The development of efficient fungal vaccines is urgent for preventing life-threatening systemic fungal infections. In this study, we prepared a synthetic, cell-based fungal vaccine for preventing systemic fungal infections using synthetic biology techniques. The synthetic cell EmEAP1 was constructed by transforming theEscherichia colichassis using a de novo synthetic fragment encoding the protein mChEap1 that was composed of theE. coliOmpA peptide, the fluorescence protein mCherry, theCandida albicansadhesin Eap1, and the C-terminally transmembrane region. The EmEAP1 cells highly exposed the mChEap1 on the cell surface under IPTG induction. The fungal vaccine was then prepared by mixing the EmEAP1 cells with aluminum hydroxide gel and CpG. Fluorescence quantification revealed that the fungal vaccine was stable even after 112 days of storage. After immunization in mice, the vaccine resided in the lymph nodes, inducing the recruitment of CD11c+dendritic cells. Moreover, the vaccine strongly activated the CD4+T splenocytes and elicited high levels of anti-Eap1 IgG. By the prime-boost immunization, the vaccine prolonged the survival time of the mice infected by theC. albicanscells and attenuated fungal colonization together with inflammation in the kidneys. This study sheds light on the development of synthetic biology-based fungal vaccines for the prevention of life-threatening fungal infections.Keywords:fungal vaccine;systemic fungal infection;Candida albicans;adhesin;synthetic biology;protection
Staphylococcus aureus readily forms biofilms on host tissues and medical devices, enabling its persistence in chronic infections and resistance to antibiotic therapy. The accessory gene regulator (Agr) quorum sensing system plays a key role in regulating S. aureus biofilm formation. This study reveals the widely used fluoroquinolone antibiotic, ciprofloxacin, strongly stimulates biofilm formation in methicillin-resistant S. aureus , methicillin-sensitive S. aureus , and clinical isolates with diverse genetic backgrounds. Crystal violet staining indicated that ciprofloxacin induced a remarkable 12.46- to 15.19-fold increase in biofilm biomass. Confocal laser scanning microscopy revealed that ciprofloxacin induced denser biofilms. Phenotypic assays suggest that ciprofloxacin may enhance polysaccharide intercellular adhesin production, inhibit autolysis, and reduce proteolysis during the biofilm development, thus promoting initial adhesion and enhancing biofilm stability. Mechanistically, ciprofloxacin significantly alters the expression of various biofilm-related genes ( icaA , icaD , fnbA , fnbB , eap , emp ) and regulators ( agrA , saeR ). Gene knockout experiments revealed that deletion of agrC , rather than saeRS , abolishes the ciprofloxacin-induced enhancement of biofilm formation, underscoring the key role of agrC . Thermal shift assays showed ciprofloxacin binds purified AgrC protein, thereby inhibiting the Agr system. Molecular docking results further support the potential interaction between ciprofloxacin and AgrC. In summary, subinhibitory concentrations of ciprofloxacin stimulate S. aureus biofilm formation via an agrC -dependent pathway. This inductive effect may facilitate local infection establishment and bacterial persistence, ultimately leading to therapeutic failure.
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