Single polyps were micro-dissected, orientated, and embedded in 1.5% low melting agarose to ease handling of these small tissue samples and to preserve any fragile areas with tissue damage. The agarose was cut into cubes, which were placed in 1% osmium tetroxide in distilled water for 1 h. A series of washes in distilled water (4 × 10 min) and ethanol (3 × 10 min in 50, 70, 90 and 100%) followed, before the samples were embedded in Spurr’s resin. Sections (both 70 nm and 500 nm in thickness) were taken from the mouth region of the polyp using a 45° diamond knife (Diatome, Hatfield, PA, USA). The pharynx of the polyp was chosen as the area of interest, because this is where pathogens were observed to accumulate, both in this study, and in previous studies [37 (link), 39 ]. The resulting thin sections (70 nm) were placed on Formvar/C–coated copper grids, for Scanning Transmission Electron microscopy (STEM), while semi-thin sections (500 nm) were mounted on round glass slides (10 mm) for NanoSIMS imaging.
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