Bowsher Alan W, Kearns Patrick J, Shade Ashley
Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan, USA.
Plant Resilience Institute, Michigan State University, East Lansing, Michigan, USA.
mSystems. 2019 Apr 2;4(2). doi: 10.1128/mSystems.00003-19. eCollection 2019 Mar-Apr.
At any given time, only a subset of microbial community members are active in their environment. The others are in a state of dormancy, with strongly reduced metabolic rates. It is of interest to distinguish active and inactive microbial cells and taxa to understand their functional contributions to ecosystem processes and to understand shifts in microbial activity in response to change. Of the methods used to assess microbial activity-dormancy dynamics, 16S rRNA/rRNA gene amplicons (16S ratios) and active cell staining with 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) are two of the most common, yet each method has limitations. Given that activity-dormancy dynamics are proxied only by laboratory methods, further study is needed to assess the level of agreement and potential complementarity of these methods. We conducted two experiments investigating microbial activity in plant-associated soils. First, we treated corn field soil with phytohormones to simulate plant soil stress signaling, and second, we used rhizosphere soil from common bean plants exposed to drought or nutrient enrichment. Overall, the 16S ratio and CTC methods exhibited similar patterns of relative activity across treatments when treatment effects were large, and the instances in which they differed could be attributed to changes in community size (e.g., cell death or growth). Therefore, regardless of the method used to assess activity, we recommend quantifying community size to inform ecological interpretation. Our results suggest that the 16S ratio and CTC methods report comparable patterns of activity that can be applied to observe ecological dynamics over time, space, or experimental treatment. Although the majority of microorganisms in natural ecosystems are dormant, relatively little is known about the dynamics of the active and dormant microbial pools through both space and time. The limited knowledge of microbial activity-dormancy dynamics is in part due to uncertainty in the methods currently used to quantify active taxa. Here, we directly compared two of the most common methods (16S ratios and active cell staining) for estimating microbial activity in plant-associated soil and found that they were largely in agreement in the overarching patterns. Our results suggest that 16S ratios and active cell staining provide complementary information for measuring and interpreting microbial activity-dormancy dynamics in soils. They also support the idea that 16S rRNA/rRNA gene ratios have comparative value and offer a high-throughput, sequencing-based option for understanding relative changes in microbiome activity, as long as this method is coupled with quantification of community size.
在任何给定时间，微生物群落中只有一部分成员在其环境中处于活跃状态。其他成员则处于休眠状态，代谢率大幅降低。区分活跃和不活跃的微生物细胞及分类群，对于了解它们对生态系统过程的功能贡献以及理解微生物活动随环境变化的转变具有重要意义。在用于评估微生物活动-休眠动态的方法中，16S rRNA/rRNA基因扩增子（16S比率）和用5-氰基-2,3-二苯基氯化四氮唑（CTC）进行的活跃细胞染色是两种最常用的方法，但每种方法都有局限性。鉴于活动-休眠动态仅通过实验室方法来推断，因此需要进一步研究以评估这些方法的一致性水平和潜在互补性。我们进行了两项关于植物相关土壤中微生物活动的实验。首先，我们用植物激素处理玉米田土壤以模拟植物-土壤胁迫信号，其次，我们使用了暴露于干旱或养分富集条件下的普通菜豆植物的根际土壤。总体而言，当处理效应较大时，16S比率和CTC方法在各处理间呈现出相似的相对活动模式，而它们出现差异的情况可归因于群落大小的变化（例如细胞死亡或生长）。因此，无论使用何种方法评估活动，我们建议对群落大小进行量化，以便进行生态学解释。我们的结果表明，16S比率和CTC方法报告的活动模式具有可比性，可用于观察随时间、空间或实验处理的生态动态。虽然自然生态系统中的大多数微生物处于休眠状态，但关于活跃和休眠微生物库在时空上的动态变化，我们所知甚少。对微生物活动-休眠动态了解有限，部分原因在于目前用于量化活跃分类群的方法存在不确定性。在此，我们直接比较了两种用于估计植物相关土壤中微生物活动的最常用方法（16S比率和活跃细胞染色），发现它们在总体模式上基本一致。我们的结果表明，16S比率和活跃细胞染色为测量和解释土壤中的微生物活动-休眠动态提供了互补信息。它们还支持这样一种观点，即16S rRNA/rRNA基因比率具有比较价值，并为理解微生物群落活动的相对变化提供了一种基于测序的高通量选择，只要该方法与群落大小的量化相结合即可。
