Unmasking the Amphotericin B Resistance Mechanisms in Candida haemulonii Species Complex

Abstract

The polyene amphotericin B (AMB) exerts a powerful and broad antifungal activity. AMB acts by (i) binding to ergosterol, leading to pore formation at the fungal plasma membrane with subsequent ion leakage, and (ii) inducing the intracellular accumulation of reactive oxygen species (ROS). Herein, we have deciphered the AMB resistance mechanisms in clinical isolates of Candida haemulonii complex (C. haemulonii, C. duobushaemulonii, C. haemulonii var. vulnera) in comparison to other clinically relevant non-albicans Candida species. Membrane gas chromatography−mass spectrometry analysis revealed that the vast majority of sterols were composed of ergosterol pathway intermediates, evidencing the absence of AMB target. Supporting this data, C. haemulonii species complex demonstrated poor membrane permeability after AMB treatment. Regarding the oxidative burst, AMB induced the formation of ROS in all species tested; however, this phenomenon was slightly seen in C. haemulonii complex isolates. Our results indicated that these isolates displayed altered respiratory status, as revealed by their poor growth in nonfermented carbon sources, low consumption of oxygen, and derisive mitochondrial membrane potential. The use of specific inhibitors of mitochondrial respiratory chain (complex I−IV) revealed no effects on the yeast growth, highlighting the metabolic shift to fermentative pathway in C. haemulonii strains. Also, C. haemulonii complex proved to be highly resistant to oxidative burst agents, which can be correlated with a high activity of antioxidant enzymes. Our data demonstrated primary evidence suggesting that ergosterol content, mitochondrial function, and fungal redox homeostasis are involved in AMB fungicidal effects and might explain the resistance presented in this multidrug-resistant, emergent, and opportunistic fungal complex.

Fingolimod Potentiates the Antifungal Activity of Amphotericin B

“…Remarkable accumulation of ROS can result in oxidative stress and cellular components damage which are toxic to the organisms, even leading to autophagy, apoptosis or necrosis (Scherz-Shouval and Elazar, 2007;Nakagawa, 2008;Yu et al, 2016). AMB binds to ergosterol at plasma membrane causing loss of ions, induces ROS accumulation in different pathogenic fungal species (Mesa-Arango et al, 2014;Silva et al, 2020) and causes lipid peroxidation through a greatly increased ROS level and reactive nitrogen species (NOS) level (Ferreira et al, 2013). The ROS induction subsequently leading to apoptosis is an important mechanism of AMB in C. albicans (Francois et al, 2006).…”

Antifungal susceptibility profile of Candida clinical isolates from 22 hospitals of São Paulo State, Brazil

“…VCZ MICs were not elevated, which differs from other studies ( 14 , 39 ). A recent Brazilian study has shown that resistance to oxidative stress along with changes in homeostasis, mitochondrial function, and ergosterol may be related to AmB resistance in the C. haemulonii complex ( 40 ). Cases of C. haemulonii complex are increasing according to recent reports, and more research is needed to understand antifungal susceptibility within the complex ( 36 ).…”

The Elevated Endogenous Reactive Oxygen Species Contribute to the Sensitivity of the Amphotericin B-Resistant Isolate of Aspergillus flavus to Triazoles and Echinocandins

“…Although AMB resistance has not been reported to be associated with the missing ergosterol in Aspergillus spp. ( Blum et al, 2008 , 2013 ), the absence of ergosterol in Candida spp., caused by mutations in genes of the ergosterol biosynthesis ( Geber et al, 1995 ; Sanglard et al, 2003 ; Martel et al, 2010a ; Vincent et al, 2013 ; Silva et al, 2020 ), leading to AMB resistance and abnormal membrane structure and function, which also drastically diminished tolerance to oxidative stress ( Vincent et al, 2013 ). These studies are consistent with the phenotypes observed in the AMB-resistant isolate of A. flavus .…”