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Coexistence and Within-Host Evolution of Diversified Lineages of Hypermutable in Long-term Cystic Fibrosis Infections


Patients with cystic fibrosis (CF) are often colonized by a single clone of the common, widespread bacterium Pseudomonas aeruginosa, resulting in chronic airway infections. Long-term persistence of the bacteria involves the emergence and selection of multiple phenotypic variants. Among these are “mutator” variants characterized by increased mutation rates resulting from the inactivation of DNA repair systems. The genetic evolution of mutators during the course of chronic infection is poorly understood, and the effects of hypermutability on bacterial population structure have not been studied using genomic approaches. We evaluated the genomic changes undergone by mutator populations of P. aeruginosa obtained from single sputum samples from two chronically infected CF patients, and found that mutators completely dominated the infecting population in both patients. These populations displayed high genomic diversity based on vast accumulation of stochastic mutations. Our results are in contrast to the concept of a homogeneous population consisting of a single dominant clone; rather, they support a model of populations structured by diverse subpopulations that coexist within the patient. Certain genes involved in adaptation were highly and convergently mutated in both lineages, suggesting that these genes were beneficial and potentially responsible for the co-selection of mutator alleles.


Vyšlo v časopise: Coexistence and Within-Host Evolution of Diversified Lineages of Hypermutable in Long-term Cystic Fibrosis Infections. PLoS Genet 10(10): e32767. doi:10.1371/journal.pgen.1004651
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1004651

Souhrn

Patients with cystic fibrosis (CF) are often colonized by a single clone of the common, widespread bacterium Pseudomonas aeruginosa, resulting in chronic airway infections. Long-term persistence of the bacteria involves the emergence and selection of multiple phenotypic variants. Among these are “mutator” variants characterized by increased mutation rates resulting from the inactivation of DNA repair systems. The genetic evolution of mutators during the course of chronic infection is poorly understood, and the effects of hypermutability on bacterial population structure have not been studied using genomic approaches. We evaluated the genomic changes undergone by mutator populations of P. aeruginosa obtained from single sputum samples from two chronically infected CF patients, and found that mutators completely dominated the infecting population in both patients. These populations displayed high genomic diversity based on vast accumulation of stochastic mutations. Our results are in contrast to the concept of a homogeneous population consisting of a single dominant clone; rather, they support a model of populations structured by diverse subpopulations that coexist within the patient. Certain genes involved in adaptation were highly and convergently mutated in both lineages, suggesting that these genes were beneficial and potentially responsible for the co-selection of mutator alleles.


Zdroje

1. BodeyGP, BolivarR, FainsteinV, JadejaL (1983) Infections caused by Pseudomonas aeruginosa. Rev Infect Dis 5: 279–313.

2. MorrisonAJJr, WenzelRP (1984) Epidemiology of infections due to Pseudomonas aeruginosa. Rev Infect Dis 6 Suppl 3: S627–642.

3. GovanJR, DereticV (1996) Microbial pathogenesis in cystic fibrosis: mucoid Pseudomonas aeruginosa and Burkholderia cepacia. Microbiol Rev 60: 539–574.

4. LyczakJB, CannonCL, PierGB (2002) Lung infections associated with cystic fibrosis. Clin Microbiol Rev 15: 194–222.

5. MahenthiralingamE, CampbellME, FosterJ, LamJS, SpeertDP (1996) Random amplified polymorphic DNA typing of Pseudomonas aeruginosa isolates recovered from patients with cystic fibrosis. J Clin Microbiol 34: 1129–1135.

6. RömlingU, WingenderJ, MullerH, TümmlerB (1994) A major Pseudomonas aeruginosa clone common to patients and aquatic habitats. Appl Environ Microbiol 60: 1734–1738.

7. MartinC, IchouMA, MassicotP, GoudeauA, QuentinR (1995) Genetic diversity of Pseudomonas aeruginosa strains isolated from patients with cystic fibrosis revealed by restriction fragment length polymorphism of the rRNA gene region. J Clin Microbiol 33: 1461–1466.

8. MowatE, PatersonS, FothergillJL, WrightEA, LedsonMJ, et al. (2011) Pseudomonas aeruginosa population diversity and turnover in cystic fibrosis chronic infections. Am J Respir Crit Care Med 183: 1674–1679.

9. WorkentineML, SibleyCD, GlezersonB, PurighallaS, Norgaard-GronJC, et al. (2013) Phenotypic heterogeneity of Pseudomonas aeruginosa populations in a cystic fibrosis patient. PLoS One 8: e60225.

10. SmithEE, BuckleyDG, WuZ, SaenphimmachakC, HoffmanLR, et al. (2006) Genetic adaptation by Pseudomonas aeruginosa to the airways of cystic fibrosis patients. Proc Natl Acad Sci U S A 103: 8487–8492.

11. HuseH, KwonT, ZlosnikJ, SpeertD, MarcotteE, et al. (2010) Parallel Evolution in Pseudomonas aeruginosa over 39,000 Generations in vivo. MBio 1: e00199–00110.

12. WongA, KassenR (2011) Parallel evolution and local differentiation in quinolone resistance in Pseudomonas aeruginosa. Microbiology 157: 937–944.

13. OliverA, CantónR, CampoP, BaqueroF, BlázquezJ (2000) High frequency of hypermutable Pseudomonas aeruginosa in cystic fibrosis lung infection. Science 288: 1251–1254.

14. OliverA, MenaA (2010) Bacterial hypermutation in cystic fibrosis, not only for antibiotic resistance. Clin Microbiol Infect 16: 798–808.

15. OliverA, BaqueroF, BlázquezJ (2002) The mismatch repair system (mutS, mutL and uvrD genes) in Pseudomonas aeruginosa: molecular characterization of naturally occurring mutants. Mol Microbiol 43: 1641–1650.

16. CiofuO, RiisB, PresslerT, PoulsenHE, HøibyN (2005) Occurrence of hypermutable Pseudomonas aeruginosa in cystic fibrosis patients is associated with the oxidative stress caused by chronic lung inflammation. Antimicrob Agents Chemother 49: 2276–2282.

17. FelizianiS, LujánAM, MoyanoAJ, SolaC, BoccoJL, et al. (2010) Mucoidy, quorum sensing, mismatch repair and antibiotic resistance in Pseudomonas aeruginosa from cystic fibrosis chronic airways infections. PLoS One 5.

18. MarvigRL, JohansenHK, MolinS, JelsbakL (2013) Genome analysis of a transmissible lineage of Pseudomonas aeruginosa reveals pathoadaptive mutations and distinct evolutionary paths of hypermutators. PLoS Genet 9: e1003741.

19. WaineDJ, HoneybourneD, SmithEG, WhitehouseJL, DowsonCG (2008) Association between hypermutator phenotype, clinical variables, mucoid phenotype, and antimicrobial resistance in Pseudomonas aeruginosa. J Clin Microbiol 46: 3491–3493.

20. TaddeiF, RadmanM, Maynard-SmithJ, ToupanceB, GouyonPH, et al. (1997) Role of mutator alleles in adaptive evolution. Nature 387: 700–702.

21. CoxEC, GibsonTC (1974) Selection for high mutation rates in chemostats. Genetics 77: 169–184.

22. SniegowskiPD, GerrishPJ, LenskiRE (1997) Evolution of high mutation rates in experimental populations of E. coli. Nature 387: 703–705.

23. MaoEF, LaneL, LeeJ, MillerJH (1997) Proliferation of mutators in A cell population. J Bacteriol 179: 417–422.

24. TenaillonO, ToupanceB, Le NagardH, TaddeiF, GodelleB (1999) Mutators, population size, adaptive landscape and the adaptation of asexual populations of bacteria. Genetics 152: 485–493.

25. GiraudA, MaticI, TenaillonO, ClaraA, RadmanM, et al. (2001) Costs and benefits of high mutation rates: adaptive evolution of bacteria in the mouse gut. Science 291: 2606–2608.

26. DenamurE, MaticI (2006) Evolution of mutation rates in bacteria. Mol Microbiol 60: 820–827.

27. OliverA, LevinBR, JuanC, BaqueroF, BlázquezJ (2004) Hypermutation and the preexistence of antibiotic-resistant Pseudomonas aeruginosa mutants: implications for susceptibility testing and treatment of chronic infections. Antimicrob Agents Chemother 48: 4226–4233.

28. MaciáMD, BlanquerD, TogoresB, SauledaJ, PérezJL, et al. (2005) Hypermutation is a key factor in development of multiple-antimicrobial resistance in Pseudomonas aeruginosa strains causing chronic lung infections. Antimicrob Agents Chemother 49: 3382–3386.

29. HenrichfreiseB, WiegandI, PfisterW, WiedemannB (2007) Resistance mechanisms of multiresistant Pseudomonas aeruginosa strains from Germany and correlation with hypermutation. Antimicrob Agents Chemother 51: 4062–4070.

30. PlasenciaV, BorrellN, MaciáMD, MoyaB, PérezJL, et al. (2007) Influence of high mutation rates on the mechanisms and dynamics of in vitro and in vivo resistance development to single or combined antipseudomonal agents. Antimicrob Agents Chemother 51: 2574–2581.

31. FerroniA, GuillemotD, MoumileK, BernedeC, Le BourgeoisM, et al. (2009) Effect of mutator P. aeruginosa on antibiotic resistance acquisition and respiratory function in cystic fibrosis. Pediatr Pulmonol 44: 820–825.

32. MenaA, SmithEE, BurnsJL, SpeertDP, MoskowitzSM, et al. (2008) Genetic adaptation of Pseudomonas aeruginosa to the airways of cystic fibrosis patients is catalyzed by hypermutation. J Bacteriol 190: 7910–7917.

33. CiofuO, MandsbergLF, BjarnsholtT, WassermannT, HøibyN (2010) Genetic adaptation of Pseudomonas aeruginosa during chronic lung infection of patients with cystic fibrosis: strong and weak mutators with heterogeneous genetic backgrounds emerge in mucA and/or lasR mutants. Microbiology 156: 1108–1119.

34. MoyanoAJ, LujánAM, ArgarañaCE, SmaniaAM (2007) MutS deficiency and activity of the error-prone DNA polymerase IV are crucial for determining mucA as the main target for mucoid conversion in Pseudomonas aeruginosa. Mol Microbiol 64: 547–559.

35. SmaniaAM, SeguraI, PezzaRJ, BecerraC, AlbesaI, et al. (2004) Emergence of phenotypic variants upon mismatch repair disruption in Pseudomonas aeruginosa. Microbiology 150: 1327–1338.

36. LujánAM, MoyanoAJ, SeguraI, ArgarañaCE, SmaniaAM (2007) Quorum-sensing-deficient (lasR) mutants emerge at high frequency from a Pseudomonas aeruginosa mutS strain. Microbiology 153: 225–237.

37. LujánAM, MaciáMD, YangL, MolinS, OliverA, et al. (2011) Evolution and adaptation in Pseudomonas aeruginosa biofilms driven by mismatch repair system-deficient mutators. PLoS One 6: e27842.

38. MoyanoAJ, SmaniaAM (2009) Simple sequence repeats and mucoid conversion: biased mucA mutagenesis in mismatch repair-deficient Pseudomonas aeruginosa. PLoS One 4: e8203.

39. MoyanoAJ, FelizianiS, Di RienzoJA, SmaniaAM (2013) Simple Sequence Repeats Together with Mismatch Repair Deficiency Can Bias Mutagenic Pathways in Pseudomonas aeruginosa during Chronic Lung Infection. PLoS One 8: e80514.

40. CramerN, KlockgetherJ, WrasmanK, SchmidtM, DavenportCF, et al. (2011) Microevolution of the major common Pseudomonas aeruginosa clones C and PA14 in cystic fibrosis lungs. Environ Microbiol 13: 1690–1704.

41. SmithEE, SimsEH, SpencerDH, KaulR, OlsonMV (2005) Evidence for diversifying selection at the pyoverdine locus of Pseudomonas aeruginosa. J Bacteriol 187: 2138–2147.

42. YangL, JelsbakL, MarvigRL, DamkiaerS, WorkmanCT, et al. (2011) Evolutionary dynamics of bacteria in a human host environment. Proc Natl Acad Sci U S A 108: 7481–7486.

43. ChungJC, BecqJ, FraserL, Schulz-TrieglaffO, BondNJ, et al. (2012) Genomic variation among contemporary Pseudomonas aeruginosa isolates from chronically infected cystic fibrosis patients. J Bacteriol 194: 4857–4866.

44. YangL, HaagensenJA, JelsbakL, JohansenHK, SternbergC, et al. (2008) In situ growth rates and biofilm development of Pseudomonas aeruginosa populations in chronic lung infections. J Bacteriol 190: 2767–2776.

45. WiehlmannL, WagnerG, CramerN, SiebertB, GudowiusP, et al. (2007) Population structure of Pseudomonas aeruginosa. Proc Natl Acad Sci U S A 104: 8101–8106.

46. DrummondAJ, SuchardMA, XieD, RambautA (2012) Bayesian phylogenetics with BEAUti and the BEAST 1.7. Mol Biol Evol 29: 1969–1973.

47. GodfreyAJ, BryanLE, RabinHR (1981) beta-Lactam-resistant Pseudomonas aeruginosa with modified penicillin-binding proteins emerging during cystic fibrosis treatment. Antimicrob Agents Chemother 19: 705–711.

48. KohlerT, Michea-HamzehpourM, HenzeU, GotohN, CurtyLK, et al. (1997) Characterization of MexE-MexF-OprN, a positively regulated multidrug efflux system of Pseudomonas aeruginosa. Mol Microbiol 23: 345–354.

49. LivermoreDM (1995) beta-Lactamases in laboratory and clinical resistance. Clin Microbiol Rev 8: 557–584.

50. MineT, MoritaY, KataokaA, MizushimaT, TsuchiyaT (1999) Expression in Escherichia coli of a new multidrug efflux pump, MexXY, from Pseudomonas aeruginosa. Antimicrob Agents Chemother 43: 415–417.

51. PooleK (2005) Aminoglycoside resistance in Pseudomonas aeruginosa. Antimicrob Agents Chemother 49: 479–487.

52. VettorettiL, PlesiatP, MullerC, El GarchF, PhanG, et al. (2009) Efflux unbalance in Pseudomonas aeruginosa isolates from cystic fibrosis patients. Antimicrob Agents Chemother 53: 1987–1997.

53. ZhangL, MahTF (2008) Involvement of a novel efflux system in biofilm-specific resistance to antibiotics. J Bacteriol 190: 4447–4452.

54. DaviesJA, HarrisonJJ, MarquesLL, FogliaGR, StremickCA, et al. (2007) The GacS sensor kinase controls phenotypic reversion of small colony variants isolated from biofilms of Pseudomonas aeruginosa PA14. FEMS Microbiol Ecol 59: 32–46.

55. SallKM, CasabonaMG, BordiC, HuberP, de BentzmannS, et al. (2014) A gacS Deletion in Pseudomonas aeruginosa Cystic Fibrosis Isolate CHA Shapes Its Virulence. PLoS One 9: e95936.

56. NelsonLK, StantonMM, ElphinstoneRE, HelwerdaJ, TurnerRJ, et al. (2010) Phenotypic diversification in vivo: Pseudomonas aeruginosa gacS- strains generate small colony variants in vivo that are distinct from in vitro variants. Microbiology 156: 3699–3709.

57. BieleckiP, LukatP, HuseckenK, DotschA, SteinmetzH, et al. (2012) Mutation in elongation factor G confers resistance to the antibiotic argyrin in the opportunistic pathogen Pseudomonas aeruginosa. Chembiochem 13: 2339–2345.

58. NyfelerB, HoepfnerD, PalestrantD, KirbyCA, WhiteheadL, et al. (2012) Identification of elongation factor G as the conserved cellular target of argyrin B. PLoS One 7: e42657.

59. OverhageJ, SchemionekM, WebbJS, RehmBH (2005) Expression of the psl operon in Pseudomonas aeruginosa PAO1 biofilms: PslA performs an essential function in biofilm formation. Appl Environ Microbiol 71: 4407–4413.

60. ValletI, OlsonJW, LoryS, LazdunskiA, FillouxA (2001) The chaperone/usher pathways of Pseudomonas aeruginosa: identification of fimbrial gene clusters (cup) and their involvement in biofilm formation. Proc Natl Acad Sci U S A 98: 6911–6916.

61. MahenthiralingamE, CampbellME, SpeertDP (1994) Nonmotility and phagocytic resistance of Pseudomonas aeruginosa isolates from chronically colonized patients with cystic fibrosis. Infect Immun 62: 596–605.

62. D'ArgenioDA, WuM, HoffmanLR, KulasekaraHD, DézielE, et al. (2007) Growth phenotypes of Pseudomonas aeruginosa lasR mutants adapted to the airways of cystic fibrosis patients. Mol Microbiol 64: 512–533.

63. HoffmanLR, KulasekaraHD, EmersonJ, HoustonLS, BurnsJL, et al. (2009) Pseudomonas aeruginosa lasR mutants are associated with cystic fibrosis lung disease progression. J Cyst Fibros 8: 66–70.

64. MartinDW, SchurrMJ, MuddMH, GovanJR, HollowayBW, et al. (1993) Mechanism of conversion to mucoidy in Pseudomonas aeruginosa infecting cystic fibrosis patients. Proc Natl Acad Sci U S A 90: 8377–8381.

65. MontanariS, OliverA, SalernoP, MenaA, BertoniG, et al. (2007) Biological cost of hypermutation in Pseudomonas aeruginosa strains from patients with cystic fibrosis. Microbiology 153: 1445–1454.

66. García-CastilloM, MáizL, MorosiniMI, Rodríguez-BañosM, SuarezL, et al. (2012) Emergence of a mutL mutation causing multilocus sequence typing-pulsed-field gel electrophoresis discrepancy among Pseudomonas aeruginosa isolates from a cystic fibrosis patient. J Clin Microbiol 50: 1777–1778.

67. WuTH, MarinusMG (1999) Deletion mutation analysis of the mutS gene in Escherichia coli. J Biol Chem 274: 5948–5952.

68. MiguelV, MontiMR, ArgarañaCE (2008) The role of MutS oligomers on Pseudomonas aeruginosa mismatch repair system activity. DNA Repair (Amst) 7: 1799–1808.

69. WinsorGL, LamDK, FlemingL, LoR, WhitesideMD, et al. (2011) Pseudomonas Genome Database: improved comparative analysis and population genomics capability for Pseudomonas genomes. Nucleic Acids Res 39: D596–600.

70. JelsbakL, JohansenHK, FrostAL, ThogersenR, ThomsenLE, et al. (2007) Molecular epidemiology and dynamics of Pseudomonas aeruginosa populations in lungs of cystic fibrosis patients. Infect Immun 75: 2214–2224.

71. RauMH, HansenSK, JohansenHK, ThomsenLE, WorkmanCT, et al. (2010) Early adaptive developments of Pseudomonas aeruginosa after the transition from life in the environment to persistent colonization in the airways of human cystic fibrosis hosts. Environ Microbiol 12: 1643–1658.

72. WinstanleyC, LangilleMG, FothergillJL, Kukavica-IbruljI, Paradis-BleauC, et al. (2009) Newly introduced genomic prophage islands are critical determinants of in vivo competitiveness in the Liverpool Epidemic Strain of Pseudomonas aeruginosa. Genome Res 19: 12–23.

73. WilderCN, AlladaG, SchusterM (2009) Instantaneous within-patient diversity of Pseudomonas aeruginosa quorum-sensing populations from cystic fibrosis lung infections. Infect Immun 77: 5631–5639.

74. TurrientesMC, BaqueroF, LevinBR, MartínezJL, RipollA, et al. (2013) Normal mutation rate variants arise in a Mutator (MutS) Escherichia coli population. PLoS One 8: e72963.

75. GalabertC, JacquotJ, ZahmJM, PuchelleE (1987) Relationships between the lipid content and the rheological properties of airway secretions in cystic fibrosis. Clin Chim Acta 164: 139–149.

76. RoseMC, VoynowJA (2006) Respiratory tract mucin genes and mucin glycoproteins in health and disease. Physiol Rev 86: 245–278.

77. BarthAL, PittTL (1996) The high amino-acid content of sputum from cystic fibrosis patients promotes growth of auxotrophic Pseudomonas aeruginosa. J Med Microbiol 45: 110–119.

78. RauMH, MarvigRL, EhrlichGD, MolinS, JelsbakL (2012) Deletion and acquisition of genomic content during early stage adaptation of Pseudomonas aeruginosa to a human host environment. Environ Microbiol 14: 2200–2211.

79. LeeDG, UrbachJM, WuG, LiberatiNT, FeinbaumRL, et al. (2006) Genomic analysis reveals that Pseudomonas aeruginosa virulence is combinatorial. Genome Biol 7: R90.

80. MareghettiL, Marquet-van der MeeN, LoulergueJ, RollandJC, AA (1998) Pseudomonas aeruginosa from cystic fibrosis patients: study using whole cell RAPD and antibiotic susceptibility. Pathol Biol 46: 319–324.

81. RömlingU, TümmlerB (2000) Achieving 100% typeability of Pseudomonas aeruginosa by pulsed-field gel electrophoresis. J Clin Microbiol 38: 464–465.

82. TenoverFC, ArbeitRD, GoeringRV, MickelsenPA, MurrayBE, et al. (1995) Interpreting chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis: criteria for bacterial strain typing. J Clin Microbiol 33: 2233–2239.

83. MoralesG, WiehlmannL, GudowiusP, van DeldenC, TümmlerB, et al. (2004) Structure of Pseudomonas aeruginosa populations analyzed by single nucleotide polymorphism and pulsed-field gel electrophoresis genotyping. J Bacteriol 186: 4228–4237.

84. ZerbinoDR, BirneyE (2008) Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18: 821–829.

85. KrawitzP, RodelspergerC, JagerM, JostinsL, BauerS, et al. (2010) Microindel detection in short-read sequence data. Bioinformatics 26: 722–729.

86. LiH, HandsakerB, WysokerA, FennellT, RuanJ, et al. (2009) The Sequence Alignment/Map format and SAMtools. Bioinformatics 25: 2078–2079.

87. DePristoMA, BanksE, PoplinR, GarimellaKV, MaguireJR, et al. (2011) A framework for variation discovery and genotyping using next-generation DNA sequencing data. Nat Genet 43: 491–498.

88. KurtzS, PhillippyA, DelcherAL, SmootM, ShumwayM, et al. (2004) Versatile and open software for comparing large genomes. Genome Biol 5: R12.

89. Balzarini MG. DRJA InfoGen versión 2013.FCA, Universidad Nacional de Córdoba, Argentina. URL http://www.info-gen.com.ar.

90. PezzaRJ, SmaniaAM, BarraJL, ArgarañaCE (2002) Nucleotides and heteroduplex DNA preserve the active conformation of Pseudomonas aeruginosa MutS by preventing protein oligomerization. Biochem J 361: 87–95.

91. JacquelínDK, MartinaMA, ArgarañaCE, BarraJL (2008) Plasmid expression of mutS, -L and/or -H gene in Escherichia coli dam cells results in strains that display reduced mutation frequency. Mutat Res 637: 197–204.

92. ChoiKH, SchweizerHP (2005) An improved method for rapid generation of unmarked Pseudomonas aeruginosa deletion mutants. BMC Microbiol 5: 30.

93. BochnerBR (2009) Global phenotypic characterization of bacteria. FEMS Microbiol Rev 33: 191–205.

94. BochnerBR (2003) New technologies to assess genotype-phenotype relationships. Nat Rev Genet 4: 309–314.

95. CooperVS, LenskiRE (2000) The population genetics of ecological specialization in evolving Escherichia coli populations. Nature 407: 736–739.

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