Abstract: Background and Objectives: Neonatal calf diarrhoea is one of the most serious problems faced livestock, causing great economic losses in the dairy industry all over the world. Several types of bacteria causing diarrhoea are resistance to the majority of antimicrobial agents’ which urgent need to search for new antimicrobial agents to treat this problem. This study aimed to evaluate the antibacterial efficacy of green Silver nanoparticles (Ag NPs) biosynthesis using natural honey against bacteria isolated from calf diarrhoea. Materials and Methods: A total of 290 faecal samples were collected from bovine calves suffering from diarrhoea at different governorates in Egypt; during the period from June 2017 to March, 2018. Bacteria were isolated from calf diarrhoea characterized by morphological, biochemical and polymerase chain reaction (PCR). Antibacterial activities of green Ag NPS on bacteria isolated from calf diarrhoea include multidrug-resistant bacteria (MDR) were determined by disc diffusion methods and minimal inhibition concentration (MIC). Results: The most bacteria isolated from calf diarrhoea were Escherichia coli (E. coli), Salmonella spp. and Staphylococcus aureus (S. aureus). All of these bacteria were susceptible to Ciprofloxacin, Gentamicin and Chloramphenicol but resistance to penicillin; Ampicillin and Amoxicillin. The antibacterial effects of green Ag NPS on MDR bacteria had similar or superior effects of Ciprofloxacin and Chloramphenicol. Conclusion: The present study proved that the green Ag NPs had marked antibacterial effect against bacteria isolated from calf diarrhoea and eco-friendly but still need deep extensive study in vivo before used in human or animals.