Honglin Li,1 Shebin Zhang,2 Qiwei Li,2 Yurong Shu,3 Song Li,2 Bo Wu,4 Zhenjie Xu2 

1Department of Clinical Laboratory, Mianyang Central Hospital, School of Medicine, University of Electronic Science and Technology of China, Mianyang, 621000, People’s Republic of China; 2Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, 510120, People’s Republic of China; 3Department of Clinical Laboratory, South China Agricultural University Hospital, Guangzhou, 510000, People’s Republic of China; 4Department of Transfusion, The Fourth Affiliated Hospital, Guangzhou Medical University, Guangzhou, 511300, People’s Republic of China

Correspondence: Zhenjie Xu, Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine, 111 Dade Road, Yuexiu District, Guangzhou, 510120, People’s Republic of China, Tel/Fax +86-02039318325, Email 15989085911@163.com Bo Wu, Department of Transfusion, The Fourth Affiliated Hospital, Guangzhou Medical University, Guangzhou, 511300, People’s Republic of China, Tel/Fax +86-02062287542, Email meteor.bullet@163.com

Purpose: It is now understood that the primary challenges in treating Pseudomonas aeruginosa (P. aeruginosa) infections are the secretion of multiple virulence factors, the formation of biofilm, and the emergence of multi-drug resistance. Small regulatory RNAs (sRNAs) and quorum sensing (QS) play an important role in regulating bacterial biofilms and multiple virulence factors, presenting potential targets for novel anti-P. aeruginosa therapies. Yinqiao Powder has demonstrated inhibitory activity against various bacteria and viruses. The objective of this study was to elucidate the precise mechanism of Yinqiao Powder’s impact on P. aeruginosa virulence and to ascertain its clinical utility.
Methods: First, the effects of Yinqiao Powder on various virulence factors of P. aeruginosa were assessed through virulence phenotype experiments, including biofilm formation assay, pyocyanin production assay, rhamnolipid assay, and motility assay. Then, a cytotoxicity assay was used to evaluate the effect of P. aeruginosa treated by Yinqiao Powder on cells. Finally, an RT-qPCR assay was used to detect the effects of Yinqiao Powder on QS system and virulence-related gene expression.
Results: This study revealed that sub-minimum inhibitory concentration (sub-MIC) levels of Yinqiao Powder significantly inhibit biofilm formation, swarming motility, pyocyanin and rhamnolipid production in a dose-dependent manner. The cytotoxicity assay also confirmed that Yinqiao Powder weakened the cytotoxicity of P. aeruginosa. Furthermore, Yinqiao Powder was found to modulate the P. aeruginosa sRNA-QS-virulence network. Specifically, it repressed the lasI, the rhlI, and sRNA P27 while upregulating sRNA PhrD. Additionally, the phzA and pqsA genes, associated with pyocyanin and rhamnolipid/biofilm regulation, respectively, were repressed by Yinqiao Powder.
Conclusion: Yinqiao Powder effectively inhibits QS system-related regulatory genes, sRNAs, biofilm formation, swarming motility, pyocyanin and rhamnolipid production at specific concentrations. These results support the potential of Yinqiao Powder as a quorum-sensing inhibitor.

Keywords: Yinqiao Powder, quorum sensing, sRNA, biofilm formation, virulence, Pseudomonas aeruginosa