Underwater light sheet hyperspectral microscopy and its applications in unicellular microalgae in-situ identification and counting

Abstract

Microalgae are a significant component of aquatic organisms and are instrumental in regulating aquatic ecosystems. But they are not always beneficial for the environment or other organisms, they can be toxic to aquatic life and humans when they produce harmful algal blooms (HABs) that release toxins and deplete oxygen in the water. A more accurate and quantitative understanding of aquatic environments can be obtained by detecting and counting microalgae. To overcome the challenges of underwater microscopic imaging, a platform integrating light sheet microscopy and a hyperspectral camera is developed to noninvasively spatially delineate and spectroscopically measure microalgae in situ at nearly unicellular resolution. It achieves a theoretical lateral resolution of 0.88 µm, providing higher signal-to-background ratio than conventional wide-field microscopy. To evaluate the performance of our instrument, individual fluorescence spectra of 15 species of microalgae cells and their paired mixtures are captured as datasets and analyzed using deep-learning algorithms for classification. The average accuracies for individual microalgae and paired mixtures are 95.67% and 78.53%, respectively. In addition, the system is applied with a single type of microalgae at different concentrations to verify the feasibility of microalgae counting. The experimental results confirm that the system allows optical measurement and can be used to assess the water quality for forecasting aquatic disasters.