Vibrio parahaemolyticus (V. parahaemolyticus) is a gram-negative species that mainly originates from seafood such as fish, shrimp and crabs (Drake et al., 2007). In pursuit of the taste of seafood, people directly consume unprocessed seafood, which increases the likelihood of being infected with V. parahaemolyticus. Symptoms such as diarrhea, nausea, vomiting, and fever usually occur after infection (Li et al., 2019). The Food and Agriculture Organization of the United Nations (FAO) and World Health Organization (WHO) report that V. parahaemolyticus cases were geographically confined to Japan until the late 1960s, but sporadic cases and outbreaks have been reported in countries in different states since 1969 (FAO and WHO, 2021). This indicated that V. parahaemolyticus has emerged as the most representative pathogen in seafood and evolved into a major concern in the field of global public health. Therefore, the exploration of a strategy that can effectively inhibit V. parahaemolyticus is of great significance in safeguarding human health and reducing the risk of foodborne diseases.

The biofilm is a community with certain structures and functions, composed of bacteria and their secreted extracellular polymeric substances (EPS), including polysaccharides, proteins, and nucleic acids (Song et al., 2017). Biofilm development is a dynamic process in which EPS forms the scaffold of the biofilm matrix, mediates surface attachment, and maintains the overall structure of the biofilm (Abiola et al., 2025). The structure of the biofilm makes it more difficult for antibacterial substances to come into contact with bacteria, which is beneficial for bacteria to grow in adverse environments. Research has proven that the biofilm community is more difficult to remove compared with planktonic V. parahaemolyticus (Yin et al., 2019). Since many types of processing equipment are used to process a variety of products, it is very likely to cause cross-contamination risks if biofilms cannot be effectively removed (Ahmed et al., 2018; Mok et al., 2021). This can lead to serious food safety issues and economic losses for enterprises. Therefore, it is of great significance to prevent and control V. parahaemolyticus and the formation of its biofilms for improving food safety.

Linalool is a naturally occurring monoterpenoid found in the essential oils of more than 400 plant species (An et al., 2021). In recent years, it has drawn increasing interest for its potent and broad-spectrum antimicrobial effects, especially in the field of alternative synthetic bacteriostatic agents. According to existing studies, linalool has an inhibitory effect on gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis), gram-negative bacteria (Escherichia coli and Shigella sonnei), and fungi (Fusarium oxysporum) (He et al., 2024; Randrianarivelo et al., 2009). It was found that the formation of V. parahaemolyticus biofilm could be inhibited by linalool at the minimum inhibitory concentration (MIC) and 1/2 MIC in our previous study (Ren et al., 2024). However, current research on linalool's inhibition of V. parahaemolyticus are largely unknown, and its potential regulatory mechanisms require further investigation. In this study, we comprehensively explored the inhibitory mechanism of linalool against biofilms using transcriptomics. Moreover, we evaluated the effectiveness of linalool in inhibiting biofilm formation on the surfaces of shrimp and shrimp processing materials.

In this study, we first determined the minimum concentration of linalool for inhibiting the biofilm formation of V. parahaemolyticus. Then, the inhibitory effect of sub-minimal inhibitory concentrations (sub-MICs) of linalool on the biofilm of V. parahaemolyticus was evaluated using crystal violet assay and field emission scanning electron microscopy (FE-SEM). The inhibitory mechanism of linalool on the biofilm was explored by measuring the content of EPS, motility, hydrophobicity, auto-aggregation, signaling molecule Autoinducer-2 (AI-2), and biofilm metabolic activity. Subsequently, transcriptomics was used to further analyze the effects of linalool on motility, signaling molecule AI-2, and biofilm metabolic activity, and to further elucidate the mechanism of linalool in inhibiting biofilm formation. In addition, we evaluated the inhibitory effect of linalool on the formation of V. parahaemolyticus biofilms on the surfaces of shrimp and shrimp processing materials. Therefore, this study aims to systematically evaluate the inhibitory effect of linalool on the biofilm formation of V. parahaemolyticus and elucidate its underlying mechanism.