An Optical Biosensor from Green Fluorescent Escherichia coli for the Evaluation of Single and Combined Heavy Metal Toxicities
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials
2.2. GFP-Modified E. coli Bacteria
2.3. GFP E. coli Bacterial Cell Culture
2.4. Development of Microbial Biosensor
2.5. Selectivity Study
2.6. Optimization of Whole Cell Biosensor Response
2.7. Reproducibility and Long Term Stability of the Biosensor
2.8. Evaluation of Single and Combined Metal Toxicities via Biosensor and Microtox Assay
3. Results and Discussion
3.1. Characteritics of the Whole Cell Biosensor Response
3.2. Selectivity Study
3.3. Optimization of Experimental Conditions
3.4. Reproducibility and Long Term Stability Studies
3.5. Biosensor Response towards Various Concentrations of Single Toxicants
Toxicants | Dynamic Range (µg/L) | LOD (µg/L) | Slopes (%RFU/µg·L−1) | EC50 (µg/L) | R2 |
---|---|---|---|---|---|
GFP cell biosensor, 2 min (n = 3) | |||||
Cu(II) | (0.05–1) | 0.04 | −45.081 | 0.9 | 0.98 |
Cd(II) | (0.50–10) | 0.32 | −5.015 | 8.9 | 0.99 |
Pb(II) | (0.70–20) | 0.46 | −2.564 | 17.4 | 0.99 |
Zn(II) | (5–100) | 2.80 | −0.506 | 84.4 | 0.99 |
Cr(VI) | (0.10–5) × 103 | 1.00 × 102 | −0.009 | 4.5 × 103 | 0.98 |
Co(II) | (0.50–7) × 103 | 2.50 × 102 | −0.006 | 6.8 × 103 | 0.99 |
Ni(II) | (0.70–10) × 103 | 4.00 × 102 | −0.005 | 9.0 × 103 | 0.98 |
Ag(II) | (1.00–20) × 103 | 7.20 × 102 | −0.002 | 2.0 × 104 | 0.99 |
Fe(III) | (5.00–70) × 103 | 2.60 × 103 | 0.001 | 6.4 × 104 | 0.99 |
Microtox assay, 15 min (n = 3) | |||||
Cu(II) | (0.03–2) × 103 | 10.12 | 38.836× 10−3 * | 1.1 × 103 | 0.98 |
Cd(II) | (0.5–80) × 103 | 0.42 × 103 | 0.953 × 10−3 * | 5.6 × 104 | 0.99 |
Pb(II) | (0.5–80) × 103 | 0.45 × 103 | 0.810 × 10−3 * | 6.1 × 103 | 0.99 |
Zn(II) | (0.5–100) × 103 | 0.46 × 103 | 0.703 × 10−3 * | 6.2 × 104 | 0.99 |
Cr(VI) | (1–150) × 103 | 0.50 × 103 | 0.446 × 10−3 * | 9.0 × 104 | 0.98 |
Co(II) | (10–150) × 103 | 5.60 × 103 | 0.500 × 10−3 * | 9.8 × 104 | 0.99 |
Ni(II) | (1–120) × 103 | 0.65 × 103 | 0.549 × 10−3 * | 6.5 × 104 | 0.98 |
Ag(II) | (1–120) × 103 | 0.52 × 103 | 0.601 × 10−3 * | 6.3 × 104 | 0.96 |
Fe(III) | (15–150) × 103 | 10.25 × 103 | 0.504 × 10−3 * | 9.5 × 104 | 0.97 |
Analyte | Biological Component | Immobilization Matrix | Dynamic Range (μg/L) | LOD (μg/L) | Time (min) | References |
---|---|---|---|---|---|---|
Cu(II) | GFP E. coli | Alginate film | (0.05–1) | 4.0 × 10−2 | 2 | This work |
H. crispa | Agar gel | (0.01–672) × 103 | 6.0 | 20 | [37] | |
A. fischeri | Alginate microspheres | (0.1–2.0) × 102 | 6.4 | 6 | [19] | |
Cd(II) | GFP E. coli | Alginate film | (0.5–10) | 0.32 | 2 | This work |
S. cereviceae | Agar gel | (4.6–45.8) × 103 | 1.83 × 103 | 8 | [37] | |
A. fischeri | Alginate microspheres | (0.2–5.0) × 103 | 1.6 × 102 | 6 | [19] | |
Pb(II) | GFP E. coli | Alginate film | (0.7–20) | 0.36 | 2 | This work |
A. fischeri | Alginate microspheres | (0.5–7) × 102 | 0.5 × 102 | 6 | [19] | |
Zn(II) | GFP E. coli | Alginate film | (5–100) | 2.80 | 2 | This work |
A. fischeri | Alginate microspheres | (0.5–7) × 102 | 0.32 × 103 | 6 | [19] | |
Cr(VI) | GFP E. coli | Alginate film | (0.1–5) × 103 | 1.0 × 102 | 2 | This work |
A.ferrooxidans | Cellulose membrane | (0.02–118) × 103 | 18 | 1 | [36] | |
A. fischeri | Alginate microspheres | (0.1–2 ) × 104 | 1.0 × 103 | 6 | [19] | |
Co(II) | GFP E. coli | Alginate film | (0.5–7) × 103 | 2.5 × 102 | 2 | This work |
A. fischeri | Alginate microspheres | (0.5–5.0) × 104 | 1.7 × 103 | 6 | [19] | |
Ni(II) | GFP E. coli | Alginate film | (0.7–10) × 103 | 4.2 × 102 | 2 | This work |
B. sphaericus | Whatman membrane | (2–40) | 0.02 | 2 | [38] | |
A. fischeri | Alginate microspheres | (0.5–7) × 104 | 2.8 × 103 | 6 | [19] | |
Ag(I) | GFP E. coli | Alginate film | (0.1–2) ×104 | 7.2 × 102 | 2 | This work |
A. fischeri | Alginate microspheres | (0.2–7) × 104 | 1.8 × 103 | 6 | [19] | |
Fe(III) | GFP E. coli | Alginate film | (5.0–70) × 103 | 2.60 × 103 | 2 | This work |
A. fischeri | Alginate microspheres | (0.5–7) × 104 | 0.31 × 104 | 6 | [19] |
Heavy Metals and Incubation Times | This Work | Futra et al. [19] |
---|---|---|
Times (min) | 2 | 6 |
Cu(II) (μg/L) | 0.9 | 1.7 × 102 |
Cd(II) (μg/L) | 8.9 | 6.3 × 103 |
Pb(II) (μg/L) | 17.4 | 0.7 × 103 |
Zn(II) (µg/L | 84.4 | 6.0 × 102 |
Cr(VI) (μg/L) | 4.5 × 103 | 1.8 × 104 |
Co(II) (μg/L) | 6.8 × 103 | 6.6 × 104 |
Ni(II) (μg/L) | 9.0 × 103 | 6.6 × 104 |
Ag(I) (μg/L) | 2.0 × 104 | 6.0 × 104 |
Fe(III) (µg/L) | 6.4 × 104 | 7.0 × 104 |
3.6. Biosensor Response towards Combined Metals
Toxicant Mixture | AI | Toxicity Rate |
---|---|---|
(1:1 w/w) | ||
Pb(II) + Zn(II) | −0.41 | Antagonistic |
Cu(II) + Zn(II) | −3.73 | Antagonistic |
Cu(II) + Pb(II) | −1.72 | Antagonistic |
Cd(II) +Zn(II) | −1.62 | Antagonistic |
Cd(II) + Pb(II) | −2.94 | Antagonistic |
Cd(II) + Cu(II) | −1.97 | Antagonistic |
(2:1 w/w) | ||
Pb(II) + Zn(II) | −0.27 | Antagonistic |
Cu(II) + Zn(II) | −1.47 | Antagonistic |
Cu(II) + Pb(II) | −1.79 | Antagonistic |
Cd(II) +Zn(II) | −0.72 | Antagonistic |
Cd(II) + Pb(II) | −2.07 | Antagonistic |
Cd(II) + Cu(II) | −1.87 | Antagonistic |
(1:2 w/w) | ||
Pb(II) + Zn(II) | −0.878 | Antagonistic |
Cu(II) + Zn(II) | −1.64 | Antagonistic |
Cu(II) + Pb(II) | −2.00 | Antagonistic |
Cd(II) +Zn(II) | −0.75 | Antagonistic |
Cd(II) + Pb(II) | −1.58 | Antagonistic |
Cd(II) + Cu(II) | −2.88 | Antagonistic |
(1:1:1 to 1:1:1:1 w/w) | ||
Cu(II) + Cd(II) + Pb(II) | −1.984 | Antagonistic |
Cu(II) + Cd(II) + Zn(II) | −1.990 | Antagonistic |
Cd(II) + Pb(II) + Zn(II) | −3.186 | Antagonistic |
Cu(II) + Cd(II) + Pb(II) + Zn(II) | −6.033 | Antagonistic |
4. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
- Duffus, J. Heavy metals—A meaningless term; chemistry and human health division clinical chemistry section commission on toxicology. Pure Appl. Chem. 2002, 74, 793–807. [Google Scholar]
- Friberg, L.; Nordberg, G.; Voulk, V. Handbook on the Toxicology of Metals; Elsevier/North-Holland Biomedical Press: Now York, NY, USA, 2005. [Google Scholar]
- Prescott, L.M.; Harley, J.; Klein, J. Microbiology, 5th ed.; McGraw-Hill Companies: New York, NY, USA, 2002. [Google Scholar]
- Paoliello, M.M.B.; Capitani, E.E.D.; da Cunha, F.G.; Matsuo, T.; Carvalho, M.D.F.; Sakuma, A.; Figueiredo, B.R. Exposure of children to lead and cadmium from a mining area of Brazil. Environ. Res. 2002, 88, 120–128. [Google Scholar] [CrossRef] [PubMed]
- Batista, B.L.; Rodrigues, J.L.; Nunes, J.A.; Tormen, L.; Curtius, A.J.; Barbosa, J.F. Simultaneous determination of Cd, Cu, Mn, Ni, Pb and Zn in nail samples by inductively coupled plasma mass spectrometry (ICP-MS) after tetramethylammonium hydroxide solubilization at room temperature: Comparison with ETAAS. Talanta 2008, 76, 575–579. [Google Scholar] [CrossRef] [PubMed]
- Manutsewee, N.; Aeungmaitrepirom, W.; Varanusupakul, P.; Imyim, A. Determination of Cd, Cu, and Zn in fish and mussel by AAS after ultrasound-assisted acid leaching extraction. Food Chem. 2007, 101, 817–824. [Google Scholar] [CrossRef]
- Liao, V.; Chien, M.; Tseng, Y.; Ou, T. Assessment of heavy metal bioavailability in contaminated sediments and soils using green fluorescence protein-based bacterial biosensors. Environ. Pollut. 2006, 142, 17–23. [Google Scholar] [CrossRef] [PubMed]
- Fulladosa, E.; Murat, J.-C.; Villaescusa, I. Study on the toxicity of binary equitoxic mixtures of metals using the luminescent bacteria Vibrio fischeri as a biological target. Chemosphere 2005, 58, 551–557. [Google Scholar] [CrossRef] [PubMed]
- Cho, J.-C.; Park, K.-J.; Ihm, H.-S.; Park, J.-E.; Kim, S.-Y.; Kang, I.; Lee, K.-H.; Jahng, D.; Lee, D.-H.; Kim, S.-J. A novel continuous toxicity test system using a luminously modified freshwater bacterium. Biosens. Bioelectron. 2004, 20, 338–344. [Google Scholar] [CrossRef] [PubMed]
- Dawson, J.J.C.; Campbell, C.D.; Towers, W.; Cameron, C.M.; Paton, G.I. Linking biosensor responses to Cd, Cu, and Zn partitioning in soils. Environ. Pollut. 2006, 142, 493–500. [Google Scholar] [CrossRef] [PubMed]
- Yim, J.H.; Kim, K.W.; Kim, S.D. Effect of hardness on acute toxicity of metal mixtures using Daphnia magna: Prediction of acid mine drainage toxicity. J. Hazard. Mater. 2006, 138, 16–21. [Google Scholar] [CrossRef] [PubMed]
- Chakraborty, T.; Babu, P.G.; Alam, A.; Chaudhari, A. GFP expressing bacterial biosensor to measure lead contamination in aquatic environmental. Curr. Sci. 2008, 94, 800–805. [Google Scholar]
- Arias-Barreiro, C.R.; Okazaki, K.; Koutsaftis, A.; Inayat-Hussain, S.H.; Tani, A.; Katsuhara, M.; Kimbara, K.; Mori, I.C. A bacterial biosensor for oxidative stress using the constitutively expressed redox-sensitive protein roGFP2. Sensors 2010, 10, 6290–6306. [Google Scholar] [CrossRef] [PubMed]
- Prachayasittikul, V.; Ayudhya, C.I.; Bulow, L. Lighting Escherichia coli cells biological sensor for Cd2+. Biotechnol. Lett. 2001, 23, 1285–1291. [Google Scholar] [CrossRef]
- Tansila, N.; Tantimongcolwat, T.; Isarankura-Na-Ayudha, C.; Nantasenamat, C.; Prachayasittikul, V. Rational design of analyte channel of the green fluorescent protein for biosensor applications. Int. J. Biol. Sci. 2007, 3, 463–470. [Google Scholar] [CrossRef] [PubMed]
- Ivask, A.; Virta, M.; Kahru, A. Construction and use of specific luminescent recombinant bacterial sensor for the assessment of bioavailable fraction of cadmium, zinc, mercury and chromium in the soil. Soil Biol. Biochem. 2002, 32, 1439–1447. [Google Scholar] [CrossRef]
- Ivask, A.; Francois, M.; Kahru, A.; Dubourguier, H-C.; Virta, M.; Douay, F. Recombinant luminescent bacterial sensors for the measurement of bioavailability of cadmium and lead in soil polluted by metal smelters. Chemosphere 2004, 55, 147–156. [Google Scholar] [CrossRef] [PubMed]
- Bondarenko, O.; Rolova, T.; Kahru, A.; Ivask, A. Bioavailability of Cd, Zn, and Hg in soil to nine recombinant luminescent metal sensor bacteria. Sensors 2008, 8, 6899–6923. [Google Scholar] [CrossRef]
- Futra, D.; Heng, L.Y.; Surif, S.; Ahmad, A.; Ling, T.L. Microencapsulated Aliivibrio fischeri in Alginate Microspheres for Monitoring Heavy Metal Toxicity in Environmental Waters. Sensors 2014, 14, 23248–23268. [Google Scholar] [CrossRef] [PubMed]
- Bomati, E.K.; Haley, J.E.; Noel, J.P.; Deheyn, D.D. Spectral and structural comparison between bright and dim green fluorescent proteins in Amphioxus. Sci. Rep. 2014, 4, 1–9. [Google Scholar] [CrossRef] [PubMed]
- Segami, S.; Makino, S.; Miyake, A.; Asaoka, M.; Maeshima, M. Dynamics of vacuoles and H+-pyrophosphatase visualized by monomeric green fluorescent protein in Arabidopsis: Artifactual bulbs and native intravacuolar spherical structures. Plant Cell 2014, 26, 3416–3434. [Google Scholar] [CrossRef] [PubMed]
- Venning-Slater, M.; Hooks, D.O.; Rehm, B.H.A. In vivo self-assembly of stable green fluorescent protein fusion particles and their uses in enzyme immobilization. Appl. Environ. Microbiol. 2014, 80, 3062–3071. [Google Scholar] [CrossRef] [PubMed]
- Bird, L.E.; Rada, H.; Verma, A.; Gasper, R.; Birch, J.; Jennions, M.; Lӧwe, J.; Moraes, I.; Owens, R.J. Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli. J. Vis. Exp. 2015, 95, 1–7. [Google Scholar]
- Dooley, C.T.; Dore, T.M.; Hanson, G.T.; Jackson, W.C.; Remington, S.J.; Tsien, R.Y. Imaging dynamic redox changes in mamalia cells with green fluorescent protein. J. Biol. Chem. 2004, 279, 22284–22293. [Google Scholar] [CrossRef] [PubMed]
- Hanson, G.T.; Aggeler, R.; Oglesbee, D.; Cannon, M.; Capaldi, R.A.; Tsien, R.Y.; Remington, S.J. Investigating mitochondrial redox potential with redox-sensitive green fluorescent protein indicators. J. Biol. Chem. 2004, 279, 13044–13053. [Google Scholar] [CrossRef] [PubMed]
- Finney, D.J. Statistical Method in Biological Assay, 3th ed.; Charles Griffin: London, UK, 1978. [Google Scholar]
- Hao, W.; Li, Q.; Zhang, J.; Jiang, Y.; Liang, W. Utility of nematode Acrobeloidesnanus for assessing subacute toxicity of heavy metals. Environ. Monit. Assess. 2009, 164, 273–278. [Google Scholar] [CrossRef] [PubMed]
- Sorensen, S.; Burmolle, M.; Hansen, L. Making bio-sensor of toxicity; new development in whole cell biosensors. Curr. Opin. Biotechnol. 2006, 17, 11–16. [Google Scholar] [CrossRef] [PubMed]
- Belkin, S. Genetically engineered microorganisms for pollution monitoring. Soil Water Pollut. Monit. Prot. Remed. 2006, 3–23, 147–160. [Google Scholar]
- Kim, B.C.; Gu, M.B. A bioluminescent sensor for high throughput toxicity classification. Biosen. Bioelectron. 2003, 18, 1015–1021. [Google Scholar] [CrossRef]
- Gil, G.C.; Mitchell, R.J.; Chang, S.T.; Gu, M.B. A biosensor for the detection of gas toxicity using a recombinant bioluminescent bacterium. Biosen. Bioelectron. 2000, 15, 23–30. [Google Scholar]
- Arica, M.Y.; Kacar, Y.; Genc, O. Entrapment of white-rot fungus Tremetes versicolor in Ca-alginate beads: Preparation and biosorption kinetic analysis for cadmium removal from an aquas solution. Bioresour. Technol. 2001, 80, 121–129. [Google Scholar] [CrossRef]
- Southward, G.M.; Klein, D. Effects of pH on Prymnesium parvum Cell Viability and Toxicity. In Management of Prymnesium Parvum at Texas State Fish Hatcheries; Texas Parks and Wildlife: Dallas, TX, USA, 2005; Chapter 6. [Google Scholar]
- Campbell, T.N.; Choy, F.Y.M. The effect of pH on green fluorescent protein: A brief review. Mol. Biol. Today 2001, 1, 1–4. [Google Scholar]
- Miller, J.N.; Miller, J.C. Statistics and Chemometrics for Analytical Chemistry, 5th ed.; Pearson Prentice Hall: London, UK, 2005. [Google Scholar]
- Quig, D. Cysteine metabolism and metal toxicity. Altern. Med. Rev. 1998, 3, 262–270. [Google Scholar] [PubMed]
- Zlatev, R.; Magnin, J.-P.; Ozil, P.; Stoytcheva, M. Bacterial sensors based on Acidithiobacillus ferrooxidans part I. Fe2+ and S2O32− determination. Biosens. Bioelectron. 2006, 21, 1493–1500. [Google Scholar] [CrossRef] [PubMed]
- Rahimi, Y.; Shrestha, S.; Banerjee, T.; Deo, S.K. Copper sensing based on the far-red fluorescent protein, HcRed, from Heteractis crispa. Anal. Biochem. 2007, 370, 60–67. [Google Scholar] [CrossRef] [PubMed]
- Verma, N.; Singh, M. A Bacillus sphaericus based biosensor for monitoring nickel ions in industrial effluents and foods. J. Autom. Methods Manag. Chem. 2006, 2006, 1–4. [Google Scholar] [CrossRef] [PubMed]
© 2015 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/4.0/).
Share and Cite
Futra, D.; Heng, L.Y.; Ahmad, A.; Surif, S.; Ling, T.L. An Optical Biosensor from Green Fluorescent Escherichia coli for the Evaluation of Single and Combined Heavy Metal Toxicities. Sensors 2015, 15, 12668-12681. https://doi.org/10.3390/s150612668
Futra D, Heng LY, Ahmad A, Surif S, Ling TL. An Optical Biosensor from Green Fluorescent Escherichia coli for the Evaluation of Single and Combined Heavy Metal Toxicities. Sensors. 2015; 15(6):12668-12681. https://doi.org/10.3390/s150612668
Chicago/Turabian StyleFutra, Dedi, Lee Yook Heng, Asmat Ahmad, Salmijah Surif, and Tan Ling Ling. 2015. "An Optical Biosensor from Green Fluorescent Escherichia coli for the Evaluation of Single and Combined Heavy Metal Toxicities" Sensors 15, no. 6: 12668-12681. https://doi.org/10.3390/s150612668
APA StyleFutra, D., Heng, L. Y., Ahmad, A., Surif, S., & Ling, T. L. (2015). An Optical Biosensor from Green Fluorescent Escherichia coli for the Evaluation of Single and Combined Heavy Metal Toxicities. Sensors, 15(6), 12668-12681. https://doi.org/10.3390/s150612668