Rutin is a natural product has various biological activities. Pasteurellosis is crucial bacterial infection of respiratory system caused by Pasteurella multocida. This study aimed to investigate the improved antibacterial effect of Rutin nanocarriers as mucoadhesive intranasal delivery against Pasteurella multocida. Different formulations of Rutin niosomes and nanostructure lipid carriers (NLCs) were formulated and well characterized. The in vivo antibacterial performance of the developed formulations against Pasteurella multocida in infected mice was conducted. Further, cytokines levels of Interferon Gamma (INF-γ) and Interlukin-12 (IL -12) in mice sera were assessed. The results revealed that developed Rutin nanocarriers were in nanosized range and exhibited high drug encapsulation. However, Rutin NLCs showed smaller particle size (240.34 ± 5.5 nm), higher encapsulation% (97.34 ± 0.15%), and higher drug release of 94.5% within 12 h comparing with Rutin niosomes. Further, Rutin NLCs presented the highest antibacterial effect against P. multocida infection compared with other treated groups. The bacterial count in lungs and livers was reduced in treated groups compared to the infected non treated one. Our results indicate that mucoadhesive Rutin nanocarriers introduce a new promising antibacterial agent for intranasal delivery against P. multocida and open vision for veterinary applications to utilize advanced nanocarriers in the management of several infections.