Insights into the global emergence of antifungal drug resistance

The global prevalence of fungal diseases has escalated in the last several decades. Currently, it is estimated that fungi infect 1.7 billion people annually and result in 1.5 million deaths every year1. Deaths due to fungal infections are increasing, with mortality often exceeding 50%, further increasing to 100% if treatment is delayed1. Despite these staggering figures, the contribution of fungal infections to the global burden of disease remains under-recognised. In Australia, over a 5-year period fungal infections cost Australia an estimated $583 million2. The median cost for one invasive fungal disease (IFD) is AU$30957, increasing to AU$80291 if the patient is admitted to an intensive care unit3. Treatment of fungal infections poses significant challenges due to the small number of safe and effective antifungal drugs available and emerging antifungal drug resistance. Resistance to every class of antifungal drugs has been described and for some drug classes is extremely common4,5.

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Antimycotic drugs and their mechanisms of resistance to Candida species

Current Drug Targets ◽

10.2174/1389450122666210719124143 ◽

2021 ◽

Vol 22 ◽

Author(s):

Sweety Dahiya ◽

Namita Sharma ◽

Aruna Punia ◽

Pooja Choudhary ◽

Prity Gulia ◽

...

Keyword(s):

Drug Resistance ◽

Candida Species ◽

Molecular Mechanisms ◽

Fungal Infections ◽

Treatment Strategies ◽

Distinct Species ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Success Rates ◽

Antifungal Drug Resistance

: Fungal infections have shown an upsurge in recent decades, mainly because of the increasing number of immunocompromised patients, and the occurrence of invasive candidiasis is found to be 7-15 folds greater than that of invasive aspergillosis. The genus Candida comprises of more than 150 distinct species; however, only a few of them are found to be pathogenic to humans. Mortality rates of Candida species are found to be around 45%, and the reasons for this intensified mortality are inefficient diagnostic techniques and unfitting initial treatment strategies. There are only a few antifungal drug classes that are employed for the remedy of invasive fungal infections, including azoles, polyenes, echinocandins, and pyrimidine analogs. During the last 2-3 decades, the usage of antifungal drugs has increased several folds, due to which the reports of escalating antifungal drug resistance have also been recorded. The resistance is mostly to the triazole-based compounds. Due to antifungal drug resistance, the success rates of treatment have been reduced and major changes have been observed in the frequency of fungal infections. In this review, we have summarized the major molecular mechanisms for the development of antifungal drug resistance.

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Fungal biofilm architecture produces hypoxic microenvironments that drive antifungal resistance

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2003700117 ◽

2020 ◽

Vol 117 (36) ◽

pp. 22473-22483 ◽

Cited By ~ 2

Author(s):

Caitlin H. Kowalski ◽

Kaesi A. Morelli ◽

Daniel Schultz ◽

Carey D. Nadell ◽

Robert A. Cramer

Keyword(s):

Drug Resistance ◽

Fungal Infections ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Drug Resistant ◽

Antifungal Drug Resistance ◽

Oxygen Gradients ◽

Biofilm Architecture

Human fungal infections may fail to respond to contemporary antifungal therapies in vivo despite in vitro fungal isolate drug susceptibility. Such a discrepancy between in vitro antimicrobial susceptibility and in vivo treatment outcomes is partially explained by microbes adopting a drug-resistant biofilm mode of growth during infection. The filamentous fungal pathogenAspergillus fumigatusforms biofilms in vivo, and during biofilm growth it has reduced susceptibility to all three classes of contemporary antifungal drugs. Specific features of filamentous fungal biofilms that drive antifungal drug resistance remain largely unknown. In this study, we applied a fluorescence microscopy approach coupled with transcriptional bioreporters to define spatial and temporal oxygen gradients and single-cell metabolic activity withinA. fumigatusbiofilms. Oxygen gradients inevitably arise duringA. fumigatusbiofilm maturation and are both critical for, and the result of,A. fumigatuslate-stage biofilm architecture. We observe that these self-induced hypoxic microenvironments not only contribute to filamentous fungal biofilm maturation but also drive resistance to antifungal treatment. Decreasing oxygen levels toward the base ofA. fumigatusbiofilms increases antifungal drug resistance. Our results define a previously unknown mechanistic link between filamentous fungal biofilm physiology and contemporary antifungal drug resistance. Moreover, we demonstrate that drug resistance mediated by dynamic oxygen gradients, found in many bacterial biofilms, also extends to the fungal kingdom. The conservation of hypoxic drug-resistant niches in bacterial and fungal biofilms is thus a promising target for improving antimicrobial therapy efficacy.

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Efflux-Mediated Antifungal Drug Resistance

Clinical Microbiology Reviews ◽

10.1128/cmr.00051-08 ◽

2009 ◽

Vol 22 (2) ◽

pp. 291-321 ◽

Cited By ~ 311

Author(s):

Richard D. Cannon ◽

Erwin Lamping ◽

Ann R. Holmes ◽

Kyoko Niimi ◽

Philippe V. Baret ◽

...

Keyword(s):

Drug Resistance ◽

Efflux Pump ◽

Fungal Infections ◽

Efflux Pumps ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Model Systems ◽

Attributable Mortality ◽

Azole Resistance ◽

Antifungal Drug Resistance

SUMMARY Fungi cause serious infections in the immunocompromised and debilitated, and the incidence of invasive mycoses has increased significantly over the last 3 decades. Slow diagnosis and the relatively few classes of antifungal drugs result in high attributable mortality for systemic fungal infections. Azole antifungals are commonly used for fungal infections, but azole resistance can be a problem for some patient groups. High-level, clinically significant azole resistance usually involves overexpression of plasma membrane efflux pumps belonging to the ATP-binding cassette (ABC) or the major facilitator superfamily class of transporters. The heterologous expression of efflux pumps in model systems, such Saccharomyces cerevisiae, has enabled the functional analysis of efflux pumps from a variety of fungi. Phylogenetic analysis of the ABC pleiotropic drug resistance family has provided a new view of the evolution of this important class of efflux pumps. There are several ways in which the clinical significance of efflux-mediated antifungal drug resistance can be mitigated. Alternative antifungal drugs, such as the echinocandins, that are not efflux pump substrates provide one option. Potential therapeutic approaches that could overcome azole resistance include targeting efflux pump transcriptional regulators and fungal stress response pathways, blockade of energy supply, and direct inhibition of efflux pumps.

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Linking Cellular Morphogenesis with Antifungal Treatment and Susceptibility in Candida Pathogens

Journal of Fungi ◽

10.3390/jof5010017 ◽

2019 ◽

Vol 5 (1) ◽

pp. 17 ◽

Cited By ~ 11

Author(s):

Jehoshua Sharma ◽

Sierra Rosiana ◽

Iqra Razzaq ◽

Rebecca Shapiro

Keyword(s):

Drug Resistance ◽

Candida Species ◽

Cellular Response ◽

Fungal Pathogens ◽

Fungal Infections ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Health Concern ◽

Biofilm Growth ◽

Antifungal Drug Resistance

Fungal infections are a growing public health concern, and an increasingly important cause of human mortality, with Candida species being amongst the most frequently encountered of these opportunistic fungal pathogens. Several Candida species are polymorphic, and able to transition between distinct morphological states, including yeast, hyphal, and pseudohyphal forms. While not all Candida pathogens are polymorphic, the ability to undergo morphogenesis is linked with the virulence of many of these pathogens. There are also many connections between Candida morphogenesis and antifungal drug treatment and susceptibility. Here, we review how Candida morphogenesis—a key virulence trait—is linked with antifungal drugs and antifungal drug resistance. We highlight how antifungal therapeutics are able to modulate morphogenesis in both sensitive and drug-resistant Candida strains, the shared signaling pathways that mediate both morphogenesis and the cellular response to antifungal drugs and drug resistance, and the connection between Candida morphology, drug resistance, and biofilm growth. We further review the development of anti-virulence drugs, and targeting Candida morphogenesis as a novel therapeutic strategy to target fungal pathogens. Together, this review highlights important connections between fungal morphogenesis, virulence, and susceptibility to antifungals.

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Dysfunction of Prohibitin 2 Results in Reduced Susceptibility to Multiple Antifungal Drugs via Activation of the Oxidative Stress-Responsive Transcription Factor Pap1 in Fission Yeast

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00860-18 ◽

2018 ◽

Vol 62 (11) ◽

Cited By ~ 1

Author(s):

Qiannan Liu ◽

Fan Yao ◽

Guanglie Jiang ◽

Min Xu ◽

Si Chen ◽

...

Keyword(s):

Oxidative Stress ◽

Drug Resistance ◽

Transcription Factor ◽

Fission Yeast ◽

Mitochondrial Protein ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Gene Encoding ◽

Content Type ◽

Antifungal Drug Resistance

ABSTRACT The fight against resistance to antifungal drugs requires a better understanding of the underlying cellular mechanisms. In order to gain insight into the mechanisms leading to antifungal drug resistance, we performed a genetic screen on a model organism, Schizosaccharomyces pombe, to identify genes whose overexpression caused resistance to antifungal drugs, including clotrimazole and terbinafine. We identified the phb2+ gene, encoding a highly conserved mitochondrial protein, prohibitin (Phb2), as a novel determinant of reduced susceptibility to multiple antifungal drugs. Unexpectedly, deletion of the phb2+ gene also exhibited antifungal drug resistance. Overexpression of the phb2+ gene failed to cause drug resistance when the pap1+ gene, encoding an oxidative stress-responsive transcription factor, was deleted. Furthermore, pap1+ mRNA expression was significantly increased when the phb2+ gene was overexpressed or deleted. Importantly, either overexpression or deletion of the phb2+ gene stimulated the synthesis of NO and reactive oxygen species (ROS), as measured by the cell-permeant fluorescent NO probe DAF-FM DA (4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate) and the ROS probe DCFH-DA (2′,7′-dichlorodihydrofluorescein diacetate), respectively. Taken together, these results suggest that Phb2 dysfunction results in reduced susceptibility to multiple antifungal drugs by increasing NO and ROS synthesis due to dysfunctional mitochondria, thereby activating the transcription factor Pap1 in fission yeast.

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Examining Signatures of Natural Selection in Antifungal Resistance Genes Across Aspergillus Fungi

Frontiers in Fungal Biology ◽

10.3389/ffunb.2021.723051 ◽

2021 ◽

Vol 2 ◽

Author(s):

Renato Augusto Corrêa dos Santos ◽

Matthew E. Mead ◽

Jacob L. Steenwyk ◽

Olga Rivero-Menéndez ◽

Ana Alastruey-Izquierdo ◽

...

Keyword(s):

Drug Resistance ◽

Natural Selection ◽

Positive Selection ◽

Resistance Genes ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Sequence Evolution ◽

Antifungal Drug Resistance ◽

Selective Pressures ◽

Substantial Heterogeneity

Certain Aspergillus fungi cause aspergillosis, a set of diseases that typically affect immunocompromised individuals. Most cases of aspergillosis are caused by Aspergillus fumigatus, which infects millions of people annually. Some closely related so-called cryptic species, such as Aspergillus lentulus, can also cause aspergillosis, albeit at lower frequencies, and they are also clinically relevant. Few antifungal drugs are currently available for treating aspergillosis and there is increasing worldwide concern about the presence of antifungal drug resistance in Aspergillus species. Furthermore, isolates from both A. fumigatus and other Aspergillus pathogens exhibit substantial heterogeneity in their antifungal drug resistance profiles. To gain insights into the evolution of antifungal drug resistance genes in Aspergillus, we investigated signatures of positive selection in 41 genes known to be involved in drug resistance across 42 susceptible and resistant isolates from 12 Aspergillus section Fumigati species. Using codon-based site models of sequence evolution, we identified ten genes that contain 43 sites with signatures of ancient positive selection across our set of species. None of the sites that have experienced positive selection overlap with sites previously reported to be involved in drug resistance. These results identify sites that likely experienced ancient positive selection in Aspergillus genes involved in resistance to antifungal drugs and suggest that historical selective pressures on these genes likely differ from any current selective pressures imposed by antifungal drugs.

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Candida antifungal drug resistance in sub-Saharan African populations: A systematic review

F1000Research ◽

10.12688/f1000research.10327.2 ◽

2017 ◽

Vol 5 ◽

pp. 2832 ◽

Cited By ~ 1

Author(s):

Charlene Wilma Joyce Africa ◽

Pedro Miguel dos Santos Abrantes

Keyword(s):

Drug Resistance ◽

Susceptibility Testing ◽

Antifungal Susceptibility ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Sub Saharan Africa ◽

African Countries ◽

Antifungal Drug Resistance ◽

Resistance Patterns ◽

Sub Saharan

Background:Candidainfections are responsible for increased morbidity and mortality rates in at-risk patients, especially in developing countries where there is limited access to antifungal drugs and a high burden of HIV co-infection. Objectives:This study aimed to identify antifungal drug resistance patterns within the subcontinent of Africa. Methods: A literature search was conducted on published studies that employed antifungal susceptibility testing on clinicalCandidaisolates from sub-Saharan African countries using Pubmed and Google Scholar. Results: A total of 21 studies from 8 countries constituted this review. Only studies conducted in sub-Saharan Africa and employing antifungal drug susceptibility testing were included. Regional differences inCandidaspecies prevalence and resistance patterns were identified. Discussion: The outcomes of this review highlight the need for a revision of antifungal therapy guidelines in regions most affected byCandidadrug resistance. Better controls in antimicrobial drug distribution and the implementation of regional antimicrobial susceptibility surveillance programmes are required in order to reduce the highCandidadrug resistance levels seen to be emerging in sub-Saharan Africa.

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Candida glabrata Drug:H+Antiporter CgQdr2 Confers Imidazole Drug Resistance, Being Activated by Transcription Factor CgPdr1

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00811-12 ◽

2013 ◽

Vol 57 (7) ◽

pp. 3159-3167 ◽

Cited By ~ 42

Author(s):

Catarina Costa ◽

Carla Pires ◽

Tânia R. Cabrito ◽

Adeline Renaudin ◽

Michiyo Ohno ◽

...

Keyword(s):

Drug Resistance ◽

Transcription Factor ◽

Multidrug Resistance ◽

Candida Glabrata ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Major Facilitator Superfamily ◽

Pathogenic Yeast ◽

Content Type ◽

Antifungal Drug Resistance

ABSTRACTThe widespread emergence of antifungal drug resistance poses a severe clinical problem. Though predicted to play a role in this phenomenon, the drug:H+antiporters (DHA) of the major facilitator superfamily have largely escaped characterization in pathogenic yeasts. This work describes the first DHA from the pathogenic yeastCandida glabratareported to be involved in antifungal drug resistance, theC. glabrata QDR2(CgQDR2) gene (ORFCAGL0G08624g). The expression ofCgQDR2inC. glabratawas found to confer resistance to the antifungal drugs miconazole, tioconazole, clotrimazole, and ketoconazole. By use of a green fluorescent protein (GFP) fusion, the CgQdr2 protein was found to be targeted to the plasma membrane inC. glabrata. In agreement with these observations,CgQDR2expression was found to decrease the intracellular accumulation of radiolabeled clotrimazole inC. glabrataand to play a role in the extrusion of this antifungal from preloaded cells. Interestingly, the functional heterologous expression ofCgQDR2in the model yeastSaccharomyces cerevisiaefurther confirmed the role of this gene as a multidrug resistance determinant: its expression was able to complement the susceptibility phenotype exhibited by itsS. cerevisiaehomologue,QDR2, in the presence of imidazoles and of the antimalarial and antiarrhythmic drug quinidine. In contrast to the findings reported for Qdr2, CgQdr2 expression does not contribute to the ability of yeast to grow under K+-limiting conditions. Interestingly,CgQDR2transcript levels were seen to be upregulated inC. glabratacells challenged with clotrimazole or quinidine. This upregulation was found to depend directly on the transcription factor CgPdr1, the major regulator of multidrug resistance in this pathogenic yeast, which has also been found to be a determinant of quinidine and clotrimazole resistance inC. glabrata.

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Antifungal Drug Development: Targeting the Fungal Sphingolipid Pathway

Journal of Fungi ◽

10.3390/jof6030142 ◽

2020 ◽

Vol 6 (3) ◽

pp. 142

Author(s):

Kyle McEvoy ◽

Tyler G. Normile ◽

Maurizio Del Poeta

Keyword(s):

Drug Resistance ◽

Drug Development ◽

Mechanism Of Action ◽

Fungal Infections ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Antifungal Compounds ◽

Metabolizing Enzymes ◽

Progressive Development

Fungal infections are becoming more prevalent and problematic due to the continual rise of immune deficient patients as well as the progressive development of drug resistance towards currently available antifungal drugs. There has been a significant increase in the development of antifungal compounds with a similar mechanism of action of current drugs. In contrast, there has been very little progress in developing compounds inhibiting totally new fungal targets or/and fungal pathways. This review focuses on novel compounds recently discovered to target the fungal sphingolipids and their metabolizing enzymes.

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The Histoplasma capsulatum DDR48 Gene Is Required For Survival Within Macrophages, Response To Oxidative Stress, And Resistance to Antifungal Drugs

10.1101/2020.10.25.354308 ◽

2020 ◽

Author(s):

Logan T. Blancett ◽

Kauri A. Runge ◽

Gabriella M. Reyes ◽

Lauren A. Kennedy ◽

Sydney C. Jackson ◽

...

Keyword(s):

Oxidative Stress ◽

Drug Resistance ◽

Stress Response ◽

Virulence Factors ◽

Histoplasma Capsulatum ◽

The United States ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Antifungal Drug Resistance

AbstractHistoplasma capsulatum (Hc) is a systemic, dimorphic fungal pathogen that affects upwards of 500,000 individuals in the United States annually. Hc grows as a multicellular mold at environmental temperatures; whereas, upon inhalation into a human or other mammalian host, it transforms into a unicellular, pathogenic yeast. This manuscript is focused on characterizing the DNA damage-responsive gene HcDDR48. HcDDR48 was originally isolated via a subtractive DNA library enriched for transcripts enriched in the mold-phase of Hc growth. Upon further analysis we found that HcDDR48 is not just expressed in the mold morphotype, but both growth programs dependent upon the environment. We found that HcDDR48 is involved in oxidative stress response, antifungal drug resistance, and survival within resting and activated macrophages. Growth of ddr48Δ yeasts was severely decreased when exposed to the reactive oxygen species generator paraquat, as compared to wildtype controls. We also found that ddr48Δ yeasts were 2-times more sensitive to the antifungal drugs amphotericin b and ketoconazole. To test HcDDR48’s involvement in vivo, we infected resting and activated RAW 264.7 murine macrophages with Hc yeasts and measured yeast survival 24-hours post-infection. We observed a significant decrease in yeast recovery in the ddr48Δ strain compared to wildtype Hc levels. Herein, we demonstrate the importance of maintaining a functional copy of HcDDR48 in order for Hc yeasts to sense and respond to numerous environmental and host-associated stressors.ImportanceHistoplasma capsulatum is an intracellular pathogen of phagocytes, where it subverts immune recognition and avoids killing by the innate immune system. Macrophages provide a permissive environment for Hc replication and killing only occurs upon the onset of the T-cell driven adaptive immune response. Hc has evolved numerous virulence factors that aid in its survival against host-derived ROS and RNS in vivo. While these virulence factors have been described in past years, only a few reports describing the regulation of these genes and how this intricate system leads to fungal survival. In this study, we characterized the stress response gene DDR48 and determined it to be indispensable for Hc survival within macrophages. HcDDR48 regulates transcript levels of superoxide dismutases and catalases responsible for detoxification of ROS and contributes to antifungal drug resistance. Our studies highlight DDR48 as a potential target to control Hc infection and decrease the severity of the disease process.

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