Balto and Togo during the cold winter of Alaska (1925): the two canine heroes in the fight against diphtheria.

In recent years, diphtheria has re-emerged in areas with inadequate vaccination coverage, and Europe has not been spared with several cases among migrants. Diphtheria is a potentially fatal infection caused mainly by toxigenic strains of Corynebacterium diphtheriae. Due to the high mortality rate, especially among young children, the fight against diphtheria is considered one of the first conquests of immunization. In the history of medicine, there is a unique case of an unconventional response to a diphtheria outbreak in which sled dogs were used to overcome the supply difficulties of diphtheria antitoxin. The mass media followed the medical response to the outbreak and raised audience awareness of public health issues. The facts of Nome, Alaska, in 1925 can serve as a catalyst to rethink conventional responses to diphtheria outbreaks in low-income countries today and promote mass media awareness of public health importance.

Diphtheria in a Swiss Asylum Seeker Reception Centre: Outbreak Investigation and Evaluation of Testing and Vaccination Strategies.

Objectives: To describe a suspected diphtheria outbreak in a Swiss asylum seeker reception centre, and to analyse its management response regarding testing and vaccination. Methods: We retrospectively analysed clinical, microbiology, and case management data of all asylum seekers tested for C. diphtheriae between 28th August and 31st December 2022 while residing at the centre. Results are reported descriptively. Results: Among 265 individuals tested, ten cases of cutaneous diphtheria, one simultaneous respiratory and cutaneous case, and nine respiratory carriers were identified. Mass throat screening, targeted throat testing and targeted wound testing yielded 4.8%, 4.3%, and 17.4% positive results, respectively. No respiratory carrier was identified among cutaneous cases undergoing a throat swab, and no symptomatic case was identified among individuals with unspecific throat symptoms. Rates of vaccination implementation of newly arriving asylum seekers before and after the outbreak were low (17.5% and 15.5%, respectively), as were rates of targeted vaccination among cases and close contacts. Conclusion: We provide evidence for transmission both prior to arrival and within the setting, suboptimal practices and timeliness of testing, and implementation gaps in vaccination.

Corynebacterium diphtheriae and Corynebacterium ulcerans: development of EUCAST methods and generation of data on which to determine breakpoints.

BACKGROUND: Evidence-based clinical susceptibility breakpoints have been lacking for antimicrobial agents used for diphtheria. OBJECTIVES: We aimed to evaluate broth microdilution and disc diffusion methods and create a dataset of MIC values and inhibition zone diameters (ZDs) from which breakpoints could be determined. METHODS: We included 400 recent clinical isolates equally distributed by species (Corynebacterium diphtheriae and Corynebacterium ulcerans) and by national surveillance programmes (France and Germany). Non-duplicate toxigenic and non-toxigenic isolates were chosen to enable the inclusion of a diversity of susceptibility levels for the 13 agents tested. Broth microdilution and disc diffusion, using EUCAST methodology for fastidious organisms, were used. RESULTS: The distributions of MIC and ZD values were largely in agreement among methods and countries. Breakpoints to allow categorization of WT isolates as susceptible, i.e. susceptible (S) or susceptible, increased exposure (I) were determined for 12 agents. The data supported a breakpoint for benzylpenicillin and amoxicillin of resistant (R) > 1 mg/L since WT isolates were inhibited by 1 mg/L or less. WT isolates were categorized as I (S ≤ 0.001 mg/L) for benzylpenicillin, emphasizing the need for increased exposure, and S (S ≤ 1 mg/L) for amoxicillin. Erythromycin breakpoints were set at S ≤ 0.06 mg/L and R > 0.06 mg/L. The corresponding ZD breakpoints were determined for all agents except amoxicillin, for which categorization was based on benzylpenicillin results. CONCLUSIONS: This work provided a large set of antimicrobial susceptibility data for C. diphtheriae and C. ulcerans, using a harmonized methodology. The dataset allowed EUCAST and experts in the diphtheria field to develop evidence-based breakpoints in January 2023.

Cluster of Nontoxigenic Corynebacterium diphtheriae Infective Endocarditis and Rising Background C. diphtheriae Cases-Seattle, Washington, 2020-2023.

BACKGROUND: Nontoxigenic Corynebacterium diphtheriae, often associated with wounds, can rarely cause infective endocarditis (IE). Five patients with C. diphtheriae IE were identified within 12 months at a Seattle-based hospital system. We reviewed prior C. diphtheriae-positive cultures to determine if detections had increased over time and evaluated epidemiologic trends. METHODS: We conducted a formal electronic health record search to identify all patients aged ≥18 years with C. diphtheriae detected in a clinical specimen (ie, wound, blood, sputum) between 1 September 2020 and 1 April 2023. We collected patient demographics, housing status, comorbidities, substance-use history, and level of medical care required at detection. We extracted laboratory data on susceptibilities of C. diphtheriae isolates and on other pathogens detected at the time of C. diphtheriae identification. RESULTS: Between 1 September 2020 and 1 April 2023, 44 patients (median age, 44 years) had a C. diphtheriae-positive clinical culture, with most detections occurring after March 2022. Patients were predominantly male (75%), White (66%), unstably housed (77%), and had a lifetime history of injecting drugs (75%). Most C. diphtheriae-positive cultures were polymicrobial, including wound cultures from 36 (82%) patients and blood cultures from 6 (14%) patients, not mutually exclusive. Thirty-four patients (77%), including all 5 patients with C. diphtheriae IE, required hospital admission for C. diphtheriae or a related condition. Of the 5 patients with IE, 3 died of IE and 1 from COVID-19. CONCLUSIONS: Findings suggest a high-morbidity outbreak disproportionately affecting patients who use substances and are unstably housed.

The basal and major pilins in the Corynebacterium diphtheriae SpaA pilus adopt similar structures that competitively react with the pilin polymerase.

Many species of pathogenic gram-positive bacteria display covalently crosslinked protein polymers (called pili or fimbriae) that mediate microbial adhesion to host tissues. These structures are assembled by pilus-specific sortase enzymes that join the pilin components together via lysine-isopeptide bonds. The archetypal SpaA pilus from Corynebacterium diphtheriae is built by the Cd SrtA pilus-specific sortase, which crosslinks lysine residues within the SpaA and SpaB pilins to build the shaft and base of the pilus, respectively. Here, we show that Cd SrtA crosslinks SpaB to SpaA via a K139(SpaB)-T494(SpaA) lysine-isopeptide bond. Despite sharing only limited sequence homology, an NMR structure of SpaB reveals striking similarities with the N-terminal domain of SpaA (N SpaA) that is also crosslinked by Cd SrtA. In particular, both pilins contain similarly positioned reactive lysine residues and adjacent disordered AB loops that are predicted to be involved in the recently proposed "latch" mechanism of isopeptide bond formation. Competition experiments using an inactive SpaB variant and additional NMR studies suggest that SpaB terminates SpaA polymerization by outcompeting N SpaA for access to a shared thioester enzyme-substrate reaction intermediate.

Molecular characterization and whole-genome sequencing of Corynebacterium diphtheriae causing skin lesion.

We present a case of skin lesion caused by nontoxigenic Corynebacterium diphtheriae. Genomic taxonomy analyses corroborated the preliminary identification provided by mass spectrometry. The strain showed a susceptible phenotype with increased exposure to penicillin, the first drug of choice for the treatment. An empty type 1 class integron carrying only the sul1 gene, which encodes sulfonamide resistance, was found flanked by transposases. Virulence factors involved in adherence and iron uptake, as well as the CRISPR-Cas system, were predicted. MLST analysis revealed the ST-681, previously reported in French Guiana, a European territory.

Pastern dermatitis outbreak associated with toxigenic and non-toxigenic Corynebacterium diphtheriae and non-toxigenic Corynebacterium ulcerans at a horse stable in Finland, 2021.

AIMS: Corynebacterium diphtheriae and Corynebacterium ulcerans, when producing toxin, are the cause of diphtheria, a potentially life-threatening illness in humans. Horses (Equus ferus caballus) are known to be susceptible to infection that may manifest clinically on rare occasions. In late 2021 and early 2022, specimens from five horses suffering from pastern dermatitis were cultured at the Laboratory of Clinical Microbiology at the Faculty of Veterinary Medicine, University of Helsinki, Finland. C. diphtheriae and/or C. ulcerans were recovered from all of these. This study aimed to (1) analyse the bacterial isolates and (2) describe the outbreak and identify possible sources of the infection and infection routes in the stable. METHODS AND RESULTS: Susceptibility testing, PCR for the tox gene, and Elek test for toxin production in PCR-positive isolates were performed. Whole genome sequencing was also conducted to achieve high-resolution strain typing. An epidemiological survey was done by means of a semi-structured interview of horses' caretaker, and contact tracing was done among people at the stable. Two tox gene-positive, toxin-producing C. diphtheriae belonged to sequence type (ST) 822. Other C. diphtheriae (n = 2, ST828) and C. ulcerans (n = 2, ST325 and ST838) isolates did not carry the tox gene. The epidemiological investigation explored numerous possible routes of transmission, but the definite source of infection was not identified. All established human contacts tested negative for diphtheriae. All horses recovered after antimicrobial treatment. CONCLUSIONS: Our study shows that C. diphtheriae and C. ulcerans may readily spread among horses at the same stable and complicate pastern dermatitis infections. These potentially zoonotic bacteria can cause outbreaks even in a country with a very low prevalence. Caretakers should be encouraged to wear gloves and practice good hand hygiene when treating infected skin lesions in horses.

Molecular characterization and phylogenetic analysis of the first Corynebacterium rouxii strains isolated in Brazil: a recent member of Corynebacterium diphtheriae complex.

BACKGROUND: Corynebacterium diphtheriae complex was formed by the species C. diphtheriae, Corynebacterium ulcerans and Corynebacterium pseudotuberculosis in the recent past. In addition to C. diphtheriae, C. ulcerans and C. pseudotuberculosis species can carry the tox gene, which encodes diphtheria toxin. Currently, three new species have been included in the complex: Corynebacterium rouxii, Corynebacterium silvaticum, and Corynebacterium belfantii. C. rouxii is derived from the ancient Belfanti biovar of C. diptheriae. We provide the complete genome sequences of two non-toxigenic strains C. rouxii isolated from a cat with a purulent infection in Brazil. The taxonomic status and sequence type, as well as the presence of resistance and virulence genes, and CRISPR-Cas system were additionally defined. RESULTS: The genomes showed an average size of 2.4 Mb and 53.2% GC content, similar to the type strain of the species deposited in Genbank/NCBI. Strains were identified as C. rouxii by the rMLST database, with 95% identity. ANI and DDH in silico were consistent with values above the proposed cut-off points for species limit, corroborating the identification of the strains as C. rouxii. MLST analyses revealed a new ST, which differs from ST-537 only by the fusA allele. No horizontal transfer resistance gene was predicted in both genomes and no mutation was detected in the constitutive genes gyrA and rpoB. Some mutations were found in the seven penicillin-binding proteins (PBPs) detected. The tox gene was not found, but its regulatory gene dtxR was present. Among the predicted virulence genes are those involved in iron uptake and adherence, in addition to the DIP0733 protein involved in epithelial cell adhesion and invasion. The CRISPR-Cas type I-E system was detected in both genomes, with 16 spacer sequences each. Of them, half are unknown according to the databases used, indicating that there is an unexplored reservoir of corynebacteriophages and plasmids. CONCLUSIONS: This is the first genomic study of C. rouxii reported in Brazil. Here we performed taxonomic analysis and the prediction of virulence factors. The genomic analyses performed in this study may help to understand the potential pathogenesis of non-toxigenic C. rouxii strains.

Outbreak of Corynebacterium diphtheriae among asylum seekers in Belgium in 2022: operational challenges and lessons learnt.

Since 2022, European countries have been facing an outbreak of mainly cutaneous diphtheria caused by toxigenic Corynebacterium diphtheriae among asylum seekers. In Belgium, between 1 March and 31 December 2022, 25 cases of toxigenic C. diphtheriae infection were confirmed among asylum seekers, mostly among young males from Afghanistan. Multi-locus sequence typing showed that most isolates belonged to sequence types 574 or 377, similar to the majority of cases in other European countries. The investigation and management of the outbreak, with many asylum seekers without shelter, required adjustments to case finding, contact tracing and treatment procedures. A test-and-treat centre was organised by non-governmental organisations, the duration of the antimicrobial treatment was shortened to increase compliance, and isolation and contact tracing of cases was not possible. A vaccination centre was opened, and mobile vaccination campaigns were organised to vaccinate a maximum of asylum seekers. No more cases were detected between end December 2022 and May 2023. Unfortunately, though, three cases of respiratory diphtheria, including one death, were reported at the end of June 2023. To prevent future outbreaks, specific attention and sufficient resources should be allocated to this vulnerable population, in Belgium and at international level.

Multidrug-resistant Corynebacterium diphtheriae in people with travel history from West Africa to France, March to September 2023.

We describe 10 unlinked cases of Corynebacterium diphtheriae infection (nine cutaneous, one respiratory) in France in 2023 in persons travelling from Guinea, Mali, Senegal, Niger or Nigeria and Central African Republic. Four isolates were toxigenic. Seven genomically unrelated isolates were multidrug-resistant, including a toxigenic respiratory isolate with high-level resistance to macrolides and beta-lactams. The high rates of resistance, including against first-line agents, call for further microbiological investigations to guide clinical management and public health response in ongoing West African outbreaks.

National public health response to an outbreak of toxigenic Corynebacterium diphtheriae among asylum seekers in England, 2022: a descriptive epidemiological study.

BACKGROUND: In July, 2022, an increase in diphtheria cases caused by toxigenic Corynebacterium diphtheriae (C diphtheriae) was reported among asylum seekers arriving by small boats to England. Rising case numbers presented challenges for case and contact management in initial reception centres, prompting changes to national guidance and implementation of population-based control measures. This study aimed to describe the outbreak of toxigenic C diphtheriae among asylum seekers arriving by small boats to England during 2022 by use of national surveillance data. METHODS: We undertook a descriptive epidemiological analysis of cases of toxigenic C diphtheriae among asylum seekers arriving by small boats to England during 2022, incorporating genomic sequencing data, antibiotic susceptibility testing results, and epidemiological data obtained through the UK Health Security Agency's national enhanced surveillance programme. Health Protection Teams conducted risk assessments, and operational data (including details regarding offer and uptake of antibiotics and vaccinations) were obtained from National Health Service partners supporting the intervention programme. FINDINGS: In 2022, C diphtheriae isolates from 86 asylum seekers arriving by small boats were submitted to the National Reference Laboratory for confirmation and testing. Toxigenic C diphtheriae was confirmed for 72 (84%) cases and one individual with typical diphtheritic lesions but from whom no C diphtheriae was isolated from clinical swabs was also included as a probable case, resulting in 73 cases of diphtheria. 71 (97%) were male, 39 (53%) were younger than 18 years, and 36 (49%) presented with cutaneous diphtheria. The prevalence of diphtheria was highest among Afghans (1·3%) compared with all other nationalities (<0·1%). Local antibiotic susceptibility testing identified six cases with a macrolide resistant strain. INTERPRETATION: The increase in diphtheria coincided with a high volume of asylum seekers arriving by small boats to England during 2022, and subsequently increased clinical awareness of the disease among this population. Long-term disruption to vaccination programmes in origin countries along with barriers to accessing health care along migrant routes puts asylum seekers arriving by small boats at risk of disease. With arrivals expected to continue in 2023, the UK Health Security Agency has recommended continuation of population-based control measures in England until October, 2023, subject to ongoing review. FUNDING: The UK Health Security Agency.

Genomic characterization of cocirculating Corynebacterium diphtheriae and non-diphtheritic Corynebacterium species among forcibly displaced Myanmar nationals, 2017-2019.

Respiratory diphtheria is a serious infection caused by toxigenic Corynebacterium diphtheriae, 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 C. diphtheriae and two co-circulating non-diphtheritic Corynebacterium (NDC) species - C. pseudodiphtheriticum and C. propinquum. C. diphtheriae 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 C. diphtheriae 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.

Imported Toxigenic Corynebacterium Diphtheriae in Refugees with Polymicrobial Skin Infections, Germany, 2022.

During August-December 2022, toxigenic Corynebacterium diphtheriae was isolated from 25 refugees with skin infections and 2 refugees with asymptomatic throat colonization at a refugee reception center in Germany. None had systemic toxin-mediated illness. Of erosive/ulcerative skin infections, 96% were polymicrobial. Erosive/ulcerative wounds in refugees should undergo testing to rule out cutaneous diphtheria.

Phenotypic and molecular characterization and complete genome sequence of a Corynebacterium diphtheriae strain isolated from cutaneous infection in an immunized individual.

Diphtheria is an infectious disease potentially fatal that constitutes a threat to global health security, with possible local and systemic manifestations that result mainly from the production of diphtheria toxin (DT). In the present work, we report a case of infection by Corynebacterium diphtheriae in a cutaneous lesion of a fully immunized individual and provided an analysis of the complete genome of the isolate. The clinical isolate was first identified by MALDI-TOF Mass Spectrometry. The commercial strip system and mPCR performed phenotypic and genotypic characterization, respectively. The antimicrobial susceptibility profile was determined by the disk diffusion method. Additionally, genomic DNA was sequenced and analyzed for species confirmation and sequence type (ST) determination. Detection of resistance and virulence genes was performed by comparisons against ResFinder and VFDB databases. The isolate was identified as a nontoxigenic C. diphtheriae biovar Gravis strain. Its genome presented a size of 2.46 Mbp and a G + C content of 53.5%. Ribosomal Multilocus Sequence Typing (rMLST) allowed the confirmation of species as C. diphtheriae with 100% identity. DDH in silico corroborated this identification. Moreover, MLST analyses revealed that the isolate belongs to ST-536. No resistance genes were predicted or mutations detected in antimicrobial-related genes. On the other hand, virulence genes, mostly involved in iron uptake and adherence, were found. Presently, we provided sufficient clinical data regarding the C. diphtheriae cutaneous infection in addition to the phenotypic and genomic data of the isolate. Our results indicate a possible circulation of ST-536 in Brazil, causing cutaneous infection. Considering that cases of C. diphtheriae infections, as well as diphtheria outbreaks, have still been reported in several regions of the world, studies focusing on taxonomic analyzes and predictions of resistance genes may help to improve the diagnosis and to monitor the propagation of resistant clones. In addition, they can contribute to understanding the association between variation in genetic factors and resistance to antimicrobials.