Skip to content
1887

Access Microbiology open research platform logo Access Microbiology

Volume 8, Issue 1

Application of the solvent effect on bioluminescent reporter bacteria as a real-time membrane toxicity assay Open Access

Abstract

Bioluminescent bioreporters are widely used across various scientific disciplines due to the well-characterized bacterial bioluminescence mechanism. However, solvent-induced membrane perturbations may confound the use of bioreporters in assessing cellular toxicity from environmental contaminants. This study investigated the solvent effect, wherein membrane damage increases intracellular availability of bioluminescent reaction precursors, increasing the light produced. A new online monitoring system was also tested with multiple bioluminescent reporters, including a newly constructed M3A strain, exposed to toluene, trichloroethylene, acetone, phenol and creosote derived from beechwood tar. Additional tests included the introduction of carbon nanotubes, fullerene and fullerenol. A solvent effect was confirmed by the detection of increased bioluminescent signal and the occurrence of fatty acid release (<0.05). Phenol (25 p.p.m.), a benchmark for bactericidal activity, demonstrated luminescence enhancement via the solvent effect. Membrane toxicity assays showed that M3A responded sensitively to sublethal and lethal membrane disruptions, whereas MJ1 did not exhibit a solvent effect, and its luminescence changes were not correlated with viability (>0.05). These results indicate that M3A is a sensitive biosensor for detecting environmental contaminants and identifying both lethal and sublethal membrane perturbations. The findings underscore essential considerations when utilizing bacterial bioluminescence as a proxy for gene expression or cellular physiology.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): Bioluminescence , bioreporter , Pseudomonas fluorescens and solvent
Funding
This study was supported by the:
  • USDA National Institute of Food and Agriculture (Award Hatch project number 1004830 and from USDA ARS Project No. 8072-42000-077)
    • Principal Award Recipient: BruceM. Applegate
© 2026 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
Loading

Article metrics loading...

/content/journal/acmi/10.1099/acmi.0.001096.v3
2026-01-08
2026-01-15

Metrics

Loading full text...

Full text loading...

/deliver/fulltext/acmi/8/1/acmi001096.v3.html?itemId=/content/journal/acmi/10.1099/acmi.0.001096.v3&mimeType=html&fmt=ahah

References

  1. Al-Hindi RR, Teklemariam AD, Alharbi MG, Alotibi I, Azhari SA et al. Bacteriophage-based biosensors: a platform for detection of foodborne bacterial pathogens from food and environment. Biosensors (Basel) 2022; 12:905 [View Article] [PubMed]
     [Google Scholar]
  2. Applegate BM, Kehrmeyer SR, Sayler GS. A chromosomally based tod-luxCDABE whole-cell reporter for benzene, toluene, ethybenzene, and xylene (BTEX) sensing. Appl Environ Microbiol 1998; 64:2730–2735 [View Article] [PubMed]
     [Google Scholar]
  3. Heitzer A, Applegate B, Kehrmeyer S, Pinkart H, Webb OF et al. Physiological considerations of environmental applications of lux reporter fusions. J Microbiol Methods 1998; 33:45–57 [View Article]
     [Google Scholar]
  4. Kelly CJ, Tumsaroj N, Lajoie CA. Assessing wastewater metal toxicity with bacterial bioluminescence in a bench-scale wastewater treatment system. Water Res 2004; 38:423–431 [View Article] [PubMed]
     [Google Scholar]
  5. Tong Z, Bischoff M, Nies LF, Myer P, Applegate B et al. Response of soil microorganisms to as-produced and functionalized single-wall carbon nanotubes (SWNTs). Environ Sci Technol 2012; 46:13471–13479 [View Article]
     [Google Scholar]
  6. Suwal S, Coronel-Aguilera CP, Auer J, Applegate B, Garner AL et al. Mechanism characterization of bacterial inactivation of atmospheric air plasma gas and activated water using bioluminescence technology. Innov Food Sci Emerg Technol 2019; 53:18–25 [View Article]
     [Google Scholar]
  7. Xu L, Yepez X, Applegate B, Keener KM, Tao B et al. Penetration and microbial inactivation by high voltage atmospheric cold plasma in semi-solid material. Food Bioprocess Technol 2020; 13:1688–1702 [View Article]
     [Google Scholar]
  8. Elasri MO, Reid T, Hutchens S, Miller RV. Response of a Pseudomonas aeruginosa biofilm community to DNA-damaging chemical agents. FEMS Microbiol Ecol 2000; 33:21–25 [View Article]
     [Google Scholar]
  9. Mioni CE, Howard AM, DeBruyn JM, Bright NG, Twiss MR et al. Characterization and field trials of a bioluminescent bacterial reporter of iron bioavailability. Marine Chem 2003; 83:31–46 [View Article]
     [Google Scholar]
  10. Delgado-Mellado N, Ayuso M, Villar-Chavero MM, García J, Rodríguez F. Ecotoxicity evaluation towards Vibrio fischeri of imidazolium- and pyridinium-based ionic liquids for their use in separation processes. SN Appl Sci 2019; 1:1–9 [View Article]
     [Google Scholar]
  11. Parvez S, Venkataraman C, Mukherji S. A review on advantages of implementing luminescence inhibition test (Vibrio fischeri) for acute toxicity prediction of chemicals. Environ Int 2006; 32:265–268 [View Article] [PubMed]
     [Google Scholar]
  12. Zadorozhnaya O, Kirsanov D, Buzhinsky I, Tsarev F, Abramova N et al. Water pollution monitoring by an artificial sensory system performing in terms of Vibrio fischeri bacteria. Sensors and Actuators B: Chemical 2015; 207:1069–1075 [View Article]
     [Google Scholar]
  13. Myer PR. Construction, Characterization, and Application of the Bioluminescent Bioreporter Pseudomonas Fluorescens M3A Purdue University; 2013
     [Google Scholar]
  14. White GP, Dunn NW. Apparent fusion of the TOL plasmid with the R91 drug resistance plasmid in Pseudomonas aeruginosa. Aust J Biol Sci 1977; 30:345–355 [View Article] [PubMed]
     [Google Scholar]
  15. Lee KG, Lee SE, Takeoka GR, Kim JH, Park BS. Antioxidant activity and characterization of volatile constituents of beechwood creosote. J Sci Food Agric 2005; 85:1580–1586 [View Article]
     [Google Scholar]
  16. Sayler GS, Puziss M, Silver M. Alkaline phosphatase assay for freshwater sediments: application to perturbed sediment systems. Appl Environ Microbiol 1979; 38:922–927 [View Article] [PubMed]
     [Google Scholar]
  17. Baglioni S. Relations between the physiological action and the chemical constitution. studies on the action of benzene derivatives on the central nervous system. Z Allgem Physiol 1904; 3:313–358
     [Google Scholar]
  18. Sikkema J, de Bont JA, Poolman B. Mechanisms of membrane toxicity of hydrocarbons. Microbiol Rev 1995; 59:201–222 [View Article] [PubMed]
     [Google Scholar]
  19. Zhu B, Xia X, Xia N, Zhang S, Guo X. Modification of fatty acids in membranes of bacteria: implication for an adaptive mechanism to the toxicity of carbon nanotubes. Environ Sci Technol 2014; 48:4086–4095 [View Article]
     [Google Scholar]
  20. Ingram LO. Changes in lipid composition of Escherichia coli resulting from growth with organic solvents and with food additives. Appl Environ Microbiol 1977; 33:1233–1236 [View Article] [PubMed]
     [Google Scholar]
  21. Ingram LO. Regulation of fatty acid composition in Escherichia coli: a proposed common mechanism for changes induced by ethanol, chaotropic agents, and a reduction of growth temperature. J Bacteriol 1982; 149:166–172 [View Article] [PubMed]
     [Google Scholar]
  22. Meighen EA. Molecular biology of bacterial bioluminescence. Microbiol Rev 1991; 55:123–142 [View Article] [PubMed]
     [Google Scholar]
  23. Pinkart HC, Wolfram JW, Rogers R, White DC. Cell envelope changes in solvent-tolerant and solvent-sensitive Pseudomonas putida strains following exposure to o-Xylene. Appl Environ Microbiol 1996; 62:1129–1132 [View Article] [PubMed]
     [Google Scholar]
  24. Tullai JW, Schaffer ME, Mullenbrock S, Sholder G, Kasif S et al. Immediate-early and delayed primary response genes are distinct in function and genomic architecture. J Biol Chem 2007; 282:23981–23995 [View Article] [PubMed]
     [Google Scholar]
  25. Rideal S, Ainslie Walker JT. Standardisation of disinfectants. J Sanit Inst 1903; 24:424–441 [View Article]
     [Google Scholar]
  26. McCann JH, Solomon KR. The effect of creosote on membrane ion leakage in Myriophyllum spicatum L. Aquat Toxicol 2000; 50:275–284 [View Article] [PubMed]
     [Google Scholar]
  27. Holmannova D, Borsky P, Svadlakova T, Borska L, Fiala Z. Carbon nanoparticles and their biomedical applications. Appl Sci 2022; 12:7865 [View Article]
     [Google Scholar]
  28. Khabashesku VN, Margrave JL, Barrera EV. Functionalized carbon nanotubes and nanodiamonds for engineering and biomedical applications. Diam Relat Mater 2005; 14:859–866 [View Article]
     [Google Scholar]
  29. Simon J, Flahaut E, Golzio M. Overview of carbon nanotubes for biomedical applications. Materials 2019; 12:624 [View Article] [PubMed]
     [Google Scholar]
  30. Weiss DR, Raschke TM, Levitt M. How hydrophobic buckminsterfullerene affects surrounding water structure. J Phys Chem B 2008; 112:2981–2990 [View Article] [PubMed]
     [Google Scholar]
  31. Bedrov D, Smith GD, Davande H, Li L. Passive transport of C60 fullerenes through a lipid membrane: a molecular dynamics simulation study. J Phys Chem B 2008; 112:2078–2084 [View Article] [PubMed]
     [Google Scholar]
  32. Arbogast JW, Foote CS, Kao M. Electron transfer to triplet fullerene C60. J Am Chem Soc 1992; 114:2277–2279 [View Article]
     [Google Scholar]
  33. Byrne H, Maser W, Mittelbach A, Roth S. Nonlinear luminescence phenomena in fullerene crystallites. Appl Phys A 1993; 56:235–239 [View Article]
     [Google Scholar]
  34. Duarte-Gómez EE, Graham D, Budzik M, Paxson B, Csonka L et al. High hydrostatic pressure effects on bacterial bioluminescence. LWT - Food Sci Technol 2014; 56:484–493 [View Article]
     [Google Scholar]
  35. Lupp C, Urbanowski M, Greenberg EP, Ruby EG. The Vibrio fischeri quorum-sensing systems ain and lux sequentially induce luminescence gene expression and are important for persistence in the squid host. Mol Microbiol 2003; 50:319–331 [View Article] [PubMed]
     [Google Scholar]
  36. King JM, Digrazia PM, Applegate B, Burlage R, Sanseverino J et al. Rapid, sensitive bioluminescent reporter technology for naphthalene exposure and biodegradation. Science 1990; 249:778–781 [View Article] [PubMed]
     [Google Scholar]
  37. Myer P, Busto-Ramos M, Hartono L, Applegate B. Non-fiber-optic bioluminescent biosensors. In Thouand G, Marks RS. eds Bioluminescent Microbial Biosensors: Design, Construction, and Implementation Jenny Stanford Publishing; 2015
     [Google Scholar]
  38. Chen Y. Use of Bioluminescence to Examine Parameters Associated with Salmonella Enterica Serotype Poona and Escherichia Coli O157: H7 Contamination of Produce. Purdue University; 2008
     [Google Scholar]
  39. Farris LA. Evaluation of Escherichia coli O157: H7 Internationalization into Romaine Lettuce Tissues and Detection of Metabolic Changes of Internalized Bacteria in Response to Bioavailable Carbon Using Developed Bioluminescent Assay. Purdue University; 2007
     [Google Scholar]
  40. Farris L, Habteselassie MY, Perry L, Chen Y, Turco R et al. Luminescence techniques for the detection of bacterial pathogens. PBD-BRRM 2008213–230 [View Article]
     [Google Scholar]
  41. Ruby EG, Nealson KH. Symbiotic association of Photobacterium fischeri with the marine luminous fish Monocentris japonica; a model of symbiosis based on bacterial studies. Biol Bull 1976; 151:574–586 [View Article] [PubMed]
     [Google Scholar]
/content/journal/acmi/10.1099/acmi.0.001096.v3
Loading
Application of the solvent effect on bioluminescent reporter bacteria as a real-time membrane toxicity assay
Access Microbiology 8, 001096.v3 (2026); https://doi.org/10.1099/acmi.0.001096.v3
/content/journal/acmi/10.1099/acmi.0.001096.v3
/content/journal/acmi/10.1099/acmi.0.001096.v3
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited Most Cited RSS feed

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