|TBTRESENSE: Tributiyltin (TBT) bioremediation and development of a biosensor to detect TBT from contaminated sites.|
Programme - Fundação para a Ciência e a Tecnologia- FEDER- COMPETE PTDC/MAR/108024/2008
Execution dates - 2010-04-01 - 2013-04-01 (36 Months)
Funding Entity - FCT
Funding for CESAM - 171830
Total Funding - 176000
Proponent Institution - Universidade de Aveiro
Tributyltin (TBT) is considered as one of the most toxic xenobiotics ever produced and deliberately introduced into the environment by Man (10). It is used in a variety of industrial processes and subsequently discharged into the environment. Once released, TBT disperses through the water and rapidly adsorb to biota and to suspended particles that latter deposit onto sediments. Its typical half life in marine waters is of several weeks, but in deeper anoxic sediments it may be of several years due to slower degradation (9). TBT has been reported to be toxic to eukaryote and prokaryotes. One of the most important biological deleterious effects of TBT is the imposex - the superimposition of male characters onto females of gastropods (1). TBT have also been identified as immune system inhibitor and endocrine disruptor in humans (7). Production, use and export of TBT was prohibited in developed countries since the 1990s by the IMO; but some countries still continue to use it (25). Consequently, TBT pollution did not decreased and has been recognized as a serious problem worldwide. Measurement of such compounds constitutes a major tool for regulatory agencies and for control and monitoring of processes in industrial areas. Traditional analytical methods are expensive, time consuming and need expertise technical personnel constituting an impediment for their application on a regular basis. Biosensors are a suitable alternative, due the possibility of identifying and quantifying specific compounds directly in air, water or sediment; their specificity of response and, in some cases, their ability to work in very dirty environments (24). However, there is still a lack of systems suitable for determination of emerging pollutants. An effort by European collaborative projects supported by the EU to produce new biosensors for endocrine disrupting compounds, such as TBT, is being made (24). However, until now, TBT biosensors developed fail to work in contaminated environments, and therefore the development of new systems is desirable. Bioremediation of a variety of contaminants has become a broadly accepted remediation technology; it often can be applied in a cost effective manner and employs naturally-occurring bacteria (12). TBT resistant bacteria have been reported (16, 18) and they constitute the basis to develop detection/remediation systems. One such bacterium (A. molluscorum isolated by our team) that is highly resistant to TBT (up to 3 mM), being the first bacteria reported to tolerate and at the same time able to degraded it to less toxic compounds (DBT and MBT) (4). A gene has also been isolated and characterized (sug-E like) that is over-expressed when cells grow in the presence of TBT. The versatility of this strain to grow in rich or poor substrates and use TBT as carbon source makes it a logical candidate to develop a biosensor and ultimately, its improvement to be employed to remediate TBT contaminated areas. The two main aims of the present proposal are the following:
To achieve these goals we will use different approaches. Firstly a pyrosequencing transcriptome analysis will reveal which other genes (apart from the one already identified) are expressed in response to TBT and involved in TBT degradation. This approach will shed more light about the mechanisms behind these two processes. Real Time-PCR analysis will complement the study of the expression of target gene(s). A whole cell-based biosensor will be constructed and its specificity, sensitivity, time to detection, repeatability and reproducibility will be evaluated. Microcosm experiments will be assembled to evaluate the biological remediation by A. molluscorum in association with the indigenous bacterial community. This will be accessed by a PCR-DGGE based approach combined with organotin analysis. Changes in bacterial community structure will be monitored and will reveal the most active strains in the remediation process. Simultaneously toxicity of the remediated water/sediments will also be evaluated over a number of indicator species. The team involved in the present proposal is able to generate all the synergisms required to carry out the project with great success, and is committed to achieve all the objectives underlined given the solid scientific knowledge and experimental investigation in areas such as: molecular biology, gene/protein expression, study of bacterial communities and bioremediation. On the other hand, some team members have preliminary results that will certainly contribute to the success of the present proposal. The proposed project will contribute substantially to the scientific knowledge and will give new insights about the mechanisms involved in the resistance/degradation of TBT. The biosensor developed will surely have immediate practical applications.
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