Data Sheet 1_Optimization of Candida tropicalis growth conditions on silicone elastomer material by response surface methodology.docx

Biofilm in the emerging pathogen Candida tropicalis and the most prevalent Non-Albicans Candida infections is linked to fouling of medical devices and virulence. The growth conditions (temperature, media pH, incubation time, inoculum size, and shaker speed) for clinical cultures of C. tropicalis were optimized on silicone elastomer material by Central composite design based on Response surface methodology. Six clinical cultures (C4, U873, U951, U1179, U1309 and U1360) and a standard culture (MTCC-184) were chosen for the study. Growth and biofilm were quantified for all the cultures by crystal violet (biofilm), MTT (cell viability), calcofluor white (cell mass), and wet and dry weight (cell mass) measurements. Among the isolates, U951 was found to fit the CCD model. The non-normal distribution and heteroscedasticity of the data favored the transformation via CCD-integrated Johnson model profiler for the prediction of the optimal growth conditions. For U951 isolate, biofilm formation was impacted by temperature and incubation time. A direct correlation was observed between biofilm formation and cell viability, with variations in the cell mass in all the cultures. This is the first of its kind study to advance an in vitro silicone elastomer-based high-throughput growth model of C. tropicalis for various applications, including the screening of potential therapeutics.

新兴病原菌热带假丝酵母（Candida tropicalis）所形成的生物膜，以及最流行的非白假丝酵母菌（Non-Albicans Candida）感染相关的生物膜，均与医疗装置结垢及致病菌毒力密切相关。本研究基于响应面法（Response Surface Methodology, RSM）的中心复合设计（Central Composite Design, CCD），在硅酮弹性体材料表面优化了热带假丝酵母临床菌株的培养生长条件，涵盖温度、培养基pH值、孵育时间、接种量及摇床转速。本研究选取6株临床菌株（C4、U873、U951、U1179、U1309及U1360）与1株标准菌株（MTCC-184）作为实验对象。采用结晶紫染色法（检测生物膜）、MTT法（检测细胞活力）、荧光增白剂（calcofluor white）染色法（检测细胞量）以及干湿重法（检测细胞量），对所有菌株的生长情况与生物膜形成能力进行定量分析。在所有受试菌株中，U951菌株的实验数据最契合中心复合设计模型。鉴于实验数据存在非正态分布与异方差性，本研究通过集成中心复合设计的约翰逊模型剖析工具完成数据转换，以预测最优生长条件。针对U951菌株，其生物膜形成能力受温度与孵育时间的显著影响。所有受试菌株的细胞量均存在差异，且生物膜形成能力与细胞活力呈直接正相关。本研究首次构建了基于硅酮弹性体的热带假丝酵母体外（in vitro）高通量生长模型，可应用于包括潜在治疗药物筛选在内的多个领域。