Periodic Reporting for period 2 - CYTO-WATER (Integrated and portable image cytometer for rapid response to Legionella and Escherichia coli in industrial and environmental waters) Periodo di rendicontazione: 2016-12-01 al 2018-05-31 Sintesi del contesto e degli obiettivi generali del progetto The CYTO-WATER project (from June 2015 to May 2018) was coordinated by Labaqua in partnership with CETaqua, ICFO, Bertin Technologies, microTEC and Memteq Ventures LTD. The objective of the project was to deploy, for the first time in industrial and environmental waters, a new imaging cytometer platform for the detection and quantification of microorganisms. The analytical process takes no more than 120 min, and involves a sample concentration step, cells labelling with specific antibodies and detection with an image cytometer, 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. The platform was validated for the detection of Legionella and E. coli but it can be easily adapted to detect other microorganisms. This innovative tool is portable and, with a wide potential market in industries, environmental agencies and private facilities. Lavoro eseguito dall’inizio del progetto fino alla fine del periodo coperto dalla relazione e principali risultati finora ottenuti The CYTO-WATER platform integrates an automatic water concentration system that includes Celltrap® concentrating filters, a microfluidic system for automatic sample preparation and a fluorescence image cytometer.Before the integration, the different modules have been adapted and validated individually in the laboratory to make sure that they meet the market specifications and comparing with conventional analysis and alternative methods, to determine whether the methods are suitable for detecting and quantifying microorganisms in water samples.Regarding the concentrator module, the objective was to adapt the existing automated liquid concentration system to the needs of the CYTO-WATER platform. The bacteria contained in the water volume to be analyzed (100 mL for E. coli and 1 L for Legionella) are concentrated in a volume from 1000 to 1500μL with a recovery rate of around 50%.For sample preparation, a microfluidic module was designed and validated. The microfluidic cartridge and tubing material was evaluated basing on the study of the optical properties, mechanical properties, hydrophilic/hydrophobic properties, and biocompatibility.The microfluidic module was evaluated comparing samples labelled with the microfluidic system and samples labelled manually. Finally, a semi-automated as well as a fully automated, one channel microfluidic labelling systems were designed and manufactured.The reader was adapted to the marker requirements carrying out several adjustments and improvements of hardware and software in the detection module. These adjustments were required to achieve the required performance in terms of limit of detection (LOD), sensitivity, repeatability, reproducibility and robustness, for the validation process.The detector was validated with spiked samples to verify the main parameters of the system. The reader was also validated with natural samples, as cooling water and seawater samples. The obtained parameters in the validation tests indicated good correlations (R2>0.9) with traditional methods (culture isolation) and alternative methods, as flow cytometry (FCM), and its suitability for detection and quantification of Legionella and E. coli in natural samples.The reader, the water concentration module and the microfluidic device (hardware and software) were integrated in an outdoor rack cabinet resulting in the CYTO-WATER platform.The operational functioning and performance of the CYTO-WATER platform in real conditions was evaluated. A detailed laboratory and on-site validations in different facilities was performed. The validation was performed comparing results with results obtained by other methodologies (culture isolation as the standard method, and FCM and qPCR as alternative methods).The three modules as separate devices are also very interest in terms of market opportunities. The main advantage of this approach is that each of the modules can be adapted to different technologies and can target other markets. Hence, this approach is more flexible and with a wider market in some cases. One clear example is the automatic labelling module that can be adapted to microorganisms detection with other techniques (different form the CYTO-WATER reader) and then adapted to the workflow in laboratories of different sectors, hospitals, universities, etc.An environmental and economic comparison between the traditional spot sampling method and the CYTO-WATER system was assessed through to a Life Cycle Assessment (LCA) and Life Cycle Costing (LCC). This comparison was undertaken for the detection of Legionella in cooling towers and hot water distribution systems from hospitals and E. coli in bathing waters. Finally, results were summarized and evaluated in the validation document.Exploitation and dissemination activities: Project brochure; 4 electronic newsletters; Roll-up; website; project video; Social media (Twitter); participation in conferences and workshops; CYTO-WATER Final Workshop was held in Barcelona; and meetings and interviews with different end-users Progressi oltre lo stato dell’arte e potenziale impatto previsto (incluso l’impatto socioeconomico e le implicazioni sociali più ampie del progetto fino ad ora) CYTO-WATER will offer the possibility of rapid detection of possible biological threats, minimizing the risk for the clients. Infectious diseases caused by microorganisms are the most common and widespread health risk associated with water, drinking and bathing. A quick detection of those microorganisms and reducing the risk of waterborne diseases minimizes the impact on the society.CYTO-WATER will also allow the reduction of cost related to water handling and as such will have a positive impact on the European industry. The developed device will be able to reduce the economic loss associated with the pathogen contamination, increasing the competitiveness of the industry. In case of outbreak, its location will be easily detected thus reducing sanitary risk and associated cost. The quick and real-time detection of pathogens and indicators of contamination into industry with water distribution systems will allow reducing the contaminated production and therefore the activity losses derived from a waterborne outbreak resulting in, for example, production interruption.The use of the CYTO-WATER platform will lead to a quicker detection of E. coli and Legionella, reducing exposure and thus, human health damages, and economic and social impacts. The proposed system will entail a higher level of protection of the natural environment (i.e. bathing waters), natural resources (through industrial effluents) and well-being. Moreover, the rapid response and access to results will benefit in terms of real-time access to information.Additionally, the project will contribute to minimize the risk of exposure to the pathogen for people, reducing the impact on the society and on the economic activities involved, as in the case of hot and cold water systems of public buildings and facilities. An analysis with LCA and LCC have ensure that the system also has other environmental benefits besides the sanitary ones exposed above and cost saving potential compared to the conventional analysis.Regarding the stimulation of sustainable economic growth, business and job creation in the water sector, CYTO-WATER will provide a rapid, low cost instrument that will be of interest of laboratories, public administrations and industries worldwide. This fact will clearly rebound to the creation of new job opportunities and, at the same time, will position Europe as the developer of an innovative product.
