Metabolomic insights into the inhibition mechanism of methyl N-methylanthranilate: A novel quorum sensing inhibitor and antibiofilm agent against Pseudomonas aeruginosa.

The quorum sensing (QS) inhibition effect of methyl N-methylanthranilate (MMA) from Pericarpium Citri Reticulatae Chachiensis against foodborne pathogen Pseudomonas aeruginosa was reported for the first time. MMA effectively attenuated QS related virulence factors production and biofilm formation, while suppressed expression of a dozen of QS related genes. Untargeted LC-MS metabolomics revealed 108 significantly altered metabolites after MMA treatment. They indicated that MMA addition reduced the efficiency of TCA cycle and antioxidant systems, disturbed amino acid and nucleotide metabolism, increased unsaturated fatty acid and decreased peptidoglycan components, which might ultimately attenuate P. aeruginosa pathogenicity and restrain biofilm formation. Physiological characterization confirmed the compromised membrane integrity and increased intracellular oxidative stress after MMA treatment. Furthermore, metabolomics data implied that MMA inhibition on QS might exert through disrupting QS autoinducer PQS biosynthesis, which was supported by molecular docking. Our data indicated that MMA could be used as a novel QS inhibitor and anti-biofilm agent to improve food safety. It also provided new insight in the possible underlying inhibition mechanism of MMA and the response of P. aeruginosa to MMA.

Probiotic Potential and Cholesterol-Lowering Capabilities of Bacterial Strains Isolated from Pericarpium Citri Reticulatae 'Chachiensis'.

Pericarpium Citri Reticulatae 'Chachiensis' (PCR-Chachiensis), the pericarps of Citri Reticulatae Blanco cv. Chachiensis, is a food condiment and traditional medicine in southeast and eastern Asia. Its rich and various bacterial community awaits exploration. The present study is the first report on probiotic screening and characterization of bacteria from PCR-Chachiensis. Based on 64 culturable bacterial isolates, 8 strains were screened out to have great survival in the simulated gastrointestinal stressful condition, being nonhemolytic and without biogenic amine formation. They were identified by 16S rRNA gene sequencing as two Bacillus, three Lactobacillus, and three strains from Bacillales. Their probiotic properties, cholesterol-lowering potential and carbohydrate utilization capability were further investigated. Though these eight strains all displayed distinct cholesterol removal potential, Bacillus licheniformis N17-02 showed both remarkable cholesterol removal capability and presence of bile salt hydrolase gene, as well as possessing most of the desirable probiotic attributes. Thus, it could be a good probiotic candidate with hypocholesterolemic potential. Bacillus megaterium N17-12 displayed the widest carbohydrate utilization profile and the strongest antimicrobial activity. Hence, it was promising to be used as a probiotic in a host and as a fermentation starter in fermented food or feed.

The central bacterial community in Pericarpium Citri Reticulatae 'Chachiensis'.

The dried and aged pericarps of Citri Reticulatae are condiments and medicinal products in southeast and eastern Asia for hundreds of years, among which Pericarpium Citri Reticulatae 'Chachiensis' (PCR-C) is the premium one with obvious health benefits. In order to explore the microbiota in PCR-C and their relationship with the chemical components during aging, culture-independent methods were applied to investigate PCR-C microbiota for the first time. Here in different PCR-C samples, 16S rRNA gene amplicon sequencing revealed common central bacterial community, which were absent or only accounted for small proportion in fresh pericarps or jute bag controls. Bacillus and Lactococcus were the top two dominant genera in PCR-C with acidic pH (4.06-4.51) and low moisture (11.48%-19.13%). Several OTUs were found to closely relate with specific compositions in essential oils and phenolics, such as d-limonene and nobiletin, which contributed to PCR-C flavor and quality. As the first study to reveal the central bacterial communities in PCR-C, it provides new insights to improve the quality and aging process of traditional Pericarpium Citri Reticulatae, and lays foundation for functional characterization of the microbes within.