Hydroquinine Possesses Antibacterial Activity, and at Half the MIC, Induces the Overexpression of RND-Type Efflux Pumps Using Multiplex Digital PCR in Pseudomonas aeruginosa

Abstract

Hydroquinine is an organic compound that is closely related to quinine-derivative drugs and contains anti-malarial and anti-arrhythmia activities. It has been also found in abundance in some natural extracts that possess antibacterial properties. However, there is little evidence demonstrating the antibacterial effect of hydroquinine. Therefore, we aimed to investigate the antibacterial properties of hydroquinine using broth microdilution methods. In addition, we evaluated the transcriptional responses of P. aeruginosa to hydroquinine-induced stress using RNA sequencing with transcriptomic analysis and validated the results using PCR-based methods. The MIC and MBC values of hydroquinine against all eight bacterial strains investigated ranged from 650 to 2500 and from 1250 to 5000 µg/mL, respectively. Transcriptomic analysis demonstrated that RND efflux pump transcripts were overexpressed (4.90–9.47 Log2 fold change). Using mRT-dPCR and RT-qPCR, we identified that mRNA levels of mexD and mexY genes were overexpressed in response to just half the MIC of hydroquinine in P. aeruginosa. In conclusion, we uncover the antimicrobial potential of hydroquinine as well as identify changes in gene expression that may contribute to bacterial resistance. Further work will be required to explore the efficacy and potential use of hydroquinine in the clinic.

High-Throughput Transcriptomic Profiling Reveals the Inhibitory Effect of Hydroquinine on Virulence Factors in Pseudomonas aeruginosa

“…No. CM0131, Oxoid, Basingstoke, UK) at 35 ± 2 °C for 18–24 h [ 31 ]. For inoculum preparation, the inoculum was adjusted in exponential growth phases to achieve turbidity equivalent to a 0.5 McFarland standard (1–2 × 10 8 CFU/mL) before being used in each experiment.…”

“…Fresh hydroquinine was made before use by dissolving in 50% DMSO and Tween 80 (1:1) solution. The hydroquinine solution was syringe-filtered (through a 0.2 μm pore size) and kept away from the light source [ 31 ].…”

“…No. 74004, QIAGEN, Hilden, Germany) as previously described by Rattanachak et al (2022) [ 31 ]. Total RNA samples were also isolated using RNase-Free DNase Set (Cat.…”

“…Each culture was incubated at 35 ± 2 °C for 1 h, with shaking at 200 rpm. The bacterial pellets were collected by centrifugation at 5000 rpm and 4 °C for 10 min [ 31 ]. The total RNA from each sample was then extracted.…”

“…Hydroquinine, an alkaloid organic compound found in particular natural products [ 28 , 29 , 30 ], was recently reported to inhibit and kill drug-sensitive (DS) and MDR strains of P. aeruginosa [ 31 ]. In addition, other alkaloid compounds have been reported to possess antimicrobial properties and anti-virulence factors in both Gram-positive and Gram-negative bacteria, such as inhibition of QS signaling and pigment production in P. aeruginosa [ 26 ], inhibiting biofilm formation in E. coli and P. aeruginosa [ 24 ] and inhibiting virulence gene expression in S. aureus [ 32 ].…”