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Abstract

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.

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2023-09-15
2024-11-09
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References

  1. Truelove SA, Keegan LT, Moss WJ, Chaisson LH, Macher E et al. Clinical and epidemiological aspects of diphtheria: a systematic review and pooled analysis. Clin Infect Dis 2020; 71:89–97 [View Article] [PubMed]
    [Google Scholar]
  2. Efstratiou A, Engler KH, Mazurova IK, Glushkevich T, Vuopio-Varkila J et al. Current approaches to the laboratory diagnosis of diphtheria. J Infect Dis 2000; 181 Suppl 1:S138–S145 [View Article] [PubMed]
    [Google Scholar]
  3. Husada D, Soegianto SDP, Kurniawati IS, Hendrata AP, Irawan E et al. First-line antibiotic susceptibility pattern of toxigenic Corynebacterium diphtheriae in Indonesia. BMC Infect Dis 2019; 19:1049 [View Article] [PubMed]
    [Google Scholar]
  4. Kneen R, Pham NG, Solomon T, Tran TM, Nguyen TT et al. Penicillin vs. erythromycin in the treatment of diphtheria. Clin Infect Dis 1998; 27:845–850 [View Article] [PubMed]
    [Google Scholar]
  5. Al-Dar AA, Al-Qassimi M, Ezzadeen FH, Qassime M, Al Murtadha AM et al. Diphtheria resurgence in Sada’a-Yemen, 2017-2020. BMC Infect Dis 2022; 22:46 [View Article] [PubMed]
    [Google Scholar]
  6. Sein C, Tiwari T, Macneil A, Wannemuehler K, Soulaphy C et al. Diphtheria outbreak in Lao People’s democratic republic, 2012-2013. Vaccine 2016; 34:4321–4326 [View Article] [PubMed]
    [Google Scholar]
  7. Strauss RA, Herrera-Leon L, Guillén AC, Castro JS, Lorenz E et al. Molecular and epidemiologic characterization of the diphtheria outbreak in Venezuela. Sci Rep 2021; 11:6378 [View Article] [PubMed]
    [Google Scholar]
  8. Weil LM, Williams MM, Shirin T, Lawrence M, Habib ZH et al. Investigation of a large diphtheria outbreak and cocirculation of Corynebacterium pseudodiphtheriticum among forcibly displaced Myanmar nationals, 2017-2019. J Infect Dis 2021; 224:318–325 [View Article] [PubMed]
    [Google Scholar]
  9. Polonsky JA, Ivey M, Mazhar MKA, Rahman Z, le Polain de Waroux O et al. Epidemiological, clinical, and public health response characteristics of a large outbreak of diphtheria among the Rohingya population in Cox’s Bazar, Bangladesh, 2017 to 2019: a retrospective study. PLoS Med 2021; 18:e1003587 [View Article] [PubMed]
    [Google Scholar]
  10. Benamrouche N, Hasnaoui S, Badell E, Guettou B, Lazri M et al. Microbiological and molecular characterization of Corynebacterium diphtheriae isolated in Algeria between 1992 and 2015. Clin Microbiol Infect 2016; 22:1005 [View Article] [PubMed]
    [Google Scholar]
  11. Both L, Collins S, de Zoysa A, White J, Mandal S et al. Molecular and epidemiological review of toxigenic diphtheria infections in England between 2007 and 2013. J Clin Microbiol 2015; 53:567–572 [View Article] [PubMed]
    [Google Scholar]
  12. du Plessis M, Wolter N, Allam M, de Gouveia L, Moosa F et al. Molecular characterization of Corynebacterium diphtheriae outbreak isolates, South Africa, march-june 2015. Emerg Infect Dis 2017; 23:1308–1315 [View Article] [PubMed]
    [Google Scholar]
  13. Grosse-Kock S, Kolodkina V, Schwalbe EC, Blom J, Burkovski A et al. Genomic analysis of endemic clones of toxigenic and non-toxigenic Corynebacterium diphtheriae in Belarus during and after the major epidemic in 1990s. BMC Genomics 2017; 18:873 [View Article] [PubMed]
    [Google Scholar]
  14. Mohd Khalid MKN, Ahmad N, Hii SYF, Abd Wahab MA, Hashim R et al. Molecular characterization of Corynebacterium diphtheriae isolates in Malaysia between 1981 and 2016. J Med Microbiol 2019; 68:105–110 [View Article] [PubMed]
    [Google Scholar]
  15. Paveenkittiporn W, Sripakdee S, Koobkratok O, Sangkitporn S, Kerdsin A. Molecular epidemiology and antimicrobial susceptibility of outbreak-associated Corynebacterium diphtheriae in Thailand, 2012. Infect Genet Evol 2019; 75:104007 [View Article] [PubMed]
    [Google Scholar]
  16. Meinel DM, Kuehl R, Zbinden R, Boskova V, Garzoni C et al. Outbreak investigation for toxigenic Corynebacterium diphtheriae wound infections in refugees from Northeast Africa and Syria in Switzerland and Germany by whole genome sequencing. Clin Microbiol Infect 2016; 22:1003 [View Article] [PubMed]
    [Google Scholar]
  17. Dangel A, Berger A, Konrad R, Bischoff H, Sing A. Geographically diverse clusters of nontoxigenic Corynebacterium diphtheriae infection, Germany, 2016-2017. Emerg Infect Dis 2018; 24:1239–1245 [View Article] [PubMed]
    [Google Scholar]
  18. Timms VJ, Nguyen T, Crighton T, Yuen M, Sintchenko V. Genome-wide comparison of Corynebacterium diphtheriae isolates from Australia identifies differences in the Pan-genomes between respiratory and cutaneous strains. BMC Genomics 2018; 19:869 [View Article] [PubMed]
    [Google Scholar]
  19. Seth-Smith HMB, Egli A. Whole genome sequencing for surveillance of diphtheria in low incidence settings. Front Public Health 2019; 7:235 [View Article] [PubMed]
    [Google Scholar]
  20. Chorlton SD, Ritchie G, Lawson T, Romney MG, Lowe CF. Whole-genome sequencing of Corynebacterium diphtheriae isolates recovered from an inner-city population demonstrates the predominance of a single molecular strain. J Clin Microbiol 2020; 58:e01651-19 [View Article] [PubMed]
    [Google Scholar]
  21. Hennart M, Panunzi LG, Rodrigues C, Gaday Q, Baines SL et al. Population genomics and antimicrobial resistance in Corynebacterium diphtheriae. Genome Med 2020; 12:107 [View Article] [PubMed]
    [Google Scholar]
  22. Xiaoli L, Benoliel E, Peng Y, Aneke J, Cassiday PK et al. Genomic epidemiology of nontoxigenic Corynebacterium diphtheriae from King County, Washington State, USA between July 2018 and May 2019. Microb Genom 2020; 6:12 [View Article] [PubMed]
    [Google Scholar]
  23. Badell E, Alharazi A, Criscuolo A, Almoayed KAA, Lefrancq N et al. Ongoing diphtheria outbreak in Yemen: a cross-sectional and genomic epidemiology study. Lancet Microbe 2021; 2:e386–e396 [View Article] [PubMed]
    [Google Scholar]
  24. Guglielmini J, Hennart M, Badell E, Toubiana J, Criscuolo A et al. Genomic epidemiology and strain taxonomy of Corynebacterium diphtheriae. J Clin Microbiol 2021; 59:e0158121 [View Article] [PubMed]
    [Google Scholar]
  25. Bernard K. The genus Corynebacterium and other medically relevant coryneform-like bacteria. J Clin Microbiol 2012; 50:3152–3158 [View Article] [PubMed]
    [Google Scholar]
  26. Maximescu P, Oprişan A, Pop A, Potorac E. Further studies on Corynebacterium species capable of producing diphtheria toxin (C. diphtheriae, C. ulcerans, C. ovis). J Gen Microbiol 1974; 82:49–56 [View Article] [PubMed]
    [Google Scholar]
  27. Sekizuka T, Yamamoto A, Komiya T, Kenri T, Takeuchi F et al. Corynebacterium ulcerans 0102 carries the gene encoding diphtheria toxin on a prophage different from the C. diphtheriae NCTC 13129 prophage. BMC Microbiol 2012; 12:72 [View Article] [PubMed]
    [Google Scholar]
  28. Heggelund L, Gaustad P, Håvelsrud OE, Blom J, Borgen L et al. Corynebacterium pseudotuberculosis Pneumonia in a veterinary student infected during laboratory work. Open Forum Infect Dis 2015; 2:fv053 [View Article] [PubMed]
    [Google Scholar]
  29. Keslin MH, McCoy EL, McCusker JJ, Lutch JS. Corynebacterium pseudotuberculosis. A new cause of infectious and eosinophilic pneumonia. Am J Med 1979; 67:228–231 [View Article] [PubMed]
    [Google Scholar]
  30. Zasada AA, Mosiej E. Contemporary microbiology and identification of Corynebacteria spp. causing infections in human. Lett Appl Microbiol 2018; 66:472–483 [View Article] [PubMed]
    [Google Scholar]
  31. Caldwell ST, Watson MT. Hospital survey of acute pesticide poisoning in South Carolina 1971-1973. J S C Med Assoc 1975; 71:249–252 [PubMed]
    [Google Scholar]
  32. Colt HG, Morris JF, Marston BJ, Sewell DL. Necrotizing tracheitis caused by Corynebacterium pseudodiphtheriticum: unique case and review. Rev Infect Dis 1991; 13:73–76 [View Article] [PubMed]
    [Google Scholar]
  33. Martaresche C, Fournier PE, Jacomo V, Gainnier M, Boussuge A et al. A case of Corynebacterium pseudodiphtheriticum nosocomial pneumonia. Emerg Infect Dis 1999; 5:722–723 [View Article] [PubMed]
    [Google Scholar]
  34. Kawasaki Y, Matsubara K, Ishihara H, Nigami H, Iwata A et al. Corynebacterium propinquum as the first cause of infective endocarditis in childhood. J Infect Chemother 2014; 20:317–319 [View Article] [PubMed]
    [Google Scholar]
  35. Jangda U, Upadhyay A, Bagheri F, Patel NR, Mendelson RI. Corynebacterium propinquum: a rare cause of prosthetic valve endocarditis. Case Rep Med 2016; 2016:1391789 [View Article] [PubMed]
    [Google Scholar]
  36. Malkoçoğlu G, Gencer H, Kaya A, Dalgıç N, Bulut ME et al. Corynebacterium propinquum bronchopneumonia in a child with ataxia telangiectasia. Turk J Pediatr 2016; 58:558–561 [View Article] [PubMed]
    [Google Scholar]
  37. Santos MR, Gandhi S, Vogler M, Hanna BA, Holzman RS. Suspected diphtheria in an Uzbek national: isolation of Corynebacterium pseudodiphtheriticum resulted in a false-positive presumptive diagnosis. Clin Infect Dis 1996; 22:735 [View Article] [PubMed]
    [Google Scholar]
  38. Izurieta HS, Strebel PM, Youngblood T, Hollis DG, Popovic T. Exudative pharyngitis possibly due to Corynebacterium pseudodiphtheriticum, a new challenge in the differential diagnosis of diphtheria. Emerg Infect Dis 1997; 3:65–68 [View Article] [PubMed]
    [Google Scholar]
  39. Indumathi VA, Shikha R, Suryaprakash DR. Diphtheria-like illness in a fully immunised child caused by Corynebacterium pseudodiphtheriticum. Indian J Med Microbiol 2014; 32:443–445 [View Article] [PubMed]
    [Google Scholar]
  40. Reddy K, Gericke S, Rabie H, Pienaar C, Maloba M. Exudative pharyngitis and Corynebacterium pseudodiphtheriticum: a case report and review of the literature. S Afr J Infect Dis 2021; 36:225 [View Article] [PubMed]
    [Google Scholar]
  41. Burkovski A. Corynebacterium pseudodiphtheriticum: putative probiotic, opportunistic infector, emerging pathogen. Virulence 2015; 6:673–674 [View Article] [PubMed]
    [Google Scholar]
  42. Xiaoli L, Peng Y, Williams MM, Cassiday PK, Nobles S et al. Complete genome sequences of four macrolide-resistant nondiphtheritic Corynebacterium isolates. Microbiol Resour Announc 2022; 11:e0049222 [View Article] [PubMed]
    [Google Scholar]
  43. Andrews S. FastQC: a quality control tool for high throughput sequencing data; 2010 https://www.bioinformatics.babraham.ac.uk/projects/fastqc/
  44. Martin M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet J 2011; 17:10 [View Article]
    [Google Scholar]
  45. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M et al. SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 2012; 19:455–477 [View Article] [PubMed]
    [Google Scholar]
  46. Gurevich A, Saveliev V, Vyahhi N, Tesler G. QUAST: quality assessment tool for genome assemblies. Bioinformatics 2013; 29:1072–1075 [View Article] [PubMed]
    [Google Scholar]
  47. Krumsiek J, Arnold R, Rattei T. Gepard: a rapid and sensitive tool for creating dotplots on genome scale. Bioinformatics 2007; 23:1026–1028 [View Article] [PubMed]
    [Google Scholar]
  48. Tatusova T, DiCuccio M, Badretdin A, Chetvernin V, Nawrocki EP et al. NCBI prokaryotic genome annotation pipeline. Nucleic Acids Res 2016; 44:6614–6624 [View Article] [PubMed]
    [Google Scholar]
  49. Inouye M, Dashnow H, Raven L-A, Schultz MB, Pope BJ et al. SRST2: rapid genomic surveillance for public health and hospital microbiology labs. Genome Med 2014; 6:90 [View Article] [PubMed]
    [Google Scholar]
  50. Gupta A, Jordan IK, Rishishwar L. stringMLST: a fast k-mer based tool for multilocus sequence typing. Bioinformatics 2017; 33:119–121 [View Article] [PubMed]
    [Google Scholar]
  51. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J et al. BLAST+: architecture and applications. BMC Bioinformatics 2009; 10:421 [View Article] [PubMed]
    [Google Scholar]
  52. Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat Methods 2012; 9:357–359 [View Article] [PubMed]
    [Google Scholar]
  53. Siguier P, Perochon J, Lestrade L, Mahillon J, Chandler M. ISfinder: the reference centre for bacterial insertion sequences. Nucleic Acids Res 2006; 34:D32–D36 [View Article] [PubMed]
    [Google Scholar]
  54. Sullivan MJ, Petty NK, Beatson SA. Easyfig: a genome comparison visualizer. Bioinformatics 2011; 27:1009–1010 [View Article] [PubMed]
    [Google Scholar]
  55. Seemann T. snippy: fast bacterial variant calling from NGS reads; 2015 https://github.com/tseemann/snippy
  56. Stamatakis A. RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 2014; 30:1312–1313 [View Article] [PubMed]
    [Google Scholar]
  57. Letunic I, Bork P. Interactive Tree Of Life (iTOL) v4: recent updates and new developments. Nucleic Acids Res 2019; 47:W256–W259 [View Article] [PubMed]
    [Google Scholar]
  58. Seemann T. SNP-dists: pairwise SNP distances from FASTA sequence alignments; 2018 https://github.com/tseemann/snp-dists
  59. Marçais G, Delcher AL, Phillippy AM, Coston R, Salzberg SL et al. MUMmer4: a fast and versatile genome alignment system. PLoS Comput Biol 2018; 14:e1005944 [View Article] [PubMed]
    [Google Scholar]
  60. Alikhan N-F, Petty NK, Ben Zakour NL, Beatson SA. BLAST Ring Image Generator (BRIG): simple prokaryote genome comparisons. BMC Genomics 2011; 12:402 [View Article] [PubMed]
    [Google Scholar]
  61. Thorvaldsdóttir H, Robinson JT, Mesirov JP. Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration. Brief Bioinform 2013; 14:178–192 [View Article] [PubMed]
    [Google Scholar]
  62. Seemann T. Prokka: rapid prokaryotic genome annotation. Bioinformatics 2014; 30:2068–2069 [View Article] [PubMed]
    [Google Scholar]
  63. Page AJ, Cummins CA, Hunt M, Wong VK, Reuter S et al. Roary: rapid large-scale prokaryote pan genome analysis. Bioinformatics 2015; 31:3691–3693 [View Article] [PubMed]
    [Google Scholar]
  64. Will RC, Ramamurthy T, Sharma NC, Veeraraghavan B, Sangal L et al. Spatiotemporal persistence of multiple, diverse clades and toxins of Corynebacterium diphtheriae. Nat Commun 2021; 12:1500 [View Article] [PubMed]
    [Google Scholar]
  65. Finger F, Funk S, White K, Siddiqui MR, Edmunds WJ et al. Real-time analysis of the diphtheria outbreak in forcibly displaced Myanmar nationals in Bangladesh. BMC Med 2019; 17:58 [View Article] [PubMed]
    [Google Scholar]
  66. Rahman MR, Islam K. Massive diphtheria outbreak among Rohingya refugees: lessons learnt. J Travel Med 2019; 26: [View Article] [PubMed]
    [Google Scholar]
  67. Hii SYF, Ahmad N, Hashim R, Liow YL, Abd Wahab MA et al. A SNP-based phylogenetic analysis of Corynebacterium diphtheriae in Malaysia. BMC Res Notes 2018; 11:760 [View Article] [PubMed]
    [Google Scholar]
  68. Dureab F, Al-Sakkaf M, Ismail O, Kuunibe N, Krisam J et al. Diphtheria outbreak in Yemen: the impact of conflict on a fragile health system. Confl Health 2019; 13:19 [View Article] [PubMed]
    [Google Scholar]
  69. Serwold-Davis TM, Groman N, Rabin M. Transformation of Corynebacterium diphtheriae, Corynebacterium ulcerans, Corynebacterium glutamicum, and Escherichia coli with the C. diphtheriae plasmid pNG2. Proc Natl Acad Sci 1987; 84:4964–4968 [View Article] [PubMed]
    [Google Scholar]
  70. Muthuirulandi Sethuvel DP, Subramanian N, Pragasam AK, Inbanathan FY, Gupta P et al. Insights to the diphtheria toxin encoding prophages amongst clinical isolates of Corynebacterium diphtheriae from India. Indian J Med Microbiol 2019; 37:423–425 [View Article] [PubMed]
    [Google Scholar]
  71. Tauch A, Bischoff N, Brune I, Kalinowski J. Insights into the genetic organization of the Corynebacterium diphtheriae erythromycin resistance plasmid pNG2 deduced from its complete nucleotide sequence. Plasmid 2003; 49:63–74 [View Article] [PubMed]
    [Google Scholar]
  72. Barraud O, Badell E, Denis F, Guiso N, Ploy M-C. Antimicrobial drug resistance in Corynebacterium diphtheriae mitis. Emerg Infect Dis 2011; 17:2078–2080 [View Article] [PubMed]
    [Google Scholar]
  73. Díez-Aguilar M, Ruiz-Garbajosa P, Fernández-Olmos A, Guisado P, Del Campo R et al. Non-diphtheriae Corynebacterium species: an emerging respiratory pathogen. Eur J Clin Microbiol Infect Dis 2013; 32:769–772 [View Article] [PubMed]
    [Google Scholar]
  74. Bernard K, Pacheco AL, Cunningham I, Gill N, Burdz T et al. Emendation of the description of the species Corynebacterium propinquum to include strains which produce urease. Int J Syst Evol Microbiol 2013; 63:2146–2154 [View Article] [PubMed]
    [Google Scholar]
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