Mumford Adam C, Adaktylou Irini J, Emerson David
Bigelow Laboratory for Ocean Sciences, East Boothbay, Maine, USA
Center for Applied Geoscience, University of Tuebingen, Tuebingen, Germany.
Appl Environ Microbiol. 2016 Oct 27;82(22):6799-6807. doi: 10.1128/AEM.01990-16. Print 2016 Nov 15.
Microbially influenced corrosion (MIC) is a major cause of damage to steel infrastructure in the marine environment. Despite their ability to grow directly on Fe(II) released from steel, comparatively little is known about the role played by neutrophilic iron-oxidizing bacteria (FeOB). Recent work has shown that FeOB grow readily on mild steel (1018 MS) incubated in situ or as a substrate for pure cultures in vitro; however, details of how they colonize steel surfaces are unknown yet are important for understanding their effects. In this study, we combine a novel continuously upwelling microcosm with confocal laser scanning microscopy (CLSM) to determine the degree of colonization of 1018 MS by the marine FeOB strain DIS-1. 1018 MS coupons were incubated with sterile seawater (pH 8) inoculated with strain DIS-1. Incubations were performed both under oxic conditions and in an anoxic-to-oxic gradient. Following incubations of 1 to 10 days, the slides were removed from the microcosms and stained to visualize both cells and stalk structures. Stained coupons were visualized by CLSM after being mounted in a custom frame to preserve the three-dimensional structure of the biofilm. The incubation of 1018 MS coupons with strain DIS-1 under oxic conditions resulted in initial attachment of cells within 2 days and nearly total coverage of the coupon with an ochre film within 5 days. CLSM imaging revealed a nonadherent biofilm composed primarily of the Fe-oxide stalks characteristic of strain DIS-1. When incubated with elevated concentrations of Fe(II), DIS-1 colonization of 1018 MS was inhibited.
These experiments describe the growth of a marine FeOB in a continuous culture system and represent direct visualizations of steel colonization by FeOB. We anticipate that these experiments will lay the groundwork for studying the mechanisms by which FeOB colonize steel and help to elucidate the role played by marine FeOB in MIC. These observations of the interaction between an FeOB, strain DIS-1, and steel suggest that this experimental system will provide a useful model for studying the interactions between microbes and solid substrates.
微生物影响的腐蚀（MIC）是海洋环境中钢铁基础设施损坏的主要原因。尽管嗜中性铁氧化细菌（FeOB）能够直接利用钢铁释放的Fe（II）生长，但人们对其作用的了解相对较少。最近的研究表明，FeOB能够在原位培养的低碳钢（1018 MS）上或作为体外纯培养的底物上轻松生长；然而，它们如何在钢铁表面定殖的细节尚不清楚，但对于理解它们的影响很重要。在本研究中，我们将一种新型的连续上升微宇宙与共聚焦激光扫描显微镜（CLSM）相结合，以确定海洋FeOB菌株DIS-1对1018 MS的定殖程度。将1018 MS试样与接种了菌株DIS-1的无菌海水（pH 8）一起培养。培养在有氧条件下以及在缺氧到有氧的梯度中进行。在培养1至10天后，将载玻片从微宇宙中取出并染色，以观察细胞和柄结构。染色后的试样安装在定制框架中以保留生物膜的三维结构，然后通过CLSM进行观察。在有氧条件下，将1018 MS试样与菌株DIS-1一起培养，导致细胞在2天内开始附着，并在5天内试样几乎完全被赭色膜覆盖。CLSM成像显示了一种主要由菌株DIS-1特有的铁氧化物柄组成的非粘附性生物膜。当与高浓度的Fe（II）一起培养时，DIS-1对1018 MS的定殖受到抑制。
这些实验描述了海洋FeOB在连续培养系统中的生长，并代表了FeOB对钢铁定殖的直接可视化。我们预计这些实验将为研究FeOB定殖钢铁的机制奠定基础，并有助于阐明海洋FeOB在MIC中的作用。对FeOB菌株DIS-1与钢铁之间相互作用的这些观察表明，该实验系统将为研究微生物与固体底物之间的相互作用提供一个有用的模型。
