Improving Biosecurity in Aquaculture using High Speed, Low cost, Lab on a Chip Micro-Cytometry for the Surveillance of Harmful Algal Blooms.

This project will develop a new seawater analysis tool to improve the biosecurity of shellfisheries, which are vulnerable to contamination with algal bio-toxins (poisons) that are associated with harmful algal blooms. It will do this by re-engineering an existing Lab on Chip (LOC) micro-cytometer that counts and characterises single cells as they flow in single file in a channel less than 0.1mm wide. It can count and discriminate specific algal cell types, including bio-toxin producing species native to the UK, directly in seawater samples. This technology represents a low cost, miniaturised alternative to existing cytometers with enhanced functionality and capability. It has the potential to significantly improve species resolution by generating synchronous fluorescence and electrical impedance measurements, the latter being absent from current systems. The generation of unique cytometric profiles for target species based on both parameters increases the accuracy with which they can be discriminated within complex samples containing non-target species with similar morphological characteristics. Other advantages of the system include the absence of a sheath flow (clean water used in standard cytometers to control cell position in the channel), which improves throughput, eliminates the need to carry or generate clean water and facilitates, where necessary, the periodic unblocking of the flow channel (by flow inversion). The project objectives were conceived with, and refined by one of our "end-users" and a project sub-contractor, the Centre for Environment, Fisheries and Aquaculture Sciences (Cefas), with whom we have a strong, long-standing collaborative relationship. Cefas carry out statutory harmful algal bloom / algal bio-toxin surveillance in line with current European Union regulations regarding the quality of shellfish produced for human consumption. This entails periodic monitoring of water samples for harmful algal species, and where necessary acts as an early warning system that stimulates the increased sampling and analysis of shellfish flesh for algal bio-toxins. Currently, water samples delivered to a centralised laboratory are analysed by microscopy. This process is time consuming and delays intervention, leading to an increased risk to human health and potential cost to the industry from loss of shellfish product. Accordingly, rapid and near-real time analysis systems are required to aid / complement microscopic analysis. To address this, the project will deliver a portable, bench-top LOC micro-cytometer for the rapid counting and discrimination of specific algal cell types in seawater samples. The system will be capable of being operated by non-specialist personnel using automated data analysis software and will be used by our project partners, Offshore Shellfish Ltd and the Environment Agency, as an in situ early warning tool, and our end-user and sub-contractor, Cefas, to complement existing water sample analysis (using microscopy), with a view to reducing analysis time. The project outputs will also complement existing algal surveillance programmes using satellite imaging by providing in situ data, uninterrupted by cloud cover. It will deliver impact by facilitating timely intervention, directly leading to economic gain and reduction in public health risks. The outputs from the project will underpin the future development of a robust, deployable system that can be operated in situ including offshore. This will provide near real-time analysis of shellfisheries waters, obviating the need to collect samples from high risk locations.

Keywords: Harmful Algal Bloom (HAB); micro-cytometer; Lab on a Chip (LOC); Micro-systems; Biosecurity; Shellfisheries; Aquaculture

In 2012, the value of shellfish landed by UK vessels constituted approximately £300M wholesale (~£1.2 B retail), whilst cultivated stocks contributed an additional ~£38 M wholesale; around 70 % of UK shellfish are exported (http://www.seafish.org/research-economics/market-insight/market-summary). Shellfish bed closures and loss of shellfish harvests due to contamination with algal bio-toxins during Harmful Algal Blooms (HABs) can lead to significant economic costs to the industry and producers. Accordingly, a key benefit from this project is the provision of a new tool, based on existing technology, that will reduce these costs by compressing the current delay between the detection of HAB species in the environment and the implementation of the appropriate intervention strategies.

Statutory shellfish monitoring authorities will benefit from the provision of re-engineered, state of the art technology that opens up new possibilities in HAB metrology. Specifically, the micro-cytometer that this project will develop will address some of the shortcomings of existing methods (microscopy and satellite imaging) by generating high resolution, in situ data unobstructed by cloud cover (better monitoring and forecasting) and a rapid analysis tool that can be used in the laboratory to streamline the analysis of water samples by informing on high risk / priority samples requiring immediate / specialist attention. The general public will benefit from improved biosecurity in shellfisheries leading to a reduction in public health risk. Over time, a reduction in the incidence of shellfish poisoning events following better forecasting and shorter interventions will benefit the industry by increasing confidence, both domestically and internationally, in UK-produced shellfish. The provision of an alternative HAB analysis tool to be used in tandem with existing methods will increase the accuracy and reliability, and ultimately the efficiency, of HAB surveillance for the UK, and is timely as the UK aquaculture industry expands including further offshore into more remote, exposed and hazardous environments. This inevitably necessitates the more stringent HAB monitoring that this project will deliver. This impact will be achieved primarily through the Centre for Environment, Fisheries & Aquaculture Science (Cefas) who will benefit from direct access to the new technology, and can rapidly and efficiently implement the project outputs for HAB suveillence.

Researchers focusing on HABs, and more broadly those focused on general phytoplankton biology and diversity, will benefit from technology that is optimised to perform state of the art phytoplankton analysis in the field, obviating the need to routinely fix samples prior to delivery to the laboratory and removing "bottling effects". The associated improvements in the understanding of HABs / phytoplankton temporal and spatial variation, function and diversity will have wider benefits to the UK and international research community. This project will develop existing micro-systems technology with clear benefit to the instrumentation industry focusing on environmental monitoring and sensing technologies.

The prototype product delivered by this project will address some limitations with commercially available cytometers (bulky, prohibitively expensive, no impedance analysis, not ruggedized, require expert users), and will create additional impact from (1) reduced cost making the technology widely available / affordable and (2) enhanced functionality for end users. We will work to commercialise the technology in the medium term with support from our innovations and enterprise teams using experience from previous patenting and commercialisation successes. The flexibility in application of the technology the measurement and discrimination of a range of cell types in complex matrices, including those with high turbidity, will generate additional academic and commercial potential.