Skip to main content
SpringerLink
Log in

Biological functions of nirS in Pseudomonas aeruginosa ATCC 9027 under aerobic conditions

  • Genetics and Molecular Biology of Industrial Organisms - Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

Through our previous study, we found an up-regulation in the expression of nitrite reductase (nirS) in the isothiazolone-resistant strain of Pseudomonas aeruginosa. However, the definitive molecular role of nirS in ascribing the resistance remained elusive. In the present study, the nirS gene was deleted from the chromosome of P. aeruginosa ATCC 9027 and the resulting phenotypic changes of ΔnirS were studied alongside the wild-type (WT) strain under aerobic conditions. The results demonstrated a decline in the formations of biofilms but not planktonic growth by ΔnirS as compared to WT, especially in the presence of benzisothiazolinone (BIT). Meanwhile, the deletion of nirS impaired swimming motility of P. aeruginosa under the stress of BIT. To assess the influence of nirS on the transcriptome of P. aeruginosa, RNA-seq experiments comparing the ΔnirS with WT were also performed. A total of 694 genes were found to be differentially expressed in ΔnirS, of which 192 were up-regulated, while 502 were down-regulated. In addition, these differently expressed genes were noted to significantly enrich the carbon metabolism along with glyoxylate and dicarboxylate metabolisms. Meanwhile, results from RT-PCR suggested the contribution of mexEF-oprN to the development of BIT resistance by ΔnirS. Further, c-di-GMP was less in ΔnirS than in WT, as revealed by HPLC. Taken together, our results confirm that nirS of P. aeruginosa ATCC 9027 plays a role in BIT resistance along with biofilm formation and further affects several metabolic patterns under aerobic conditions.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Arai H, Kodama T, Igarashi Y (1999) Effect of nitrogen oxides on expression of the nir and nor genes for denitrification in Pseudomonas aeruginosa. FEMS Microbiol Lett 170:19–24

    CAS  PubMed  Google Scholar 

  2. Barraud N, Hassett DJ, Hwang S-H, Rice SA, Kjelleberg S, Webb JS (2006) Involvement of nitric oxide in biofilm dispersal of Pseudomonas aeruginosa. J Bacteriol 188:7344–7353

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Barraud N, Schleheck D, Klebensberger J, Webb JS, Hassett DJ, Rice SA, Kjelleberg S (2009) Nitric oxide signaling in Pseudomonas aeruginosa biofilms mediates phosphodiesterase activity, decreased cyclic Di-GMP levels, and enhanced dispersal. J Bacteriol 191:7333–7342

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Chapman JS (2003) Biocide resistance mechanisms. Int Biodeterior Biodegrad 51:133–138

    CAS  Google Scholar 

  5. Chapman JS, Diehl MA (1995) Methylchloroisothiazolone-induced growth inhibition and lethality in Escherichia coli. J Appl Microbiol 78:134–141

    CAS  Google Scholar 

  6. Chen YC, Xie XB, Shi QS, Ouyang YS, Chen YB (2010) Species identification of industry spoilage microorganism and the resistance analysis. Microbiol China 37:1558–1565

    Google Scholar 

  7. Chuanchuen R, Beinlich K, Hoang TT, Becher A, Karkhoff-Schweizer RR, Schweizer HP (2001) Cross-resistance between triclosan and antibiotics in Pseudomonas aeruginosa is mediated by multidrug efflux pumps: exposure of a susceptible mutant strain to triclosan selects nfxB mutants overexpressing MexCD-OprJ. Antimicrob Agents Chemother 45:428–432

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Davies KJP, Lloyd D, Boddy L (1989) The effect of oxygen on denitrification in Paracoccus denitrificans and Pseudomonas aeruginosa. Microbiology 135:2445–2451

    CAS  Google Scholar 

  9. De Kievit TR, Parkins MD, Gillis RJ, Srikumar R, Ceri H, Poole K, Iglewski BH, Storey DG (2001) Multidrug efflux pumps: expression patterns and contribution to antibiotic resistance in Pseudomonas aeruginosa biofilms. Antimicrob Agents Chemother 45:1761–1770

    PubMed  PubMed Central  Google Scholar 

  10. Drenkard E (2003) Antimicrobial resistance of Pseudomonas aeruginosa biofilms. Microbes Infect 5:1213–1219

    CAS  PubMed  Google Scholar 

  11. Germ M, Yoshihara E, Yoneyama H, Nakae T (1999) Interplay between the efflux pump and the outer membrane permeability barrier in fluorescent dye accumulation in Pseudomonas aeruginosa. Biochem Biophys Res Commun 261:452–455

    CAS  PubMed  Google Scholar 

  12. Hancock RE (1998) Resistance mechanisms in Pseudomonas aeruginosa and other non-fermentative gram-negative bacteria. Clin Infect Dis 27:S93–S99

    CAS  PubMed  Google Scholar 

  13. Huang CY, Hsieh SP, Kuo PA, Jane WN, Tu J, Wang YN, Ko CH (2009) Impact of disinfectant and nutrient concentration on growth and biofilm formation for a Pseudomonas strain and the mixed cultures from a fine papermachine system. Int Biodeterior Biodegrad 63:998–1007

    CAS  Google Scholar 

  14. Huang YH, Lin JS, Ma JC, Wang HH (2016) Functional characterization of triclosan-resistant enoyl-acyl-carrier protein reductase (FabV) in Pseudomonas aeruginosa. Front Microbiol 7:1903

    PubMed  PubMed Central  Google Scholar 

  15. Su JJ, Liu BY, Liu CY (2001) Comparison of aerobic denitrification under high oxygen atmosphere by Thiosphaera pantotropha ATCC 35512 and Pseudomonas stutzeri SU2 newly isolated from the activated sludge of a piggery wastewater treatment system. J Appl Microbiol 90:457–462

    CAS  PubMed  Google Scholar 

  16. Köhler T, Michea-Hamzehpour M, Plesiat P, Kahr A-L, Pechere J-C (1997) Differential selection of multidrug efflux systems by quinolones in Pseudomonas aeruginosa. Antimicrob Agents Chemother 41:2540–2543

    PubMed  PubMed Central  Google Scholar 

  17. Kohler T, Michea-Hamzehpour M, Henze U, Gotoh N, Curty LK, Pechere JC (1997) Characterization of MexE-MexF-OprN, a positively regulated multidrug efflux system of Pseudomonas aeruginosa. Mol Microbiol 23:345–354

    CAS  PubMed  Google Scholar 

  18. Krüger A, Grüning N-M, Wamelink MM, Kerick M, Kirpy A, Parkhomchuk D, Bluemlein K, Schweiger M-R, Soldatov A, Lehrach H (2011) The pentose phosphate pathway is a metabolic redox sensor and regulates transcription during the antioxidant response. Antioxid Redox Signal 15:311–324

    PubMed  Google Scholar 

  19. Lambert P (2002) Mechanisms of antibiotic resistance in Pseudomonas aeruginosa. J R Soc Med 95:22–26

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Laopaiboon L, Smith RN, Hall SJ (2001) A study of the effect of isothiazolones on the performance and characteristics of a laboratory-scale rotating biological contactor. J Appl Microbiol 91:93–103

    CAS  PubMed  Google Scholar 

  21. Li XZ, Zhang L, Poole K (1998) Role of the multidrug efflux systems of Pseudomonas aeruginosa in organic solvent tolerance. J Bacteriol 180:2987–2991

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCt method. Methods 25:402–408

    CAS  Google Scholar 

  23. Lyczak JB, Cannon CL, Pier GB (2000) Establishment of Pseudomonas aeruginosa infection: lessons from a versatile opportunist. Microbes Infect 2:1051–1060

    CAS  PubMed  Google Scholar 

  24. Lyczak JB, Cannon CL, Pier GB (2002) Lung infections associated with cystic fibrosis. Clin Microbiol Rev 15:194–222

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Ma JC, Wu YQ, Cao D, Zhang WB, Wang HH (2017) Only acyl carrier protein 1 (AcpP1) functions in Pseudomonas aeruginosa fatty acid synthesis. Front Microbiol 8:2186

    PubMed  PubMed Central  Google Scholar 

  26. Morita Y, Tomida J, Kawamura Y (2014) Responses of Pseudomonas aeruginosa to antimicrobials. Front Microbiol 4:422

    PubMed  PubMed Central  Google Scholar 

  27. O’Toole G, Kaplan HB, Kolter R (2000) Biofilm formation as microbial development. Annu Rev Microbiol 54:49–79

    PubMed  Google Scholar 

  28. Poole K (2011) Pseudomonas aeruginosa: resistance to the max. Front Microbiol 2:65

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Pucci MJ, Podos SD, Thanassi JA, Leggio MJ, Bradbury BJ, Deshpande M (2011) In vitro and in vivo profiles of ACH-702, an isothiazoloquinolone, against bacterial pathogens. Antimicrob Agents Chemother 55:2860–2871

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Römling U, Galperin MY, Gomelsky M (2013) Cyclic di-GMP: the first 25 years of a universal bacterial second messenger. Microbiol Mol Biol Rev 77:1–52

    PubMed  PubMed Central  Google Scholar 

  31. Rashid MH, Kornberg A (2000) Inorganic polyphosphate is needed for swimming, swarming, and twitching motilities of Pseudomonas aeruginosa. Proc Natl Acad Sci USA 97:4885–4890

    CAS  PubMed  Google Scholar 

  32. Rinaldo S, Giardina G, Castiglione N, Stelitano V, Cutruzzola F (2011) The catalytic mechanism of Pseudomonas aeruginosa cd(1) nitrite reductase. Biochem Soc T 39:195–200

    CAS  Google Scholar 

  33. Robinson MD, Oshlack A (2010) A scaling normalization method for differential expression analysis of RNA-seq data. Genome Biol 11:R25–R25

    PubMed  PubMed Central  Google Scholar 

  34. Roy AB, Petrova OE, Sauer K (2013) Extraction and quantification of cyclic di-GMP from P. aeruginosa. Bio Protoc 3:e828

    PubMed  PubMed Central  Google Scholar 

  35. Ryan RP, Lucey J, O’Donovan K, McCarthy Y, Yang L, Tolker-Nielsen T, Dow JM (2009) HD-GYP domain proteins regulate biofilm formation and virulence in Pseudomonas aeruginosa. Environ Microbiol 11:1126–1136

    CAS  PubMed  Google Scholar 

  36. Sandoe JAT, Wysome J, West AP, Heritage J, Wilcox MH (2006) Measurement of ampicillin, vancomycin, linezolid and gentamicin activity against enterococcal biofilms. J Antimicrob Chemother 57:767–770

    CAS  PubMed  Google Scholar 

  37. Silvestrini MC, Galeotti CL, Gervais M, Schininà E, Barra D, Bossa F, Brunori M (1989) Nitrite reductase from Pseudomonas aeruginosa: sequence of the gene and the protein. FEBS Lett 254:33–38

    CAS  PubMed  Google Scholar 

  38. Singh PK, Schaefer AL, Parsek MR, Moninger TO, Welsh MJ, Greenberg E (2000) Quorum-sensing signals indicate that cystic fibrosis lungs are infected with bacterial biofilms. Nature 407:762–764

    CAS  PubMed  Google Scholar 

  39. Stepanović S, Vuković D, Dakić I, Savić B, Švabić-Vlahović M (2000) A modified microtiter-plate test for quantification of staphylococcal biofilm formation. J Microbiol Methods 40:175–179

    PubMed  Google Scholar 

  40. Su S, Panmanee W, Wilson JJ, Mahtani HK, Li Q, VanderWielen BD, Makris TM, Rogers M, McDaniel C, Lipscomb JD, Irvin RT, Schurr MJ, Lancaster JR, Kovall RA, Hassett DJ (2014) Catalase (KatA) plays a role in protection against anaerobic nitric oxide in Pseudomonas aeruginosa. PLoS One 9:e91813

    PubMed  PubMed Central  Google Scholar 

  41. Tang QY, Feng MG (2007) DPS data processing system: experimental design, statistical analysis and data mining. Science Press, Beijing

    Google Scholar 

  42. Vicentini CB, Romagnoli C, Manfredini S, Rossi D, Mares D (2011) Pyrazolo [3,4-c] isothiazole and isothiazolo [4,3-d] isoxazole derivatives as antifungal agents. Pharm Biol 49:545–552

    CAS  PubMed  Google Scholar 

  43. Williams TM (2007) The mechanism of action of isothiazolone biocide. Power Plant Chem 9:14–22

    CAS  Google Scholar 

  44. Zhou G, Li LJ, Shi QS, Ouyang YS, Chen YB, Hu WF (2013) Effects of nutritional and environmental conditions on planktonic growth and biofilm formation for Citrobacter werkmanii BF-6. J Microbiol Biotechnol 23:1673–1682

    CAS  PubMed  Google Scholar 

  45. Zhou G, Li LJ, Shi QS, Ouyang YS, Chen YB, Hu WF (2014) Efficacy of metal ions and isothiazolones in inhibiting Enterobacter cloacae BF-17 biofilm formation. Can J Microbiol 60:5–14

    CAS  PubMed  Google Scholar 

  46. Zhou G, Shi QS, Huang XM, Xie XB (2016) Comparison of transcriptomes of wild-type and isothiazolone-resistant Pseudomonas aeruginosa by using RNA-seq. Mol Biol Rep 43:527–540

    CAS  PubMed  Google Scholar 

  47. Zhou G, Shi QS, Ouyang YS, Chen YB (2014) Involvement of outer membrane proteins and peroxide-sensor genes in Burkholderia cepacia resistance to isothiazolone. World J Microbiol Biotechnol 30:1251–1260

    CAS  PubMed  Google Scholar 

  48. Zumft WG (1997) Cell biology and molecular basis of denitrification. Microbiol Mol Biol Rev 61:533–616

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was funded by the GDAS’ Project of Science and Technology Development (No. 2019GDASYL-0104006), the National Natural Science Foundation of China (Nos. 31770091 and 31500036), and Natural Science Foundation of Guangdong Province (No. 2015A030313713). We are grateful to Prof. Hai-hong Wang of South China Agricultural University for generously providing us the plasmids of pK18-GM and pSRK-GM.

Author information

Author notes

  1. Gang Zhou and Hong Peng contributed equally to this paper.

Authors and Affiliations

  1. Guangdong Open Laboratory of Applied Microbiology, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, State Key Laboratory of Applied Microbiology Southern China, Guangdong Institute of Microbiology, Guangdong Academy of Sciences, Guangzhou, 510070, Guangdong, People’s Republic of China

    Gang Zhou, Hong Peng, Ying-si Wang, Cai-ling Li, Peng-fei Shen, Xiao-mo Huang, Xiao-bao Xie & Qing-shan Shi

Authors
  1. Gang Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hong Peng
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ying-si Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Cai-ling Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Peng-fei Shen
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Xiao-mo Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Xiao-bao Xie
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Qing-shan Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Xiao-bao Xie or Qing-shan Shi.

Ethics declarations

Conflict of interest

The authors declare that they have no conflicts of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Additional file 1.

Detection of differently expressed genes between WT and ΔnirS transcriptomes using RNA-seq technique. Differences with FDR ≤ 0.05 and log2FC absolute value ≥ 1 were set as the threshold for significant differences in gene expression. (XLS 767 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zhou, G., Peng, H., Wang, Ys. et al. Biological functions of nirS in Pseudomonas aeruginosa ATCC 9027 under aerobic conditions. J Ind Microbiol Biotechnol 46, 1757–1768 (2019). https://doi.org/10.1007/s10295-019-02232-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10295-019-02232-z

Keywords

Search

Navigation