1887

Abstract

An SNP is a spontaneous genetic change having a potential to modify the functions of the original genes and to lead to phenotypic diversity of bacteria in nature. In this study, a phylogenetic analysis of serovar Enteritidis, a major food-borne pathogen, showed that eight strains of . Enteritidis isolated in South Korea, including FORC_075 and FORC_078, have almost identical genome sequences. Interestingly, however, the abilities of FORC_075 to form biofilms and red, dry and rough (RDAR) colonies were significantly impaired, resulting in phenotypic differences among the eight strains. Comparative genomic analyses revealed that one of the non-synonymous SNPs unique to FORC_075 has occurred in , which encodes a sensor kinase of the EnvZ/OmpR two-component system. The SNP in leads to an amino acid change from Pro248 (CG) in other strains including FORC_078 to Leu248 (CG) in FORC_075. Allelic exchange of between FORC_075 and FORC_078 identified that the SNP in is responsible for the impaired biofilm- and RDAR colony-forming abilities of . Enteritidis. Biochemical analyses demonstrated that the SNP in significantly increases the phosphorylated status of OmpR in . Enteritidis and alters the expression of the OmpR regulon. Phenotypic analyses further identified that the SNP in decreases motility of . Enteritidis but increases its adhesion and invasion to both human epithelial cells and murine macrophage cells. In addition to an enhancement of infectivity to the host cells, survival under acid stress was also elevated by the SNP in . Together, these results suggest that the natural occurrence of the SNP in could contribute to phenotypic diversity of . Enteritidis, possibly improving its fitness and pathogenesis.

Funding
This study was supported by the:
  • Ministry of Science, ICT and Future Planning (Award 2017R1E1A1A01074639)
    • Principle Award Recipient: SangHo Choi
  • Ministry of Food and Drug Safety (Award 19162MFDS037)
    • Principle Award Recipient: SangHo Choi
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2021-05-05
2021-06-12
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References

  1. Arber W. Genetic variation: molecular mechanisms and impact on microbial evolution. FEMS Microbiol Rev 2000; 24:1–7 [View Article][PubMed]
    [Google Scholar]
  2. Yang L, Jelsbak L, Marvig RL, Damkiær S, Workman CT et al. Evolutionary dynamics of bacteria in a human host environment. Proc Natl Acad Sci U S A 2011; 108:7481–7486 [View Article][PubMed]
    [Google Scholar]
  3. Lieberman TD, Michel J-B, Aingaran M, Potter-Bynoe G, Roux D et al. Parallel bacterial evolution within multiple patients identifies candidate pathogenicity genes. Nat Genet 2011; 43:1275–1280 [View Article][PubMed]
    [Google Scholar]
  4. Kelly BG, Vespermann A, Bolton DJ. The role of horizontal gene transfer in the evolution of selected foodborne bacterial pathogens. Food Chem Toxicol 2009; 47:951–968 [View Article][PubMed]
    [Google Scholar]
  5. Sokurenko EV, Hasty DL, Dykhuizen DE. Pathoadaptive mutations: gene loss and variation in bacterial pathogens. Trends Microbiol 1999; 7:191–195 [View Article][PubMed]
    [Google Scholar]
  6. Collery MM, Kuehne SA, McBride SM, Kelly ML, Monot M et al. What’s a SNP between friends: The influence of single nucleotide polymorphisms on virulence and phenotypes of Clostridium difficile strain 630 and derivatives. Virulence 2017; 8:767–781
    [Google Scholar]
  7. Mikheecheva NE, Zaychikova MV, Melerzanov AV, Danilenko VN. A nonsynonymous SNP catalog of Mycobacterium tuberculosis virulence genes and its use for detecting new potentially virulent sublineages. Gen Biol Evol 2017; 9:887–899
    [Google Scholar]
  8. Scaltriti E, Sassera D, Comandatore F, Morganti M, Mandalari C et al. Differential single nucleotide polymorphism-based analysis of an outbreak caused by Salmonella enterica serovar Manhattan reveals epidemiological details missed by standard pulsed-field gel electrophoresis. J Clin Microbiol 2015; 53:1227–1238 [View Article][PubMed]
    [Google Scholar]
  9. Chen H-M, Wang Y, Su L-H, Chiu C-H. Nontyphoid Salmonella infection: microbiology, clinical features, and antimicrobial therapy. Pediatr Neonatol 2013; 54:147–152 [View Article][PubMed]
    [Google Scholar]
  10. European Food Safety Authority and European Centre for Disease Prevention and Control (EFSA and ECDC) The European Union summary report on trends and sources of zoonoses, zoonotic agents and food-borne outbreaks in 2017. Efsa J 2018; 16:e05500 [View Article][PubMed]
    [Google Scholar]
  11. Capra EJ, Laub MT. Evolution of two-component signal transduction systems. Annu Rev Microbiol 2012; 66:325–347 [View Article][PubMed]
    [Google Scholar]
  12. Delgado J, Forst S, Harlocker S, Inouye M. Identification of a phosphorylation site and functional analysis of conserved aspartic acid residues of OmpR, a transcriptional activator for ompF and ompC in Escherichia coli. Mol Microbiol 1993; 10:1037–1047 [View Article][PubMed]
    [Google Scholar]
  13. Wang LC, Morgan LK, Godakumbura P, Kenney LJ, Anand GS. The inner membrane histidine kinase EnvZ senses osmolality via helix-coil transitions in the cytoplasm. Embo J 2012; 31:2648–2659
    [Google Scholar]
  14. Chakraborty S, Winardhi RS, Morgan LK, Yan J, Kenney LJ. Non-canonical activation of OmpR drives acid and osmotic stress responses in single bacterial cells. Nat Commun 2017; 8:1587 [View Article][PubMed]
    [Google Scholar]
  15. Head CG, Tardy A, Kenney LJ. Relative binding affinities of OmpR and OmpR-phosphate at the ompF and ompC regulatory sites. J Mol Biol 1998; 281:857–870 [View Article][PubMed]
    [Google Scholar]
  16. Aiba H, Mizuno T. Phosphorylation of a bacterial activator protein, OmpR, by a protein kinase, EnvZ, stimulates the transcription of the ompF and ompC genes in Escherichia coli. FEBS Lett 1990; 261:19–22 [View Article][PubMed]
    [Google Scholar]
  17. Dorman CJ, Chatfield S, Higgins CF, Hayward C, Dougan G. Characterization of porin and ompR mutants of a virulent strain of Salmonella typhimurium: ompR mutants are attenuated in vivo. Infect Immun 1989; 57:2136–2140 [View Article][PubMed]
    [Google Scholar]
  18. Quinn HJ, Cameron ADS, Dorman CJ. Bacterial regulon evolution: distinct responses and roles for the identical OmpR proteins of Salmonella typhimurium and Escherichia coli in the acid stress response. PLoS Genet 2014; 10:e1004215 [View Article][PubMed]
    [Google Scholar]
  19. Gerstel U, Park C, Romling U. Complex regulation of csgD promoter activity by global regulatory proteins. Mol Microbiol 2003; 49:639–654
    [Google Scholar]
  20. Hammar M, Arnqvist A, Bian Z, Olsen A, Normark S. Expression of two csg operons is required for production of fibronectin- and Congo red-binding curli polymers in Escherichia coli K-12. Mol Microbiol 1995; 18:661–670
    [Google Scholar]
  21. Romling U, Rohde M, Olsen A, Normark S, AgfD RJ. the checkpoint of multicellular and aggregative behaviour in Salmonella typhimurium regulates at least two independent pathways. Mol Microbiol 2000; 36:10–23
    [Google Scholar]
  22. Austin JW, Sanders G, Kay WW, Collinson SK. Thin aggregative fimbriae enhance Salmonella enteritidis biofilm formation. Fems Microbiology Letters 1998; 162:295–301
    [Google Scholar]
  23. Solano C, Garcia B, Valle J, Berasain C, Ghigo JM et al. Genetic analysis of Salmonella enteritidis biofilm formation: critical role of cellulose. Mol Microbiol 2002; 43:793–808
    [Google Scholar]
  24. Römling U. Characterization of the rdar morphotype, a multicellular behaviour in Enterobacteriaceae. Cell Mol Life Sci 2005; 62:1234–1246 [View Article][PubMed]
    [Google Scholar]
  25. Xu S, Zou X, Sheng X, Zhang H, Mao L et al. Expression of fljB:z66 on a linear plasmid of Salmonella enterica serovar Typhi is dependent on fliA and flhDC and regulated by OmpR. Braz J Microbiol 2010; 41:729–740 [View Article][PubMed]
    [Google Scholar]
  26. Ellermeier CD, Ellermeier JR, Slauch JM. Hild, HilC and RtsA constitute a feed forward loop that controls expression of the SPI1 type three secretion system regulator hilA in Salmonella enterica serovar typhimurium. Mol Microbiol 2005; 57:691–705 [View Article][PubMed]
    [Google Scholar]
  27. Feng X, Oropeza R, Kenney LJ. Dual regulation by phospho-OmpR of ssrA/B gene expression in Salmonella pathogenicity island 2. Mol Microbiol 2003; 48:1131–1143 [View Article][PubMed]
    [Google Scholar]
  28. Lee AK, Detweiler CS, Falkow S. OmpR regulates the two-component system SsrA-SsrB in Salmonella pathogenicity island 2. J Bacteriol 2000; 182:771–781
    [Google Scholar]
  29. Galán JE. Molecular genetic bases of Salmonella entry into host cells. Mol Microbiol 1996; 20:263–271 [View Article][PubMed]
    [Google Scholar]
  30. Groisman EA, Ochman H. Cognate gene clusters govern invasion of host epithelial-cells by Salmonella typhimurium and Shigella flexneri. Embo J 1993; 12:3779–3787
    [Google Scholar]
  31. Groisman EA, Ochman H. How Salmonella became a pathogen. Trends Microbiol 1997; 5:343–349 [View Article][PubMed]
    [Google Scholar]
  32. Shea JE, Hensel M, Gleeson C, Holden DW. Identification of a virulence locus encoding a second type III secretion system in Salmonella typhimurium. Proc Natl Acad Sci U S A 1996; 93:2593–2597 [View Article][PubMed]
    [Google Scholar]
  33. Bang IS, Kim BH, Foster JW, Park YK. OmpR regulates the stationary-phase acid tolerance response of Salmonella enterica serovar Typhimurium. J Bacteriol 2000; 182:2245–2252
    [Google Scholar]
  34. Chakraborty S, Mizusaki H, Kenney LJ. A FRET-based DNA biosensor tracks OmpR-dependent acidification of Salmonella during macrophage infection. Plos Biology 2015; 13:
    [Google Scholar]
  35. Chakraborty S, Kenney LJ. A new role of OmpR in acid and osmotic stress in Salmonella and E. coli. Front Microbiol 2018; 9:
    [Google Scholar]
  36. Richter M, Rosselló-Móra R. Shifting the genomic gold standard for the prokaryotic species definition. Proc Natl Acad Sci U S A 2009; 106:19126–19131 [View Article][PubMed]
    [Google Scholar]
  37. Croucher NJ, Page AJ, Connor TR, Delaney AJ, Keane JA et al. Rapid phylogenetic analysis of large samples of recombinant bacterial whole genome sequences using Gubbins. Nucleic Acids Res 2015; 43:e15
    [Google Scholar]
  38. Page AJ, Taylor B, Delaney AJ, Soares J, Seemann T et al. SNP-sites: rapid efficient extraction of SNPs from multi-FASTA alignments. Microbial Genomics 2016; 2:
    [Google Scholar]
  39. Stamatakis A. RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics 2006; 22:2688–2690
    [Google Scholar]
  40. Seemann T. Prokka: rapid prokaryotic genome annotation. Bioinformatics 2014; 30:2068–2069
    [Google Scholar]
  41. 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
    [Google Scholar]
  42. Kurtz S, Phillippy A, Delcher AL, Smoot M, Shumway M et al. Versatile and open software for comparing large genomes. Genome Biology 2004; 5:
    [Google Scholar]
  43. Choi Y, Chan AP. PROVEAN web server: a tool to predict the functional effect of amino acid substitutions and indels. Bioinformatics 2015; 31:2745–2747
    [Google Scholar]
  44. Hecht M, Bromberg Y, Rost B. Better prediction of functional effects for sequence variants. BMC Genomics 2015; 16:S1
    [Google Scholar]
  45. Sim NL, Kumar P, Hu J, Henikoff S, Schneider G et al. SIFT web server: predicting effects of amino acid substitutions on proteins. Nucleic Acids Res 2012; 40:W452–W7
    [Google Scholar]
  46. Neiger MR, Gonzalez JF, Gonzalez-Escobedo G, Kuck H, White P et al. Pathoadaptive alteration of Salmonella biofilm formation in response to the gallbladder environment. J Bacteriol 2019; 201:
    [Google Scholar]
  47. Park JH, Jo Y, Jang SY, Kwon H, Iriecurrency Y et al. The cabABC operon essential for biofilm and rugose colony development in Vibrio vulnificus. Plos Pathog 2015; 11:
    [Google Scholar]
  48. Datsenko KA, Wanner BL. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 2000; 97:6640–6645
    [Google Scholar]
  49. Philippe N, Alcaraz JP, Coursange E, Geiselmann J, Schneider D. Improvement of pCVD442, a suicide plasmid for gene allele exchange in bacteria. Plasmid 2004; 51:246–255
    [Google Scholar]
  50. Lee ZW, Hwang SH, Choi G, Jang KK, Lee TH et al. A MARTX Toxin rtxA Gene Is Controlled by Host Environmental Signals through a CRP-Coordinated Regulatory Network in Vibrio vulnificus. mBio. 2020; 11:
    [Google Scholar]
  51. Kim SM, Lee DH, Choi SH. Evidence that the Vibrio vulnificus flagellar regulator FlhF is regulated by a quorum sensing master regulator SmcR. Microbiology-Sgm 2012; 158:2017–2025
    [Google Scholar]
  52. Yoshida CE, Kruczkiewicz P, Laing CR, Lingohr EJ, Gannon VPJ et al. The Salmonella in silico Typing Resource (SISTR): an open web-accessible tool for rapidly typing and subtyping draft Salmonella genome assemblies. PLoS One 2016; 11:1 [View Article][PubMed]
    [Google Scholar]
  53. MacKenzie KD, Palmer MB, Koster WL, White AP. Examining the link between biofilm formation and the ability of pathogenic Salmonella strains to colonize multiple host species. Front Vet Sci 2017; 4:
    [Google Scholar]
  54. Desai SK, Kenney LJ. To ~P or not to ~P? non-canonical activation by two-component response regulators. Mol Microbiol 2017; 103:203–213
    [Google Scholar]
  55. Desai SK, Kenney LJ. Switching lifestyles is an in vivo adaptive strategy of bacterial pathogens. Front Cell Infect Microbiol 2019; 9:421 [View Article][PubMed]
    [Google Scholar]
  56. Desai SK, Winardhi RS, Periasamy S, Dykas MM, Jie Y et al. The horizontally-acquired response regulator SsrB drives a Salmonella lifestyle switch by relieving biofilm silencing. elife 2016; 5:
    [Google Scholar]
  57. Serra DO, Richter AM, Hengge R. Cellulose as an architectural element in spatially structured Escherichia coli biofilms. J Bacteriol 2013; 195:5540–5554 [View Article][PubMed]
    [Google Scholar]
  58. Klumpp J, Fuchs TM. Identification of novel genes in genomic islands that contribute to Salmonella typhimurium replication in macrophages. Microbiology 2007; 153:1207–1220
    [Google Scholar]
  59. Gal-Mor O, Gibson DL, Baluta D, Vallance BA, Finlay BB. A novel secretion pathway of Salmonella enterica acts as an antivirulence modulator during salmonellosis. Plos Pathog 2008; 4:
    [Google Scholar]
  60. Prehna G, Li Y, Stoynov N, Okon M, Vuckovic M et al. The zinc regulated antivirulence pathway of Salmonella is a multiprotein immunoglobulin adhesion system. J Biol Chem 2012; 287:32324–32337 [View Article][PubMed]
    [Google Scholar]
  61. Theoret JR, Cooper KK, Zekarias B, Roland KL, Law BF et al. The Campylobacter jejuni Dps homologue is important for in vitro biofilm formation and cecal colonization of poultry and may serve as a protective antigen for vaccination. Clin Vaccine Immunol 2012; 19:1426–1431 [View Article][PubMed]
    [Google Scholar]
  62. Prigent-Combaret C, Brombacher E, Vidal O, Ambert A, Lejeune P et al. Complex regulatory network controls initial adhesion and biofilm formation in Escherichia coli via regulation of the csgD gene. J Bacteriol 2001; 183:7213–7223
    [Google Scholar]
  63. Gerstel U, Kolb A, Romling U. Regulatory components at the csgD promoter-additional roles for OmpR and integration host factor and role of the 5 ' untranslated region. Fems Microbiol Lett 2006; 261:109–117
    [Google Scholar]
  64. Thomson NR, Clayton DJ, Windhorst D, Vernikos G, Davidson S et al. Comparative genome analysis of Salmonella Enteritidis PT4 and Salmonella Gallinarum 287/91 provides insights into evolutionary and host adaptation pathways. Genome Res 2008; 18:1624–1637 [View Article]
    [Google Scholar]
  65. Stabler RA, He M, Dawson L, Martin M, Valiente E et al. Comparative genome and phenotypic analysis of Clostridium difficile 027 strains provides insight into the evolution of a hypervirulent bacterium. Genome Biol 2009; 10:R102 [View Article][PubMed]
    [Google Scholar]
  66. Tomomori C, Tanaka T, Dutta R, Park H, Saha SK et al. Solution structure of the homodimeric core domain of Escherichia coli histidine kinase EnvZ. Nat Struct Biol 1999; 6:729–734 [View Article][PubMed]
    [Google Scholar]
  67. Horstmann N, Saldana M, Sahasrabhojane P, Yao H, XP S et al. Dual-Site phosphorylation of the control of virulence regulator impacts group A streptococcal global gene expression and pathogenesis. Plos Pathog 2014; 10:
    [Google Scholar]
  68. Ichikawa M, Minami M, Isaka M, Tatsuno I, Hasegawa T. Analysis of two-component sensor proteins involved in the response to acid stimuli in Streptococcus pyogenes. Microbiology-Sgm 2011; 157:3187–3194
    [Google Scholar]
  69. Flores AR, Jewell BE, Yelamanchili D, Olsen RJ, Musser JM. A single amino acid replacement in the sensor kinase LiaS contributes to a carrier phenotype in Group A Streptococcus. Infect Immun 2015; 83:4237–4246 [View Article][PubMed]
    [Google Scholar]
  70. Horstmann N, Tran CN, Brumlow C, DebRoy S, Yao H et al. Phosphatase activity of the control of virulence sensor kinase CovS is critical for the pathogenesis of group A streptococcus. Plos Pathog 2018; 14:
    [Google Scholar]
  71. White AP, Gibson DL, Grassl GA, Kay WW, Finlay BB et al. Aggregation via the red, dry, and rough morphotype is not a virulence adaptation in Salmonella enterica serovar typhimurium. Infect Immun 2008; 76:1048–1058 [View Article][PubMed]
    [Google Scholar]
  72. Tükel C, Nishimori JH, Wilson RP, Winter MG, Keestra AM et al. Toll-like receptors 1 and 2 cooperatively mediate immune responses to curli, a common amyloid from enterobacterial biofilms. Cell Microbiol 2010; 12:1495–1505 [View Article][PubMed]
    [Google Scholar]
  73. Pontes MH, Lee E-J, Choi J, Groisman EA. Salmonella promotes virulence by repressing cellulose production. Proc Natl Acad Sci U S A 2015; 112:5183–5188 [View Article][PubMed]
    [Google Scholar]
  74. Ahmad I, Lamprokostopoulou A, Le Guyon S, Streck E, Barthel M et al. Complex c-di-GMP signaling networks mediate transition between virulence properties and biofilm formation in Salmonella enterica serovar typhimurium. PLoS One 2011; 6:e28351 [View Article][PubMed]
    [Google Scholar]
  75. Winter SE, Winter MG, Godinez I, Yang HJ, Russmann H et al. A rapid change in virulence gene expression during the transition from the intestinal lumen into tissue promotes systemic dissemination of Salmonella. Plos Pathog 2010; 6:
    [Google Scholar]
  76. Miao EA, Leaf IA, Treuting PM, Mao DP, Dors M et al. Caspase-1-induced pyroptosis is an innate immune effector mechanism against intracellular bacteria. Nat Immunol 2010; 11:1136–1142 [View Article][PubMed]
    [Google Scholar]
  77. Fàbrega A, Vila J. Salmonella enterica serovar Typhimurium skills to succeed in the host: virulence and regulation. Clin Microbiol Rev 2013; 26:308–341 [View Article][PubMed]
    [Google Scholar]
  78. Yuk HG, Schneider KR. Adaptation of Salmonella spp. in juice stored under refrigerated and room temperature enhances acid resistance to simulated gastric fluid. Food Microbiol 2006; 23:694–700
    [Google Scholar]
  79. Álvarez-Ordóñez A, Prieto M, Bernardo A, Hill C, López M. The acid tolerance response of Salmonella spp.: an adaptive strategy to survive in stressful environments prevailing in foods and the host. Food Res Int 2012; 45:482–492 [View Article]
    [Google Scholar]
  80. Hammarlof DL, Kroger C, Owen SV, Canals R, Lacharme-Lora L et al. Role of a single noncoding nucleotide in the evolution of an epidemic African clade of Salmonella. Proc Natl Acad Sci U S A 2018; 115:E2614–E23
    [Google Scholar]
  81. MacKenzie KD, Wang Y, Musicha P, Hansen EG, Palmer MB et al. Parallel evolution leading to impaired biofilm formation in invasive Salmonella strains. PLoS Genet 2019; 15:e1008233
    [Google Scholar]
  82. Letunic I, Bork P. Interactive tree of life (iTOL) V3: an online tool for the display and annotation of phylogenetic and other trees. Nucleic Acids Res 2016; 44:W242–W5
    [Google Scholar]
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