In-Vitro and In-Vivo Antimicrobial Activity of Protoberberine Alkaloids as Novel Therapeutic Candidates Against Mycoplasma hyopneumoniae

Mycoplasma hyopneumoniae (M. hyopneumoniae, Mhp), the primary causative agent of swine enzootic pneumonia, poses significant threats to the pork industry, challenging food safety and sustainable development of the livestock industry. Here, four protoberberine alkaloids–epiberberine, jatrorrhizine, berberine, and coptisine–were identified by screening multiple natural compounds and their anti-Mhp activity evaluated. All alkaloids exhibited potent inhibitory effects against the virulent Mhp strain ES-2, with minimum inhibitory concentrations from 8‒32 µg/mL. Their bactericidal activities were time- and concentration-dependent. At the cellular level, all protoberberine alkaloids significantly enhanced infected cell viability, suppressed pro-inflammatory cytokine expression (TNF-α, IL-6, and IL-1β), and exhibited low cytotoxicity toward host cells. In a Chang Da binary cross breeding pig infection model, jatrorrhizine markedly alleviated clinical symptoms, reduced pulmonary pathogen loads, and mitigated histopathological damage in lung tissues, with therapeutic efficacy comparable to that of florfenicol. Our findings demonstrated that protoberberine alkaloids possessed potent anti-Mhp activity, high safety profiles, and promising therapeutic potential.