Helbling Damian E, Hammes Frederik, Egli Thomas, Kohler Hans-Peter E
Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Microbiology, Dübendorf, Switzerland.
Appl Environ Microbiol. 2014 Feb;80(4):1306-13. doi: 10.1128/AEM.03622-13. Epub 2013 Dec 6.
The fundamentals of growth-linked biodegradation occurring at low substrate concentrations are poorly understood. Substrate utilization kinetics and microbial growth yields are two critically important process parameters that can be influenced by low substrate concentrations. Standard biodegradation tests aimed at measuring these parameters generally ignore the ubiquitous occurrence of assimilable organic carbon (AOC) in experimental systems which can be present at concentrations exceeding the concentration of the target substrate. The occurrence of AOC effectively makes biodegradation assays conducted at low substrate concentrations mixed-substrate assays, which can have profound effects on observed substrate utilization kinetics and microbial growth yields. In this work, we introduce a novel methodology for investigating biodegradation at low concentrations by restricting AOC in our experiments. We modified an existing method designed to measure trace concentrations of AOC in water samples and applied it to systems in which pure bacterial strains were growing on pesticide substrates between 0.01 and 50 mg liter(-1). We simultaneously measured substrate concentrations by means of high-performance liquid chromatography with UV detection (HPLC-UV) or mass spectrometry (MS) and cell densities by means of flow cytometry. Our data demonstrate that substrate utilization kinetic parameters estimated from high-concentration experiments can be used to predict substrate utilization at low concentrations under AOC-restricted conditions. Further, restricting AOC in our experiments enabled accurate and direct measurement of microbial growth yields at environmentally relevant concentrations for the first time. These are critical measurements for evaluating the degradation potential of natural or engineered remediation systems. Our work provides novel insights into the kinetics of biodegradation processes and growth yields at low substrate concentrations.
在低底物浓度下发生的与生长相关的生物降解的基本原理目前还知之甚少。底物利用动力学和微生物生长产量是两个至关重要的过程参数，它们可能会受到低底物浓度的影响。旨在测量这些参数的标准生物降解测试通常忽略了实验系统中普遍存在的可同化有机碳（AOC），其浓度可能超过目标底物的浓度。AOC的存在实际上使在低底物浓度下进行的生物降解测定成为混合底物测定，这可能会对观察到的底物利用动力学和微生物生长产量产生深远影响。在这项工作中，我们引入了一种新方法，通过在实验中限制AOC来研究低浓度下的生物降解。我们改进了一种现有的用于测量水样中痕量AOC浓度的方法，并将其应用于纯细菌菌株在0.01至50毫克/升的农药底物上生长的系统。我们通过高效液相色谱-紫外检测（HPLC-UV）或质谱（MS）同时测量底物浓度，并通过流式细胞术测量细胞密度。我们的数据表明，从高浓度实验估计的底物利用动力学参数可用于预测在AOC限制条件下低浓度下的底物利用情况。此外，在我们的实验中限制AOC首次实现了在环境相关浓度下对微生物生长产量的准确直接测量。这些是评估天然或工程修复系统降解潜力的关键测量。我们的工作为低底物浓度下生物降解过程的动力学和生长产量提供了新的见解。
