Table_1_Matrix glycoconjugate characterization in multispecies biofilms and bioaggregates from the environment by means of fluorescently-labeled lectins.docx

Environmental biofilms represent a complex mixture of different microorganisms. Their identity is usually analyzed by means of nucleic acid-based techniques. However, these biofilms are also composed of a highly complex extracellular matrix produced by the microbes within a particular biofilm system. The biochemical identity of this extracellular matrix remains in many cases an intractable part of biofilms and bioaggregates. Consequently, there is a need for an approach that will give access to the fully hydrated structure of the extracellular matrix or at least a major part of it. A crucial compound of the matrix identified as carbohydrate-based polymers represents major structural and functional constituents. These glycoconjugates can be characterized by using fluorescently-labeled lectins in combination with confocal laser scanning microscopy. The lectin approach is defined previously, as fluorescence lectin barcoding (FLBC) and fluorescence lectin-binding analysis (FLBA), where FLBC is equal to the screening of a particular sample with all the commercially available lectins and FLBA is the actual analysis of the matrix throughout an experiment with a selected panel of lectins. As the application of immune-based techniques in environmental biofilm systems is impossible, the lectin approach is currently the only option for probing lectin-specific glycoconjugates in complex biofilms and bioaggregates. From all the commercially available lectins tested, the lectins such as AAL, HAA, WGA, ConA, IAA, HPA, and LEA showed the highest binding efficiency. Furthermore, 20 of the overall lectins tested showed an intermediate signal intensity, nevertheless very useful for the assessment of matrix glycoconjugates. With the data compiled, we shall virtually shed more light on the dark matter of the extracellular matrix and their 3-dimensional distribution in environmental biofilm systems. The results will be helpful in future studies with a focus on the extracellular matrix glycoconjugates present in environmental microbial communities.

环境生物膜代表了一种由不同微生物构成的复杂混合体。其身份通常通过基于核酸的技术进行分析。然而，这些生物膜亦由特定生物膜系统中微生物产生的极其复杂的细胞外基质组成。该细胞外基质的生化身份在许多情况下成为生物膜和生物聚集体的难以解决的组成部分。因此，迫切需要一种能够获取细胞外基质完全水合结构或至少其主要部分结构的途径。基质中鉴定出的以碳水化合物为基础的多聚物是主要的结构和功能成分。这些糖基化合物的特征可以通过结合荧光标记的凝集素与共聚焦激光扫描显微镜进行表征。凝集素方法已被定义为荧光凝集素条形码（FLBC）和荧光凝集素结合分析（FLBA），其中FLBC等同于对所有商业可用凝集素的特定样本的筛选，而FLBA则是通过选择一组凝集素在实验过程中对基质进行实际分析。由于在环境生物膜系统中应用基于免疫的技术是不可能的，凝集素方法目前是探究复杂生物膜和生物聚集体内凝集素特异性糖基化合物的唯一选择。在测试的所有商业凝集素中，AAL、HAA、WGA、ConA、IAA、HPA和LEA等凝集素显示出最高的结合效率。此外，20种总体测试的凝集素显示出中等信号强度，尽管如此，对于评估基质糖基化合物仍极为有用。凭借所收集的数据，我们将在环境生物膜系统中细胞外基质的暗物质及其三维分布方面提供更多的洞察。这些结果将有助于未来针对环境微生物群中存在的细胞外基质糖基化合物的相关研究。