Potent Activity of Venom from a Single Captive-Bred Bitis rhinoceros Specimen Against Planktonic Clinical MRSA and Early Biofilm Development

Background: Antimicrobial resistance represents a major global health threat, particularly due to the increasing prevalence of multidrug-resistant pathogens such as methicillin-resistant Staphylococcus aureus (MRSA). Snake venoms have emerged as promising sources of bioactive molecules with potential antimicrobial applications. However, their activity against clinical MRSA isolates and bacterial biofilms remains insufficiently characterized. This study evaluated the antibacterial, biofilm formation inhibitory, and hemocompatibility properties of venom obtained from a captive-bred Bitis rhinoceros specimen against S. aureus strains and clinical MRSA isolates. Methods: Minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), minimum biofilm inhibitory concentration (MBIC), minimum biofilm eradication concentration (MBEC), and biofilm formation inhibition assays were performed. In addition, direct and indirect hemolytic and hemagglutinating activities were evaluated to assess venom selectivity and biocompatibility. Results: Proteomic characterization revealed that the venom proteome is dominated by C-type lectin-like proteins (46.7%), with minor contributions from PIII snake venom metalloproteinase, phospholipase A2, and L-amino acid oxidase toxin families. B. rhinoceros venom exhibited potent bactericidal activity against planktonic S. aureus cells, with minimum inhibitory and bactericidal concentration values ranging from 4 to 16 mg/L. Notably, the two clinical MRSA isolates showed lower values than the reference strain, although this observation should be interpreted with caution given the limited number of isolates tested. The venom also strongly inhibited biofilm formation at the concentrations tested, with inhibition exceeding 97% and remaining statistically significant across all three strains and MIC multiples when analyzed at the level of the independent experiment (n = 3; paired t-test, Benjamini–Hochberg-adjusted p < 0.001), although limited activity was observed against preformed mature biofilms. Hemocompatibility assays demonstrated low direct and indirect hemolytic activity together with favorable selectivity indices against bacterial cells. Conclusions: These findings suggest that captive-bred B. rhinoceros venom may represent a promising source of antimicrobial compounds against planktonic MRSA infections and early-stage biofilm development. The favorable hemocompatibility profile supports its potential applicability as a topical antimicrobial candidate. Nevertheless, this study is a proof of concept and, therefore, further assays are required to isolate and functionally characterize the specific venom fractions responsible for the antimicrobial activity.