scholarly journals Population genomics and antimicrobial resistance in Corynebacterium diphtheriae

2020 ◽  
Vol 12 (1) ◽  
Author(s):  
Melanie Hennart ◽  
Leonardo G. Panunzi ◽  
Carla Rodrigues ◽  
Quentin Gaday ◽  
Sarah L. Baines ◽  
...  
Keyword(s):  
Soviet Union ◽  
Antimicrobial Resistance ◽  
Diphtheria Toxin ◽  
Population Genomics ◽  
Bacterial Species ◽  
Toxin Production ◽  
Multidrug Resistant ◽  
Corynebacterium Diphtheriae ◽  
Toxin Gene ◽  
Phylogenetic Structure

Abstract Background Corynebacterium diphtheriae, the agent of diphtheria, is a genetically diverse bacterial species. Although antimicrobial resistance has emerged against several drugs including first-line penicillin, the genomic determinants and population dynamics of resistance are largely unknown for this neglected human pathogen. Methods Here, we analyzed the associations of antimicrobial susceptibility phenotypes, diphtheria toxin production, and genomic features in C. diphtheriae. We used 247 strains collected over several decades in multiple world regions, including the 163 clinical isolates collected prospectively from 2008 to 2017 in France mainland and overseas territories. Results Phylogenetic analysis revealed multiple deep-branching sublineages, grouped into a Mitis lineage strongly associated with diphtheria toxin production and a largely toxin gene-negative Gravis lineage with few toxin-producing isolates including the 1990s ex-Soviet Union outbreak strain. The distribution of susceptibility phenotypes allowed proposing ecological cutoffs for most of the 19 agents tested, thereby defining acquired antimicrobial resistance. Penicillin resistance was found in 17.2% of prospective isolates. Seventeen (10.4%) prospective isolates were multidrug-resistant (≥ 3 antimicrobial categories), including four isolates resistant to penicillin and macrolides. Homologous recombination was frequent (r/m = 5), and horizontal gene transfer contributed to the emergence of antimicrobial resistance in multiple sublineages. Genome-wide association mapping uncovered genetic factors of resistance, including an accessory penicillin-binding protein (PBP2m) located in diverse genomic contexts. Gene pbp2m is widespread in other Corynebacterium species, and its expression in C. glutamicum demonstrated its effect against several beta-lactams. A novel 73-kb C. diphtheriae multiresistance plasmid was discovered. Conclusions This work uncovers the dynamics of antimicrobial resistance in C. diphtheriae in the context of phylogenetic structure, biovar, and diphtheria toxin production and provides a blueprint to analyze re-emerging diphtheria.

scholarly journals Population genomics and antimicrobial resistance in Corynebacterium diphtheriae

2020 ◽  
Author(s):  
Melanie Hennart ◽  
Leonardo G. Panunzi ◽  
Carla Rodrigues ◽  
Quentin Gaday ◽  
Sarah L. Baines ◽  
...  
Keyword(s):  
Soviet Union ◽  
Antimicrobial Resistance ◽  
Diphtheria Toxin ◽  
Population Genomics ◽  
Bacterial Species ◽  
Toxin Production ◽  
Multidrug Resistant ◽  
Corynebacterium Diphtheriae ◽  
Phylogenetic Structure ◽  
Overseas Territories

ABSTRACTCorynebacterium diphtheriae, the agent of diphtheria, is a genetically diverse bacterial species. Although antimicrobial resistance has emerged against several drugs including first-line penicillin, the genomic determinants and population dynamics of resistance are largely unknown for this neglected human pathogen.Here we analyzed the associations of antimicrobial susceptibility phenotypes, diphtheria toxin production and genomic features in C. diphtheriae. We used 247 strains collected over several decades in multiple world regions, including the 163 clinical isolates collected prospectively from 2008 to 2017 in France mainland and overseas territories.Phylogenetic analysis revealed multiple deep-branching sublineages, grouped into a Mitis lineage strongly associated with diphtheria toxin production, and a tox-negative Gravis lineage with few tox+ exceptions including the 1990s ex-Soviet Union outbreak strain. The distribution of susceptibility phenotypes allowed proposing ecological cutoffs for most of the 19 agents tested, thereby defining acquired antimicrobial resistance. Penicillin resistance was found in 17.2% of prospective isolates. Four isolates were multidrug resistant (>8 agents), including to penicillin and macrolides. Homologous recombination was frequent (r/m = 5) and horizontal gene transfer contributed to the emergence of antimicrobial resistance in multiple sublineages. Genome-wide association mapping uncovered genetic factors of resistance, including an accessory penicillin-binding protein (PBP2m) located in diverse genomic contexts. Gene pbp2m is widespread in other Corynebacterium species and its expression in C. glutamicum demonstrated its effect against several beta-lactams. A novel 73-kb C. diphtheriae multi-resistance plasmid was discovered.This work uncovers the dynamics of antimicrobial resistance in C. diphtheriae in the context of phylogenetic structure, biovar and diphtheria toxin production, and provides a blueprint to analyze re-emerging diphtheria.

scholarly journals Assessing the Genetic Diversity of Austrian Corynebacterium diphtheriae Clinical Isolates, 2011-2019

2020 ◽  
Author(s):  
Justine Schaeffer ◽  
Steliana Huhulescu ◽  
Anna Stoeger ◽  
Franz Allerberger ◽  
Werner Ruppitsch
Keyword(s):  
Genetic Diversity ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Resistance Genes ◽  
Diphtheria Toxin ◽  
Corynebacterium Diphtheriae ◽  
Toxin Gene ◽  
Whole Genome ◽  
Skin Infections ◽  
Antimicrobial Resistance Genes

Background: Diphtheria is a vaccine preventable disease with a high potential for re-emergence. One of its causative agent is Corynebacterium diphtheriae, some strains producing the diphtheria toxin. From 2011 to 2019, 57 clinical C. diphtheriae strains were isolated in Austria, either from the respiratory track or from skin infections. Objectives: The aim of the study was to investigate the genetic diversity of these C. diphtheriae isolates using whole genome sequencing. Methods: Isolates were characterized by genome wide comparison using single nucleotide polymorphism or core genome multilocus sequence typing, and by searching sequence data for antimicrobial resistance genes and genes involved in diphtheria toxin production. Results: Genetic diversity between the isolates was high, with no clear distribution over time or place. C. belfantii isolates were separated from other strains, and were strongly associated with respiratory infections (OR = 57). Two clusters, limited in time and space, were identified. Almost 40% of strains carried resistance genes against tetracycline or sulfonamides, mostly from skin infections. Microbiological tests showed that 55% of isolates were resistant to penicillin, but did not carry genes conferring β-lactam resistance. Diphtheria toxin gene with no non-synonymous mutation was found in three isolates only. Conclusion: This study showed that sequencing can provide valuable information complementing routine microbiological and epidemiological investigations. It allowed us to identify unknown clusters, evaluate antimicrobial resistances more broadly and support toxigenicity results obtained by PCR. For these reasons, C. diphtheriae surveillance could strongly benefit from the routine implementation of whole genome sequencing.

scholarly journals Construction and Characterization of Transposon Insertion Mutations in Corynebacterium diphtheriae That Affect Expression of the Diphtheria Toxin Repressor (DtxR)

2002 ◽  
Vol 184 (20) ◽  
pp. 5723-5732 ◽  
Author(s):  
Diana Marra Oram ◽  
Ana Avdalovic ◽  
Randall K. Holmes
Keyword(s):  
Oxidative Stress ◽  
Diphtheria Toxin ◽  
Corynebacterium Diphtheriae ◽  
Insertion Mutant ◽  
Toxin Gene ◽  
Wild Type ◽  
Transposon Insertion ◽  
Diphtheria Toxin Repressor ◽  
Mutant Strains

ABSTRACT Transcription of the bacteriophage-borne diphtheria toxin gene tox is negatively regulated, in response to intracellular Fe2+ concentration, by the chromosomally encoded diphtheria toxin repressor (DtxR). Due to a scarcity of tools, genetic analysis of Corynebacterium diphtheriae has primarily relied on analysis of chemically induced and spontaneously occurring mutants and on the results of experiments with C. diphtheriae genes cloned in Escherichia coli or analyzed in vitro. We modified a Tn5-based mutagenesis technique for use with C. diphtheriae, and we used it to construct the first transposon insertion libraries in the chromosome of this gram-positive pathogen. We isolated two insertions that affected expression of DtxR, one 121 bp upstream of dtxR and the other within an essential region of the dtxR coding sequence, indicating for the first time that dtxR is a dispensable gene in C. diphtheriae. Both mutant strains secrete diphtheria toxin when grown in medium containing sufficient iron to repress secretion of diphtheria toxin by wild-type C. diphtheriae. The upstream insertion mutant still produces DtxR in decreased amounts and regulates siderophore secretion in response to iron in a manner similar to its wild-type parent. The mutant containing the transposon insertion within dtxR does not produce DtxR and overproduces siderophore in the presence of iron. Differences in the ability of the two mutant strains to survive oxidative stress also indicated that the upstream insertion retained slight DtxR activity, whereas the insertion within dtxR abolished DtxR activity. This is the first evidence that DtxR plays a role in protecting the cell from oxidative stress.

scholarly journals Respiratory Illness Caused by Corynebacterium diphtheriae and C. ulcerans, and Use of Diphtheria Antitoxin in the United States, 1996–2018

2020 ◽  
Author(s):  
John O Otshudiema ◽  
Anna M Acosta ◽  
Pamela K Cassiday ◽  
Stephen C Hadler ◽  
Susan Hariri ◽  
...  
Keyword(s):  
United States ◽  
Diphtheria Toxin ◽  
Healthcare Providers ◽  
Respiratory Illness ◽  
The United States ◽  
Corynebacterium Diphtheriae ◽  
Toxin Gene ◽  
Corynebacterium Ulcerans ◽  
Diphtheria Antitoxin ◽  
Control And Prevention

Abstract Background Respiratory diphtheria is a toxin-mediated disease caused by Corynebacterium diphtheriae. Diphtheria-like illness, clinically indistinguishable from diphtheria, is caused by Corynebacterium ulcerans, a zoonotic bacterium that can also produce diphtheria toxin. In the United States, respiratory diphtheria is nationally notifiable: specimens from suspected cases are submitted to the Centers for Disease Control and Prevention (CDC) for species and toxin confirmation, and diphtheria antitoxin (DAT) is obtained from CDC for treatment. We summarize the epidemiology of respiratory diphtheria and diphtheria-like illness and describe DAT use during 1996–2018 in the United States. Methods We described respiratory diphtheria cases reported to the National Notifiable Diseases Surveillance System (NNDSS) and C. ulcerans-related diphtheria-like illness identified through specimen submissions to CDC during 1996–2018. We reviewed DAT requests from 1997 to 2018. Results From 1996 to 2018, 14 respiratory diphtheria cases were reported to NNDSS. Among these 14 cases, 1 was toxigenic and 3 were nontoxigenic C. diphtheriae by culture and Elek, 6 were culture-negative but polymerase chain reaction (PCR)-positive for diphtheria toxin gene, 1 was culture-positive without further testing, and the remaining 3 were either not tested or tested negative. Five cases of respiratory diphtheria-like illness caused by toxigenic C. ulcerans were identified. DAT was requested by healthcare providers for 151 suspected diphtheria cases between 1997 and 2018, with an average of 11 requests per year from 1997 to 2007, and 3 per year from 2008 to 2018. Conclusions Respiratory diphtheria remains rare in the United States, and requests for DAT have declined. Incidental identification of C. ulcerans-related diphtheria-like illness suggests surveillance of this condition might be warranted.

Coordinate regulation of siderophore and diphtheria toxin production by iron in Corynebacterium diphtheriae

1990 ◽  
Vol 9 (4) ◽  
pp. 267-273 ◽  
Author(s):  
Shih-Peng S. Tai ◽  
Amy E. Krafft ◽  
Padmaja Nootheti ◽  
Randall K. Holmes
Keyword(s):  
Diphtheria Toxin ◽  
Toxin Production ◽  
Corynebacterium Diphtheriae ◽  
Coordinate Regulation

Optimization of diphtheria toxin production by Corynebacterium diphtheriae using a casein-based medium in a fermenter

2016 ◽  
Vol 21 (4) ◽  
pp. 537-543 ◽  
Author(s):  
Yong-Ju Chung ◽  
Jin-A Lee ◽  
Mi-Young Jung ◽  
Sang-Mi Lee ◽  
Tae-Yeon Kim ◽  
...  
Keyword(s):  
Diphtheria Toxin ◽  
Toxin Production ◽  
Corynebacterium Diphtheriae

scholarly journals Tracking of Antibiotic Resistance Transfer and Rapid Plasmid Evolution in a Hospital Setting by Nanopore Sequencing

mSphere ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
Silke Peter ◽  
Mattia Bosio ◽  
Caspar Gross ◽  
Daniela Bezdan ◽  
Javier Gutierrez ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Antimicrobial Resistance ◽  
Resistance Gene ◽  
Bacterial Species ◽  
Hospital Setting ◽  
Multidrug Resistant ◽  
Plasmid Transfer ◽  
Nanopore Sequencing ◽  
Content Type ◽  
Antimicrobial Resistance Gene

ABSTRACT Infections with multidrug-resistant bacteria often leave limited or no treatment options. The transfer of antimicrobial resistance genes (ARG) carrying plasmids between bacterial species by horizontal gene transfer represents an important mode of expansion of ARGs. Here, we demonstrate the application of Nanopore sequencing in a hospital setting for monitoring transfer and rapid evolution of antibiotic resistance plasmids within and across multiple species. In 2009, we experienced an outbreak with extensively multidrug-resistant Pseudomonas aeruginosa harboring the carbapenemase-encoding blaIMP-8 gene. In 2012, the first Citrobacter freundii and Citrobacter cronae strains harboring the same gene were detected. Using Nanopore and Illumina sequencing, we conducted comparative analysis of all blaIMP-8 bacteria isolated in our hospital over a 6-year period (n = 54). We developed the computational platform plasmIDent for Nanopore-based characterization of clinical isolates and monitoring of ARG transfer, comprising de novo assembly of genomes and plasmids, plasmid circularization, ARG annotation, comparative genome analysis of multiple isolates, and visualization of results. Using plasmIDent, we identified a 40-kb plasmid carrying blaIMP-8 in P. aeruginosa and C. freundii, verifying the plasmid transfer. Within C. freundii, the plasmid underwent further evolution and plasmid fusion, resulting in a 164-kb megaplasmid, which was transferred to C. cronae. Multiple rearrangements of the multidrug resistance gene cassette were detected in P. aeruginosa, including deletions and translocations of complete ARGs. In summary, plasmid transfer, plasmid fusion, and rearrangement of the ARG cassette mediated the rapid evolution of opportunistic pathogens in our hospital. We demonstrated the feasibility of near-real-time monitoring of plasmid evolution and ARG transfer in clinical settings, enabling successful countermeasures to contain plasmid-mediated outbreaks. IMPORTANCE Infections with multidrug-resistant bacteria represent a major threat to global health. While the spread of multidrug-resistant bacterial clones is frequently studied in the hospital setting, surveillance of the transfer of mobile genetic elements between different bacterial species was difficult until recent advances in sequencing technologies. Nanopore sequencing technology was applied to track antimicrobial gene transfer in a long-term outbreak of multidrug-resistant Pseudomonas aeruginosa, Citrobacter freundii, and Citrobacter cronae in a German hospital over 6 years. We developed a novel computational pipeline, pathoLogic, which enables de novo assembly of genomes and plasmids, antimicrobial resistance gene annotation and visualization, and comparative analysis. Applying this approach, we detected plasmid transfer between different bacterial species as well as plasmid fusion and frequent rearrangements of the antimicrobial resistance gene cassette. This study demonstrated the feasibility of near-real-time tracking of plasmid-based antimicrobial resistance gene transfer in hospitals, enabling countermeasures to contain plasmid-mediated outbreaks.

scholarly journals Distribution of carbapenem resistance determinants among epidemic and non-epidemic types of Acinetobacter species in Japan

2014 ◽  
Vol 63 (6) ◽  
pp. 870-877 ◽  
Author(s):  
Mari Matsui ◽  
Satowa Suzuki ◽  
Kunikazu Yamane ◽  
Masato Suzuki ◽  
Toshifumi Konda ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Bacterial Species ◽  
Carbapenem Resistance ◽  
Multidrug Resistant ◽  
The Other ◽  
Fluoroquinolone Resistance ◽  
Pasteur Institute ◽  
Acinetobacter Species ◽  
Resistance Determinants ◽  
Sequence Types

We performed a comparative molecular analysis on three types of clinically isolated Acinetobacter spp.: epidemic sequence types (STs) of Acinetobacter baumannii (epidemic ST-AB), non-epidemic sequence types of A. baumannii (non-epidemic ST-AB) and non-baumannii Acinetobacter spp. A total of 87 isolates – 46 A. baumannii, 25 A. pittii and 16 A. nosocomialis – from 43 hospitals were analysed. Of these, 31 A. baumannii isolates were ST1 or ST2 according to the Pasteur Institute multilocus sequence typing scheme and were defined as epidemic ST-AB. The other 15 A. baumannii isolates were defined as non-epidemic ST-AB. The epidemic ST-AB isolates harboured the bla OXA-23-like gene or had an ISAba1 element upstream of bla OXA-51-like, or both, whereas non-epidemic ST-AB and non-baumannii Acinetobacter spp. isolates harboured bla OXA-58-like or metallo-β-lactamase genes, or both. The proportion of multidrug-resistant isolates was significantly higher in the epidemic ST-AB isolates (48 %) than that in the other types of Acinetobacter isolates (5 %) (P<0.05). In addition, epidemic ST-AB isolates exhibited a relatively higher proportion of fluoroquinolone resistance. We demonstrated that, in terms of genotypes and phenotypes of antimicrobial resistance, non-epidemic ST-AB isolates shared more similarity with non-baumannii Acinetobacter spp. isolates than with epidemic ST-AB isolates, regardless of bacterial species. In addition, this study revealed that, even in Japan, where IMP-type metallo-β-lactamase producers are endemic, epidemic ST-AB harbouring bla IMP have not yet emerged.

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