Appl Opt. 2022 Feb 10;61(5):B331-B338. doi: 10.1364/AO.444976.
A study on locomotion in a 3D environment of Tetraselmis microalgae by digital holographic microscopy is reported. In particular, a fast and semiautomatic criterion is revealed for tracking and analyzing the swimming path of a microalga (i.e., Tetraselmis species) in a 3D volume. Digital holography (DH) in a microscope off-axis configuration is exploited as a useful method to enable fast autofocusing and recognition of objects in the field of view, thus coupling DH with appropriate numerical algorithms. Through the proposed method we measure, simultaneously, the tri-dimensional paths followed by the flagellate microorganism and the full set of the kinematic parameters that describe the swimming behavior of the analyzed microorganisms by means of a polynomial fitting and segmentation. Furthermore, the method is capable to furnish the accurate morphology of the microorganisms at any instant of time along its 3D trajectory. This work launches a promising trend having as the main objective the combined use of DH and motility microorganism analysis as a label-free and non-invasive environmental monitoring tool, employable also for in situ measurements. Finally, we show that the locomotion can be visualized intriguingly by different modalities to furnish marine biologists with a clear 3D representation of all the parameters of the kinematic set in order to better understand the behavior of the microorganism under investigation.
本文报道了利用数字全息显微镜研究微藻（即三角褐指藻）在 3D 环境中的运动。特别是，提出了一种快速半自动的标准，用于跟踪和分析微藻（即三角褐指藻）在 3D 体积中的游动路径。离轴配置的数字全息术（DH）被用作一种有用的方法，能够在视场中快速自动对焦和识别物体，从而将 DH 与适当的数值算法相结合。通过所提出的方法，我们可以同时测量鞭毛微生物的三维路径和描述所分析微生物的游动行为的整套运动学参数，方法是通过多项式拟合和分割。此外，该方法能够在其 3D 轨迹的任何时刻提供微生物的精确形态。这项工作开创了一种很有前景的趋势，其主要目标是将 DH 与运动微生物分析相结合，作为一种无标记和非侵入性的环境监测工具，也可用于原位测量。最后，我们展示了可以通过不同的方式来生动地可视化运动，为海洋生物学家提供运动学参数集的所有参数的清晰 3D 表示，以便更好地理解所研究的微生物的行为。
