Screening and characterization of a novel Antibiofilm polypeptide derived from filamentous Fungi

Abstract

In the present study, 120 fungal isolates were locally isolated from soil and selected according to their ability to antimicrobial activity. Then, selected isolates were tested for their ability to prevent biofilm formation and only one isolate (A01) showed an antibiofilm effect. The isolate A01 identified as Aspergillus tubingensis by sequencing of the 18S ITS region and a segment of β-tubulin gene. Then, 5 fractions were prepared from the culture filtrate of A. tubingensis A01 using the ultrafiltration technique to find active polypeptide fraction. The experiments revealed that one of them had an antibiofilm activity. The MALDI-TOF/MS analyses demonstrated that this polypeptide composed of 92 amino acids and had a molecular mass of 10,087 Da. The sequence alignment showed homology with hypothetical protein (OJI81679.1). The gene coding for this polypeptide consisting of 279 nucleotides, herein we called astucin, was cloned and sequenced from A. tubingensis A01 to confirm results. The MIC of the purified polypeptide was 32 m/L and 128 μg/mL and the MBIC was 2 and 8 μg/mL against Staphylococcus aureus and MRSA, respectively. The results demonstrated that the antimicrobial and antibiofilm activity of astucin, together with its lack of cytotoxicity, makes it an alternative for application in medicine. SIGNIFICANCE: Antibiotic resistance is a global problem and the emergence of antibiotic resistant bacteria reduce the effect the current treatment approaches. In this context, antimicrobial peptides stand out as potentional agents to combat bacterial infection especially, biofilm related infections. Importantly, this study have greatly considered our understanding for fungal derived antibiofilm polypeptides. In this study, traditional selection method combined with crystal violet assay is used to investigate antibiofilm polypeptides. We identified antibiofilm polypeptides purified from A. tubingensis A01. This protein shows antimicrobial and antibiofilm activity against S. aureus.

Keywords: Antibiofilm polypeptide; Aspergillus tubingensis; Extracellular protein; Staphylococcus aureus.