Evaluation of Genomic Sequence-Based Growth Rate Methods for Synchronized Synechococcus Cultures
Evaluation of Genomic Sequence-Based Growth Rate Methods for Synchronized Synechococcus Cultures
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DOI：
10.1128/aem.01743-21
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发表时间：
2021-10
影响因子：
4.4
通讯作者：
J. Carroll;N. Van Oostende;B. Ward
中科院分区：
文献类型：
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作者：
J. Carroll;N. Van Oostende;B. Ward
小但丰富的蓝细菌菌株，如光合聚球藻属。海洋生态系统之所以重要，是因为它们对海洋的初级生产力作出了重大贡献。这些细菌产生氧气，并提供生物可利用的碳，这是生物体在更高的营养水平所必需的。通过使用替代物（如原位荧光、细胞周期或细胞计数）计算微生物生长速率（μ）的标准方法对于确定细菌在海洋碳（C）和氮（N）循环中所起作用的大小至关重要。在混合组合的分类特定的增长率将是有用的归因于单个物种的生态地球化学过程和理解相关的分支，如聚球藻和原绿球藻之间的生态位分化。我们测试了三种新的基于DNA测序的方法（iRep，bPTR和GRiD），用于评估不同光照强度和温度下光同步聚球藻培养物的生长。使用体内荧光和细胞周期分析来获得生长速率的标准估计值，以与基于序列的方法（SBM）进行比较。SBM值均与通过标准技术计算的生长速率不相关，尽管所有三个SBM与在昼夜周期中处于S期（DNA复制）的细胞的百分比相关。确定最大DNA复制时间的不准确性不太可能完全解释SBM和生长率之间缺乏关系的原因，但海洋表层大多数微生物表现出一定程度的昼夜循环性这一事实是应用这些方法的一个警告。SBM与DNA复制相关，但不能用生长速率定量解释。重要性小，但丰富，蓝藻菌株，如光合聚球藻属。海洋生态系统之所以重要，是因为它们对海洋的初级生产力作出了重大贡献。这些细菌产生氧气，并提供生物可利用的碳，这是生物体在更高的营养水平所必需的。天然微生物群落的小规模和多样性意味着分类特异性活动（例如，增长率）在该领域很难获得。有人建议，基于序列的方法（SBM）可能能够解决这个问题。然而，我们发现，SBM可以检测到DNA复制，并与细胞周期的阶段，但不能解释在绝对增长率聚球藻培养下生长的昼夜周期，如在海洋中经历的。
Small but abundant, cyanobacterial strains such as the photosynthetic Synechococcus spp. are important because they contribute significantly to primary productivity in the ocean. These bacteria generate oxygen and provide biologically available carbon, which is essential for organisms at higher trophic levels. ABSTRACT Standard methods for calculating microbial growth rates (μ) through the use of proxies, such as in situ fluorescence, cell cycle, or cell counts, are critical for determining the magnitude of the role bacteria play in marine carbon (C) and nitrogen (N) cycles. Taxon-specific growth rates in mixed assemblages would be useful for attributing biogeochemical processes to individual species and understanding niche differentiation among related clades, such as found in Synechococcus and Prochlorococcus. We tested three novel DNA sequencing-based methods (iRep, bPTR, and GRiD) for evaluating the growth of light-synchronized Synechococcus cultures under different light intensities and temperatures. In vivo fluorescence and cell cycle analysis were used to obtain standard estimates of growth rate for comparison with those of the sequence-based methods (SBM). None of the SBM values were correlated with growth rates calculated by standard techniques despite the fact that all three SBM were correlated with the percentage of cells in S phase (DNA replication) over the diel cycle. Inaccuracy in determining the time of maximum DNA replication is unlikely to account entirely for the absence of a relationship between SBM and growth rate, but the fact that most microbes in the surface ocean exhibit some degree of diel cyclicity is a caution for application of these methods. SBM correlate with DNA replication but cannot be interpreted quantitatively in terms of growth rate. IMPORTANCE Small but abundant, cyanobacterial strains such as the photosynthetic Synechococcus spp. are important because they contribute significantly to primary productivity in the ocean. These bacteria generate oxygen and provide biologically available carbon, which is essential for organisms at higher trophic levels. The small size and diversity of natural microbial assemblages mean that taxon-specific activities (e.g., growth rate) are difficult to obtain in the field. It has been suggested that sequence-based methods (SBM) may be able to solve this problem. We find, however, that SBM can detect DNA replication and are correlated with phases of the cell cycle but cannot be interpreted in terms of absolute growth rate for Synechococcus cultures growing under a day-night cycle, like that experienced in the ocean.