Dose-Response Study of a Fenugreek-Based Antibiotic Alternative in Bábolna Tetra-SL Chicks (1-42 Days Old) with Mixed Bacterial Infections

Adam Kerek^1,2*Abel Szabo³Peter Ferenc Dobra⁴Krisztina Bardos^2,5Bettina Paszerbovics⁶Zsofia Bata⁷Viviana Molnar-Nagy⁷Akos Jerzsele^2,3László Ózsvári^2,5

• ¹University of Veterinary Medicine Budapest, Budapest, Hungary
• ²National Laboratory of Infectious Animal Diseases, Antimicrobial Resistance, Veterinary Public 7 Health and Food Chain Safety, University of Veterinary Medicine Budapest, Budapest, Hungary
• ³Department of Pharmacology and Toxicology, University of Veterinary Medicine, Budapest, Hungary
• ⁴Department of Pathology, University of Veterinary Medicine Budapest, Budapest, Hungary
• ⁵Department of Veterinary Forensics and Economics, Institute of Economics and Biostatistics, University of Veterinary Medicine, Budapest, Hungary
• ⁶Department of Biostatistics, Institute of Economics and Biostatistics, University of Veterinary Medicine, budapest, Hungary
• ⁷Dr. Bata Zrt, ócsa, Hungary

Combating antimicrobial resistance is one of the most pressing public health challenges of our time. The rapid spread of resistant, zoonotic bacterial strains in livestock farming is increasingly raising concerns about the need to reduce antibiotic use. Because of this, there is an urgent need for safe and effective alternatives in animal husbandry. This study aimed to perform an in vivo the dose-response analysis of fenugreek (Trigonella foenum-graecum), as a plant-based antibiotic alternative feed supplement in Bábolna Tetra-SL chicks (1-42 days old) with a 1:1 sex ratio. A total of 270 chicks were randomly assigned to 18 groups (15 birds per group) and subjected to six different treatment groups in three replicates: fenugreek at 1×, 10×, and 100× doses, an antibiotic-treated group (enrofloxacin), a positive control group (infection only), and a negative control group (no infection or treatment). The infection was induced using mixed Salmonella Enteritidis and Escherichia coli, administered via gavage on days 3 and 4 of life. The birds were monitored for clinical symptoms, body weight, feed intake, and Salmonella shedding through cloacal swab samples. Statistical analyses included mixedeffect logistic regression for mortality, mixed-effect linear models for weight gain, two-way ANOVA for feed efficiency, and random effects continuation ratio models for Salmonella isolation. Significant interactions for Group:Day and Sex:Day in weight gain were identified (p < 0.0001 for both). Additionally, the 1× dose group showed significantly reduced Salmonella shedding compared to the positive control group on day 33 (p = 0.0031). The low-dose group (1×) demonstrated the most promising results, showing a 63% reduction in Salmonella shedding on day 10 and 31% on day 17. This group exhibited the fewest clinical symptoms, no diarrhea, and the lowest individual and specific feed intake up to day 24. The findings of this study suggest that low-dose fenugreek supplementation could be a viable strategy for reducing Salmonella shedding in poultry, potentially contributing to reduced antibiotic use in poultry farming and thus playing a role in the global effort to combat antimicrobial resistance. Future research will involve large-scale industrial trials and next-generation sequencing to evaluate the additive's impact on gut microbiota composition.