Jee 400

Thirty microliters of purified phage suspension was adsorbed to carbon-coated copper grids (400-mesh) in a vacuum evaporator (JEE400, JEOL Ltd. Tokyo, Japan), allowed to air dry and then negatively stained with 2% phosphotungstic acid (pH 7.2). The excess solution was absorbed with filter paper, and samples were observed with a transmission electron microscope (JEM-1011, JEOL Ltd. Tokyo, Japan) operating at 80 kV (López-Cuevas et al., 2011 (link)).
Bacteriophage plaques formed on a TSA plate during the process of propagation (using dilutions that generated 15–30 plaques per plate) were analyzed according to the procedure described by Gallet, Kannoly & Wang (2011) (link) with minor modifications. Briefly, images of ten plates were captured by a supersensitive high-resolution 16-bit camera that was deeply cooled for faint image detection (Bio-Rad Laboratories), and the image of five plaques for each plate were displayed with the ImageJ software (developed at the National Institutes of Health, Bethesda, Maryland). The plates were then incubated for 18–24 h at 37 °C before plaque size determination. To calculate the surface area (expressed in square millimeters) corresponding to each pixel, a graticule of 1 mm² was used as the reference scale for the simplified measurement of the lysis plaques. According to the analysis, each pixel corresponded to 0.5 mm².

Electron micrographs of purified phage particles were obtained according to standard method. Suspension phage sample was dropped (approximately 30 μL) onto 400-mesh carbon-coated Formvar covered grids placed in a vacuum evaporator (JEE400, JEOL Ltd. Tokyo, Japan), stained with 2% (wt/vol) phosphotungstic acid (pH 7.2), and air dried. Samples were examined in a transmission electron microscope (JEM-1011, JEOL Ltd. Tokyo, Japan) at an acceleration voltage of 80 kV, and phage particles were examined at 15,000–25,000 times magnification.

The surface of the coverslip of a glass-bottom chamber was coated with carbon by vapor deposition using a vacuum evaporator (JEOL, JEE-400). The thickness of the carbon layer was approximately 20 nm. To make the surface hydrophilic, the surface of the carbon-coated coverslip was activated with a plasma treatment. The chamber was sterilized with 70% ethanol and dried if necessary. The cells were settled on the surface of the carbon-coated coverslip, and they were slightly compressed with agarose block (1.5%, dissolved in BSS, 1 mm thick) to observe the ventral cell surface^{52 (link)}.

Blueberry fragments were dried by critical point drying (CPD 030, BAL-TEC, Balzers, Liechtenstein). The following procedure was adopted to prepare the samples for drying: (1) the fruits were cut perpendicular to the long axis with a fresh razor blade, (2) each sample was placed in a fixative containing glutardialdehyde (2.5 g = 100 g in 0.1 mol/L phosphate buffer, pH = 7.2) for 48 h at a temperature of 4 °C, (3) the sample was rinsed in Milli-Q water, and (4) the sample was dehydrated for 15 min in a graded ethanol series (30–99.8 mL = 100 mL). The specimens were mounted on aluminum stubs using silver paste and coated with gold in a vacuum evaporator (JEE 400, JEOL, Tokyo, Japan) to make their surfaces electrically conductive for SEM analysis. The micrographs of berry cross-sections were acquired with a scanning electron microscope (JEOL, model 5200, Tokyo, Japan), and accelerating voltage was set at 10 kV.