Underwater microorganisms observation with off-axis digital holography microscopy using partially coherent illumination

Nowadays, the plankton net is replaced by underwater imaging systems. With the advent of digital technologies, detection and classification of plankton and subcentimetre-scale pelagic organisms may be largely accelerated even if still requiring human expertise at some processing stages. Moreover, the non intrusive aspect of in situ imaging systems are preferable for preserving the organisms in their natural habitat to avoid damaging those most fragile. This paper presents experimental results of a new digital holography microscope (DHM) prototype, Holoflow@Sea, based on an off-axis configuration. Using a partially coherent laser illumination, the DHM has been developed for in situ observation with and without microscope objectives. The holograms can be acquired with a frame rate up to 15 images per second on a CCD array of 2456×2058 pixels. Reconstruction algorithms enable then refocusing and tracking of objects in the probed volume with a resolution up to a few micrometers when working with the highest magnification setup. The use of partially coherent light source is shown to efficiently reduce the speckle noise that comes with fully coherent illumination. Although an off-axis configuration results in a lower resolution than that of an equivalent in-line configuration, the approach is shown to be relevant for in situ underwater applications. Based on the results, this paper discusses the prototype performance in different natural waters.