Weaning is the most crucial period associated with increased stress and susceptibility to diseases in rabbits. Methicillin-resistant Staphylococcus aureus (MRSA), a historic emergent pathogen related to post weaning stressors, adversely affects rabbit’s growth rate and productive cycle. Since MRSA is rapidly evolving antibiotics resistance, natural products are desperately required to tackle the public health threats posed by antimicrobial resistance. Thus, this study aimed to screen the in vitro antibacterial activity of Nigella sativa extract (NSE) and its interactions with antibiotics against MRSA isolates. Moreover, 200 weaned rabbits were divided into 4 groups to investigate the in vivo superiority of NSE graded levels towards growth performance, tight junction integrity, immune responsiveness and resistance against MRSA. Herein, NSE showed promising antimicrobial activities against MRSA isolates from animal (77.8%) and human (64.3%) origins. Additionally, MRSA isolates exposed to NSE became sensitive to all antimicrobials to which they were previously resistant. Our results described that the growth-promoting functions of NSE, especially at higher levels, were supported by elevated activities of digestive linked enzymes. Post-NSE feeding, rabbits’ sera mediated bactericidal activities against MRSA. Notably, upregulated expression of occludin, CLDN-1, MUC-2 and JAM-2 genes was noted post NSE supplementation with maximum transcriptional levels in 500 mg/kg NSE fed group. Our data described that NSE constitutively motivated rabbits’ immune responses and protected them against MRSA-induced experimental infection. Our results suggest the antimicrobial, growth stimulating and immunomodulation activities of NSE to maximize the capability of rabbits for disease response.