Barbara Salerno ¹ Matteo Cornaggia ¹ Raffaella Sabatino ² Andrea Di Cesare ² Claudio Mantovani ¹ Lisa Barco ¹ Benedetta Cordioli ¹ Luca Bano ¹ Carmen Losasso ^1*

• ¹ Food Safety/Risk Analysis and Public Health, Experimental Zooprophylactic Institute of the Venezie (IZSVe), Legnaro, Italy
• ² CNR-IRSA,, Verbania, Italy

Farms are significant hotspots for the dissemination of antibiotic resistance into the environment and directly to humans. This underscores the need for comprehensive strategies to reduce the prevalence of antibiotic-resistant bacteria and genes (ARGs) within farms. A guideline focused on "best practices for farming" to reduce the spread of antibiotic resistance has been developed and provided to the owners of selected dairy farms (N=10) and veterinarians. This guideline focuses on the prudent use of antibiotics, the adoption of selective therapy for animals, and the implementation of hygienic and immune-prophylactic practices. Fecal samples were randomly collected from animals across various breeding categories, including lactating cows, dry cows, and calves, on the selected farms. These samples were taken both before and after the implementation of the farming guidelines. The presence of selected ARGs was initially screened using end-point PCR. Positive ARGs (blaTEM, ermB, sul2, and tetA) were then quantified using digital droplet PCRCodice campo modificato and expressed in relative terms by dividing the copy number of the ARGs by the copy number of the 16S rRNA gene. The ARG abundances in the samples collected in 2022 were higher in the lactating cow category. Interestingly, although the overall amount of antibiotics administered (based on data from veterinary prescriptions in the sampled farms) was similar in both sampling campaigns, the total abundance of the selected ARGs, as well as individual ARGs (specifically blaTEM and tetA), significantly decreased after the adoption of the good farming practices. This result underscores the importance of the prudent use of antibiotics and represents an initial step toward addressing the spread of antibiotic resistance in farm environments. Additionally, it strengthens previous findings, highlighting that the lactating cow category is a crucial point in farming where efforts should be focused to reduce the abundance of ARGs.