ANTIMICROBIAL ACTIVITIES OF GRAPHENE OXIDE AGAINST BIOFILM AND INTRACELLULAR STAPHYLOCOCCUS AUREUS ISOLATED FROM BOVINE MASTITIS

Abstract

Intro: Staphylococcus aureus (S. aureus) is one of the causative agents of bovine mastitis. The treatment using conventional antimicrobial has been hampered due to the development of antimicrobial resistance, the ability of the bacteria to form biofilms and localize inside the host cells Methods: GO antibiofilm properties were evaluated based on the disruption of biofilm structure, and the intracellular antimicrobial activities were determined by the survival of S. aureus in infected bovine mammary cells following GO exposure. The mechanism for GO intracellular antimicrobial activities were investigated using endocytosis inhibitors. GO toxicity towards the host cells were assessed using resazurin assay. Findings: At 100 mg/L GO reduced between 30-70% of S. aureus biofilm mass, suggesting GO's ability to disrupt the biofilm structure. At 200 mg/L GO killed almost 80% of intracellular S. aureus and the antimicrobial activities were inhibited when cells were pre-treated with cytochalasin D, suggesting GO intracellular antimicrobial activities were dependent on the actin-polymerisation of the cell membrane. At < 250 mg/L, GO enhanced the Mac-T cells viability, and cells were only affected at higher dosages Discussion: Here the study demonstrated antibacterial and antibiofilm efficacy of GO, a carbon-based nanomaterial against S. aureus isolated from subclinical cases in Malaysia. GO was effective against the extracellular, intracellular and biofilms form of S. aureus. And the intracellular antimicrobial activities appeared to be dependent on actin polymerisation of the membrane cell that control the macropinocytosis molecule uptake pathway of the cells. Finally, the study showed that growth of Mac-T cells was enhanced when exposed to low concentrations of GO and toxicity was only observed when the cell was exposed at very high concentration of GO. Conclusion: GO efficacy against S. aureus in vitro suggested the compound could be further tested in vivo to recognise its potential as one of the bovine mastitis therapy.