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Evolving moldy murderers: Aspergillus section Fumigati as a model for studying the repeated evolution of fungal pathogenicity

Evolving moldy murderers: Aspergillus section Fumigati as a model for studying the repeated evolution of fungal pathogenicity

  • Antonis Rokas, 
  • Matthew E. Mead, 
  • Jacob L. Steenwyk, 
  • Nicholas H. Oberlies, 
  • Gustavo H. Goldman
PLOS
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Biology is more like history than it is like physics. You have to know the past to understand the present.”–Carl Sagan (1980)

Introduction

Species in the genus Aspergillus are saprophytic filamentous fungi that are most commonly found in soil and litter environments of subtropical and warm temperate latitudes [1]. Inhalation of asexual spores produced by Aspergillus fumigatus and a few other species in the genus cause a group of diseases collectively referred to as aspergillosis [2]. The most severe form of aspergillosis is invasive aspergillosis, which primarily affects individuals with compromised immune systems or preexisting lung conditions [3]. Since drugs targeting invasive aspergillosis are not always effective due to our lack of understanding of how they function inside the human host [4] and the evolution of drug resistance [5, 6], infected individuals suffer high morbidity and mortality [7]. Collectively, Aspergillus fungi affect millions of patients and cause hundreds of thousands of life-threatening infections every year [8, 9].

Not all pathogenic Aspergillus species exhibit the same infection rates [10, 11]. Approximately 70% of all Aspergillus infections are caused by A. fumigatus, whereas the remaining 30% of infections stem from other species in the genus [12]. Some of these other pathogenic species–for example, Aspergillus flavus, Aspergillus niger, and Aspergillus terreus–are distantly related to A. fumigatus [12]; each of these three species belong to different Aspergillus sections (note “section” is a taxonomic rank in-between the genus and species ranks) and show extensive genomic divergence [13]. However, there are also pathogenic Aspergillus species, such as Aspergillus lentulus and Aspergillus udagawae, that belong to the same section as A. fumigatus (section Fumigati) and are much more closely related [1416]. In contrast, the vast majority of Aspergillus species, including many close relatives of the pathogens mentioned above, are either not pathogenic or very rarely cause disease [16, 17]. Interestingly, A. fumigatus and a few other species in the genus can also cause opportunistic infections, mainly in other mammals and birds, and occasionally in other vertebrates and invertebrates [18].

Even though a great deal is known about some aspects of A. fumigatus pathogenicity [2, 19, 20], we have only recently begun to examine why pathogenicity varies so dramatically across the entire genus and the traits and genetic elements that contributed to this variation. Addressing this question requires that we consider the fact that pathogenic Aspergillus species are not dependent on their hosts for survival and their pathogenic effects are entirely accidental or opportunistic. Thus, understanding the evolution of pathogenicity in the genus requires that we understand how variation in the traits that enable Aspergillus species to survive in their natural soil and litter environments has rendered a few of these species capable to establish infections inside human hosts.

In this pearl article, we focus on section Fumigati, a lineage of ~60 species that includes A. fumigatus and its close relatives [16, 21], to discuss the latest advances in our understanding of the evolution of pathogenicity in Aspergillus. More broadly, given that the ability to cause human disease has repeatedly evolved across the fungal tree of life, and that the vast majority of human fungal pathogens have non-pathogenic close relatives [22], understanding the evolution of Aspergillus pathogenicity can serve as a model for studying fungal pathogenicity in general.

Pathogenicity in Aspergillus section Fumigati fungi has evolved multiple times independently

Only a handful of species in section Fumigati are considered pathogenic [10, 16, 21], and the distribution of these pathogens on the section’s phylogeny [16] suggests that the ability to cause human disease has evolved at least 5 times independently (Fig 1). For example, whereas A. fumigatus infects >300,000 humans per year [9], its close evolutionary relative A. fischeri, whose protein sequences exhibit, on average, 95% similarity to their A. fumigatus orthologs, has only rarely been reported to cause human disease and is not considered clinically relevant [23, 24]. Interestingly, evolutionary reconstruction shows that A. fumigatus pathogenicity likely evolved after the species diverged from either A. fischeri or from its even closer non-pathogenic relative A. oerlinghausenensis, suggesting that the last common ancestor of these three species was non-pathogenic (Fig 1). The same is true for most other pathogens in the section. For example, the pathogenic A. udagawae accounts for a few thousand infections per year [2527], but its close relatives (e.g., Aspergillus aureolus, Aspergillus acrensis, and Aspergillus wyomingensis) are not considered clinically relevant (Fig 1), suggesting that the pathogenicity of A. udagawae evolved independently and that the common ancestor of A. udagawae and its close relatives was non-pathogenic.

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Fig 1. Reconstruction of the ability to cause human disease on the Aspergillus section Fumigati phylogeny suggests that pathogenicity evolved multiple times independently in the lineage.

For the trait reconstruction inference, Biosafety Level (BSL) 2 organisms were considered pathogenic and BSL1 organisms or organisms that so far lack BSL labelling were considered non-pathogenic; these transitions to a pathogenic lifestyle (i.e., from BSL1 to BSL2) are labelled by red bars on the figure. Note that clinical isolates from humans or other mammals from a few additional species in the section have been identified [10, 28]; this handful includes relatively newly described species that some authors consider to have pathogenic potential (e.g., Aspergillus novofumigatus [29]) as well as organisms thought to be on the non-pathogenic end of the spectrum (e.g., A. fischeri [30, 31]). The phylogeny of the section was redrawn from Hubka et al. [16].

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The observed spectrum of pathogenicity cannot be explained by ecology or ascertainment bias and is likely to have a genetic component

Several ecological attributes, such as the global ubiquity of their small and readily airborne asexual spores, are thought to contribute to the pathogenicity of Aspergillus molds in general [11], and to the pathogenicity of A. fumigatus in particular [19, 32]. Although these ecological attributes are undoubtedly important for infecting and causing disease in a human host, the observed spectrum of pathogenicity among section Fumigati species cannot be solely explained by the known differences in species’ ecologies [1, 3335]. For example, previous studies have shown that A. fischeri, one of the closest non-pathogenic relatives of A. fumigatus, can be frequently isolated from a variety of locales, including soils, fruits, and hospitals [3335]. Case in point, approximately 2% of the fungi isolated from the Beijing Hospital environment were A. fischeri [34], but only a handful of infections caused by this fungus have ever been reported [10, 23, 24].

Another ecological attribute known to be associated with fungal pathogenicity is thermal tolerance [36]. However, all species in section Fumigati that have been tested can grow, dependent on the growth medium used, at 37°C [14]. These data suggest that pathogenicity in Aspergillus section Fumigati is not simply due to species’ abilities to grow at the human body temperature. Nevertheless, species in the section do show substantial differences in how well they can grow at 37°C [14], but these differences are likely to have a genetic basis (see below). It would be highly interesting for future studies to examine growth curves of closely related pathogenic and non-pathogenic species in specific stressful and human infection-relevant conditions (e.g., at 37°C, with limited nutrient availability, low levels of oxygen and high levels of oxidative stress).

Another potential explanation for some of the observed differences in the spectrum of pathogenicity among species is ascertainment bias. In the context of Aspergillus pathogenicity, ascertainment bias is a term that describes systematic deviations from the true incidence of disease caused by a given species. These systematic deviations stem from the methods used to taxonomically identify (ascertain) clinical isolates and estimate how often they cause disease, i.e., from our failure to measure the true numbers of infections caused by so-called cryptic species, namely organisms that are morphologically similar to major pathogens, such as A. fumigatus, but genetically distinct from them [10]. The true burden some of these cryptic species, including species currently thought to not be clinically relevant, place on human health is unknown and may be in several cases underestimated [22]. However, numerous molecular typing studies of clinical isolates from diverse countries routinely identify the known pathogens in section Fumigati (Fig 1), but not the non-pathogens [2527], indicating that the observed variation in pathogenicity is not solely an artifact of species misdiagnosis. These data suggest that the differences in pathogenicity observed across Aspergillus section Fumigati have, at least partially, a genetic basis.

Support for the role of genetic differences in contributing to the observed spectrum of pathogenicity is provided by the numerous traits, and their underlying genes and pathways, that are required for pathogenicity in A. fumigatus [2, 19, 20, 37] and have been found to exhibit substantial genetic and phenotypic diversity among section Fumigati species. These traits include thermotolerance, the ability to respond to multiple environmental stresses, including antifungal drugs, and the capacity to biosynthesize a range of structurally diverse secondary metabolites [14, 30, 38, 39].

Two models for the evolution of pathogenicity in Aspergillus molds

One useful approach for gaining insights into the genetic foundations of the multiple, independent origins of pathogenicity in Aspergillus section Fumigati is the development of conceptual models that describe the differences that we would expect to observe in genomic comparisons involving pathogenic and non-pathogenic species. We propose two alternative, although not necessarily mutually exclusive, models, which we have named the “conserved pathogenicity” model and the “species-specific pathogenicity” model (Fig 2). The conserved pathogenicity model posits that A. fumigatus and other pathogenic species in section Fumigati share common pathogenicity traits and genetic elements (or shared differences in genetic elements) that are absent in non-pathogens (e.g., traits / elements E1 and E2 in Fig 2) or vice versa (e.g., trait / element E3 in Fig 2). In contrast, the species-specific model posits that each pathogen contains a unique suite of traits and genetic elements (or unique differences in genetic elements) that distinguish it from its non-pathogenic relatives; these traits / elements could be ones that are uniquely present in a given pathogen but absent in related pathogens and non-pathogens (e.g., traits / elements E4 and E5 in Fig 2) or vice versa (e.g., trait / element E6 in Fig 2). Note that these shared or species-specific genetic elements (or differences in genetic elements) are not limited to differences in gene content but to any type of genetic variation that alters pathogenicity trait values. These variants can range from, for example, differences in a single or in a handful of nucleotides within otherwise conserved protein-coding or non-coding (regulatory) regions to larger-scale differences concerning the presence of entire genetic pathways and networks.

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Fig 2. Graphic illustration of the “conserved pathogenicity” and the “species-specific” pathogenicity models.

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Discerning which model explains the repeated evolution of pathogenicity is key for developing research strategies to understand the underlying molecular mechanisms in the genus and, more broadly, in filamentous fungi and beyond. For example, the conserved pathogenicity model would predict that pathogenicity stems from the action of conserved genetic elements, suggesting that known genetic determinants of virulence in A. fumigatus [40] would be great candidates for involvement in virulence in other pathogenic Aspergillus species. It is the adoption of the conserved pathogenicity model that underlies recent examinations of the degree to which genetic elements known to contribute to A. fumigatus pathogenicity are conserved in other species [29, 30, 41]. In contrast, the species-specific model would predict the opposite, namely that the genetic determinants of virulence are unique to each pathogen, suggesting that extrapolations of knowledge on virulence mechanisms from one pathogenic species to another would be futile. Genomic comparisons between the major pathogen A. fumigatus and its close non-pathogenic relative A. fischeri [30, 31] as well as broad comparisons of select species across the genus [29, 41] have begun to shed light on the validity of, and provide support for, both of these models.

Support for the conserved pathogenicity model

Consistent with one of the predictions of the conserved pathogenicity model, examinations of dozens of A. fumigatus genes known to be associated with virulence [40] have shown that most of these genetic determinants of virulence are highly conserved in closely related species [29, 30]. For example, a recent genomic comparison of the major pathogen A. fumigatus with its close, non-pathogenic relative A. fischeri showed that 48 of 49 known genetic determinants of A. fumigatus virulence (e.g., CrzA, the C2H2-type zinc finger transcription factor involved in calcium ion homeostasis, or LaeA, a methyltransferase and master regulator of secondary metabolism) were highly conserved in A. fischeri [30]. However, these results also suggested that the differences in virulence among organisms spanning the pathogenicity spectrum may not be primarily due to differences in gene content, which virtually all genomic comparisons of fungal pathogens and non-pathogens in the genus [2931, 41] and beyond [4244] have heavily focused on. How these conserved genetic determinants of virulence function in other pathogenic, as well as in non-pathogenic, species is an interesting future direction of inquiry.

Support for the species-specific pathogenicity model

Comparisons of gene content between closely related Aspergillus species have also identified numerous genes that appear to be species-specific. For example, a broad scale comparison of A. fumigatus strains Af293 and A1163 against A. fischeri and A. clavatus found that approximately 8.5% of genes were unique to A. fumigatus and absent from the other two species [31]. These A. fumigatus-specific genes tend to reside near the ends of chromosomes (i.e., are subtelomeric) and have functions associated with metabolism (e.g., secondary metabolism, transport, and detoxification), raising the hypothesis that some of them may aid A. fumigatus survival inside the human host [31]. For example, more than two thirds of A. fumigatus biosynthetic gene clusters are absent from the closely related non-pathogen A. fischeri [30, 38]; however some of these biosynthetic gene clusters are found in other species in section Fumigati, suggesting that they were lost in A. fischeri rather than originated within A. fumigatus. Several other clusters and their secondary metabolites appear to have uniquely evolved in A. fumigatus, or their loss in species closely related to A. fumigatus was so widespread that they are now uniquely present in A. fumigatus.

One recent, striking example in support of the species-specific model was the discovery of the gene hypoxia-responsive morphology factor A or hrmA (Afu5g14900) [45]. While homologs of hmrA are present in other distantly related fungi, this subtelomeric gene is polymorphic within A. fumigatus and is absent from the genomes of all sequenced Aspergillus section Fumigati species. Investigation of hrmA function shows that it likely regulates a cluster of genes, which also appear to be absent from the genomes of other Aspergillus section Fumigati species, that collectively contribute to the generation of a morphotype that facilitates adaptation to very low oxygen conditions encountered by the fungus inside human lungs [45].

Concluding remarks

This pearl has focused on Aspergillus section Fumigati, outlining two general models for the repeated evolution of pathogenicity in the section and how they could aid in the design of experiments aimed at elucidating the underlying molecular changes responsible. The same approach could also be employed for developing strategies for the burgeoning problem of drug resistance (e.g., to what extent are mechanisms of drug resistance conserved across pathogenic species?). But the utility of studying the evolution of Aspergillus section Fumigati pathogenicity goes beyond, as Carl Sagan so aptly put it, understanding “the present”. By posing questions such as “are species that are currently considered non-pathogenic but contain conserved genetic determinants of virulence likely to emerge as new pathogens?”, we believe that an evolutionary approach–by identifying the presence of constellations of genes and traits associated with pathogenicity in non-pathogens–also holds promise for predicting the emergence of new pathogens in the future.

Acknowledgments

We thank our collaborators and members of the Rokas, Goldman, and Oberlies labs for their feedback and support in our studies on the evolution of Aspergillus pathogenicity.

References

  1. 1. Klich MA. Biogeography of Aspergillus species in soil and litter. Mycologia. 2002;94(1):21–7. ISI:000173697600003. pmid:21156474
  2. 2. Latge JP, Steinbach WJ, editors. Aspergillus fumigatus and Aspergillosis. Washington, DC: ASM Press; 2009.
  3. 3. Segal BH. Aspergillosis. N Engl J Med. 2009;360(18):1870–84. Epub 2009/05/01. 360/18/1870 [pii] pmid:19403905.
  4. 4. Rosowski EE, He J, Huisken J, Keller NP, Huttenlocher A. Efficacy of voriconazole against A. fumigatus infection depends on host immune function. Antimicrob Agents Chemother. 2019. pmid:31740552.
  5. 5. Fisher MC, Hawkins NJ, Sanglard D, Gurr SJ. Worldwide emergence of resistance to antifungal drugs challenges human health and food security. Science. 2018;360(6390):739–42. pmid:29773744.
  6. 6. Revie NM, Iyer KR, Robbins N, Cowen LE. Antifungal drug resistance: evolution, mechanisms and impact. Curr Opin Microbiol. 2018;45:70–6. pmid:29547801; PubMed Central PMCID: PMC6135714.
  7. 7. Brown GD, Denning DW, Gow NA, Levitz SM, Netea MG, White TC. Hidden killers: human fungal infections. Sci Transl Med. 2012;4(165):165rv13. pmid:23253612.
  8. 8. Denning DW. Calling upon all public health mycologists: To accompany the country burden papers from 14 countries. Eur J Clin Microbiol Infect Dis. 2017;36(6):923–4. pmid:28150046.
  9. 9. Bongomin F, Gago S, Oladele RO, Denning DW. Global and multi-national prevalence of fungal diseases-estimate precision. J Fungi (Basel). 2017;3(4):57. pmid:29371573; PubMed Central PMCID: PMC5753159.
  10. 10. Lamoth F. Aspergillus fumigatus-related species in clinical practice. Front Microbiol. 2016;7:683. pmid:27242710; PubMed Central PMCID: PMC4868848.
  11. 11. Paulussen C, Hallsworth JE, Alvarez-Perez S, Nierman WC, Hamill PG, Blain D, et al. Ecology of aspergillosis: insights into the pathogenic potency of Aspergillus fumigatus and some other Aspergillus species. Microb Biotechnol. 2017;10(2):296–322. pmid:27273822; PubMed Central PMCID: PMC5328810.
  12. 12. Steinbach WJ, Marr KA, Anaissie EJ, Azie N, Quan SP, Meier-Kriesche HU, et al. Clinical epidemiology of 960 patients with invasive aspergillosis from the PATH Alliance registry. J Infect. 2012;65(5):453–64. pmid:22898389.
  13. 13. Rokas A, Galagan JE. The Aspergillus nidulans genome and a comparative analysis of genome evolution in Aspergillus. In: Goldman GH, Osmani SA, editors. The Aspergilli: Genomics, Medical Applications, Biotechnology, and Research Methods. Boca Raton, FL: CRC Press; 2008. p. 43–55.
  14. 14. Samson RA, Hong S, Peterson SW, Frisvad JC, Varga J. Polyphasic taxonomy of Aspergillus section Fumigati and its teleomorph Neosartorya. Stud Mycol. 2007;59:147–203. pmid:18490953; PubMed Central PMCID: PMC2275200.
  15. 15. Steenwyk JL, Shen XX, Lind AL, Goldman GH, Rokas A. A robust phylogenomic time tree for biotechnologically and medically important fungi in the genera Aspergillus and Penicillium. mBio. 2019;10(4). pmid:31289177; PubMed Central PMCID: PMC6747717.
  16. 16. Hubka V, Barrs V, Dudova Z, Sklenar F, Kubatova A, Matsuzawa T, et al. Unravelling species boundaries in the Aspergillus viridinutans complex (section Fumigati): opportunistic human and animal pathogens capable of interspecific hybridization. Persoonia. 2018;41:142–74. pmid:30728603; PubMed Central PMCID: PMC6344812.
  17. 17. Kocsube S, Perrone G, Magista D, Houbraken J, Varga J, Szigeti G, et al. Aspergillus is monophyletic: Evidence from multiple gene phylogenies and extrolites profiles. Stud Mycol. 2016;85:199–213. pmid:28082760; PubMed Central PMCID: PMC5220211.
  18. 18. Seyedmousavi S, Guillot J, Arne P, de Hoog GS, Mouton JW, Melchers WJ, et al. Aspergillus and aspergilloses in wild and domestic animals: a global health concern with parallels to human disease. Med Mycol. 2015;53(8):765–97. pmid:26316211.
  19. 19. Kwon-Chung KJ, Sugui JA. Aspergillus fumigatus-what makes the species a ubiquitous human fungal pathogen? PLoS Pathog. 2013;9(12). e1003743 10.1371/journal.ppat.1003743. WOS:000330535400002.
  20. 20. Raffa N, Keller NP. A call to arms: Mustering secondary metabolites for success and survival of an opportunistic pathogen. PLoS Pathog. 2019;15(4):e1007606. pmid:30947302.
  21. 21. Sugui JA, Peterson SW, Figat A, Hansen B, Samson RA, Mellado E, et al. Genetic relatedness versus biological compatibility between Aspergillus fumigatus and related species. J Clin Microbiol. 2014;52(10):3707–21. pmid:25100816; PubMed Central PMCID: PMC4187787.
  22. 22. Konopka J, Casadevall A, Taylor J, Heitman J, Cowen L. One Health: Fungal Pathogens of Humans, Animals, and Plants. American Academy of Microbiology, 2019.
  23. 23. Lonial S, Williams L, Carrum G, Ostrowski M, McCarthy P Jr. Neosartorya fischeri: an invasive fungal pathogen in an allogeneic bone marrow transplant patient. Bone Marrow Transplant. 1997;19(7):753–5. pmid:9156256.
  24. 24. Coriglione G, Stella G, Gafa L, Spata G, Oliveri S, Padhye AA, et al. Neosartorya fischeri var fischeri (Wehmer) Malloch and Cain 1972 (anamorph: Aspergillus fischerianus Samson and Gams 1985) as a cause of mycotic keratitis. European journal of epidemiology. 1990;6(4):382–5. pmid:2091938.
  25. 25. Balajee SA, Kano R, Baddley JW, Moser SA, Marr KA, Alexander BD, et al. Molecular identification of Aspergillus species collected for the Transplant-Associated Infection Surveillance Network. J Clin Microbiol. 2009;47(10):3138–41. Epub 2009/08/14. pmid:19675215; PubMed Central PMCID: PMC2756904.
  26. 26. Alastruey-Izquierdo A, Mellado E, Pelaez T, Peman J, Zapico S, Alvarez M, et al. Population-based survey of filamentous fungi and antifungal resistance in Spain (FILPOP Study). Antimicrob Agents Chemother. 2013;57(7):3380–7. pmid:23669377; PubMed Central PMCID: PMC3697314.
  27. 27. Negri CE, Goncalves SS, Xafranski H, Bergamasco MD, Aquino VR, Castro PT, et al. Cryptic and rare Aspergillus species in Brazil: prevalence in clinical samples and in vitro susceptibility to triazoles. J Clin Microbiol. 2014;52(10):3633–40. pmid:25078909; PubMed Central PMCID: PMC4187744.
  28. 28. Sugui JA, Kwon-Chung KJ, Juvvadi PR, Latge JP, Steinbach WJ. Aspergillus fumigatus and related species. Cold Spring Harb Perspect Med. 2014;5(2):a019786. pmid:25377144; PubMed Central PMCID: PMC4315914.
  29. 29. Kjaerbolling I, Vesth TC, Frisvad JC, Nybo JL, Theobald S, Kuo A, et al. Linking secondary metabolites to gene clusters through genome sequencing of six diverse Aspergillus species. Proc Natl Acad Sci U S A. 2018;115(4):E753–E61. pmid:29317534; PubMed Central PMCID: PMC5789934.
  30. 30. Mead ME, Knowles SL, Raja HA, Beattie SR, Kowalski CH, Steenwyk JL, et al. Characterizing the pathogenic, genomic, and chemical traits of Aspergillus fischeri, a close relative of the major human fungal pathogen Aspergillus fumigatus. mSphere. 2019;4(1):e00018–19. pmid:30787113; PubMed Central PMCID: PMC6382966.
  31. 31. Fedorova ND, Khaldi N, Joardar VS, Maiti R, Amedeo P, Anderson MJ, et al. Genomic islands in the pathogenic filamentous fungus Aspergillus fumigatus. PLoS Genet. 2008;4(4):e1000046. Epub 2008/04/12. pmid:18404212; PubMed Central PMCID: PMC2289846.
  32. 32. Tekaia F, Latge JP. Aspergillus fumigatus: saprophyte or pathogen? Curr Opin Microbiol. 2005;8(4):385–92. Epub 2005/07/16. S1369-5274(05)00092-5 [pii] pmid:16019255.
  33. 33. Hong SB, Kim DH, Park IC, Samson RA, Shin HD. Isolation and identification of Aspergillus section Fumigati strains from arable soil in Korea. Mycobiology. 2010;38(1):1–6. pmid:23956617; PubMed Central PMCID: PMC3741588.
  34. 34. Tong XL, Xu HT, Zou LH, Cai M, Xu XF, Zhao ZT, et al. High diversity of airborne fungi in the hospital environment as revealed by meta-sequencingbased microbiome analysis. Sci Rep-Uk. 2017;7. ARTN 39606 WOS:000391026800001. pmid:28045065
  35. 35. Frąc M, Jezierska-Tys S, Yaguchi T. Occurrence, detection, and molecular and metabolic characterization of heat-resistant fungi in soils and plants and their risk to human health. Advances in Agronomy. 2015;132:161–204.
  36. 36. Robert V, Cardinali G, Casadevall A. Distribution and impact of yeast thermal tolerance permissive for mammalian infection. BMC Biol. 2015;13:18. pmid:25762372; PubMed Central PMCID: PMC4381509.
  37. 37. Brown NA, Goldman GH. The contribution of Aspergillus fumigatus stress responses to virulence and antifungal resistance. J Microbiol. 2016;54(3):243–53. pmid:26920884.
  38. 38. Frisvad JC, Larsen TO. Extrolites of Aspergillus fumigatus and other pathogenic species in Aspergillus section Fumigati. Front Microbiol. 2015;6:1485. pmid:26779142; PubMed Central PMCID: PMC4703822.
  39. 39. Alastruey-Izquierdo A, Alcazar-Fuoli L, Cuenca-Estrella M. Antifungal susceptibility profile of cryptic species of Aspergillus. Mycopathologia. 2014;178(5–6):427–33. pmid:24972670.
  40. 40. Abad A, Victoria Fernandez-Molina J, Bikandi J, Ramirez A, Margareto J, Sendino J, et al. What makes Aspergillus fumigatus a successful pathogen? Genes and molecules involved in invasive aspergillosis. Revista Iberoamericana de Micologia. 2010;27(4):155–82. Epub 2010/10/27. S1130-1406(10)00089-6 [pii] pmid:20974273.
  41. 41. de Vries RP, Riley R, Wiebenga A, Aguilar-Osorio G, Amillis S, Uchima CA, et al. Comparative genomics reveals high biological diversity and specific adaptations in the industrially and medically important fungal genus Aspergillus. Genome Biol. 2017;18(1):28. pmid:28196534.
  42. 42. Butler G, Rasmussen MD, Lin MF, Santos MA, Sakthikumar S, Munro CA, et al. Evolution of pathogenicity and sexual reproduction in eight Candida genomes. Nature. 2009;459(7247):657–62. Epub 2009/05/26. nature08064 [pii] pmid:19465905.
  43. 43. Martinez DA, Oliver BG, Graser Y, Goldberg JM, Li W, Martinez-Rossi NM, et al. Comparative genome analysis of Trichophyton rubrum and related dermatophytes reveals candidate genes involved in infection. mBio. 2012;3(5):e00259–12. pmid:22951933; PubMed Central PMCID: PMC3445971.
  44. 44. Sharpton TJ, Stajich JE, Rounsley SD, Gardner MJ, Wortman JR, Jordar VS, et al. Comparative genomic analyses of the human fungal pathogens Coccidioides and their relatives. Genome Res. 2009;19(10):1722–31. pmid:19717792; PubMed Central PMCID: PMC2765278.
  45. 45. Kowalski CH, Kerkaert JD, Liu KW, Bond MC, Hartmann R, Nadell CD, et al. Fungal biofilm morphology impacts hypoxia fitness and disease progression. Nat Microbiol. 2019. pmid:31548684.