Departamento de Física Aplicada, Instituto de Física Gleb Wataghin, Universidade Estadual de Campinas, Campinas, São Paulo, Brasil.
Departamento de Física Aplicada, Instituto de Física Gleb Wataghin, Universidade Estadual de Campinas, Campinas, São Paulo, Brasil.
Biophys J. 2021 Apr 20;120(8):1443-1453. doi: 10.1016/j.bpj.2021.02.009. Epub 2021 Feb 17.
Microchannels can be used to simulate xylem vessels and investigate phytopathogen colonization under controlled conditions. In this work, we explore surface functionalization strategies for polydimethylsiloxane and glass microchannels to study microenvironment colonization by Xylella fastidiosa subsp. pauca cells. We closely monitored cell initial adhesion, growth, and motility inside microfluidic channels as a function of chemical environments that mimic those found in xylem vessels. Carboxymethylcellulose (CMC), a synthetic cellulose, and an adhesin that is overexpressed during early stages of X. fastidiosa biofilm formation, XadA1 protein, were immobilized on the device's internal surfaces. This latter protocol increased bacterial density as compared with CMC. We quantitatively evaluated the different X. fastidiosa attachment affinities to each type of microchannel surface using a mathematical model and experimental observations acquired under constant flow of culture medium. We thus estimate that bacterial cells present ∼4 and 82% better adhesion rates in CMC- and XadA1-functionalized channels, respectively. Furthermore, variable flow experiments show that bacterial adhesion forces against shear stresses approximately doubled in value for the XadA1-functionalized microchannel as compared with the polydimethylsiloxane and glass pristine channels. These results show the viability of functionalized microchannels to mimic xylem vessels and corroborate the important role of chemical environments, and particularly XadA1 adhesin, for early stages of X. fastidiosa biofilm formation, as well as adhesivity modulation along the pathogen life cycle.
微通道可用于模拟木质部导管，并在受控条件下研究植物病原体的定殖。在这项工作中，我们探索了聚二甲基硅氧烷和玻璃微通道的表面功能化策略，以研究木质部导管中 Xylella fastidiosa subsp. pauca 细胞的微环境定殖。我们密切监测了细胞在微流控通道内的初始粘附、生长和迁移，作为模拟木质部导管中发现的化学环境的函数。羧甲基纤维素 (CMC) 是一种合成纤维素，也是 X. fastidiosa 生物膜形成早期过度表达的一种黏附素，被固定在设备的内表面上。与 CMC 相比，后一种方案增加了细菌密度。我们使用数学模型和在培养基恒流条件下获得的实验观察结果，定量评估了不同类型的微通道表面对 X. fastidiosa 的不同附着亲和力。因此，我们估计细菌细胞在 CMC 和 XadA1 功能化通道中的附着率分别提高了约 4%和 82%。此外，变流量实验表明，与聚二甲基硅氧烷和玻璃原始通道相比，XadA1 功能化微通道中的细菌粘附力对剪切应力的数值大约增加了一倍。这些结果表明，功能化微通道具有模拟木质部导管的可行性，并证实了化学环境，特别是 XadA1 黏附素，在 X. fastidiosa 生物膜形成的早期阶段以及病原体生命周期中黏附性的调节方面的重要作用。
