Oral Sciences Research Group, School of Medicine, Dentistry and Nursing, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom.
Institute of Healthcare, Policy and Practise, University of the West of Scotland, Paisley, United Kingdom.
mSphere. 2018 Jul 11;3(4):e00334-18. doi: 10.1128/mSphere.00334-18.
has emerged as a significant global nosocomial pathogen. This is primarily due to its antifungal resistance profile but also its capacity to form adherent biofilm communities on a range of clinically important substrates. While we have a comprehensive understanding of how other species resist and respond to antifungal challenge within the sessile phenotype, our current understanding of biofilm-mediated resistance is lacking. In this study, we are the first to perform transcriptomic analysis of temporally developing biofilms, which were shown to exhibit phase- and antifungal class-dependent resistance profiles. A transcriptome assembly was performed, where sequenced sample reads were assembled into an ~11.5-Mb transcriptome consisting of 5,848 genes. Differential expression (DE) analysis demonstrated that 791 and 464 genes were upregulated in biofilm formation and planktonic cells, respectively, with a minimum 2-fold change. Adhesin-related glycosylphosphatidylinositol (GPI)-anchored cell wall genes were upregulated at all time points of biofilm formation. As the biofilm developed into intermediate and mature stages, a number of genes encoding efflux pumps were upregulated, including ATP-binding cassette (ABC) and major facilitator superfamily (MFS) transporters. When we assessed efflux pump activity biochemically, biofilm efflux was greater than that of planktonic cells at 12 and 24 h. When these were inhibited, fluconazole sensitivity was enhanced 4- to 16-fold. This study demonstrates the importance of efflux-mediated resistance within complex communities and may explain the resistance of to a range of antimicrobial agents within the hospital environment. Fungal infections represent an important cause of human morbidity and mortality, particularly if the fungi adhere to and grow on both biological and inanimate surfaces as communities of cells (biofilms). Recently, a previously unrecognized yeast, , has emerged globally that has led to widespread concern due to the difficulty in treating it with existing antifungal agents. Alarmingly, it is also able to grow as a biofilm that is highly resistant to antifungal agents, yet we are unclear about how it does this. Here, we used a molecular approach to investigate the genes that are important in causing the cells to be resistant within the biofilm. The work provides significant insights into the importance of efflux pumps, which actively pump out toxic antifungal drugs and therefore enhance fungal survival within a variety of harsh environments.
已经成为一种重要的全球医院病原体。这主要是由于其抗真菌耐药性，但也由于其能够在一系列临床重要的基质上形成附着生物膜群落。虽然我们对其他物种在静止表型中如何抵抗和应对抗真菌挑战有了全面的了解，但我们对生物膜介导的耐药性的了解还很缺乏。在这项研究中，我们首次对随时间发展的生物膜进行了转录组分析，结果表明生物膜表现出与阶段和抗真菌类别的耐药性有关。进行了转录组组装，其中测序样本的读取被组装成一个大约 1150 兆碱基的转录组，由 5848 个基因组成。差异表达（DE）分析表明，生物膜形成和浮游细胞中分别有 791 和 464 个基因上调，最低 2 倍变化。在生物膜形成的所有时间点，粘附素相关的糖基磷脂酰肌醇（GPI）锚定细胞壁基因都上调。随着生物膜发展为中间和成熟阶段，许多编码外排泵的基因上调，包括 ATP 结合盒（ABC）和主要易化剂超家族（MFS）转运蛋白。当我们用生物化学方法评估外排泵活性时，生物膜的外排大于浮游细胞在 12 和 24 小时时的外排。当这些被抑制时，氟康唑的敏感性增强了 4 到 16 倍。这项研究表明了在复杂的生物膜中，外排介导的耐药性的重要性，并且可能解释了在医院环境中对多种抗菌剂的耐药性。真菌感染是人类发病率和死亡率的一个重要原因，特别是如果真菌附着并在生物和无生命的表面上作为细胞群落（生物膜）生长。最近，一种以前未被识别的酵母，已经在全球范围内出现，由于现有的抗真菌药物难以治疗，引起了广泛关注。令人震惊的是，它也能够作为一种生物膜生长，这种生物膜对抗真菌药物高度耐药，但我们不清楚它是如何做到的。在这里，我们使用分子方法来研究在生物膜中导致细胞产生耐药性的重要基因。这项工作为外排泵的重要性提供了重要的见解，外排泵主动泵出有毒的抗真菌药物，从而增强了真菌在各种恶劣环境中的生存能力。
