Abstract
Bovine mastitis is a major threat to animal health and the dairy industry. Staphylococcus aureus is a contagious pathogen that is usually associated with persistent intramammary infections, and biofilm formation is a relevant aspect of the outcome of these infections. Several biological activities have been described for snake venoms, which led us to screen secretions of Bothrops jararacussu for antibiofilm activity against S. aureus NRS155. Crude venom was fractionated by size-exclusion chromatography, and the fractions were tested against S. aureus. Biofilm growth, but not bacterial growth, was affected by several fractions. Two fractions (15 and 16) showed the best activities and were also assayed against S. epidermidis NRS101. Fraction 15 was identified by TripleTOF mass spectrometry as a galactose-binding C-type lectin with a molecular weight of 15 kDa. The lectin was purified from the crude venom by D-galactose affinity chromatography, and only one peak was observed. This pure lectin was able to inhibit 75% and 80% of S. aureus and S. epidermidis biofilms, respectively, without affecting bacterial cell viability. The lectin also exhibited a dose-dependent inhibitory effect on both bacterial biofilms. The antibiofilm activity was confirmed using scanning electron microscopy. A pre-formed S. epidermidis biofilm was significantly disrupted by the C-type lectin in a time-dependent manner. Additionally, the lectin demonstrated the ability to inhibit biofilm formation by several mastitis pathogens, including different field strains of S. aureus, S. hyicus, S. chromogenes, Streptococcus agalactiae, and Escherichia coli. These findings reveal a new activity for C-type lectins. Studies are underway to evaluate the biological activity of these lectins in a mouse mastitis model.
Highlights
MethodsThis study required no approval by the University Ethics Committee on Animal Use (CEUA) based on reasoning that the study dealt with venom milked from snakes of the wild and no laboratory experimentation was conducted with animals
A bacterial biofilm is defined as a complex and structured community of organisms enclosed in a self-produced polymeric matrix that contains exopolysaccharides, proteins, teichoic acids, PLOS ONE | DOI:10.1371/journal.pone.0120514 March 26, 2015Disruption of Staphylococcal Biofilms by a C-Type Lectin
A microplate biofilm assay and a growth curve analysis were performed to evaluate whether fractions 15 and 16 were affecting bacterial growth in a way that could promote a reduction in biofilm production (Fig. 3)
Summary
This study required no approval by the University Ethics Committee on Animal Use (CEUA) based on reasoning that the study dealt with venom milked from snakes of the wild and no laboratory experimentation was conducted with animals. The committee understands that the research was previously released by the Chico Mendes Institute for Biodiversity Conservation (ICMBio), an administrative arm of the Ministry of the Environment (MMA), that has the overall responsibility of supervising all wildlife research in the country (permit no 39126–1). This permit allowed snake capture and milking that were carried out by a herpetologist from the Department of Animal Biology. The samples were collected at the Mata da Biologia, a University research land used by students and faculty of the Universidade Federal de Viçosa
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