-
Journal of Clinical Microbiology Nov 2021Corynebacterium diphtheriae is highly transmissible and can cause large diphtheria outbreaks where vaccination coverage is insufficient. Sporadic cases or small clusters...
Corynebacterium diphtheriae is highly transmissible and can cause large diphtheria outbreaks where vaccination coverage is insufficient. Sporadic cases or small clusters are observed in high-vaccination settings. The phylogeography and short timescale evolution of C. diphtheriae are not well understood, in part due to a lack of harmonized analytical approaches of genomic surveillance and strain tracking. We combined 1,305 genes with highly reproducible allele calls into a core genome multilocus sequence typing (cgMLST) scheme. We analyzed cgMLST gene diversity among 602 isolates from sporadic clinical cases, small clusters, or large outbreaks. We defined sublineages based on the phylogenetic structure within C. diphtheriae and strains based on the highest number of cgMLST mismatches within documented outbreaks. We performed time-scaled phylogenetic analyses of major sublineages. The cgMLST scheme showed high allele call rate in C. diphtheriae and the closely related species C. belfantii and C. rouxii. We demonstrate its utility to delineate epidemiological case clusters and outbreaks using a 25 mismatches threshold and reveal a number of cryptic transmission chains, most of which are geographically restricted to one or a few adjacent countries. Subcultures of the vaccine strain PW8 differed by up to 20 cgMLST mismatches. Phylogenetic analyses revealed a short-timescale evolutionary gain or loss of the diphtheria toxin and biovar-associated genes. We devised a genomic taxonomy of strains and deeper sublineages (defined using a 500-cgMLST-mismatch threshold), currently comprising 151 sublineages, only a few of which are geographically widespread based on current sampling. The cgMLST genotyping tool and nomenclature was made publicly accessible (https://bigsdb.pasteur.fr/diphtheria). Standardized genome-scale strain genotyping will help tracing transmission and geographic spread of C. diphtheriae. The unified genomic taxonomy of C. diphtheriae strains provides a common language for studies of ecology, evolution, and virulence heterogeneity among C. diphtheriae sublineages.
Topics: Corynebacterium diphtheriae; Diphtheria; Genome, Bacterial; Genomics; Humans; Multilocus Sequence Typing; Phylogeny
PubMed: 34524891
DOI: 10.1128/JCM.01581-21 -
Journal of Bacteriology Sep 2021Corynebacterium diphtheriae is the causative agent of a severe respiratory disease in humans. The bacterial systems required for infection are poorly understood, but the...
Corynebacterium diphtheriae is the causative agent of a severe respiratory disease in humans. The bacterial systems required for infection are poorly understood, but the acquisition of metals such as manganese (Mn) is likely critical for host colonization. MntR is an Mn-dependent transcriptional regulator in C. diphtheriae that represses the expression of the genes, which encode a putative ABC metal transporter. However, other targets of Mn and MntR regulation in C. diphtheriae have not been identified. In this study, we use comparisons between the gene expression profiles of wild-type C. diphtheriae strain 1737 grown without or with Mn supplementation and comparisons of gene expression between the wild type and an deletion mutant to characterize the C. diphtheriae Mn and MntR regulon. MntR was observed to both repress and induce various target genes in an Mn-dependent manner. Genes induced by MntR include the Mn-superoxide dismutase, , and the putative ABC transporter locus, . DNA binding studies showed that MntR interacts with the promoter regions for several genes identified in the expression study, and a 17-bp consensus MntR DNA binding site was identified. We found that an mutant displayed increased sensitivity to Mn and cadmium that could be alleviated by the additional deletion of the transport locus, providing evidence that the MntABCD transporter functions as an Mn uptake system in C. diphtheriae. The findings in this study further our understanding of metal uptake systems and global metal regulatory networks in this important human pathogen. Mechanisms for metal scavenging are critical to the survival and success of bacterial pathogens, including Corynebacterium diphtheriae. Metal import systems in pathogenic bacteria have been studied as possible vaccine components due to high conservation, critical functionality, and surface localization. In this study, we expand our understanding of the genes controlled by the global manganese regulator, MntR. We determined a role for the MntABCD transporter in manganese import using evidence from manganese and cadmium toxicity assays. Understanding the nutritional requirements of C. diphtheriae and the tools used to acquire essential metals will aid in the development of future vaccines.
Topics: Bacterial Proteins; Biological Transport; Cloning, Molecular; Corynebacterium diphtheriae; DNA, Bacterial; Gene Expression Regulation, Bacterial; Manganese; Membrane Transport Proteins; Mutation; Oligonucleotide Array Sequence Analysis; Protein Binding; RNA, Bacterial; Regulon; Repressor Proteins
PubMed: 34370555
DOI: 10.1128/JB.00274-21 -
Current Protocols in Microbiology Sep 2020This article describes several established approaches for genetic manipulation of Corynebacterium diphtheriae, the causative agent of diphtheria that is known to have...
This article describes several established approaches for genetic manipulation of Corynebacterium diphtheriae, the causative agent of diphtheria that is known to have provided key evidence for Koch's postulates on the germ theory. First, it includes a detailed gene deletion method that generates nonpolar, in-frame, markerless deletion mutants, utilizing the levansucrase SacB as a counter-selectable marker. Second, it provides a thorough protocol for rescuing deletion mutants using Escherichia coli-Corynebacterium shuttle vectors. Finally, a Tn5 transposon mutagenesis procedure is described. In principle, these protocols can be used for other Corynebacterium species, including Corynebacterium glutamicum and Corynebacterium matruchotii. © 2020 Wiley Periodicals LLC Basic Protocol 1: Gene deletion in Corynebacterium diphtheriae Basic Protocol 2: Complementation of a mutant strain Basic Protocol 3: Tn5 transposon mutagenesis of Corynebacterium diphtheriae.
Topics: Corynebacterium; Corynebacterium diphtheriae; DNA Transposable Elements; DNA, Bacterial; Diphtheria; Escherichia coli; Gene Deletion; Genetic Complementation Test; Genetic Vectors; Hexosyltransferases; Mutagenesis, Insertional
PubMed: 32865881
DOI: 10.1002/cpmc.111 -
Infection, Genetics and Evolution :... Sep 2016Diphtheria is a debilitating disease caused by toxigenic Corynebacterium diphtheriae strains and has been effectively controlled by the toxoid vaccine, yet several... (Review)
Review
Diphtheria is a debilitating disease caused by toxigenic Corynebacterium diphtheriae strains and has been effectively controlled by the toxoid vaccine, yet several recent outbreaks have been reported across the globe. Moreover, non-toxigenic C. diphtheriae strains are emerging as a major global health concern by causing severe pharyngitis and tonsillitis, endocarditis, septic arthritis and osteomyelitis. Molecular epidemiological investigations suggest the existence of outbreak-associated clones with multiple genotypes circulating around the world. Evolution and pathogenesis appears to be driven by recombination as major virulence factors, including the tox gene and pilus gene clusters, are found within genomic islands that appear to be mobile between strains. The number of pilus gene clusters and variation introduced by gain or loss of gene function correlate with the variable adhesive and invasive properties of C. diphtheriae strains. Genomic variation does not support the separation of C. diphtheriae strains into biovars which correlates well with findings of studies based on multilocus sequence typing. Genomic analyses of a relatively small number of strains also revealed a recombination driven diversification of strains within a sequence type and indicate a wider diversity among C. diphtheriae strains than previously appreciated. This suggests that there is a need for increased effort from the scientific community to study C. diphtheriae to help understand the genomic diversity and pathogenicity within the population of this important human pathogen.
Topics: Biological Evolution; Clone Cells; Corynebacterium diphtheriae; Diphtheria; Diphtheria Toxoid; Disease Outbreaks; Genetic Variation; Genome, Bacterial; Genomic Islands; Genotype; Humans; Molecular Epidemiology; Multilocus Sequence Typing; Phylogeny; Virulence Factors
PubMed: 27291708
DOI: 10.1016/j.meegid.2016.06.024 -
Genome Medicine Nov 2020Corynebacterium diphtheriae, the agent of diphtheria, is a genetically diverse bacterial species. Although antimicrobial resistance has emerged against several drugs...
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.
Topics: Anti-Bacterial Agents; Corynebacterium diphtheriae; DNA, Bacterial; Diphtheria; Diphtheria Toxin; Drug Resistance, Bacterial; Genes, Bacterial; Genome-Wide Association Study; Genomics; Humans; Macrolides; Metagenomics; Microbial Sensitivity Tests; Multilocus Sequence Typing; Phylogeny; Prospective Studies
PubMed: 33246485
DOI: 10.1186/s13073-020-00805-7 -
International Journal of Molecular... Mar 2022, the etiological agent of diphtheria, is a re-emerging pathogen, responsible for several thousand deaths per year. In addition to diphtheria, systemic infections, often... (Review)
Review
, the etiological agent of diphtheria, is a re-emerging pathogen, responsible for several thousand deaths per year. In addition to diphtheria, systemic infections, often by non-toxigenic strains, are increasingly observed. This indicates that besides the well-studied and highly potent diphtheria toxin, various other virulence factors may influence the progression of the infection. This review focuses on the known components of responsible for adhesion, invasion, inflammation, and cell death, as well as on the cellular signaling pathways activated upon infection.
Topics: Corynebacterium; Corynebacterium diphtheriae; Diphtheria; Diphtheria Toxin; Humans; Virulence Factors
PubMed: 35328715
DOI: 10.3390/ijms23063298 -
Revista Chilena de Infectologia :... Apr 2018
Topics: Animals; Bacterial Typing Techniques; Chile; Corynebacterium diphtheriae; Diphtheria; Erythromycin; Penicillin G
PubMed: 29912257
DOI: 10.4067/s0716-10182018000200189 -
Microbial Genomics Sep 2023Respiratory diphtheria is a serious infection caused by toxigenic , and disease transmission mainly occurs through respiratory droplets. Between 2017 and 2019, a large...
Respiratory diphtheria is a serious infection caused by toxigenic , and disease transmission mainly occurs through respiratory droplets. Between 2017 and 2019, a large diphtheria outbreak among forcibly displaced Myanmar nationals densely settled in Bangladesh was investigated. Here we utilized whole-genome sequencing (WGS) to characterize recovered isolates of and two co-circulating non-diphtheritic (NDC) species - and isolates recovered from all 53 positive cases in this study were identified as toxigenic biovar mitis, exhibiting intermediate resistance to penicillin, and formed four phylogenetic clusters circulating among multiple refugee camps. Additional sequenced isolates collected from two patients showed co-colonization with non-toxigenic biovar gravis, one of which exhibited decreased susceptibility to the first-line antibiotics and harboured a novel 23-kb multidrug resistance plasmid. Results of phylogenetic reconstruction and virulence-related gene contents of the recovered NDC isolates indicated they were likely commensal organisms, though 80.4 %(45/56) were not susceptible to erythromycin, and most showed high minimum inhibition concentrations against azithromycin. These results demonstrate the high resolution with which WGS can aid molecular investigation of diphtheria outbreaks, through the quantification of bacterial genetic relatedness, as well as the detection of virulence factors and antibiotic resistance markers among case isolates.
Topics: Humans; Corynebacterium diphtheriae; Diphtheria; Myanmar; Phylogeny; Corynebacterium; Genomics
PubMed: 37712831
DOI: 10.1099/mgen.0.001085 -
The New England Journal of Medicine Mar 2018
Topics: Child, Preschool; Corynebacterium diphtheriae; Diphtheria; Female; Humans; Skin Diseases, Bacterial; Staphylococcus aureus
PubMed: 29590546
DOI: 10.1056/NEJMicm1701825 -
Microbial Genomics Oct 2022Diphtheria is a potentially fatal respiratory disease caused by toxigenic forms of the Gram-positive bacterium . Despite the availability of treatments (antitoxin and...
Diphtheria is a potentially fatal respiratory disease caused by toxigenic forms of the Gram-positive bacterium . Despite the availability of treatments (antitoxin and antimicrobials) and effective vaccines, the disease still occurs sporadically in low-income countries and in higher income where use of diphtheria vaccine is inconsistent. Diphtheria was highly endemic in Vietnam in the 1990s; here, we aimed to provide some historical context to the circulation of erythromycin resistant organisms in Vietnam during this period. After recovering 54 . isolated from clinical cases of diphtheria in Ho Chi Minh City between 1992 and 1998 we conducted whole genome sequencing and analysis. Our data outlined substantial genetic diversity among the isolates, illustrated by seven distinct Sequence Types (STs), but punctuated by the sustained circulation of ST67 and ST209. With the exception of one isolate, all sequences contained the gene, which was classically located on a corynebacteriophage. All erythromycin resistant isolates, accounting for 13 % of organisms in this study, harboured a novel 18 kb )-carrying plasmid, which exhibited limited sequence homology to previously described resistance plasmids in . Our study provides historic context for the circulation of antimicrobial resistant in Vietnam; these data provide a framework for the current trajectory in global antimicrobial resistance trends.
Topics: Humans; Corynebacterium diphtheriae; Diphtheria; Erythromycin; Vietnam; Corynebacterium; Diphtheria Toxoid; Antitoxins
PubMed: 36259695
DOI: 10.1099/mgen.0.000861