Background: Avian pathogenic Escherichia coli (APEC) is a leading cause of colibacillosis in poultry. Piper betle L. is a medicinal plant rich in bioactive compounds including hydroxychavicol that possess potent antibacterial activity. This study aimed to investigate the efficacy of a P. betle L. leaf nanoemulsion (NEPE) and hydroxychavicol against multidrug-resistant APEC isolates. Methods: In vitro and in silico analysis of NEPE and hydroxychavicol against APEC were determined. Results: The nanoemulsion exhibited potent antibacterial activity, with MIC and MBC values of 0.06–0.25% v/v and 0.125–0.25% v/v, respectively. The MIC and MBC values of hydroxychavicol against isolates ranged from 0.25 to 1.0 mg/mL. A time–kill assays revealed rapid bactericidal effects of both compounds, achieving a ≥3-log reduction within 4 h at 4 × MIC. Scanning electron microscopy demonstrated that APEC cells treated with hydroxychavicol exhibited filamentous cells with incomplete septa. Molecular docking and dynamics simulations of hydroxychavicol against APEC cell division proteins were investigated. According to the binding energy, hydroxychavicol exhibited the highest affinity with ZapE, FtsW, FtsX, FtsZ, and FtsA, respectively. However, the FtsA protein showed the least protein conformational change throughout the 5000 ns simulation, reflecting a highly stable conformation. Conclusions: These confirm the potential stability of protein and ligand, as supported by molecular dynamics simulation. The results suggested the potential of NEPE and hydroxychavicol, which may have promising antibacterial potential that can be used to inhibit APEC growth.

Non-absorbable sutures provide a site for bacterial attachment and increase the risk of surgical site infections. An alternative prevention of infections requires plant-extract coatings on sutures. The objectives of this study were to develop P. betle leaf extract-coated non-absorbable sutures and to investigate their activities on staphylococci. P. betle leaves were extracted and analyzed for the phytochemicals. P. betle extract was coated on sutures, including polyester and polypropylene. The stability of hydroxychavicol on coated sutures was evaluated. Four treatments were designed, including (1) uncoated, (2) antibiotic/extract-free-coated, (3) extract-coated, and (4) gentamicin-coated sutures. Each treatment was tested for antibacterial, antibiofilm, and anti-adhesion activities on Staphylococcus aureus and Staphylococcus pseudintermedius. In addition, the cytotoxicity of extract-coated sutures was tested. Analysis of the extract identified hydroxychavicol (40.07%) as the primary phytochemical. Stability tests indicated higher hydroxychavicol on Day 1 of extract-coated polyester compared to polypropylene, and the levels decreased on the subsequent days (p < 0.05). Antibacterial activity of extract-coated polyester showed antibacterial effects during the experiment period (5.16 ± 2.35 mm), while polypropylene showed no effectiveness. Additionally, biofilm inhibition was found to be 36.63 ± 27.08% and 37.34 ± 26.98% in tested staphylococci for extract-coated polyester and polypropylene, respectively. Anti-adhesion showed that the extract-coated sutures had a higher ability to decrease tested bacteria attachment (56.25–60.42% living cell reduction). The cytotoxicity study revealed that extract-coated sutures of ≤1.5 mg/1.5 cm had a 99% survival rate. The findings indicate that the coated sutures showed antibacterial, antibiofilm, and anti-adhesion effects against staphylococci causing canine skin infections and might lead to alternative surgical use in veterinary medicine.

Background: Avian pathogenic Escherichia coli (APEC) contributes substantially to colibacillosis outbreaks in chickens. Because APEC cells readily attach to surfaces and develop biofilms, they pose a notable hazard to poultry production and food safety. This study investigated the antibiofilm and anti-adhesion activities of deep eutectic solvent-based emulsion containing Piper betle L. extract (DEPE) and hydroxychavicol, a pure compound isolated from P. betle leaves against APEC. Methods: Antibiofilm and anti-adhesion activities of DEPE and hydroxychavicol against APEC were investigated. Molecular docking and dynamics simulation of DEPE and hydroxychavicol was conducted. In addition, anti-adhesion activity of DEPE on chicken meat during storage was evaluated. Results: DEPE and hydroxychavicol significantly inhibited biofilm formation at sub-MIC, with DEPE achieving up to 80% inhibition and hydroxychavicol up to 69%. At 8 × MIC, DEPE and hydroxychavicol diminished the viability of both early and established biofilms. Furthermore, DEPE and hydroxychavicol reduced APEC adhesion on the surface as observed by SEM. In silico analyses demonstrated the stable binding of hydroxychavicol to adhesion-related proteins, particularly EcpA and FimH, suggesting a possible mechanism for its anti-adhesion activity. At day 5, DEPE at 4 × MIC significantly reduced 63% bacterial adhesion to chicken meat surfaces during storage, while maintaining the meat’s color. Conclusions: These findings indicate that DEPE and hydroxychavicol are promising candidates for limiting APEC biofilm formation and surface attachment and may serve as alternative antibacterial agents in poultry-related food safety applications.