Sampling and yeast isolation
The sampling for this study
took place in the Atacama Desert near the San Pedro de Atacama village
(22° 55′ S, 68° 12′ W) in January 2021. The sampling strategy involved
identifying tree species and collecting various samples from each tree.
Briefly, 5 g of bark, bark-exuded gum, pods, and flowers of Algarrobo
(Neltuma chilensis (Molina) C.E.Hughes & G.P.Lewis), Tamarugo
(Strombocarpa tamarugo (Phil) C.E.Hughes & G.P.Lewis) and Chañar
(Geoffroea decorticans were collected. A total of 22 samples were
collected in aseptic conditions and transferred to tubes containing 10
mL of selective enrichment medium composed of YNB (Yeast Nitrogen Base)
supplemented with 2% (w/v) glucose and 4% (v/v) ethanol. This medium
was specifically designed to exclusively select for ethanol-tolerant
yeast species (Villarreal et al. , 2022). The tubes were incubated
at 25°C without agitation for a period of 14 days. After incubation, 100
µl aliquots were spread onto yeast-extract peptone dextrose (YPD) agar
plates containing chloramphenicol (20 µg ml−1) and incubated at 25°C
until yeast colonies emerged. Representative colonies of each distinct
morphotype were purified through streak inoculation on YPD agar for
further characterization. To ensure long-term preservation, yeast
cultures were stored at -80°C in a broth culture supplemented with 20%
(w/v) glycerol.