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Integrated and portable image cytometer for rapid response to Legionella and Escherichia coli in industrial and environmental waters

Periodic Reporting for period 1 - CYTO-WATER (Integrated and portable image cytometer for rapid response to Legionella and Escherichia coli in industrial and environmental waters)

Reporting period: 2015-06-01 to 2016-11-30

The project was launched in June 2015 and is expected to be completed in May 2018. The project is coordinated by Labaqua in partnership with CETaqua, ICFO, Bertin Technologies, microTEC and Memteq Ventures LTD. The objective of the project is to deploy, for the first time in industrial and environmental waters, a new imaging cytometer platform for the detection and quantification of microorganisms. This will allow quantify Legionella and E. coli population within 120 minutes from obtaining the sample, overcoming in this way the main disadvantage of traditional methods used in laboratories, i.e. long time-to results which can currently last up to 12 days in the case of Legionella and 1 day for E. coli. This tool will be an easy-to-handle portable form, which will increase its versatility and widen the possibilities of onsite applications.
The work performed and main results achieved per work package were:
WP1 Project Management: During this first half of the project, two internal 2 follow-up reports were provided: M1 to M7 & M8 to M13 about technical and financial information of the project. In addition this first official periodic reporting has been done with information from M0 to M18 (01/06/2015 to 30/11/2016). This report includes an overview of the activities carried out, a description of project progress and an identification of problems encountered and corrective actions taken
WP2 Adaptation of Cytometry Reader to Market-Specific Requirements: ICFO’s reader was upgraded and adapted for market compliance; by carrying out proper adjustments and improving the optical system and software platform. Four beta prototypes of the optical reader (CYT-Beta) were assembled. Off calibration of the readers was performed using sample series with different microorganism concentrations provided by Labaqua. A benchmarking at Labaqua’s facilities was carried out. More validation data should be obtained in order to adjust the system to the necessities of the market. Additional comparison with traditional method will be performed in the coming month to finish the adjustment of the device.
WP3 Microfluidic System Set-up for Water Sampling: The objectives are to select and validate materials for microfluidics, to develop and test microfluidics solutions, to compose and manufacture the microfluidic set-up for the sample treatment, and to design and construct the interface to, e.g. concentrator for integration into the platform, by the moment the work performed has been: the materials have successfully been validated; the specific properties of the concentrated liquid to be analysed as well as the volumes to be processed inside the cartridge made it impossible to use capillary forces for liquid transport, so active pumping is now required; a new cartridge respecting this new situation has been designed and produced by microTEC; for the supply of the cartridge with the sample, reagents, buffer etc. a concept has been developed based on various pumps, valves, controllers etc.
WP4 Automated Water Concentration Module Adaptation: The objective is to adapt the automated liquid concentration module to the needs of microfluidic module. The bacteria contained in the water volume to be analysed (100 mL for E. coli and 1 L for Legionella) will be concentrated in a volume from 100 μL to 600 μL with a recovery rate as high as possible, and in an appropriate buffer in order to facilitate the analysis. Some major orientations have led to important evolutions brought to the initial concept. They are: the decision to perform up to 4 operations without human intervention; the decision to use a decontamination protocol in order to simplify the concept and hence to reduce the manufacturing cost; the need of a sedimentation step before the labelling step in order to minimize the clogging risks and to preserve the microfluidics unit of sample preparation.
WP5 Integration of the whole CYTO-WATER Sensor System: The main objective is to achieve the integration of the reader, the concentrator and the microfluidic cartridge to form the new CYTO-WATER platform. During this period, different tasks have been performed: the Bertin software integration has been agreed upon. In relation to microfluidics module, mechanical issues were discussed in several meetings in order to fit the microfluidic cartridges holder with the moving part of the reader. The CYT-Beta reader has been coupled to a stepper motor so that it can automatically reach each cartridge to measure. This WP is in the preliminary stages: the concentrator is ready to be integrated and the main structure of the complete system is ready too.
WP6 Validation and First Application in Real Environment: No specific activities have been performed yet in this WP
WP7 Project Dissemination and Market Communication: In the early stages, Project Diss
Objectives: The overall objective of the proposed project is to deploy, for the first time in industrial and environmental waters, a new imaging cytometer platform for the detection of microorganisms. The platform will result from the integration of proprietary technologies already available to the consortium partners: an automatic water concentration cartridge combined with a microfluidic cell will provide an adequate sample to a newly designed fluorescence image cytometer whose readings will be recorded and processed by a Global Control System.
CYTO-WATER will be validated for quantifying Legionella and Escherichia coli population within 2 hours from obtaining the sample, overcoming in this way the main disadvantage of traditional methods.
The main activities to overcome the objective will be: The main activities to overcome the objective will be: Validation of the existing fluorescence microscope reader for industrial use. The newly designed reader will be validated for high-throughput, rapid and sensitive detection and quantification of microorganisms.
Adaptation of the reader to the automatic concentration system already validated in the lab for in-situ concentration. The concentration system that integrates Celltrap® filters will be coupled to the custom-made microfluidics cartridge in order to provide an adequate entry of the final elution volume from the concentrator to the detection device.
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