Bristol Dental School, University of Bristolgrid.5337.2, Bristol, United Kingdom.
Wolfson Bioimaging Facility, Biomedical Sciences Building, University of Bristolgrid.5337.2, Bristol, United Kingdom.
Appl Environ Microbiol. 2022 Jul 12;88(13):e0069822. doi: 10.1128/aem.00698-22. Epub 2022 Jun 13.
Extracellular DNA (eDNA) is an important component of biofilm matrix that serves to maintain biofilm structural integrity, promotes genetic exchange within the biofilm, and provides protection against antimicrobial compounds. Advances in microscopy techniques have provided evidence of the cobweb- or lattice-like structures of eDNA within biofilms from a range of environmental niches. However, methods to reliably assess the abundance and architecture of eDNA remain lacking. This study aimed to address this gap by development of a novel, high-throughput image acquisition and analysis platform for assessment of eDNA networks within biofilms. Utilizing Streptococcus gordonii as the model, the capacity for this imaging system to reliably detect eDNA networks and monitor changes in abundance and architecture (e.g., strand length and branch number) was verified. Evidence was provided of a synergy between glucans and eDNA matrices, while it was revealed that surface-bound nuclease SsnA could modify these eDNA structures under conditions permissive for enzymatic activity. Moreover, cross talk between the competence and hexaheptapeptide permease systems was shown to regulate eDNA release by S. gordonii. This novel imaging system can be applied across the wider field of biofilm research, with potential to significantly advance interrogation of the mechanisms by which the eDNA network architecture develops, how it can influence biofilm properties, and how it may be targeted for therapeutic benefit. Extracellular DNA (eDNA) is critical for maintaining the structural integrity of many microbial biofilms, making it an attractive target for the management of biofilms. However, our knowledge and targeting of eDNA are currently hindered by a lack of tools for the quantitative assessment of eDNA networks within biofilms. Here, we demonstrate use of a novel image acquisition and analysis platform with the capacity to reliably monitor the abundance and architecture of eDNA networks. Application of this tool to Streptococcus gordonii biofilms has provided new insights into how eDNA networks are stabilized within the biofilm and the pathways that can regulate eDNA release. This highlights how exploitation of this novel imaging system across the wider field of biofilm research has potential to significantly advance interrogation of the mechanisms by which the eDNA network architecture develops, how it can influence biofilm properties, and how it may be targeted for therapeutic benefit.
细胞外 DNA（eDNA）是生物膜基质的重要组成部分，有助于维持生物膜的结构完整性、促进生物膜内的基因交换，并为抗菌化合物提供保护。显微镜技术的进步为来自各种环境小生境的生物膜中的 eDNA 的蛛网或晶格状结构提供了证据。然而，仍然缺乏可靠评估 eDNA 丰度和结构的方法。本研究旨在通过开发一种新颖的、高通量的图像采集和分析平台来解决这一差距，用于评估生物膜内的 eDNA 网络。利用酿脓链球菌作为模型，验证了该成像系统可靠检测 eDNA 网络并监测丰度和结构变化（例如，链长和分支数）的能力。证据表明，葡聚糖和 eDNA 基质之间存在协同作用，同时表明在允许酶活性的条件下，表面结合的核酸酶 SsnA 可以修饰这些 eDNA 结构。此外，还表明了感受态和六肽透酶系统之间的串扰可调节酿脓链球菌释放 eDNA。这种新型成像系统可应用于更广泛的生物膜研究领域，具有显著推进 eDNA 网络结构如何发展、它如何影响生物膜特性以及如何针对其进行治疗获益的机制研究的潜力。细胞外 DNA（eDNA）对于维持许多微生物生物膜的结构完整性至关重要，因此成为管理生物膜的有吸引力的目标。然而，我们对 eDNA 的了解和靶向目前受到缺乏用于定量评估生物膜内 eDNA 网络的工具的限制。在这里，我们展示了一种具有可靠监测 eDNA 网络丰度和结构能力的新型图像采集和分析平台的应用。将该工具应用于酿脓链球菌生物膜，提供了有关 eDNA 网络如何在生物膜内稳定以及可以调节 eDNA 释放的途径的新见解。这突出了在更广泛的生物膜研究领域中利用这种新型成像系统具有显著推进研究 eDNA 网络结构如何发展、它如何影响生物膜特性以及如何针对其进行治疗获益的机制的潜力。
