Understanding how bacteria grow and divide requires insight into both the molecular-level dynamics of ultrastructure and the chemistry of the constituent components. Atomic force microscopy (AFM) can provide near molecular resolution images of biological systems but typically provides limited chemical information. Conversely, while super-resolution optical microscopy allows localization of particular molecules and chemistries, information on the molecular context is difficult to obtain. Here, we combine these approaches into STORMForce (stochastic optical reconstruction with atomic force microscopy) and the complementary SIMForce (structured illumination with atomic force microscopy), to map the synthesis of the bacterial cell wall structural macromolecule, peptidoglycan, during growth and division in the rod-shaped bacterium Bacillus subtilis. Using “clickable” d-amino acid incorporation, we fluorescently label and spatially localize a short and controlled period of peptidoglycan synthesis and correlate this information with high-resolution AFM of the resulting architecture. During division, septal synthesis occurs across its developing surface, suggesting a two-stage process with incorporation at the leading edge and with considerable in-filling behind. During growth, the elongation of the rod occurs through bands of synthesis, spaced by ∼300 nm, and corresponds to denser regions of the internal cell wall as revealed by AFM. Combining super-resolution optics and AFM can provide insights into the synthesis processes that produce the complex architectures of bacterial structural biopolymers.

中文翻译：

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相关超分辨率光学和原子力显微镜揭示了枯草芽孢杆菌细菌细胞壁结构与合成之间的关系

了解细菌如何生长和分裂需要深入了解超微结构的分子水平动力学和组成成分的化学。原子力显微镜 (AFM) 可以提供生物系统的近分子分辨率图像，但通常提供有限的化学信息。相反，虽然超分辨率光学显微镜可以定位特定分子和化学物质，但很难获得有关分子背景的信息。在这里，我们将这些方法结合到 STORMForce（使用原子力显微镜的随机光学重建）和互补的 SIMForce（使用原子力显微镜的结构化照明）中，以绘制细菌细胞壁结构大分子肽聚糖在生长和分裂过程中的合成图。棒状细菌枯草芽孢杆菌。使用“可点击”的d-氨基酸掺入，我们荧光标记和空间定位肽聚糖合成的短且受控周期，并将此信息与所得结构的高分辨率 AFM 相关联。在分裂过程中，隔膜合成发生在其发育表面，这表明一个两阶段的过程，在前缘合并，在后面有大量的填充。在生长过程中，杆的伸长通过合成带发生，间隔约 300 nm，并且对应于 AFM 显示的内部细胞壁的更密集区域。结合超分辨率光学和 AFM 可以深入了解产生细菌结构生物聚合物复杂结构的合成过程。