Antifungal and Antibiofilm Activities of B-Type Oligomeric Procyanidins From Commiphora leptophloeos Used Alone or in Combination With Fluconazole Against Candida spp.

Sylvie Michel,Ana Caroline Zanatta,Thiago Antǒnio De Sousa Araújo,Guilherme Maranhão Chaves,Matheus De Freitas Fernandes-Pedrosa,Wagner Vilegas,Silvana Maria Zucolotto,Renato Dantas-Medeiros,Luanda Bárbara Ferreira Canário De Souza,Raphaël Grougnet,Manoela Torres-Rêgo,Bruno Amorim-Carmo,Júlia Morais Fernandes

doi:10.3389/fmicb.2021.613155

Abstract

Commiphora leptophloeos (Burseraceae) is a medicinal plant native to Brazil which is popularly used for treating oral and vaginal infections. There has been no scientific evidence pointing to its efficacy in the treatment of these infections. Thus, this study sought to investigate the cytotoxic, antifungal, and antibiofilm activity of C. leptophloeos against Candida spp. and to isolate, identify, and quantify the content of B-type oligomeric procyanidins (BDP) in the extract of C. leptophloeos stem bark. The extract and the n-butanol fraction were obtained by maceration and liquid-liquid partition, respectively. Phytochemical analysis performed by HPLC-PDA/ELSD and FIA-ESI-IT-MS/MS allowed the identification and quantification of BDP in the samples. The application of centrifugal partition chromatography helped isolate BDP, which was identified by 1H NMR and MS analyses. Candida spp. reference strains and clinical isolates (including fluconazole-resistant strains) derived from the blood cultures of candidemic patients and the vaginal secretion of patients with vulvovaginal candidiasis were used for evaluating the antifungal and antibiofilm effects. Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) were determined by the microdilution technique, and biofilm inhibition was evaluated through crystal violet and XTT assays. The combined action of BDP with fluconazole was determined by the checkerboard method. The extract, the n-butanol fraction, and the BDP exhibited antifungal activity with MIC values ranging from 312.5 to 2500 μg/mL and were found to significantly reduce the biofilm formed in all the Candida strains investigated. BDP showed a fungicidal potential against strains of Candida spp. (especially against fluconazole-resistant strains), with MIC and MFC values ranging from 156.2 to 2500 μg/mL. In addition, the combined application of BDP and fluconazole produced synergistic antifungal effects against resistant Candida spp. (FICI = 0.31–1.5). The cytotoxic properties of the samples evaluated in human erythrocytes through hemolytic test did not show hemolytic activity under active concentrations. The findings of the study show that C. leptophloeos has antifungal and antibiofilm potential but does not cause toxicity in human erythrocytes. Finally, BDP, which was isolated for the first time in C. leptophloeos, was found to exhibit antifungal effect against Candida spp. either when applied alone or in combination with fluconazole.

Highlights

Several microorganisms are part of the normal human microbiota, and these include the fungi from the Candida genus
C. albicans is the most virulent and isolated species from the bloodstream of hospitalized patients, in recent years, studies published have pointed to a noticeable increase in the occurrence of infections caused by clinically relevant non-C. albicans Candida (NCAC) species, such as Candida tropicalis, Candida dubliniensis, Candida krusei, Candida glabrata, and Candida parapsilosis (Biswal et al, 2017; Banerjee et al, 2019)
The phytochemical profile of HECL and butanolic fraction (BF) was initially characterized by mass spectrometry which indicated the presence of quinic acid and oligomeric procyanidins derived exclusively from flavan-3-ol

Summary

INTRODUCTION

Several microorganisms are part of the normal human microbiota, and these include the fungi from the Candida genus. There appears to be a reasonable number of antifungal drugs in the market, very few of these drugs, including amphotericin B and echinocandins, are effective against fungal infections associated with biofilm formation (Sherry et al, 2017; De Cássia Orlandi Sardi et al, 2018) These antifungal drugs have been found to have some limitations, which include increased antifungal resistance (mainly due to emerging Candida species), high costs, and high degree of toxicity in humans (Colombo et al, 2017; Spivak and Hanson, 2018). Considering the importance of C. leptophloeos in traditional medicine, coupled with the lack of scientific studies on this plant species, the present work sought to investigate the cytotoxicity and antifungal and antibiofilm activities of B-type oligomeric procyanidins from Commiphora leptophloeos alone or in combination with fluconazole against Candida spp. The study aimed to isolate, characterize, and quantify the active compounds present in the stem bark of C. leptophloeos

MATERIALS AND METHODS

RESULTS

DISCUSSION

CONCLUSION

DATA AVAILABILITY STATEMENT