Citation: Rosales-López, C.; Vargas-López, A.; Monge-Artavia, M.; Rojas-Chaves, M. Evaluation of Conditions to Improve Biomass Production by Submerged Culture of Ganoderma sp.. Microorganisms 2022, 10, 1404. https://doi.org/10.3390/ microorganisms10071404 Academic Editor: Edward A. Bayer Received: 19 May 2022 Accepted: 2 July 2022 Published: 12 July 2022 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). microorganisms Article Evaluation of Conditions to Improve Biomass Production by Submerged Culture of Ganoderma sp. Catalina Rosales-López * ,† , Alejandro Vargas-López , Mariana Monge-Artavia and Miguel Rojas-Chaves Centro de Investigación en Biotecnología, Instituto Tecnológico de Costa Rica (ITCR), Cartago 159-7050, Costa Rica; alevalo.1392@gmail.com (A.V.-L.); marige97@gmail.com (M.M.-A.); mirojas@itcr.ac.cr (M.R.-C.) * Correspondence: crosales@itcr.ac.cr; Tel.: +506-2550-9303 † PhD. Doctorado en Ciencias Naturales para el Desarrollo (DOCINADE) ITCR-UNA-UNED, Costa Rica. Abstract: In the present investigation, the conditions for in vitro submerged culture of a native strain of Ganoderma sp. were evaluated. Different culture medium ingredients, inoculum concentrations, inoculation methods, configuration, and airflows were evaluated to improve biomass production. The addition of thiamine and olive oil to the culture medium increased biomass production, as well as inoculating 6.6 g/L since there are no significant differences in biomass growth according to inoculum origin (pre-inoculum, discs or with spores). The best configuration of the 3 L stirred tank bioreactor was using three impellers and a porous air diffuser of 0.25 volume per volume per minute (vvm), the dry biomass concentration was 22.6 g/L after 12 days of cultivation at 30 ◦ C, much higher than other investigations. This study provides relevant information for pilot-scale production of this fungus for future secondary metabolites. The culture medium was optimized, and it was defined that the concentration and origin of the inoculum did not influence the growth of Biomass, but the aeration and the configuration of the system allowed the establishment of protocols for the cultivation of Ganoderma sp. Keywords: Ganoderma; culture medium; bioreactor; biomass; inoculum; secondary metabolites 1. Introduction The genus Ganoderma contains many large species of bracket fungi belonging to the phylum Basidiomycota. Most of the species are saprophytes and grow on decomposing trunks [1]. Over 250 species of Ganoderma have been described worldwide, most of them classified based on pleomorphic characteristics [2]. In Costa Rica, nine species of this genus have been identified morphologically: G. australe, G. amazonense, G. dorsale, G. longistipi- tatum, G. oerstedii, G. orbiforme, G. perzonatum, G. resinaceum, and G. stipitatum [3,4]. The main taxonomic characteristic of this genus is the double-walled basidiospore. The inner wall is thick and yellowish-brown with numerous endosporic projections, and the external wall is thin, smooth, and hyaline [5]. Further characterization of Ganoderma is based on its microstructures; a molecular analysis is difficult because of the lack of gene sequences from neotropical collection zones. Ganoderma is recognized for its medicinal value. It is used to treat diseases, including gastric ulcers, chronic hepatitis, hypertension, nephritis, asthma, arthritis, bronchitis, in- somnia, cancer, diabetes, and anorexia [6]. Among the many bioactive compounds present are steroids, lignins, lectins, ganomycins, vitamins, nucleosides, nucleotides, alkaloids, amino acids, polysaccharides, and triterpenes [7]. Special attention has been given to the last two groups of compounds mentioned due to their effects on conditions relevant to human health. Ganoderic acids are a group of oxygenated triterpenes derived from lanosterol, which have been isolated from fruiting bodies, mycelium, and spores of Ganoderma sp. This family of compounds is numerous due to the possible substitutions on carbons 3, 7, 11, 12, 15, 22, Microorganisms 2022, 10, 1404. https://doi.org/10.3390/microorganisms10071404 https://www.mdpi.com/journal/microorganisms