Why focus on atmospheric pollutants and especially VOC ?

Atmospheric pollution is characterized by the presence of gas and particles (nitrogen oxides, carbon oxides, ozone, particles PM10 and PM25, Volatile Organic Compounds, black smoke) in the outside air with harmful impact on human health and/or on the environment. [1] [2].

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, equal to one eighth of the world annual deaths. [3] The estimated cost of these premature deaths in WHO countries is $ 1.6 trillion per year. [4]
In France there are 3.5 million people with asthma, 50 000 people suffering from serious respiratory deficiencies, and 150 000 deaths of patients suffering from cardiovascular diseases. [5] All those diseases originate or at least are aggravated by atmospheric pollution.
In addition to public health issues, these pollutants can have a harmful effect on the environment as they acidify water and soil and decrease plant growth. These changes trigger a decline in agricultural yields and alter aquatic ecosystems. [5]

Atmospheric pollution is an accumulation of complex phenomena. To understand local and regional emissions, meteorological conditions, transport, and pollutant transformation must be considered. [2]

Our team decided to focus on VOCs for several reasons. VOCs are organic chemicals with a high vapor pressure (0.01kPA) at room temperature (293.15K = 20°C). Their high vapor pressure results from a low boiling point, which causes large numbers of molecules to evaporate from the liquid or sublimate from the solid form of the compound and enter the ambient air, a trait known as volatility. [1,6] Over the past ten years the average toluene concentration in the periphery of Paris was 6.1 µg/m3 with a maximum of 24 µg/m3 in traffic area. [2]
These primary pollutants originate from fuel evaporation, car traffic, industrial processes, heating systems in residential areas, domestic use of solvents and also from vegetation. They have a role in secondary particle and ozone formation [1] [6].

Although air pollution contains only 2% VOCs (see figure) [2], the impact of those pollutants on health and environment is major.
These pollutants are persistent in the environment, they bio-accumulate in living tissues and are able spread over long distances. Although their effect on human health are only partially known, scientists have demonstrated their systemic effects (hepatic, hematologic, immunologic), their toxicity on the reproductive systems, their genetic toxicity. VOCs are known to be carcinogenic.

Therefore, to protect populations and the environment against harmful pollutants, regulations were defined and air quality is supervised according to European directives [1]. At the European level, the 2008/50/CE directive of May 21st 2008, specifies air quality standards in Europe. This text was transposed into French law by decree n°2010-1250 of October 21st 2010 on air quality. Specific directives are applied to VOCs, such as the European 2004/107/CE directive, dating from December 20th 1994, concerning VOC emission from fuel storage and fuel distribution in gas stations.
Even if pollutant concentrations are getting lower, their final concentrations do not respect the threshold of 5 μg/m3 defined by the law. The objective to reach for the annual mean concentration is 2 μg/m3.

Bioluminescence has been recognized as an alternative to fluorescence [21] and has become widely preferred for quantitative bioanalysis [22]. It is assumed that the light intensity is proportional to the concentrations of the targeted molecules. [22]

In bio analysis, photon emission chemistries including fluorescence and bioluminescence are popular due to their inherently high sensitivity and simplicity. [22] Fluorescence relies on photons as the energy source whereas in bioluminescence, the photon emission is based on natural biochemistry.
Fluorescence is brighter due to the high rates of photon excitation. However, this high influx of photon raises the background levels. The number of photons introduced is much more important than the number produced by the reporter gene and photodetectors could not be able to discriminate between excitation and emission photons. Others fluorophores can be present within the samples. [22] As no photons are introduced into the sample, background levels in bioluminescence assay are much lower. [22][21]

As a results, bioluminescent chemistries present 10- to 1,000-fold higher sensitivity than fluorescence assays [23] Bioluminescence can be measured to zeptomole levels (few molecules per cell) and linearity extended over six to eight logs. [22]

Moreover, as bioluminescence naturally evolved within a biological context, it is more compatible with bioluminescence systems. [22]

To conclude, bioluminescence is greatly used in assay methodologies and these assays are known for their quantitative precision, low inherent backgrounds, and low sample interference. [22] For all these reasons, we decided to base our biosensor on the use of bioluminescence as the reporter gene.

References :

NB: Links are provided when available

1. Bilan de la qualité de l’air en France en 2014 et principales tendances observées sur la période 2000-2014, Laurence Monnoyer-Smith, Anne Bottin, Commissariat Général au développement durable, September 2015

2. AIRPARIF, association for air quality monitoring in Ile de France (France)

3. 7 million premature deaths annually linked to air pollution, WHO , 2015

4. Air pollution costs European economies US$ 1.6 trillion a year in diseases and deaths, new WHO study says, WHO

5. « Bilan de la qualité de l’air en France en 2012 et principales tendances observées sur la période 2000-2012 », Energy and climate executive management

6. Le bulletin de la mesure et de la caractérisation des polluants dans les rejets atmosphériques : Mesure des Composés Organiques Volatils », Jean Poulleau, INERIS

7. Active sampling measurement (norme NF EN 14662-1, november 2005)

8. Active sampling measurement (norme NF EN14662-4, november 2005)

9. RAPPORT D’ÉTUDE 05/10/2010 N° INERIS-DRC-10-112289-10754A: Stratégie de mesure des niveaux de concentration en benzène, toluène, éthylbenzène et xylènes dans l’air ambiant autour d’installations classées

10. Jarque, S., Bittner, M., Blaha, L. & Hilscherova, K. Yeast Biosensors for Detection of Environmental Pollutants: Current State and Limitations. Trends in Biotechnology (2016).

11. 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.

12. Rawson, D.M., Willmer, A.J., and Turner, A.P. (1989). Whole-cell biosensors for environmental monitoring. Biosensors 4, 299–311.

13. iGEM Peking 2013 Wiki

14. iGEM Cambridge 2009

15. 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.

16. X. Liu, K. J. Germaine,D. Ryan, and D.N.Dowling, “Whole-cell fluorescent biosensors for bioavailability and biodegradation of polychlorinated biphenyls,” Sensors, vol. 10, no. 2, pp. 1377–1398, 2010.

17. 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).

18. Luc BOUSSE, « Whole cell biosensors », sensors and actuators, 1996

19. 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.

20. Nunes-Halldorson, V. da S., and Duran, N.L. (2003). Bioluminescent bacteria: lux genes as environmental biosensors

21. Promega corporation. Welsh, D.K., and Noguchi, T. (2012). Cellular Bioluminescence Imaging. Cold Spring Harbor Protocols 2012, pdb.top070607

22. Wood, K (2011), The bioluminescence advantage, laboratorynews

23. Fan, F. and Wood, K. (2007) Bioluminescent Assays for High-Throughput Screening. Assay Drug Dev. Technol. 5, 127–36.