Citation: Digut , ˘ a, C.F.; Mihai, C.; Toma, R.C.; Cîmpeanu, C.; Matei, F. In Vitro Assessment of Yeasts Strains with Probiotic Attributes for Aquaculture Use. Foods 2023, 12, 124. https://doi.org/10.3390/ foods12010124 Academic Editors: Arun K. Bhunia, Joana S. Amaral, Derek V. Byrne, Theodoros Varzakas, Esther Sendra and Benu P. Adhikari Received: 10 November 2022 Revised: 18 December 2022 Accepted: 22 December 2022 Published: 26 December 2022 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/). foods Article In Vitro Assessment of Yeasts Strains with Probiotic Attributes for Aquaculture Use Camelia Filofteia Digut , ă 1,† , Constant , a Mihai 2, *, Radu Cristian Toma 1,† , Carmen Cîmpeanu 2 and Florentina Matei 1 1 Faculty of Biotechnologies, University of Agronomic Sciences and Veterinary Medicine of Bucharest, 59, Mărăs , ti Blvd., District 1, 011464 Bucharest, Romania 2 Faculty of Land Reclamation and Environmental Engineering, University of Agronomic Sciences and Veterinary Medicine of Bucharest, 59, Mărăs , ti Blvd., District 1, 011464 Bucharest, Romania * Correspondence: mihai_tania@yahoo.com † These authors contributed equally to this work. Abstract: This study aimed to investigate in vitro the probiotic potential of three yeasts strains (BB06, OBT05, and MT07) isolated from agro-food natural sources. Screening was performed, including several functional, technological, and safety aspects of the yeast strains, in comparison to a reference Saccharomyces boulardii, to identify the ones with suitable probiotic attributes in aquaculture. The yeast strains were identified by 5.8S rDNA-ITS region sequencing as Metschnikowia pulcherrima OBT05, Saccharomyces cerevisiae BB06, and Torulaspora delbrueckii MT07. All yeast strains were tolerant to different temperatures, sodium chloride concentrations, and wide pH ranges. S. cerevisiae BB06 showed a strong and broad antagonistic activity. Moreover, the S. cerevisiae strain exhibited a high auto-aggregation ability (92.08 ± 1.49%) and good surface hydrophobicity to hexane as a solvent (53.43%). All of the yeast strains have excellent antioxidant properties (>55%). The high survival rate in the gastrointestinal tract (GIT) can promote yeast isolates as probiotics. All yeast strains presented a resistance pattern to the antibacterial antibiotics. Non-hemolytic activity was detected. Furthermore, freeze-drying with cryoprotective agents maintained a high survival rate of yeast strains, in the range of 74.95–97.85%. According to the results obtained, the S. cerevisiae BB06 strain was found to have valuable probiotic traits. Keywords: Saccharomyces; non-Saccharomyces; probiotic properties; safety issue 1. Introduction Fish production in farm aquaculture has increased significantly in the past years. Nevertheless, semi-intensive or intensive production systems have exposed the fish to prolonged stressful conditions with a negative impact on their well-being by suppressing their immunity, thereby increasing the fish’s susceptibility to pathogens, all of which can translate into low performance [1–3]. However, the uncontrolled and prolonged application of antibacterial antibiotics to treat bacterial diseases that occur most frequently in the fish leads to significant changes in the microflora of the fish, their accumulation in the tissues [4,5], a decrease in immunity, and the appearance of antibiotic-resistant bacteria, which represent a serious threat [6–9]. The consumption of contaminated fish products could have a negative impact on human health [10,11]. Currently, the use of probiotics has been promoted as a viable, safe alternative method for sustainable aquaculture [5,12–19]. Probiotics are viable microorganisms which, when administered in adequate amounts, provide health benefits to the host [20,21]. Lactic bacteria and Bacillus spp. have been the most researched probiotics and widely administered in aquaculture via diet or water, either by themselves, as a mix of different species, or combined with other ingredients such as prebiotics to improve performance and health status of fishes [14,19,22–27]. However, the possible risk to transfer antibiotic resistance genes through bacterial probiotics to Foods 2023, 12, 124. https://doi.org/10.3390/foods12010124 https://www.mdpi.com/journal/foods