Department of Environmental Science, University of Arizona, Tucson, Arizona, USA.
School of Animal and Comparative Biomedical Science, University of Arizona, Tucson, Arizona, USA.
Microbiol Spectr. 2024 May 2;12(5):e0365023. doi: 10.1128/spectrum.03650-23. Epub 2024 Mar 19.
Anaerobic microbes play crucial roles in environmental processes, industry, and human health. Traditional methods for monitoring the growth of anaerobes, including plate counts or subsampling broth cultures for optical density measurements, are time and resource-intensive. The advent of microplate readers revolutionized bacterial growth studies by enabling high-throughput and real-time monitoring of microbial growth kinetics. Yet, their use in anaerobic microbiology has remained limited. Here, we present a workflow for using small-footprint microplate readers and the Growthcurver R package to analyze the kinetic growth metrics of anaerobic bacteria. We benchmarked the small-footprint Cerillo Stratus microplate reader against a BioTek Synergy HTX microplate reader in aerobic conditions using DSM 28618 cultures. The growth rates and carrying capacities obtained from the two readers were statistically indistinguishable. However, the area under the logistic curve was significantly higher in cultures monitored by the Stratus reader. We used the Stratus to quantify the growth responses of anaerobically grown and DSM 29485 to different doses of the toxin sodium arsenite. The growth of and was sensitive to arsenite doses of 1.3 µM and 0.4 µM, respectively. Complete inhibition of growth was achieved at 38 µM arsenite for and 338 µM in . These results show that the Stratus performs similarly to a leading brand of microplate reader and can be reliably used in anaerobic conditions. We discuss the advantages of the small format microplate readers and our experiences with the Stratus.
We present a workflow that facilitates the production and analysis of growth curves for anaerobic microbes using small-footprint microplate readers and an R script. This workflow is a cost and space-effective solution to most high-throughput solutions for collecting growth data from anaerobic microbes. This technology can be used for applications where high throughput would advance discovery, including microbial isolation, bioprospecting, co-culturing, host-microbe interactions, and drug/toxin-microbial interactions.
厌氧微生物在环境过程、工业和人类健康中发挥着关键作用。监测厌氧菌生长的传统方法，包括平板计数或取部分培养液进行光密度测量，既费时又费资源。微孔板读数器的出现彻底改变了细菌生长研究，使其能够高通量实时监测微生物生长动力学。然而，它们在厌氧微生物学中的应用仍然有限。在这里，我们提出了一种使用小足迹微孔板读数器和 Growthcurver R 包分析厌氧菌动力学生长指标的工作流程。我们在有氧条件下使用 DSM 28618 培养物对小足迹 Cerillo Stratus 微孔板读数器与 BioTek Synergy HTX 微孔板读数器进行了基准测试。从两个读数器获得的生长率和承载能力在统计学上没有区别。然而，Stratus 监测的培养物中逻辑曲线下的面积显著更高。我们使用 Stratus 来量化在不同剂量的毒素亚砷酸钠下厌氧生长的 和 DSM 29485 的生长反应。 和 的生长对亚砷酸钠剂量分别为 1.3µM 和 0.4µM 敏感。亚砷酸钠浓度为 38µM 时 和 的生长完全受到抑制，而在 中为 338µM。这些结果表明 Stratus 的性能与领先品牌的微孔板读数器相似，可以在厌氧条件下可靠使用。我们讨论了小格式微孔板读数器的优势以及我们使用 Stratus 的经验。
我们提出了一种工作流程，该流程使用小足迹微孔板读数器和 R 脚本方便地为厌氧微生物生成和分析生长曲线。该工作流程是一种经济高效的解决方案，可用于从厌氧微生物收集生长数据的大多数高通量解决方案。该技术可用于需要高通量来推进发现的应用，包括微生物分离、生物勘探、共培养、宿主-微生物相互作用和药物/毒素-微生物相互作用。
