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Atractylodes chinensis is one of the most commonly used bulk herbs in East Asia; however, root rot can seriously affect its quality and yields. In contrast to chemical pesticides, biological control strategies are environmentally compatible and safe. For this study, 68 antagonistic bacterial strains were isolated from the rhizospheres of healthy Atractylodes chinensis. Strain SY42 exhibited the most potent fungicidal activities, with inhibition rates against F. oxysporum , F. solani , and F. redolens of 67.07 %, 63.40 % and 68.45 %, respectively. Through morphological observation and molecular characterization, strain SY42 was identified as Paenibacillus polymyxa. The volatile organic components (VOCs) produced by SY42 effectively inhibited the mycelial growth of pathogenic fungi through diffusion. SY42 significantly inhibited the germination of pathogenic fungal spores. Following co-culturing with SY42, the mycelium of the pathogenic fungus was deformed, folded, and even ruptured. SY42 could produce cellulases and proteases to degrade fungal cell walls. Pot experiments demonstrated the excellent biocontrol efficacy of SY42. This study revealed that P. polymyxa SY42 inhibited pathogenic fungi through multiple mechanisms, which verified its utility as a biocontrol agent for the control of A. chinensis root rot. • Paenibacillus polymyxa SY42 was an antagonistic bacteria isolated from the rhizosphere of healthy Atractylodes chinensis. • Strain SY42 could significantly inhibit the root rot pathogenic fungi of Atractylodes chinensis. • Volatile organic compounds (VOCs) produced by strain SY42 could inhibit pathogenic fungi. • Strain SY42 could effectively inhibit spore germination and germ tube growth of the pathogenic fungus. • Strain SY42 could produce a variety of hydrolytic enzymes and antimicrobial substances. [ABSTRACT FROM AUTHOR]