Antibacterial and antibiofilm properties of 1,3,4-oxadiazoles against multidrug-resistant Pseudomonas aeruginosa isolated from burn infections – an in vitro study

Abstract

Pseudomonas aeruginosa is a major causative agent of burn wound infections, frequently characterised by multidrug resistance (MDR) and robust biofilm formation, which limit the efficacy of conventional antibiotics. 1,3,4-Oxadiazole derivatives have emerged as potential antimicrobial and antibiofilm agents; however, comprehensive evaluations against clinical MDR isolates from burn patients remain limited. The objective of this study was to investigate the in vitro antibacterial and antibiofilm activities of a synthesised 1,3,4-oxadiazole compound against clinical MDR P. aeruginosa isolates from burn wounds. In addition, the study sought to assess its synergistic potential with gentamicin and imipenem, and to examine its effects on biofilm-associated gene expression. A total of 75 non-duplicate clinical P. aeruginosa isolates were collected from patients admitted to educational hospitals in Hamedan, Iran, including 49 burn-wound isolates and 26 respiratory isolates. From the initial burn-wound collection, nine isolates were selected for detailed antimicrobial and antibiofilm analyses based on their clinical relevance and biofilm phenotype; these comprised seven MDR and two non-MDR isolates, all demonstrating strong or intermediate biofilm formation. Antimicrobial susceptibility and biofilm formation were assessed using the disk diffusion and microtiter crystal violet assays. MIC, MBC, MBIC, and MBEC values were determined for 1,3,4-oxadiazole, gentamicin, and imipenem. Checkerboard assays were performed to evaluate the combined effects of 1,3,4-oxadiazole with gentamicin or imipenem against planktonic and biofilm-associated cells. The effects of sub-inhibitory concentrations of 1,3,4-oxadiazole on lasR and algD expression were evaluated using real-time PCR. Among the 75 clinical isolates, 57 (76%) were classified as MDR and 74 (98.7%) produced biofilms. From the 49 burn-wound-derived isolates, nine were selected for downstream analysis, including seven MDR and two non-MDR isolates, all exhibiting strong or intermediate biofilm formation. The geometric mean MIC, MBC, MBIC, and MBEC values of 1,3,4-oxadiazole against these selected burn isolates were 20.54, 41.21, 52.56, and 105.11 µg/mL, respectively. In the checkerboard assays, 1,3,4-oxadiazole showed enhanced activity in combination with gentamicin and imipenem. For planktonic cells, the gentamicin–oxadiazole combination yielded geometric mean FIC and FBC indices of 0.54 and 0.50, respectively, whereas the imipenem–oxadiazole combination produced corresponding values of 0.72 and 0.62. Synergistic interactions were observed in 3 of the 9 selected isolates (33.3%). Against biofilm-associated cells, the gentamicin–oxadiazole combination resulted in geometric mean FBIC and FBEC indices of 0.55 and 0.59, while the imipenem–oxadiazole combination showed values of 0.62 and 0.61, indicating a similar improvement in antibiofilm activity. Exposure to sub-inhibitory concentrations of 1,3,4-oxadiazole resulted in concentration-dependent downregulation of algD and lasR expression. 1,3,4-Oxadiazole has been demonstrated to possess potent antibacterial and antibiofilm properties against MDR and biofilm-forming P. aeruginosa from burn wounds. In addition, it has been shown to enhance the efficacy of conventional antibiotics and effectively downregulate key biofilm-associated genes. These findings lend support to the hypothesis that the substance under investigation has the potential to function as a promising alternative or adjunctive therapeutic strategy for the management of biofilm-related MDR P. aeruginosa infections in patients suffering from burns.