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Induction of the viable but nonculturable state of Salmonella enterica serovar Enteritidis deficient in (p)ppGpp synthesis

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Abstract

Salmonella enterica enters a viable but nonculturable (VBNC) state in adverse environmental conditions. Under nutritional stress, RelA and SpoT proteins synthesize (p)ppGpp, a molecule that controls the expression of genes important for the survival of a cell under those conditions. This study aimed to verify the involvement of (p)ppGpp in the entry of S. enterica serovar Enteritidis PT4 578 cells into the VBNC state by evaluating ΔrelA and ΔrelAΔspoT mutants. The wild-type and mutant strains deficient in (p)ppGpp synthesis were subjected to osmotic, nutritional and cold stress, after which the cells entered a VBNC state at different time periods, concurrent with reductions in cell diameter, volume and length, and conversion from a bacillary to a coccoid form. No difference in the culturability or cell viability of the wild-type and single and double mutants was observed. Expression of the rpoS gene was increased in the double mutant, while both mutant strains presented a reduction in mreB gene expression after 25 days under conditions of nutritional and cold stress. Surprisingly, (p)ppGpp was not necessary for induction of the VBNC state in Salmonella PT4 578 cells, but may be associated with regulation of genes that control septum formation during cell division, maintaining the bacillary cell morphology (mreB) and stress response (rpoS). Our findings contribute to the understanding of the mechanisms of resistance and survival of Salmonella under adverse conditions.

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References

  • Albertini MC, Accorsi A, Teodori L, Pierfelici L, Uguccioni F, Rocchi MBL, Burattini S, Citterio B (2006) Use of multiparameter analysis for Vibrio alginolyticus viable but nonculturable state determination. Cytometry A 69:260–265

    Article  PubMed  Google Scholar 

  • AOAC (1998) Association of Official Analytical Chemistis International. Bacteriological Analytical Manual. 8rd edn. Gaithersburg, Md, USA

  • Brown L, Gentry D, Elliot T, Cashel M (2002) DksA affects ppGpp induction of RpoS at a translational level. J Bacteriol 184:4455–4465

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Cashel M, Gallant J (1969) Two compounds implicated in the function of the RC gene of Escherichia coli. Nature 221:838–841

    Article  CAS  PubMed  Google Scholar 

  • Cashel M, Gentry DR, Hernandez VJ, Vinella D (1996) The stringent response. In: Neidhardt FC, Curtiss IR, Ingraham JL, Lin ECC, Low KB, Magasanik B, Reznikoff WS, Riley M, Schaechter M, Umbarger HE (eds) Escherichia coli and Salmonella: cellular and molecular biology, 2nd edn. ASM Press, Washington D.C

    Google Scholar 

  • CDC (2012) Trends in Foodborne Illness in the United States 1996–2011

  • Chatterji D, Ojha AK (2001) Revisiting the stringent response, ppGpp and starvation signaling. Curr Opin Microbiol 4:160–165

    Article  CAS  PubMed  Google Scholar 

  • Chiu SW, Chen SY, Wong HC (2008) Localization and expression of MreB in Vibrio parahaemolyticus under different stress. Appl Environ Microbiol 74:7016–7022

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Doble AC, Bulmer DM, Kharraz L, Karavolos MH, Anjam Khan CM (2012) The function of the bacterial cytoskeleton in Salmonella pathogenesis. Virulence 3:446–449

    Article  PubMed Central  PubMed  Google Scholar 

  • Eymann C, Homuth G, Scharf C, Hecker M (2002) Bacillus subtilis functional genomics: global characterization of the stringent response by proteome and transcriptome analysis. J Bacteriol 184:2500–2520

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Hengee R (2009) Proteolysis of σS (RpoS) and the general stress response in Escherichia coli. Res Microbiol 160:667–676

    Article  Google Scholar 

  • Hirsch M, Elliot T (2005) Fis Regulates Transcriptional induction of RpoS in Salmonella enterica. J Bacteriol 187:1568–1580

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Jishage M, Kvint K, Shingler V, Nystrom T (2002) Regulation of σ factor competition by the alarmone ppGpp. Genes Dev 16:1260–1270

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Magnusson LU, Gummersson B, Joksimovic P, Farewell A, Nystrom T (2007) Identical, independent, and opposing roles of ppGpp and DksA in Escherichia coli. J Bacteriol 189:5193–5202

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Mata GMSC (2012) Dissertation, Federal University of Viçosa, Viçosa, MG, Brazil

  • Miller J (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • Morton RD (2001) Aerobic plate count. In: Downes FP, Ito K (eds) Compendium of methods for the microbiological examination of foods, 4th edn. American Public Health Association-APHA, Washington, DC

    Google Scholar 

  • Munro PM, Flatau GN, Clement RL, Gauthier MJ (1995) Influence of the RpoS (KatF) sigma factor on maintenance of viability and culturability of Escherichia coli and Salmonella typhimurium in seawater. Appl Environ Microbiol 61:1853–1858

    PubMed Central  CAS  PubMed  Google Scholar 

  • Neidhardt FC, Ingraham JL, Schaechter M (1990) Physiology of the bacterial cell. A molecular approach. Sinauer Associates, Massachusetts

    Google Scholar 

  • Oliver JD (2000) The public health significance of viable but nonculturable bacteria. In: Colwell RR, Grimes DJ (eds) Nonculturable microorganisms in the environment. American Society for Microbiology Press, Washington, DC, pp 277–299

    Chapter  Google Scholar 

  • Oliver JD (2010) Recent findings on the viable but nonculturable state in pathogenic bacteria. FEMS Microbiol Rev 34:415–425

    Article  CAS  PubMed  Google Scholar 

  • Paul BJ, Barker MM, Ross W, Schneider DA, Webb C, Foster JW, Gourse RL (2004) DksA: a critical component of the transcription initiation machinery that potentiates the regulation of rRNA promoters by ppGpp and the initiating NTP. Cell 118:311–322

    Article  CAS  PubMed  Google Scholar 

  • Perederina A, Vassvlveva MN, Tahirov TH, Yokoyama S, Artsimovitch I, Vassvlveva DG (2004) Regulation through the secondary channel—structural framework for ppGpp-DksA synergism during transcription. Cell 118:297–309

    Article  CAS  PubMed  Google Scholar 

  • Powell BS, Court DL (1998) Control of ftsZ expression, cell division, and glutamine metabolism in luria bertani medium by the alarmone ppGpp in Escherichia coli. J Bacteriol 180:1053–1062

    PubMed Central  CAS  PubMed  Google Scholar 

  • Ramey WD, Ishiguro EE (1978) Site of inhibition of peptidoglycan biosynthesis during the stringent response in Escherichia coli. J Bacteriol 135:71–77

    PubMed Central  CAS  PubMed  Google Scholar 

  • Shen S, Fang FC (2012) Integrated stress responses in Salmonella. Int J Food Microbiol 152:75–81

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Spector MP, Kenyon WJ (2012) Resistance and survival strategies of Salmonella enterica to environmental stress. Rev Food Res Int 45:455–481

    Article  CAS  Google Scholar 

  • Thompson LJ, Merrell DS, Neilan BA, Mitchell H, Lee A, Falkow S (2003) Gene expression profiling of Helicobacter pylori reveals a growth-phase-dependent switch in virulence gene expression. Infect Immun 71:2643–2655

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Vogt SL, Green C, Stevens KM, Day B, Erickson DL, Woods E, Storey DG (2011) The stringent response is essential for Pseudomonas aeruginosa virulence in the rat lung agar bead and Drosophila melanogaster feeding models of infection. Infect Immun 79:4094–4104

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Weber H, Polen T, Heuveling J, Wendisch V, Hengge R (2005) Genome-wide analysis of the general stress response network in Escherichia coli: σS-dependent genes, promoters and sigma factor selectivity. J Bacteriol 187:1591–1603

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Wu J, Xie J (2009) Magic spot: (p)ppGpp. J Cell Physiol 220:297–302

    Article  CAS  PubMed  Google Scholar 

  • Xiao H, Kalman M, Keharaz K, Zemel S, Glaser G, Cashel M (1991) Residual guanosine 3’,5’-bispyrophosphate synthetic activity of reZA null mutants can be eliminated by spoT null mutations. J Biol Chem 266:5980–5990

    CAS  PubMed  Google Scholar 

  • Yu XC, Margolin W (1999) FtsZ ring clusters in min and partition mutants: role of both the Min system and the nucleoid in regulating FtsZ ring localization. Mol Microbiol 32:315–326

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by a CAPES-Brazil fellowship.

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Authors and Affiliations

  1. LUNAM Université, Oniris, University of Nantes, Nantes, France

    Ramila Cristiane Rodrigues & Evandro Martins

  2. Institut National de la Recherche Agronomique (INRA), Nantes, France

    Ramila Cristiane Rodrigues & Evandro Martins

  3. Department of Microbiology, Federal University of Viçosa (UFV), Viçosa, MG, Brazil

    Maria Cristina Dantas Vanetti & Míriam Teresinha dos Santos

  4. Department of Food, Federal University of Ouro Preto (UFOP), Viçosa, MG, Brazil

    Uelinton Manoel Pinto

  5. Avenida Peter Henry Rolfs, s/n - Campus Universitário, Viçosa, MG, 36570-000, Brazil

    Míriam Teresinha dos Santos

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  1. Ramila Cristiane Rodrigues
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  2. Evandro Martins
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  3. Maria Cristina Dantas Vanetti
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  4. Uelinton Manoel Pinto
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Correspondence to Míriam Teresinha dos Santos.

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Rodrigues, R.C., Martins, E., Vanetti, M.C.D. et al. Induction of the viable but nonculturable state of Salmonella enterica serovar Enteritidis deficient in (p)ppGpp synthesis. Ann Microbiol 65, 2171–2178 (2015). https://doi.org/10.1007/s13213-015-1057-6

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  • DOI: https://doi.org/10.1007/s13213-015-1057-6

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