Abstract
BackgroundCorynebacterium 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.MethodsHere, 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.ResultsPhylogenetic 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.ConclusionsThis 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.
Highlights
Corynebacterium diphtheriae, the agent of diphtheria, is a genetically diverse bacterial species
All isolates were confirmed as C. diphtheriae based on an average nucleotide identity (ANI) value higher than 96% with the C. diphtheriae type strain NCTC11397T
Our phylogenetic analyses showed that C. diphtheriae is highly diverse and comprises multiple sublineages that are grouped into two major lineages, characterized by distinctive associations with biovars Mitis and Gravis and with the diphtheria toxin gene
Summary
Corynebacterium diphtheriae, the agent of diphtheria, is a genetically diverse bacterial species. Diphtheria, if untreated, is one of the most severe bacterial infections of humans. It typically affects the upper respiratory tract causing pseudomembrane formation, sometimes leading to suffocation and death. The infection can be complicated by systemic symptoms, caused by the diphtheria toxin. The diphtheria toxin, encoded by the tox gene, is carried by lysogenized corynephages within the chromosome of some C. diphtheriae strains [5, 6]. Concern exists about the possibility of lysogenic conversion of previously nontoxigenic strains during colonization, infection, or transmission chains [7]. The high genetic diversity of C. diphtheriae strains underlies their variable colonization, adhesion, and pathogenicity properties [8–10]. Three main biovars (Mitis, Gravis, and Belfanti) are distinguished since the 1950s, their phylogenetic relationships are poorly defined [11–13]
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