Beyenal Haluk, Donovan Conrad, Lewandowski Zbigniew, Harkin Gary
Center for Biofilm Engineering, P.O. Box 173980, Room 366 EPS, Montana State University, Bozeman, MT 59717-3980, USA.
J Microbiol Methods. 2004 Dec;59(3):395-413. doi: 10.1016/j.mimet.2004.08.003.
Quantitative parameters describing biofilm physical structure have been extracted from three-dimensional confocal laser scanning microscopy images and used to compare biofilm structures, monitor biofilm development, and quantify environmental factors affecting biofilm structure. Researchers have previously used biovolume, volume to surface ratio, roughness coefficient, and mean and maximum thicknesses to compare biofilm structures. The selection of these parameters is dependent on the availability of software to perform calculations. We believe it is necessary to develop more comprehensive parameters to describe heterogeneous biofilm morphology in three dimensions. This research presents parameters describing three-dimensional biofilm heterogeneity, size, and morphology of biomass calculated from confocal laser scanning microscopy images. This study extends previous work which extracted quantitative parameters regarding morphological features from two-dimensional biofilm images to three-dimensional biofilm images. We describe two types of parameters: (1) textural parameters showing microscale heterogeneity of biofilms and (2) volumetric parameters describing size and morphology of biomass. The three-dimensional features presented are average (ADD) and maximum diffusion distances (MDD), fractal dimension, average run lengths (in X, Y and Z directions), aspect ratio, textural entropy, energy and homogeneity. We discuss the meaning of each parameter and present the calculations in detail. The developed algorithms, including automatic thresholding, are implemented in software as MATLAB programs which will be available at site prior to publication of the paper.
描述生物膜物理结构的定量参数已从三维共聚焦激光扫描显微镜图像中提取出来，并用于比较生物膜结构、监测生物膜发育以及量化影响生物膜结构的环境因素。研究人员此前曾使用生物体积、体积与表面积比、粗糙度系数以及平均和最大厚度来比较生物膜结构。这些参数的选择取决于进行计算的软件的可用性。我们认为有必要开发更全面的参数来描述三维异质生物膜形态。本研究提出了从共聚焦激光扫描显微镜图像计算得出的描述三维生物膜异质性、大小和生物量形态的参数。本研究扩展了先前的工作，即从二维生物膜图像中提取关于形态特征的定量参数到三维生物膜图像。我们描述了两种类型的参数：(1) 显示生物膜微观异质性的纹理参数，以及 (2) 描述生物量大小和形态的体积参数。所呈现的三维特征包括平均（ADD）和最大扩散距离（MDD）、分形维数、平均游程长度（在X、Y和Z方向）、纵横比、纹理熵、能量和均匀性。我们讨论了每个参数的含义并详细介绍了计算方法。所开发的算法，包括自动阈值处理，在软件中作为MATLAB程序实现，将在论文发表前在网站上提供。
