1887

Abstract

is the clinically most important species within the genus and, as a result of the continuous emergence of multi-drug resistant (MDR) strains, the cause of severe nosocomial infections. The decline in the effectiveness of antibiotic treatments for infections caused by MDR bacteria has generated particular interest in the study of bacteriophages. In this study, we characterized a total of 40 temperate bacteriophages (prophages) with a genome range of 11.454–84.199 kb, predicted from 16 carbapenemase-producing clinical strains of belonging to different sequence types, previously identified by multilocus sequence typing. These prophages were grouped into the three families in the order (27 prophages belonging to the family , 10 prophages belonging to the family and 3 prophages belonging to the family ). Genomic comparison of the 40 prophage genomes led to the identification of four prophages isolated from different strains and of genome sizes of around 33.3, 36.1, 39.6 and 42.6 kb. These prophages showed sequence similarities (query cover >90 %, identity >99.9 %) with international Microbe Versus Phage (MVP) (http://mvp.medgenius.info/home) clusters 4762, 4901, 3499 and 4280, respectively. Phylogenetic analysis revealed the evolutionary proximity among the members of the four groups of the most frequently identified prophages in the bacterial genomes studied (33.3, 36.1, 39.6 and 42.6 kb), with bootstrap values of 100 %. This allowed the prophages to be classified into three clusters: A, B and C. Interestingly, these temperate bacteriophages did not infect the highest number of strains as indicated by a host-range assay, these results could be explained by the development of superinfection exclusion mechanisms. In addition, bioinformatic analysis of the 40 identified prophages revealed the presence of 2363 proteins. In total, 59.7 % of the proteins identified had a predicted function, mainly involving viral structure, transcription, replication and regulation (lysogenic/lysis). Interestingly, some proteins had putative functions associated with bacterial virulence (toxin expression and efflux pump regulators), phage defence profiles such as toxin–antitoxin modules, an anti-CRISPR/Cas9 protein, TerB protein (from ter operon) and methyltransferase proteins.

Funding
This study was supported by the:
  • Instituto de Salud Carlos III, http://dx.doi.org/10.13039/501100004587 (Award PI19/00878)
  • Instituto de Salud Carlos III, http://dx.doi.org/10.13039/501100004587 (Award PI16/01163)
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License.
Loading

Article metrics loading...

/content/journal/mgen/10.1099/mgen.0.000369
2020-04-29
2024-05-05
Loading full text...

Full text loading...

/deliver/fulltext/mgen/6/5/mgen000369.html?itemId=/content/journal/mgen/10.1099/mgen.0.000369&mimeType=html&fmt=ahah

References

  1. Shabbir MAB, Hao H, Shabbir MZ, Wu Q, Sattar A et al. Bacteria vs. bacteriophages: parallel evolution of immune arsenals. Front Microbiol 2016; 7:1292 [View Article]
    [Google Scholar]
  2. Bergh Ø, Børsheim KY, Bratbak G, Heldal M. High abundance of viruses found in aquatic environments. Nature 1989; 340:467–468 [View Article]
    [Google Scholar]
  3. Wommack KE, Colwell RR. Virioplankton: viruses in aquatic ecosystems. Microbiol Mol Biol Rev 2000; 64:69–114 [View Article]
    [Google Scholar]
  4. Taylor VL, Fitzpatrick AD, Islam Z, Maxwell KL. The diverse impacts of phage morons on bacterial fitness and virulence. Adv Virus Res 2019; 103:1–31
    [Google Scholar]
  5. Breitbart M, Rohwer F. Here a virus, there a virus, everywhere the same virus?. Trends Microbiol 2005; 13:278–284 [View Article]
    [Google Scholar]
  6. Brister JR, Le Mercier P, Hu JC. Microbial virus genome annotation — mustering the troops to fight the sequence onslaught. Virology 2012; 434:175–180 [View Article]
    [Google Scholar]
  7. Pope WH, Bowman CA, Russell DA, Jacobs-Sera D, Asai DJ et al. Whole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversity. eLife 2015; 4:e06416 [View Article]
    [Google Scholar]
  8. López M, Rueda A, Florido JP, Blasco L, Fernández-García L et al. Evolution of the Quorum network and the mobilome (plasmids and bacteriophages) in clinical strains of Acinetobacter baumannii during a decade. Sci Rep 2018; 8:2523 [View Article]
    [Google Scholar]
  9. Kwan T, Liu J, DuBow M, Gros P, Pelletier J. Comparative genomic analysis of 18 Pseudomonas aeruginosa bacteriophages. J Bacteriol 2006; 188:1184–1187 [View Article]
    [Google Scholar]
  10. Grose JH, Jensen GL, Burnett SH, Breakwell DP. Genomic comparison of 93 Bacillus phages reveals 12 clusters, 14 singletons and remarkable diversity. BMC Genomics 2014; 15:855 [View Article]
    [Google Scholar]
  11. Deveau H, Labrie SJ, Chopin M-C, Moineau S. Biodiversity and classification of lactococcal phages. Appl Environ Microbiol 2006; 72:4338–4346 [View Article]
    [Google Scholar]
  12. Gregory AC, Solonenko SA, Ignacio-Espinoza JC, LaButti K, Copeland A et al. Genomic differentiation among wild cyanophages despite widespread horizontal gene transfer. BMC Genomics 2016; 17:930 [View Article]
    [Google Scholar]
  13. Switt AI, Sulakvelidze A, Wiedmann M, Kropinski AM, Wishart DS et al. Salmonella phages and prophages: genomics, taxonomy, and applied aspects. Methods Mol Biol 2015; 1225:237–287
    [Google Scholar]
  14. Kalatzis PG, Rørbo NI, Castillo D, Mauritzen JJ, Jørgensen J et al. Stumbling across the same phage: comparative genomics of widespread temperate phages infecting the fish pathogen Vibrio anguillarum. Viruses 2017; 9:122 [View Article]
    [Google Scholar]
  15. Grose JH, Casjens SR. Understanding the enormous diversity of bacteriophages: the tailed phages that infect the bacterial family Enterobacteriaceae. Virology 2014; 468-470:421–443 [View Article]
    [Google Scholar]
  16. Hendrix RW, Smith MCM, Burns RN, Ford ME, Hatfull GF. Evolutionary relationships among diverse bacteriophages and prophages: all the world's a phage. Proc Natl Acad Sci USA 1999; 96:2192–2197 [View Article]
    [Google Scholar]
  17. Dion MB, Oechslin F, Moineau S. Phage diversity, genomics and phylogeny. Nat Rev Microbiol 2020; 18:125–138
    [Google Scholar]
  18. Hatfull GF. Dark matter of the biosphere: the amazing world of bacteriophage diversity. J Virol 2015; 89:8107–8110 [View Article]
    [Google Scholar]
  19. de Jonge PA, Nobrega FL, Brouns SJJ, Dutilh BE. Molecular and evolutionary determinants of bacteriophage host range. Trends Microbiol 2019; 27:51–63 [View Article]
    [Google Scholar]
  20. Argov T, Azulay G, Pasechnek A, Stadnyuk O, Ran-Sapir S et al. Temperate bacteriophages as regulators of host behavior. Curr Opin Microbiol 2017; 38:81–87 [View Article]
    [Google Scholar]
  21. Pratama AA, van Elsas JD. Gene mobility in microbiomes of the mycosphere and mycorrhizosphere – role of plasmids and bacteriophages. FEMS Microbiol Ecol 2019; 95:fiz053 [View Article]
    [Google Scholar]
  22. Wiedenbeck J, Cohan FM. Origins of bacterial diversity through horizontal genetic transfer and adaptation to new ecological niches. FEMS Microbiol Rev 2011; 35:957–976 [View Article]
    [Google Scholar]
  23. Brüssow H, Canchaya C, Hardt W-D. Phages and the evolution of bacterial pathogens: from genomic rearrangements to lysogenic conversion. Microbiol Mol Biol Rev 2004; 68:560–602 [View Article]
    [Google Scholar]
  24. Gorrie CL, Mirceta M, Wick RR, Judd LM, Wyres KL et al. Antimicrobial-resistant Klebsiella pneumoniae carriage and infection in specialized geriatric care wards linked to acquisition in the referring hospital. Clin Infect Dis 2018; 67:161–170 [View Article]
    [Google Scholar]
  25. Pitout JDD, Peirano G, Kock MM, Strydom K-A, Matsumura Y. The global ascendency of OXA-48-type carbapenemases. Clin Microbiol Rev 2019; 33:e00102-19 [View Article]
    [Google Scholar]
  26. Pendleton JN, Gorman SP, Gilmore BF. Clinical relevance of the ESKAPE pathogens. Expert Rev Anti Infect Ther 2013; 11:297–308 [View Article]
    [Google Scholar]
  27. Vuotto C, Longo F, Balice M, Donelli G, Varaldo P. Antibiotic resistance related to biofilm formation in Klebsiella pneumoniae. Pathogens 2014; 3:743–758 [View Article]
    [Google Scholar]
  28. Bengoechea JA, Sa Pessoa J. Klebsiella pneumoniae infection biology: living to counteract host defences. FEMS Microbiol Rev 2019; 43:123–144 [View Article]
    [Google Scholar]
  29. Duprilot M, Decre D, Genel N, Drieux L, Sougakoff W et al. Diversity and functionality of plasmid-borne VagCD toxin-antitoxin systems of Klebsiella pneumoniae. J Antimicrob Chemother 2017; 72:1320–1326
    [Google Scholar]
  30. Esteban-Cantos A, Aracil B, Bautista V, Ortega A, Lara N et al. The carbapenemase-producing Klebsiella pneumoniae population is distinct and more clonal than the carbapenem-susceptible population. Antimicrob Agents Chemother 2017; 61:e02520-16 [View Article]
    [Google Scholar]
  31. 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]
    [Google Scholar]
  32. Gao NL, Zhang C, Zhang Z, Hu S, Lercher MJ et al. MVP: a microbe–phage interaction database. Nucleic Acids Res 2018; 46:D700–D707 [View Article]
    [Google Scholar]
  33. 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]
    [Google Scholar]
  34. Black LW. Old, new, and widely true: the bacteriophage T4 DNA packaging mechanism. Virology 2015; 479-480:650–656 [View Article][PubMed][PubMed]
    [Google Scholar]
  35. Kumar S, Stecher G, Li M, Knyaz C, Tamura K. MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol 2018; 35:1547–1549 [View Article]
    [Google Scholar]
  36. Bardina C, Colom J, Spricigo DA, Otero J, Sánchez-Osuna M et al. Genomics of three new bacteriophages useful in the biocontrol of Salmonella. Front Microbiol 2016; 7:545 [View Article]
    [Google Scholar]
  37. Zabarovskiĭ ER, Turina OV. Rapid isolation of phage lambda DNA. Mol Biol 1988; 22:1451–1455
    [Google Scholar]
  38. Raya RR, H'bert EM. Isolation of phage via induction of lysogens. Methods Mol Biol 2009; 501:23–32
    [Google Scholar]
  39. Zimmermann L, Stephens A, Nam S-Z, Rau D, Kübler J et al. A completely reimplemented MPI bioinformatics toolkit with a new HHpred server at its core. J Mol Biol 2018; 430:2237–2243 [View Article]
    [Google Scholar]
  40. Hildebrand A, Remmert M, Biegert A, Söding J. Fast and accurate automatic structure prediction with HHpred. Proteins 2009; 77:128–132 [View Article]
    [Google Scholar]
  41. Meier-Kolthoff JP, Göker M. VICTOR: genome-based phylogeny and classification of prokaryotic viruses. Bioinformatics 2017; 33:3396–3404 [View Article]
    [Google Scholar]
  42. Cahill J, Young R. Phage lysis: multiple genes for multiple barriers. Adv Virus Res 2019; 103:33–70
    [Google Scholar]
  43. Rivera-Calzada A, Fronzes R, Savva CG, Chandran V, Lian PW et al. Structure of a bacterial type IV secretion core complex at subnanometre resolution. EMBO J 2013; 32:1195–1204 [View Article]
    [Google Scholar]
  44. Nguyen KB, Sreelatha A, Durrant ES, Lopez-Garrido J, Muszewska A et al. Phosphorylation of spore coat proteins by a family of atypical protein kinases. Proc Natl Acad Sci USA 2016; 113:E3482–E3491 [View Article]
    [Google Scholar]
  45. Kim DJ, Park K-S, Kim J-H, Yang S-H, Yoon JY et al. Helicobacter pylori proinflammatory protein up-regulates NF-kappaB as a cell-translocating Ser/Thr kinase. Proc Natl Acad Sci USA 2010; 107:21418–21423 [View Article]
    [Google Scholar]
  46. Deochand DK, Grove A. MarR family transcription factors: dynamic variations on a common scaffold. Crit Rev Biochem Mol Biol 2017; 52:595–613 [View Article]
    [Google Scholar]
  47. Yao J, Guo Y, Wang P, Zeng Z, Li B et al. Type II toxin/antitoxin system ParE. Environ Microbiol 2018; 20:1224–1239
    [Google Scholar]
  48. Harms A, Brodersen DE, Mitarai N, Gerdes K. Toxins, targets, and triggers: an overview of toxin-antitoxin biology. Mol Cell 2018; 70:768–784 [View Article]
    [Google Scholar]
  49. Chiang S-K, Lou Y-C, Chen C. NMR solution structure of KP-TerB, a tellurite-resistance protein from Klebsiella pneumoniae. Protein Sci 2008; 17:785–789 [View Article]
    [Google Scholar]
  50. Bailly-Bechet M, Vergassola M, Rocha E. Causes for the intriguing presence of tRNAs in phages. Genome Res 2007; 17:1486–1495 [View Article]
    [Google Scholar]
  51. Casjens SR. Comparative genomics and evolution of the tailed-bacteriophages. Curr Opin Microbiol 2005; 8:451–458 [View Article]
    [Google Scholar]
  52. Adriaenssens E, Brister JR. How to name and classify your phage: an informal guide. Viruses 2017; 9:70 [View Article]
    [Google Scholar]
  53. Maciejewska B, Roszniowski B, Espaillat A, Kęsik-Szeloch A, Majkowska-Skrobek G et al. Klebsiella phages representing a novel clade of viruses with an unknown DNA modification and biotechnologically interesting enzymes. Appl Microbiol Biotechnol 2017; 101:673–684 [View Article]
    [Google Scholar]
  54. López M, Rueda A, Florido JP, Blasco L, Gato E et al. Genomic evolution of two Acinetobacter baumannii clinical strains from ST-2 clones isolated in 2000 and 2010 (ST-2_clon_2000 and ST-2_clon_2010). Genome Announc 2016; 4:e01182-16 [View Article]
    [Google Scholar]
  55. Folimonova SY. Superinfection exclusion is an active virus-controlled function that requires a specific viral protein. J Virol 2012; 86:5554–5561 [View Article]
    [Google Scholar]
  56. Feiner R, Argov T, Rabinovich L, Sigal N, Borovok I et al. A new perspective on lysogeny: prophages as active regulatory switches of bacteria. Nat Rev Microbiol 2015; 13:641–650 [View Article]
    [Google Scholar]
  57. Bobay L-M, Rocha EPC, Touchon M. The adaptation of temperate bacteriophages to their host genomes. Mol Biol Evol 2013; 30:737–751 [View Article]
    [Google Scholar]
  58. Roszniowski B, McClean S, Drulis-Kawa Z. Burkholderia cenocepacia prophages - prevalence, chromosome location and major genes involved. Viruses 2018; 10:297
    [Google Scholar]
  59. Hatfull GF, Hendrix RW. Bacteriophages and their genomes. Curr Opin Virol 2011; 1:298–303 [View Article]
    [Google Scholar]
  60. Van den Bossche A, Ceyssens P-J, De Smet J, Hendrix H, Bellon H et al. Systematic identification of hypothetical bacteriophage proteins targeting key protein complexes of Pseudomonas aeruginosa. J Proteome Res 2014; 13:4446–4456 [View Article]
    [Google Scholar]
  61. Klumpp J, Fouts DE, Sozhamannan S. Bacteriophage functional genomics and its role in bacterial pathogen detection. Brief Funct Genomics 2013; 12:354–365 [View Article]
    [Google Scholar]
  62. Tabassum R, Shafique M, Khawaja KA, Alvi IA, Rehman Y et al. Complete genome analysis of a Siphoviridae phage TSK1 showing biofilm removal potential against Klebsiella pneumoniae. Sci Rep 2018; 8:17904 [View Article]
    [Google Scholar]
  63. Xu Y, Yu X, Gu Y, Huang X, Liu G et al. Characterization and genomic study of phage vB_EcoS-B2 infecting multidrug-resistant Escherichia coli. Front Microbiol 2018; 9:793 [View Article]
    [Google Scholar]
  64. Blasco L, Ambroa A, Lopez M, Fernandez-Garcia L, Bleriot I et al. Combined use of the Ab105-2φΔCI lytic mutant phage and different antibiotics in clinical isolates of multi-resistant Acinetobacter baumannii. Microorganisms 2019; 7:556 [View Article]
    [Google Scholar]
  65. Dedrick RM, Guerrero-Bustamante CA, Garlena RA, Russell DA, Ford K et al. Engineered bacteriophages for treatment of a patient with a disseminated drug-resistant Mycobacterium abscessus. Nat Med 2019; 25:730–733 [View Article]
    [Google Scholar]
  66. Fokine A, Rossmann MG. Molecular architecture of tailed double-stranded DNA phages. Bacteriophage 2014; 4:e28281 [View Article]
    [Google Scholar]
  67. Feiss M, Rao VB. The bacteriophage DNA packaging machine. Adv Exp Med Biol 2012; 726:489–509
    [Google Scholar]
  68. Boyd EF, Carpenter MR, Chowdhury N. Mobile effector proteins on phage genomes. Bacteriophage 2012; 2:139–148 [View Article]
    [Google Scholar]
  69. Liekniņa I, Kalniņš G, Akopjana I, Bogans J, Šišovs M et al. Production and characterization of novel ssRNA bacteriophage virus-like particles from metagenomic sequencing data. J Nanobiotechnol 2019; 17:61 [View Article]
    [Google Scholar]
  70. Palacios M, Miner TA, Frederick DR, Sepulveda VE, Quinn JD et al. Identification of two regulators of virulence that are conserved in Klebsiella pneumoniae classical and hypervirulent strains. mBio 2018; 9:e01443-18 [View Article]
    [Google Scholar]
  71. Yang QE, Walsh TR. Toxin–antitoxin systems and their role in disseminating and maintaining antimicrobial resistance. FEMS Microbiol Rev 2017; 41:343–353 [View Article]
    [Google Scholar]
  72. Bondy-Denomy J, Pawluk A, Maxwell KL, Davidson AR. Bacteriophage genes that inactivate the CRISPR/Cas bacterial immune system. Nature 2013; 493:429–432 [View Article]
    [Google Scholar]
  73. Maxwell KL. The anti-CRISPR story: a battle for survival. Mol Cell 2017; 68:8–14 [View Article]
    [Google Scholar]
  74. Hwang S, Maxwell KL. Meet the anti-CRISPRs: widespread protein inhibitors of CRISPR-Cas systems. CRISPR J 2019; 2:23–30 [View Article]
    [Google Scholar]
  75. Zhu Y, Gao A, Zhan Q, Wang Y, Feng H et al. Diverse mechanisms of CRISPR-Cas9 inhibition by type IIC anti-CRISPR proteins. Mol Cell 2019; 74:296–309 [View Article]
    [Google Scholar]
  76. Borges AL, Davidson AR, Bondy-Denomy J. The discovery, mechanisms, and evolutionary impact of anti-CRISPRs. Annu Rev Virol 2017; 4:37–59 [View Article]
    [Google Scholar]
  77. Hudaiberdiev S, Shmakov S, Wolf YI, Terns MP, Makarova KS et al. Phylogenomics of Cas4 family nucleases. BMC Evol Biol 2017; 17:232 [View Article]
    [Google Scholar]
  78. Chen Y-T, Chang H-Y, Lai Y-C, Pan C-C, Tsai S-F et al. Sequencing and analysis of the large virulence plasmid pLVPK of Klebsiella pneumoniae CG43. Gene 2004; 337:189–198 [View Article]
    [Google Scholar]
  79. Whelan KF, Colleran E, Taylor DE. Phage inhibition, colicin resistance, and tellurite resistance are encoded by a single cluster of genes on the IncHI2 plasmid R478. J Bacteriol 1995; 177:5016–5027 [View Article]
    [Google Scholar]
  80. Murphy J, Mahony J, Ainsworth S, Nauta A, van Sinderen D. Bacteriophage orphan DNA methyltransferases: insights from their bacterial origin, function, and occurrence. Appl Environ Microbiol 2013; 79:7547–7555 [View Article]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/mgen/10.1099/mgen.0.000369
Loading
/content/journal/mgen/10.1099/mgen.0.000369
Loading

Data & Media loading...

Supplements

Supplementary material 1

EXCEL
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error