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Why focus on atmospheric pollutants and especially VOC ?

It is known that pollution has harmful effects on human health. According to the world health organization (WHO), 7 million premature deaths are due to atmospheric pollution, which equals to one eighth of the world annual deaths. The cost of those premature deaths in the WHO countries is $ 1.431 trillion per year. In France, there are 3.5 million asthmatic people, 50 000 people suffering from serious respiratory deficiency, 150 000 deaths of patients suffering from cardiovascular diseases . In addition of public health issues, those pollutants can have a harmful effect on the environment as they acidify waters and soils and decrease vegetal growth. Those effects trigger a decline in agricultural yield and alter aquatic ecosystems.

Why using a biosensor instead of existing methods ?

Why realizing a Biosensor? Air and water pollution due to a growing industrialization is on a major concern and the need for continuous on-line monitoring is growing, as explain previously. To address this need, scientists are developing living organisms able to detect the presence of pollutants, and whole-cell biosensors are seen to have particular advantages in such environmental monitoring (8,10). Those advantages are explained in details below. On the one hand, whole cell biosensors for environmental application require selection of a biocatalyst sensitive to the pollutant or group of pollutants to be detected, a transducer capable of following those metabolic events disturbed by the pollutant; and a suitable method of presentation of the biocatalyst to the transducer. (10) As the recognition profile of a promoter, used as the detector element, is usually made up of only a few molecules, which makes biosensors a precise and accurate measurement tool. In addition, according to the strength of the output signals, the quantity of initial environment signals (quantity of pollutants) can be determined (2).

Also, by changing the genetic construction, the targeted pollutant can be change as well as the sensibility of a biosensor, i.e. the importance of its response to a given stimulus, can be modulated by modifying its genetic construction. It was demonstrated by the 2009 Cambridge iGEM team in their E.chromi project (5). The purpose of that project was to create kits of parts that would help the design and construction of future biosensors. They developed a sensitivity tuner that could affect the output by changing the input-sensitive promoter’s response behavior, and report a response, using clear, user-friendly outputs.

Another major advantages of biosensors are their ability, depending on the genetic construction to detect several environmental signals simultaneously and to transduce those signals into different outputs (different bioluminescence color, fluorescence emission). Then, different kinds of bacteria able to detect two different pollutants, could be introduced in the same device and respond with different color output. On the other hand, as biosensors use cellular machinery, they do not require any other energy input than water, sugar and salt. This fact presents a major advantages compare to actual pollution detection systems which are energy and water consumers. Results obtain from biosensor can be analyze without expensive machinery2. Also, when carry in the appropriate container that do not allow the release, biosensors are very suitable for field applications as they are more easily transportable than actual chemical physical measurement device. Thus, accurate and sensitive whole cell biosensors for pollutants detection and quantification provide useful alternatives to direct analytical methods because of their low cost, rapid response, reproducibility, and ease of production. (11)

In addition, living cells do not only give analytical information but also functional information on the effect of a stimulus on an organism. In the opposite of traditional analytical chemistry detection, biosensors are able to distinguish pollutants that are available to biological systems from those that exist in the environment in inert, unavailable forms and therefore the real impact of pollutants on living organisms (3)(4). This information is important and relevant for environmental measures1, 12.

Even if it is known that biosensors present a good alternative for continuous on-line monitoring, only few tools using biosensors are currently commercialize. Aware of this fact and of the numerous advantages that present biosensors in environmental monitoring, our team had decided to realize one to detect atmospheric pollutions.

(1) Luc BOUSSE, « Whole cell biosensors », sensors and actuators, 1996 (2) Website of iGEM Peking 2013 available : http://2013.igem.org/Team:Peking/Project/BioSensors (3) X. Liu, K. J.Germaine,D. Ryan, andD.N.Dowling, “Whole-cell fluorescent biosensors for bioavailability and biodegradation of polychlorinated biphenyls,” Sensors, vol. 10, no. 2, pp. 1377–1398, 2010. (4) Wong, L. S., Lee, Y. H. & Surif, S. Whole Cell Biosensor Using Anabaena torulosa with Optical Transduction for Environmental Toxicity Evaluation. Journal of Sensors 2013, 1–8 (2013). (5) Website of iGEM Cambridge 2009 available on : http://2009.igem.org/Team:Cambridge (8) Hernandez C.A., Osma, J.F. (2014). Whole cell biosensors. In M. Stoytcheva & J.F. Osma (Eds.). Biosensors: Recent Advances and Mathematical Challenges. Barcelona: OmniaScience, pp. 51-96. (9) Charrier, T., Jos, M., Affi, M., Jouanneau, S., Gezekel, H., and Thou, G. (2010). Bacterial Bioluminescent Biosensor Characterisation for On-line Monitoring of Heavy Metals Pollutions in Waste Water Treatment Plant Effluents. In Biosensors, P. Andrea, ed. (InTech) (10) Rawson, D.M., Willmer, A.J., and Turner, A.P. (1989). Whole-cell biosensors for environmental monitoring. Biosensors 4, 299–311. (11) Behzadian, F., Barjeste, H., Hosseinkhani, S., and Zarei, A.R. (2011). Construction and Characterization of Escherichia coli Whole-Cell Biosensors for Toluene and Related Compounds. Current Microbiology 62, 690–696. (12) Selifonova, O., Burlage, R., and Barkay, T. (1993). Bioluminescent sensors for detection of bioavailable Hg(II) in the environment. Appl. Environ. Microbiol. 59, 3083–3090.

Why choosing Bioluminescence ?

Bioluminescence is currently mainly use for transcription study and cell imaging. We want to develop a new application for bioluminescence that should allow us to create a biosensor of a new kind, different from the existing devices. This is why we decided to based our biosensor on the use of bioluminescence as the reporter gene. This method has become increasingly popular for quantitative analysis . The transcription of luciferase in the presence of specific pollutants and the use of its substrate as a manner to assess pollutant concentration is a new, easy and innovative approach , yet poorly studied.

Nunes-Halldorson, V. da S., and Duran, N.L. (2003). Bioluminescent bacteria: lux genes as environmental biosensors. Brazilian Journal of Microbiology 34. Wood, K (2007).« The bioluminescence advantage », Promega corporation. Welsh, D.K., and Noguchi, T. (2012). Cellular Bioluminescence Imaging. Cold Spring Harbor Protocols 2012, pdb.top070607. http://www.labnews.co.uk/features/the-bioluminescence-advantage-13-09-2011/ BIOLUMINESCENT BACTERIA: LUX GENES AS ENVIRONMENTAL BIOSENSORS Keith WOOD, « The bioluminescence advantage », Promega corporation, 2007 Welsh DK, Noguchi T. (2012), “Cellular Bioluminescence Imaging”, in Cold Spring Harb Protoc; 2012 http://parts.igem.org/Protein_coding_sequences/Reporters https://france.promega.com/products/reporter-assays-and-transfection/reporter-assays/nanoluc-luciferase-redefining-reporter-assays/