Identificação molecular de leveduras não-Saccharomyces isoladas de arbóreas nativas do Brasil

Abstract

The bioprospection of fungal biodiversity is fundamental for the advancement of biotecnology, especially in the Search for non-conventional microorganisms, that may present superior phylogenetic characteristics under stress conditions and that remain poorly studied compared to comercial strains of Saccharomyces cerevisiae. The main objective of this study was the molecular identification of eight non-Saccharomyces yeast strains previously isolated from the phyllosphere of native Brazilian arboreal species, including jabuticaba, butiá, mulberry, loquat, hog plum tree, and gabiroba. The isolates deposited in the the Microbiological Collection of the Paraná Network (CMRP), a laboratory at the Federal Technological University of Paraná (UTFPR), Ponta Grossa campus, where they were reactivated in YPD culture medium for verification of isolate purity and analysis of macro and micromorphology. For the molecular study, genomic DNA extraction was first performed, followed by the amplification of conserved and viable regions of ribossomal DNA (rDNA) using primers V9G and LS266 for the ITS (Internal Transcribed Spacer) region and primers NL1 and NL4 for the D1/D2 region of the 28S rDNA. Taxonomic identification at the species level was achieved by Sanger sequencing of the purified amplicons, followed by smilarity analysis using BLAST on the NCBI (National Center for Biothecnology Information) database and confirmation through the construction of a multilocus phylogenetic tree. The analysis of BLAST allowed the identification of three distinct taxonomic groups, all with notable biotechnological potential: two strains of Lodderomyces elongisporus (100% identity), one strain of Meyerozyma caribbica (100% identity), and five strains belonging to the genus Torulaspora, which presented species-level divergence between the ITS (T. delbrueckii) and 28S (T. pretoriensis) markers, which was subsequently evaluated and confirmed in the multilocus phylogenetic tree as being T. pretoriensis. The identified species represent genetic resources with great potencial for industrial applications, including the prodution of enzymes (celulases and pectinases), bioenergy, and the modulation of the organoleptic profile in high-value fermentations, reforcing the importance of bioprospecting non-convencional microrganisms from native Brazilian arboreal species.