Sadanandan Bindu, Vaniyamparambath Vijayalakshmi, Lokesh K N, Shetty Kalidas, Joglekar Amruta P, Ashrit Priya, Hemanth Beena
Department of Biotechnology, M S Ramaiah Institute of Technology, Bangalore, Karnataka, India.
Department of Plant Science, North Dakota State University, Fargo, North Dakota, USA.
J Appl Microbiol. 2022 Apr;132(4):3277-3292. doi: 10.1111/jam.15402. Epub 2022 Jan 25.
Optimization of Candida albicans growth and biofilm formation is essential for understanding the recalcitrance of this pathogen to advance functional analysis on hospital tools and material surfaces. Optimization and quantification of biofilm have always been a challenge using the conventional one variable at a time (OVAT) method. The present study uses central composite design-based response surface methodology for optimization of conditions to induce growth and biofilm formation in Candida albicans on polystyrene microtiter plates.
Statistical software package, Stat Soft®, STASTICA version 12.6 was used for data analysis. The variables considered in the design matrix were media pH, temperature, incubation period, shaker speed and inoculum size. A four-pronged quantification approach with XTT assay (cell viability), crystal violet assay (biofilm), calcofluor white assay and wet/dry weight measurements (cell mass) was used to understand different aspects of biofilm formation. Cell viability and cell mass were inversely related; however, biofilm was independent of these two factors. The study also highlighted the fact that foetal bovine serum does not significantly contribute to cell adhesion and in turn in vitro biofilm formation in some of the cultures.
A high-throughput optimization of C. albicans growth and biofilm formation on polystyrene microplate has been developed and validated.
This is a first time approach to optimize the interaction of parameters for C. albicans biofilm formation using RSM. Heterogeneity in growth conditions for local strains of C. albicans clinical isolates was observed. This microtiter plate-based method can be used for future screening of therapeutics for the control of C. albicans.
优化白色念珠菌的生长和生物膜形成对于理解该病原体对医院工具和材料表面的顽固性至关重要，这有助于推进功能分析。使用传统的一次单变量（OVAT）方法对生物膜进行优化和定量一直是一项挑战。本研究采用基于中心复合设计的响应面方法，优化在聚苯乙烯微量滴定板上诱导白色念珠菌生长和生物膜形成的条件。
使用统计软件包Stat Soft®，STASTICA 12.6版进行数据分析。设计矩阵中考虑的变量有培养基pH值、温度、孵育时间、振荡器速度和接种量。采用四管齐下的定量方法，即XTT法（细胞活力）、结晶紫法（生物膜）、钙荧光白法以及湿/干重测量法（细胞量），以了解生物膜形成的不同方面。细胞活力与细胞量呈负相关；然而，生物膜与这两个因素无关。该研究还强调了一个事实，即胎牛血清对某些培养物中的细胞黏附以及体外生物膜形成没有显著贡献。
已开发并验证了一种高通量优化聚苯乙烯微孔板上白色念珠菌生长和生物膜形成的方法。
这是首次使用响应面法优化白色念珠菌生物膜形成参数相互作用的方法。观察到白色念珠菌临床分离株本地菌株生长条件的异质性。这种基于微量滴定板的方法可用于未来控制白色念珠菌的治疗药物筛选。
