Ellipse 80i light microscope

For morphological studies, we used three different media: cornmeal dextrose agar (CMD, Difco, BD Science, USA), PDA (Difco, BD Science, USA), and synthetic low nutrient agar (SNA, Difco, BD Science, USA) (Chaverri et al. 2015 (link)). Each strain was first cultured on an SNA plate for 3 days and a small agar piece of 0.5 cm diameter with mycelium was then transferred, respectively, to new CMD, PDA, and SNA plates. Strains were incubated in 9 cm diam with three replicates. Petri dishes at 25 °C with a 12 h natural light and 12 h darkness interval. Colony diameter at 25 °C was measured three days after inoculation, and the time when mycelium entirely covered the surface of the agar plate was also recorded. Micromorphological characters were examined from the cultures of one-week-old colonies on SNA (Chaverri et al. 2015 (link)). A Nikon Ellipse 80i light microscope, equipped with differential interference contrast (DIC) optics, was used to capture digital images.

Surface scrapings were obtained from the border of the skin lesions on the patient’s cheek for direct microscopic examination of potassium hydroxide (KOH). Surface scrapings were sterilized for 15 s with 75% alcohol and blood with sterilized water for 1 min and then inoculated on the Sabouraud medium. Finally, the subculture was incubated on yeast extract peptone dextrose agar (YPD) at 30 and 37°C, respectively, for 7 days. Our newly obtained isolate was deposited in the Air Force Medical Center Culture Collection (AFMCC) and was preserved in the China General Microbiological Culture Collection Center (CGMCC) as well. A Nikon Ellipse 80i light microscope, equipped with differential interference contrast (DIC) optics, was used to capture digital images. Tarosoft (R) v.0.9.7 Image Frame Work was used to measure the morphological structures and the Adobe Photoshop CS6 image 22 software (Adobe Systems, United States) to edit the photographic plates.
For observation by SEM, the fungus was inoculated on a PDA medium at 30°C. After 7 days, two patches (0.3 cm × 0.3 cm) of the fungal colony were fixed in 2.5% glutaraldehyde in 0.1 M phosphate-buffered saline (BPS, pH 7.2) at 4°C. Then, the samples were dehydrated and sputter-coated by following the method described by Sun et al. (2023) (link). Samples were loaded on the SEM (SU8010, Hitachi, Tokyo, Japan) and observed and photographed.