4

Fungicide Resistance in Plant Protection Use

The third panel of the workshop explored the development of antifungal resistance in response to the agricultural use of fungicides. Marin Brewer, associate professor of Mycology and Plant Pathology at the University of Georgia, and Tom Chiller, chief of the Mycotic Diseases Branch at the Centers for Disease Control and Prevention, co-moderated the session. Matthew Fisher, professor of Fungal Disease Epidemiology at Imperial College London, discussed the history, genetics, and mechanisms of antifungal resistance. He described the prevalence of resilience in Aspergillus fumigatus (A. fumigatus) and challenges in developing antifungal treatments. Kevin Doughty, senior stewardship manager at Bayer AG Crop Science division, outlined the conditions capable of promoting the selection and amplification of resistant isolates of A. fumigatus, as well as the role these settings can play in the distribution and frequency of azole-resistant fungal strains.

GENETICS AND MECHANISMS OF FUNGICIDE RESISTANCE

Fisher discussed the history of antifungals, the distinction between resistance and tolerance, the polymorphism processes involved in resistance, the prevalence of azole-resistant A. fumigatus (ARAf), and the development of new antifungal clinical treatments.

The Proliferation of Antifungals, Tolerance, and Resistance

Humans have been battling blights and molds since agricultural practices began, said Fisher. Over the past 400 years, methods of combating

fungal infections have ranged from urine and brining in the seventeenth century, arsenic and copper sulphate in the eighteenth century, and a Bordeaux mixture of grapes in the nineteenth century. The era of modern fungicides began in the twentieth century with the introduction of a panoply of fungicides. During the current century, new methods such as RNA interference are being developed. Although numerous fungicides are used in agriculture, there are far fewer classes of antifungals available for animal use, due to their toxicity. At present, only four classes of antifungal drugs have been developed for use on animals.

Fungi are eukaryotes with flexible genomes that have the ability to respond adeptly to natural selection, Fisher explained. A large and comprehensive literature review found that fungi develop resistance to all classes of drugs used against plant and animal fungal infections (Fisher et al., 2018) (see Figure 4-1). Fungi utilize a spectrum of mechanisms to avoid the toxic effects of antifungals. Resistance mechanisms that have been identified thus far include (1) changing the structure of the target site, (2) overexpressing genes to produce more copies of the target site, (3) deleting the target site or molecule, (4) removing the drugs out of the organism through efflux pumps, (5) regulating stress response pathways that counteract the drug effects, and (6) utilizing genomic plasticity to alter regions of the genome (e.g., aneuploidy, hypermutation).

Fisher drew a distinction between resistance and tolerance, noting that the term “resistance” is often inaccurately used in cases of tolerance. Resistance is a hardwired trait—heritable and often genetically encoded—that is acquired due to changes that directly or indirectly affect the drug-target interaction (Fisher et al., 2022). Tolerance describes the ability of a fungus to grow at drug concentrations above a set point minimum inhibitory concentration of a target fungal pathway (Berman and Krysan, 2020). This more complex trait involves a wide range of epigenetic and/or general stress response pathways. A fungus growing in the presence of an antifungal drug does not necessarily indicate resistance, as a tolerance mechanism could be at play.

The relatively limited number of mode-of-action targets available in fungi has led agriculturalists to iterate on the same class of chemicals used in clinics, said Fisher. Azoles best exemplify this dual use. Azoles have been used as fungicides since the 1970s, with at least 20 varieties of azoles in use today. He remarked that large quantities of azoles are utilized in the environment (e.g., in agricultural products, paints, antifouling agents, and others) and that they may constitute the most widely used chemicals on the planet. These include both first-generation and newer generations of azoles. Pulmazole and opelconazole are examples of newly available antifungals.

Azole Resistance in Aspergillus

Farmers defend crops against a wide spectrum of blights using azole fungicides that target the ergosterol biosynthetic pathway, said Fisher. Azoles disrupt the pathway by inducing steric inhibition of the sterol 14-alpha-demethylase (CYP51), an enzyme within the cytochrome P450 superfamily (Bhattacharya et al., 2018). Polymorphisms in CYP51 take place within a number of plant pathogens, leading to resistant pathogens that affect a broad range of crops essential for the human food supply (Price et al., 2015). As these plant pathogens evolve resistance, farmers must respond by shifting to new formulations of azoles or modifying the crop through trait breeding or genetic modification to increase its resilience to the evolved fungal pathogens.

Fisher’s literature review revealed a sizable increase in azole resistance in both the human and plant spheres (Fisher et al., 2018), which raises the question of how much of the increasing resistance seen in patients is acquired through fungal adaptations to fungicides. Multiple research groups have focused on answering that question, Fisher noted. For instance, A. fumigatus is a ubiquitous thermophilic fungus found throughout the planet. Patients with chronic fungal infections are typically prescribed long-term azole treatment, and A. fumigatus evolves in response to become resistant to repeat exposures of the azole drugs over time (Snelders et al., 2011). Fisher stated, however, the polymorphisms that develop in these clinical isolates in response to long-term azole treatment tend to be single nucleotide polymorphisms in the cyp51A locus (Gsaller et al., 2016). These differ from a set of mutations that occurs in ARAf in the wild, in which tandem repeats occur in a promoter region that is the domain for the cap-binding complex (CBC) transcription factor binding. These tandem repeats increase the transcription of cyp51A. When these tandem repeats occur with polymorphisms that induce conformational changes that confer resistance, the result are the widely distributed alleles—TR34 L98H and TR46 Y121F T289A—found in the environment (Verweij et al., 2009).

Fisher described a study in which he and colleagues performed whole genome sequencing of A. fumigatus samples from across the United Kingdom (Rhodes et al., 2022). This population genetic analysis confirmed acquisition of azole resistance in Aspergillus environmental and clinical isolates throughout the United Kingdom. The dense clustering on the phylogenetic analysis suggests that azole resistance in the Aspergillus population analyzed arose through a relatively recent origin instead of spreading via recombination throughout the entire phylogeny. The genomes featured strong signs of selection on the cyp51A locus. Fisher noted that the resistant isolates from the environment were nearly identical to resistant clinical isolates, suggesting that patients are being infected by ARAf from the environment.

The Prevalence of Resistant Aspergillus fumigatus

Fisher remarked on the outstanding question of how patients were becoming infected with A. fumigatus from the environment. This fungus is ubiquitous in the soil and air. Jennifer Shelton, an applications scientist at Oxford Nanopore Technologies, developed a community science (also known as “citizen science”) approach to examine community exposure to ARAf during her doctoral research at Imperial College London (Brackin et al., 2020; Shelton, 2021; Shelton et al., 2020). Generating interest through social media, Shelton enlisted participants from across the United Kingdom. Sterile sticky films and envelopes were mailed to participants who then exposed the adhesive films to the open air for an 8-hour period, then mailed the samples back to the research team. Shelton received 1,894 returned samples from which she cultivated 2,366 A. fumigatus samples collected from the open air and screened these samples for resistance to a commonly used azole fungicide, tebuconazole (Brackin et al., 2020; Shelton, 2021; Shelton et al., 2020). She found that, of the samples screened, 1 in 20 were resistant to tebuconazole, 1 in 25 to itraconazole, 1 in 40 to voriconazole, 1 in 25 isavuconazole, and 1 in 150 were resistant to all tested medical azoles (Shelton, 2021). Fisher described that these findings constitute a broad countrywide exposure to multidrug-resistant A. fumigatus bioaerosols. Shelton then sequenced the cyp51A locus and found that the majority of the azole-resistant varieties contained the TR34/L98H allele, which is found broadly across Europe and the rest of the world, said Fisher (Shelton, 2021). He stated that by his own quick calculations, a person in the United Kingdom is on average exposed to 83,000 A. fumigatus within an 8-hour period, and nearly 4,000 of these are azole-resistant. Fisher pointed out that even if his calculations are incorrect by two orders of magnitude, an average person would be exposed to 40 azole-resistant A. fumigatus spores across an 8-hour period.

New Antifungal Drug Development

The prevalence of resistant fungi requires new antifungals, and phase II and phase III trials are currently testing promising clinical antifungals with novel modes of action, Fisher noted (Fisher et al., 2022). These include fosmanogepix, an inhibitor of the Gwt1 enzyme (an acyltransferase necessary in glycosyl-phosphatidylinositol biosynthesis); ibrexafungerp, rezafungin, and opelconazole, which are antifungal triterpenoids; and olorofim, a dihydroorotate dehydrogenase (DHODH) inhibitor. However, agricultural fungicides are also being developed that potentially share the same target site as these clinical antifungals, and thus concerns of developing cross-resistance persist (EPA, 2022; Fungicide Resistance Action Committee, 2022). For example,

the U.S. Environmental Protection Agency (EPA) has proposed registration for ipflufenoquin, a DHODH inhibitor fungicide (EPA, 2022; Fungicide Resistance Action Committee, 2022). Ipflufenoquin may or may not share the same target as the clinical DHODH inhibitor, olorofim. Fisher remarked that if the clinical and agricultural antifungals have different targets, they can both be safely used. In cases where antifungals share the same target, this overlap in use cases may impede the ability of the next generation, novel mode-of-action clinical antifungals to effectively treat the broad range of patients in need of new treatment options.

AZOLE-RESISTANT ASPERGILLUS FUMIGATUS IN AGRONOMIC SETTINGS: HOTSPOTS AND COLDSPOTS

Doughty discussed the agricultural uses of azoles, the processes and ideal conditions for the development of ARAf, the features of agronomic settings that can contribute to ARAf selection and amplification, in particular the importance of plant waste management in addressing ARAf. Representing CropLife International, the industry association of the major crop protection research and development companies, Doughty described the agronomic context as including broad acre crops, horticultural crops, and plantation crops. Given the link between environmentally derived A. fumigatus isolates found in patients and the use of azole fungicides in agricultural and agronomic settings, the development of mitigation strategies depends on understanding the locations where selection and amplification of resistance are taking place.

Azole-Resistant Aspergillus fumigatus in Context

Doughty echoed previous speakers that ARAf presents a significant, widespread hindrance to treatment of invasive aspergillosis with medical azoles. In addition to medical use, azoles (known as demethylation inhibitors or DMI fungicides) are used in the environment in the fields of agriculture/horticulture, material protection, and veterinary medicine. Within agriculture, azoles are the backbone of crop protection strategies that have been used for decades to ensure stable and reliable food security. He described azoles as an essential tool in avoiding resistance to fungicides with other modes of action and the primary tool for fighting mycotoxins that can be produced by Fusarium species in wheat and maize. Thus, their use mitigates health issues mycotoxins can cause in humans. He noted that azoles used in agriculture can be—but are not necessarily—active against A. fumigatus. While agricultural azole fungicides that are active against A. fumigatus can foster cross-resistance with medical azole fungicides, Doughty specified that A. fumigatus is not a

target pathogen in the crop protection use of agricultural azoles. However, collateral exposure of the fungus within agronomic settings can occur, particularly in the form of residues that remain after agricultural azoles are applied. He added that the selection and amplification of pre-existing resistance genotypes in these settings is most likely attributable to A. fumigatus coming into contact with residues in plant waste piles.

Agronomic “Hotspots” for Azole-Resistant Aspergillus fumigatus

The potential medical ramifications of ARAf underscores the value of determining the characteristics of agronomic settings that contribute to ARAf selection and amplification processes, said Doughty. He defined a “hotspot” for ARAf as an agronomic setting in which the following criteria are fulfilled: (1) favorable conditions for growth and multiplication of A. fumigatus, (2) exposure of A. fumigatus to residual concentrations of DMI fungicides that are selective for resistant genotypes, and (3) mass release of airborne spores of A. fumigatus into the environment. Selection and amplification cannot take place in the absence of residues of a DMI fungicide that is active against A. fumigatus. For A. fumigatus to develop resistance, the DMI fungicide must be effective against the fungus at concentrations that add selective pressure to the organism. Furthermore, sufficiently high concentrations of DMI residues (in relation to the minimum inhibitory concentration for wild-type A. fumigatus) are required to increase the proportion of the resistant genotypes at the expense of the wild type. The result is an amplification of the resistant portion of the A. fumigatus population. A hotspot then needs a method for mass release of airborne ARAf, predominantly in spore form. According to current understanding of the process, this mass release of predominantly resistant spores is a precondition for the link between the hotspot and the patient.

Patterns of ARAf Distribution and Frequency in Agronomic Settings

Doughty and colleagues conducted a literature review to examine which agronomic settings might contribute to ARAf selection and amplification (Doughty et al., 2021). The existence of a background proportion of ARAf within the environmental A. fumigatus population (including agronomic settings) has been reported, although the proportion of resistant isolates varied between settings. No distinction was detected between the frequency of ARAf in agronomic settings and in samples from urban settings. Furthermore, researchers found no clear distinction in the recovery of ARAf between azole-treated crops and organic soils and crops. Doughty noted that these comparative studies are particularly useful in understanding hotspots because much of the research on ARAf has been conducted

by sampling soils without a comparative element in the survey. Given that A. fumigatus has a background level of resistance that is presumably reflected in the air spora, and thus deposition of spores onto agronomic settings, comparative work sheds more light on the role of different agronomic settings.

The growing crop itself is not a particularly conducive habitat for A. fumigatus, said Doughty. The fungus has been found and recovered from tomato plants, but it does not thrive on the plant surface and does not attack the plant. However, because A. fumigatus is a saprobic fungus, the waste derived from crops that do not necessarily harbor ARAf can become hotspots for ARAf. A growing crop and surrounding soil may harbor limited proportions of A. fumigatus, but a large increase in the incidence of resistant isolates can occur in the waste created when the crops are harvested. Doughty stated that waste piles can fulfill all of the requirements of a hotspot and warrant focused attention in efforts to address ARAf.

Flower bulb waste is a hotspot example that has been extensively investigated. Research indicates that stockpiling plant waste can create a hotspot, depending on how the waste is managed (Zhang et al., 2021d). In addition to supporting large populations of A. fumigatus, plant waste piles also provide the conditions for sexual reproduction of the fungus, making them an ideal environment for the generation of new genotypes. Doughty noted that wide variety of cyp51A mutations of ARAf isolates have been identified in flower bulb waste piles, representing genetic variability (Zhang et al., 2021b).

In contrast to flower waste hotspots, Doughty described cereal crops—one of the main targets of azole fungicide use—as ARAf “coldspots.” Large populations of A. fumigatus or ARAf are not found in the cereal crop, soil, grain, or straw. The most frequent environmental ARAf genotypes that were detected are TR₃₄/L98H and TR₄₆/Y121F/T289A, which have been found in soils for cereal crops, but in low quantities. The frequencies of ARAf genotypes present in azole-treated cereal crop soils are similar to frequencies found in urban air. Additionally, a U.K. comparative trial examining the relative frequency of ARAf in long-term treated soils and in soils untreated with azoles found similar low proportions of resistant isolates in both treated and untreated soils (Fraaije et al., 2020). A German study compared the relatively low frequency of Aspergillus-resistant isolates in cereal fields and apple orchards in both organic and azole-treated fields (Barber et al., 2020). Once again, both settings were found to be a cold spot.

Approaches to Addressing Azole-Resistant Aspergillus fumigatus

Doughty stated that waste management is a major area of concern due to the large numbers of A. fumigatus spores that can be generated in plant

waste. Although ARAf is the central target for investigation, mitigation, and avoidance measures, reducing the release of A. fumigatus could generally be a supplemental aim. In positioning fungicides in agronomic settings, the crop protection industry should consider the risk of selecting resistance in human pathogenic fungi, he suggested. Informed by the expanding knowledge base around azole resistance, the industry could apply lessons learned to avoid potential additional selection and amplification of resistant isolates. Similar measures can be taken with fungicides with new modes of action in agronomic settings. He added that the integrated disease management approach taken in addressing fungicide resistance for target pathogens in the crop itself is unlikely to address hotspots in agricultural waste. For example, the alternation of using fungicides with different modes of action against target pathogens in crops will not necessarily extend to effective management of resistance in waste piles of non-target organisms. Doughty maintained that understanding crop waste management processes and options will be foundational in developing prevention and mitigation strategies and in informing future decisions of how fungicides are positioned in agronomic settings.

DISCUSSION

Overlap in Agriculture and Clinical Use Considerations

Brewer asked whether the use of antifungals in the clinic and environment can be safe under specific conditions. Fisher replied that only a small change in the binding site can confer resistance. Therefore, it is essential to identify where the fungicide is binding to in comparison to the clinical antifungal drug. For example, cross-resistance problems can occur if a DHODH inhibitor in both the clinical and agricultural product is affecting the exact same space on the enzyme. However, any difference in the drug-target binding could potentially avoid issues that can arise with this overlap in use. Binding studies are critical in understanding the drug-target interaction, and experimental evolution can be used to proactively learn about potential cross-resistance, said Fisher. In cases where cross-resistance is not observed, the overlapping use in agricultural and clinical settings can be safe. Doughty noted the dilemma of achieving twin aims of human health protection and food security. New modes of action are pressing needs in both medicine and agriculture to maintain resistance management strategies and to have effective tools to address infection. Doughty stated that if a DHODH inhibitor is introduced into the fungicides market, it will be important to know whether it is active against A. fumigatus and whether the specific intended use of the DHODH fungicide will create residues to which A. fumigatus will be exposed. Understanding both the resistance

profile of the human pathogenic fungi and exposure scenarios are the starting point for avoiding problematic use cases, he added.

Mutations in Aspergillus fumigatus

Given that tandem repeat (TR) mutations have not been described for plant pathogens occurring in crops regularly treated with azole fungicides, Brewer asked for an explanation about why A. fumigatus have the TR₃₄ and TR₄₆ mutations that lead to pan-azole resistance. Fisher acknowledged that he does not have an explanation, but that it is too soon to say that no tandem repeats are occurring in plant pathogens afflicting crops. Far more genome sequencing has been performed on human fungal pathogens than on agricultural ones, thus tandem repeats may exist in crops but have not yet been discovered. He noted that A. fumigatus has a wide array of tandem repeats in environmental settings, indicating a proclivity for this activity, yet this does not seem to occur in de novo evolution in the lungs during infection. The reason for this is not yet understood, Fisher added.

Fungicide Risk Assessment for Resistance

Given that A. fumigatus is not a plant pathogen and that issues of A. fumigatus resistance are a downstream effect of the introduction of fungicide to the environment, Chiller asked how fungicides can be evaluated for risk of resistance and effects that ultimately affect human health. Doughty replied that the regulation of new agricultural tools should be based on understanding of the intrinsic activity of a new molecule and the risk of exposure of human patients. The European Commission has issued a mandate to review this situation for azole fungicides with regard to ARAf. In coming years, discussions about how policy will approach the regulation of pesticides may include consideration of the risk of selection of resistance for human pathogens, said Doughty. Chiller commented on the importance of these discussions in light of the balance needed in safeguarding human health and food supply. He added that experts in human health who are concerned about the potential for new fungicides to cross-react with new antifungals—or even antibacterials—need to take the critical role of fungicides in agriculture into account.

Routes of Transmission of Azole-Resistant Aspergillus fumigatus

Brewer asked about the relationship between hotspots and transmission—e.g., whether people working or living near resistance hotspots are at an increased risk of contracting a resistant fungal infection. Doughty replied that he is not aware of direct transmission to humans

working in tulip fields (see Chapter 8 for further detail). Fisher stated that Shelton’s community science survey results were used to build numerous statistical models with environmental variables to use in identifying associations (Shelton, 2021). The only variable that demonstrated significance in the risk of potential transmission was industrial composters. Green waste recycling is now taking place on a large scale, with approximately 350 industrial composters in the United Kingdom. However, no association was found in terms of the amount of ARAf. Fisher said this may be due to the low number of azole-resistant isolates, which warrants further research given the association of industrial composters with high burdens of A. fumigatus.

Brewer asked about the routes of transmission of ARAf to humans, whether these routes include sporulation occurring on food, and whether azole-resistant isolates are associated with different types of food. Fisher stated that since agricultural waste piles have been identified as hotspots, he is curious about whether small countertop compost bins in household kitchens could also become hotspots. Relatively high fungicide residues could be expected in a bin containing fruit and vegetable peelings. Given that many people spend up to 90 percent of their lives indoors, Fisher and colleagues conducted a small community science indoor surveillance and found more azole resistance indoors than outdoors (Shelton et al., 2022). The numbers were small, but the finding suggests more research on indoor air is warranted. The relative abundance of spores was very similar in indoor and outdoor air, with the former heavily biased to molds and the latter biased to yeasts. Thus, moving from outdoors to indoors involves exposure to both yeast-dominated and mold-dominated aerobiomes. Fisher noted that this surveillance was conducted in the United Kingdom, a damp climate that promotes mold in house. During a recent trip to Singapore, a humid climate, he noticed that indoor air filtration is common. In the United Kingdom, air cleaning filtration systems were uncommon until they grew in popularity in response to the COVID-19 pandemic. Although air filtration systems can remove spores from the air, filters that are not regularly cleaned can act as growth surfaces; the systems then pump new spores into the air.

Doughty stated that Aspergillus—including resistant genotypes—is found on food commodities. For instance, resistant Aspergillus can be found on fruits in the supermarket. Moreover, Aspergillus can be found in coffee samples in spite of the coffee having been roasted at several hundred degrees. This link is possibly disconnected from fungicide use in the field, as food commodities could come into contact with Aspergillus during storage, transport, or processing. For example, coffee is particularly susceptible to Aspergillus contamination during sea transport. Doughty remarked on the importance of understanding the relation between the use of azole

fungicides in crops and the appearance of resistant isolates on food commodities at point of sale.

Chiller asked whether the development of resistant strains in humans is related to locations where azoles have been applied in the environment and whether foods that have been treated with azole during production carry resistant strains through the supply chain. Fisher responded that research on aerosols indicates that dispersal from hotspots happens rapidly, and sampling in the United Kingdom indicates broad, low levels across the country. Precise epidemiology is required to locate hotspots, although certain crops such as bulbs are well known for heavy azole use and Aspergillus presence between their layers. A huge global industry ships bulbs around the world, with the Netherlands exporting billions of bulbs annually. International trade enables resistant strains to become globalized easily; Aspergillus spores then become aerosolized and spread throughout the importing countries. Doughty also described research on strains isolated from individuals and from their immediate surroundings that found no correlation between genotypes from patients and from nearby fields in terms of tandem repeat and single nucleotide polymorphism profile (Rocchi et al., 2014). However, a clear link was found between the genotypes recovered from patients and from their garden compost heaps. This carries the implication that the compost heap is a potential source of infection, whereas the surrounding fields of maize, wheat, and barley are not.

Coldspot Crop Management

Chiller asked how the cereal crop coldspots in the United Kingdom or other testing locations managed crops in terms of crop residue and whether strategies such as no-till were used. Doughty recalled research in which samples from the long-term Rothamsted field trials—which began in the 1800s—were compared with various types of treatment on cereal plots (Fraaije et al., 2020). The comparison examined samples from plots treated with azoles since their market introduction, plots that had not been treated, and grassland. With regard to whether no-till strategy was used with straw crops, Doughty was uncertain.

Factors Promoting Azole Resistance

Brewer asked whether some azoles are more inclined to promote resistance than others, given that agricultural azole use began in the 1970s, but ARAf has increased significantly in the past two decades. Fisher remarked that fungicide azoles vary widely: their chemical structure can be long-tailed or short-tailed; they can have slightly different modes of action; and their resistance profiles vary in similarity to clinical azoles. In addition to these

factors, azoles are used in much higher quantities now than they were in the 1990s and decades prior. Thus, the force of selection is greater and more widely applied. Doughty referenced a study examining a group of azoles most structurally similar to medical triazoles (Snelders et al., 2012). The study found that homogeneity in the mode of action at the active site seemed to indicate that this group was particularly relevant to the selection of resistant isolates in the environment. Individual azoles vary greatly in terms of Aspergillus activity, thus the properties of each individual compound should be examined, said Doughty.

REFLECTIONS ON DAY ONE

Paige Waterman, interim chair of medicine and vice chair for clinical research at the F. Edward Herbert School of Medicine at the Uniformed Services University of the Health Sciences, Bethesda, and Jeff LeJeune, food safety officer in the Food Systems and Food Safety Division of the Food and Agriculture Organization of the United Nations, offered reflections on the first day of the workshop. The workshop began with an overview of fungal diseases within humans and the effect of rising environmental temperatures—in conjunction with lower human temperatures—as a natural driver of the increasingly adapted fungal pathogens with the potential to wreak havoc. Efforts of the global Quadripartite Joint Secretariat on Antimicrobial Resistance (AMR) to connect work, and in particular surveillance, in the One Health, AMR, and antifungal resistance sphere were highlighted. Existing mechanisms for fungicide tracking, use, and resistance within the United States were described. Meaningful responses to antifungal use and resistance—whether coordinated or not—are lagging in connection with the human tendency to focus on familiar or recent threats rather than on emerging ones. Speakers offered examples of deadly consequences of fungal infections and highlighted the extent to which fungi strains resistant to antifungals remain problematic to diagnose and treat.

The second panel discussed the complexities inherent in developing new antifungal drugs, including the “antihuman” properties of antifungals due to the similarities between the fungal and animal kingdoms, said Waterman. Increasingly, fungal pathogens cause systemic infections with high morbidity and mortality that can affect people who are not immunocompromised, fueling the need for new drugs. Antifungal resistance is also rising, driven in some countries by the environment. The reliance on azole fungicides for crops such as corn, soy, and wheat and the spread of resistance are factors to balance. Additionally, challenges in diagnosis factor into stewardship efforts in both the agricultural and medical settings. Speakers discussed the evolving nature of fungi and specific fungal pathogens, including the somewhat unexpected emergence of Candida auris.

Azole resistance in general and A. fumigatus were the focus of the third panel of the workshop. Community science research and compost studies have demonstrated the ubiquity of aerosolized exposure of Aspergillus. Waterman remarked that the large number of settings meeting the requirements of resistance hotspots warrants further study. Fungal pathogens are ubiquitous in nature and commonly develop resistance—driven by both environmental factors as well as the frequent use of azoles—with implications for both food security and human health. The fungal pathogen transfer from plants to humans may not be as uncommon as previously thought, further fueling the need to coordinate and develop surveillance, detection, diagnosis, risk assessment, mitigation, and therapeutics efforts.

LeJeune commented on the disease triad involving effects of pathogens, humans, and the environment on disease manifestation. The first day of the workshop discussed fungal diseases in humans, fungal agents, the ramifications of aspergillosis and candidiasis, and modern molecular epidemiology and medicine used to characterize and treat these infections. Changes in human susceptibility, immunity, and even temperature factor into increasing infection rates. Environmental hotspots may be generating fungal pathogens, and the United States is currently monitoring agricultural use of antifungal agents, accumulation of residues, and crude commodities. Speakers explored the interplay of the environment, climate crisis, and the manifestation of fungal diseases, as well as the underrepresentation of fungal diseases in research and public awareness.