Whole transcriptome sequencing analysis of synergistic combinations of plant-based antimicrobials and zinc oxide nanoparticles against Campylobacter jejuni

The emergence of antibiotic resistance among animal farms impels the development of novel antimicrobials or strategies for agri-food production. The combinational use of agents to achieve a synergistic antimicrobial effect provides many advantages such as dosage reduction, shortened treatment time, and avoidance of antimicrobial resistance. In this study, we evaluated the killing efficacy of single agent or combinational use of three antimicrobials, including cinnamon oil, encapsulated curcumin and zinc oxide nanoparticles (ZnO NPs), against a leading foodborne pathogen Campylobacter jejuni. We then investigated the antimicrobial mechanism using whole transcriptome sequencing analysis (RNA-Seq). The single-agent treatment of cinnamon oil, encapsulated curcumin, or ZnO NPs showed a significant antimicrobial effect against C. jejuni by generating more than 8-log reduction within 3 h. The transcriptional signatures of C. jejuni in response to these agents varied, indicating that these agents shared distinct mechanisms of action and were likely to generate synergistic effects. Cinnamon oil affected the integrity of cell membrane, which might lead to an increase in cell permeability. Encapsulated curcumin and ZnO NPs disrupted bacterial outer membranes and cell membranes against the same membrane protein targets. The combinational use of these agents showed synergistic antimicrobial effects and distinct mechanisms of action compared to single treatment. The combination of cinnamon oil and encapsulated curcumin provoked the expression of cellular signaling, but repressed the chemotaxis-associated genes. The antimicrobial resistance associated genes showed a low expression level in the combination of encapsulated curcumin and ZnO NPs. The tri-combination treatment systematically overexpressed genes involved in the amino acid synthesis, protein translation, and membrane protein synthesis. This study provides new insights in combating Campylobacter with minimizing the development of antimicrobial resistance in long-term usage.