Welcome to IMC 2018 International Mycological Congress
Fungi of the National Botanic Gardens, Glasnevin, Ireland – A taxonomic species inventory.
- H. Fox
- M. Cullen
The National Botanic Gardens habitat is a mature, dynamic arboretum with tended lawns growing a wide range of Temperate Zone garden plants, over calcareous and alluvial soils on Carboniferous limestone bedrock. There are old limestone perimeter walls and some 100-year-old quarried siliceous rock surfaces, with limited habitat for aquatic fungi. The site has experienced land use continuity as a public garden since 1795. The educational use of a mycological inventory, for this 19-hectare urban Botanical Garden, is to guide visitors to specific places in the garden where they can relocate and view perennial, ephemeral or transient species of fungi and lichens. This fungal species re-finder tool empowers our visitor guide staff for outdoor educational demonstration of the visual macroscopic features and other taxonomic characters of these species to visitors to Glasnevin. Macrofungi, microfungi on land plants, and lichenised fungi were identified using mycological literature, with growth visible to the naked eye using a x10 hand lens in the field and compound microscopy. Genetic techniques have not been employed as yet. This site inventory catalogues the contributions of over 100 different people and covers the time between 1795 and 2017. The entire Irish mycological chorological literature and distribution catalogues listing species record abstracts from County Dublin was filtered. Herbarium archives and a card index maintained by the late Maura J.P. Scannell from 1970 to 1989 formed the nucleus of pre-1990 records of 150 species. Original fieldwork since 2011 to extend the inventory was undertaken. Specimens were studied using standard taxonomic literature. An inventory of 540 species was prepared from over 1800 records of fungi and lichens in the National Botanic Gardens. Most species are known from one to a few dates per decade. The very low temporal and spatial density of agaric fungi in this city garden inventory is an interesting phenomenon, with a trend for a recent increase. The increase in corticolous lichen ascomycete diversity and density since the 1970s is shown clearly in the data, and this relates well with the big improvement in air quality in the city since 1989, and by the increased crown diameters of trees in the arboretum. Records of perennial and ephemeral fungi and lichens on site provide an interpretative window to 40% of 1500 species now known from County Dublin. For a student visiting and studying fungi and lichens at Glasnevin, in any week of the year, not more than 20% of the County Dublin mycota can be shown to them in situ. The general spatial rarity, temporal infrequency, and small biomass of colonies of most of the 540 species of fungi in this urban habitat merits further ecological explantion.
The National Institute of Science and Technology Virtual Herbarium of Brazil: a successful network for sharing herbarium data and images
- L. Maia
- A. Peixoto
- A. Vieira
- D. Canhos
- J. Stehmann
- M. Barbosa
- M. Menezes
Brazil’s flora is considered the richest in the world, and its mycota is also estimated as among the most diverse on the planet. This immense natural wealth constitutes a scientific, economic and cultural heritage that must be scientifically recognized, preserved, and used in a sustainable way. Brazil has more than 200 herbaria; 65% of them have less than 20,000 specimens and only 9% contain more than 100,000 exsiccatae. The National Institute of Science and Technology (INCT) Virtual Herbarium of Brazil project has brought together data from more than 100 of these herbaria, from all States of the country, and from 21 foreign herbaria, by an intensive and extensive networking effort of curators, technicians and researchers. Its mission is to: Expand Brazil’s knowledge base on flora and fungal diversity; Improve the quality of Brazilian herbaria; Encourage public policies to ensure sustainability of herbaria, training of taxonomists, and support to biodiversity studies; Make species occurrence data a fundamental instrument for decision making and for the formulation of public policies; Encourage free and open data and information sharing in a comprehensive, useful and usable format; Offer data and information to make environmental sustainability just as important as social and economic development in the formulation and assessment of public policies. In eight years, the INCT-Virtual Herbarium achieved its initial objectives and changed a paradigm by stimulating institutions and curators to share information while reinforcing the networking of collaborators that are willing to improve the collections and data quality, as well as in training personnel. The activities of the project include improvement and upgrading of collections’ management and data quality (mainly the accurate identification of fungi and plants – 200 visits were carried out by 80 specialists to 70 herbaria, some more than once); training of students, technicians, and curators (60 courses were given by > 80 specialists, benefiting > 930 people), and technology development (with a range of digital tools: data cleaning, annotation system, lacunas, biogeo and statistics of data use) to make herbarium data and images available on-line. The INCT-Virtual herbarium adopted the speciesLink platform as the basis for its information system and the data are also available in other platforms (GBIF, IDigBio, and SiBBr). Through this network it has been possible to increase the knowledge about Brazil’s biodiversity, disseminating about 6 million records and 1.9 million images of fungi and plants (http://inct.florabrasil.net). The INCT-Virtual Herbarium of Brazil represents a successful multi-institutional data-sharing network that allows access to the entire world, in a free and open way, to most fungi and plants recorded in the country. The Project, with its continuity, expansion and improvement recently approved for six more years, is supported by: Conselho Nacional de Ciência e Tecnologia (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), and Fundação de Amparo à Ciência e Tecnologia de Pernambuco (FACEPE).
Utilization gaps: mushrooms as a means of poverty alleviation and forest conservation in the Chin State, Myanmar
- P. Mortimer
- S. Karunarathna
The Greater Mekong Subregion (GMS) is well known for the high diversity and abundance of macrofungi, this diversity is coupled to the varied of landscapes and forest types across the region. The GMS also includes numerous rural areas, characterized by low income levels, poor access to markets, poor infrastructure, and high levels of forest degradation. Within this context, we have selected a number of villages in the Chin State of Myanmar to study the extent of traditional knowledge relating to the use of mushrooms, and how mushrooms can be used as a means of poverty alleviation and forest conservation. Preliminary studies on the diversity of mushroom species in the forest systems (fallow forest and natural forests) surrounding the villages and workshops with the villagers to assess their knowledge of the mushrooms collected have been carried out. Dominant tree species in both the fallow and natural forests included Alnus nepalensis, Pinus kesiya, Quercus spp., Castonopsis sp, and Rhododendron arboretum. We collected 55 mushroom species of which 19 were edible, and 9 were of commercial interest - Thelephora ganbajun, Termitomyces clypeatus, Auricularia auricula-judae, Ganoderma leucocontextum, Cantherellus sp., Lactarius sp., Russula albida, and R. virescens. The 30 (14 women, 16 men) villagers interviewed in our study had almost no knowledge regarding the use of the mushrooms collected, and only two individuals had a basic knowledge (they recognized 3 species that could be eaten). All participants expressed a desire for further training and saw the potential of mushrooms as a resource, both for household nutrition and trade. All participants expressed a strong fear of mushrooms poisoning. In order to help address the fears and potential dangers of mushroom poisoning we are currently developing a social media platform for the uploading of images of collected mushrooms to assist in identification of collected specimens. Currently the biggest hurdles to trade are lack of knowledge and limited market access.
Teaching mycology by the methodology of problematization with an Antarctic lecture
- D. Dos Santos
- I. Leite Filho
There are few researchers in fungi field comparing to the others biological fields. Learning about the Fungi Kingdom in high school is essential for stimulating new researchers and new scientific investigations concerning these living beings. In fact, there are many teaching methods to guide teachers in and out the classroom. However, both novice and experienced scientists must be able to apply the scientific method to investigations. Students learning regarding mycological scientific concepts through the observation of reality, providing key points that lead to theorization and hypothesis collection and later, application to reality which can be an effective procedure. This study aimed to build knowledge about fungi, from an Antarctic experience for the scientific literacy of high school students by the Methodology of Problematization. To achieve this goal, four theoretical-practical classes were developed and delivered, in and out the classroom, using the science-teaching laboratory. This classes were categorized and analyzed by words clouds thru transcription of the classes recorded audios and logbooks. All students in the classroom were separated into three groups providing the learning of the fungi by research, among which the scientific contents were conducted in the form of experiences beginning with a lecture-class, followed by three theoretical-practical classes. These activities show many possible indicators of scientific literacy. Thus, the results of the sequential classes indicates the importance of social interaction between students-teacher and students-students to promote learning, but also contributed to the continuous improvement in the methods of teaching learning process. In fact, interesting investigations points can be stimulating for student’s scientific learning. In short and in conclusion, there is a strong need to learn science in a scientific way, contributing to scientific literacy and, consequently, the substantial learning of scientific concepts, so that future generations can apply such knowledge acquired in basic education to their research in higher education, and may even to leverage exponentially the Brazilian research developed in the Antarctic continent.
New edible fungi from Southeast Asia: discovery to production
- S. Karunarathna
- K. Hyde
- P. Mortimer
- J. Xu
- A. Bandara
The forests of Southeast Asia have the potential to be a rich source of cultivatable edible fungi. Although significant amounts of research on the taxonomy and phylogeny of edible mushrooms have been carried out, far fewer studies have focused on the domestication of wild fungi. Today, the most commonly cultivated strains are temperate species, but tropical and subtropical mushrooms are both abundant and highly diverse, with many species having long histories of human consumption. In addition, many new species have recently been introduced to science, including numerous species of high nutritional and medicinal value. The domestication and cultivation of tropical mushrooms therefore provides an enormous opportunity for Southeast Asian countries. Due to the difficulties of cultivating mycorrhizal species, we have concentrated on saprobic species. Most tropical and subtropical mushrooms, if provided with appropriate conditions, grow and produce fruiting bodies more quickly than temperate species. Tropical and subtropical mushrooms can be produced using cheap, readily available waste products such as sawdust, corn cobs, rice straw, sugarcane bagasse, and other forest and agricultural residues, making them an ideal crop for smallholder farmers. We have collected and isolated numerous strains of wild mushroom species from Southeast Asian forests, and have published some initial results documenting our progress in domesticating these species. Using a variety of steps including sample collection, isolation, spawn production and fruiting body production in sawdust and compost media, we showed for the first time that it is possible to domesticate the following fungi: Pleurotus giganteus; a new Thai-French hybrid strain of Agaricus subrufescens; A. flocculosipes; A. subtilipes; Auricularia thailandica; A. cornea (white); Panus roseus; Macrolepiota dolichaula; Ganoderma australe; and G. leucocontextum. These discoveries may create new opportunities for the mushroom growing industry and for smallholder farmers in Southeast Asia in particular.
Economically valuable mushrooms of Armutlu (Yalova)
- S. Candar Erol
- H. Alli
- I. Akata
In this study totally 123 economically valuable macrofungi specimens have been collected from different localities of Armutlu (Yalova) province between 2010 and 2014. As a result of field and laboratory studies, 23 species within the 16 families and 6 orders were identified. Two of them belong to the division Ascomycota and 21 to Basidiomycota. The species list is given with the informations on localities, habitats, collecting dates and collection numbers.
Edible Fungi of woodlands and plantation forests of Ireland
- M. Cullen
Sustainable development of edible mushroom foraying and the planting of trees in natural, native mycorrhizal associations with edible fungi in Ireland are explored.
A compilation of geographical, botanical and temporal occurrence data on edible species of mushrooms and their forest vegetation contexts from historical and original observations, plus data analysis thereof, were chief objectives. Knowledge development from a low base was important as an educational resource and to contribute to effective policies for fungal conservation and good forest management. The development of safe and respectful ways to interact with the beauty, diversity and flavours of wild, edible Irish fungi were priorities.
Ireland has no Red Data List for fungi or any legislative protections for fungal species, apart from one lichen. Care in publication of localities of edible fungi is required as the resource is patchily distributed and vulnerable to greed.
Along with extensive personal records since 1999, a nationwide survey of forests was undertaken. Recording of 100 plots of 50x2m were monitored across the main woodland and plantation forest types and ecological settings of Ireland on several occasions between September and December in 2007, 2008 and 2009. Carpophore counts and masses of edible species as well as ground flora and tree information from plots were databased. Historical literature detailing all records of edible species in Ireland was reviewed and herbarium voucher material examined.
Russula ochroleuca (marginally edible) and Laccaria laccata were the most widespread species found. While results were generally positive, a high proportion of the exotic, coniferforested area in Ireland was found to be unproductive for prized edible fungi.Awareness and training for the opportunities are still unsupported. Craterellus species in the Winter Chanterelle group currently offer potential and some genetics work was carried out on this group.
Continuous cover forest methods would improve continuity of edible fungi in Irish forests. Forest pathology expertise has been limited. Care needs to be taken not to transfer parasitic organisms with nursery stock soil or on wood-cutting and chipping equipment.
Truffles are more common than previously thought. Beech provides an important mycorrhizal host association for Tuber aestivum in Ireland as well as in England, Sweden and north-western France.
The impact of Armillaria is of concern for the health of deciduous forests. The lack of control measures for this and other parasitic fungi are challenges for securing healthy populations of edible fungi and their host trees.
Imported, inoculated nursery stock for plantation forestry, as well as treatments involving mycorrhizal products have influenced mycorrhizal populations. Alien mycorrhizal inoculants and soil additives with fungal components are potential threats to native fungal biodiversity. It is proposed that inoculated stock and fungal “ameliorants” be subject to legislation for myco- and phytosanitary risks.
Lessons were learned from experiences at the National Poisons Information Centre and from serving on the panel for fungal expertise there, as well as from experiences of mushroom poisoning cases in other countries. There have been improvements to education for the public, with medical professionals and staff in advisory positions.
Augmentation of nutritive value of rice using Rhizopus oligosporus through fermentation
- R. Hafeez
The production of fermented food is one of the oldest food processing technology. Many of these foods have been manufactured because their unique flavor, aroma and texture attributes are much appreciated by the consumer. Moreover, because filamentous fungi simultaneously decrease anti-nutrient compounds and partially hydrolyzed substrate biopolymers, pre-fermented bio-product may be used as an inexpensive food and feed supplement that may support marketing claims. The results of fermented rice (variety super 85) with Rhizopus oligosporus at 30ºC for time period of 28 hours was a new food product consisting of soft rice grains with fruity pleasant smell and sweet taste. Chemical composition of fermented and un-fermented rice in respect of %age of moisture contents, protein, crude fat and crude fiber was 42.43, 1.75, 4.1, 9.106, 1.84 and 7.3. In case of calcium highest value was noted in fermented aerobic rice sample 19.36mg/100g.The highest value of phosphorous was calculated in processed control sample i.e. 3.328mg/100g. The highest value of phytic acid was calculated in simple rice sample 0.856%. GABA (aminobutyric acid) was detected in fermented aerobic and anaerobic rice samples with highest Rf value as compared to simple and control rice samples. Rf value of fermented anaerobic and fermented aerobic rice samples was 7.22 and 7.11 respectively.
Key Words: Food biotechnology, Fermentation, Rice, Rhizopus oligosporus, GABA.
Heterologous gene expression of Aspergillus oryzae strain isolated from Korean traditional fermented foods
- S. Jun
- B. Zhu
- M. Lee
- J. Yu
- J. Kim
- K. Han
It is important to find an efficient way to enhance the expression of heterologous gene(s) of interest in a fungal expression system for giving the high potential of fungi as genetic resources. Protein expression systems that produce the heterologous gene products using fungi are important for various aspects and it is recommended to use the GRAS strain as a host to ensure product safety. Therefore, here we constructed a heterologous gene expression system for producing foreign gene product including bacterial origin one such as bacterial β-glucosidase by using a GRAS fungus Aspergillus oryzea. The produced β-glucosidase is a hydrolytic enzyme and the expression of the gene was stimulated by placing it under the control of the constitutively activated gpdA gene promoter or threonine-inducing alcA gene promoter of Aspergillus nidulans. The pyrithiamine-resistant gene, ptrA, was used as the selection marker for Aspergillus transformation. The signal peptide of A. oryzae α-amylase AmyB was linked to the N-terminus of the bacterial β-glucosidase protein, and 3X FLAG was tagged at the C-terminus. A. oryzae transformants successfully overexpressed the β-glucosidase gene, and expression level was monitored by western blot analysis with anti-FLAG antibody. The functional activity of the protein was detected by esculin hydrate converting test and pNP-β-D-glucopyranoside (pNP-Glc) measurement assay. The expression system of A. oryzae could be beneficial for industrial applications.
The historic and current ethnomycology of Egypt and Middle East countries
- A. Abdel-Azeem
Egypt is considered the cradle of mycology. Ancient Egyptians documented the use of fungi on walls and pillars of temples, within hieroglyphic texts, ear studs and medical prescriptions since 5619 B.C. Ancient Egyptians believed that some mushrooms were plants of immortality and called them “a gift from the God Osiris” and symbolized as Was, Djed pillar of Osiris, and ankh (crux ansata). Egyptian pharaohs proclaimed mushrooms to be food reserved only for royalty; common people were not even allowed to touch them. The Pharaohs of ancient Egypt believed they had magical powers. Egyptian crowns (white and triple) were inspired from the primordia of Psilocybe cubensis. The most ancient historical use of truffles probably originated prehistorically in the mideastern and North African cradles of civilization. Species of desert truffles (Terfezia, Tirmania and Phaeangium) probably served to the Pharaohs. Better descriptions of the kind of desert truffles that the pharaohs of Egypt may have consumed, along with an ancient version of traditional truffle preparations still popular in North Africa and the Middle East, can be found in the Bible. In the seventh century Prophet Muhammad peace be upon him (صلى الله عليه وسلم) said a hadith to his followers “Truffles are a part of manna and its juice is healing for the eyes”. This study consists of a survey focusing on the knowledge, use and ethnomycological practices of mushrooms and desert truffles among the native people of the Middle Eastern countries. The presentation will highlight their application in traditional medicine in this part of the world. This work also explores the biology and ecology of truffles in the Middle East, their importance in fragile desert ecosystems, assess their conservation status and effects of various cultivation practices on sustaining truffle populations. General management principles and considerations to sustain this valuable fungal resource will be discussed.
First record of edibility and ethnomicological notes of Ustilago maydis in Ecuador
- J. Gamboa
Ustilago maydis is known by the Ecuadorian local communities with the vernacular name of "Cuscungo", for a long time its use was unknown for Science, however in the last decade ethnomicological studies have been carried out in corn crops, located in the Ecuadorian mountain range, where informal surveys were applied to people from 40 to 80 years of age to know ethnomicological data. In this way, its uses were found with the categories: edible human, edible animal, ludic among the most representative. This fungi represents balance for human communities, traditional areas of cultivation and ecosystems.
Key words: Ustilago maydis, ethnomicolgy, Ecuador.
Evaluation of antimicrobial and cytotoxic activites of secondary metabolites produced by Aspergillus flavus isolated from Cerrado Goiano
- E. Couto
- R. Silva
- T. Silva
- F. Almeida
- V. Monteiro
Microorganisms represent an important source of bioactive compounds with high added economic value. Among the microorganisms, fungi have many advantages of use due to their potential as producers of secondary metabolites. Due to their bioactive properties, many secondary metabolites have been adopted for pharmaceutical use as antibiotics, tumor inhibitors and immunosuppressants among other properties.The goal of this study was to investigate the antimicrobial and cytotoxic activities of secondary metabolites produced by Aspergillus flavus fungus isolated from the Cerrado Goiano soil.The production of the secondary metabolites was evaluated by fermentation in submerged culture in TLE medium and after fermentation, the medium was extraction by liquid-liquid partition with ethyl acetate. The secondary metabolites were analyzed by high performance liquid chromatography (HPLC). The isolates of A. flavus were used to evaluate the toxicity in Artemia salina, evaluation of susceptibility to antimicrobial, and cytotoxic evaluation by the MTT. In the evaluation of the toxicity Artemia salina in The compounds isolated from the fungi were tested in concentrations of 2400, 1200, 300, 75 e 18,75 µg. mL-1. The analysis of the tests tested gives high toxicity (LC50 = 11.49 μg mL mL -1) to potassium dichromate, within the standards presented in the literature, attesting to the suitability of the experimental conditions. According to the classification of Nguta et al. (2012), in this assay the extract of secondary metabolites presented LC50 equivalent to 81 μg.mL-1 this result was considered of high toxicity according to the parameter adopted. To evaluate the antibacterial activity of the isolated compounds, the inhibition test was carried out by dilution in broth using the concentrations of 40 µg.mL-1; 80 µg.mL-1 e 160 µg.mL-1 was tested against standard bacteria ATCC (S. aures e E. coli) e MRSA (Methicillin-resistant Staphylococcus aureus). As a positive control, Chloramphenicol at 100mg.mL-1 concentration for E. coli and Oxacillin at 100mg.mL-1 concentration for S. aureus were used. The concentration of 80 μg.mL-1 was able to inhibit 100% E. coli and 95% S.aureus and 98% MRSA. The concentration of 160 μg.mL-1 was able to inhibit 100% the bacteria tested. A. flavus was subjected to the cytotoxicity bioassay against WM1366 and A375 tumor cells. WM1366 and A375 lineage cells (5x105 / ml), and human mononuclear cells (1.5 x106 cells / ml) were cultured in the presence of A.flavus extracts at concentrations of 6 to 48 μg / mL in 96-well plates. After the 24 h incubation period, the enzymatic activity was determined by the enzymatic reduction of 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Secondary metabolites were efficient in promoting cytotoxic effect on WM1366 and A375 cells in a dose-dependent manner. Thus mitochondrial activity was decreased by 98% and in WM1366 cells mitochondrial activity was decreased by 100%. In conclusion, the metabolites produced by Aspergillus flavus presented potent antimicrobial and antitumor activities, evidencing the importance of future studies.
Elevated CO2 concentration expands the range of temperature and water activity conditions wherein Aspergillus flavus produces aflatoxin B1
- M. Gilbert
- B. Mack
- G. Obrian
- A. Rodriguez
- D. Bhatnagar
- G. Payne
- N. Magan
- A. Medina
There is a significant interest on understanding the impacts that climate change factors [CO2 concentration, water availability (aw) and temperature increases (T)] may have on mycotoxigenic fungi. We have, for the first time, examined on conducive media and maize grain the impact that three-way interactions between these factors (aw, T and elevated CO2) have on: (i) growth, (ii) the relative expression of all genes in the aflatoxin gene cluster using both RT-qPCR and RNAseq, and (iii) the phenotypic aflatoxin B1 (AFB1) production by Aspergillus flavus. On conducive media, interactions between water stress (aw; 0.97, 0.95, 0.92), temperature (34, 37oC) and CO2 exposure (350, 650, 1000 ppmv) were considered and the growth, AFB1 production and expression of biosynthetic genes (aflD, aflR) studied by RT-qPCR. For maize grains, interactions between water stress (aw; 0.99, 0.91), T (30, 37oC) and CO2 exposure (350, 650, 1000 ppmv) were included. Fungal growth, AFB1 production and expression of the all genes in the aflatoxin gene cluster were studied by RNAseq. The results showed that for growth there was relatively little effect. In contrast, the three-way interacting conditions (elevated CO2, aw, T) had a profound effect on AFB1 production both in media and maize grains. Importantly, under slightly elevated CO2 conditions there was a stimulation of aflatoxin B1 production. Although under current CO2 concentrations at 37oC there was no or minimal production of aflatoxins, our results show that, in conducive media and 650 and 1000 ppm CO2, there was a significant increase in expression of both aflD and aflR at 0.95 and 0.92 aw. There was an associated increase in AFB1 in these treatments. Similar observations were made in stored maize grain. Differential expression of several genes was found by RNAseq in the aflatoxin gene cluster in relation with these interacting factors. The range of aw and Ts in which aflatoxin B1 is produced increased under elevated CO2 conditions. This is the first study to examine these three-way interacting climatic factors on growth and mycotoxin production in A.flavus. This provides data which is necessary to help a better and more accurate prediction of the real impacts that climate change is and will be having on important staple food and feed chains due to mycotoxigenic fungal spoilage and toxin contamination.
New methodology to improvement the sanitary quality of stored –in pod peanut by in situ application of pure BHA (2 (3) ter-butil-4 hidroxianisol) and their formulation
- D. Garcia
- N. Girardi
- M. Passone
- A. Nesci
- J. Garcia
- M. Etcheverry
In Argentina in-pod peanut intended for use as seed for planting are stored from 3 to 6 months (from harvest to planting) where fungal contamination could increase resulting in a decrease of the grain quality during storage. However, only germinative power and vigor of the grains are normally evaluated, while sanitary quality is not considered. For this, the aim of this work was to evaluate the effect of pure BHA and in combination with their formulation (F-BHA) on peanut seeds stored in open containers in the field.
The experiment was carried out in a storage company of south of Córdoba, Argentina during 2017. Two hundred kilograms of in-pod peanut treated with: Treatment 1 (T1) pure BHA (10mM); Treatment 2 (T2) the mix of pure BHA and F-BHA (5mM: 5mM); Treatment 3 (T3) pure BHA (5mM) at the bottom of the peanut container and F-BHA mix in the peanut and Treatment 4 (T4) untreated grains, were used. Fungal level, residues of BHA and fluctuations of temperature and humidity during 4 months were determined.
Fungal count was significantly affected by treatment and time according to ANOVA test (p<0.05). This effect was strongly observed after 30 days where control grains showed fungal level 1.6 times higher respect with the treated grains. At this time, no significant differences (p<0.05) were found for CFU/g levels between T1 and T2. However, T3 showed fungal levels 20% higher respect T1 and T2. Among the most frequently isolated fungal genera were Penicillium, Cladosporium, Alternaria and Fusarium.
For the both treatment, BHA residual levels showed a pick of the amount of the antioxidant after 30 days of the assay. At this time values of 924.28, 748.72 and 193.2 ng/g were found for T1, T2 and T3 respectively. At the final of the experiment levels decreased to reach 110 ng/g for T1; 330.81 ng/g for T2 and 28.10 ng/g for the T3. The temperatures recorded during the sampling fluctuated between 23 and 35 ° C according to the month in which the sampling was taken, while the humidity of the peanut grains remained around 5% during all assay.
The treatments used in this study on in-pod peanut, reduced significantly (p<0.05) fungal development of the seeds stored in field. This reduction was strongly observed during the first months of storage coinciding with the highest levels of antioxidant detected in the samples of peanut. On the other hand, the highest levels of BHA present in peanut kernels were observed for the combined treatment (T2) where the final amounts were around 3 times higher compared with T1 (pure antioxidant) and 12 times higher compared with T3. The combined treatment (T2) could be an alternative for control of spoilage fungal on in-pod peanut stored in the field where conditions may not be suitable for storage.
Biodiversity of marine-derived Trichoderma spp. and their exploitable biological activities
- Y. Heo
- K. Kim
- S. Jang
- S. Kwon
- M. Park
- Y. Lim
- J. Kim
- H. Lee
Natural metabolites derived from microorganisms are an important source of novel medicinal compounds. Since 1950s, secondary metabolites from marine-derived fungi have been discovered, many of which have unique structures and have remarkable biological and pharmaceutical properties. A genus Trichoderma is found in a variety of environments and has been reported to produce diverse biologically active secondary metabolites. However, studies on the secondary metabolites of Trichoderma derived from the ocean are relatively rare. This study investigated the diversity and physiological activity of 35 Trichoderma spp. isolated from various marine resources such as sea sand, seaweed, tidal flat sediment, and Arctoscopus japonicus eggs in South Korea. They were identified using the DNA sequence of the translation elongation factor region and the 50% majority conformational tree was constructed through Bayesian analysis. To make fungal extract, all fungi were cultivated in potato dextrose agar for a week and extracted with MeOH. The MeOH extract was dissolved in EtOAc and D.W. (1:1), and the EtOAc layer was taken and dried using a rotary evaporator. The final extract was dissolved in dimethyl sulfoxide (10 mg/mL) for subsequent bioassays. The antioxidant activity of the extract was measured by an analytical methods using 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) and 2,2-diphenyl-1-phenylhydrazide. As a result, they were identified as 18 different Trichoderma species including three new species candidates and six species not reported in Korea. The fungal extracts of Trichoderma bissettii and Trichoderma longibrachiatum exhibited high radical-scavenging activities in both assays. It is expected that marine-derived Trichoderma spp. can be a great resource for bioprospecting useful natural compounds. The antifungal activities of the extracts are currently being investigated.
Exploring the diversity and bioactivity of cultivable Arctic marine fungi from the Svalbard archipelago
- O. Hagestad
- T. Rämä
- B. Altermark
- E. Hansen
- J. Andersen
The diversity of marine fungi in Polar Regions, especially Arctic, is severely understudied and is a potential source for novel marine natural products. It can be hypothesized that due to extreme conditions, such as low temperatures, Arctic marine fungi possess unique chemical adaptions of biotechnological relevance. The aim of the study was to get insights into the diversity of marine fungi at high latitudes and screen fungal extracts for the presence of novel bioactive compounds. Fungi were isolated on three media from driftwood, sediments, macro-algae and pelagic water around the Arctic archipelago of Svalbard in 2016. Subculturing was conducted until pure culture isolates were obtained and these were sequenced using ITS5-4 primer pair amplifying the ITS rRNA region. The sequenced isolates were grouped into operational taxonomic units (OTUs) using a 98% sequence similarity cutoff value and a representative isolate from each OTU was sequenced for SSU and LSU rRNA genes. As a preliminary test of the isolates’ potential to produce bioactive compounds, an antibacterial plug assay was performed by testing each OTU against five different pathogenic bacteria (Escherichia coli, Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus aureus and Streptococcus group B). In total, more than 200 isolates were sequenced, and 95 of these yielded fungal ITS sequences. These were grouped into 20 OTUs. The diversity was highest in Ascomycota (17 OTUs), followed by Zygomycota (2) and Basidomycota (1). Within Ascomycota, Lulworthiales was the most frequent order with 6 OTUs. Other orders with several OTUs included Eurotiales (3), Hypocerales (3) and Helotiales (2). Identity of the obtained ITS sequences to sequences in Genbank ranged from 85-100%, indicating that some species have not been sequenced for ITS or represent novel species. Representative isolates from Eurotiales (3), Helotiales (1), Hypocerales (1) and one order of incertae sedis as well as one order from Zygomycota showed inhibiting zones against gram-positive bacteria ranging from 8-17 mm. These preliminary results will be used to select strains for further bioactivity screening and potential scale-up cultivation of isolates producing new compounds. This work will shed light on the Arctic fungal diversity and the potentially undiscovered chemical space they possess. In addition, the results of the project will be used to prioritize strains for genome sequencing with the aim of linking produced bioactive compounds with the biosynthetic gene clusters responsible for their production.
The Mediterranean Sea: an untapped source of fungal diversity
- E. Bovio
- L. Garzoli
- G. Gnavi
- P. Marchese
- A. Poli
- G. Varese
The Mediterranean Sea, with almost 12,000 endemic species, is among the 25 global biodiversity hotspots recognised worldwide. Nonetheless, this estimate does not take into account the microbial diversity that is still largely unknown. In the last years, the Mycotheca Universitatis Taurinensis (MUT) carried out several research programs to assess the marine fungal diversity in the Mediterranean Sea to better understand the specificity of the fungal community for different substrates (animals, seagrasses, algae, wood, ecc), and to find new species valuable for biotechnological exploitation (i.e. new drugs, new antifouling agents, ecc). Biotic and abiotic marine substrates were sampled during several campaigns. A culturomic approach was employed for the isolation of marine fungi; specific culture media that mimicked both the marine environment and the original substrates of isolation were developed in order to increase the number of cultivable fungi. Fungal strains were identified by combining morpho-physiological and molecular features with deep phylogenetic analysis. Today, the MUT is the largest public collection of marine Mediterranean fungi, hosting almost 1400 taxa from more than 20 different matrixes. Our results show that Ascomycota are ubiquitous in the Mediterranean Sea, while Basidiomycota and Zygomicota are less frequent. Several species were reported for the first time in marine environment, and each substrate was shown to host a specific fungal community. Interestingly, numerous species displayed unique morphological and molecular features and are currently under investigation. Remarkably, these strains could produce enzymes and molecules relevant in the interaction with the host and the surrounding environment. With the present work, we contribute to increase the knowledge on marine fungal diversity.
Assessment of antioxidant activity of some medicinal fungi harvested in forested and mountainous regions of Armenia
- V. Gevorgyan
- A. Gilovyan
- T. Seferyan
- L. Margaryan
- S. Nanagulyan
Many fungal strains show strong antioxidant activity, comparable to classic antioxidants, such as Vitamin C or E. In presented research we have investigated some major compounds responsible for antioxidant activity for 6 different species of fungi (Hypholoma fasciculare, Agaricus bisporus, Pleurotus ostreatus, Trichaptum abietinum, Polyporus squamosus, Schizoplyllum commune), growing in Republic of Armenia. All species of fungi were harvested in June-August 2015 in forested and mountainous regions of Armenia. Antioxidant activity was assessed via potentiometric method based on oxidation/reduction of Fe2+/Fe3+ in acetate buffer media (pH=3.6). Phenolic content of fungi was determined by spectrophotometry, using gallic acid as standard, according to the method described by the International Organization for Standardization (ISO) 14502-1. Flavonoid content was determined by spectrophotometry via reaction with AlCl3 in methanol at 410nm wavelength, using rutin as standard. β-carotene and lycopene contents were determined by spectrophotometry at 453nm, 505nm and 663nm wavelength. Results indicate that all investigated species show antioxidant activity except Trichaptum abietinum. Agaricus bisporus and Pleurotus ostreatus show highest antioxidant activity (8.8 x 10-3 g/l and 6.8 x 10-3 g/l accordingly) among investigated species. Phenolic compounds have a significant role, but there are other antioxidants present as well. Hypholoma fasciculare has highest phenolic content (0.665 g/l). It also has highest β-carotene and flavonoid content (0.324 g/l and 2.505 g/l accordingly), and Schizophyllum commune and Polyporus squamosus contain trace amounts of lycopene. Further researches of antioxidant activity on lipid membranes and biological objects, such as tissue, cell cultures are in progress.
Biologically active compounds in lichenized fungi
- M. Temina
- V. Dembitsky
Lichenized fungi (usually referred to as lichens) are unique symbiotic organisms that grow in spots on substrates that are too harsh or limited for most other organisms such as bare rock, desert sand, cleared soil, dead wood, animal bones, rusty metal, living bark, and so on. Lichens consist of two organisms: a fungus (mycobiont) and an intimately associated photosynthetic partner (photobiont). The photobiont can be either a green alga or a cyanobacterium, and some lichens can contain both alga and cyanobacterium. As adaptations for life in specific habitats, lichens produce a large number of chemical compounds (metabolites); almost all of them are unique as they are unknown in other plant sources. Lichen metabolites possess numerous biological activities including antiviral, antibacterial, anti-inflammatory, analgesic, antitumor, etc. Thus, lichens may be of interest for pharmaceutical sciences and the industry itself. The goal of the present research was to study the metabolites of lichens distributed in Israel. We collected different lichen species dwelling on trees, soil and rocks in the Mediterranean territory of Israel and investigated the composition and structure of the chemical compounds isolated from them. All studied species had a high diversity of lipids, fatty acids, aromatic compounds and other metabolites. Metabolites of two fruticose epiphytic species (Ramalina lacera and Tornabea scutellifera) and four foliose epilithic species from the genus Collema (C. cristatum, C. callopismum, C. fuscovirens and C. flaccidum) were studied in detail the most. It is to be noted that photobionts of Ramalina lacera and Tornabea scutellifera are green algae, while photobionts of species from the genus Collema are cyanobacteria. Four compounds belonging to monotetrahydrofuranic acetogenins (Tornabeatins A, B, C, and D) have been isolated as new natural products from Tornabea scutellifera, and their biological activities were tested. The studied metabolites showed modest activity against different microorganisms and significant antitumor activity. Testing of biological activities of rare fatty acids isolated from Ramalina lacera and Collema species detected significant antibacterial and antifungal activities of these compounds. A photo protective mycosporine (Collemin A) was found in calcicolous lichens Collema cristatum and C. callopumum. The study of this compound showed that it prevents UV-B induced cell destruction in a dose-dependent manner, partially prevents pyrimidine dimer formation, and completely prevents UV-B induced erythema when applied to the skin prior to irradiation. Thus, our studies revealed that lichens distributed in Israel might be good potential sources of bioactive phytochemicals.
Differentiation of fungal communities between different crops and substrates in an intercropping trial
- R. Tonjock Kinge
- E. Casson
- M. Gryzenhout
Isolation based methods have previously revealed that fungal communities occurring in soil, the plant rhizosphere, within plants and often between plant species are usually different with some fungi having the ability to occur across niches and plant species. However, very few of these studies consistently and comprehensively compared fungal communities between all of these variable origins, most likely due to the vast number of isolations and morphological or DNA sequence based identifications that would be needed. The advent of environmental sequencing and high throughput sequencing technologies, however, made such large scale comparisons doable. This study represents comparisons of the mycobiomes present as endophytes within four legumes and sorghum planted in an intercropping trial. These plant endophyte mycobiomes were also compared with the fungi occurring in the rhizospheres of the plants, as well as bulk soil from the various plots. As expected a great deal of overlap as well as differences could be observed between the mycobiomes of the different crops. Similarly, fungal communities from bulk soil, rhizosphere soil and plant communities were mostly distinct. Moreover, these profiles also differed between crops. The approach developed in this study enables tracking interactions and development of fungal communities during intercropping experiments, as well as the mycobiomes present in different niches. This is important when such planting trials aims to promote beneficial organisms and deter detrimental pathogens.
The maize fungal community under various agricultural practices.
- L. Grubisha
- Z. Gates
- L. Deweese
Fungal and bacteria communities play important roles in biogeochemical processes. Understanding the role of microbial diversity in agricultural systems is important for developing long-term management plans. While numerous studies have examined forest and grassland ecosystems, less is known about agricultural systems in the Midwest USA. Furthermore, even less is known about microbial communities of conventional and organic agriculture. A goal of this study is to compare fungal diversity in organic vs convention corn rotation cropping systems using different tillage regimes. The University of Wisconsin Extension Farm at Arlington has been assessing various farming practices for several decades. We sampled five cropping treatments, each with four replications. Three soil samples per field were collected using 15 cm x 2.5 cm soil cores. DNA was isolated from 60 soil samples. Microbial communities will be identified using next generation DNA sequencing. PCR was performed using Illumina barcodes and ITS primers for the ITS1 region of the ribosomal DNA repeat. Analysis of fungal communities under different agricultural methods associated with maize farming in Wisconsin will be presented. These results will provide information on how current agricultural practices affect soil fungal biodiversity. Corn is a major crop of Wisconsin and thus this data will provide important information for Wisconsin farmers.
Characterization of the fungal phytobiome of sorghum (Sorghum bicolor) using environmental sequencing
- G. Pambuke
- R. Tonjock Kinge
- E. Casson
- M. Nyaga
- M. Gryzenhout
The phytobiome consists of microbes associated with external and internal areas of a particular plant. These organisms may interact with each other and with the plant, which may have beneficial of detrimental effects. Environmental sequencing using next generation sequencers enables intensive characterization of the phytobiomes of plants. In this study we characterized the fungal component occurring as endophytes from all plant tissues of sorghum (Sorghum bicolor), as well as fungi from the rhizosphere and surrounding bulk soil across two years. Sorghum is one of the most cultivated crops in the world, and widely used in rural agriculture. Complete plants, rhizosphere soil and bulk soil were sampled from Potchefstroom, South Africa, and processed for environmental sequencing of the ribosomal internal transcribed spacer regions using the MiSeq Illumina sequencer. Results indicated significantly different communities of fungi from the different substrates (plant, rhizosphere, soil) and a level of organization between plant tissues. A number of plant pathogenic genera were also detected. Using environmental sequence, these taxa can be monitored more easily over time and their interactions with agricultural practices and environmental factors can be characterized.
Assembly of Sorghum’s mycobiome
- C. Gao
- L. Montoya
- L. Xu
- M. Madera
- J. Hollingsworth
- R. Hutmacher
- J. Dahlberg
- D. Colemann-Derr
- P. Lemaux
- J. Taylor
Fungal ecologists have long debated whether the assembly of the Mycobiome is deterministic, stochastic or strongly influenced by historical contingency, and are especially curious about the processes responsible for the observed patterns. We investigated the assembly of the Mycobiome from weekly sampled leaves, roots, rhizosphere and soils over the growth season of sorghum [Sorghum bicolor (L.) Moench] that had been subjected to pre-flowering drought, post-flowering drought or no drought (control), using fungal ITS2 DNA sequence (Illumina, MiSeq). Stochasticity (Raup-Crick index between -0.95 and 0.95) was demonstrated early in the growth season, when fungi are rare on newly emerged leaves (weeks 1-7) and roots (week 2). The observed stochasticity was attributed to ecological drift because its strength was strongly negatively correlated with fungal community size. The stochasticity could not be attributed to dispersal or historical contingency because there was no evidence of distance-decay, stochastic colonization, or priority effects. Stochasticity gave way to homogenous selection for assembly of the Mycobiome in roots at week 3, in leaves at week 8, and was never prominent in rhizosphere or soil. Consequently, the strong stochasticity increased the observed fungal richness of leaves in weeks 1-7 and roots at week 2, and alleviated the effect of habitat selection for Mycobiomes between leaves and belowground compartments. In contrast to the hypothesis that release from stress favors stochasticity, release from drought (control) did not increase stochasticity of community assembly for the Mycobiome in this study. Instead, drought deterministically caused deviation of fungal community composition from control, and release from preflowering drought (rewatering) reduced root community dissimilarity between preflowering drought and control. Besides, drought respectively increased, not changed and decreased fungal richness in leaves, roots and rhizospheres, which might be attributed to the drought-induced differential changes in fitness of fungal OTUs.
Diversity and phylogenetic assessment of fungal communities in sugarcane plantation
- M. Raza
- Y. Diao
- Z. Zhang
- L. Cai
Sugarcane (Saccharum officinarum L.) is monocotyledonous cash crop, which belongs to Andropogoneae tribe in family Poaceae. It is widely grown in tropical and subtropical areas and cultivated in nearly 60 countries of the world. China is the third largest sugar cane producing country after Brazil and India. Fungi are most destructive pest of sugarcane worldwide and responsible for yield reduction and economical losses. In the recent few years, a few species have been firstly reported on sugarcane in China. However, exclusive fungal species are cause of disease development and few are still unknown with accurate morphological and molecular identification. Here, we collected 370 diseased root and leaf samples of sugarcane from 8 sites located at south of China and obtained 762 fungal strains via single spore and tissue isolation and our preliminary phylogenetic analysis based on ITS locus suggested that they belong to 141 species in 52 different genera. Among these species, 81% (135 species) belongs to Ascomycota; 0.6% (3 species) belongs to Basidiomycota and 0.5% (3 species) belongs to Zygomycota while 17% were not identified to species level. At the family level, most of isolated species belongs to Apiosporaceae, Chaetomiaceae, Didymellaceae, Hypocreaceae, Nectriaceae, Pleosporaceae and Trichocomaceae. The most common genera associated with sugarcane included Bipolaris, Chaetomium, Colletotrichum, Curvularia, Epicoccum, Fusarium, Nigrospora and Trichoderma. Further multi-locus phylogenic analysis revealed that at least 22 previously unknown species belongs to genus Chaetomium, Curvularia, Epicoccum, Nigrospora and two gen. novel in Chaetomiaceae. B. bicolor, B. setariae, C. geniculata, C. muehlenbeckiae, C. hominis and C. verruculosa, N. Lacticolonia, N. aurantiaca, N. gorlenkoana, N. camelliae-sinensis and N. pyriformis were reported first time on sugarcane. Present work significantly improved our understanding of mycobiota associated with sugarcane that dominantly grown in Guangxi (South of China) which provide scientific references for controlling sugarcane diseases.
Monograph of fungi on Pandanaceae
- S. Tibpromma
- K. Hyde
- S. Karunarathna
- P. Mortimer
Members of the plant family Pandanaceae are ecologically and economically important. Studies of micro-fungi on Pandanaceae based on morphology and phylogeny in Southeast Asia have been poorly done. Most of previous micro-fungi studies on Pandanaceae were carried out based on only morphology. The saprobes and endophytic fungi on Pandanaceae in Southeast Asia are provided here based on the current morphological and phylogenetic data. This study will be continued to produce a world monograph of fungi on Pandanaceae.
Getting to the grassroots of DSE symbiosis: case study from native and non-native perennial C4 grasses
- A. Kazarina
- K. Mandyam
- A. Nanjundaswamy
- G. Panicker
Grass roots host a suite of microbes of which Dark septate endophytes (DSE) are common non-mycorrhizal fungal endophytes. DSE are abundant in North American and European grasslands with Periconia macrospinosa, Microdochium sp., and Darksidea sp. being the most commonly isolated DSE fungi. Perennial C4 grasses namely Switchgrass (Panicum virgatum) and Giant Miscanthus (Miscanthus X giganteus) are top bioenergy feedstocks due to their abundant biomass yields. While the former is a native North American grass widely used for ecological restoration and soil conservation, the latter is a non-native sterile hybrid. Since Long-term and large-scale cultivation of these two grasses is expected by 2030 to meet U.S. bioenergy mandates, it is interesting to evaluate and compare their DSE symbioses. The objective of this study was to isolate DSE fungi from four native switchgrass varieties and a commercial variety of giant miscanthus and evaluate the outcome of their symbiosis. DSE fungi were isolated from Alamo, Bomaster, Colony and Kanlow varieties of switchgrass and Freedom Giant Miscanthus (FGM) cultivated in Alcorn State University’s Lorman Experiment Station. As expected Periconia macrospinosa was the most common DSE from both grasses. To test the outcome of the symbiosis, Miscanthus sinensis the parent of M. giganteus and switchgrass varieties were cultivated with P. macrospinosa isolated from respective hosts. A total of 20 isolates with 10 each from FGM and switchgrass were evaluated. Seedlings were grown in sand: soil mixture with and without DSE fungus for six weeks. Past studies have indicated outcome of DSE symbiosis is dependent both on host and fungal genotypes and is along the mutualism-parasitism continuum. Our preliminary data also indicate this trend. Plant biomass and N, P, K levels will be recorded as proxy for DSE symbiosis. Data from cultivated grasses like in this study and those from native grasslands seem to indicate that Periconia macrospinosa is the most commonly isolated DSE fungus from grasses and could very well serve as model DSE fungus.
Temporal dynamics of endophytic fungi in grassland systems
- G. May
- L. Kinkel
The seasonal dynamics of foliar endophytic fungal communities in the iconic prairie grass, Andropogon gerardii (big blue stem), were investigated to ask how these communities shift over time and ecological context. If the many and diverse fungi occupying leaves have similar functions as endophytes, we expect that their distribution through seasonal time and over the development of individual leaves and plants should reflect only a stochastic sampling process from the regional pool of taxa available. We sampled leaves of A. gerardii from marked plants throughout one season and over two years, in field plots that had received additonal nutrients or restricted large herbivores or both ecological treatments. We utilized both NexGen and culturing approaches to estimate fungal communities. Our findings demonstrate the importance of stochastic sampling of taxa available at one time in the environment nonetheless, fungal taxa shift through seasonal time and over the development of leaves on individual plants. In addition, the arrival of novel late-season taxa apparently caused major shifts in these communities. Functional analyses using experimental communities are underway to investiagate causes of these shifts in taxa over time and plant development.
Ectomycorrhizal (ECM) fungal community of mixed forest of endemic pines and broad leaves in BiDoup – NuiBa National Park, Vietnam
- H. Pham
- T. Dang
- Q. Dang
- D. Nguyen
- K. Nara
Two 4 hectares - plots were set up in mixed forest between endemic pines Pinus dalatensis, Pinus krempfii and broad leaves in Bi Doup Nui Ba National Park, Vietnam. The first plot was set up in the dominant of pines P. dalatensis forest and the second plot was in Fagaceae dominant forest. Two underground ECM fungal constituent along a 500 meter transect in each plot were identified by ITS regions. Each transect included 50 soil samples in 25 collected sites. In pine dominant plot, transect was through P. dalatensis vegetation. Totally, 4966 ECM root-tips belonged to 76 fungal species were recorded. In Fagaceae dominant plot, 4354 ECM root-tips belonged to 83 fungal species were recorded. In both plot, the community structure of underground ECM fungi is evenly high diversity described by diversity index (Simpson 1/D=19.62 and Shannon-Wiener H’=3.5), species evenness (Pielou J about 0.8 in each plot). Russula spp. is a dominant fungal group. In each plot, a 1.5 km length x 1 meter width transect will be set up as the perimeter of 3 smaller concentric squares of 4 has, 1 ha and 0.25ha. All observability fungal reproductive structures along transect will be recorded and collect in each 2 weeks of year 2015 (27 times). Totally, 1320 sporocarps were collected and identified belonged to more than 150 fungal species by ITS regions. The ECM fungal species were propose from reference. The comparison between the underground ECM underground fungal community and terrestrial showed that the similarity of total species and content of species. This is the first study investigates the ECM fungal community of endemic P. dalatensis, the most southern population of 5 needles pine in the world and the very narrow distribution pine P. krempfii in both underground and terrestrial. The additional transect through P. krempfii vegetation is setting up.
Diversity of temperate species of Clavulina and new insights about their ecological role in central Mexico
- E. Pérez-Pazos
- M. Villegas Ríos
- R. Garibay-Orijel
- R. Salas-Lizana
Clavulina J. Schröt. (Clavulinaceae, Basidiomycota) is a genus of macroscopic fungi, mostly with clavarioid basidiocarps. In temperate ecosystems, the most commonly recognized species are: C. amethystina, C. coralloides, C. cinerea, and C. rugosa. Although species of Clavulina are often categorized as ectomycorrhizal fungi (ECMF), growing evidence suggests that categories such as ECMF or saprotrophic fungi (SAPF) are loose and some plasticity may exist. The aim of this work was to describe the diversity of Clavulina species in two sites of a continuous Abies religiosa forest, with contrasting disturbance conditions and test their potential role as ECMF/SAPF. Species diversity was explored using basidiocarps and mycorrhizae, which were collected during the rainy seasons from 2011-2015 in La Sierra de las Cruces, in Central Mexico. Basidiocarps were described macro and microscopically, by focal and electron microscopy. DNA was extracted from mycorrhizae and basidiocarps, the nrDNA ITS (ITS) region was amplified, and the products were sequenced by Sanger platform. The sequences were edited in the Geneious program and together with those available in GenBank database, were aligned (MUSCLE) in the Aliview program. A consensus phylogenetic tree was obtained using Bayesian inference and Hydnum repandum as outgroup. To test the potential role of the sampled Clavulina species we obtained ∂13C and ∂15N isotopic values from Clavulina basidiocarps, SAPF, ECMF and fir leaves by means of mass spectrometry. Isotopic mean values were compared with one-way ANOVA. Morphological and phylogenetic analyzes were coincident and species were delimited as Clavulina reae, C. sp. nov. 1, C. sp. nov. 2, y C. sp. nov. 3. Clavulina reae and C. sp. nov. 3 are close to species that have been described in Europe, while C. sp. nov. 1 y C. sp. nov. 2 do not correspond to taxa previously described for the genus. Morphologically, the size of the basidiospores, the pigmentation of the basidiocarps and the presence of cystidia clearly delimited the four species. All Clavulina species showed ∂13C and ∂15N values intermediate between ECMF and SAPF at both sampling sites; in ∂13C Clavulina species are closer to ECMF (P<0.01), meanwhile ∂15N values, groups them together with SAPF (P<0.001). Even if ∂13C values strength the position of Clavulina in ECMF guild, ecological functions of Clavulina species at these sampling sites, points towards a role in the transference of nutrients different from nitrogen. Integrating taxonomic, systematic and ecological data is crucial to understand the whole biology of fungal species like Clavulina.
Growth of three mycorrhizal mushrooms from Sierra Tarahumara of Chihuahua, Mexico, under two pH conditions and six nutritive media
- L. Gómez Flores
- M. Quiñonez Martínez
- J. Flores Margez
- F. Jiménez Vega
- N. Martínez Ruíz
- F. Garza Ocañas
In the Chihuahua forest more than 50 species of mycorrhizal mushrooms are known, some of them with particular characteristics. Astraeus hygrometricus is an ectomycorrhizal mushroom that is found mainly in juvenile forests and is considered as a disturbance indicator species in Chihuahua. Laccaria laccata is an ectomycorrhizal mushroom associated with a large number of pine and oak species and Pisolithus tinctorius is a frequently used species in the inoculation of plants of forest importance. However, there are only few studies of its mycorrhizal efficiency of these three mushrooms with Pinus arizonica, one of the pine species with the highest economic interest in Chihuahua. Knowing the nutritional and environmental needs that these species require to obtain their mycelial production is fundamental in principle to improve the cultivation conditions and offer an alternative to mycorrhization in greenhouse conditions. In the present work, the growth of three mushrooms were evaluated in six growing media (Potato dextrose agar (PDA), Melin-Norkrans (MN), oats (AV), corn (MZ), trypticase soy (ATS) and dextrose sabouraud (ADS)), with the purpose of establishing the best growing conditions as part of the first stage of investigation related to mycorrhization with P. arizonica. Mycelial development for Astraeus hygrometricus and Pisolithus tinctorius was carried out from spores planted in PDA medium, for Laccaria laccata was realized using internal tissue until pure mycelium was obtained. Then mycelial fragments (1 cm2) were transferred into the six growing media under two pH conditions (4.8 and 5.8) with four replicates per treatment. Colonial morphology was determined considering macromorphological characteristics, growing media and pH. The highest colonial diameter for A. hygrometricus was obtained in ADS medium at pH 5.8 (8 cm) where it was observed the highest growth rate (16 mmd-1) after seven days of incubation. The highest values for P. tinctorius occurred in MN medium at pH 5.8 (8 cm) after seven days of incubation. However, AV at pH 4.8 was the medium where the highest growth rate was reached (12 mmd-1). The colonial morphology of A. hygrometricus was particularly different in each medium. Finally, for Laccaria laccata the highest values occurred in three growing media ADS, AV and MZ (8 cm) after six days of treatment with the highest growth rate of 16 mmd-1, it was also observed that there were no differences between pH.
Ectomycorrhizal effect of the mushroom Astraeus hygrometricus (From Sierra Tarahumara of Chihuaua, Mexico) on the growth of Pinus arizonica Engelman seedlings
- L. Gómez Flores
- M. Quiñonez Martínez
- J. Flores Margez
- F. Garza Ocañas
- F. Jiménez Vega
- N. Martínez Ruíz
Astraeus hygrometricus is an ectomycorrhizal mushroom easily recognized in the field for its star shape, is frequently consumed in countries such as Thailand and India, and it has been used in the inoculation of different species of pine around the world, but mainly in Asia. In Mexico, specially in Chihuahua, we can found in juvenile forest and it is considered as a disturb indicator specie. Chihuahua is one of the states with the highest forest production in Mexico, Pinus arizona is the most economically important due to wood use, however it has also been overexploited, so strategies for reforestation have been proposed. One of them is through the inoculation of seedlings destined for reforestation with ectomycorrhizal mushrooms but until now there are not studies with both species. In the present work the ectomycorrhizal capacity of Pinus arizonica with Astraeus hygrometricus was evaluated using the spore inoculation technique under three different inoculum volumes as treatments (10, 25 and 50 mL) with the porpouse of establishing the best dose of inoculation through the registration of morphometric variables in the pine seedlings. The main variables evaluated were height of the complete seedling, height and width of the foliage and percentage of mycorrhization of the root. Likewise the mycorrhiza was morphological and histologically characterized. Six months after inoculation the two volumes of spore inoculation (10 and 25 mL) gave similar results compared with the control, however the best treatment was 50 mL, the average height of the plant was 6.33 cm in contrast to the control whose value was 4.99 cm, while the average height of the foliage was 6.03 cm in contrast to the control value 4.74 cm and 5.81 cm for width of the foliage in contrast with 4.75 cm to the control. The percentage of mycorrhiza colonization was 64%, the mycorrhizas of Astraeus hygrometricus in Pinus arizonica presented a coralloid morphology and white color that was smaller at the tips, the mantle surface was granular and the Hartig net was observed.
Scleroderma meridionale ectomycorrhizae on Halimium halimifolium: expanding the Mediterranean symbiotic repertoire
- M. Leonardi
- M. Neves
- O. Comandini
- A. Rinaldi
AbstractScleroderma is a gasteroid genus in the Boletales (Basidiomycota), with a cosmopolitan distribution. Species of Scleroderma establish ectomycorrhizal (ECM) symbiosis with a range of coniferous and non-coniferous trees and shrubs, both in temperate and tropical regions, with little tendency to host specificity, a feature that might have facilitated the wide distribution of the genus. With the contribute of confocal laser scanning microscopy, we describe the morpho-anatomical features of the ectomycorrhizae formed by Scleroderma meridionale on Halimium halimifolium, a cistaceous plant belonging to a small group of woody shrubs occurring in open vegetation types in the Mediterranean region. The mycobiont and host plant identity in ECM was verified through molecular tools. Mycorrhizal system is very small, up to 1.9 mm, mostly coralloid to irregularly pinnate. The mantle surface is felty, whitish with silver patches. Differentiated rhizomorphs occur infrequently. Mantle surface is characterized by a network of branched hyphae organized in hyphal boundles. Hyphae are frequently covered by granules or warts. These characters, except for the presence of granules, are similar to those reported for the only two naturally-occurring Scleroderma ECM described so far, i.e. S. bovista on Populus and S. citrinum on Betula and Pinus. On the other side, the peculiarity of S. meridionale + Halimium ECM is the particularly small dimension of mycorrhizal system, a character shared with ECM formed by Cistaceae. At the best of our knowledge, this is the first description of an ectomycorrhiza on Halimium, a plant whose mycorrhizal biology deserves to be explored in greater detail.
Spatial patterns of ectomycorrhizal fungi in Soudanian woodlands
- B. Furneaux
- R. Houdanon
- M. Bahram
- N. Yorou
- M. Ryberg
Trees which form ectomycorrhizal (ECM ) symbiosis with fungi are important in the Soudanian zone of West Africa, including Central and Northern Benin, where they are often the dominant canopy trees in savannah woodlands. Many of the fungal partners produce edible mushrooms during the rainy season, which are harvested by the local population for culinary use. Our ongoing study investigates the processes which spatially structure the ECM fungal community in these woodlands. We exhaustively sample ECM fruitbodies in nine 50x50m plots, each divided into 25 10x10m subplots, in Central Benin during the mushroom season, from June-October. Sampling began in 2015 and will continue at least through 2018. In addition to fruitbodies, we are also characterizing the soil fungal community through DNA metabarcoding. We consider the influence of soil characteristics, size and proximity of four species of ECM host trees, canopy closure, ground cover, and microclimate on the productivity, diversity, and community composition of ECM fungi both above and below-ground. In addition, we compare the results obtained from both short (ITS2) and long (ITS2 + partial LSU) amplicons using both the IonTorrent S5 (short) and Pacific Biosciences RSII (short and long) platforms, to compare tradeoffs between cost, sequencing bias, and the ability to place reads taxonomically. Preliminary results show that the fungal fruitbody community has strong spatial structure with high correlation between years (r2=0.54), about half of which is explained by variation in soil characteristics, including total Phosphorus, pH, total Nitrogen, and clay fraction. Other environmental variables, including the presence of different host species, explain only a small amount of variance. The total biomass of fruitbodies produced is negatively correlated with several factors that may represent human impacts, including elevated Nitrogen, the incursion of grasses, and evidence of charcoal burning.
Diversity of ectomycorrhizal fungi as an indicator of the forest structure of Chihuahua, Mexico.
- M. Quiñónez Martínez
- F. Garza Ocanas
- P. Lavin Murcio
- I. Enriquez Anchondo
- C. Vital Garcia
- M. Olivas Sanchez
The forests of the state of Chihuahua have an area of 2,801.80 km2, representing 1.13% of the state territorial extension. This is dominated by pine and pine-oak forests, characterized by a wide variety of fungi species that perform different ecological functions. However, the fungi considered of greater ecological importance in the forests are those associated with the roots of the trees forming mycorrhizas. The ectomycorrhizal fungi (EMF) are a key functional group in the regulation of nutrients circulating between the soil and the plants in the most terrestrial ecosystems. Numerous studies describe the relationship between the recovery of disturbed ecosystems and the EMF community. Some research has shown a total reduction in the richness of ectomycorrhizal species and significant changes in the composition of species after felling, as well as a positive correlation between mortality of EMF, increased by fire intensity and trees mortality. However, some results suggest that the composition of the EMF community is not substantially altered by the low intensities of fires or by restoration of burning or scarceness of the organic layer that remains unchanged.
In the municipality of Bocoyna, there are highly disturbed areas due mainly to excessive wood production and overexploitation of their forests, generating the deforestation of some natural areas within the pine and oak-oak communities, causing the formation of felling areas and burning, which in turn is also one of the determining factors of the heterogeneous structure of the landscape.
The objective of this study was to analyze the changes in the richness and abundance of EMF sporomes along a gradient of forest condition: secondary succession by burning, natural area and forest regeneration.
In each site three quadrants of 2500 m2 were located based on the homogeneity of the site, at intervals of ten days during the months of July to September 2016 and 2017. The number of sporomes per species was counted for each site to obtain ecological parameters.
In addition, soil samples were taken for the analysis of texture and mineral elements by study area. The results show that the natural zone presented the greatest richness (n = 39) and abundance of EMF species, highlighting Amanita muscaria, A. rubescens and Cantharellus cibarius. In the area of forest regeneration, Laccaria laccata stands out for its abundance and finally in the zone of secondary succession only three different species were recorded, dominating Astraeus hygrometricus with a record of more than 500 sporomes. Soil analyzes showed that the area where Astraeus hygrometricus was abundant with high levels of P, K, Ca, Mn and Zinc and a pH of 4.9 considered an acid soil in this forest.
Ectomycorrhiza of pecan in Brazil
- T. Grebenc
- M. Sulzbacher
- D. Fronza
- A. Giachini
- Z. Antoniolli
- R. Jacques
Ectomycorrhiza studies on pecan (Carya illinoinensis (Wangenh) K. Koch) are becoming more common in recent years, both globally and in Brazil. In greenhouse conditions pecan can be mycorrhized easily with different truffles (Tuber spp.) species including several commercial species. On the other hand, the growth dynamic and optimal ecological conditions for pecan nuts production in plantations is not in favor of required truffle ecology. We aim to review the biology of both partners in mycorrhiza, pecan seedling/tree and selected commercial truffle species to foreseen optimal co-cultivation and management conditions. The ectomycorrhiza diversity was reviewed and analyzed in selected pecan plantations in Brazil and the growth requirements of pecan in native sites and commonly used management practices in commercial pecan plantations were reviewed. The main challenge was the bringing closer optimal soil conditions that would be in favor for commercial truffle species used in study (T. aestivum, T. borchii and T. brumale) and conditions required for normal growth, development, and fruiting of pecans. The ectomycorrhizal community on pecan in plantations in southern Brazil was relative rich, with species common for carbonate-poor sites. Besides other species a novel truffle species was discovered, presumably introduced from N. America and recently described by Grupe et al. (2018) as Tuber floridanum spec. nov. The southern regions of Brazil have the appropriate conditions for the development of truficulture, a market niche that does not yet exist in Brazil, but which can become highly profitable, and pecan can be a beneficial host as revealed by Sulzbacher et al, 2018 in recently submitted paper. We propose a time- and management-sensitive production chain: truffle – pecan seeds – pecan wood, which can be feasible and economically sustainable, if the plantation is properly managed throughout its life-cycle.
Teamwork makes the dream work: disentangling cross-taxon congruence across soil biota in Pinus nigra plantations
- D. Barbato
- C. Perini
- S. Maccherini
- G. Bacaro
- E. Tordoni
- S. Mocali
- I. De Meo
- E. Bianchetto
- S. Landi
- G. Bettini
- B. Gargani
- M. Marchi
- P. Cantiani
- L. Gardin
- E. Salerni
Soil plays a fundamental role in many ecological processes, interacting with above-ground biodiversity in a complex network of ecosystem functions. Recently, there has been an increasing interest in the use of correlates for biodiversity assessment with evidence showing that the use of surrogate taxa in conservation planning is substantially more effective than that of proxies based on environmental data alone. A large body of literature deals with co-variation of species richness and composition in grassland soils whilst only few studies were developed on forests and even less in forest plantations. In this context, a multidisciplinary EU-Life project (SelPiBioLife, LIFE13 BIO/IT/000282) was established in 2014 in two mountain areas of the Apennines (Italy) aiming at evaluating the application of an innovative forest management technique along with its effects on soil biodiversity in Pinus nigra plantations. Based on data collected before the silvicultural treatment, main aim of our study was to test the robustness of cross-taxon congruence across vascular plants, fungi (macrofungi, microfungi, ectomycorrhizae), carabids, microarthropods, nematodes and bacteria, also exploring how abiotic (soil and spatial-topograhic variables) and biotic (dendrometric variables) predictors drive the community concordances among taxa. Abundance data were obtained at the available taxonomic resolution for each biological group. Correlations between taxa were performed through Mantel and partial Mantel tests, while variation partitioning analysis was used to assess the total variance of each dependent taxon in the pure effect of another taxon, pure effect of soil, pure effect of spatial-topographic factors, pure effects of dendrometric variables, partial shared variation and total shared effect among all sets of predictors. In general, the distribution pattern of almost all the groups analysed showed highly supported inter-group congruence, while nematodes were not significantly correlated with other taxon. All biological groups also detained close relationship with the overall dataset of environmental-spatial variables. Considering the variation partitioning results, the variance attributed solely to pure effect of biotic or abiotic predictors was significant only in some cases (e.g. bacteria); remarkably, in all dependent taxa, total shared and partial shared effect of all sets of predictors always explained the highest portion of total variation. In conclusion, we can affirm that it is not a unique factor but rather the mutualistic relationship of all variables, both biotic and abiotic, to regulate the above-below ground subsystems in Pinus nigra plantations.
Wood decomposing fungal diversity explains enzyme patterns and element bioavailability in a long-term field experiment of the German Biodiversity Exploratories
- H. Kellner
- S. Leonhardt
- E. Stengel
- J. Moll
- B. Hoppe
- C. Bässler
- F. Buscot
- M. Hofrichter
Deadwood is an important structural component in forest ecosystems and its degradation is mainly controlled through microorganisms (fungi and bacteria) and their secreted digestion enzymes, as well as by xylobiontic arthropods. While the biochemical processes of wood decay are fairly well understood, in situ data about the factors that explain the heterogeneous distribution patterns of wood-decomposing fungi, their corresponding lignocellulolytic enzyme activities and the resulting changes of wood physico-chemical parameters and element bioavailability are rather scarce. We investigated the deadwood decomposition in a long-term experiment, which has been started within the German Biodiversity Exploratories in 2009. Here, we especially address the following hypotheses: i) the structure of fungal communities corresponds to the decomposing tree species and significantly differs between, sap- and heartwood, ii) specific ecological groups of fungi (e.g. white-rot Basidiomycota) explain the presence of particular enzymes and their distinct spatial patterns; and iii) fungal depolymerization processes increase the bioavailability of essential elements in decomposing wood.
Samples were taken from deadwood logs of 13 different tree species (n=3) exposed to decomposition for six years in beech forests in Central Germany. Fungal communities were amplicon-sequenced using Illumina MiSeq. Fifteen extracellular enzyme activities (hydrolases and oxidoreductases) as well as wood parameters (e.g. water-soluble lignin fragments) and element contents were measured.
i) The fungal community structure and species richness significantly differed between the 13 decomposing tree species (specifically between deciduous and coniferous tree species), which can partially be explained by the different physico-chemical traits of the logs. For example, the lignin content, pH, organic extractives and element contents were found to considerably vary between deciduous and coniferous wood and thus correlated to distinct fungal sub-communities. However, we found no differences in fungal species richness and community structure between sap- and heartwood, six years after log deployment. In this experiment, fungal succession has seemingly reached the optimal phase of decomposition for several tree species. It means that with few exceptions (e.g. Ascocoryne sarcoides), the fungi has grown and colonized the logs from the outer to the inner side.
ii) The presence of particular fungal families (e.g. Meruliaceae or Coniochaetaceae) was found to correlate with the secretion of characteristic lignin-modifying and/or (hemi)cellulolytic enzymes. Specifically, the abundance of white-rot basidiomycetes was positively correlated with the manganese peroxidase activity, which is the crucial enzyme of incipient lignin degradation.
iii) Fungal depolymerisation activity evaluated by qualitative and quantitative analysis of small to medium-sized, water-soluble lignin fragments (0.5-5 kDa) significantly correlated with element bioavailability, which may be an important result regarding flux processes in forest ecosystems. Causal agent of this relation are ligninolytic white-rot basidiomycetes.
Altogether we gathered an in-depth dataset of the microbial diversity, activity and resulting changes in decaying wood, which enables us to mechanistically understand the complex processes in the course of the biodegradation of woody biomass. In addition we currently analyse the co-occurring bacterial community in the logs.
Wood decomposition and fungal communities: the interactive influences of soil and substrate nutrients
- J. Jones
- A. Ferrer
- K. Heath
- P. Zalamea
- J. Dalling
Wood decomposition is fundamental to carbon and nutrient cycling. Fungi play a critical role in wood decay because they produce lignocellulolytic enzymes that degrade wood polymers. The identity of these decomposers may have a profound effect on wood decomposition rate, as the ability to degrade lignin efficiently is restricted to a subset of fungal lineages. However, decomposition rate may also be influenced by nutrient availability. Increased nutrient availability can speed decay either by allowing increased production of nutrient-rich wood decay enzymes, or by shifting community composition towards taxa with intrinsically higher enzyme production and activity. In turn, enhanced enzymatic decay may also depend on nutrient source, as fungi can use nutrients from both wood and soil.
This project explores: (1) Whether nutrients influence decay rate directly, by limiting enzyme production, or indirectly, by affecting fungal community composition; (2) The relative importance of wood and soil nutrient availability as determinants of decay rate. We used a novel experimental approach where saplings of four tree species were grown in the greenhouse under different soil nutrient treatments to induce 4-fold variation in wood phosphorus, nitrogen, and potassium concentrations. Wood samples from these saplings were then placed in different soil nutrient treatments established as part of a long-term soil fertilization experiment in a seasonally moist tropical forest in Panama. We measured the effects of wood nutrients and soil nutrients on wood decay rate and included fungal community analyses and assays of enzymatic activity.
We found that wood nutrient treatment explained more variation in decomposition rate than soil nutrient treatment. Cellulose, hemicellulose, and chitin related enzyme activity responded to both wood nutrients and soil nutrients. However, acid phosphatase activity only responded to wood nutrient treatment. Completion of fungal environmental amplicon sequencing will allow us to determine whether increases in enzyme activity in response to increased nutrient availability represents up-regulation of enzyme production by similar fungal communities, or by shifts in fungal community composition in response to substrate and soil conditions. A more complete understanding of how fungal communities and nutrient limitation interact to influence wood decay, has implications for predicting how global change phenomena, including nitrogen deposition, will impact one of the largest compartments of global carbon cycle.
Life after death: Dynamics of beech wood-associated communities over a gradient of life and decay stages
- L. Jackson Iii
- J. Heilmann-Clausen
Wood is an important renewable resource that stores massive amounts of carbon globally, and supports complex food webs comprised of diverse, highly specialized organisms. As such, it is important to understand the processes that control wood decomposition to more accurately predict global carbon cycles and protect biodiversity under increasing pressure due to climate change effects, or habitat loss or fragmentation. Wood-inhabiting endophytes sometimes act as latent saprotrophs and recent experiments suggest that priority effects imposed at the endophyte-saprotroph transition may impact community assembly dynamics that affect species richness, decay rate, decay type, litter quality and carbon release in natural settings. Results will be presented from a study in which amplicon sequencing and physicochemical data are being used to assess community dynamics across a gradient of living (5 growth stages) and dead (6 decay stages) European beech (Fagus sylvatica) wood from a nature reserve in Denmark. We aim to reconstruct wood-associated community assembly histories using network analysis methods to identify key species or abiotic factors that determine decay progression. We expect that species or factors with high betweenness centrality scores will link modules that are associated with distinct decay types. Further, these modules are likely to be characterized by species or factors exhibiting high scores for closeness centrality.
Fungal carbon utilization in a temperate forests
- T. Martinović
- T. Mašínová
- M. Štursová
- P. Baldrian
Being one of the major terrestrial carbon pools, forests are extremely important for carbon sequestration and greenhouse gas fluxes. The forest soil processes carried out by microorganisms are thus critical for global carbon cycle and consequently global climate. Boreal and temperate forests provide niches for a diverse community of fungi, both saprotrophic and mycorrhizal. Fungi produce a variety of enzymes responsible for decomposition of even the most recalcitrant plant biopolymers. Furthermore, they play a crucial role in carbon sequestration and mediate allocation of plant-derived carbon into soils. This experiment aimed to track the incorporation of carbon derived from different sources in forest soil into the fungal biomass using stable isotope probing. Six substrates labeled with 13C isotope were added to microcosms containing homogenized soil from a temperate forest in the Czech Republic. The substrates varied in complexity – from simple sugars and organic acids present in root exudates (glucose, citric acid) to more complex carbon sources (cellulose, hemicellulose) and substrates resembling total plant biomass (maize leaf) and fungal biomass (chitin). Microcosms were incubated for 1 and 3 weeks, together with associated controls. Substrate respiration rates were determined by the analysis of the CO2 released in microcosms. Fungal taxa incorporating the 13C into their biomass were identified after DNA extraction, 13C-DNA separation and amplicon sequencing of ITS2 on Illumina MiSeq. The highest respiration rate (mmol/g 13CO2) after 1 week of incubation was observed in microcosms supplemented with citric acid, followed by glucose, hemicellulose, maize leaf, cellulose and finally chitin. After incubation of 3 weeks, a considerable increase in chitin respiration rate was observed, while the increase in respiration of other substrates occurred at a more similar rate. While all substrates were highly incorporated by the Ascomycota, Zygomycota were detected in chitin microcosms in higher relative abundances compared to the other substrates. In the case of chitin, the most dominant 13C-accummulating genus was Mortierella, while in other substrates Geomyces seemed to be the most dominant genus. Other highly enriched genera included Pseudocosmospora (chitin), Trichosporon (citric acid), Pencillium (glucose, hemicellulose, citric acid) and Aureobasidium (maize leaf). The results of this study show that multiple fungal taxa are capable of utilization of all carbon sources regardless of their complexity. However, the results also indicate a difference in fungal community responsible for chitin decomposition, confirming the specificity and importance of fungal biomass as carbon source in temperate forests.
Assessment of dead wood fungi biodiversity in urban parks and natural reserves across New Jersey
- S. Blue
- M. Shumskaya
- A. Vindas-Cruz
Ecosystems with high species richness such as forests rely on biodiversity of decomposers to boost availability of forest elements. Dead wood is an important component of any forest. It is a decomposition site, it protects against erosion, improves water retention in the ecosystem, and creates ecological niches. Dead wood-inhabiting fungi are saproxylic taxa that decompose cellulose and/or lignin.
Here, we assess biodiversity of deadwood-inhabiting fungi in areas with varying degrees of human modification (removal of foliage, snags, logs). We hypothesize that removal of dead wood from urban parks diminishes the diversity of dead wood fungi which may affect the stability of ecosystems. Our goal is to compose a comprehensive database of dead wood inhabiting fungi and compare areas with different forest management practices to assess human impact.
We present preliminary results from assessment of different locations spanning across northern to southern NJ. Fungal fruit bodies were collected for two hours at each sampling site and photos and GPS coordinates were recorded and shared via iNaturalist.org. Online database is available for the world community.
Fungi identification was performed using both morphology and DNA sequence analysis. The nuclear ribosomal Internal Transcribed Spacer (ITS1, ITS2) region was used as a DNA barcoding marker, allowing identification of fungal species via DNA sequencing and bioinformatics methods.
Plant decapitation alters decomposition processes and initiates complex restructuring of fungal communities in tree roots
- P. Kohout
- P. Baldrian
- J. Albrechtova
Although root biomass represents a significant pool of organic matter in terrestrial ecosystems, information on the factors influencing root decomposition is scarce, in contrast to the vast body of data on aboveground litter decomposition. Here, we assessed changes in fungal community structure, lignin and cellulose concentrations, fungal biomass and decomposition activity in the roots of a Picea abies stand for a two-year period following decapitation of Picea abies seedlings compared to data from non-decapitated seedlings. We found that the termination of photosynthate flow is associated with profound changes in decomposition processes, fungal biomass as well as fungal community composition. We found no support for the involvement of ectomycorrhizal fungi in the decomposition of roots, but we found some evidence that root endophytic fungi may have an important role in the early stages of this process.
Where mycorrhizal fungi meet leaf-endophytes: Fungal community dynamics during early decomposition of translocated leaf litter
- D. Persoh
Litter decomposition is a key process in nutrient cycling and therefore crucial for ecosystem functioning. The major decomposers in forest ecosystems are considered to be saprotrophic soil fungi. However, recent evidence suggests that previously endophytic as well as mycorrhizal fungi are involved in the degradation of organic matter. To assess the changes in fungal community composition and the dynamics of different functional guilds (i.e. endophytic, saprotrophic and mycorrhizal fungi) during early decomposition, we designed a litter translocation experiment. Beech trees from a tree nursery were planted at different altitudes (500 m and 1000 m asl) at the ‘Untersberg’ mountain to exclude effects by genetic pre-adaptation of the hosts. After one growing season the trees accumulated site specific endophytic fungal communities. The leaf-litter of each of five trees was collected separately in the following autumn, portioned and incubated at each altitude. Litterbags were collected regularly for 15 month. Content and stable isotopes of carbon and nitrogen were measured and nucleic acids extracted. The present and active fungal communities were assessed by Next Generation Sequencing of the ITS rRNA gene (DNA) and precursor ITS rRNA (RNA), respectively. Isotopic signatures indicated that decomposition was initially faster at the lower, but after summer faster at the higher altitude. Composition of fungal communities (DNA) started to differ between incubation sites in the following May, while the community activity (RNA) already differed significantly in March, i.e. when the sites were still covered with snow. Origin of the samples significantly shaped the communities throughout the first year of incubation. Only after the second winter, origin of the decaying leaves was no longer reflected by the fungal decomposer communities. Proportion of the previously endophytic fungi decreased only slowly in the first year, while ectomycorrhizal fungi occurred at noteworthy contributions from July on and increased in abundance in most samples until October. Community dynamics will be discussed in dependence from incubation altitude, with a focus on the interaction of saprotrophic soil fungi, previously endophytic fungi and ectomycorrhizal fungi.
Laetiporus lobatus (Basidiomycota, Polyporales), a new fungal species from Costa Rica
- J. Kout
- J. Vlasák
- D. Linder
The genus Laetiporus with one species L. sulphureus (Bull.:Fr.) Murrill was erected by Murrill for a common and very distinct wood-rotting fungus characterized by poroid hymenophore, dimidiate to flabellate pilei of bright orange to yellow color, soft and fleshy context, dimitic hyphal system with simple septate generative hyphae and characteristic, broad and interlocking binding hyphae, and causing a brown rot. For a long time the only member of the genus has been considered a cosmopolitan species .However, incompatibility studies revealed 5–6 incompatibility groups/species only in the USA and this result was confirmed by molecular studies. New American species have been described, all often very similar to L. sulphureus, but differing by sequence, ecology and geographical distribution and showing also slight differences in basidiospores size and shape. We have recently collected and studied specimen from Costa Rica showing ITS sequence and morphological traits different from other Laetiporus sp., we describe it as new species, Laetiporus lobatus. Laetiporus lobatus is similar to L. sulphureus but with much smaller pores, 7–8 per mm, small spores 3.9–4.4 × 2.8–3.2 μm, strikingly lobed pileus margin and unique ribosomal ITS sequence. Laetiporus taxonomic problems are briefly discussed and a key to described species is provided.
New poroid species of Hymenochaetaceae (Hymenochaetales, Basidiomycota) from southern Chile
- M. Rajchenberg
- M. Pildain
- J. Becerra
- R. Reinoso Cendoya
- D. Cajas Madriaga
Polyporoid fungi from Chile are poorly known. Most of the species present in the Patagonian Andes forests of Argentina are also found there, but southern Chile is distinguished by particular phytogeographic formations that host a high diversity of tree plants. Field work in the Valdivian rainforest and the temperate durifoliate forests revealed specimens of four taxa that were studied morphologically, through culture characterization and phylogenetic evidence (sequences comparisons of molecular markers) and that are here shown to represent new taxa in the genera Fomitiporia (ITS, LSU, EF, rpb2), Phylloporia (ITS, LSU) and Fomitiporella (ITS, LSU) (Hymenochaetaceae, Hymenochaetales, Basidiomycota). We describe, characterize and show the phylogenetic position of the following proposed new taxa: Fomitiporia “chilensis” (on Cryptocarya alba and Peumus boldus), Phylloporia “boldus” (on living Peumus boldus), Fomitiporella “muriforme” (on undetermined fallen trunks) and Fomitiporella “podocarpus” (on living Podocarpus nubigena). Fomitiporia “chilensis” is phylogenetically associated to the resupinate South American lineage of the Fomitiporia punctata species complex and is morphologically similar to Fomitiporia dryophila. Phylloporia “boldus” is distinguished by relatively large, dull chestnut basidiospores that lack the typical yellowish color of the wall in other species of the genus; it is related to Phylloporia dependens, a taxon described from tropical China that is phylogenetically weakly related to the core species group in Phylloporia. Fomitiporella “muriforme” and Fomitiporella “podocarpus” are represented by single specimens but are otherwise distinguished phylogenetically. We also report the wide presence and distribution of Fomitiporella americana, a taxon recently described from SE USA; this taxon is morphologically similar to Fomitiporella inermis, it decays trunks of many hosts in southern Chile and Argentina and is responsible of the white heart rot present in standing Austrocedrus chilensis. The combined ITS and 28S concatenated analysis showed that the genera Inocutis, Phellinotus and Arambarria are included or fall within the gross phylogeny of Fomitiporella. The analyses confirmed that these genera are closely related and form a strong monophyletic group (BA 100, ML 100) and that Fomitiporella taxa appear across multiple branches of the phylogeny. Funding: Fondecyt 1151028 and Cooperación bilateral MinCyT (Argentina)-CONICyT (Chile) CH13/06.
Diversity and taxonomy of Ganoderma species in South Africa, inferred from morphology and a multi-locus phylogeny
- J. Tchotet Tchoumi
- M. Coetzee
- M. Rajchenberg
- J. Roux
Ganoderma is a cosmopolitan genus of Polypores that encompasses species important for forestry, medicine and cultural traditions. Despite the importance of this genus, knowledge pertaining to the species diversity of Ganoderma in South Africa is limited. This study aimed at elucidating the identity and phylogenetic placements of a large collection of Ganoderma samples (including basidiomes and wood samples) obtained during a wood-rot fungus survey in the Garden Route National Park (GRNP) of South Africa and during earlier fieldwork at other localities. Identification was facilitated by phylogenetic analyses of DNA sequences obtained from the ITS regions, a region of the β-tubulin and Translation Elongation Factor 1-alpha (TEF) genes, respectively, as well as morphological characters. Results from these analyses revealed that isolates from the collections belong to eight Ganoderma species. Of these, G. applanatum, G. austroafricanum, G. destructans and G. enigmaticum have previously been reported from South Africa, while G. cupreum and G. resinaceum are new records for the country. The remaining two species are novel taxa belonging to subgen. Elfvingia and described as G. acacicola sp. nov. and G. knysnamense sp. nov. Ganoderma acacicola occurs on native and non-native hosts, including Acacia cyclops, Celtis africana, Prunus africana and an unknown palm species in four provinces of the country. The fungus is characterised by a perennial, triquetrous and broadly attached basidiome, a sulcate up to zonate yellowish brown to brown pilear surface, and ovoid to ellipsoid basidiospores. Ganoderma knysnamense was collected only in the GRNP where it was also the most abundant fungus among the species identified. It is distinguished by its applanate to ungulate, sometimes convex, and dimidiate to broadly attached basidiome, its chocolate-brown pilear surface covered with a hard woody-like crust and ellipsoid, broadly ellipsoid to ovoid basidiospores. The discovery of two new Ganoderma species as well as the two newly-recorded species raises the total known Ganoderma species in South Africa to 15. The continual discovery of new species, as is shown in this study and other recent studies, suggest that many more Ganoderma species are likely to be discovered in South Africa and indicates that further research is warranted on this important genus in the country and in Africa.
Ectomycorrhizal associations with Salix humboldtiana in Southern South America: An ancient cross-continental exchange.
- A. Mujic
- E. Nouhra
- F. Kuhar
- C. Truong
- M. Smith
Ectomycorrhizal (ECM) forests in southern South America (SSA) are dominated by Nothofagaceae (southern beech) trees. Based upon the fossil record, Nothofagaceae trees and their ECM fungi have been present in SSA for 60 – 100 million years with little evidence for co-occurring ECM hosts during this time. A second native ECM host tree, Salix humboldtiana (Humboldt’s willow), colonized SSA from the north between 3 – 15 MYA. This event followed the formation of the Isthmus of Panama during the Great American Biotic Interchange. While Nothofagaceae species and S. humboldtiana overlap in latitudinal range, these trees represent different lineages, occupy different niches, and seldom inhabit the same forests. Both Salix and Nothofagaceae associate with a diverse assemblage of ECM fungi but the biogeographic origins of these communities is expected to be different (Salix from the Northern Hemisphere and Nothofagaceae from the Southern Hemisphere). However, it is possible the two hosts have exchanged ECM fungi since the colonization of SSA by Salix. In this study we characterize ECM communities associated with S. humboldtiana in Argentina and compare these communities with Nothofagaceae to investigate the potential for ECM host jumping between these taxa. We sampled rhizosphere soil, ECM root tips, and ECM fungal sporocarps from stands of S. humboldtiana at 18 sites throughout its range in Argentina from Parque Nacional Calilegua (Northern Argentina) to the Chubut River (Central Patagonia). Since exotic Eurasian Salix spp. were present along the same watercourses throughout much of our sampling range, we also sampled rhizosphere soil and ECM root tips from exotic Salix at 5 sites to investigate the potential for shared ECM communities between exotic Salix and S. humboldtiana . Fungal communities from soil DNA and ECM root tips were identified using Illumina Miseq meta-barcoding of the ITS1 region. Operational taxonomic units (OTUs) were sorted by putative ecological niche (saprobic, pathogenic, mycorrhizal) using a combination of FUNGUILD and Blast searches of NCBI GenBank and our extensive in-house database. Results of our comparative metagenomic analyses indicate that ECM fungal communities of S. humboldtiana are composed of fungi primarily from northern hemisphere ECM lineages and lack representatives from endemic southern hemisphere ECM fungal lineages (eg. Descolea, Phaeohelotium, Austropaxillus). There are several species of exotic ECM Basidiomycota that were likely introduced into S. humboldtiana ECM communities from exotic European Salix. Some lineages of ECM host generalist fungi may also have moved between S. humboldtiana stands and Nothofagaceae forests. Exemplar lineages from our dataset include species of ECM host generalists Hebeloma (Agaricales), which are likely to have moved into Nothofagaceae forests from S. humboldtiana stands or exotic Northern Hemisphere hosts, and Tomentella (Thelephorales) which may have undergone bidirectional host shifts between Nothofagaceae and S. humboldtiana.
Evolutionary relationships of Gloeoporus (Irpicaceae, Basidiomycota) with emphasis on the G. dichrous complex
- V. Motato-Vásquez
- A. De Mello Gugliotta
- O. Miettinen
Gloeoporus is a cosmopolitan genus with species able to decay a wide range of substrates. The genus include species characterized mainly by the continuous gelatinous hymenium along tube surface and dissepiments, shallow pores, whitish context contrasting with a brightly colored hymenophore, monomitic hyphal system and allantoid basidiospores. Currently, 13 species of Gloeoporus are accepted by most mycologist but only four of these [viz., G. citrinoalbus, G. dichrous G. hainanensis and G. pannocinctus] have been included in phylogenetic analyses. Despite a long history of formal recognition and research, several aspects of Gloeoporus phylogeny have yet to be uncovered. This is because the phylogenetic position of Polyporus conchoides (= G. thelephoroides, type species of the genus) has long been uncertain, and the diversity of Gloeoporus is in large part composed of geographically widespread species. The objective of our study was to assess species limits and infer species-level phylogenetic relationships within Gloeoporus, with special emphasis in G. dichrous complex, through a combination of detailed morphological studies of type/original collections and molecular phylogenetic analyses of specimens collected in North America, South America, Europe and Southeast Asia. Different optimality criteria were used to analyze combined molecular matrices of two genes (ITS and nLSU). The dataset includes 74 ITS and nLSU original sequences in addition to 140 ITS and nLSU sequences retrieved from GenBank. Our preliminary results suggest the monophyly of Gloeoporus within the Irpicaceae family with high support. Sequences of specimens morphologically similar with or identified in GenBank as G. dichrous were recovered in three different sub-clades, as follows: 1) a sub-clade of widely distributed specimens, including European specimens collected next to the type locality (G. dichrous s.s.), together with North and South American specimens; 2) a sub-clade of Asian specimens representing three different lineages independent from G. dichrous s.s.; and 3) a well-supported clade of Gloeoporus specimens from Brazil. Gloeoporus thelephoroides was included for the firsts time in a phylogenetic analysis. Our analyses have shown that the species as currently defined represents a complex of species, with at least four different lineages independent of G. thelephoroides s.s. Additional collections, meticulous study of morphology and more molecular evidence will allow clarification of the evolutionary history of these species. In light of the large number of synonyms and names available in Gloeoporus we will discuss how to best apply the existing names to the clades recovered in the molecular phylogenies.
The genus Trichaptum (Agaricomycetes, Basidiomycota) in São Paulo state, Brazil
- L. Lima
- R. Pires
- V. Motato-Vásquez
- M. Westphalen
- A. Gugliotta
The genus Trichaptum was described by Murril in 1904 based on specimens collected in India, Central and South America. In its original description, it was characterized by annual basidiomes, sessile, dimidiate, with brownish color, short tubes that become labyrinthiform with age, with darkest coloration than the context and smooth spores. Nowadays, the diagnoses of the genus expanded, encompassing widely distributed species, characterized by resupinate and effused-reflexed basidiomes, with poroid to irpicoid or lamellate hymenophore, di- to trimitic hyphal system, generative hyphae with clamps and hymenial cystidia. Sixty-six species names are associated with Trichaptum in online mycological databases. Nine accepted species were recorded in Brazil and six in São Paulo state, i.e. T. abietinum, T. biforme, T. byssogenum, T. fumosoavellaneum, T. perrottetii and T. sector. This study aims to understand the species distribution on the Southeast of the Brazilian Atlantic forest and its relationships through morphological and phylogenetic studies. Macro and microscopic analyses based on specimens collected in different Conservation Units of the state and on specimens deposited on SP herbarium are in progress. The study of type specimens is also being carried out in order to compare and infer important characters to delimit the species and genus. An identification key of the studied species will be presented. For phylogenetic studies, DNA extraction and amplification of ribosomal regions Internal transcribed spacer (ITS1, 5.8S and ITS2) and Large subunit (nLSU) of fresh or recently collected materials are being made. Phylogenetic analyses were constructed using Maximum Likelihood and Bayesian Inference methods. Initial analysis indicates that Trichaptum Murril as currently defined is not monophyletic. The species T. byssogenum was described from Java but the type specimen was never sequenced and as far as is known there are no recent collections in the type locality. Sequences of specimens from Brazil and sequences retrieved from GenBank morphologically identified as T. byssogenum nest in a separate clade independent from Trichaptum s.s., but related to Hyphodontia species. This new clade represents a possible new genus including at least two different lineages, one of them formed by Brazilian specimens and other from New Zealand and Sri Lanka specimens. Other specimen recently collected and examined indicates morphological characteristics intermediary between T. biforme and T. sector. Phylogenetic results indicate that it could be a new species, but more comparisons as well as type studies are required to elucidate its proper taxonomic placement. In addition, the morphological studies of the herbarium materials showed that T. fumosoavellaneum does not occur on São Paulo state and that T. sector has a pseudo-trimitic hyphal system, without true binding hyphae, which differs from descriptions by other authors. The sequences produced in this study are important new additions to the knowledge of the genetic diversity of specimens in the Brazilian Atlantic forest. This project is an important addition to the knowledge of the genus Trichpatum in the Neotropics, allowing a better understanding of its taxonomic placement as well as re-appraisal of morphological characters for species differentiation.
Hydnophanerochaete and Odontoefibula, two new genera of phanerochaetoid fungi (Polyporales, Basidiomycota) from East Asia
- C. Chen
- S. Wu
- C. Chen
The broad generic concept of Phanerochaete is one of the largest genera in corticoid fungi, decomposing various woody substrata and causing a white rot. Two new genera of Phanerochaete s.l. are presented, namely Hydnophanerochaete and Odontoefibula. The type species of Hydnophanerochaete is Phanerochaete odontoidea. Odontoefibula is established based on a new species, O. orientalis (generic type), and O. deflectens (≡ Grandinia deflectens). Both genera are characterized by having an effused basidiocarp with odontioid hymenial surface, simple-septate generative hyphae, cystidia lacking, clavate basidia, ellipsoid basidiospores which are smooth, thin-walled, inamyloid, non-dextrinoid, and acyanophilous. Hydnophanerochaete is diagnostic by its compact texture of subiculum with thick-walled subicular hyphae, and presence of quasi-binding hyphae. Odontoefibula is featured by its basidiocarp turning dark brown in KOH, and dense texture of subiculum with thin- to slightly thick-walled subicular hyphae. Morphological study and multigene phylogenetic analyses based on sequences inferred from two combined datasets (nrITS+nrLSU+rpb1 and nrITS+nrLSU) respectively, indicate that Hydnophanerochaete and Odontoefibula are respectively placed in Meruliaceae and Donkia clade of Phanerochaetaceae. Phanerochaete subodontoidea is a synonym of P. odontoidea, according to morphological and molecular evidence.
How many Steccherinum (Polyporales) species are there in the Neotropics?
- M. Westphalen
- A. Gugliotta
The genus Steccherinum, typified by S. ochraceum, encompasses species with hydnoid and poroid hymenophore mainly characterized by the presence of a dimitic hyphal system, large and usually heavily encrusted skeletocystidia and small globose to cylindrical basidiospores. Even though the genus is widely accepted, and about 10 species have been recorded in the Neotropics, many of these are identified with names originally given to temperate species or representing synonyms. Furthermore, there is no study specifically focused on this group in the Neotropical region combining morphological and molecular evidence. In order to contribute to this picture, twelve hydnoid specimens of Stecchetinum collected in southern Brazil throughout 2017 were studied. DNA extraction was performed using CTAB 2% lysis buffer. The Internal Transcribed Spacer (ITS1, 5.8S and ITS2) and the Large Subunit (LSU) of the nuclear ribosomal RNA (rRNA) regions were amplified and used to infer phylogenetic analysis. The PCR products obtained were sent to Macrogen (Korea) for sequencing. A dataset including the sequences obtained, as well as others available on Genbank database, was prepared. Phylogenetic trees were constructed using Bayesian Inference and Maximum Likelihood methods. The macro and micromorphological analyses revealed the presence of three morphogroups, none of them fitting the concept of species previously registered from the region. However, the phylogenetic analysis showed that at least five different lineages in Steccherinum can be recovered with high support, evidencing a greater diversity and the possible occurrence of cryptic species. One of the morphogroups identified encompasses three similar species, morphologically related to S. ochraceum, which are very difficult to discern form one another. While one of them can be distinguished by the formation of small pilei, the other two are completely resupinate. Microscopically, all three species present very small basidiospores (2.6–3.0(3.5) × 1.7–2.5 µm), which can also differentiate them from other species in the genus. The other two species recovered present larger basidiospores (3.5–4.5 × 3–3.8 µm), one of them having strictly resupinate pinkish basidiomes while the other is characterized by effused-reflexed basidiomes with brown pilear surface and whitish hymenophore. The high diversity found shows that many Neotropical Steccherinum may have been overlooked, suggesting the possibility of new species in the genus. Further studies, including the addition of more specimens to the molecular analyses and a more detailed morphological comparison of recent collections with type specimens, are being carried out in order to try to elucidate the actual diversity of the genus in the Neotropics. The data obtained shows the importance of critical analyses combining all possible empirical evidence before implementing new taxonomic decisions. In an era of fast changes for the systematics, it is always important to supplement molecular data with a detailed morphological study in order to find characteristics of taxonomic value and keep on reviewing and updating the knowledge of Fungi in the world.
Comparative Genomics of the Genus Amanita
- J. Herr
The genus Amanita has long been an iconic Basidiomycete lineage – to the point that when most people think of a mushroom the envision the red and white cap of Amanita muscaria. There are dazzling arrays of biochemical pathways that are induced across the genus – with clades with edible, psychoactive, and deadly poisonous members. Additionally, previous research has shown the loss of genes responsible for a saprophytic lifestyle and the transition to ectomycorrhizal association with woody plant hosts. In order to understand both life history and phylogenomics of the group, we have initiated a study to compare the genus using genome sequence data. Phylogenomic analysis reveal the relative position of each species across the genus using a suite of genes culled from annotated coding regions. Gene annotation methods were used to identify candidate genes for interaction with plants (effectors and other small secreted protein production), novel biochemical pathways, and secondary metabolite production. Pathway analysis has identified numerous biochemical pathways that have emerged across the Amanita lineage.
Phylogenetic position and taxonomy of Kusaghiporia usambarae gen. et sp. nov. (Polyporales)
- J. Hussein
- J. Hussein
- D. Tibuhwa
- S. Tibell
Polyporales form a large group of Basidiomycota, containing more than 1,800 species in over 216 genera and 37 families. Seven clades are recognized in Polyporales: the ‘antrodia’, ‘core polyporoid’, ‘residual polyporoid’, ‘phlebioid’, ‘tyromyces’, ‘gelatoporia’ and ‘fragiliporia’ clades. Currently, the ‘antrodia clade’ contains more than 26 genera, which are of economical importance as food as well as a source of pharmaceutical and biotechnological products. However, some species are plant pathogens. ‘Kusaghizi’ is a local name of a large polyporoid mushroom from the West Usambara Mountains in Tanzania. The mushroom produces large dark brown fruiting bodies up to 60 cm wide, which at maturity may weigh more than 10 kg. It has a high rate of mycelial growth and regeneration, and was found growing on both dry and green leaves of shrubs and attached to the bases of living trees. It was also observed to degrade snakes and insects coming into contact with it. This mushroom has a long tradition of being used as food and medicine by local communities although no scientific description of this has been carried out. This study describes the species and infers its phylogenetic position. Morphologically, the mushroom produces dark brown basidiomata with globose to subglobose basiodiospores. Phylogenetic analyses based on individual and concatenated data sets of nrLSU, nrSSU, RPB2 and TEF1 genes grouped the mushroom together with Laetiporus and Wolfiporia, with strong support to form a monophyletic group in the ‘antrodia clade’. A new genus and species is proposed: Kusaghiporia usambarae gen. et sp. nov. to accommodate this species.
Macroevolutionary analyses of fruiting body forms and nutritional modes in Agaricomycetes based on an 8500-species phylogeny
- M. Sánchez-García
- M. Ryberg
- F. Kalsoom
- D. Hibbett
The Agaricomycetes contains approximately 35,000 described species and a wide variety of fruiting body morphologies such as resupinate, coralloid/clavarioid, pileate-stipitate, sessile and gasteroid forms, as well as diverse nutritional modes, such as saprotrophy, parasitism and mutualistic symbiosis. Previous studies have hypothesized that the resupinate fruiting body form is plesiomorphic, there is a directional trend that favors pileate-stipitate forms, the gasteroid form is irreversible and pileate-sessile and clavarioid forms are labile. The development of new methodological tools and the increasing amount of public DNA sequences provide an opportunity to test these and other hypotheses about the evolution and diversification dynamics in the Agaricomycetes. The aims of this research are to identify broad evolutionary patterns within this group, and to study the effects of morphological transitions and nutritional strategies on diversification rates. We reconstructed a six-gene phylogeny that contains 8,500 species of Agaricomycetes and estimated trait-independent diversification rates using BAMM, as well as trait-dependent diversification rates using BiSSE and MuSSE. We tested several coding regimes that include transitions from non-gasteroid to gasteroid forms, and multi-state transitions such as resupinate, clavarioid, pileate-stipitate, sessile and gasteroid. Our results suggest that there have been 30-40 major shifts in diversification rates, most of them in the Agaricales, Russulales, and Polyporales, with at least 120 transitions from non-gasteroid to gasteroid forms, from both clavarioid/coralloid and pileate-stipitate ancestors. Gasteromycetation does not affect diversification rates. Analyses of diversification dynamics of nutritional modes are ongoing.
Species boundaries in Mycosphaerellaceae s. lat.
- J. Groenewald
- M. Bakhshi
- C. Nakashima
- S. Videira
- U. Braun
- P. Crous
The Mycosphaerellaceae (Capnodiales, Dothideomycetes) contains thousands of species, and currently includes 213 generic names according to MycoBank. These genera encompass many important plant pathogens, saprobes and endophytes, as well as extremotolerant species. Co-occurrence of multiple species or genotypes within the same lesion is a common phenomenon, as is the movement of species between diverse hosts in different plant families. While some species are highly host-specific, others can occur on several hosts or can temporarily occupy a substrate in search of their optimal host (“pogo-stick hypothesis”). Homo- and heterothallic species occur in the same genus, with species observed as being strictly sexual, asexual, or being able to readily produce both morphs in culture. In the past, morphology and host association played a major role in species identification and naming. However, the decrease in cost of Sanger sequencing and the availability of universal primers for numerous fungal house-keeping genes have changed the way we approach species discovery and identification. The ITS nrDNA was accepted as the official barcode gene for Fungi. However, this locus lacks resolution for species identification in several genera in the Mycosphaerellaceae, such as Cercospora, Pseudocercospora, Ramularia and Septoria. Commonly used secondary barcode genes for these genera include partial actin, beta-tubulin, calmodulin, histone H3, translation elongation factor 1-alpha and DNA-directed RNA polymerase II second largest protein subunit. What is evident from the resulting individual gene trees is that no single protein-coding gene is able to distinguish all species known from DNA data; and ITS nrDNA is consistently the worst locus for phylogenetic species recognition for several genera in this family. A combination of at least two protein-coding loci is routinely required to resolve most of the species. Although the use of multilocus sequence data provides better resolution for species delineation, there are three major constraints which are encountered when screening new loci. These are 1) ease of amplification for all isolates of the genus, 2) level of resolution, and 3) the availability of material for DNA comparisons. However, the increasing number of publicly available genomes of especially Dothideomycetes species could provide a source of alternative phylogenetic markers which could lead to the identification of genes that are easier to amplify universally, and have improved resolution at the species level. The impact of multilocus DNA data on generic and species concepts in Mycosphaerellaceae are discussed, along with future challenges.
Re-evaluation of the genera in the Chaetomiaceae
- X. Wang
- F. Bai
- R. Samson
- P. Crous
- J. Houbraken
The Chaetomiaceae (Ascomycota) are well known for their cellulolytic activity and the production of various biologically active secondary metabolites that have great potential in agriculture, medicine and industry. On the other hand, many species are also able to grow in the indoor environment, causing adverse health effects, or are reported as causal agents of fungal infections in humans. Most genera occurring in Chaetomiaceae have not been revised based on molecular data. In this study, genera of Chaetomiaceae are re-evaluated on the basis of morphology and a four-locus DNA phylogeny. The asexual genera Botryotrichum, Humicola and Trichocladium are redefined to include species that reproduce sexually, and Chaetomium longicolleum is shown to represent the sexual morph of Staphylotrichum. Furthermore, most of the other known genera in Chaetomiaceae are re-circumscribed. The present study is a first attempt to establish an inclusive modern classification of the Chaetomiaceae composed of monophyletic genera.
Colletotrichum gloeosporioides s.l.: a multilocus species tree and a flexible metabarcoding framework for species identification and diversity exploration
- V. Doyle
- C. Baer
- R. Gazis
- S. Rehner
Colletotrichum gloeosporioides s.l.: a multilocus species tree and a flexible metabarcoding framework for species identification and diversity exploration
The Colletotrichum gloeosporioides (Cg) complex is an ecologically and economically important lineage of plant-associated fungi that includes diverse pathogen and endophyte species. Prior misperceptions that Cg are host specific yielded a proliferation of species recognized primarily on the basis of host affiliation. However, molecular phylogenetic characterization of Cg associated with single and multiple sympatric plant hosts have shown that many pathogen and endophyte species possess multi-host ranges, while also demonstrating that Cg encompasses extensive terminal phylogenetic diversity. While the community of biologists investigating Cg is in consensus that multilocus sequencing is essential for improved accuracy of species delimitation, the suite of multilocus markers in current usage often vary among studies and individual markers differ widely in phylogenetic informativeness. Based on demonstrations that the Apn2/Matigs nuclear intergenic spacer is one of the most informative phylogenetic markers yet developed for Cg, we introduce eight novel nuclear intergenic markers for Cg selected by comparative genomic analysis, rank their phylogenetic informativeness, and present a multilocus phylogeny of ~80 Cg terminals representative of pathogen and endophyte diversity from the Neotropics, many of which represent lineages with pantropical/global distributions. This new set of markers, which possess robust signal for Cg phylogenetics, will improve our ability to accurately delimit species boundaries and explore the roles of lineage sorting and hybridization as sources of topological conflict in the Cg phylogeny. In addition, the multilocus species tree can serve as a flexible tool for species identification and diversity discovery, which can be achieved by use of any individual marker for metabarcode analyses.
Insights into Claussenomyces Kirschst.: Past, Present and Future
- L. Quijada
- N. Matočec
- I. Kušan
- H. Baral
- J. Mitchell
- J. Karakehian
- J. Kout
- D. Pfister
The Leotiomycetes are recognized as one of the most diverse classes of Ascomycota. Despite the high diversity in this class, certain orders and families have a high proportion of taxa that have not been studied using molecular methods. A good example of the paucity of molecular data is found in Tympanidaceae (Baral 2015), a poorly studied family containing nine genera, which have previously been placed in the Bulgariaceae, Dermateaceae, and Helotiaceae: Claussenomyces, Collophora, Durandiella, Grovesiella, Holwaya, Micraspis, Myriodiscus, Pragmopora, and Tympanis. Some of them are associated with diseases such as bark and xylem lesions, internal necrosis, vascular streaking, cankers and crown damage (Quijada 2015, Baral 2016). The genus Claussenomyces was erected by Kirschstein in 1923 for a single species (C. jahnianus). Korf & Abawi (1971) drastically widened the concept of the genus combining and adding three species: Holwaya salicis E. Müll. & S. Ahmad, Corynella prasinula (P. Karst.) Boud., and Corynella atrovirens (Pers.) Boud. Currently, Claussenomyces is one of the most diverse genera in Tympanidaceae with 16, mostly conifer-associated species, with corticolous, lignicolous, fungicolous, and resinicolous lifestyles. A monographic treatment of the genus does not exist, and only a few species — C. kirschsteinianus, C. prasinulus, C. olivaceus, C. cf. hydnicola, Claussenomyces sp. or uncultured Claussenomyces — have publically available sequences (NCBI). Baral & Marson (2005) provided for first time a key to species of the genus, but also a revision of several species and types (C. jahnianus, C. pusillus, Tympanis xylophila). In 2013, the first author started to work in the genus and reviewed more than 40 collections, including most of the types with the exception of C. kirschsteinianus, C. simplex and C. pleomorphicus. Another additional 15 collections have been treated in detail jointly by the second and third author previous to the present work. Current results of our morphological and phylogenetic analyses revealed that Claussenomyces is a polyphyletic genus that should be split into at least four genera represented as: C. jahnianus, C. atrovirens group, C. prasinulus group, and C. kirschsteinianus group. The three former are related to Phacidiales whereas the latter is related to Helotiales. Although C. jahnianus still lacks molecular data, we consider it different from all remaining species based on its deviating morphology, but tentatively place it in the Phacidiales. In addition, several species currently included in Claussenomyces belong to other genera (i.e. Claussenomyces pini in Durandiella, C. tympanoides = C. pusillus in Holwaya), or are synonyms of other species in the Claussenomyces (i.e. C. canariensis and C. clavatus being conspecific with C. atrovirens). In the future, the 16 current binominals will be reduced to 10 species, divided into four genera: Claussenomyces, restricted to its type species (C. jahnianus), and three newly erected genera defined by phylogenic, microscopic, and ontogenetic features as follows: (1) fungicolous, lignicolous and resinicolous species with true ascoconidia (C. atrovirens group, 6 species); (2) lignicolous species without ascoconidia (C. prasinulus, 1 species) and (3) resinicolous species without ascoconidia (C. kirschsteinianus group, 2 species).
Divergence on the side of speciation: a peek at ergot alkaloid gene evolution in Claviceps purpurea and relatives
- W. Findlay
- K. Dadej
- O. Pochopsky
- P. Shoukouhi
- H. Nguyen
- J. Menzies
- K. Seifert
- M. Liu
Ergotism, a gruesome malady of humans and animals, is caused by consumption of grains contaminated with sclerotia of Claviceps (ergot), which produce a wide spectrum of potent mycotoxins namely ergot alkaloids (EA). These alkaloids have caused significant health, social and economic concerns at different times in history, but have also provided potential cures for various stubborn diseases. The contrary effects of these compounds have attracted a great interest in understanding their production by fungi. Molecular biology has brought to light the ergot alkaloid synthesis (EAS) genes and the pathways responsible for EA production. The presence or absence of certain EAS genes can be used to predict the production of EA by different species and the variance of EAS gene sequences could be used to develop DNA-based assays to predict the production of EA substrates. The objective of this study was to examine the sequence variances and to gain an understanding of the evolutionary patterns of EAS genes of Claviceps purpurea and close relatives to provide background knowledge for developing DNA-based detection tools for EA substrates. The whole genomes of 29 strains of C. purpurea and close relatives were sequenced by Illumina platform and assembled. EAS gene clusters were extracted from each genome using the reference sequences of C. purpurea strain 20.1 (GenBank: JN186799.1) and in-house pipelines. Full span DNA sequences of 11 EAS genes were recovered. DNA polymorphism of each individual gene was analyzed by using DnaSP. DNA matrices of each gene were subjected to three approaches for phylogenetic analysis: maximum parsimony (MP), bayesian (BA) and maximum likelihood (ML). The results indicate that EAS genes evolved with different patterns at different rates, and that possible gene duplication events happened before and after speciation.
Rearrangement of Pestalotiopsis sensu lato by anastomosis grouping
- S. Nozawa
- K. Watanabe
Filamentous fungi have been classified by molecular phylogenetic analysis. In 2014, the genera of Pestalotiopsis sensu strict, Neopestalotiopsis and Pseudopestalotiopsis which differ minimally in terms of conidial morphs and sexual morphs, were separated from Pestalotiopsis sensu lato based on a phylogenetic analysis of the 28S nrRNA gene (LSU) region. Simultaneously, a data set combining the internal transcribed spacer region (ITS), β-tubulin, and the partial translation elongation factor 1-alpha (TEF1) were introduced as better molecular set for understanding each species within the genera Pestalotiopsis and Pseudopestalotiopsis, respectively. However, in the phylogenetic tree of LSU, the boot strap value on the node connected with Neopestalotiopsis and Pseudopestalotiopsis was low, and both genera appeared to fall into one clade. Regarding species classification, phylogeny based on the combined data set loses stability as the number of found new species increases.
Within the concept of natural classification, we have been searching for useful indicators of fungal relationships, other than the molecular data sets conceived for the genera and/or species classification. When classifying Rhizoctonia, the lineage obtained by anastomosis grouping reflected that obtained by molecular phylogenetic analysis, using the ITS region (Sharon et al., 2008). We used hyphal anastomosis to classify Pestalotiopsis s. lat. within the concept of biological species, since this fungal group does not produce sexual morphs and anastomosis is the first event to distinguish self from non-self in the sexual reproduction process. In our preliminary test, we found that within the same genus the hyphae could fuse within the same species and between different ones. Thus, all species within one genus have an ability for hyphal anastomosis with each other, which expanded the understanding of lineage at the genus level. Hyphal anastomosis experiments were conducted using species from the same genus and from different ones, where evaluation of hyphal anastomosis, was based on the transformation of GFP or RFP into strains of each genus. Additionally, we investigated whether molecular the results of phylogenetic analysis reflected those of anastomosis grouping. For molecular phylogenetic analysis, the LSU, ITS, TEF1 etc. were used, and the evolutionary distance was estimated by phylogenetic trees, using the methods of maximum likelihood and neighbor joining. Finally, we rearranged the classification of Pestalotiopsis s. lat.
Pseudopestalotiopsis hydeae sp. nov. a new species from Diospyros sp.
- H. Ariyawansa
- W. Chang
- S. Maharachchikumbura
- I. Tsai
We illustrate a taxon of Pseudopestalotiopsis that is new to science from the culture collection of National Taiwan University (NTUCC). The genus Pseudopestalotiopsis consist of 14 formerly described species and are well recognised for their ability to produce unique medicinal compounds that may have pharmaceutical and agricultural applications. We established multi-disciplinary analyses using single- and multi-gene (ITS, β–tubulin and tef1) phylogenies together with morphology to assess the taxonomic status of the Pseudopestalotiopsis species isolated from Diospyros sp. in Taiwan. The new taxon compatible with the species of Pseudopestalotiopsis in having dark concolourous median cells with knobbed apical appendages. The results suggest that our Pseudopestalotiopsis isolate is morphologically and genetically distinct from its closely related species Ps. camelliae-sinensis and should be recognized as Ps. hydeae sp. nov. Further, this study increases the base of information regarding the diversity of Pseudopestalotiopsis species that occur in Taiwan and to the best of our knowledge, this is the first record of Pseudopestalotiopsis species associated with Diospyros sp. in Taiwan.
Keywords: New record, New species, Pestalotioid fungi, Phylogeny, Taxonomy
Penicillium gravinicasei, a new species isolated from Apulian cave cheese
- P. Anelli
- S. Peterson
- M. Haidukowski
- A. Logrieco
- A. Moretti
- F. Epifani
- A. Susca
Several species of the genus Penicillium were isolated during a survey of the mycobiota of Apulian cave cheeses ripened in a cave in Gravina di Puglia, Italy. A novel species, Penicillium gravinicasei, is described in Penicillium section Cinnamopurpurea. Its taxonomic novelty was determined using a polyphasic approach, combining phenotypic, molecular (β-tubulin, calmodulin, ITS and DNA dependent RNA polymerase) DNA sequences and mycotoxin production data. The type strain of Penicillium gravinicasei is ITEM 17411 = NRRL 66733. Phylogenetic analyses of the RPB2 data showed that isolates of the novel species form a clade most closely related to Penicillium cinnamopurpureum and P. parvulum with high bootstrap support. The fungus did not produce ochratoxin A, citrinin, patulin, sterigmatocystin or aflatoxin B1 on PDA, MEA and YES. The novel species had a high growth rate on agar media supplemented with 5% NaCl, and could be distinguished from other section Cinnamopurpurea species by phenotypic and molecular characteristics.
Daldinia sp. nov, from a community forest in Northern Thailand
- S. Wongkanoun
- P. Srikitikulchai
- L. Went
- M. Stadler
- J. Luangsa-Ard
Hypoxylaceae is one of the well-known and widely distributed families of pyrenomycetous fungi with unitunicate asci and pigmented ascospores. Our survey and collection of Xylariaceae and Hypoxylaceae in the northern part of Thailand has found many specimens and one of these may be a new species of the genus Daldinia. The genus Daldinia was described by Cesati & De Notaris in 1863 (Hypoxylaceae, Xylariales, Xylariomycetidae, Sordariomycetes, Pezizomycotina, Ascomycota). In this study Daldinia sp. nov was found on decaying wood from a community forest in Chiang Dao district, Chiang Mai province, northern area of Thailand. It is characterized by productions of superficial small to widely effused, pulvinate stromata, with mouse grey to pale mouse grey surface, inconspicuous perithecial mound. The perithecia are monostichous lanceolate with black drop, umbilicate ostiole, containing unitunicate cylindrical asci, unicellular ellipsoid dark brown to blackish brown ascospores, and with straight to slightly curved germ slit much less than the spore length on convex size, smooth perispore dehiscent in 10% KOH. The morphological characteristic of this fungus is very similar to that of D. placentiformis in the shape of stromata, but is different in KOH extraction in producing dark vinaceous pigment. However, phylogenetic analysis of the multiple loci including internal transcript spacer region (ITS), large subunit of the rDNA (LSU), second largest subunit of the RNA polymerase II (RPB2), and beta-tubulin (TUB2) shows this fungus is placed in the Hypoxylaceae and is closely related to D. korfii. It is clear that this fungus showed inconspicuous horizontal zones but D. korfii has conspicuous zones. The stromatal acetonitrile extraction contains BNT, and 2 new compounds (Cytochalasin). Critical examination of the phenotypic characters indicates that this fungus represents a new species in Thailand.
The segregation of Hypoxylaceae as a family: A good example of a profound revision within the ascomycota
- L. Wendt
- E. Sir
- E. Kuhnert
- S. Heitkämper
- C. Lambert
- A. Hladki
- A. Romero
- J. Luangsa-Ard
- P. Srikitikulchai
- D. Peršoh
- M. Stadler
Stromatic Xylariales exhibit an extraordinary diversity in almost every aspect: They are distributed over the entire globe and colonize various habitats – although primarily known as predominant endophytes, they show a tremendous morphological plasticity, and produce a wide variety of secondary metabolites. A multi-gene phylogeny was conducted using partial ribosomal (ITS, LSU) and protein coding (RPB2 & TUB2) gene sequences of over 100 fungal strains. The topology of the phylogenetic tree backed-up previous research on morphology and chemotaxonomy of stromatic Xylariales and justified the segregation of the Hypoxylaceae from the Xylariaceae as its own family. Furthermore, two new genera were established within the Hypoxylaceae, Jackrogersella (formerly Annulohypoxylon) and Pyrenopolyporus (formerly Hypoxylon). Additionally, the genera Biscogniauxia, Camillea and Obolarina were expelled from the Xylariaceae and included into the Graphostromataceae – due to their high affinities to Graphostroma platystomum. In conclusion, the current classification of the Xylariales is in accordance with the asexual morphs and secondary metabolite profiles, rather than with the traditional concept that was based on ascospore morphology as the predominant criterion.
Diversity and distribution of order Pezizales (Ascomycetes) in Pakistan
- A. Niazi
- A. Khalid
- N. Afshan
Pakistan is a square stretch of land between Arabian Sea and Karakorum Mountains covering an area of 87.98 million hectares. The forest covers an area of about 4.2 million hectares which is equivalent to 4.8 % of the total land area. About 40% of the forests of this area are present in one of the province Khyber Pakhtunkhwa due to the moisture of the Himalayan area. These moist areas provide conducive environment for the fungal growth like ascomycetous fungi. Ascomycota is a division or phylum of the kingdom Fungi. Its members are commonly known as the sac fungi or ascomycetes. This is the largest phylum of Fungi which contains 15 classes, 68 orders, 327 families, 6255 genera and 64163 species worldwide. From Pakistan, so far 66 families, 309 genera and 1214 species of ascomycota have been reported. The 14 families & 34 genera belong to Archaeascomycetes lichenized ascomycetes, 2 families and 3 genera from unitunicate lichenized ascomycetes and 50 families 272 genera from bitunicate lichenized ascomycetes. The order Pezizales is the most diverse group of ascomycetes represented in Pakistan. The defining feature of this fungal group is the operculate ascus which is microscopic sexual structure in which non-motile spores, called ascospores, are formed. Most of these Ascomycetes are found in Himalayan moist temperate forests which occupy Kashmir, Murree & Hazara hills, lower Dir, upper reaches of Kurram Agency, moist parts of upper Swat, Gilgit and Baltistan because of moisture and dense vegetation cover. The maximum occurrence of these fungi is recorded from Khyber Pakhtunkhwa due to high rainfall and humidity. The data has been collected from different research articles as well as from Fungi of Pakistan.
Phylogenetic overview of Basidiomycota with divergence times of higher taxa and a phyloproteomics perspective
- R. Zhao
- S. Sánchez-Ramírez
- M. Stata
- J. Moncalvo
- Z. Yang
- Y. Dai
- K. Hyde
We provide a phylogenetic overview of Basidiomycota and related phyla in relation to ten years of DNA based phylogenetic studies since 2007. We selected 529 species to address phylogenetic relationships of higher-level taxa using a maximum-likelihood framework and sequence data from six genes (nrLSU, nrSSU, 5.8S, tef1-a, rpb1 and rpb2). These species represent 18 classes, 62 orders, 183 families, and 392 genera from the phyla Basidiomycota (including the newly recognized subphylum Wallemiomycotina) and Entorrhizomycota, and 13 species of Ascomycota as outgroup taxa. We also conducted a molecular dating analysis for 116 species representing 17 classes and 54 orders of Basidiomycota and Entorrhizomycota. Finally we performed a phyloproteomics analysis from 109 Basidiomycota species and 6 outgroup taxa using aminoacid sequences retrieved from 396 orthologous genes. The time-tree indicates that the mean of stem ages of phyla are ca. 530 Ma; subphyla of Basidiomycota are 406–490 Ma; most classes are 358–393 Ma for those of Agaricomycotina and 245–356 Ma for those of Pucciniomycotina and Ustilaginomycotina; most orders of those subphyla split 120–290 Ma. Most higher-level taxa of Basidiomycota are generally supported. However, the younger divergence times of Leucosporidiales (Microbotryomycetes) indicate that its order status is not supported, thus we propose combining it under Microbotryales. On the other hand, the families Buckleyzymaceae and Sakaguchiaceae (Cystobasidiomycetes) are raised to Buckleyzymales and Sakaguchiales due to their older divergence times. In general, the six-gene phylogenies are in agreement with the phyloproteomics tree except for the placements of Wallemiomycotina, six orders. These conflicting placements in the six-gene phylogeny vs the phyloproteomics tree are discussed. This leads to future perspectives for assessing gene orthology and problems in decipheringtaxon ranks using divergence times.
Diversity and host specificity in the genus Sarea Fr. (Ascomycota)
- J. Mitchell
- I. Garrido-Benavent
- L. Quijada
- J. Karakehian
- D. Pfister
First published by Fries in 1825, the genus Sarea today comprises two accepted species of resinicolous discomycetes. Both species have a very broad range, with S. difformis reported from North America, Europe, and northwestern Africa, and S. resinae reported from North America, Europe, northern and central Africa, and central and eastern Asia. Both species have also been reported in southern hemisphere locations, such as New Zealand, on non-native trees. Both species also have a broad range of hosts in the Pinaceae, with S. difformis reported on Cedrus atlantica and both Sarea species reported on species of Pinus, Picea, Larix, Pseudotsuga, Abies and Tsuga. In addition, S. resinae has been reported on species in the Cupressaceae, including members of the genera Cupressus, Chamaecyparis, Juniperus and Taxodium. With few exceptions, specimens of each Sarea species share a very similar macro- and micromorphology, with specimens from multiple hosts fitting the specific concepts published by Hawksworth and Sherwood in 1981. Some molecular work has been done on the genus, but in almost all cases sequences are not associated with a vouchered herbarium specimen including the sexual morph. The objective of this study is to determine the degree of relatedness of geographically distant specimens collected in North America, Europe, and Macaronesia on different native and non-native host species. With permission, collections have been made of both species of Sarea from California, Georgia and the New England states in the USA, Northern and Southern Europe and Macaronesia. In addition to detailed measurements of the micromorphological features of specimens, ITS and LSU sequences have been generated using Sanger sequencing for analysis and comparison with published sequences. In contrast to the generally only slight morphological differences noted among specimens, ITS sequences from Europe, Asia, and North America not only differ by about 4% from each other, but also when submitted to phylogenetic analyses form multiple well-supported clades for each continent. These patterns are supported by similar analyses using the LSU sequences and ITS+LSU sequences. These clades also point to host specificity at the host family or genus level. In conclusion, the composition of the genus Sarea seems much more complicated than previously reported, with the possibility of multiple cryptic species in both accepted taxa; additional work must be done to further expand geographical and host range sampling of specimens to include in these analyses in order to approach a full picture of the diversity in Sarea.
Status of Rust fungi on Polygonaceae in Pakistan: Some new reports and an updated checklist of recorded species
- N. Afshan
- A. Ishaq
- A. Niazi
- A. Khalid
In the present research work, we study Polygonaceae parasitized by three taxa of rust fungi in the Deosai plains, Pakistan. Puccinia shikotsuensis and P. polygoni-alpini are new records for Pakistan; Uromyces polygoni-avicularis is reported first time from Deosai Plains. Original illustrations, line drawings, descriptions and data on distribution and host range of these rust species worldwide along with a checklist of rust fungi of Polygonaceae in Pakistan are given. This work will not only be a baseline for further studies in the selected site but will also help in selection of means to protect this economically important plant family from described specific parasites.
New records of rust fungi (Pucciniales, Basidiomycota) from the Himalayan Forests of Pakistan
- A. Ishaq
- M. Aime
- N. Afshan
- A. Khalid
Rust fungi are well-known plant pathogens that cause diseases in both agricultural crops and wild vegetation. There are approximately 8,000 described species worldwide. The Himalayan Mountains of Pakistan are among the top 25 hotspots of biodiversity in the world. Here we present results from ongoing exploration of the rust fungi in the region. Many species have been documented for the first time from Pakistan, including Coleosporium tussilaginis, Melampsora dimorphospora, Puccinia phaeopoda and Puccinia polygoni-alpini and Macruropyxis fulva sp. nov. discovered. Full host data have been compiled, morphological features have been recorded from multiple collections, and DNA sequence data generated and analyzed. These and other new records will be presented. The documentation of new taxon from this region will enhance our understanding of fungal biodiversity in Himalayan ecosystems.
Mysterious species of Thekopsora (Pucciniales) producing dimorphic uredinial sori and spores from Jilin, northeast of China
- J. Ji
- Z. Li
- Y. Li
- M. Kakishima
Jilin Province is located in the northeast of China and the east mountainous areas including Changbai mountain ranges are rich in vegetation. However, the inventory and ecology of rust fungi have not been sufficiently investigated in Jilin Province. Therefore, we surveyed rust fungi in several locations in Jilin Province from 2013 to 2017 and collected about 1000 rust specimens. Among these specimens, we found specimens on species of Galium (Rubiaceae), Aster and Kalimeris (Compositae) producing two different types of sori (uredinia?) on the same plant leaves. One type was similar to uredinial sori of Thekopsora or Pucciniastrum, another one was similar to those of Coleosporium. We suspected that two different species of rust fungi infected the same host plants because several rust species belonging to these genera have been recorded on these host plant genera. For clarification of the rust species on these specimens we carried out morphological observations with LM and SEM, and molecular analyses with 28S and ITS regions of rDNA. As the results, different types of uredinial sori on the same host plants were phylogenetically identical and were produced by one species of rust fungi, not caused by contamination of two species, although the morphological structures of these types were different each other. We also found two different types of spores (urediniospores?); echinulate and verrucose spores, inside of the same sori. The phylogenetic analyses showed that these specimens included into the group of Thekopsora, but they were separated into two distinct clades, which were different from other species of Thekopsora including species reported on the same host genera. Therefore, we suspect that they are new members of Thekopsora producing dimorphic uredinial sori and spores. The functions of these spores in their life cycles are still unknown.
Crossopsora byrsonimae, a possible type species of a new genus of Cerrado rust fungi
- E. Souza
- S. Elias
- D. Pinho
- R. Miller
- M. Aime
- J. Dianese
Phakopsoraceae is a well known polyphyletic family of rust fungi. Most of the ca. 13 genera currently placed in Phakopsoraceae s.l. have not been subject to phylogenetic analyses. In this study we examined the placement of the genus Crossopsora within Pucciniales based on morphological traits and newly generated sequences of rDNA nucLSU. Both the type species, C. ziziphi, and 5 specimens of C. byrsonimae were phylogenetically compared with representatives of the order Pucciniales. The phylogenetic reconstructions using Bayesian Inference and Maximum Likelihood analyses showed that C. ziziphi does not belong to Phakopsoraceae s.l., and remains in an uncertain phylogenetic position in Pucciniales. Besides that, the specimens of C. byrsonimae were not congeneric with the type species C. ziziphi and, furthermore, not allocated in Phakopsoraceae s.l.. Thus, our data clearly indicated that C. byrsonimae can be considered as the type species of a new genus. They show in addition to the phylogenetic divergence, the two species show clear phenotypic dissimilarities that include diverse spermogonial morphology (group VI type 7 in the former and group VI type 5 in last), aecial type (Caeoma-like in C, ziziphi and Aecidium-like in C. byrsonimae), and different geographical distributions.
Morphological review and phylogeny of Cerradoa palmaea, the rust fungus of Arecaceae from the Brazilian Cerrado
- S. Elias
- E. Souza
- M. Ebinghaus
- D. Begerow
- J. Dianese
Cerradoa palmaea was described in 1976 by Ono & Hennen based on the holotype collected in Brasília, DF, Brazil. This was the first rust fungus reported on Arecaceae with well documented teliosporic and uredinosporic phases, although the host species indicated was incorrectly identified as Attalea ceraensis (sic) (correct spelling: Attalea geraensis). As shown by reexamination of the holotype, PUR F18664, it became clear that the real host is Syagrus commosa. In the original description, using light microscopy and excellent drawings, the new genus was compared with morphologically close genera such as: Edythea, Prospodium, Hemileia, and Desmella; all four also showing supra-stomatal telia and uredia. However, the differences in spore shape, sporogenesis, and hosts were then sufficient to recognize Cerradoa as a good genus. However, Cerradoa was considered a synonym of Edythea in the 4rd Edition of the Illustrated Genera of Rust Fungi-2003, without a solid justification. Here additional morphological data is inserted using SEM images and more updated light microscopy supported by Nomarski´s optics. Now, for the first time a molecular phylogeny is presented using rDNA nucLSU and Cox3 sequences of Cerradoa palmaea resulting in the allocation the genus within the limits of the Pucciniaceae, the largest family of the order Pucciniales. Based on the only available nucLSU sequences, Prospodium, and Desmella were completely segregated from Cerradoa when submitted to phylogenetic reconstrution using Bayesian Inference and Maximum Likelihood analyses; the same result was obtained for Hemileia using both nucLSU and Cox3. Lacking comparable Edythea sequences in GenBank it was impossible to arrive at a conclusion as far as its relationship with Cerradoa.
Identity and disease cycle of a smut fungus on wiregrass in a longleaf pine-grassland ecosystem in the southeastern USA
- J. Kerrigan
- A. Alqurashi
- K. Savchenko
- J. Walker
A smut fungus that hinders wiregrass restoration efforts in longleaf pine-grassland ecosystems is being investigated in North and South Carolina and Georgia. These ecosystems are unique to the southeastern USA; they are characterized by an open canopy of primarily longleaf pine (Pinus palustris) and a dense ground layer of herbaceous species. Wiregrasses, Aristida stricta and A. beyrichiana, are perennial bunchgrasses and the dominant grass found in longleaf pine forests. Once the predominant forest type in the southeast, longleaf pine forests have been reduced to a fraction of area they once covered due to land use changes and fire suppression. Seeds of Aristida species are required for regeneration efforts, but seed production has been affected adversely by a smut fungus. Smut fungi can be damaging pathogens of grasses and typically infect inflorescences of host plants, replacing the seeds with teliospores. Our objectives are to identify the smut species from A. beyrichiana and A. stricta, and to investigate the disease cycles. Based on microscopic examinations and comparisons of DNA sequences of the ITS, LSU, and GADPH regions, it is a previously undescribed species of Langdonia, which is a monophyletic genus found on Aristida species. Investigations are underway to study infection and colonization of the host by this smut pathogen. Understanding this host-pathogen system will help management efforts to increase the availability of Aristida spp. seeds.
Emphasizing South African rust fungi of the genus Ravenelia
- M. Ebinghaus
- W. Maier
- M. Kemler
- A. Brachmann
- M. Wingfield
- D. Begerow
The rust fungi are the most diverse group of obligate plant parasitic fungi and commonly infect the prevailing number of land plant lineages throughout major climatic regimes. The genus Ravenelia, however, appears to be confined to members of the legume family with tropical and sub-tropical distributions. With more than 250 species, it is the third largest genus of rusts and can easily be recognized due to its notably complex and distinguished teliospores. Here, we will present recent research on the diversity and phylogeny of this genus but emphasizing South African representatives. Our studies encompassed light- and electron microscopy as well as molecular phylogenetic techniques based on ribosomal DNA. This enabled us to present the first molecular phylogenetic analyses of Ravenelia and to describe 9 new species. The phylogenetic reconstructions show evidence for rust lineages specifically associated to monophyletic host groups but furthermore indicate a polyphyletic origin of Ravenelia. These studies serve as a basis to analyse and discuss phylogeographic patterns as well as aspects of the ecology and evolution in major lineages of Ravenelia rusts in South Africa. This study will be complemented by population genomic analyses using ddRAD sequencing.
Endophytic Cryphonectriaceae in the Melastomataceae threaten Eucalyptus plantation foresty in Colombia and globally.
- C. Rodas
- G. Granados
- A. McTaggart
- J. Roux
- M. Wingfield
The Cryphonectriaceae includes 23 genera, including numerous important tree pathogens of global and economic importance. In recent years, some species of Cryphonectriaceae have been shown to live as asymptomatic endophytes in Melastomataceae trees and shrubs in Africa, Asia and Latin America. Since early 2000s, host switches have been revealed where species of Cryphonectriaceae have moved from Melastomataceae to infect plantation grown Eucalyptus spp. In this study, we identified species of Cryphonectriaceae that occur in asymptomatic tissues of six different species in three genera of Melastomataceae that grow naturally alongside Eucalyptus (Myrtaceae) plantations in Colombia. Healthy branch segments were collected in five provinces of Colombia and incubated for two to four weeks to promote sporulation of endophytes. Fruiting bodies of Cryphonectriaceae that emerged were isolated and identified based on DNA sequence data for the Internal Transcribed Spacer and β-tubulin. Three species of Cryphonectriaceae, Aurapex penicillata, Chrysoporthe cubensis and C. inopina were identified. The results showed that a relatively large number of species of Melastomataceae are natural hosts of the Cryphonectriaceae in Colombia. These include the aggressive Eucalyptus pathogen C. cubensis that threatens plantation forestry in Colombia. The results of this study also emphasise the risks involved in moving apparently healthy stem tissue of Melastomataceae to new environments where some of these plants are popular amenity trees and shrubs.
Recent horizontal transfer of eight mitochondrial introns between two unrelated Ceratocystis pathogens of ohia in Hawaii
- C. Mayers
- T. Harrington
Two undescribed Ceratocystis species are associated with a new and alarming disease of native ohia trees (Meterosideros polymorpha) in Hawaii, rapid ohia death (ROD). Ceratocystis sp. A causes a severe wilt disease and is the primary cause of mortality, and Ceratocystis sp. B causes a canker-stain disease. Phylogenetic analyses of mitochondrial and nuclear genes place sp. A in the Latin American Clade (LAC) of Ceratocystis, where many other aggressive tree pathogens reside, whereas sp. B is clearly placed in the less-aggressive Asian-Australian Clade (AAC). The two pathogens often co-occur in diseased sapwood of ohia but appear to be sexually incompatible. To explore the possibility of genetic interaction between these two unrelated pathogens, we sequenced the genomes of eight sp. A isolates, one sp. B isolate, and twelve Ceratocystis relatives in the LAC and AAC. Circularized mitochondrial genomes extracted from the de novo assemblies of each isolate were all relatively large for fungi (97kb–160kb), and larger in sp. B (155kb) than in sp. A (127–143kb), with variation mostly due to accumulated introns. We identified 87 introns in 16 mitochondrial genes across 25 Ceratocystis mitochondrial genomes and compared intron distribution among isolates. The majority of introns in sp. A are shared by other LAC isolates with reduced identity, suggesting ancestral LAC introns. However, eight introns in sp. A appear to be of AAC origin, because they are not found in other members of the LAC but are found with 100% identity in sp. B and most are also found with 99%–100% identity in AAC representative C. uchidae. These eight introns vary in size (1082–3079bp) and in the mobile machinery they encode in intronic open reading frames, including reverse transcriptase (RT), maturase, and homing endonuclease genes (HEG). Four of the eight introns (in rnl, cox1, nad2, and nad6) are typical Group II introns that encode RT and maturase, one is a Group II intron in cob that encodes two HEGs but no RT or maturase, one is a Group IB intron in nad4 that encodes two HEGs, one is an unclassified twintron that encodes two HEGs and inserts into a Group ID intron of cox2, and one is a Group II intron in rnl with no apparent mobile machinery. Diagnostic primers designed for each of the eight introns were tested against 63 sp. A isolates and 16 sp. B isolates from Hawaii. All eight introns are present in all sp. B isolates, but they are haphazardly-distributed among populations of sp. A. The intron-laden sp. B appears to predate the recent introduction of sp. A to Hawaii, and as populations of sp. A spread across the island, transient hyphal anastomosis may have allowed the transfer of mobile introns from sp. B mitochondria to those of sp. A during their close proximity in diseased ohia sapwood. If true, this suggests that introgression of additional genetic elements between the two fungal species could have occurred, which might include pathogenicity factors and other parasitic DNA and have implications for the origin and future of ROD.
Ceratocystidaceae exhibit high levels of recombination at the mating-type (MAT) locus
- M. Simpson
- M. Coetzee
- M. Van Der Nest
- M. Wingfield
- B. Wingfield
Sexual reproduction is an important part of many fungal life cycles and is controlled by genes at the mating-type (MAT) locus. Two idiomorphs are present at this locus and each contain distinct MAT genes. The presence of particular MAT genes are responsible for whether the fungus is self-sterile (heterothallic) or self-fertile (homothallic). The Ceratocystidaceae is a family of Ascomycetes that includes many economically important plant pathogens and saprobes. These species display different mating strategies, which makes the MAT loci in this family particularly interesting. The aim of this study was to determine and compare the gene content and structure of the regions flanking the MAT locus in 12 species of Ceratocystidaceae representing five genera. Our results showed that the genes typically found flanking the MAT locus, such as SLA2 and APN2, in the Sordariomycetes were present in the genomes of the Ceratocystidaceae analysed in this study. There were, however, differences in gene order and presence of these genes around the MAT locus in the various species examined. This was especially evident when synteny in the regions immediately flanking the MAT locus was assessed. For example, species of Huntiella and Endoconidiophora were highly syntenic outside of the MAT locus, while the three species of Ceratocystis did not display much synteny. Ceratocystis spp. also had more transposable elements in the regions flanking this locus compared to other genera in the Ceratocystidaceae. We hypothesise that the transposable elements Ceratocystis may have facilitated recombination in the mating type region.
Characterization of the MAT locus in species of Ceratocystidaceae utilizing uni-directional mating-type switching
- P. Wilken
- E. Steenkamp
- Z. De Beer
- M. Wingfield
- B. Wingfield
Sexual reproduction is an important part of the life-cycle of most Ascomycetes. This process is controlled by the genes present at a single genomic position, the mating-type or MAT1 locus. Characterization of the genes at this locus has made it possible to elucidate many unique sexual strategies. These include uni-directional mating-type switching, a form of homothallism characterised by the loss of DNA from the MAT1 locus. Although uni-directional mating-type switching is known only in a small number of Ascomycetes, three genera in the family Ceratocystidaceae utilize this switching mechanism as reproductive strategy. This represents the largest assemblage of related species known to undergo uni-directional mating-type switching. In this study, the structure of the MAT1 locus of nine species in the genera Ceratocystis, Endoconidiophora, and Davidsoniella were elucidated using full genome sequences. The results showed that the structure of the locus is conserved across all three genera. For each species, four genes made up the MAT1 locus: two MAT1-1 genes (MAT1-1-1 and MAT1-1-2) and two MAT1-2 genes (MAT1-2-1 and MAT1-2-7). In addition, two copies of a direct repeat flanked the MAT1-2 genes in each species. These repeat elements would anchor a recombination event where the MAT1-2 genes would be deleted from the genome. The loss of MAT1-2 is a hallmark of uni-directional mating-type switching, allowing self-fertile isolates to produce both self-fertile and self-sterile progeny. The availability of complete MAT1 loci for these species provides a crucial link to understanding the evolution of sexual reproduction in the Ceratocystidaceae.
Dissecting pathogenicity and virulence in Ceratocystis albifundus using a GWAS approach
- M. Van Der Nest
- N. Soal
- E. Steenkamp
- L. De Vos
- B. Wingfield
The African fungus Ceratocystis albifundus is an important pathogen of commercially propagated Acacia mearnsii and Protea cynaroides. It also infects a wide range of native tree species asymptomatically. Although populations of C. albifundus vary dramatically in terms of fitness traits such as pathogenicity and virulence, the molecular basis of these phenotypes remains unknown. We aimed to identify the possible pathways and processes encoded in genomic regions associated with pathogenicity and virulence. Accordingly, a collection of 36 genetically diverse C. albifundus isolates, originating from a wide range of host species and geographic regions, was used in a Genome Wide Association Study (GWAS). The isolates were phenotyped using pathogenicity assays on one year old A. mearnsii seedlings. To determine their genotypes, individual isolates were subjected to low-coverage genome sequencing using Ion Torrent technology. These sequences were mapped to a high-quality hybrid reference genome for the fungus, which was determined using the PacBio RS II and Illumina HiSeq sequencing platforms. Following quality and frequency-based filtering, ca. 17 000 single nucleotide polymorphisms were identified within the collection of isolates. Correlation analysis using these polymorphisms allowed identification of a number of genomic regions significantly associated with pathogenicity and virulence. Sequence analysis revealed that the genes occurring in these regions encode products involved in various functions that likely influence pathogenicity and virulence. Subsequent functional characterization of these genes will confirm their involvement in these fitness traits of C. albifundus. Our findings demonstrated the power of GWAS for determining the molecular basis of important phenotypic characters in fungal pathogens.
Co-evolutionary patterns in Phragmidium (Pucciniales) are blurred by the reticulate evolutionary history of its Rosaceae hosts
- M. Ebinghaus
- B. Xu
- S. Lotze-Engelhard
- M. Kemler
- D. Begerow
- Z. Jiang
Rust fungi of the genus Phragmidium infect genera and species within the Rosaceae, including the economically important genera Rosa and Rubus. As rust fungi are obligate plant pathogens most species exhibit pronounced host specificity and cospeciation with their hosts is traditionally thought to be common. In Phragmidium, however, several species are able to successfully infect a broad number of congeneric hosts. It has been known for a long time that a number of species within the host genera Rosa and Rubus are prone to hybridization that is reflected by difficulties in terms of morphological species recognition. Recent molecular based studies confirmed not only the tendency for hybridization in nature but also suggested that hybridization was a main driver in the evolution of Roses and Rubus species. This process is termed reticulate evolution instead of a strict bifurcating evolution. For the species pair of dogrose rusts, Ph. mucronatum and Ph. tuberculatum, it was postulated that their broad host ranges may be a result of the reticulate history of its hosts by enabling the rusts to shift from one host to another via a “hybrid bridge” of shared infection relevant traits. Due to the common phenomenon of reticulation in Roses and Rubus, we hypothesized to find widespread evidence for hybrid bridges in this host – parasite system. In the present study, we focused on rusts collected in China and analyzed a set of 56 specimens that represent 24 species of Phragmidium. Molecular phylogenetic analyses of 28S rDNA revealed highly polyphyletic host associations that indicate several independent host colonization events and thus strict cospeciation is unlikely. Our analyses further support the current view that the reticulate evolutionary history of the host species enables the rust pathogen to infect several different host species. This process may also facilitate jumps to even more distantly related hosts and which offers an explanation for the blurred co-evolutionary patterns observed within this genus but also for the broad host ranges frequently found in Phragmidium.
Taxonomy and mating strategy of Thielaviopsis basicola - a globally important pathogen of agricultural crops
- W. Nel
- T. Duong
- A. Hammerbacher
- B. Wingfield
- M. Wingfield
- Z. De Beer
Thielaviopsis basicola (Ascomycota: Ceratocystidaceae) is a well-known pathogen that causes the disease commonly known as black root rot on multiple important plant species. The species has been extensively studied since its first description more than 150 years ago, but important questions regarding its taxonomic placement and mating behaviour remained to be answered. The aims of this study were to resolve the taxonomic placement of the species using a multi-gene phylogenetic approach, and to determine the sexual reproductive strategy of the species. Phylogenetic analyses of sequence data from six gene regions from a relatively large number of isolates from 13 geographic regions showed that the species groups as a distinct generic lineage in the Ceratocystidaceae, which we subsequently described as Berkeleyomyces gen. nov. Additionally, our analyses also separated this collection of T. basicola isolates into two well-supported lineages, which were recognized as two distinct species. One of these lineages was selected to represent the originally described species and was described under the new combination B. basicola. The second lineage was described as a novel sister species named B. rouxiae. To investigate the sexual reproductive strategy of the two species, we sequenced and assembled the genomes of two isolates of B. basicola. Genome analyses following with mating gene specific PCR indicated that B. basicola and its sister species have a typical heterothallic mating system in which individual isolates of both species harbour either the MAT1-1 or the MAT1-2 idiomorphs. Mating type diagnostics using newly developed PCR-based mating type markers revealed that both mating types of both species are present in nature, although a sexual state for either of the species remains to be discovered.
Intraspecies genetic variability and mating compatibility in basidiomycete fungus, Fomitopsis pinicola
- A. Shnyreva
- A. Shnyreva
The polypore fungus, Fomitopsis pinicola, is a widespread basidiomycete species in forests of central Russia. Polypore fungi are known to be the main decomposers of lignocellulose litter in nature. Besides, they are applied in traditional Chinese and Russian medicine due to a wide range of bioactive metabolites. That is why the investigation of genetic resources of polypore fungi is of importance. The red belt conk, F. pinicola, represents an excellent organism for genetic study because of the heterothallic type of sexual compatibility and a haploid-dikaryotic life cycle which can be reproduced under the laboratory conditions. Mating criteria are easily testable by clamps formed on dikaryotic mycelia. Somatic incompatibility mechanism, which regulates intraspecific recognition of self or non-self-partner under somatic contacts between heterokaryotic mycelia, is also easily testable on Petri plates. In this study, genotype diversity, sexual and somatic compatibility were analyzed in F. pinicola natural populations. Fruit bodies were collected in forests of Moscow suburbs through summer and autumn in 2013-2015. Mycelial cultures were isolated from fruit bodies, and a total 37 isolates of F. pinicola were deposited in the collection MSU_BIO_EBF together with 9 strains from Finland and France. For screening mating compatibility, dikaryotic isolates were subjected to analysis in mon-mon matings against the tester monokaryons. To proceed mon-mon matings, we obtained the fertile hymenium producing sterile basidiospore prints on Petri plates. Monokaryotic testers with different mating type alleles (AxBx, AyBy, AxBy, AyBx) were deposited in the collection. Intraspecies genetic polymorphism was analyzed by using variable ITS-region of rRNA gene cluster. Minisatellite DNA SSR analysis was performed on genomic DNA with four different primers (ISP1, ISP2, ISP4 and ISP6). Dendrograms of genetic similarity between the strains were constructed based on ITS sequences and minisatellites markers using MEGA7 and TREECON program accordingly. F. pinicola was shown to have the tetrapolar mating compatibility system which is controlled by two unlinked loci with multiple alleles. All strains possessed different alleles except those grown within the same substrate (a log) at short distance, not more than 3 meters. Test on somatic compatibility between the natural isolates revealed somatic clones only within the same log. In the F. pinicola populations, moderate antagonistic (somatically incompatible) responses were predominated (frequency p=0.48) with the overall diversity index Hvc=0.782. The species showed very low ITS sequence divergence, as well as homogeneity of morphological characters among the strains from geographically distant origins, e.g. Russia, Finland, and France. Similarity between the strains were up to 99-100%, that suggests rather low polymorphism of the species. This trend was also reflected on the phylogram demonstrating little genetic differentiation within the species F. pinicola, proving that this is a highly outcrossing heterothallic fungus with panmictic populations. Assessment of genotypic variability based on minisatellite DNA markers showed low genetic polymorphism as well: genetic distances between the strains ranged from 0.05 to 0.18. However, the strains of different origins were separated on the dendrogram and were clustered according their geographical origin. The research was supported by RSF grant No. 14-50-00029 and RFBR grant 15-54-05065.
Population genomics of the globetrotter Serpula lacrymans reveal tight bottleneck and local adaptations
- I. Skrede
- C. Murat
- J. Hess
- J. Sønstebø
- S. Maurice
- A. Kohler
- D. Barry
- N. Högberg
- D. Eastwood
- F. Martin
- H. Kauserud
The bolete Serpula lacrymans is likely the most devastating fungal wood-decayer of construction materials in temperate regions worldwide. From its native range in North East Asia is has spread to most other continents, including Europe and Southern Oceania, as well as other parts of Asia (Japan). In this study, we performed population genomics analyses of 39 isolates from these three regions in order to assess differences across the invasive populations. More specific, we wanted to analyse the demographic history and level of intra-geographic structuring and local adaptations in the invasive populations. The three populations, Europe, Japan and New Zealand (the latter only represented by three isolates) were strongly differentiated. The European population had clearly gone through a tight bottleneck during its establishment while the Japanese population is far more diverse. Long genomic blocks without genetic diversity in Europe suggest a single founding event, with no secondary migration from the native regions. In both regions, there was a clear geographic sub-structuring, indicating dispersal limitations. Coalescence simulations was used to study the time since the populations diverged and the changes in effective population size. In scans for genetic adaptations in the two populations, we identified three outlier loci subject to purifying selection. These were linked to transport and decay, and support that these functions are particularly important in the human-made habitats of S. lacrymans worldwide. Of the three isolates from New Zealand, one had an admixed genomic make up with genetic elements from both the European and the Japanese populations. Since the original ‘out of Asia’ dispersal events, our analyses indicate that the dry rot fungus has travelled between continents, also giving rise to admixed genotypes.
Tracing the naturalization of golden oysters in the United States
- A. Bruce
- T. Osmundson
Wild fruitings of non-native golden oyster mushrooms (Pleurotus citrinopileatus) began appearing in North America approximately 5 years ago. Golden oysters are indigenous to eastern Russia, northern China, and Japan, but appear to have naturalized into American woodlands rapidly. The large fruitings observed by mushroom collectors suggest that indigenous saprobes are being outcompeted and displaced, presenting a potential threat to fungal biodiversity. I aim to infer how P. citrinopileatus populations have become naturalized, using population genomic data to infer migration pathways and relatedness between wild and cultivated strains. I will test two competing hypotheses for the spread of this organism: 1) multiple introductions have occurred (i.e., that each naturalized isolate is most closely related to a commercial strain; or 2) a single escape of a commercial strain occurred, followed by spread of wild populations. I will also attempt to determine whether naturalization via spores (i.e., involving recombination and more likely due to accidental escape of basidiospores) or clonal spread (i.e., more likely due to intentional introduction) is better supported by genetic evidence. Genome sequencing of 29 wild specimens from 6 American states, as well as 7 cultivated strains, is currently underway. These data will be used to construct a single-nucleotide polymorphism (SNP) dataset to infer the most likely pathway of naturalization and spread.
Mating and infection strategies in the Botryosphaeriaceae: emerging insights from genome and transcriptome studies
- B. Slippers
- J. Nagel
- S. Naidoo
- A. Marsberg
- L. Bezuidenhout
- A. Postma-Smidt
- M. Wingfield
Species in the Botryosphaeriaceae are amongst the most common fungi found in tree microbiomes. While these fungi are common in asymptomatic tissues, they can also cause disease when the plants are under stress. Disease symptoms commonly include dieback and cankers and can have severe negative impacts in forestry and agriculture, as well as in urban and natural forests. DNA sequence-based phylogenetic studies have revealed substantial species diversity in the Botryosphaeriaceae with many species rare and seemingly endemic to certain regions. In contrast, these studies have also revealed that some species have a near global distribution and extensive host ranges. Despite their importance, many aspects of the biology of these fungi remain poorly understood, including their mating strategies and infection biology. In this study, we interrogated 29 of the genomes of Botryosphaeriaceae that have thus far been sequenced, including 12 that were added as part of the study. The analyses revealed that heterothallism is the likely ancestral mating strategy, but that there has been at least five transitions to homothallism in the Botryosphaeriaceae. Unlike other latent pathogens, species in the Botryosphaeriaceae appear to have expanded CAZyme repertoires, possibly contributing to their ability to access a range of carbohydrate resources in a broad range of hosts. Furthermore, transcriptome studies revealed the initiation of a necrotrophic infection, yet without expression of symptoms. While the latter studies are still at a preliminary stage, they are beginning to reveal the mechanisms that mediate a state of tolerance that make it possible for these latent pathogens to be so common in woody plants.
Inheritance of putative virulance genes in Microbotryum by artificial hybridisation
- S. Klenner
- B. Büker
- D. Begerow
Biotrophic fungi are characterized by intimate interactions with highly specific hosts and often cause devastating plant diseases. Growth and proliferation during biotrophic phases necessitates an adapted set of genes to survive in planta. Hybridisation of distantly related species should lead to new arrangements of virulence factors like effector and transporter genes enabling fungal hybrids to new or formerly resistant host plant species. We use the model genus Microbotryum infecting highly specific species of Caryophyllaceae to provide insights into processes of host adaptation and genome arrangements during hybridisation processes. A total of 2600 Silene latifolia POIRET seedlings were inoculated with wildtype, cross and hybrid strains of Microbotryum species to analyze the hybridisation potential and the viability of Microbotryum. Our preliminary results indicate that homospecific crosses and interspecific hybrids cause infection at lower disease rates than infections with the coevolved parasite M. lychnidis-dioicae. Interspecific hybrids do mostly not affect morphological plant traits except the count of buds that was significantly altered by fungal infection. Our data indicate that hybrid infected plants have an altered blooming period compared to the adapted parasite M. lychnidis-dioicae. The inheritance of putative virulence factors and its potential impact on host specificity as well as the role in host-pathogen interaction will be discussed.
'Missing links' to the fungi domesticated by termites
- L. Van De Peppel
- M. Nieuwenhuis
- A. Grum-Grzhimaylo
- M. Cárdenas Toquica
- D. Lodge
- A. Franco-Molano
- T. Baroni
- D. Aanen
The old-world fungus-growing termites (Macrotermitinae) cultivate fungi of the basidiomycete genus Termitomyces. The termites propagate their fungal symbionts via asexual spores inoculated on plant material upon gut passage, and most species transmit the fungal symbionts horizontally to new colonies via the occasional production of sexual fruiting bodies, the mushrooms. Phylogenetic reconstructions indicate that the symbiotic fungi all descend from a single domestication event some 30 million years ago, and that no ‘escapes’ of fungal symbionts to a free-living lifestyle have occurred afterwards. In 2007, three non-symbiotic species closely related to Termitomyces have been discovered. Those species have some interesting traits that appear to make them ‘missing links’ between the domesticated and free-living fungi. First, they combine asexual and sexual reproduction on a single morphological structure, which is exceptional for basidiomycete fungi. Second, they grow on fecal pellets of an as yet unknown animal, probably an insect, which is similar to the way termites cultivate their fungi, viz. on termite dung. We present the results of phylogenetic analyses of mitochondrial genomes to reconstruct the phylogenetic position of those newly described species relative to the domesticated fungi, to reconstruct character-state evolution and attempts to identify the animal species producing the dung on which those fungi are growing.
The echo of a distant time in labyrinths of coral caves: Genomic analysis of Apterostigma-farmed Pterulaceae and free-living counterparts
- C. Leal-Dutra
- B. Dentinger
- A. Rodrigues
- G. Griffith
Approximately 60 million years ago, the common ancestor of the fungus-farming ants (subfamily Myrmicinae, tribe Attini, subtribe Attina; hereinafter referred to as “attine”) swapped its hunting-gathering behaviour to fungal agriculture. Since then, five different types of fungiculture have emerged, the attine leaf-cutters being the most specialised and advanced. The latter are the most studied group of fungus-farming ants due to the economic loss it causes in crops. However, the poorly studied Apterostigma "pilosum" group, an early diverging group of attine ants, also presents some clues that lead us to believe their crops are domesticated. Approximately at the same time the leaf-cutters started to domesticate their cultivar, Leucoagaricus gongylophorus, these Apterostigma switched to cultivation of the very distant coral mushroom family Pterulaceae (Agaricales). As in L. gongylophorus, the two Pterula cultivars do not produce fertile basidiomes and were never reported growing outside the symbiosis. Therefore, after extensive fieldwork in Brazil, several samples of the pterulaceous cultivars and their free-living counterparts were isolated into axenic cultures. Genome sequencing of these cultures of Apterostigma cultivars and free-living Pterula has allowed us to explore the evolutionary consequences of domestication in these fungi.
Co-divergence of slimy salamanders (Plethodon spp.) and microbial communities of the skin and gut
- D. Walker
- M. Albecker
- M. McCoy
- C. Camp
- C. Kelehear
- A. Hill
- J. Wooten
- J. Rheubert
- P. Tinker
- D. Talbert
- S. Graham
Recent research recognizes a multicellular host and its symbiotic microorganisms as the ‘holobiont’—a single unit of evolution. This study used high-throughput sequencing to characterize the complete skin and gut microbiome including both bacteria and fungi of nine closely related terrestrial slimy salamanders (family Plethodontidae). We characterized the salamanders as a holobiont to determine if microbial communities correlate with the evolutionary divergence of host nuclear and mitochondrial gene trees. We hypothesized that the skin and gut microbiome become more specialized to the host species over time, and that evolutionary divergences in the microbiome occur in tandem with host speciation events. Five host clades were compared within a phylogenetic context, and nine sub-clades were assessed for differences in microbial assemblage across sub-clade and geographic location. We found that the complete microbiome is similar in α- and β-diversity between host sub-clade but differs by geographic collection location. To infer microbiome function, we identified antifungal species in the skin- and gut-bacterial communities. In the fungal-gut microbiome, saprotrophs, pathotrophs, and symbiotrophs were the most abundant functional groups. Using a novel approach based on machine learning, we accurately determined host clade using the microbiome and identified indicator taxa specific to each host clade. We found that salamander individuals with similar mitochondrial gene sequences share similar gut-bacterial, gut-fungal, and skin-bacterial communities, indicating possible codivergence of host-microbial communities. Lastly, phylogenetic divergence of 20 microbial operational taxonomic units correlated with divergence of the salamander mitochondrial gene tree, indicating microbial symbionts may share evolutionary history with their host.
How common is mycovirus host-switching in nature?
- J. Myers
- T. James
Emergent viral diseases are often a result of host-switching, which has conventionally been thought a rare phenomenon. Host-switching in fungal RNA viruses(mycoviruses) is thought to be especially rare since mycovirus transmission is presumably restricted to intracellular strategies. Extracellular transmission is the primary transmission strategy of nearly all other viruses however, and we hypothesize that the absence of extracellular transmission in mycoviruses is unlikely. Further, recent evidence demonstrates that host-switching is more frequent in RNA viruses than previously recognized. Are viruses in fungi really an exception to this? If so, how do they defend themselves from extracellular invasion?Whether, and to what extent, host-switching of mycoviruses occurs in nature has not yet been tested. If mycoviral transmission is as restricted as currently thought, theory predicts coevolution of mycovirus and host. Thus, we take a phylogenetic approach to test our hypothesis that host-switching of mycoviruses occurs. Soils have been collected in Michigan from environments that are similar in composition but spatially distant. Common fungal species are being selectively isolated from each soil, screened for mycoviruses, and the phylogenetic relatedness of mycoviruses infecting those fungi will be compared. If mycoviruses from the same soils, regardless of fungal host, are more genetically similar than viruses in the same species from different soils, then host-switching is supported. The guiding questions of this research address our basic understanding of disease and have implications important to human, wildlife, agricultural, and ecosystem health.
Identification and initial characterization of RNA mycoviruses infecting the chestnut pathogen Cryphonectria naterciae
- C. Cornejo
- H. Bragança
- D. Rigling
The ascomycete genus Cryphonectria includes known tree pathogens. The most studied species is C. parasitica – the causal agent of the destructive chestnut blight disease, which was introduced in North America and in Europe in the early 20th century. Some strains of C. parasitica however exhibit reduced levels of virulence due to a viral infection. The so-called Cryphonectria hypovirus (CHV) attenuates the pathogenicity of C. parasitica reducing its parasitic growth and sporulation capacity. CHVs are cytoplasmatic, unencapsidated dsRNA viruses with genome ranges between 9 and 13 kbp. CHVs have no extracellular phase and are transmitted mainly from infected to non-infected fungal strains via hyphal anastomosis. The discovery of CHV sparked off great interest in mycoviruses as natural biocontrol agents. The aim of this study is to screen additional species of Cryphonectria for the presence of CHV or other RNA viruses, (1) to characterize them molecularly and (2) to test their capacity in hypovirulence on chestnuts trees. In a pilot study, we detected the presence of conserved CHV-sequences in some C. naterciae isolates. These strains were used to extract dsRNA directly from lyophilized mycelium and to generate an Illumina cDNA library by reverse-transcription including specific sequencing adaptors. Currently, a de novo assembly of the reads is being aligned to viral sequences to characterize them molecularly. The potential for using these Cryphonectria viruses as agents for the biological control on chestnuts trees will be presented.
Phylogenetic inference of coevolution between fungi of the Asterina genus (Asterinaceae, Asterinales) and their host plants
- A. Firmino
- O. Pereira
- M. Berbee
Species of Asterina genus was proposed by Léveillé in 1845, being Asterina melastomatis Lév. collected in living leaves of a Melastomataceae in Brazil, the type species. The genus is characterized by having circular to irregular with a star-shapped fissure, superficial mycelium with lateral appressoria, bitunicate asci, and bicelular pigmented ascospores. Furthermore, the genus is biotrophic, growing superficially on living leaves of their hosts and are distributed on tropical and subtropical regions. The Asterina genus and the other members of the Asterinaceae family form specific interactions with the live cells of their hosts and are probably host-specific. Although this specificity is assumed by many mycologists, this hypothesis has never been tested in any experiment or checked by molecular data. Phylogenetic analysis involving Asterina spp. were constructed using nuc LSU rDNA (28S) sequences. While for the plants, ribulose-1,5-bisphosphate (RbcL) sequences were used. The phylogeny of the Asterina species and their respective hosts were used to generate the co-phylogenetic analysis using the Jane program. The phylogenetic analysis obtained through Bayesian Inference for the Asterina species generated a tree with high values of posterior probability. As well as for the Asterina species, the phylogenetic analysis obtained through Bayesian Inference for the host plants also generated a tree with high values of posterior probability, which corroborates with previous phylogenies. The Jane program found 1488 possible solutions that group the phylogeny of the parasites (Asterina spp.) to the phylogeny of the hosts. All possible solutions have the same cost (cost = 0). These 1488 solutions are very similar between them, and can be grouped into a single isomorphic solution (solutions with the same cost and same number of events, changing just the relative time - before or after a given node). All solutions demonstrate the occurrence of coevolution between the Asterina species and their host plants. The results obtained by molecular analysis in the present study corroborate with the history of the taxonomy of Asterinales that, since the first description of the type species in 1845 by Léveillé, strongly takes into account the host in the identification of the fungal species.
Phylogenetic delineation and geographic distribution of Laccaria nobilis and phenotypic relatives.
- C. Charley-Suarez
- G. Muller
- A. Wilson
Laccaria nobilis Smith is one of the larger and more charismatic species of the genus, originally described from the Rocky Mountains of Colorado. However, its species distribution includes Alaska, the Pacific Northwest, Mexico, the Midwestern United States, and Eastern Canada. Cryptic species within Laccaria is a very real possibility so the question is whether this distribution accurately reflects the distribution of L. nobilis or whether there are other morphological similar Laccaria species within this distribution. The purpose of this paper is to establish the phylogenetic identity of L. nobilis from its home range and compare this to specimens of L. nobilis from other parts of the continent. Molecular sequence data from the nrITS region, as well as the single protein coding genes RPB2 and EF1-alpha are used for phylogenetic evaluation. Maximum likelihood analysis of molecular sequence data for specimens of L. nobilis will identify a monophyletic clade of specimens from Colorado and then evaluate which specimens from a broader geographic range fit within this clade. This will ultimately help evaluate the hypothesis that large specimens of L. nobilis outside of Colorado, actually represent non L. nobilis species.
Non-concerted genomic evidence and the Ophiocordyceps sinensis ITS pseudogene hypothesis
- J. Zhu
- Y. Li
- Y. Yao
- Z. Zhang
- W. Xie
The “ITS pseudogene” hypothesis for Ophiocordyceps sinensis (Os) was raised by Li et al. on the basis of detections of the ITS1-5.8S-ITS2 DNA sequences of Genotypes #1,#5 Os (Groups A,C) in cultured mycelial genomic DNA of the single ascospore of natural Cordyceps sinensis (Cs) and the 5.8S gene cDNA of Genotype #1 (Hirsutella sinensis, or Hs) in a cDNA library constructed from total RNA of the ascosporic mycelia, but non-detection of 5.8S gene cDNA of Genotype #5. However the following scientific findings are inconsistent with the hypothesis. (1)Zhang et al., Mao et al. and Wei et al. reported the detections of Genotypes #3,#4,#5 Os in natural or artificial Cs, but non-detection of Genotype #1 Hs. (2)Three genome sequences (ANOV00000000, LKHE00000000, &LWBQ00000000) of Hs strains Co18, 1229, & ZJB12195 contain the ITS sequences of only Genotype #1, but not Genotypes #3-#6,#15-#17 (transition mutants), Genotypes #7-#11 (transversion mutants), and Genotype #12 (insertion/deletion mutants), and Genotypes #13-#14 (DNA segment reciprocal substitution hereditary variations). (3)PCR with using genotype-specific primers, amplicon-cloning sequencing and SNP mass spec genotyping revealed differential coexists of multiple genotypes of Os in the stroma, caterpillar body, ascocarps and ascospores of Cs during maturation. (4)EcoRI digestion assay, Southern blotting and RFLP revealed that the biomasses of the EcoRI-sensitive Os (GC-biased Genotypes #1-#3,#7-#14) and the EcoRI-resistant Os (AT-biased Genotypes #4-#6,#15-#17) are changing in a dynamic, asynchronous fashion during the Cs maturation. (5)Southern blotting revealed a single DNA moiety (EcoRI-sensitive Os) in the mycelia of Genotype #1 Hs, but a doublet (EcoRI-sensitive and -resistant Os) in Cs [Zhu et al.2010]. These results indicate that the ITS sequences of mutant Genotypes #2-#17 belong not to the genome of Genotype #1 Hs, but to the genomes of independent Os fungi. The 5.8S-F/R primers used for amplification of 5.8S transcripts [Li et al.2013] also called into questions. (i)The 5.8S-F/R are not Genotype #1 Hs-specific, and have similarities of 61.9%-100% and 31.8%-95.5% with the 17 genotype Os. (ii)Similarities between the 5.8S-F/R primers and the 5.8S gene of the 3 Hs genome sequences are 95.2% & 59.1%, respectively. (iii)Similarities between the 5.8S-F and Tolypocladium sinense(KX082970), Paecilomyces hepiali(EF555097), and Fusarium sp.(KJ735013) are equally high, 95.2%, while P. hepiali and Fusarium sp. were found in the multicellular heterokaryotic ascospores of Cs [Zhu et al.2016]. (vi)The steric conformational differences of 5.8S rRNA of Genotypes #1,#4,#5 [Li et al. 2013] may significantly affect the reverse-transcription of mutant Os 5.8S genes and the subsequent cloning and sequencing, while consistent failures have been reported [Gao et al.2011,2012]. Thus the detected 5.8S cDNA by Li et al. needs to be verified to truly belong to Genotype #1 Hs, and non-detection of the 5.8S cDNA of the mutant Os genotypes need to be confirmed with using different sets of primers, or different molecular approaches. In conclusion, [Li et al.2013] provides incomplete and controversial evidence to support their “ITS pseudogene” hypothesis, and encourages further studies. (Supported by Grant #2017-SF-118 from the Science-Technology Department of Qinghai Province)
Functional study on a non-ribosomal peptide synthetase in an entomopathogenic fungus Cordyceps bassiana
- J. Han
- M. Hyun
- J. Han
- J. Kim
- J. Oh
- B. Shrestha
- J. Cho
- S. Lee
- K. Lee
- W. Kong
- G. Sung
- C. Lee
Nonribosomal peptides (NRPs) are a group of fungal and microbial secondary metabolites with diverse properties as immunosuppressant, pigments, antibiotics, medicines, etc. NRPs are synthesized by nonribosomal peptide synthetase (NRPS) enzymes, which consist of repeated domains of adenylation, thiolation and condensation, and some modifying elements. Fungal NRPSs usually cooperate with several regulatory genes in gene cluster to produce a certain peptide. In most cases, however, their functions and biosynthesizing pathways have been still remained unknown due to their cryptic expression and recalcitrant genetics. Cordyceps bassiana is an ascomycetous fungus that parasites various arthropod species. It produces the NPRs such as beauvericin and bassianolide that are insecticidal virulence factor. Our research group analyzed the genomic data of C. bassina strain C101 to discover novel genes biosynthesizing useful compounds. Using fungiSMASH, the fungal version of antiSMASH, 35 NRPS-encoding genes and their gene clusters were found. Among them our study focused on understanding the function of Cbnrps6. We generated the Cbnrps6 gene deletion and over-expressed mutants, respectively, via Agrobacterium tumefaciens-mediated transformation (ATMT). And then the mutants were experimentally verified using qRT-PCR and Southern blotting. The 100-fold increase of Cbnrps6 gene expression was shown in the overexpression mutant. In comparative metabolic profiling between the n-butanol extracts of normal strain and two mutants, a unique peak which is suspected that of the final product of Cbnrps6 gene was detected at retention times of 25.1 min. Its chemical structure was determined using GC-MS and NMR analyses.
Exploring the transcriptome-methylome dynamics in Termitomyces mushroom
- F. Hsu
- H. Hsieh
- J. Lu
- Y. Lee
- P. Chen
- C. Lin
Termitomyces mushrooms are found to be superior to all other mushrooms globally because of their biting aroma and fullness of nutrients. In Taiwan, the fruiting body of Termitomyces eurhizus is a gourmet basidiomycetous mushroom to many people, produced through mutualistically symbiotic association with black-winged subterranean termite (Odontotermes formosanus). Until now, the fruiting body can only be acquired from the wild, but not with the cultivated conditions. While the mystery of fruiting in another basidiomycete Coprinopsis cinerea is well studied it is never clear with T. eurhizus, possibly because of the precise environmental conditions implying epigenetic co-regulation, confounded with a complicated symbiotic relationship with genotypically unique termite. To understand the fruiting dynamics and the life cycle of T. eurhizus, we profiled the transcriptome and genome-wide DNA methylation across the developmental stages of T. eurhizus. As a preliminary result, we found the genome of T. eurhizus is approximately 76M bp with DNA methylation mostly in CpG context; comparing to black truffle it is smaller and more methylated. We also found the fruiting tissues show different global methylation profiles different from the other tissues; suggesting DNA methylation may actually play a role in the fruiting formation. Lastly, we found the expression of genes and transposons are anti-correlated with their methylation levels; indicating epigenetic regulation in T. eurhizus. This is also different from truffle that the gene expression is independent of the methylation at gene body. Our study provides valuable insight into the epigenome regulation of Termitomyce with a great potential to uncover its mystery of fruiting.
Gene regulation changes during heterokaryosis in Agaricomycetes inferred from mRNA-Seq and methylome patterning
- R. Powers
- T. James
The process of heterokaryosis in the mushroom-forming fungi of the Agaricomycetes involves plasmogamy (fusion of cytoplasm) between two mating-compatible haploid mycelium (n). However, unlike most organisms, the fusion of nuclei (karyogamy) does not occur until immediately before meiosis, and much of the mushroom life cycle occurs as a heterokaryon (n+n), in which two unique nuclear genomes coexist in each cell, providing opportunities for conflict or cooperation between nuclei. While there is evidence that the two nuclei in a heterokaryon are actively communicating with each other to modulate gene expression, the genetic basis of gene regulation between nuclei during this important and long-lasting part of the life-cycle remains largely unknown. For this study, we looked at changes in gene regulation during the transition from two individual haploid mycelia to a heterokaryon using a combination of mRNA-Seq and whole-genome bisulfite sequencing (WGBS). Transcriptomes and genome-wide methylation profiles of two haploid homokaryotic isolates of five taxa (Coprinopsis cinerea, Heterobasidion irregulare, Wolfiporia cocos, Coprinellus disseminatus and Cyathus stercoreus) along with a mated heterokaryon for each strain were generated and correlated. Here, we report on the variation in gene regulation and methylation patterns both between homokaryotic and heterokaryotic isolates of the same species, as well at the general pattern of mRNA changes and methylome profiles during heterokaryosis across the species examined.
Uncovering the role of pseudouridylation in a fungal pathogen Cryptococcus neoformans
- S. Lee
- J. Kim
- Y. Bahn
Cryptococcus neoformans is an opportunistic fungal pathogen that causes cryptococcosis in both immunocompromised and immunocompetent individuals. Due to its clinical importance, revealing the factors that can affect its life cycle is critical. Among the various factors, pseudouridylation of RNA is the most abundant type of post-transcriptional modification. Pseudouridylases isomerize uridine into pseudouridine, therefore can affect the stability of RNA structure. In Saccharomyces cerevisiae, 8 proteins exist as stand-alone pseudouridylases, and each protein has specific pseudouridylation sites and roles. To discover the features of pseudouridylases, we aim to identify 6 putative pseudouridylases in C. neoformans. We sorted out the enzymes based on the database from FungiDB and NCBI. We used BLAST search with protein sequences to find out any corresponding orthologs in multiple organisms, such as S. cerevisiae, Candida albicans, Aspergillus fumigatus and Neurospora crassa. To characterize the function of pseudouridylases, we constructed 10 mutant strains representing 5 putative pseudouridylases and we examined their phenotypic traits under various conditions so far. By using pseudouridylation RNA sequencing, we will identify pseudouridylated RNA transcripts and characterize their role in pathogenicity of C. neoformans.
Complex multicellularity in fungi: in search of a minimal genetic toolkit of fruiting body development
- L. Nagy
- K. Krizsan
Mushroom formation is one of the most spectacular and complex processes in the fungal world, comparable to the development of higher plants and animals in terms of its complexity. Yet, its genetic bases, in particular, conserved developmental genetic events are hardly known. Here we set out to identify conserved developmentally regulated genes in mushroom-forming fungi. We identify developmentally regulated genes by comparing transcriptome data across three to eight developmental stages of six Agaricales, Polyporales and Hymenochaetales species. One of the species included in our analysis is Phanerochaete chrysosporium, which procudes simple, crust-like (resupinate) fruiting bodies, presumably resembling the ancestral fruiting body morphologies in the Agaricomycotina. We find that 10-40% of the genes are differentially regulated during fruiting body development in the examined species, comprising functions related to cell wall synthesis and modification, mRNA stability, cell growth and regulation of transcription. By studying the conservation of developmentally genes and their evolution through a comparative genomics analysis of 202 fungal genomes, we aim to understand the origin of fruiting body-related genes and to zoom in on the minimal gene set required to initiate and develop basidiomycete complex fruiting bodies.
Evolutionary transitions in fungal epigenomics
- S. Mondo
- I. Grigoriev
- R. O'Malley
With an estimated age of up to 1 billion years old, the Fungi have evolved a variety of diverse and creative ways to regulate gene activity. In addition to histone modifications, fungi utilize DNA modifications such as 5-methylcytosine (5mC) and 6-methyladenine (6mA). However, with few exceptions their utilization is taxonomically restricted. Recently, we discovered that the 6mA is highly abundant in early fungi, but is largely absent from Dikarya. 6mA plays a positive role in gene expression and is preferentially deposited at gene promoters based on their function. Interestingly, in the Dikarya and early-diverging fungi lacking 6mA, 5mC is the predominant modification. This indicates that in fungi these modifications cannot readily coexist with one-another. Similar to other studies, we found that 5mC was primarily restricted to repetitive sequence and was involved in transposon suppression. These findings suggest that the two modifications are near complete opposites, differing both in location and role in gene expression, raising interesting questions as to why this pattern has emerged. Here we explore evolutionary transitions in fungi that may have led to this pattern, including selective pressure from transposable elements and gene gain/loss. We have identified a variety of methyltransferases that are significantly different in their presence across fungi which may be involved in 6mA regulation. Lastly, we explore factors involved in nucleosome placement and how DNA modification act to manipulate them.