Abstract
Background:
Antibiotic resistance in Escherichia coli and Salmonella spp. poses a significant threat to poultry health and productivity. Overuse of antibiotics in poultry farming has led to the emergence of multidrug-resistant (MDR) pathogens, which have contributed to food safety concerns and limited treatment options. Natural antimicrobial alternatives, particularly herbal agricultural products such as purple onions and chives, have gained interest as sustainable solutions. Furthermore, fermentation with probiotic bacteria enhances the antimicrobial properties of botanical extracts, improving their efficacy against pathogenic bacteria.

Aim:
This study aimed to evaluate the antibacterial efficacy of fermented purple onion (FPO) and chive (FC) extracts against Escherichia coli and Salmonella spp., both causative agents of diarrhea, in broiler chickens. The research also compared their effectiveness with conventional antibiotics used in poultry farming.

Methods:
We fermented fresh purple onion and chive extracts using Lactobacillus plantarum and Bacillus subtilis for 72 hours under anaerobic conditions. Pathogenic E. coli (22) and Salmonella spp. (9) were isolated from 30 diarrheal chicken fecal samples and identified through biochemical tests, in vivo virulence testing in chicks, PCR detection of toxin genes (Stx1, Stx2, and Stn), and 16S rRNA sequencing. The antibacterial activity of the fermented extracts was evaluated using disk diffusion assays, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) tests. The bioactive compound content, including phenolics, flavonoids, and sulfur compounds, was quantified using an aluminum chloride colorimetric assay to assess their contribution to antimicrobial efficacy.

Results:
In vivo virulence assessment of chickens revealed that 4 E. coli strains (18.2%) and 5 Salmonella strains (26.3%) induced diarrhea. Of these, 2 E. coli strains (50%) harbored the Stx1 toxin gene, and all 5 Salmonella strains (100%) carried the Stn gene. 16S rRNA gene sequencing identified 2 E. coli strains and 4 Salmonella strains. All bacterial isolates exhibited MDR, particularly beta-lactam and macrolide antibiotics. Both fermented extracts exhibited significant antibacterial activity against MDR E. coli and Salmonella spp., with FC exhibiting superior efficacy compared to FPO. The inhibition zones of FC (12.5–17.6 mm) were significantly larger than those of FPO (10.1–16.8 mm), particularly against E. coli FG31-1 and Salmonella pullorum NCTC10705. The MIC and MBC values indicated that FC had a stronger bactericidal effect, requiring lower concentrations to inhibit and kill bacterial growth. FC contained higher levels of flavonoids and allicin, which were correlated with its enhanced antibacterial properties.

Conclusion:
FPO and FC, particularly FPO, show strong antibacterial potential as natural alternatives to antibiotics in poultry farming. Their ability to inhibit MDR pathogens suggests their role in reducing antibiotic dependency and improving poultry health. Further, in vivo studies and commercial formulation strategies are required to validate the practical application of these formulations in the poultry industry.

Key words: Antibiotic alternative, Chive, E. coli, Fermentation, Purple onion, Salmonella spp.