analysis , Drug Resistance, Multiple, Bacterial , Hospitals , Rivers/microbiology , beta-Lactamases/analysis , Anti-Bacterial Agents/pharmacology , Austria , Bacteria/drug effects , Bacteria/metabolism , Carbapenems/pharmacology , Multilocus Sequence Typing , Pilot Projects , Whole Genome SequencingABSTRACT
The Carba NP test is a simple confirmation method for carbapenemase-producing Enterobacteriaceae (CPE) but reagents have to be freshly prepared as imipenem sodium salt is unstable. We evaluated the Carba NP test performance based on a commercially available 10-fold cheaper drug formulation containing cilastatin against 217 CPE and 78 non-CPE isolates with reduced meropenem susceptibility. Specificity and sensitivity were 100 % and 98.6 %, respectively and 3 false negative results of blaVIM-1-producing Proteus mirabilis were reproducible with the RAPIDEC® Carba NP test. Cilastatin does not disturb test performance provided that the imipenem drug quantity is doubled.
Subject(s)
Bacterial Proteins/metabolism , Bacterial Typing Techniques/methods , Cilastatin/administration & dosage , Enterobacteriaceae/enzymology , Enterobacteriaceae/isolation & purification , beta-Lactamases/metabolism , Bacterial Proteins/analysis , Enterobacteriaceae/drug effects , Reproducibility of Results , Sensitivity and Specificity , beta-Lactamases/analysisABSTRACT
Application of high pressure to biological macromolecules can be used to find new structural states with a smaller specific volume of the system. High pressure NMR spectroscopy is a most promising analytical tool for the study of these states at atomic resolution. High pressure quartz cells are difficult to handle, high quality sapphire high pressure cells are difficult to obtain commercially. In this work, we describe the use of high pressure ceramic cells produced from yttrium stabilized ZrO(2) that are capable of resisting pressures up to 200 MPa. Since the new cells should also be usable in the easily damageable cryoprobes a completely new autoclave for these cells has been constructed, including an improved method for pressure transmission, an integrated safety jacket, a displacement body, and a fast self-closing emergency valve.