Novel Taxa of Thermally Dimorphic Systemic Pathogens in the Ajellomycetaceae (Onygenales)
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Received: 3 November 2016 | Revised: 30 December 2016 | Accepted: 30 December 2016 DOI: 10.1111/myc.12601 ORIGINAL ARTICLE Novel taxa of thermally dimorphic systemic pathogens in the Ajellomycetaceae (Onygenales) Karolina Dukik1,2† | Jose F. Muñoz3,4,5† | Yanping Jiang1,6 | Peiying Feng1,7 | Lynne Sigler8 | J. Benjamin Stielow1,9 | Joanna Freeke1,9 | Azadeh Jamalian1,9 | Bert Gerrits van den Ende1 | Juan G. McEwen4,10 | Oliver K. Clay4,11 | Ilan S. Schwartz12,13 | Nelesh P. Govender14,15 | Tsidiso G. Maphanga15 | Christina A. Cuomo3 | Leandro F. Moreno1,2,16 | Chris Kenyon14,17 | Andrew M. Borman18 | Sybren de Hoog1,2 1CBS-KNAW Fungal Biodiversity Centre, Utrecht, The Netherlands 2Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, Amsterdam, The Netherlands 3Broad Institute of MIT and Harvard, Cambridge, MA, USA 4Cellular and Molecular Biology Unit, Corporación para Investigaciones Biológicas (CIB), Medellín, Colombia 5Institute of Biology, Universidad de Antioquia, Medellín, Colombia 6Department of Dermatology, The Affiliated Hospital, Guizhou Medical University, Guiyang, China 7Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China 8University of Alberta Microfungus Collection and Herbarium and Biological Sciences, Edmonton, AB, Canada 9Thermo Fisher Scientific, Landsmeer, The Netherlands 10School of Medicine, Universidad de Antioquia, Medellín, Colombia 11School of Medicine and Health Sciences, Universidad del Rosario, Bogotá, Colombia 12Faculty of Medicine and Health Sciences, Epidemiology for Global Health Institute, University of Antwerp, Antwerp, Belgium 13Rady Faculty of Health Sciences, Max Rady College of Medicine, University of Manitoba, Winnipeg, MB, Canada 14University of Cape Town, Cape Town, South Africa 15National Institute for Communicable Diseases, Johannesburg, South Africa 16Basic Pathology Department, Federal University of Paraná, Curitiba, Brazil 17Sexually Transmitted Infection Unit, Institute of Tropical Medicine, Antwerp, Belgium 18PHE UK Mycology Reference Laboratory, Bristol, UK Correspondence Yanping Jiang and Sybren de Hoog, Summary CBS-KNAW Fungal Biodiversity Centre, Recent discoveries of novel systemic fungal pathogens with thermally dimorphic yeast- Utrecht, The Netherlands. Emails: [email protected] and like phases have challenged the current taxonomy of the Ajellomycetaceae, a family [email protected] currently comprising the genera Blastomyces, Emmonsia, Emmonsiellopsis, Helicocarpus, Funding Information Histoplasma, Lacazia and Paracoccidioides. Our morphological, phylogenetic and phy- This project has been funded in whole or in logenomic analyses demonstrated species relationships and their specific phenotypes, part with Federal funds from the National Institute of Allergy and Infectious Diseases, clarified generic boundaries and provided the first annotated genome assemblies to National Institutes of Health, Department support the description of two new species. A new genus, Emergomyces, accommo- of Health and Human Services, under Grant Number U19AI110818 to the Broad Institute. dates Emmonsia pasteuriana as type species, and the new species Emergomyces afri- This work was partly supported by Colciencias canus, the aetiological agent of case series of disseminated infections in South Africa. via the grants “A Gene Atlas for Human Pathogenic Fungi,” 122256934875. Both species produce small yeast cells that bud at a narrow base at 37°C and lack CBS- KNAW dedicated a grant from the †Contributed equally to this work. 296 | © 2017 Blackwell Verlag GmbH wileyonlinelibrary.com/journal/myc Mycoses 2017;60:296–309 DUKIK ET AL. | 297 EC- funded SYNTHESYS project for systematic resources (http://www.synthesys. adiaspores, classically associated with the genus Emmonsia. Another novel dimorphic info/) to Benjamin Stielow to perform Ion pathogen, producing broad- based budding cells at 37°C and occurring outside North Torrent PGM next generation sequencing studies and novel experimental designs to America, proved to belong to the genus Blastomyces, and is described as Blastomyces advance fungal molecular phylogenetics. One percursus. part of the Ion Torrent PGM next generation sequencing studies was funded by The KEYWORDS Institute of Tropical Medicine in Antwerp, Belgium. Ajellomycetaceae, Blastomyces, Emergomyces, Emmonsia, genomics, phylogeny 1 | INTRODUCTION of these novel taxa are opportunistic pathogens of immunocom- promised hosts, primarily persons infected with HIV.1 An important Recent discoveries of novel systemic human pathogens with a ther- emerging species associated with disease in advanced HIV infection mally dimorphic pathogenic phase that consist of budding yeast cells was found in South Africa with at least 56 cases reported since being have challenged the current taxonomy of the family Ajellomycetaceae correctly identified in 2008.8,9 The agent causing this mycosis was (order Onygenales).1 For nearly 100 years, only four genera of clas- closely related to Emmonsia pasteuriana, which is also known to infect sical systemic pathogens, each containing just one or two species, patients with AIDS and patients with other immune disorders.10–12 were recognised in the order Onygenales, ie Coccidioides, Blastomyces, The present work studies the relationships of unclassified iso- Histoplasma and Paracoccidioides. All these fungi reside in their fila- lates from clinical sources with existing Emmonsia and Blastomyces mentous forms in soil or guano, and upon inhalation by the host, they species and other members of the Ajellomycetaceae by means of morphologically shift to an invasive spherule (Coccidioides) or a yeast- comprehensive morphological and phylogenetic analyses involving like form in the host’s pulmonary system. While phylogenetic anal- both multilocus and whole genome sequencing. We describe the new yses have placed the genus Coccidioides in the family Onygenaceae, genus Emergomyces to include Emmonsia pasteuriana as type species Blastomyces, Histoplasma and Paracoccidioides proved to be members (Emergomyces pasteurianus comb. nov.) and Emergomyces africanus sp. of the family Ajellomycetaceae.2 nov. for a fungus formerly mentioned by Schwartz et al. as Emmonsia Another documented genus within the Ajellomycetaceae is sp. 5.1 Another dimorphic human pathogen is described as Blastomyces Emmonsia, until recently known mainly for species that cause pulmo- percursus sp. nov. (formerly named Emmonsia sp. 31). We completed nary infections in small mammals. Until the description of Emmonsia the first annotated genome assemblies for these novel species, which pasteuriana in 1998, the genus Emmonsia contained two species: the may guide the development of new diagnostics. genetically homogeneous Emmonsia crescens and a more diverse spe- cies, Emmonsia parva.3 These are the aetiological agents of adiaspiro- 2 | MATERIALS AND METHODS mycosis, a pulmonary disease of terrestrial mammals and occasionally of humans.4,5 They differ from classical dimorphic pathogenic fungi by 2.1 | Strains and phenotypes their pathogenic phase consisting of large, thick- walled adiaspores in- stead of budding yeast cells. Emmonsia crescens has adiaspores often Reference strains were taken from the collection of the over 100 μm in diameter and a maximum growth temperature of 37°C, Centraalbureau voor Schimmelcultures (CBS) of CBS- KNAW Fungal while the adiaspores of Emmonsia parva are mostly 15- 25 μm in diam- Biodiversity Centre, Utrecht, The Netherlands, the University of eter and the fungus grows up to 40°C. Multilocus phylogenetic analy- Alberta Microfungus Collection and Herbarium (UAMH), Devonian sis suggested these species to be less closely related than anticipated. Botanic Garden, Edmonton, Canada (now UAMH Centre for Global Emmonsia parva clustered with Blastomyces dermatitidis/B. gilchris- Microfungal Biodiversity, Toronto, Canada) and the National Collection tii, while Emmonsia crescens took a rather isolated position.1 Recent of Pathogenic Fungi (NCPF), Mycology Reference Laboratory, Bristol, phylogenomic analysis supported Emmonsia crescens as a sister group U.K., supplemented by kind donations of individual researchers. to the clade including Histoplasma, Emmonsia parva and B. dermatiti- Twenty- four strains were selected for detailed morphological and dis/B. gilchristii.6 Taken together, the genetic evidence suggests that, molecular study (Table 1). These were part of a larger dataset com- in spite of striking morphological, ecological and pathophysiological prising 109 strains, including outgroup (Table S1) used in multilocus similarities, aetiological agents of adiaspiromycosis are polyphyletic. analyses. Reference strains belonging to Coccidioides, Paracoccidioides, Since the 1970s, novel pathogens have emerged with phylo- Blastomyces and Histoplasma as well as the novel taxa described here genetic, morphological and clinical similarities to known members were handled in biosafety level 3 (BSL- 3) laboratories; Emmonsia cre- of the Ajellomycetaceae. Schwartz et al. [1] summarised reports of scens and Emmonsia parva were handled at BSL- 2. Strains were cul- numerous additional human cases due to novel species in the fam- tured on 2% Malt Extract Agar (MEA, Oxoid) plates using inoculum ily Ajellomycetaceae, most of which remained undescribed. Recently, from lyophilised, cryo- preserved or fresh mycelium. Cultures were Wang et al. [7] reported on another novel species from China. Several incubated for 28 days at a temperature of 24°C. 298 | TABLE 1 Strains