Quattro s environmental

Walnut hull (exocarp) discs were fixed overnight in 0.1-M sodium phosphate buffer with 2.5% glutaraldehyde and 2% paraformaldehyde. Tissues were rinsed three times in 0.1M NaH2PO4 for 10 min and dehydrated in increasing concentrations of ethanol (EtOH) for 10 min each (30, 50, 70, 95, and 100% EtOH). Dehydration with 95% and 100% EtOH was repeated three times. Samples were dried using Tousimis 931 GL Super Critical Autosamdri, mounted onto aluminum stubs and coated with gold using Pelco Auto Sputter Coater SC-7. Micrographs were produced on a Thermo Fisher Quattro S Environmental operating at 15 kV at the UC Davis Advanced Materials Characterization and Testing laboratory (AMCaT).

To analyze the fungal pellet structure, pellets were observed using a Thermo Fisher Quattro S Environmental scanning electron microscope (Thermo Scientific, Rochester, NY, USA) in high vacuum mode using 5 kV accelerating voltage to capture pellets before a brief increase to 15 kV for imaging. Prior to imaging, pellets and yeast biocapsules were freeze-dried and gold-sputter-coated. To observe the inner area of the fungal pellets or yeast biocapsules, pellets were cut in half with a scalpel. Yeast cells in images obtained from SEM were later colored using Adobe Photoshop CS6 13.1.3 by Adobe Inc. (San Jose, CA, USA).