Rattling Bugs Away: The potential Antibacterial Properties of Crotalus Mictlantecuhtli against S. aureus and E. coli

Introduction

Mexico is the country with the highest diversity of rattlesnakes worldwide, and their venom represents a valuable source of molecules with biomedical potential. Documented bioactivities include antiproliferative, analgesic and microbicide properties. Following that, the World Health Organization considers antibiotic resistance as a global threat. Therefore, studying alternatives to these drugs is imperative. The Veracruz neotropical rattlesnake (Crotalus mictlantecuhtli) is an endemic species from Central Mexico whose venom has been characterized before, but with no reports of its antibacterial properties, to the best of our knowledge.

Methods

Four protein fractions from C. mictlantecuhtli venom were obtained by size exclusion chromatography (SE-LC). The fractions were lyophilized and stored at -20°C until analysis. Protein quantification was performed using a Bicinchoninic Acid (BCA) Protein Assay, with bovine serum albumin (BSA) as a standard. All fractions were adjusted by protein content, diluted with saline solution (NaCl 0.9%) and added to 96-well microplates with Müller–Hinton (MH) in concentrations ranging from 10 to 0.001 µg/µL. To evaluate the antimicrobial activity, the Minimal Inhibitory Concentration (MIC) was determined via the microdilution method, which was performed by inoculating each well with ATCC strains E. coli (25922) or S. aureus (25923) standardized in brain–heart infusion to 0.5 McFarland. The results were corroborated using the agar well diffusion method in MH. Additionally, blood samples were drawn and anticoagulated with calcium-balanced lithium heparin to evaluate indirect hemolysis in 5% blood agar. The data was analyzed by two-way ANOVA and direct observation.

Results

Venom fractions 1, 3 and 4 did not differ significantly from the gentamicin control below 1 µg/µL for E. coli and S. aureus (P > 0.05). Fraction 1 exhibited hemolytic halos with diameters of 0.7 mm and 14 mm, respectively.

Conclusions

C. mictlantecuhtli venom contains proteins of medical interest, showing antimicrobial potential; hence, it is imperative to continue studying it.