AFM imaging of extracellular polymer release by marine diatom Cylindrotheca closterium (Ehrenberg) Reiman & J.C. Lewin

Abstract

Extracellular polysaccharide production by marine diatoms is a significant route by which photosynthetically produced organic carbon enters the trophic web and may influence the physical environment in the sea. This study highlights the capacity of atomic force microscopy (AFM) for investigating diatom extracellular polysaccharides with a subnanometer resolution. Here we address a ubiquitous marine diatom Cylindrotheca closterium, isolated from the northern Adriatic Sea, and its extracellular polymeric substance (EPS) at a single cell level. We applied a simple procedure for AFM imaging of diatom cells on mica under ambient conditions (in air) to achieve visualization of their EPS with molecular resolution. The EPS represents a web of polysaccharide fibrils with two types of cross-linking: fibrils association forming junction zones and fibril-globule interconnections with globules connecting two or more fibrils. The fibril heights were 0.4-2.6 nm while globules height was in the range of 3-12 nm. Polymer networks of native gel samples from the Northern Adriatic and the network formed by polysaccharides extracted from the C. closterium culture share the same features regarding the fibril heights, pore openings and the mode of fibril association, proving that the macroscopic gel phase in the Northern Adriatic can be formed directly by the self-assembly of diatom released polysaccharide fibrils.

Effects of light intensity on components and topographical structures of extracellular polysaccharides from the cyanobacteria Nostoc sp.

“…Pletikapic et al (2011) confirmed the surface morphology of EPS isolated from Cylindrotheca closterium appeared as networks at 10 μg/ml. The AFM image of EPS from Amphidinium carterae appeared as different particle sizes, with 181 nm average roughness (Mandal et al, 2011).…”

Diatom Polysaccharides: Extracellular Production, Isolation and Molecular Characterization

“…However, the segments forming the network shown in Figure 9c are significantly wider (50 vs. 140 nm), suggesting side-by-side associations once the maximum height of individual fibrils is reached. If we take the value of 2.6 nm observed using AFM analysis as the maximum fibril height, then up to six monomolecular fibrils can constitute a single polysaccharide fibril [79]. The absence of globules in the AFM images of isolated polysaccharides also indicates that the globules, which appeared as a constitutive component in the EPS of single cells (Figures 9b,c), are not polysaccharides.…”

“…Protocols for marine sample AFM imaging have been developed only recently: single diatom cells and released polymers [79]; isolated polysaccharides from diatom cultures [3,79]; marine gel polymers and networks [80]. The samples were prepared using the drop deposition method (5-10 µL aliquots) and mica as a substrate.…”

Cell adhesion and spreading at a charged interface: Insight into the mechanism using surface techniques and mathematical modelling

“…The force was kept at the lowest possible value in order to minimize the forces of interaction between the tip and the surface. Measurements were performed in air, at room temperature and 50-60 % relative humidity, which leaves the samples with a small hydration layer, helping to maintain the structure [26] using freshly cleaved mica as a substrate. All images are presented as raw data except for the first-order two-dimensional flattening.…”