Fungal diversity of the hypersaline Inland Sea in Qatar
-
Rashmi Fotedar
Rashmi Fotedar works as an expert, Department of Genetic Engineering, Ministry of Municipality and Environment, Doha, Qatar. She earned her Master’s in molecular biology from University of Technology, Sydney, Australia and her PhD from All India Institute of Medical Sciences New Delhi, India. Her research interests include marine microbiology and molecular characterization of fungi and bacteria from environment and humans.
,
Anna Kolecka
Anna Kolecka is a researcher at Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands. From 2010 to 2017 she was working as a postdoc in the Yeast Research Group, led by Teun Boekhout. During those years she extensively studied the implementation of MALDI-TOF mass spectrometry for yeast identification and DNA-based molecular approaches for fungal taxonomy. She further explored the application of proteomics for fungal discrimination, recently by using high resolution accurate mass-mass spectrometry (HRAM-MS). She completed her MSc Eng studies in environmental protection at Gdansk University of Technology, Poland in 2005 and later received her PhD in microbiology at Comenius University, Bratislava, Slovakia in 2011.
Teun Boekhout, works as a principal investigator at Westerdijk Fungal Biodiversity Institute (previously known as CBS Fungal Biodiversity Centre, CBS-KNAW), Utrecht, and Institute of Biodiversity and Ecosystem Dynamics, University of Amsterdam, Amsterdam, The Netherlands. He studied biology in Utrecht where he also graduated. Since 1982 he is working in mycology. In 2018 he was elected Fellow of the American Academy of Microbiology. In 2011 he was editor of
The yeasts, a taxonomic study and presently he is editor in chief ofThe yeasts , an open access platform on yeast diversity.Jack W. Fell, PhD, Professor Emeritus, University of Miami. Chairman, Science Advisory Board, Bahamas Marine EcoCentre. Dr Fell a marine scientist, with his colleagues and students, studied microbial ecology from the tropics to Antarctic, mangroves to deep sea. His lab research included basidiomycetous yeast mating genetics, he was a pioneer in basidiomycete molecular phylogeny and developed molecular probes for rapid species detection of marine microbes and human pathogens.
Ameena Al-Malki works as a specialist in Genetic Engineering Department, Biotechnology Centre, Ministry of Environment, Doha, Qatar. She is involved in studies related to isolation and molecular identification of microorganism from marine environment, detection of GMOs in agricultural and food products. Her fields of interest include the molecular characterization of bacteria and fungi from environmental samples.
Aisha Zeyara works as an expert in Department of Genetic Resources, Ministry of Municipality and Environment, Doha, Qatar. She has a degree in microbiology and molecular biology from Qatar University, Doha, Qatar. Aisha is a molecular microbiologist and has been involved in research studies including isolation, identification and characterization of microbial pathogens from Qatari marine environment.
Masoud Al Marri, works as Director, Agricultural Research Department, Ministry of Municipality and Environment, Doha, Qatar. He earned his Master’s degree from University of Florida, USA and has been involved in research projects related to marine environment and environmental monitoring of marine, ground water and air in Qatar.
Abstract
The hypersaline Inland Sea in Qatar constitutes a unique ecosystem characterized by salinities up to saturation, extreme temperature fluctuations, and limited rainfall. To reveal the fungal diversity of this environment, we isolated fungi from water samples collected at the Inland Sea. Taxonomic identification of the isolates was done via DNA barcoding of the ITS1 and ITS2 ribosomal DNA (rDNA) domains and the D1/D2 domains of the nuclear large subunit rDNA. Additional genes, including glyceraldehyde-3-phosphate dehydrogenase (gapdh) and translation elongation factor 1-alpha (tef1), were included for isolates of Alternaria, actin (Act) for Cladosporium, part of the beta-tubulin (BenA) and calmodulin (CaM) genes for Aspergillus and Penicillium. In total, 159 fungal isolates, including 85 Ascomycota and 74 Basidiomycota, were obtained from the water samples collected during four samplings in the winter and summer seasons. About 14% (22/159) of the strains, presumably novel species, were preliminarily identified to the genus level. This is the first report highlighting the diversity of fungi from the hypersaline Inland Sea in Qatar.
Introduction
Fungi are ubiquitous microorganisms that are part of the microbiota occurring in natural ecosystems, including fresh and marine waters ranging from the ocean surface to the deep sea, eutrophic to ultra-oligotrophic lakes, lagoons, rivers, ground waters, melting water and ice of glaciers (Fell et al. 2011, Turchetti et al. 2013, Gunde-Cimerman and Zalar 2014, Grum-Grzhimaylo et al. 2016, Mokhtarnejad et al. 2016). Fungal diversity in so-called extreme environments may be affected by a variety of abiotic and biotic factors, including temperature, pressure, UV radiation (UVR), salinity, presence of fauna, flora and other microorganisms, run-off from soils and spills, and anthropogenic effluents. In these extreme habitats the interactions between abiotic factors, for instance temperature, salinity, and pH, result in the colonization by microorganisms adapted to these environmental conditions (Gunde-Cimerman et al. 2000, Pedros-Alio et al. 2000, Pedros-Alio 2004).
High concentrations of salt (hypersalinity) result in an extreme environment with limited fungal and other microbial growth. Due to this strong selection pressure the diversity of halophilic and halotolerant fungi decreases with increasing salt concentrations (Gunde-Cimerman et al. 2000, Butinar et al. 2005a, Gunde-Cimerman and Plemenitaš 2005, Grum-Grzhimaylo et al. 2016). Many genera of fungi such as Aspergillus P. Micheli ex Halle, Alternaria Nees, Aureobasidium Viala et G. Boyer, Cladosporium Link, Penicillium Link and Hortaea Nishim. et Miyaji, have been reported from hypersaline environments (Kis-Papo et al. 2001, Diaz-Munoz and Montalvo-Rodrigue 2004, Gunde-Cimerman et al. 2004, Butinar et al. 2005a, Coelho et al. 2010, Al-Musallam et al. 2011, Nazareth et al. 2012, Jaouani et al. 2014, Mokhtarnejad et al. 2016).
The Inland Sea is a hypersaline lagoon at the inlet of the Arabian Gulf located in southeast Qatar, bordering Saudi Arabia. This lagoon extends 15 km from north to south and 12 km from east to west and is connected to the Arabian Gulf by a 10-km long channel that allows exchange of water mass and subsequent interaction with bottom substrata (Figure 1). The area surrounding the Inland Sea is characterized by the presence of large mobile sand dunes, a tidal embayment system, inland and coastal saltpans, salt hills, stony deserts, elevated areas, and rocky ridges. The Inland Sea is a largely uninhabited area of global importance (Schwarze et al. 2005), which was nominated for the World Natural Heritage List (http://whc.unesco.org/en/tentativelists/5317/). In 1993 Inland Sea was declared a water sanctuary by Ministerial Decree No.78 of the Ministry of Municipal Affairs and Agriculture of Qatar that banned all commercial fishing activities. Because of its connectivity to the Arabian Gulf, Inland Sea is characterized by a pronounced salt gradient with salinities ranging from 57 to 75 in the tidal embayment area with saturated salt conditions in the saltpans near the coastal zones. These characteristics offer a unique opportunity to investigate microbial communities along a natural pronounced salinity gradient, with the original microbiota recruited from the Arabian Gulf.
Map of Inland Sea of Qatar and location of sampling sites. Image Map courtesy: Centre for GIS, Ministry of Municipality and Environment. www.mme.gov.qa.
The aim of this investigation was to assess the fungal diversity in the hypersaline ecosystem of Inland Sea. Sequencing of different genes was done via DNA barcoding of the ITS1 and ITS2 ribosomal DNA (rDNA) domains and the D1/D2 domains of the nuclear large subunit rDNA. Additional genes, including glyceraldehyde-3-phosphate dehydrogenase (gapdh) and translation elongation factor 1-alpha (tef1), were included for isolates of Alternaria, actin (Act) for Cladosporium, part of the beta-tubulin (BenA) and calmodulin (CaM) genes for Aspergillus and Penicillium.
Materials and methods
Inland Sea sampling sites and collection of water samples
The Inland Sea is located at the inlet of the Arabian Gulf in southeast Qatar (N 24.625308; E051.297259; Figure 1). This area is an arid region characterized by high solar radiation, high salinities ranging from 57 to 75, and limited rainfall (75.2 mm per year). Sampling at eight locations (Figure 1, Table 1) took place during the winter (December 2013) and summer (September 2014) seasons. Seven coastal sites (INS1-INS7) were sampled near the littoral zone in the vicinity of salt pans at a depth of 0.5 m. One site (INS8) was sampled at a depth of 2.5 m in the pelagic zone at approximately 250 m from the coast (Table 1). INS8 was located at the entrance of the 10-km-long channel that allows exchange of water from the Arabian Gulf to the lagoon. Sites INS2, INS6, and INS7 were influenced by human activity due to the proximity of recreational areas and camps, which are allowed in the area from October to April.
Details of sampling sites at the Inland Sea, characteristics of water samples and fungal counts.
| Station/depth (m) | Coordinates | Sampling season/month | Salinity | pH | Temp. (°C) | Yeast count (CFUa/l) | Filamentous fungi count (CFUa/l)<<#Hvind>> |
|---|---|---|---|---|---|---|---|
| INS1/0.5 | N 24.6706; E 051.2884 | Winter/Dec 2013 | 73 | 8.06 | 24–25 | 70 | 100 |
| INS2/0.5 | N 24.6171; E 051.3573 | Winter/Dec 2013 | 75 | 8.06 | 24–25 | 140 | 100 |
| INS3/0.5 | N 24.65787; E 51.28368 | Summer/Sept 2014 | 72 | 8.07 | 30–32 | 90 | 15 |
| INS4/0.5 | N 24.66417;E 51.29282 | Summer/Sept 2014 | 73 | 8.2 | 30–32 | 0 | 15 |
| INS5/0.5 | N 24.64786; E51.30041 | Summer/Sept 2014 | 75 | 8.1 | 30–32 | 0 | 15 |
| INS6/0.5 | N 24.64357; E 51.33078 | Summer/Sept 2014 | 74 | 8.2 | 30–32 | 710 | 10 |
| INS7/0.5 | N 24.62436; E 51.34179 | Summer/Sept 2014 | 72 | 8.1 | 32 | 910 | 10 |
| INS8/2.5 | N 24.5522; E 051.3325 | Summer/Sept 2014 | 57.3 | 7.99 | 32 | 69 | 8 |
aColony forming units (CFU) based on gross morphology of colonies seen on the filter paper. The yeast column included all the yeast isolates and also Aureobasidium spp. and Hortaea werneckii which grew on the medium as yeasts.
The distances between the sampling sites (INS1-INS7) ranged from approximately 1–3 km, whereas INS8 was further away, approximately 8.2 km from INS2 (Figure 1). Water samples were collected in a 1.7-l Niskin Water Sampler (model 1010) with survey outboard motor boats. The water samples were collected in sterile glass bottles of 1 l that were transferred to the laboratory within 2–3 h after collection, and processed immediately for the isolation of fungi with standard microbiological methods. Marine water parameters, such as temperature, salinity, and pH, were measured in three replicate readings with a 650 MDS Data Logger and 6920 V2 multiparameter, water Quality Sonde (Xylem Analytics, UK) calibrated against standard solutions.
Isolation, identification, and characterization of yeasts and filamentous fungi
Culture methods
For the isolation of fungi, five volumes of 200 ml of water per site were filtered through sterile nitrocellulose membranes (0.45 μm pore size; 47 mm diameter, Merck-Millipore, Ltd., Ireland) with a sterilized filtration device. The process was replicated three times for each sampling site. The filters were placed on modified glucose-yeast extract-peptone agar (GYPA) medium (2% glucose (Conda laboratories, Spain), 1% peptone (Conda laboratories, Spain), 0.5% yeast extract (Conda lab, Spain), 0.05% chloramphenicol (Sigma Aldrich, USA), 1.7% agar, 50% filtered water collected from Inland Sea). The medium was prepared with Inland Sea water obtained from the same sites where the water samples were collected. Initial experiments with the incubation temperatures of >25°C and 30°C for summer and winter seasons, respectively, showed an overgrowth of filamentous fungi, which obscured the yeast colonies. The plates were consequently incubated at 17°C and 25°C for 8–10 weeks for the winter and summer samples, respectively (Fell et al. 2011).
The plates were inspected for fungal growth at intervals of 48–72 h and cell counts (CFU (colony forming unit) l−1) of yeasts and filamentous fungi were made with a stereoscopic microscope (Olympus SZX 16, Japan). The fungal count was scored after 3, 5, 7, 14, 30 and 60 days of incubation and the averages (per site/per sampling) of the total viable yeast and filamentous fungi were calculated. Microscopic and morphological observations were made with an Olympus SZX 16 stereomicroscope (Olympus, Japan). At least one colony of each macro-morphological colony type was picked and subcultured onto a fresh medium for further purification. For yeasts, purification of isolates was performed on modified GYPA plates, whereas potato dextrose agar (PDA, Conda Laboratories, Spain) plates were used for filamentous fungi. The isolates were purified by repeated sub-culturing at least three to five times and stored in 50% glycerol (Avonchem, Cheshire, UK) in sterile distilled water at −20°C.
PCR amplification, sequencing, and phylogenetic analyses
For molecular identification, genomic DNA was extracted in 96-well plates following the PrepMan TM Ultra (Applied Biosystems, UK) protocol (Stielow et al. 2015). The internal transcribed spacer 1 and 2 regions (ITS), including the 5.8S rDNA and the D1/D2 domains of the nuclear large subunit (LSU) rDNA, were used for molecular identification (Fell et al. 2000, Scorzetti et al. 2002, Vu et al. 2016). The primers ITS4 and ITS5 (White et al. 1990) were used to amplify the ITS regions. The LSU region was amplified with the primers LR0R (Rehner and Samuels 1994) and LR5 (Vilgalys and Hester 1990). PCR reactions for amplification were performed in volumes of 12.5 μl containing 2.5 μl of the genomic DNA as reported by Stielow et al. (2015). The PCR product was checked by agarose gel electrophoresis, purified with Fast AP, the alkaline phosphatase (Fermentas, Thermo Fisher Scientific, USA) and cycle-sequenced with the ABI Big Dye™ v3.1 technology (Thermo Fisher Scientific, USA). To increase the taxonomic resolution for some groups of filamentous fungi, additional barcodes were used. For Cladosporium isolates, part of the actin (act) gene was amplified with the primer set ACT-512F and ACT-783R (Carbone and Kohn 1999) as described in Groenewald et al. (2013). For Alternaria species, part of the gapdh and tef1 genes were amplified with the primer sets gpd1 and gpd2 (Berbee et al. 1999) and EF1-728F and EF1-986R (Carbone and Kohn 1999), respectively (Woudenberg et al. 2013). For Aspergillus and Penicillium species, part of the beta-tubulin (BenA) and calmodulin (CaM) genes amplified with the primer sets Bt2a-F and Bt2b-R (Glass and Donaldson 1995) and Cmd5-F and Cmd6-R (Hong et al. 2005), respectively, were sequenced as described in Samson et al. (2014) and Visagie et al. (2014). Forward and reverse sequences were manually corrected with SeqMan software v 8.0.2 (DNA Star Inc., Madison, WI, USA). The resulting consensus sequences were compared by Basic Local Alignment Search Tool (BLASTn) (Altschul et al. 1990) with sequences available in the GenBank database for primary identifications (https://blast. ncbi.nlm.nih.gov/Blast.cgi?PAGE_TYPE=BlastSearch). All sequences from the Qatar isolates were grouped by genus and aligned with Clustal W in MEGA v.6 software (Tamura et al. 2011, 2013) with sequences of relevant type strains. Phylogenetic analysis of the LSU and ITS regions followed the procedures of Fell et al. (2000) and Scorzetti et al. (2002). Phylogenetic trees based on the D1/D2 sequence were constructed by using the program provided within the MEGA6 software package. Phylogenetic placement of the novel species was analyzed with the maximum likelihood method (Tamura et al. 2011, 2013). Based on this analysis and the initial BLAST scores, the identity of the isolates at genus, section or species (-complex) level is listed in Table 2.
Species of fungi isolated from the Inland Sea, Qatar.
| Group | Species recovered | Site/No. of isolates | Strains sequenced b | ITS-GenBank accession numbers | LSU-GenBank accession numbers | Gapdh-GenBank accession numbers | Tef1-GenBank accession numbers | Actin (A)/β-tubulin (BT) Calmodulin (C) GenBank accession numbers | No. of isolates in Winter | No. of isolates in Summer | Total no. (%) |
|---|---|---|---|---|---|---|---|---|---|---|---|
| Ascomycota | Alternaria spp.a | INS8 (4) | 2M108A | KY387606 | KY781812 | KY387604 | KY387608 | – | 2 | 4 | 6 (3.8) |
| INS1 (2) | INM7 | KY387605 | KY781811 | KY387603 | KY387607 | – | |||||
| Alternaria sect. Alternaria D.P. Lawrence, P.B. Gannibal, T.L. Peever, B.M. Pryor | INS1 (3) INS2 (5) INS3 (1) INS8 (1) | INM4 INM10 | KY744118 – | KY744136 – | MF084400 MF084401 | MF084404 MF084405 | – – | 8 | 2 | 10 (6.3) | |
| Alternaria sect. Ulocladioides Woudenberg et Crous | INS1 (1) INS8 (1) | INM5 2M109 | – – | – – | MF084402 MF084403 | MF084406 – | – – | 1 | 1 | 2 (1.2) | |
| Aspergillus citrinoterreus | INS8 (1) | 2M110B | KY781744 | KY781745 | – | – | BT-MF084376 C-MF084381 | 0 | 1 | 1 (0.6) | |
| J. Guinea, M. Sandoval-Denis, P. Escribano, E. Bouza et J. Guarro | |||||||||||
| Aureobasidium iranianum | INS1 (1) | YINS1 | KY781746 | KY781747 | – | – | – | 1 | 0 | 1 (0.6) | |
| Arzanlou et Khodaei | |||||||||||
| Aureobasidium melanogenum (Hermanides-Nijhof) Zalar, Gostincar et Gunde-Cim. | INS8 (2) | 2Y211 2Y215A | KY781748 KY781750 | KY781749 KY781751 | – | – | – | 0 | 2 | 2 (1.2) | |
| Aureobasidium pullulans | INS8 (1) | 2Y101 | – | KY781752 | – | – | – | 0 | 1 | 1 (0.6) | |
| (de Bary) G. Arnaud | |||||||||||
| Zalaria obscura Visagie, Z. Humphries et Seifert | INS3 (2) INS7 (1) INS8 (1) | INY20 INY28 2Y102 | KY781753 KY781755 KY781757 | KY781754 KY781756 KY781758 | – – – | – – – | – – – | 0 | 4 | 4 (2.5) | |
| Candida metapsilosis Tavanti, A. Davidson, Gow, M. Maiden et Odds | INS7 (1) | INY21 | – | KY695002 | – | – | – | 0 | 1 | 1 (0.6) | |
| Candida orthopsilosis Tavanti, A. Davidson, Gow, M. Maiden et Odds | INS8 (1) | 2Y191A | – | – | – | – | – | 0 | 1 | 1 (0.6) | |
| Candida parapsilosis (Ashford) Langeron et Talice | INS8 (1) | 2Y213 | KY694996 | KY695003 | – | – | – | 0 | 1 | 1 (0.6) | |
| Cladosporium cladosporioides(Fresen.) G.A. de Vries | INS2 (2) | INM21 | KY781759 | KY781760 | – | – | A-MF084386 | 2 | 0 | 2 (1.3) | |
| INM29 | – | – | – | – | – | ||||||
| Cladosporium halotoleransZalar, de Hoog et Gunde-Cim. | INS1 (3) INS2 (1) INS5 (1) | INM1 INM14 | KY781786 KY781788 | KY781787 KY781789 | – – | – – | A-MF084398 A-MF084399 | 4 | 1 | 5 (3.2) | |
| Cladosporium limoniforme Bensch, Crous et U. Braun | INS1 (2) INS2 (1) INS4 (1) INS5 (1) INS6 (1) INS7 (2) INS8 (1) | INM3 INM6 2INM1 | KY781780 KY781782 KY781784 | KY781781 KY781783 KY781785 | – – – | – – – | – – A-MF084397 | 3 | 6 | 9 (5.7) | |
| Cladosporium spp.a | INS1 (2) | INM18 | KY781769 | KY781770 | – | – | A-MF084391 | 6 | 1 | 7 (4.4) | |
| INM24 | KY781774 | KY781775 | – | – | A-MF084394 | ||||||
| INS2 (3) | INM23 | – | KY781773 | – | – | A-MF084393 | |||||
| INM37 | KY781776 | KY781777 | – | – | A-MF084395 | ||||||
| INS3 (2) | INM34 | KY781771 | KY781772 | – | – | A-MF084392 | |||||
| 2INM9 | KY781778 | KY781779 | – | – | A-MF084396 | ||||||
| Cladosporium tenuissimum Cooke | INS1 (5) | INM8 INM12 | KY781761 KY781763 | KY781762 KY781764 | – – | – – | A-MF084387 A-MF084388 | 5 | 0 | 5 (3.2) | |
| Cladosporium xantochromaticum Sandoval-Denis, Gené et Cano | INS1 (2) INS2 (2) | INM11 INM16 | KY781765 KY781767 | KY781766 KY781768 | – – | – – | A-MF084389 A-MF084390 | 4 | 0 | 4 (2.5) | |
| Debaryomyces hansenii (Zopf) Lodder et Kreger | INS2 (1) | YINM36B | KY694997 | KY695004 | – | – | – | 1 | 0 | 1 (0.6) | |
| Debaryomyces nepalensis | INS1 (2) INS2 (3) | YINS3 | KY694998 | KY695005 | – | – | – | 5 | 0 | 5 (3.2) | |
| Goto et Sugiy | |||||||||||
| Fusarium chlamydosporum Wollenw. et Reinking | INS3 (1) | 2INM10B | KY781790 | KY781791 | – | – | – | 0 | 1 | 1 (0.6) | |
| Hyphozyma spp.a | INS6 (2) | INY32C | KY695000 | KY695007 | – | – | – | 0 | 2 | 2 (1.3) | |
| INY36 | KY695001 | KY695008 | – | – | – | ||||||
| Hortaea werneckii (Horta) Nishim. et Miyaji | INS8 (5) | 2Y193B | KY781792 | KY781793 | – | – | – | 0 | 5 | 5 (3.2) | |
| 2Y210 | KY781794 | KY781795 | – | – | – | ||||||
| Trichoderma placentula Jaklitsch | INS6 (1) INS7 (2) | INY33, INY10B | – | KY781797 | – | – | – | 0 | 3 | 3 (1.9) | |
| KY781796 | – | – | – | – | |||||||
| Knufia petricola | INS8 (1) | 2Y215B | KY781798 | KY781799 | – | – | – | 0 | 1 | 1 (0.6) | |
| (Wollenz. et de Hoog) Gorbushina et Gueidan | |||||||||||
| Penicillium oxalicum | INS5 (1) | 2INM6A | KY781804 | KY781805 | – | – | C-MF084384 | 0 | 2 | 2 (1.3) | |
| Currie et Thom | BT-MF084379 | ||||||||||
| INS8 (1) | 2M110A | KY781806 | KY781807 | – | – | C-MF084385 | |||||
| BT-MF084380 | |||||||||||
| Penicillium chrysogenumThom | INS3 (1) INS4 (1) | 2INM8 | KY781802 | KY781803 | – | – | C-MF084383 | 0 | 2 | 2 (1.3) | |
| 2INM4 | KY781800 | KY781801 | – | – | BT-MF084378 C-MF084382 BT-MF084377 | ||||||
| Sarocladium bacillisporum | INS1 (1) | YINS5 | KY781808 | – | – | – | – | 1 | 0 | 1 (0.6) | |
| (Onions et G.L. Barron) Summerb. | |||||||||||
| Basidiomycetes | Kondoa spp.a | INS8 (4) | 2Y188 2Y189 2Y209 2Y214 | KY744105 – KY744106 KY744107 | KY744120 | – | – | – | 0 | 4 | 4 (2.5) |
| KY744121 | |||||||||||
| KY744122 | |||||||||||
| KY744123 | |||||||||||
| Naganishia albida (Saito) X.Z. Liu, F.Y. Bai, M. Groenew. et Boekhout | INS7 (1) | INY12 | KY744108 | KY744124 | – | – | – | 0 | 1 | 1 (0.6) | |
| Naganishia albidosimilis (Vishniac et Kurtzman) X.Z. Liu, F.Y. Bai, M. Groenew. et Boekhout | INS3 (1) INS6 (2) INS7 (1) | INY14 INY25 | KY744109 KY744110 | KY744125 KY744126 | – – | – – | – – | 0 | 4 | 4 (2.5) | |
| Naganishia qatarensisa R. Fotedar, A. Kolecka, T. Boekhout, J.W. Fell A. Anand, A. Al Malaki, A. Zeyara, M. Al Marri. | INS6 (1) INS7 (1) | INY13 INY29 | KY744111 MG852088 | KY744127 KY744128 | – – | – – | – – | 0 | 2 | 2 (1.3) | |
| Papiliotrema laurentii | INS6 (1) | INY35 | KY744112 | KY744129 | – | – | – | 0 | 1 | 1 (0.6) | |
| (Kuff.) X.Z. Liu, F.Y. Bai, M. Groenew. et Boekhout | |||||||||||
| Rhodotorula diobovata (S.Y. Newell et I.L. Hunter) Q.M. Wang, F.Y. Bai, M. Groenew. et T. Boekhout. | INS6 (1) | INY34 | – | KY744130 | – | – | – | 0 | 1 | 1 (0.6) | |
| Rhodotorula mucilaginosa(A. Jörg.) F.C. Harrison | INS3 (6) INS6 (18) INS7(23) | INY2 INY6 – | KY744113 KY744114 – | KY744131 KY744132 – | – – – | – – – | – – – | 0 | 47 | 47 (29.6) | |
| Symmetrospora marina (Phaff, Mrak et O.B. Williams) Q.M. Wang, F.Y. Bai, M. Groenew. et Boekhout | INS8 (12) | 2Y186 2Y190A | KY744115 KY744116 | KY744133 KY744134 | – – | – – | – – | 0 | 12 | 12 (7.6) | |
| Symmetrospora spp.a | INS8 (1) | 2Y207 | KY744117 | KY744135 | – | – | – | 0 | 1 | 1 (0.6) | |
| Tremellales sp. | INS2 (1) | INM28 | KY781809 | KY781810 | – | – | – | 1 | 0 | 1 (0.6) | |
| Total isolates (number) | 44 | 115 | 159 (100) |
aStrains preliminarily identified at the genus level on the basis of the LSU, ITS, TEf1, Gapdh 1 and 2 sequence analyses.
bStrains sequenced and sequences deposited in GenBank.
Results
The physicochemical characteristics of the water samples are summarized in Table 1. The range of pH and salinity for the coastal sites, i.e. INS1-INS7, was similar, whereas INS8 showed a lower pH and salinity. During the winter and summer seasons the seawater temperatures varied between 24–25°C and 30–32°C, respectively (Table 1).
The total average yeast and filamentous fungus counts for each sampling site are shown in Table 1. The average number of isolates (including both yeasts and filamentous fungi) ranged from 15 to 920 CFU l−1 (Table 1), with the highest values corresponding to the sites INS7, INS6 and INS2 that were coastal zones in the vicinity of salt pans.
One hundred and fifty-nine isolates (yeasts and filamentous fungi) were obtained from the water samples (Table 2). Seventy-two percent of the isolates (115/159) were recovered during the summer season and 28% (44/159) during the winter season. Selected sequences of the fungal isolates were deposited in the GenBank database (www.ncbi.nlm.nih.gov.), and their GenBank accession numbers are listed in Table 2. Taxonomic assignments of yeasts were done according to the published literature (Kurtzman et al. 2011, Liu et al. 2015a,b, Wang et al. 2015a,b). Twenty-two strains (22/159; 13.8%) in six genera [Alternaria spp. (6 isolates), Cladosporium spp. (7 isolates), Hyphozyma spp. (2 isolates), Kondoa spp. (4 isolates), Naganishia qatarensis (2 isolates), Symmetrospora sp. (1 isolate)] could not be identified to the species level on the basis of the D1/D2 domains of the LSU and ITS rDNA. These strains may represent novel species (Table 2, Figures 2–4). The novel species of Naganishia Goto has been described as N. qatarensis by Fotedar et al. (2018). Formal taxonomic descriptions of the remaining novel species are in progress.
![Figure 2: Maximum likelihood multi-gene tree based on ITS, gapdh and tef1 sequences of Alternaria sp. nov. (marked in bold), including all reference isolates from Alternaria sect. Chalastospora and the closest relatives from Alternaria sect. Infectoriae with Alternaria penicillata CBS 116607 (Alternaria sect. Crivellia) as outgroup.The RAx ML bootstrap support values ≥75% (BS) and Bayesian posterior probabilities ≥0.95 (PP) are given at the nodes. Thickened lines indicate a BS of 100% and a PP of 1.0. The GenBank accession numbers of the studied sequences are given in brackets [ITS; gapdh; tef1] after the species names.](/document/doi/10.1515/bot-2018-0048/asset/graphic/j_bot-2018-0048_fig_002.jpg)
Maximum likelihood multi-gene tree based on ITS, gapdh and tef1 sequences of Alternaria sp. nov. (marked in bold), including all reference isolates from Alternaria sect. Chalastospora and the closest relatives from Alternaria sect. Infectoriae with Alternaria penicillata CBS 116607 (Alternaria sect. Crivellia) as outgroup.
The RAx ML bootstrap support values ≥75% (BS) and Bayesian posterior probabilities ≥0.95 (PP) are given at the nodes. Thickened lines indicate a BS of 100% and a PP of 1.0. The GenBank accession numbers of the studied sequences are given in brackets [ITS; gapdh; tef1] after the species names.
Phylogenetic tree showing the placement of Kondoa sp. nov. (marked in bold) and some closely related species based on the analysis of the combined ITS region and D1/D2 domains of LSU rDNA.
Type strains indicated by superscript T. Sequences not generated during this study were obtained from GenBank (accession numbers are shown in parentheses). The tree was constructed by maximum-likelihood method based on the Kimura 2-parameter model using MEGA6 software. Numbers on branches represent bootstrap values from 1000 replicates. Scale bar represents 0.05 substitutions per nucleotide position.
Phylogenetic tree showing the placement of Naganishia qatarensis (marked in bold) and some closely related species based on the analysis of the combined ITS region and D1/D2 domains of LSU rDNA.
Type strains indicated by superscript T. Sequences not generated during this study were obtained from GenBank (accession numbers are shown in parentheses). The tree was constructed by maximum-likelihood method based on the Kimura 2-parameter model using MEGA6 software. Numbers on branches represent bootstrap values from 1000 replicates. Scale bar represents 0.02 substitutions per nucleotide position (Fotedar et al. 2018).
Rhodotorula mucilaginosa, a halo- and psychrotolerant red-pigmented yeast species, represented almost 30% (47/159 isolates) of all fungal isolates and was obtained at a sampling depth of 0.5 m from three sites (INS3, INS6, INS7) with a salinity of 72–74 and temperatures ranging from 30 to 32°C.
The majority (40.2%; 64/159 isolates) of the fungal isolates consisted of melanized fungi belonging to the genera Alternaria, Aureobasidium, Cladosporium, Hortaea, Knufia L.J. Hutchison et Unter. and Zalaria Visagie, Z. Humphries et Seifert. These melanized fungi were isolated from all sites sampled. Among the melanized fungi, Cladosporium species (20%; 32/159 isolates) were most commonly isolated, followed by Alternaria species (11.3%; 18/159 isolates).
Discussion
The Inland Sea of Qatar is a unique ecosystem with extreme environmental conditions. During sampling we encountered temperatures of 24–32°C, below average rainfall (winter, 0.1 mm; summer, 0 mm) and high salinities (57–75). For sites INS1-7, no trend was observed between the fungal isolates obtained and the environmental factors measured, such as temperature, pH, salinity, and season of isolation. Sites INS7, INS6 and INS2 (approx. 2–3 km apart from each other) located at the east side had the highest fungal counts, and this could be related to the proximity of these sites to the coastal zone, and the recreational area and camps. Sites INS4, INS5, and INS3 showed low fungal counts that could be due to the pristine nature of these sites with absence of human activity. INS8 (salinity 57, pH 7.9, depth 2.5 m) showed a higher species diversity (Table 2), when compared to the other sites (salinity >70, pH 8.02–8.35 and 0.5 m depth).
The yeast communities in Inland Sea were similar to those reported from other hypersaline environments (Coelho et al. 2010, Yurkov et al. 2011, 2012, França et al. 2016, Mokhtarnejad et al. 2016). Several studies reported a predominance of basidiomycetous fungi in hypersaline aquatic environments (Libkind et al. 2003, Butinar et al. 2005a, de Garcia et al. 2007, Brandao et al. 2011, Mokhtarnejad et al. 2016), but in our study ascomycetous and basidiomycetous yeasts occurred almost with equal abundance (Ascomycota 52.8%; Basidiomycota 47.2%). Some psychrotolerant yeasts, such as species of Naganishia, Rhodotorula F.C. Harrison and Debaryomyces Lodder et Kreger-van Rij ex Kreger-van Rij, were isolated in this study. These yeast species have also been reported from cold ecosystems, e.g. the Calderone glacier, Alpine glaciers and high Arctic glaciers (Butinar et al. 2007, Turchetti et al. 2008, 2013, Branda et al. 2010, Zalar and Gunde-Cimerman 2014).
Rhodotorula mucilaginosa was the most frequently isolated yeast species (29.5% of the total isolates) from the sites adjacent to the salt pans (INS6 and INS7), which were impacted by human activity, due to their proximity to the recreational camps. Our results are in accordance with other reports highlighting R. mucilaginosa as the dominant yeast in hypersaline environments with a wide range of salinities and pH (Lahav et al. 2002, Burgard et al. 2010). Symmetrospora marina was the second most dominant yeast species with 12 isolates and was isolated from the pelagic site INS8 (salinity 57, 32°C). This species was reported from an aquatic environment in Japan (Urano et al. 2001), from the surface washing of shrimp (Sampaio 2011), and the Florida Everglades (Fell et al. 2011). This is the first report of S. marina from a hypersaline environment.
Naganishia albidosimilis (formerly Cryptococcus albidosimilis Vishniac et Kurtzman), Naganishia albida and Naganishia qatarensis (Fotedar et al. 2018) were isolated from three sites with temperatures ranging from 30 to 32°C. Naganishia albidosimilis was reported from hypersaline lakes in Iran (Mokhtarnejad et al. 2016). Strains of Kondoa species that belong to the Agaricostilbum clade of Pucciniomycotina (Fonseca et al. 2000, Sampaio 2011, Wang et al. 2015a) were isolated in low frequencies from INS8 only. The Qatari isolates are related to other marine Kondoa species, such as K. aeria Á. Fonseca, J.P. Samp. et Fell, and K. malvinella (Fell et I. L. Hunter) Y. Yamada, Nakagawa et I. Banno, on the basis of the phylogenetic analyses, but may present an undescribed species (Figure 3). Kondoa aeria is known from terrestrial habitats (Fonseca et al. 2000), waters of the North Island of New Zealand (Francis et al. 2016), and deep sea sediments of Sagami Bay, Japan (Minegishi et al. 2006). To our knowledge, this is the first report of the isolation of Kondoa spp. from a hypersaline lagoon.
Species of the genus Debaryomyces, i.e. D. hansenii and D. nepalensi, were recovered during the winter seasons from two sites with a salinity range of 73–75, a depth of 0.5 m and a temperature of 24–25°C. Debaryomyces hansenii has a broad salinity tolerance (Yadav and Loper 1999) and the isolation of this halotolerant species has been documented from oceanic and aquatic environments (Hagler and Ahearn 1987, Nagahama 2006, Coelho et al. 2010), high salinity coastal and oceanic environments (Almeida 2005), and natural hypersaline lakes that are exposed to seasonal low temperatures (Butinar et al. 2005a).
Candida metapsilosis and C. orthoparapsilosis were isolated in low numbers (2/159 isolates). A single isolate of Candida parapsilosis was cultured from INS8. This species was previously reported from aquatic and hypersaline environments (Soares et al. 1997, Butinar et al. 2005a, Hagler 2006, Nagahama 2006).
The largest group (40.2%; 64/159 isolates) of the filamentous fungal isolates belonged to the melanized fungal genera Alternaria, Aureobasidium, Cladosporium, Hortaea, Knufia and Zalaria. Cladosporium species (20%; 32/159 isolates) were the most dominant, followed by Alternaria species (11.3%; 18/159 isolates). These melanized isolates were recovered from different sampling sites (with varying salinities and temperatures) across the Inland Sea suggesting a wide adaptability and ability to survive in various environments of high salinities and high temperatures. The majority of these black fungi (75%; 48/64 isolates) were isolated from the coastal sites (INS1-7) near the vicinity of salt pans, indicating that they are able to grow in hypersaline waters and this observation is in agreement with those made elsewhere (Gunde-Cimerman et al. 2000, Nazareth et al. 2012).
Cladosporium was the most species-rich genus and was present in almost all sampling sites with high salinity of 72–75, and this observation is in accordance with reports of Cladosporium occurring in salterns in Slovenia and Puerto Rico (Gunde-Cimerman et al. 2000, Cantrell et al. 2006), Great Salt Lake, Utah (Cronin and Post 1977), sabkhas (coastal salt flats) and salterns (Gunde-Cimerman et al. 1997, Butinar et al. 2005b, Cantrell et al. 2006, Al-Musallam et al. 2011), deserts, and salty environments (Kis-Papo et al. 2001, Grishkan et al. 2003, Conley et al. 2006, Smolyanyuk and Bilanenko 2011). Several studies have shown that species of Alternaria are predominant in deserts and salty environments (Gunde-Cimerman et al. 2000, Kis-Papo et al. 2001, Conley et al. 2006, Smolyanyuk and Bilanenko 2011). These fungi produce thick-walled and strongly melanized spores, which are important to protect against UVR and increase desiccation tolerance (Sterflinger et al. 2012).
Hortaea werneckii, another melanized fungus, was recovered from the water samples collected from the deep water site INS8 (2.5 m depth; salinity 57.3). This yeast-like fungus was previously reported from drying salty pools (de Hoog and van den Ende 1992), hypersaline waters in Europe (Gunde-Cimerman et al. 2000, Plemenitas et al. 2008), saline water of crystallization ponds (Gunde-Cimerman et al. 2003) and solar salterns in Puerto Rico (Diaz-Munoz and Montalvo-Rodriguez 2004), suggesting that hypersaline water is an ecological habitat of this species (Gunde-Cimerman et al. 2000, Kogej et al. 2005). Hortaea werneckii thrives in extreme environments characterized by scarce nutrients, low oxygen tension, high temperature, high UVR, high osmotic stress, as well as a mixture of these conditions (Sterflinger 2006). In our study we did not isolate this species from sites near the salt pans, but in contrast H. werneckii was isolated from the deeper water samples of INS8.
Based on the D1/D2 LSU and ITS rDNA sequence analyses, 13.8% (22/159) of the isolates, presumably novel species, were preliminarily identified to the genus level. These species include Alternaria spp., Cladosporium spp., Hyphozyma spp., Kondoa spp., Naganishia qatarensis (Fotedar et al. 2018) and Symmetrospora sp. (Table 2; Figures 2–4). Taxonomically undefined yeast and filamentous fungi have been isolated in various extreme environments, including glaciers (Butinar et al. 2007, Turchetti et al. 2013), alkaline soils (Grum-Grzhimaylo et al. 2013, 2016), and saline as well as hypersaline environments (Buchalo et al. 1998, Jones and Abdel-Wahab 2005, Zalar et al. 2007, Al-Musallam et al. 2011).
This is the first report highlighting the isolation of halotolerant fungi from hypersaline waters of the Inland Sea in Qatar. Due to the rising sea levels this study will be useful as a reference for future monitoring studies in Qatar.
About the authors
Rashmi Fotedar works as an expert, Department of Genetic Engineering, Ministry of Municipality and Environment, Doha, Qatar. She earned her Master’s in molecular biology from University of Technology, Sydney, Australia and her PhD from All India Institute of Medical Sciences New Delhi, India. Her research interests include marine microbiology and molecular characterization of fungi and bacteria from environment and humans.
Anna Kolecka is a researcher at Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands. From 2010 to 2017 she was working as a postdoc in the Yeast Research Group, led by Teun Boekhout. During those years she extensively studied the implementation of MALDI-TOF mass spectrometry for yeast identification and DNA-based molecular approaches for fungal taxonomy. She further explored the application of proteomics for fungal discrimination, recently by using high resolution accurate mass-mass spectrometry (HRAM-MS). She completed her MSc Eng studies in environmental protection at Gdansk University of Technology, Poland in 2005 and later received her PhD in microbiology at Comenius University, Bratislava, Slovakia in 2011.
Teun Boekhout, works as a principal investigator at Westerdijk Fungal Biodiversity Institute (previously known as CBS Fungal Biodiversity Centre, CBS-KNAW), Utrecht, and Institute of Biodiversity and Ecosystem Dynamics, University of Amsterdam, Amsterdam, The Netherlands. He studied biology in Utrecht where he also graduated. Since 1982 he is working in mycology. In 2018 he was elected Fellow of the American Academy of Microbiology. In 2011 he was editor of The yeasts, a taxonomic study and presently he is editor in chief of The yeasts, an open access platform on yeast diversity.
Jack W. Fell, PhD, Professor Emeritus, University of Miami. Chairman, Science Advisory Board, Bahamas Marine EcoCentre. Dr Fell a marine scientist, with his colleagues and students, studied microbial ecology from the tropics to Antarctic, mangroves to deep sea. His lab research included basidiomycetous yeast mating genetics, he was a pioneer in basidiomycete molecular phylogeny and developed molecular probes for rapid species detection of marine microbes and human pathogens.
Ameena Al-Malki works as a specialist in Genetic Engineering Department, Biotechnology Centre, Ministry of Environment, Doha, Qatar. She is involved in studies related to isolation and molecular identification of microorganism from marine environment, detection of GMOs in agricultural and food products. Her fields of interest include the molecular characterization of bacteria and fungi from environmental samples.
Aisha Zeyara works as an expert in Department of Genetic Resources, Ministry of Municipality and Environment, Doha, Qatar. She has a degree in microbiology and molecular biology from Qatar University, Doha, Qatar. Aisha is a molecular microbiologist and has been involved in research studies including isolation, identification and characterization of microbial pathogens from Qatari marine environment.
Masoud Al Marri, works as Director, Agricultural Research Department, Ministry of Municipality and Environment, Doha, Qatar. He earned his Master’s degree from University of Florida, USA and has been involved in research projects related to marine environment and environmental monitoring of marine, ground water and air in Qatar.
Acknowledgments
This research was supported by research grant NPRP 6-647-1-127 from the Qatar National Research Fund, a member of Qatar Foundation to R. Fotedar, (Ministry of Environment, Qatar), T. Boekhout (Westerdijk Fungal Biodiversity Institute Uppsalalaan, Utrecht, Netherlands) and J.W. Fell (Rosenstiel School of Marine and Atmospheric Sciences, University of Miami, Key Biscayne, FL, USA). The authors are solely responsible for the content of the manuscript.
The publication of this article was funded by the Qatar National Library.
References
Almeida, J.M.G.C.F. 2005. Yeast community survey in the Tagus estuary. FEMS Microbiol. Ecol. 53: 295–303.10.1016/j.femsec.2005.01.006Search in Google Scholar
Al-Musallam, A., H. Al-Sammar and N. Al-Sané. 2011. Diversity and dominance of fungi inhabiting the sabkha area in Kuwait. Bot. Mar. 54: 83–94.10.1515/bot.2010.069Search in Google Scholar
Altschul, S.F., W. Gish, W. Miller, E.W. Myers and D.J. Lipman. 1990. Basic local alignment search tool. J. Mol. Biol. 215: 403–410.10.1016/S0022-2836(05)80360-2Search in Google Scholar
Berbee, M.L., M. Pirseyed and S. Hubbard. 1999. Cochliobolus phylogenetics and the origin of known, highly virulent pathogens, inferred from ITS and glyceraldehyde-3-phosphate dehydrogenase gene sequences. Mycologia 91: 964–977.10.2307/3761627Search in Google Scholar
Branda, E., B. Turchetti, G. Diolaiuti, M. Pecci, C. Smiraglia and P. Buzzini. 2010. Yeast and yeast-like diversity in the southernmost glacier of Europe (Calderone glacier, Apennines, Italy). FEMS Microbiol. Ecol. 72: 354–369.10.1111/j.1574-6941.2010.00864.xSearch in Google Scholar PubMed
Brandao, R.L., D. Libkind, B.M.A. Vaz, L.C. Espirito Santo, M. Moline, V. de Garcia, M. van Broock and C. Rosa. 2011. Yeasts from an oligotrophic lake in Patagonia (Argentina): diversity, distribution, and synthesis of photoprotective compounds and extracellular enzymes. FEMS Microbiol. Ecol. 76: 1–13.10.1111/j.1574-6941.2010.01030.xSearch in Google Scholar PubMed
Buchalo, A.S., E. Nevo, S.P. Wasser, A. Oren and H.P. Molitoris. 1998. Fungal life in the extremely hypersaline water of the Dead Sea: first records. Proc. R. Soc. London. 265: 1461–1465.10.1098/rspb.1998.0458Search in Google Scholar PubMed PubMed Central
Burgard, G., D. Arzur, L. Durand, M.A. Cambon-Bonavita and G. Barbier. 2010. Marine culturable yeasts in deep-sea hydrothermal vents: species richness and association with fauna. FEMS Microbiol. Ecol. 72: 121–133.10.1111/j.1574-6941.2010.00881.xSearch in Google Scholar PubMed
Butinar, L., S. Santos, I. Spencer-Martins, A. Oren and N. Gunde-Cimerman. 2005a. Yeast diversity in hypersaline habitats. FEMS Microbiol. Lett. 244: 229–234.10.1016/j.femsle.2005.01.043Search in Google Scholar PubMed
Butinar, L., S. Sonjak, P. Zalar, A. Plemenitas and N. Gunde-Cimerman. 2005b. Melanized halophilic fungi are eukaryotic members of the microbial communities’ in hypersaline waters of solar salterns. Bot. Mar. 48: 73–79.10.1515/BOT.2005.007Search in Google Scholar
Butinar, L., I. Spencer-Martins and N. Gunde-Cimerman. 2007. Yeasts in high Arctic glaciers: the discovery of a new habitat for eukaryotic microorganisms. Antonie van Leeuwenhoek 91: 277–289.10.1007/s10482-006-9117-3Search in Google Scholar PubMed
Cantrell, A., L. Casillas-Martinez and M. Molina. 2006. Characterization of fungi from hypersaline environments of solar salterns using morphological and molecular techniques. Mycol. Res. 110: 962–970.10.1016/j.mycres.2006.06.005Search in Google Scholar PubMed
Carbone, I. and L.M. Kohn. 1999. A method for designing primer sets for speciation studies in filamentous ascomycetes. Mycologia 91: 553–556.10.2307/3761358Search in Google Scholar
Coelho, M.A., J.M. Almeida, I.M. Martins, A.J. de Silva and J.P. Sampaio. 2010. The dynamics of the yeast community of the Tagus river estuary: testing the hypothesis of the multiple origins of estuarine yeasts. Antonio van Leeuwenhoek 98: 331–342.10.1007/s10482-010-9445-1Search in Google Scholar PubMed
Conley, C.A., G. Ishkhanova, C.P. McKay and K. Cullings. 2006. A preliminary survey of non-lichenized fungi cultured from the hyper arid Atacama Desert of Chile. Astrobiol. 6: 521–526.10.1089/ast.2006.6.521Search in Google Scholar PubMed
Cronin, A.E. and F.J. Post. 1977. Reports of a dematiaceous hyphomycete from Great Salt Lake, Utah. Mycologia 69: 846–847.10.2307/3758878Search in Google Scholar
de Garcia, V., S. Brizzio, D. Libkind, P. Buzzini and M. van Broock. 2007. Biodiversity of cold-adapted yeasts from glacial meltwater rivers in Patagonia, Argentina. FEMS Microbiol. Ecol. 59: 331–341.10.1111/j.1574-6941.2006.00239.xSearch in Google Scholar PubMed
de Hoog, G.S. and A.H.G. van den Ende. 1992. Nutritional pattern and eco-physiology of Hortaea werneckii, agent of human tinea nigra. Antonie van Leeuwenhoek 62: 321–329.10.1007/BF00572601Search in Google Scholar PubMed
Diaz-Munoz, G. and R. Montalvo-Rodriguez. 2004. Halophilic black yeast Hortaea werneckii in the Cabo Rojo solar salterns: its first record for this extreme environment in Puerto Rico. Caribb. J. Sci. 41: 360–365.Search in Google Scholar
Fell, J.W., T. Boekhout, A. Fonseca, G. Scorzetti and A. Statzell-Tallman. 2000. Biodiversity and systematics of basidiomycetous yeasts as determined by large-subunit rDNA D1/D2 domain sequence analysis. Int. J. Syst. Evol. Microbiol. 50: 1351–1371.10.1099/00207713-50-3-1351Search in Google Scholar PubMed
Fell, J.W., A. Statzell-Tallman, G. Scorzetti and M.H. Gutiérrez. 2011. Five new species of yeasts from freshwater and marine habitats in the Florida Everglades. Antonie van Leeuwenhoek 9: 533–549.10.1007/s10482-010-9521-6Search in Google Scholar PubMed
Fonseca, I., P. Sampaio, J. Inacio and J.W. Fell. 2000. Emendation of the basidiomycetous yeast genus Kondoa and the description of Kondoa aeria sp. nov. Antonie van Leeuwenhoek 77: 293–302.10.1023/A:1002453628455Search in Google Scholar
Fotedar, R., A. Kolecka, T. Boekhout, J.W. Fell, A. Anand, A. Al Malaki, A. Zeyara and M. Al Marri. 2018. Naganishia qatarensis sp. nov., a novel basidiomycetous yeast species from a hypersaline marine environment in Qatar. Int. J. Syst. Evol. Micobiol. 68: 2924–2929.10.1099/ijsem.0.002920Search in Google Scholar PubMed
França, L., C. Sanninom, B. Turchetti, P. Buzzini and R. Margesin. 2016. Seasonal and altitudinal changes of culturable bacterial and yeast diversity in Alpine forest soils. Extremophiles 20: 855–873.10.1007/s00792-016-0874-2Search in Google Scholar PubMed PubMed Central
Francis, M.M., V. Webb and G.C. Zuccarello. 2016. Marine yeast biodiversity on seaweeds in New Zealand waters. N. Zeal. J. Bot. 54: 30–47.10.1080/0028825X.2015.1103274Search in Google Scholar
Glass, N.L. and G.C. Donaldson. 1995. Development of primer sets designed for use with the PCR to amplify conserved genes from filamentous ascomycetes. Appl. Environ. Microbiol. 61: 1323–1330.10.1128/aem.61.4.1323-1330.1995Search in Google Scholar PubMed PubMed Central
Grishkan, I., E. Nevo and S.P. Wasser. 2003. Soil micromycete diversity in the hypersaline Dead Sea coastal area (Israel). Mycol. Progr. 2: 19–28.10.1007/s11557-006-0040-9Search in Google Scholar
Groenewald, J.Z., C. Nakashima, J. Nishikawa, H.D. Shin, J.H. Park, A.N. Jama, M. Groenewald, U. Braun and P. Crous. 2013. Species concepts in Cercospora: spotting the weeds among the roses. Stud. Mycol. 75: 115–170.10.3114/sim0012Search in Google Scholar PubMed PubMed Central
Grum-Grzhimaylo, A.A., M.L. Georgieva, A.J.M. Debets and E.N. Bilanenko. 2013. Are alkalitolerant fungi of the Emericellopsis lineage (Bionectriaceae) of marine origin? IMA Fungus 4: 213–228.10.5598/imafungus.2013.04.02.07Search in Google Scholar PubMed PubMed Central
Grum-Grzhimaylo, A.A., M.L. Georgieva, S.A. Bondarenko, A.J.M. Debets and E.N. Bilanenko. 2016. On the diversity of fungi from soda soils. Fungal Divers. 76: 27–74.10.1007/s13225-015-0320-2Search in Google Scholar
Gunde-Cimerman, N. and A. Plemenitaš. (eds). 2005. Adaptation to life at high salt concentrations in Archaea, Bacteria, and Eukarya. Springer-Verlag, Heidelberg. pp. 576.10.1007/1-4020-3633-7Search in Google Scholar
Gunde-Cimerman, N. and P. Zalar. 2014. Extremely halotolerant and halophilic fungi inhabit brine in solar salterns around the globe. Food. Technol. Biotechnol. 52: 170–179.Search in Google Scholar
Gunde-Cimerman, N., P. Zalar and A. Cimerman. 1997. Diversity of fungal community in high salt marine environments. Proc. Int. Symp. Environ. Biotech (ISEB). Oostende, Belgium. pp. 189–191.Search in Google Scholar
Gunde-Cimerman, N., P. Zalar, G.S. de Hoog and A. Plemenitaš. 2000. Hypersaline waters in salterns-natural ecological niches for halophilic black yeasts. FEMS Microbiol. Ecol. 32: 235–240.10.1111/j.1574-6941.2000.tb00716.xSearch in Google Scholar PubMed
Gunde-Cimerman, N., S. Sonjak, P. Zalar, J.C. Frisvad, B. Diderichsen and A. Plemenitas. 2003. Extremophilic fungi in arctic ice: the relationship between adaptation to low temperature and water activity. Phy. Chem. Earth. A/B/C. 28: 1273–1278.10.1016/j.pce.2003.08.056Search in Google Scholar
Gunde-Cimerman, N., P. Zalar, U. Petrovic, M. Turk, T. Kogej, G.S. de Hoog and A. Plemenitas. 2004. Fungi in the Salterns. In: (A. Ventosa, ed) Halophilic microorganisms. Springer-Verlag, Heidelberg. pp. 103–113.10.1007/978-3-662-07656-9_7Search in Google Scholar
Gunde-Cimerman, N., J. Ramos and A. Plemenitas. 2009. Halotolerant and halophilic fungi. Mycol. Res. 113: 1231–1241.10.1016/j.mycres.2009.09.002Search in Google Scholar PubMed
Hagler, A.N. 2006. Yeasts as indicators of environmental quality. In: (C.A. Rosa and P. Gábor, eds) Biodiversity and ecophysiology of yeasts. Springer-Verlag, Berlin. pp. 519–536.10.1007/3-540-30985-3_21Search in Google Scholar
Hagler, A.N. and D.G. Ahearn. 1987. Ecology of aquatic yeasts. In: (A.H. Rose and J.S. Harrison, eds) The yeasts, Vol 2, Yeasts and the environment. Academic Press, London. pp. 181–205.Search in Google Scholar
Hong, S.B., S.J. Go, H.D. Shin, J.C. Frisvad and R.A. Samson. 2005. Polyphasic taxonomy of Aspergillus fumigatus and related species. Mycologia 97: 1316–1329.10.1080/15572536.2006.11832738Search in Google Scholar
Jaouani, A., M. Neifar, V. Prigione, A. Ayari, I. Sbissi, S.B. Amor, S.B. Tekaya, G.C. Varese, A. Cherif and M. Gtari. 2014. Diversity and enzymatic profiling of halotolerant micromycetes from Sebkha El Melah, a Saharan salt flat in southern Tunisia. Biomed. Res. Internat.2014: 1–11.10.1155/2014/439197Search in Google Scholar PubMed PubMed Central
Jones, E.B.G. and M.A. Abdel-Wahab. 2005. Marine fungi from the Bahamas Islands. Bot. Mar. 48: 356–364.10.1515/bot.2005.047Search in Google Scholar
Kis-Papo, T., I. Grishkan, A. Oren, S.P. Wasser and E. Nevo. 2001. Spatiotemporal diversity of filamentous fungi in the hypersaline Dead Sea. Mycol. Res. 105: 749–756.10.1017/S0953756201004129Search in Google Scholar
Kogej, T., J. Ramos, A. Plemenita and N. Gunde-Cimerman. 2005. The halophilic fungus Hortaea werneckii and the halotolerant fungus Aureobasidium pullulans maintain low intracellular cation concentrations in hypersaline environments. Appl. Environ. Microbiol. 71: 6600–6605.10.1128/AEM.71.11.6600-6605.2005Search in Google Scholar PubMed PubMed Central
Kurtzman, C.P., T. Boekhout and J.W. Fell. 2011. The Yeasts: a taxonomic study, 5th ed. Amsterdam, Elsevier. pp. 2354.Search in Google Scholar
Lahav, R., P. Fareleira, A. Nejidat and A. Abeliovich. 2002. The identification and characterization of osmotolerant yeast isolates from chemical wastewater evaporation ponds. Microbiol. Ecol. 43: 388–396.10.1007/s00248-002-2001-4Search in Google Scholar PubMed
Libkind, D., S. Brizzio, A. Ruffini, M. Gadanho, M. van Broock and J.P. Sampaio. 2003. Molecular characterization of carotenogenic yeasts from aquatic environments in Patagonia, Argentina. Antonie van Leeuwenhoek 84: 313–322.10.1023/A:1026058116545Search in Google Scholar
Liu, X.Z., Q.M. Wang, B. Theelen, M. Groenewald, F.Y. Bai and T. Boekhout. 2015a. Phylogeny of tremellomycetous yeasts and related dimorphic basidiomycetes reconstructed from multigene sequence analyses. Stud. Mycol. 81: 1–16.10.1016/j.simyco.2015.08.001Search in Google Scholar
Liu, X.Z., Q.M. Wang, M. Göker, M. Groenewald, A.V. Kachalkin, H.T. Lumbsch, A.M. Millanes, M. Wedin, A.M. Yurkov, T. Boekhout and F.Y. Bai. 2015b. Towards an integrated phylogenetic classification of the Tremellomycetes. Phylogeny of tremellomycetous yeasts and related dimorphic basidiomycetes reconstructed from multigene sequence analyses. Stud. Mycol. 81: 85–147.10.1016/j.simyco.2015.12.001Search in Google Scholar
Minegishi, H., T. Miura, Y. Yoshida, R. Usami and F. Abe. 2006. Phylogenetic analysis of pectin degrading yeasts from deep-sea environments. J. Japan. Soc. Extremop. 5: 21–26.10.3118/jjse.5.21Search in Google Scholar
Mokhtarnejad, L., M. Arzanlou, A. Babai-Ahari, Di Mauro, A. Onofri, P. Buzzini and B. Turchetti. 2016. Characterization of basidiomycetous yeasts in hypersaline soils of the Urmia Lake National Park, Iran. Extremophiles 20: 915–928.10.1007/s00792-016-0883-1Search in Google Scholar
Nagahama, T. 2006. Yeast biodiversity in freshwater, marine and deep-sea environments. In: (C.A. Rosa and P. Gábor, eds) The yeast handbook biodiversity and ecophysiology of yeasts. Springer, Berlin. pp. 241–262.10.1007/3-540-30985-3_12Search in Google Scholar
Nazareth, S., V. Gonsalves and A. Shweta. 2012. First record of obligate halophilic Aspergilli from the Dead Sea. Ind. J. Microbiol. 52: 22–27.10.1007/s12088-011-0225-zSearch in Google Scholar
Pedros-Alio, C., J.I. Calderon-Paz, M.L. MacLean, G. Medina, C. Marrase, J.M. Gasol and N. Guixa-Boixereu. 2000. The microbial food web along salinity gradients. FEMS Microbiol. Ecol. 32: 143–155.10.1016/S0168-6496(00)00025-8Search in Google Scholar
Pedros-Alio, C. 2004. Trophic ecology of solar salterns. In: (A. Ventosa, ed) Halophilic microorganisms. Springer-Verlag, Heidelberg. pp. 33–48.10.1007/978-3-662-07656-9_2Search in Google Scholar
Plemenitas, A., T. Vaupotic, M. Lenassi, T. Kogej and N. Gunde-Cimerman. 2008. Adaptation of extremely halotolerant black yeast Hortaea werneckii to increased osmolarity: a molecular perspective at a glance. Stud. Mycol. 61: 67–75.10.3114/sim.2008.61.06Search in Google Scholar PubMed PubMed Central
Rehner, S.A. and G.J. Samuels. 1994. Taxonomy and phylogeny of Gliocladium analyzed from nuclear large subunit ribosomal DNA sequences. Mycol. Res. 98: 625–634.10.1016/S0953-7562(09)80409-7Search in Google Scholar
Sampaio, J.P. 2011. Rhodotorula. In: (C.P. Kurtzman, J.W. Fell and T. Boekhout, eds) The Yeasts, a taxonomic study. Elsevier, Amsterdam. pp. 1873–1927.10.1016/B978-0-444-52149-1.00155-5Search in Google Scholar
Samson, R.A., C.M. Visagie, J. Houbraken, S.B. Hong, V. Hubka, C.H. Klaassen, G. Perrone, K.A. Seifert, A. Susca, J.B. Tanney, J. Varga, S. Kocsubé, G. Szigeti, T. Yaguchi and J.C. Frisvad. 2014. Phylogeny, identification, and nomenclature of the genus Aspergillus. Stud. Mycol. 78: 141–173.10.1016/j.simyco.2014.07.004Search in Google Scholar
Schwarze, H.H., K. Barth, W. Al-Malki and I. Minz. 2005. Tourism in the Khor Al-Adaid Area, State of Qatar. Report to Qatar Tourism Authority. pp. 29.Search in Google Scholar
Scorzetti, G., J.W. Fell, A. Fonseca and A. Statzell-Tallman. 2002. Systematics of basidiomycetous yeasts: a comparison of large subunit D1/D2 and internal transcribed spacer rDNA regions. FEMS Yeast Res. 2: 495–517.10.1016/S1567-1356(02)00128-9Search in Google Scholar
Smolyanyuk, E.V. and E.N. Bilanenko. 2011. Communities of halotolerant micromycetes from the areas of natural salinity. Microbiology 80: 877–883.10.1134/S002626171106021XSearch in Google Scholar
Soares, C.A.G., M. Maury, F.C. Pagnocca, F.V. Araujo and L.C. Mendoca-Hagler. 1997. Ascomycetous yeast from tropical intertidal dark mud of southeast Brazilian estuaries. J. Gen. Appl. Microbiol. 4: 265–272.10.2323/jgam.43.265Search in Google Scholar PubMed
Sterflinger, K. 2006. Black yeasts and meristematic fungi: ecology, diversity and identification. In: (C.A. Rosa and P. Gábor, eds) The Yeast Handbook: Biodiversity and ecophysiology of yeasts. Springer, Berlin, Heidelberg. pp. 501–514.10.1007/3-540-30985-3_20Search in Google Scholar
Sterflinger, K., D. Tesei and K. Zakharova. 2012. Fungi in hot and cold deserts with particular reference to microcolonial fungi. Fungal Ecol. 5: 453–462.10.1016/j.funeco.2011.12.007Search in Google Scholar
Stielow, J.B., C.A. Lévesque and K.A. Seifert. 2015. One fungus, which genes? Development and assessment of universal primers for potential secondary fungal DNA barcodes. Persoonia 35: 242–263.10.3767/003158515X689135Search in Google Scholar PubMed PubMed Central
Tamura, K., D. Peterson, N. Peterson, G. Stecher, M. Nei and S. Kumar. 2011. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 28: 2731–2739.10.1093/molbev/msr121Search in Google Scholar PubMed PubMed Central
Tamura, K., G. Stecher, D. Peterson, A. Filipski and S. Kumar. 2013. MEGA6: molecular Evolutionary Genetics Analysis version 6.0. Mol. Biol. Evol. 30: 2725–2729.10.1093/molbev/mst197Search in Google Scholar
Turchetti, B., P. Buzzini, M. Goretti, E. Branda, G. Diolaiuti, C. D’Agata, C. Smiraglia and A. Vaughan-Martini. 2008. Psychrophilic yeasts in glacial environments of Alpine glaciers. FEMS Microb. Ecol. 63: 73–83.10.1111/j.1574-6941.2007.00409.xSearch in Google Scholar
Turchetti, B., M. Goretti, E. Branda, G. Diolaiuti, C. D’Agata, C. Smiraglia, A. Onofri and P. Buzzini. 2013. Influence of abiotic variables on culturable yeast diversity in two distinct Alpine glaciers. FEMS Microb. Ecol. 86: 327–340.10.1111/1574-6941.12164Search in Google Scholar
Urano, N., M. Yamazaki and R. Ueno. 2001. Distribution of halotolerant and/or fermentative yeasts in aquatic environments. J. Tokyo. Univ. Fish. 87: 23–29.Search in Google Scholar
Vilgalys, R. and M. Hester. 1990. Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species. J. Bacteriol. 172: 4238–4246.10.1128/jb.172.8.4238-4246.1990Search in Google Scholar
Visagie, C.M., J. Varga, J. Houbraken, M. Meijer, S. Kocsubé, N. Yilmaz, R. Fotedar, K.A. Seifert, J.C. Frisvad and R.A. Samson. 2014. Ochratoxin production and taxonomy of the yellow aspergilli (Aspergillus section Circumdati). Stud. Mycol. 78: 1–6.10.1016/j.simyco.2014.07.001Search in Google Scholar
Vu, D., M. Groenewald, S. Szöke, G. Cardinali, U. Eberhardt, B. Stielow, M. de Vries, G. Verkleij, P.W. Crous, T. Boekhout and V. Robert. 2016. DNA barcoding analysis of more than 9000 yeast isolates contributes to quantitative thresholds for yeast species and genera delimitation. Stud. Mycol. 85: 91–105.10.1016/j.simyco.2016.11.007Search in Google Scholar
Wang, Q.M., A.M. Yurkov, M. Göker, H.T. Lumbsch, S.D. Leavitt, M. Groenewald, B. Theelen, X.Z. Liu, T. Boekhout and F.Y. Bai. 2015a. Phylogenetic classification of yeasts and related taxa within Pucciniomycotina. Stud. Mycol. 81: 149–189.10.1016/j.simyco.2015.10.001Search in Google Scholar
Wang, Q.M., M. Groenewald, M. Takashima, B. Theelen, P.J. Han, X.Y. Liu, T. Boekhout and F.Y. Bai. 2015b. Phylogeny of yeasts and related filamentous fungi within Pucciniomycotina determined from multigene gene sequence analyses. Stud. Mycol.81: 27–53.10.1016/j.simyco.2015.08.001Search in Google Scholar
White, T.J., T. Bruns, S. Lee and J. Taylor. 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: (M.A. Innis, D.H Gelfand, J.J Sninsky and T.J. White, eds) PCR protocols: a guide to methods and applications. Academic Press, San Diego, California, USA. pp. 315–322.10.1016/B978-0-12-372180-8.50042-1Search in Google Scholar
Woudenberg, J.H.C., J.Z. Groenewald, M. Binder and P.W. Crous. 2013. Alternaria redefined. Stud. Mycol. 75: 171–212.10.3114/sim0015Search in Google Scholar
Yadav, J.S. and J.C. Loper. 1999. Multiple P450alk (cytochrome P450 alkane hydroxylase) genes from the halotolerant yeast Debaryomyces hansenii. Gene 226: 139–146.10.1016/S0378-1119(98)00579-4Search in Google Scholar
Yurkov, A.M., M. Kemler and D. Begerow. 2011. Species accumulation curves and incidence-based species richness estimators to appraise the diversity of cultivable yeasts from beech forest soils. PLoS One 6: e23671.10.1371/journal.pone.0023671Search in Google Scholar PubMed PubMed Central
Yurkov, A.M., M. Kemler and D. Begerow. 2012. Assessment of yeast diversity in soils under different management regimes. Fungal Ecol. 5: 24–35.10.1016/j.funeco.2011.07.004Search in Google Scholar
Zalar, P. and N. Gunde-Cimerman. 2014. Cold-adapted yeasts in Antarctic habitats. In: (P. Buzzini and R. Margesin, eds) Cold-adapted yeasts: biodiversity, adaptation strategies and biotechnological significance. Springer, Berlin, Heidelberg. pp. 49–73.10.1007/978-3-662-45759-7_3Search in Google Scholar
Zalar, P., G.S. de Hoog, H.J. Shroers, P.W. Crous, J.W. Groenewald and N. Gunde-Cimerman. 2007. Phylogeny and ecology of the ubiquitous saprobe Cladosporium sphaerospermum, with descriptions of seven new species from hypersaline environments. Stud. Mycol. 58: 157–183.10.3114/sim.2007.58.06Search in Google Scholar PubMed PubMed Central
©2019 Rashmi Fotedar et al., published by De Gruyter, Berlin/Boston
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 License.
Articles in the same Issue
- Frontmatter
- In this issue
- Physiology and ecology
- The future of marine biodiversity and marine ecosystem functioning in UK coastal and territorial waters (including UK Overseas Territories) – with an emphasis on marine macrophyte communities
- Thalassodendron ciliatum (Cymodoceaceae) in Palau: occurrence, typhoon impacts and changes over time
- Influence of habitat complexity on fish density and species richness in structurally simple forereef seagrass meadows
- Estuarine algal responses to increasing nitrate concentrations during closed mouth conditions of oligotrophic systems: a laboratory microcosm experiment
- Taxonomy/phylogeny and biogeography
- Achnanthales from historical Grunow collection in Porto Subzanski, Croatia
- Fungal diversity of the hypersaline Inland Sea in Qatar
- Annual reviewer acknowledgement
- Reviewer acknowledgement Botanica Marina volume 61 (2018)
Articles in the same Issue
- Frontmatter
- In this issue
- Physiology and ecology
- The future of marine biodiversity and marine ecosystem functioning in UK coastal and territorial waters (including UK Overseas Territories) – with an emphasis on marine macrophyte communities
- Thalassodendron ciliatum (Cymodoceaceae) in Palau: occurrence, typhoon impacts and changes over time
- Influence of habitat complexity on fish density and species richness in structurally simple forereef seagrass meadows
- Estuarine algal responses to increasing nitrate concentrations during closed mouth conditions of oligotrophic systems: a laboratory microcosm experiment
- Taxonomy/phylogeny and biogeography
- Achnanthales from historical Grunow collection in Porto Subzanski, Croatia
- Fungal diversity of the hypersaline Inland Sea in Qatar
- Annual reviewer acknowledgement
- Reviewer acknowledgement Botanica Marina volume 61 (2018)
