1. Accurately quantifying rates and patterns of biological nitrogen fixation (BNF) in terrestrial ecosystems is essential to characterize ecological and biogeochemical interactions, identify mechanistic controls, improve BNF representation in conceptual and numerical modelling, and forecast nitrogen limitation constraints on future carbon (C) cycling.
2. While many resources address the technical advantages and limitations of different methods for measuring BNF, less systematic consideration has been given to the broader decisions involved in planning studies, interpreting data, and extrapolating results. Here, we present a conceptual and practical road map to study design, study execution, data analysis and scaling, outlining key considerations at each step.
3. We address issues including defining N-fixing niches of interest, identifying important sources of temporal and spatial heterogeneity, designing a sampling scheme (including method selection, measurement conditions, replication, and consideration of hotspots and hot moments), and approaches to analysing, scaling and reporting BNF. We also review the comparability of estimates derived using different approaches in the literature, and provide sample R code for simulating symbiotic BNF data frames and upscaling.
4. Improving and standardizing study design at each of these stages will improve the accuracy and interpretability of data, define limits of extrapolation, and facilitate broader use of BNF data for downstream applications. We highlight aspects—such as quantifying scales of heterogeneity, statistical approaches for dealing with non-normality, and consideration of rates versus ecological significance—that are ripe for further development.

中文翻译：

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陆地生态系统中生物固氮取样和规模化的路线图

1. 准确量化陆地生态系统中生物固氮 (BNF) 的速率和模式对于表征生态和生物地球化学相互作用、确定机械控制、改进概念和数值建模中的 BNF 表示以及预测未来碳 (C) 循环的氮限制约束至关重要。
2. 虽然许多资源解决了测量 BNF 的不同方法的技术优势和局限性，但对涉及规划研究、解释数据和推断结果的更广泛决策的系统考虑较少。在这里，我们提出了研究设计、研究执行、数据分析和扩展的概念和实践路线图，概述了每一步的关键考虑因素。
3. 我们解决的问题包括定义感兴趣的 N-fixing 生态位、确定时间和空间异质性的重要来源、设计采样方案（包括方法选择、测量条件、复制以及热点和热点时刻的考虑），以及分析、缩放的方法并报告 BNF。我们还回顾了文献中使用不同方法得出的估计值的可比性，并提供了用于模拟共生 BNF 数据帧和放大的示例 R 代码。
4. 在每个阶段改进和标准化研究设计将提高数据的准确性和可解释性，定义外推的限制，并促进 BNF 数据在下游应用中的更广泛使用。我们强调了一些方面——例如量化异质性的尺度、处理非正态性的统计方法，以及考虑速率与生态重要性——这些都适合进一步发展。