Monohexosylceramides from Rhizopus Species Isolated from Brazilian Caatinga: Chemical Characterization and Evaluation of Their Anti-Biofilm and Antibacterial Activities.

Edson Vieira,Eliana Barreto-Bergter,Celuta Alviano,Milagre Pele,Daniela Alviano,Mariana Xisto,Galba De Campos-Takaki

doi:10.3390/molecules23061331

Abstract

Monohexosylceramides (CMHs) are highly conserved fungal glycosphingolipids playing a role in several cellular processes such as growth, differentiation and morphological transition. In this study, we report the isolation, purification and chemical characterization of CMHs from Rhizopus stolonifer and R. microspores. Using positive ion mode ESI-MS, two major ion species were observed at m/z 750 and m/z 766, respectively. Both ion species consisted of a glucose/galactose residue attached to a ceramide moiety containing 9-methyl-4,8-sphingadienine with an amidic linkage to a hydroxylated C16:0 fatty acid. The antimicrobial activity of CMH was evaluated against Gram positive and Gram negative bacteria using the agar diffusion assay. CMH from both Rhizopus species inhibited the growth of Bacillus terrae, Micrococcus luteus (M. luteus) and Pseudomonas stutzeri (P. stutzeri) with a MIC50 of 6.25, 6.25 and 3.13 mg/mL, respectively. The bactericidal effect was detected only for M. luteus and P. stutzeri, with MBC values of 25 and 6.25 mg/mL, respectively. Furthermore, the action of CMH on the biofilm produced by methicillin-resistant Staphylococcus aureus (MRSA) was analyzed using 12.5 and 25 mg/mL of CMH from R. microsporus. Total biofilm biomass, biofilm matrix and viability of the cells that form the biofilm structure were evaluated. CMH from R. microsporus was able to inhibit the MRSA biofilm formation in all parameters tested.

Highlights

Fungi of the order Mucorales are commonly found in soil as saprophytes and decomposing organic matter all around the world
The Folch lower layer containing neutral glycolipids was fractionated by silica gel chromatography and the fractions monitored by HPTLC on silica plates developed with chloroform: methanol: 2 M NH4 OH (40:10:1 v/v)
A major lithiated ion species at m/z 750 was observed in R. microsporus grown in peptone-dextrose dextrose broth (PDB) and YEPD media and in R. stolonifer grown in YEPD medium, respectively

Summary

Introduction

Fungi of the order Mucorales are commonly found in soil as saprophytes and decomposing organic matter all around the world. In Brazil these fungi are widely found in the northeastern region in arid and semi-arid ecosystems such as the Caatinga, which is an exclusively Brazilian domain in the semi-arid region of Brazil [1,2]. Molecules 2018, 23, 1331 distributed in soils of the Caatinga in the Brazilian Northeast [2]. Little was known about the cell wall glycoconjugates of the Rhizopus species and, their specific functions in the fungal cell and in their interaction with the environment. CMH are important in promoting alkaline tolerance in vitro due to the involvement of CMH in the regulation of membrane fungal lipid domains that lead to the redistribution of CMH in the membrane in an alkaline environment [11,12,13]. CMH interact with defensins isolated from insects and plants [14]

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