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In the present study, we screened for potential probiotic yeast that could survive under extreme frozen conditions. The antimicrobial and heat-stable properties of the isolated yeast strains <i>Saccharomyces boulardii</i> (<i>S. boulardii</i>) (<i>KT000032</i>, <i>KT000033</i>, <i>KT000034</i>, <i>KT000035</i>, <i>KT000036</i>, and <i>KT000037</i>) was analyzed and compared with commercial probiotic strains. The results revealed that the tested <i>S. boulardii KT000032</i> strain showed higher resistance to gastric enzymes (bile salts, pepsin, and pancreatic enzyme) at low pH, with broad antibiotic resistance. In addition, the strain also showed efficient auto-aggregation and co-aggregation abilities and efficient hydrophobicity in the in-vitro and in-vivo <i>C. elegens</i> gut model. Further, the KT000032 strain showed higher antimicrobial efficiency against 13 different enteropathogens and exhibited commensal relationships with five commercial probiotic strains. Besides, the bioactive compounds produced in the cell-free supernatant of probiotic yeast showed thermo-tolerance (95 °C for two hours). Furthermore, the thermo-stable property of the strains will facilitate their incorporation into ready-to-eat food products under extreme food processing conditions.