Welcome to IMC 2018 International Mycological Congress
- K. Shrivastava
Knowledge and use of medicinal and edible mushrooms of the Sierra Tarahumara of Chihuahua, Mexico.
- M. Quiñónez Martínez
- M. Olivas Sanchez
- J. Valero Galván
- R. Gonzalez Fernandez
- E. Rico Escobar
- A. Corral Avitia
- I. Enriquez Anchondo
Chihuahua is the largest state in Mexico. It takes 12.6% of the country’s area. The southwestern portion of the Sierra Madre Occidental in this state also known as Sierra Tarahumara named as it is occupied by an ethnic group known as the Raramuri or Tarahumara, which means “light footed people”. The territory consists of canyons and ravines with pine, oak and pine-oak forests in the higher plateaus. There is a great diversity in edible and medicinal mushrooms all-around of the Chihuahua state counties. Their residents are the only consumers of wild mushrooms in the Northern Mexico; they have a long tradition of collecting, using and eating them during the “rainy season.” However, despite the wide diversity of edible mushrooms that grow in these areas, residents have a selective preference. This paper aims to record evidence of the knowledge and use of wild potentially edible and medicinal mushroom species by inhabitants of towns in Bocoyna and Urique municipalities of Chihuahua, Mexico. In the forests of the Sierra, there are records of around 450 species of fungi; 50 of them with edible importance at nationwide and apparently only 16 fungi species of those 50 are being consumed by the inhabitants of the municipalities of Bocoyna and Urique with Amanita caesarea complex being the most preferred by mestizos and Raramuris. We observed no apparent differences in the population studied in terms of gender, occupation, or language, regarding the recognition and consumption of species; however, this is not conclusive and so it is important to continue with a greater number of such studies to check whether this knowledge and use is differential. Forty eight percent of the people surveyed reported to collect mushrooms directly from the field or forest areas while the others buy them either from the Raramuris who sell them on the side roads, or at their home as a result of door to door to selling. Seven percent mentioned the “Fungus Fair,” which is carried out every year in the month of August provides them with a good opportunity to buy mushrooms and reassures them they are edible. Three percent mentioned that besides selling them, they teach other people how to handle mushrooms in the place known as The Valley of the Mushrooms. As medicinal mushrooms, three species are used for the purpose of healing wounds of the skin, and to remove pimples in the face: Calvatia, Lycoperdon and Astraeus hygrometricus.
Edible mushrooms in Mizoram, India: Occurrence and perception in the region
- J. Zothanzama
- L. Hmar
- J. Vabeikhokhei
- Z. Zohmangaiha
- B. Held
- R. Blanchette
Mizoram is regarded as one of the biodiversity hotspots of the world. A diverse group of flora and fauna have been documented. Knowledge on the edible mushrooms is very limited in the region. Moreover, the local knowledge and perception on the edible species of the region is very low. Knowledge of edible mushrooms among the Mizo people has been there for a long time. However, it has been perceived that the number of edible species known by the people is very few. A study of the occurrence of the edible mushrooms growing in Mizoram was undertaken. From the study a total of 32 species of edible mushrooms was identified from different districts of Mizoram. Study on the local knowledge and perception on the edible mushrooms was also undertaken from different sections of the local communities. It is found that a small percentage of edible species are presently known to be edible by the Mizos despite the existence of other edible species and distribution in the region.
Determining seasonality and local abundance of edible mushrooms in Puerto Rico using social media as a tool
- K. Miller
- Y. Rivera-Torres
AbstractThe knowledge of edible mushrooms in Puerto Rico is limited at least in part due to cultural attitudes, which could be largely characterized as locally “mycophobic”. For the past three years, I collected information on the mushrooms that are found in the island, especially edible species in the genera Cantharellus, Calvatia, Lentinula, and Laetiporus. These genera are easy to identify based on macromorphological traits and therefore lend themselves well to analyzing photos uploaded by users of a Facebook group dedicated to local mushrooms. These data are supplemented by observations from mushroomobserver.org, as well as inventories on cybertruffle.org. The data are therefore a blend, ranging from local residents’ photos to established scientific inventories with a total of 150 observations between the four genera. The goal is to answer questions of seasonality, location and abundance of local edible mushrooms, the harvesting of which is of potentially significant commercial interest. The downfall to this “crowdsourced” data is, of course, demographics, as much of the population lives in the Northeast. It is therefore to be expected that a disproportionate number of observations will come from this region. Puerto Rico is divided into 78 municipalities, which is noted, along with the date and ID, for each observation. For each species of edible mushroom, the data are displayed showing where on the island and during which months a species is most likely encountered. The data shows that Cantharellus coccolobae occur mainly in coastal beaches, associated with Coccoloba uvifera, but they were also observed growing in karstic areas on the north coast of the island, in the presence of other Coccoloba species, such as C. diversifolia. Although C. coccolobae occurred year-round, they were especially abundant during the “rainy season” (April-November), a pattern repeated for all species treated here. Calvatia cyathiformis is the only recorded Calvatia species on the island. Its distribution is cosmopolitan within Puerto Rico, but the records show that it prefers grassy pastures and it has adapted well to the metropolitan area and its abundance of maintained lawns. Lentinula boryana, or Florida shiitake, proves exceedingly rare or at least is not well documented. This is most likely due to its occurrence in inland municipalities with late successional tropical forests, such as El Yunque; habitat which is not common on the island, and is at once sparsely populated and difficult to traverse, and more observations are needed for this species to determine its seasonality. Laetiporus caribaea is observed in similar habitats as Lentinula boryana, but are more frequently documented, which could be due to their striking orange-red fruiting bodies.
The knowledge of future teachers of science and biology in basic education about the fungi
- S. Xavier-Santos
- A. Persijn
Although mycology integrates a component of great relevance in basic education, being present in different moments of the national curricular guidelines or the reference curriculum of the states, recent studies show that this content has been neglected in the courses of teacher training for that level of education, in Brazil. The objective of this study was to investigate the knowledge about fungi among the final students of the Biological Sciences Degree courses in the state of Goiás, Brazil. We sampled 10 of the 32 courses existing in the state, so that the sample universe totaled 123 students, belonging to the last period of these courses. The data collection was done from a semi-structured questionnaire applied to the participants. The analysis of the data shows that most of the future teachers have less knowledge than they should have, lacking in depth, with conceptual inaccuracies, presenting difficulty in developing critical and logical reasoning in formulating answers, besides they showing an anthropocentric view, in which fungi are at the service of the human species, with little attention to their interactions and their ecological role. The best performance was observed among students from courses that offer specific discipline for Mycology content. These frailties are incompatible with an efficient scientific training. Since the quality of the teaching in basic education is intrinsically linked to the quality of the training of the teachers who work in it, the courses in Biological Sciences require special attention in regard to the approach to this subject, in order to promote the consolidation of mycological knowledge among the educators they make available to society.
Cultivation of Polyporus squamosus on substrates from residues of wood processing industries in Finland
- M. Cortina Escribano
- P. Veteli
- R. Linnakoski
- V. Möttonen
- H. Vanhanen
Demand for non-meat protein sources has grown extensively in Finland as well as in rest of Europe. In Finland wood processing industries produces vast amounts of residues, which could be suitable for production of edible mushrooms thus increasing the local protein production as well as harboring circular economy. Our aim in this study was to test cultivation of Polyporus squamosus on substrates from wood processing industries, and evaluate the feasibility of cultivation. Populus tremula and Betula pendula sawdust and chips were used as substrates. Two strains of P. squamosus originating from Finland were used in this study. The substrate bags of 1 kg dry weight were filled as follows: 1) 40% of wood sawdust and 60% of wood chips in the case of P. tremula species 2) B. pendula substrate bags were filled with 100% wood chips 3) Used coffee and rye bran were tested as nutritional additives, 20% of the total substrate bag weight for both nutrients and wood chips from the two species. A total of ten replicates per each substrate formula were inoculated with 150 ml of two strains of P. squamosus barley spawn. When the substrate bags were completely colonized, they were kept at 26°C with a relative humidity of 80% to support fruiting body formation. The biological efficiency was calculated considering the kilograms of fresh fruit body per kilogram of dry substrate. First flush was first observed one month after inoculation, suggesting that crops can be obtained within 2 months. The substrate formula that was found to be the most suitable for fruit body formation was the one containing 20% of rye addition in both B. pendula and P. tremula wood residues. Differences between mycelium growth rates were observed between the fungal strains, emphasizing the importance for strain selection most suitable for commercial cultivation in future. Our results suggest that B. pendula and P. tremula wood residues serve as equally suitable substrates which can be utilized for the cultivation of P. squamosus.
Pleurotus pulmonarius production optimized on oil palm bunch
- N. Chinwendu
This study was conducted to study the yield and some macro-morphological characters of Pleurotus pulmonarius fruit bodies cultivated on Hydrochloric acid (HCl) optimized oil palm bunch (OPB) substrate. Concentrated HCl was diluted in tap water at 0.1%, 0.2%, 0.3%, 0.4% and were used to induce changes on the initial pH (9.5) of OPB to 8.9, 8.2, 7.9, 6.2 and control (9.1) respectively; after soaking for 48hrs. One way Analysis of variance (ANOVA) and Correlation test were adopted for data analysis. Mean separation was also done by Duncan Multiple Range Test (DMRT) at probability level of 5%. Results showed that 0.1%, 0.2%, 0.3% and 0.4% HCl treated OPB substrates produced P. pulmonarius primordia after 9, 9, 10, 11 and control (12days) respectively. Results further revealed that 0.4% HCl treated OPB substrate induced the highest (900g/kg) fruit body yield and Biological Efficiency (90%) while control (493g/kg and B.E 49.3%) respectively, produced the lowest quantity of fruit bodies. Some macro-morphological characters of harvested fruit bodies revealed that mean cap size (C.Scm) and Weight (wt.g/kg) of fruit bodies were highest (3.83cm and 3.5g/kg) in 0.4% HCl treated OPB respectively. Mean Stipe Length (S.Lcm) was highest (2.77cm) in 0.3% OPB substrate and was significant at p ≤ 0.05. S.L and C.S of fruit bodies as well as C.S and Wt. were significantly correlated while there was no correlation between S.L and Wt. of fruit bodies. HCl was found as a suitable acid buffer for the optimization of the pH of the highly alkaline OPB for cultivation of P. pulmonarius fruit bodies. Oil palm bunch should therefore be adopted in the commercial production of the Oyster mushroom if certified safe for human consumption.
Utilization of Amazonian waste wood for the production of the edible mushroom Pleurotus ostreatus and Lentinus strigosus in the Amazon
- F. Pessoa
- C. Sales-Campos
- L. Rolim
Found in the Amazon, Pleurotus ostreatus and Lentinus strigosus are not widely cultivated. Like other edible mushrooms, the use of these fungi to convert regional waste is of little cost and is recommended. They are able to grow in the wood waste of Simaboura amara (marupá) and Anacardium gigantium (cajuí), supplemented with a mixture of bran: 75% rice (Oryza spp.), 20% wheat (Triticum spp.) and 5% corn (Zea mays). The following formulation was made for the two substrate of wood waste: sawdust (68%), mixture of bran (30%) and calcium carbonate (2%), the same being homogenized and humidified to 75%. All material was kept in a growth chamber in the dark at a constant temperature of 25°C and humidity of 80% until full establishment. Then, a photoperiod of eight hours created the stimulating condition for the production of the mushroom. The following were main areas of evaluation: biological efficiency (BE,%), yield (g kg-1) and organic matter loss (OML%). The results showed that the formulation made with cajuí obtained better results with EB at 221.17% (P. ostreatus) and 104.88% (L. strigosus); Yield: an average of 220.23g kg-1 (P. ostreatus) and an average of 72.5g kg-1 (L. strigosus); OML was not significantly different for the two types of substrates used. The cultivation of P. ostreatus and L. strigosus reveals that this is a promising raw material due to its great local availability at low to no cost. In addition, production of the mushroom reduces pollution of the environment.
DNA-based identification of consumer-relevant mushrooms: A partial solution for product certification?
- H. Raja
- T. Baker
- J. Little
- N. Oberlies
Attributing the correct scientific name to dietary ingredients made from fungal materials remains a challenge, in part due to difficulties in species authentication by chemical means and the nature of fungal taxonomic names, which are undergoing numerous taxonomic revisions with the application of molecular methods. This can be particularly difficult for samples that contain fungal mycelia, where morphological characteristics do not present sufficient variation to differentiate species. This challenge is compounded by the fact that many of those materials maybe heavily processed, including drying, milling, and even extraction, prior to analysis. However, monitoring the safety and quality of such products is a requirement for the protection of consumer health. The main goal of the study, which was performed as a collaboration of academic researchers (University of North Carolina at Greensboro) and industry scientists (Procter & Gamble), was to demonstrate that Sanger sequencing of the ITS region is an appropriate means for verification of species identities. We generated ITS barcodes for 33 representative fungi, which are being used by consumers for food and dietary supplement purposes. After generating ITS barcodes utilizing standard procedures accepted by the Consortium for the Barcode of Life, we tested the utility of the ITS by performing a BLAST search against NCBI GenBank. In some cases, we also downloaded published, homologous sequences of the ITS region of fungi inspected in this study and examined the phylogenetic relationships of barcoded fungal species in light of modern taxonomic and phylogenetic studies. In the majority of cases, we were able to sequence the ITS region from powdered mycelium samples, grocery store mushrooms, and capsules from commercial dietary supplements. Results demonstrated that the ITS region was able to identify the mushrooms used in the present study to species-level. We anticipate that these data will motivate a discussion on DNA based species identification, particularly as it applies to the verification/certification of fungal containing products.
Characteristics of a new cultivar Grifola frondosa "Daebak" with bottle cultivation
- L. Yunhae
- J. Dae-Hoon
- C. Jong-In
- G. Hee-Min
- H. Hye-Jeong
- J. Kab-Yeul
We aim to introduce a new cultivar of Grifola frondosa by crossing of mono-spore. The name of this cultivar is ‘Daebak’, it means jackpot. As in the case of control cultivar ‘Cham’, temperature of mycelial growth and fruiting of the ‘Daebak’ were also same at 25℃ and 18℃, respectively. The incubation period was 57days, two days shorter than that of the ‘Cham’ by bottle cultivation. The rate of fruiting for the ‘Daebak’ was 98.4%, which was 24.8%p higher than that of the 'Cham'. In addition, the coefficient of variation for the ‘Daebak’ was 0.6, which was lower than the ‘Cham’ 5.3, resulting uniform fruiting. The L-value of pileus for the ‘Daebak’ was lower than that of the ‘Cham’. The diameter of pileus and length of stem for the ‘Daebak’ were larger and higher than those of the ‘Cham’, respectively. Physical properties (strength, springness, and brittleness) of this cultivar were lower than those of the 'Cham'. The fresh weight of this cultivar was 139g/1,100㎖ and was 28% higher than that of the ‘Cham’. Additionally the new cultivar has greater uniformity due to the coefficient of variation in the quantity being lower than the ‘Cham’. Shelf life of this cultivar at 4 ℃ was 42 days and 6 days longer than that of the 'Cham'. In conclusion, new cultivar ‘Daebak’ of Grifola frondosa was in quantity, quality and storage compared to the previous cultivar ‘Cham’, but also need to be bred with a more physically strong cultivar for the future.
Analysis of nutritional and neutraceutical properties of selected wild-grown mushrooms of Nepal.
- J. Upadhyaya
Mushrooms are the fleshy spore-bearing fruiting bodies of fungi. Wild mushrooms are source of many different nutraceuticals such as unsatured fatty acids, phenolic compounds, tocopherols, ascorbic acid, carotenoids and alkaloids and nutrients such as proteins, fats, ash, fiber, moisture and carbohydrates. Nepal possesses diverse phytogeographical zones related to altitude and other factors, and rich in wild mushrooms. The information and knowledge about nutritional and nutraceutical values of wild mushrooms is limited and poor. Therefore, the present study is undertaken to document the use of wild edible mushrooms and analyze their nutritive values. Herein, it was reported and compared the nutritional value and neutraceutical values of the wild mushroom species; Laetiporus sulphureus, Polypore sp., Trametes elegans , Trichaptum biforme, Lenzites betulina, Stereum complicatum, Trametes versicolor, Trichaptum subchartaceum, and Ganoderma Lucidium. The nutritional and nutraceutical properties analyzed according Association of Official Analytical Chemists (AOAC) and spectrophotometrically respectively. These mushrooms (samples) were rich in proteins (6.8- 60.23%) and fibers were range from 0.174 - 36.38% and contained fat range from 3.642- 14.6%. The carbohydrate contents ranged from 7.058 to 59% (on the basis of dry weight). Similarly; ash content and moisture content ranges from 10- 19% and10-16% respectively. The protein content was highest in Ganoderma Lucidium. (G. lucidium) and lowest in Trametes elegans (T. elegans). The fat content was highest in L. sulphureu and lowest in G. lucidium. . The analysis revealed that the total phenolic contents ranged from 3.95 to 10.05 mg ml-1. Similarly, the total flavonoid contents ranged from 2.149 to 11.36 mg ml-1. The result indicated the high levels of antioxidants activity thus making mushrooms suitable to be used as functional foods or nutraceutical sources. Therefore, this study provides new information regarding chemical properties of wild mushrooms, which is very important for the biodiversity characterization.
Productivity of edible Amanita at Phusing Agricultural Development Center, Sisaket, Thailand
- U. Pinruan
- S. Somrithipol
- S. Sommai
- P. Khamsuntorn
- T. Boonpratuang
- S. Takhiankling
- K. Nissa
The cultivation of edible ectomycorrhizal mushrooms associated with forest trees is becoming popular in Thailand. It is currently applied to reforestation projects by forestry officials, and in agroforestry situations by farmers. However, information on the mushroom productivity and sustainability is unavailable. This study investigated and reports on the yield of edible Amanita and other wild edible mushrooms in a 1280m2 plot at Phusing Agriculture Extension Center, Sisaket Province between 2014 and 2016. The plot was on an area mainly covered by Dipterocarpus alatus trees, which were planted on bare land in 2003 and were inoculated with Amanita during 2004-2005. In 2014 eight edible mushroom species (total weight 72.6 kg) were found in the plot. Three Amanita spp. (60.2 kg, 83%) were the dominant group: red Amanita (Amanita cf. hemibapha; 43.2 kg, 59%), yellow Amanita (Amanita cf. hemibapha; 15.7 kg, 22%), and white Amanita (Amanita cf. princeps; 1.3 kg, 2%). These Amanita species morphologically resemble Amanita hemibapha, and A. princeps, but molecular data based on ITS and LSU show that they are taxonomically new to science, and are currently being described. Other inferior mushrooms found in the plot were Russula nigricans (8.1 kg), Termitomyces microcarpus (2.4 kg), Lactarius sp. (1.5 kg), and Russula emetica and Russula sp. (less than 1 kg). In 2015, two additional mushroom species, Russula virescens and Termitomyces sp., were also found, but the total yield of the plot was stable (72.1 kg). Yellow and white Amanita increased their yields (40 kg, 56% and 6.3 kg, 9%) but red Amanita sharply decreased (2.5 kg, 3%) resulting in decline of the total Amanita yield (48.8 kg, 68%). In 2016, nine edible ectomycorrhizal mushrooms were found with the total yield 73.6 kg. Russula nigricans became the dominant species (38.4 kg, 52%) while the Amanita group (31.6 kg, 43%) decreased: yellow Amanita (25.9 kg, 35%), white Amanita (3.6 kg, 5%), and red Amanita (2.1 kg, 3%). In 2017, seven edible mushrooms were found with the total yield only 17.1 kg. Yellow Amanita became the dominant species (6.7 kg, 39%), red Amanita (4.5 kg, 26%), white Amanita (3.0 kg, 17%), while Russula nigricans decreased (0.8 kg, 5%). The presentation will show monthly yields of each mushroom species and some environmental factors throughout the last four years. Declining trend of the Amanita productivity, as well as possibility of rehabilitation in correlation of environmental factors, will be also discussed.
Mushroom poisonings in South China and study on the toxin genes of lethal Amanita
- W. Deng
- T. Li
Poisonous mushrooms are the main factor causing the fatal disasters in food poisoning incidents in China. According to the statistical data of Chinese Center for Disease Control and Prevention, the numbers of deaths from mushroom poisonings were 25-55% of the total numbers of deaths from food poisonings from 2004 to 2014. South China is a high risk area for mushroom poisonings, the authors investigated and analyzed 113 mushroom poisoning cases in South China from 2000 to 2014, which involved 325 patients and 52 deaths, with an overall mortality of 16 %. About 200 poisonous mushrooms have been reported from South China. More than 50% poisoning cases were caused by Amanita exitialis Z.L. Yang & T. H. Li and Chlorophyllum molybdites (Meyer : Fr.) Mass., but all the poisoned deaths were caused by lethal Amanita. Cyclopeptides are the main fatal toxins in lethal Amanita, which encoded by MSDIN family. Based on the transcriptome and genome sequencing with Illumina HiSeq 2000, the author studied the toxin gene family and POPB involved in the toxin biosynthesis of the lethal Amanita from South China. The results showed that 70 different toxic gene family members were obtained from three lethal Amanita species, which encoded 3 toxic peptides (α-amanitin, β-amanitin and phallacidin) and 45 new unknown peptides. The research showed that lethal Amanita can produce abundant toxic peptides and related peptides, which will lay a strong foundation for the toxic peptide gene expression and exploitation of new cyclopeptide resources. This work was funded by grants from the National Natural Science Foundation of China (Nos. 31470155 and 31670018).
A study of the biodiversity and secondary metabolites of fungal endophytes from medicinal plants in Guizhou
- J. Kang
- T. Wen
- Y. Qian
- S. Zhou
- X. Ma
In southwest China, Guizhou province has a well-known fame for its prolific biodiversity of medicinal plants. For solving the problems of plant shortage, environmental disruption and obtaining novel structural as well as bio-active natural products, we have been studying the fungal endophytes and their secondary metabolites from medicinal plants in Guizhou province for a number of years. We isolated over five thousand fungal strains from medicinal plants including Artemisia carvifolia, Artemisia japonica, Blumea balsamifera, Camptotheca acuminate, Dendrobium orchids, Ginkgo biloba, Nothapodytes pittosporoides, Reineckia carnea, Taxus brevifolia. Dozens even hundreds of different fungal isolates were obtained from each plant species. Most of the fungal endophytes belong to Ascomycota which mainly distribute in Pezizomycete, Dothediomycete and Sordariomycete. A few of them are classified into Basidiomycota and Zygomycota. Fungal endophyte taxa were subjected to vary kinds of factors, for example, the age of host, organ, humidity of sample site, altitude, season, surrounding plant, extent of environmental contamination and so on. We combined bio-activity analysis in vitro, chemical composition identification and gene detection in fungal endophyte to screen and evaluate 1003 fungal strains. Fifty-six of them showed anti-inflammatory, antitumor, antioxidant, anti-pathogen and P-gp inhibitory bio-activity to different extent. At present, we accomplished the study for secondary metabolites of ten fungal strains that possess high bio-activity. More than 200 bio-active compounds were isolated. Ten of them have new structures. However, there is a long way to produce bio-active compounds in large scale because of the low production. Therefore, it is necessary to improve cultivation methods or alter inner gene in fungal endophyte by genetic engineering for achieving novel chemical structure as potential new drug.
Diversity of Medicinal Mushrooms of South India
- M. Kumar
AbstractTropics are considered as rich repositories of mushroom diversity and most of the new mushrooms reported in recent years are from tropics (Hawksworth, 2001). India is one such tropical country with diverse ecological characteristics for species richness. In India, the survey of literature indicates a total number of 1,160 species are only described in these 2 orders viz., Agaricales and Boletales until now (Manjula, 1983, Lakhanpal 1995 and Natarajan et al., 2005).
Basidiomycetes especially mushrooms are unlimited sources of biologically active compounds. There are over 700 species of higher basidiomycetes that have been found to possess significant pharmacological activities (Wasser, 2002). Studies on medicinal mushrooms have exponentially increasing in the last two decades. Attempts have been made in many parts of the world to explore the use of mushrooms and their metabolites for the treatment of a variety of human ailments (Jong and Birmingham, 1992). In the present study, biodiversity of medicinal mushroom from South India especially in the Eastern Ghats region was taken up in order to fill up the lacuna. The study resulted in documentation of many medicinal mushrooms from these region which include Ganoderma lucidum, Phellinus badius, Gymopilus dilepis, Gymnopilus palmata, Lentinus tuberregium, Calocybe Indica, Pleurotus ostreatus, L. squarrosulus, L. cladopus, Mycena pura, Macrolepiota rhacodes, Termitomyces microcarpa, Termitomyces eurrhizus, Auricularia polytricha and so on and evaluation of few.
Distribution and ethnomycology of wild edible fungi of capital complex area of Arunachal Pradesh, India
- K. Shrivastava
- K. Sherpa
AbstractWild Edible Fungi (WEM) or macrofungi are central components of forest ecosystems that influence the other life forms. The mapping of macrofungi is essential component of fungal diversity research that help obtaining baseline information and distribution records needed for the assessment of changes in biological diversity. Arunachal Pradesh, the Indian northeastern most state, is one amongst world’s top 35 global biodiversity hotspots and an important sub-center for the origin of wide variety of mushrooms. In this paper, results of macrofungi collected from four study areas of capital complex of Itanagar, Papum pare district of Arunachal Pradesh, India during winter and spring season of 2015-16 are presented and discussed. In all, 641 macrofungal fruiting bodied were collected during the course of investigation and identified according to their morphological, microscopic and spore prints features. They were categorized under 60 species, 45 genera, 22 families and 7 orders. The Phylum Basidiomycota representing 58 species while Phylum Ascomycota and Myxomycota with 1 species each. Polyporales and Agaricales were the two most dominant orders reported with maximum number of species (26 and 22 respectively) and Polyporaceae was most dominant family (19 species). Family Polyporaceae also contributed maximum (19%) followed by Auriculariaceae, Hymenchaetaceae and Tricholomataceae (4% each) and Marasmiaceae (3%). The species Pycnoporus cinnabarinus was reported as most dominant species. The maximum frequency of occurrence was exhibited by Trametes versicolor and Schizophyllum commune (83.33%), while maximum density was recorded for Schizophyllum commune (126.67%) followed by Trametes versicolor (120%) and Xylaria polymorpha (93.33%). To document the ethnic acquaintance indigenous beliefs and uses of WEM of Papum pare district, ethonomycological information was collected through questionnaire survey and personal interviews of local knowledge holders. The Papum pare district is dominated by Nyshing community who use 16 species of wild mushrooms (15 edibles and 1 poisonous) as food. WEM are commonly called here as ‘Tayin’ however they are also named as Ikhnuro, Indhi, Lolam tayin, Pukhaso, Rubhj tayin, Inya, Huhiun etc. for specific WEM. Aside from utilizing mushrooms as food, a few species are also used as medicine. The local name of species used in medicine is Heimey Yoru which could not be identified but especially used in the treatment of blood pressure. In a nutshell, this survey revealed the highest number of macrofungi consumed to be of family Agaricaceae, followed by Tricholomataceae, and Russulaceae. The current data can pave the way for future research work and also make people aware of the many varieties of wild edible fungi available in the state.
Salt stress on Ganoderma lucidum: morphological, physiological and biochemistry aspects
- M. Yarina
- L. Krasnopolskaya
- A. Usov
- A. Marakhonov
- Y. Shipilov
Ganoderma lucidum strains are able to produce biologically active polysaccharides. It is widely known that stresses are inducing alteration of metabolome. However, the salt stress effect on G. lucidum has not been studied yet. The purpose of this work was to assess the impact of sodium chloride stress on the mycelia G. lucidum growth and polysaccharide production. We have examined several strains from our samples for the main work objective determination. An identification of ITS rDNA sequences has been conducted with a purpose of specifying the taxonomic position of four strains. For further work we selected G. lucidum strains capable of forming an alkali-soluble and water-soluble polysaccharides with high antitumor activity and capability to cytokines induction. Preclinical trials of an alkali-soluble highly branched xylomannan are in progress now.NaCl effect on the growth of G. lucidum was studied during its introduction into the dense and liquid nutrient culture medium in an amount of 0.5, 1.0, 1.5 and 2.0%. The sharp reduction of fungus colony diameter was observed in medium containing 1.0% or more of sodium chloride. When we added 2.0% NaCl to the dense medium G. lucidum colonies diameter was 21.4% of that of the control. IC50 NaCl estimated concentration was 1.45%.When concentration of chloride in the liquid medium was from 0.5 to 2.0%, we observed the gradual decrease G. lucidum biomass from 13.3 to 76.5%, respectively. The process was linear. Estimated concentration of the IC50 NaCl for submerged cultivation was 1.48%. In medium containing 12.5 g/l of sodium chloride and above, we observed the formation of brown pigment.The total polysaccharides in G. lucidum mycelium have been gradually reduced to 84.6% with 1,0% NaCl, and then have been increased to 171, 6% with 2,0% NaCl to the control. We assessed the changes of the content of monosaccharides as part of studied polysaccharides. Increase of sodium chloride concentrations in the liquid medium to 1.0% has been accompanied by an increase of the total protein in the mycelium to 167.2% compared with that in the control. Further increase of NaCl in the liquid medium to 2.0% was accompanied by a gradual decrease in protein up to 137.7 % to the benchmark. We used the method of light micrsoscopy and method of scanning electron microscopy to reveal micromorphological changes of immersed G. lucidum mycelium when it is grown in medium containing NaCl. In particular, we observed «ball-like» structures, an increase of chlamydospores and hyphae crystals with increase of sodium chloride concentration.
Biodiversity and enzymes bioprospection of Antarctic filamentous fungi
- M. Martorell
- L. Ruberto
- W. Mac Cormack
- L. Castellanos De Figueroa
Antarctica is one of the most suitable places for the bioprospecting of psychrotrophic fungi. The aim of this study was to investigate the diversity of filamentous fungi from 25 De Mayo Island, Antarctica and their ability to produce extracellular hydrolytic enzymes at low temperature. A total of 51 fungi isolates were obtained from 31 different samples. We identified twelve different genera, seven taxa belonged to the Ascomycota phylum (Cadophora, Helotiales, Monographella, Oidodendron, Penicillium, Phialocephala, Phialophora, Phoma and Pseudogymnoascus), one taxa to the Basidiomycota phylum (Irpex) and two taxa to the Mucoromycota phylum (Mortierella and Mucor). Some taxa not previously reported in Antarctica, as Monographella lycopodina, Mucor zonatus and Penicillium kojigenum, were identify. Nine isolates could not be identified to genus level, and could be representing novel species. Most of the fungi were psychrotrophic (76.5%) rather than psychrophilc. Nevertheless, only five isolates were able to grow at 35ºC, and the optimal temperature for growth was 15ºC for 65% of the fungal isolates. Results from enzymes production (amylase, cellulase, xylanase, lipase, esterase, laccase, protease) at low temperatures revealed that the Antarctic environment contains metabolically diverse cultivable fungi, which represent potential tools for biotechnological applications in cold regions.
Bioremediation of contaminated land by autochtonous fungi: Life-Biorest strategy
- F. Spina
- G. Spini
- A. Poli
- A. Zanellati
- T. Reigner
- A. Blieux
- A. Echairi
- E. Puglisi
- J. Ortega-Calvo
- G. Varese
Soil degradation is a serious issue in the European Union, causing the loss of more than 340,000 areas. LIFE BIOREST (LIFE15 ENV/IT/000396, www.lifebiorest.com) is a UE funded project in the framework of the LIFE Project, aimed to treat a soil contaminated by PHAs, BTEX and alkanes. This site (about 80,000 m2 wide) is located in Italy (Fidenza, Emilia Romagna) and has a long history of industrial exploitation. The project aimed to optimize a biomerediation method where the transformation made by consortia of fungi and bacteria is finalized by the final step of re-vegetation. The first phase of the project is indeed focused to characterize the microbial community that naturally populate this extreme environment and isolate those microbes capable of growing in the presence of pollutants as sole C source. The best performing strains will be used to set up consortia working in microcosms and mesocosms before up-scaling the process at in-situ level (biopiles). A solid screening and a liquid enrichment using few selected contaminants (naphthalene, pyrene, phenanthrene, benzene, alkanes and oil extracted from the soil) were carried out to identify the strains with the best adaptation and degradation skills. Despite the strong contamination, microbial communities was consistently developed: more than 220 fungi belonging to 70 species have been identified. Most of the fungal strains belonged to Ascomycetes (mainly to the genera Aspergillus, Cladosporium, Fusarium and Scedosporium ) even though almost 20 Basidiomycetes were also isolated. A further screening was based on an innovative miniaturized approach in 96 multiwell plates in order to evaluate the growth rate of each strain in the presence of 6 contaminants. During the 3 weeks experiments, several strains were capable of growing on the pollutants (at 200 ppm and 1% v/v) as much as positive controls with glucose, highlighting their capability to exploit complex source of nourishment as far as simple and bioavailable ones. Since the bioavailability of organic pollutants in soil is a recognized issue that often limit the efficiency of bioremediation approaches, strains were also screened for their capability to produce biosurfactants. Some fungi were found capable of producing extracellular broths with both emulsifier and biosurfactants activity. Almost 30 fungi and 30 bacteria have been selected and will be tested in micro and mesocosms singly and in consortia with selected bacteria in order to evaluate also their capability to grow and colonize the contaminated soil, and ultimately decontaminate it within 3 months treatment. According to the degradation skills, one consortia was selected for biopile trials. Fungi demonstrated that they could be successfully coupled with bacteria and plants to sensibly reduce the environment hazard of contaminated land, ultimately restoring their ecological functions.
Biodegradation of BTEX by fungi isolated from the hypersaline lagoon Las Salinas, Punta Cuchara Natural Reserve, Ponce, Puerto Rico
- J. Acevedo
- J. Pérez-Jiménez
Benzene, toluene, ethylbenzene, and the isomers of xylene (BTEX) are volatile anthropogenic pollutants derived from petroleum products that cause harmful effects in humans and other organisms. Fungi were isolated from a 35-ha hypersaline coastal lagoon, Punta Cuchara Natural Reserve, Ponce, Puerto Rico, which is an important nursery for marine species and serves as an avian refuge. Coastal lagoons frequently are contaminated with BTEX. A total of 25 culture-dependent fungal species from the lagoon included the following: Aspergillus sp., Penicillium sp., Cladosporium sp., Trichoderma sp. Fusarium sp., Curvularia sp., Chaetomium sp., Blastoschizomyces capitatus, Candida albicans, C. glabrata, C. rugosa, C. parapsilosis, C. tropicalis, C. zeylanoides, Cryptococcus neoformans, C. albidus, and C. uniguttulatus, Geotrichum sp., Kluyveromyces sp., Prototheca zopfii, Rhodotorula minuta, R. mucilaginosa, Saccharomyces cerevisiae, Trichosporon cutaneum, and Yarrowia lipolytica. A static culture system, each consisting of a serum bottle capped with Teflon Mininert™ valves was used in each of three successive trials to determine degradation of BTEX by each fungi species. An aqueous mineral medium stock solution was prepared with NaNO3 (3.0 g/L), KCl (0.5 g/L), MgSO4 (0.5 g/L), FeSO4 (0.01 g/L), and K2HPO4 (1.0 g/L). Each fungal species was inoculated at a concentration of 1 x 104 yeast cells/mL into the static culture system with a mixture of 118.75 mL of the mineral stock solution and 6.25 mL of BTEX. The BTEX was the only source of energy and carbon. The inoculation was incubated for 150 hr at 25oC. HPLC-DAD method was used to determine the biodegradation of the BTEX in each system by each fungus. The majority of the fungi species degraded the BTEX completely, although traces of BTEX were detected for some species. Thus, funguses species appear to serve as biodegradation organisms in hypersaline lagoons, many of which could be contaminated from petroleum spills.
Screening macrofungi for antibacterial compounds
- S. Qaraghuli
- M. Brown
- U. Streher
- L. Burgoyne
- D. Catcheside
The widespread development of resistance to antibiotics amongst bacteria pathogenic to humans has led to interest in finding new antimicrobials. We hypothesised that Macrofungi are a likely source of novel antibiotics since their mode of nutrition makes them vulnerable to competition. Amongst the macrofungi, saprotrophs export enzymes to digest macromolecules and must compete with bacteria for the breakdown products and ectomycorrhizal species need to protect the zone of nutrient exchange with plant roots. Accordingly, the production of antimicrobial compounds to control competition for nutrients would provide a competitive advantage to macrofungi. To test our hypothesis, extracts of 170 species have been screened for antibiotics including compounds that block the bacterial efflux pumps that expel antibiotics and compounds that impede the formation of bacterial biofilms. We have screened fruiting bodies and, where possible, also cultures for compounds active against a panel of sixteen bacterial species, including the ESKAPE group of pathogens that cause troublesome hospital acquired infections. We found that a substantial proportion of Australian macrofungi screened do produce antibacterial compounds, including some that inhibit the formation of biofilms or a Staphylococcus aureus efflux pump. One of the new antibacterial compounds identified is non-toxic to human tissue cultures and is amenable to synthesis, offering the possibility of a new family of antibiotics
Neuroprotective metabolites from Hericium
- Z. Rupcic
- K. Wittstein
- M. Rascher
- S. Kanaki
- R. Köster
- M. Stadler
Antibiotic resistance and neurodegenerative diseases are two major medical issues we have to face and which will become even more serious in the future. In order to cope with them and to develop improved therapies new classes of bioactive natural products with different modes of action are needed urgently. Hericium spp. of the phylum basidiomycota are among the most praised medicinal and edible mushrooms, and they have been known to produce secondary metabolites, such as hericenones and erinacines, which were isolated from the fruiting bodies and cultured mycelium, respectively. Many of these compounds were found to promote nerve growth factor (NGF) biosynthesis. Recently, corallocin A-C were isolated from basidiomes of the species Hericium coralloides and were the first members of this compound family that were found to be able to modulate both, NGF and brain-derived neurotrophic factor (BDNF) production. This prompted us to extend our studies to the metabolites from cultures of Hericium species, where metabolite profiles of a strain of the Lion’s Mane mushroom (Hericium erinaceus) and a strain of the rare species, Hericium flagellum (synonym H. alpestre) were examined. Highly similar metabolites were observed in both strains, with cyathane diterpenoids being the predominant ones. Seven metabolites obtained from H. erinaceus and H. flagellum were evaluated regarding their neurotrophin inducing effects. Although none of the tested compounds showed intrinsic neurothrophic activity, erinacines A, B, C, CJ14.258 and the new derivative Z1 clearly enhanced the neurotrophin production in astrocytic cells. Moreover, for the first time we observed a promoting effect of cyathane diterpenoid derivatives on BDNF expression. As they are able to stimulate the transcription of both neurotrophins, they may act upstream on a common molecular target of both pathways or via a third independent pathway.
Investigation of neurite stimulatory properties of Hericium erinaues (Monkey’s head mushroom) on rat pheochromocytoma cells (PC12 cells)
- Y. Tan
- C. Lim
- K. Wong
- V. Sabaratnam
Dementia is a neurodegenerative disease that has become one of the major issues in the 21th century. However, till date there is no specific treatment that can cure or prevent dementia from occurring in humans. Commercial drugs for dementia can only reduce the symptoms of the disease and even worse long term consumptions might pose adverse effects to other bodily functions. Therefore, this study aims to investigate the potential of Hericium erinaceus extracts to stimulate neurite outgrowth in PC12 cells. Briefly, fresh H. erinaceus was cut to cube, freeze dried and extracted with 20% w/v of 95% ethanol and overnight macerated with deionized water followed by 30min hot water extraction respectively. The crude extracts were rotary evaporated and freeze dried before the assay. PC12 cells were treated with 20ug/ml, 40ug/ml, 60 ug/ml and 80ug/ml of the extracts respectively. Neurite bearing score ranged between 2.5-29.6% with the highest score being 50ng/ml NGF treated cells. Pearson’s correlation showed positive correlation between extract concentration and neurite bearing score. Immunofluorescence assay was performed to confirm the neurite outgrowth via staining with rabbit anti-neurofilament 200 polyclonal antibody. The RNAs of the treated cells were also extracted to check the expression level of neurite outgrowth related gene, neuritin via qPCR. This would be the first report of neuritin gene expression following neurite outgrowth induced by mushroom extracts. Further test will be conducted to investigate the effect of co-incubation of H. erinaceus extract with 5ng/ml of NGF. In conclusion, H. erinaceus may be applied as potential health and functional food source in management of neuronal related diseases.
Ganoderma lucidum as a promising source of anti-virals against non-enveloped enteroviruses
- D. Reshamwala
- R. Linnakoski
- J. Hellström
- P. Veteli
- M. Cortina Escribano
- H. Vanhanen
- V. Marjomäki
Enterovirus infections are amongst the most common infections affecting people worldwide. These viruses cause severe outbreaks especially among children, with symptoms varying from mild to severe including aseptic meningitis, heart muscle damage and paralysis. They have recently been revealed to contribute to chronic diseases, such as type 1 diabetes, as well as with cardiomyopathies and atherosclerosis. Despite efforts of developing anti-virals against enteroviruses that have been going on for years, no vaccines (except for poliovirus) and drugs have made it past the clinical phase and into the market. So far, antiviral effects of extracts obtained from fruiting bodies and/or mycelia in liquid cultures (fermentation) have been described from several mushroom species, of which Ganoderma lucidium is one of the most extensively studied. Crude extracts as well as fractioned compounds including triterpenes and ganoderic acids from G. lucidum have been noted to exhibit inhibitory effects against enterovirus 71 (EV71), herpes simplex virus type 1 and 2 (HSV-1 & 2), as well hepatitis B virus (HBV). Our aim was to evaluate the antiviral effects of crude extracts (water and alcohol extract) from fruiting bodies collected from wild, and liquid fermentation of different strains of G.lucidum isolated from various geographical locations in Finland against enterovirus B group viruses, such as coxsackievirus B3 (CVB3). The Ganoderma extracts and sterile-filtered liquid culture media were incubated for 1 h or shorter periods with the purified virus, and thereafter added on lung carcinoma cells (A549). In addition to infectivity tests, a more detailed investigation to decipher the mechanism of action were performed using confocal microscopy, gradient fractionation and TEM. Our results show a direct inhibitory effect of liquid cultures on enterovirus (CVB3) without cytotoxicity in human cells. We noted a clear difference in the intensity of inhibitory effect between different strains of G. lucidum. In the liquid fermentation, the duration also clearly affected on the results, longer fermentation time giving higher inhibitory effect. These results demonstrate the importance of screening and selection of fungal strains with desired intensity of effect. This is of utmost importance if aiming for production of pharmaceutical products in bioreactors.
Taxonomic analysis and medicinal properties of macromycetes in Shikahogh State Reserve (Republic of Armenia)
- L. Margaryan
- S. Nanagulyan
- Y. Hovhannisyan
- R. Matevosyan
- V. Gevorgyan
Biological diversity is one of the powerful reservoirs of resources. The main tasks of a modern science is the study and preservation of biodiversity and its conservation in separate regions, including special protected nature areas (SPNA), which is one of the main key for humanity sustainable development. The goal of this work is a comprehensive study of macroscopic fungi species composition in Shikahogh State Reserve of Armenia. The reserve, with an area of 12.137 ha, is located in the south-eastern part of Syunik Marz, on the northern macroslope of the Meghri Range, at an altitude of 700 to 2400 meters above the sea level. Shikahogh reserve is represented by different types of landscape due to the difficult mountain terrain, vertical zonation, from the forest to the mountain steppes and alpine meadows ending. As an experimental material was chosen macroscopic fungi, collected from the territory of the Shikahogh Reserve as well as the samples from the herbarium of the Department of Botany and Mycology of Yerevan State University. The studies were carried out during the vegetation period using the route-expedition method from 2009 to 2016. Within the studied area 436 species of macrofungi that belong to 176 genus, 74 families, 22 orders, 7 classes, 2 subdivisions and 2 divisions of Ascomycota and Basidiomycota were revealed. The taxonomic analysis of the main families of macroscopic fungi in Shikahogh Reserve area showed that the following families are characterized with rich number of species: Polyporaceae (47 species), Tricholomataceae (38), Agaricaceae (26), Russulaceae (26), Strophariaceae (23), Hymenochaetaceae (21), Cortinariaceae (19), Inocybaceae (17), Mycenaceae (12) and the rest 65 families including 1-9 species (47.5%). In our studies we also registered 99 species of macromycetes with medicinal properties. They belong to classes of Agaricomycetes, Sordariomycetes and Tremellomycetes and 9 orders. The main part of the fungi with medicinal properties refers to the order of Agaricales (52 species). Most of the species found on the territory of the Reserve have antibacterial, antiviral, antifungal properties, which is associated with such chemicals as terpenoids, purines, phenol derivatives isolated from fruit bodies and fungal mycelium (Agaricus campestris, Coriolus versicolor, Ganoderma applanatum). For example, according to our data, the local population engaged in animal breeding uses Lycoperdon perlatum for the treatment of purulent diseases of rabbit ears. Thus, this study indicates that the collected data can be used for an update of the data passports of protected areas of Republic of Armenia and it is giving potential opportunities for the usage of macromycetes in medicine and the national household.
A Taxonomic Survey of Commercially Available Reishi Products: A Buyer Beware Market
- A. Loyd
- M. Jusino
- C. Truong
- R. Blanchette
- B. Richter
- M. Smith
- J. Smith
The genus Ganoderma contains species that occur commonly around the world and function as primary wood decay fungi of a wide array of tree species. In addition to ecological functions, species of Ganoderma have been used in traditional medicine in Asia for thousands of years. They are often referenced with common names such as reishi or lingzhi. Ganoderma species contain suites of triterpenes and polysaccharides that have been reported to have medicinal value, and have gained interest from the pharmaceutical industries in recent years. Globally, the taxonomy of Ganoderma species is chaotic, and the taxon name G. lucidum (sensu lato) has been used for most laccate (shiny) Ganoderma species. However, it is now known that G. lucidum (sensu stricto) has a limited native distribution in temperate climates of Europe and some parts of China. Furthermore, reishi or lingzhi products, sold as medicine, are not strictly regulated by the Food and Drug Administration in the United States. To determine what species are being sold in commercially-available products marketed as reishi or lingzhi, twenty products labeled as containing G. lucidum were purchased, DNA was extracted and the internal transcribed spacer (ITS) region was sequenced using Sanger sequencing. Based on Sanger sequencing, the majority of the products (93%) were identified as G. lingzhi, which is native to Asia and the most widely cultivated taxon. Microscopic analysis of the products revealed different spore morphologies within individual products, and Illumina sequencing of the ITS1 region was performed on all products to determine if multiple Ganoderma species could be present. Of the twenty products tested, none contained the species G. lucidum. Similar to the Sanger sequencing results, the Illumina results confirmed that G. lingzhi was in most products, but other Ganoderma species were also present, including the taxa G. applanatum G. gibossum, G. sessile, and G. sinense. It is likely that there are differences in the quality and quantity of medicinally-relevant chemicals produced by different Ganoderma species. Furthermore, if fruiting bodies are cultivated and/or formulations are manufactured outside of the United States, regulations focusing on the content and quality of these products should be re-evaluated before they are ingested or marketed as medicine.
Temporal stability of the fungal community in a tropical rainforest canopy
- K. Cook
- J. Sharma
- A. Taylor
- D. Taylor
The spatio-temporal dynamics of fungal communities are largely unknown, despite their ecological importance. Changes in fungal species composition and relative abundance can have impacts on ecosystem processes, such as decomposition rates. They also affect macroscopic plant communities, through gains or losses of mutualistic or pathogenic fungi. Many ecological studies only capture a single snapshot of the fungal community or compare single time points before and after a major disturbance or experimental manipulation. It is often not known if these communities are stable over time. More research is needed to see if observations at one time-point can be used to make inferences about the future. To address this problem, we examined the temporal variability of the diverse fungal community in a tropical rainforest canopy. We selected five branches of Saurauia montana trees in a low montane rainforest in Tapanti National Park, Costa Rica and sampled 64 points across these branches each July for three years. Samples included tree bark and any litter or living bryophytes present on the branch surface. ITS sequencing with Illumina technology was used to assess fungal diversity. We found that there is a small, but statistically significant, effect of year on fungal community. Despite this, the fungal community at a given point was more similar to communities at the same point in different years than to other points in the same year, even when these points are nearby on the same tree branch. In previous research on community spatial structure in this system, we found high turnover between samples at the same time-point at sub-meter spatial scales. Here we found that species composition is consistent at a point between years. We conclude that the fungal communities in this system are structured at a very fine spatial scale but are persistent over time. Thus, observations at one time-point can be representative of points in the future.
Characterizing microbiomes of fungal fruiting bodies across functional guilds and growing habits
- G. Benucci
- G. Bonito
Over the past decade, culture-independent approaches employing next-generation DNA sequencing have revolutionized our capacity to investigate the composition and dynamics of microbial communities (microbiomes). Major advances have been made on understanding human, animal, and plant microbiomes, however, less attention has been given to the microbial communities of fungi. Bacteria perform various functions in fungi and their reproductive structures (fruiting bodies), spanning from pathogenicity (e.g., mycoparasites) to growth promotion (e.g., mycorrhizal helper bacteria). However, how mushroom-associated bacteria assemble and function within fungi belonging to different ecological guilds and growing habits has yet to be addressed. To this end, we used Illumina high-throughput 16S rRNA amplicon sequencing to characterize bacterial communities in thirty-two fungal genera across Ascomycota and Basidiomycota. Through bioinformatic analyses of the ~6.4M raw sequences generated we recovered 1384 Operational Taxonomic Units (OTUs) across 160 samples. Proteobacteria (71.8%), Bacteroidetes (23.3%) Firmicutes (1.9%), and Actinobacteria (0.7%) were the most abundant phyla overall. Bacterial communities structured according to different fungal functional guilds (mycorrhizal, endophytic, saprophytic) as well as growing habitus (hypogeous, epigeous, wood-growing). The highest bacterial richness was recorded in Tuber, Chantarellus, Hydnum, and Morchella; the lowest in Ganoderma, Hericium, Paxillus, and Lyophyllum. The genus Bradyrhizobium was abundant in Tuber, but also present in Scleroderma, Elaphomyces, and Lycoperdon, all grouped by a similar sequestrate fruiting structure. Pseudomonas was highly abundant in Coprinus, Leatiporus, and Suillus, while Burkholderia were dominated inside Clavariadelphus, Grifola, and Hydnum. Bacteria belonging to Polaromonas, Pedobacter, and Janthinobacterium were all abundant in Helvella, Gyromitra, and Morchella which all share a similar fruiting morphology and belong to Pezizales. Our data demonstrate that fruiting bodies microbiomes relate to the functional guild of their fungal host, which raises the question of whether these bacteria provide symbiotic functions or depend upon specific ecological requirements. A wider sampling of fungi belonging to the same trophic guild, in different geographic areas, will improve the separation of community variations attributable to ecological habitat filtering from metabolic-based symbiotic interactions.
Potential distribution of South American species of the lichen genus Parmotrema (Parmeliaceae, Lecanorales): implications for conservation
- A. Michlig
- N. Niveiro
- N. Ramírez
- M. Rodríguez
Species distribution models have become an important tool to assess different issues in fields as biogeography, ecology, evolution, and climatic change. Although they have been commonly used in other organisms, just a limited number of studies about lichen-forming fungi have included modeling approaches. Species distribution models allow us to recognized those areas with favorable ecological conditions to species could develop outside their known localities, and to determine rare or threatened species, contributing thus to the development of conservation strategies. The aim of this study is to estimate the potential geographic distribution of South American species of Parmotrema, to identify which are the environmental variables influencing their distribution, and to analyze their conservation implications. For this, six species were selected: P. cristobaliae, P. flavomedullosum, P. homotomum, P. laciniellum, P. masonii, and P. melanochaetum. A database with recorded localities in literature for each species and also specimens deposited at CTES herbarium was made. Localities for which no geographic coordinates were available were georeferenced with Google EarthTM. The potential distribution was modeled with Maxent version 3.3.k, using the 19 climatic variables of temperature and precipitation and altitude of the WorldClim database, estimating their potential influence in these species distributions. The results were visualized and analyzed using DIVA-GIS version 7.5. The potential distribution maps are presented and the influence of environmental variables of each considered species are analyzed. This study has shown that P. homotomum and P. masonii has a restricted potential distribution area. The distribution of P. masonii, as P. cristobaliae, could be related to the distribution of the seasonally dry forests, as other species of the genus. The distribution of P. laciniellum has also a similar biogeographic pattern, but it would be adapted to more wide environmental conditions. Based on these results, we could identify two priority areas for conservation.
Mapping fungal diversity at the global scale
- C. Lepinay
- A. Machač
- P. Baldrian
- P. Kohout
- D. Storch
- T. Větrovský
Fungi represent a fundamental component of ecosystems due to their major ecological and economic roles. The fungal distribution worldwide is not only governed by dispersal limitation but also by the suitability of ecological niches determined by both the abiotic environmental factors, i.e., climate and soil characteristics, and the biotic factors, i.e., the plant communities. While most organisms show a gradient of diversity that increases near the tropics, the gradient of fungal diversity is not yet fully described. Therefore, we aimed to map the biogeographic patterns of soil fungal diversity on global scale. Our objectives were to characterize the fungal taxa richness gradient worldwide and to determine which climatic factors could best explain such a gradient. To explore the geographic patterns of fungal diversity, we assigned the soil samples to 1 x 1 degree grid cells covering the globe. Grid-based, rather than locality-based, analyses standardize the geographic scale of the analysis. Altogether we identified fungal diversity in more than 200 grid cells. We used multiple approaches to evaluate the patterns of global diversity. First, we used non-parametric smoothing to investigate the changes in species diversity (number of taxa) with latitude, using a second-degree polynomial function fitted locally to 75% of the data points. Second, we investigated the changes in species diversity across different regions of the world, using climate-based generalized linear model (GLM). Specifically, we characterized the climate within each grid cell using 19 bioclimatic variables and net primary production. These variables capture temperature, precipitation, seasonality, their mean and variation over the course of the year. To identify the most parsimonious model of fungal diversity, we evaluated 4.5 million candidate GLMs that contained different combinations of the previously compiled variables using an exhaustive search algorithm in the R package leaps, but limiting the maximum size of the model to 10 variables to ensure tractability. The best model was selected based on adjusted R2, AIC, and BIC and used to predict fungal diversity across all grid cells covering the globe. Namely, we predicted the expected diversity of species, and the upper and lower bounds on the expected diversity (95% prediction interval). Finally, we evaluated the global patterns of species diversity using kriging. Kriging extrapolates the patterns of species diversity from the inferred degree of geographic autocorrelation in species diversity across the sampled regions. The results, across different methods and statistical setups, consistently indicate that fungal diversity declines from high latitudes toward the tropics. Non-parametric smoothing reveals that fungi reach their highest diversity at high latitudes of the northern hemisphere. More detailed results, derived from climate-based GLMs, confirm that fungal diversity peaks in the boreal forests of Eurasia and North America. Tropical diversity is limited in comparison, except for several putative biodiversity hotspots in the Amazon and African savannahs. These results indicate different global diversity patterns between soil fungi and other previously studied organisms such as plants or animals. Our study should help fungal ecologists to advance the knowledge on fungal diversity and ecology worldwide.
Host specificity differs along the life cycle of Tremella hypogymniae
- A. Millanes
- V. Tuovinen
- P. Diederich
- M. Westberg
- H. Johannesson
- M. Wedin
Although traditionally considered the symbiosis between a fungus and green algae or cyanobacteria, lichens are complex systems that harbour a diversity of additional organisms. How these organisms are distributed within the thallus is often poorly known. A variety of bacterial and fungal taxa inhabit the lichen thalli, including Alphaproteobacteria, Acidobacteria, Actinobacteria or Betaproteobacteria, and both ascomycetes and basidiomycetes in fungi. Among the basidiomycetes, the “heterobasidiomycetes” show a high specificity towards their lichenized hosts. These fungi are dimorphic: they have both a dikaryotic filamentous phase and a haploid yeast phase in their life cycle. The filamentous phase is usually host specific in the lichen-inhabiting species but little is known on the requirements of the yeast phase. Recently, it has been shown that both Cyphobasidium and at least one Tremella species are able to complete their life cycle within the lichen thallus, where the yeast phase grows in the lichen cortex. In this work we further investigated the frequency and host-specificity of the yeast phase of tremellalean fungi growing on lichens. For this we chose Tremella hypogymniae, a species that is restricted to one of the most common lichens in boreal and temperate forests of the northern hemisphere (Hypogymnia physodes). We used highly specific PCR primers to selectively amplify T. hypogymniae, avoiding other organisms including other tremellalean taxa. We searched for T. hypogymniae in H. physodes and in other common Parmeliaceae species more or less closely related to H. physodes in different coniferous forests in Sweden and Spain. In addition, we performed fluorescent in situ hybridization (FISH) to identify the location of the different phases of the life cycle of Tremella within the host lichen thalli. Our results suggest that the yeast phase of T. hypogymniae is very frequent – although not always detected – in the cortex of several closely related lichens, whereas meiosis and subsequent formation of basidia seem to occur only when Tremella hypogymniae grows in Hypogymnia physodes. This implies that host specificity is related to sexual reproduction, whereas the asexual yeast-phase can develop asymptomatically in a wider range of hosts. Our study represents an important step in the understanding of the life cycle and biology of lichen-inhabiting basidiomycete fungi.
Lichenicolous ascomycetes on Siphula-like lichens
- J. Motiejunaite
- A. Suija
- M. Zhurbenko
- G. Kantvilas
A survey of collections of Siphula-like lichens [Siphula (Icmadophilaceae) and Parasiphula (Coccotremataceae)] in the herbaria of H, HO, TNS and UPS revealed a rich and highly specialised lichenicolous biota of c. 20 taxa. Of these, we have been able to identify c.10 to species rank, including five that we describe as new to science in the genera Cercidospora, Endococcus and Pyrenidium. In addition, we significantly expand the geographical distribution and the number of host species for siphulicolous fungi. By far the greatest species richness is found in the Southern Hemisphere, the centre of speciation of the host genera. The only lichenicolous species recorded from the Northern Hemisphere is Sphaerellothecium siphulae. No lichenicolous fungi are shared between Siphula sensus stricto and the morphologically similar genus, Parasiphula, providing further support for their separation.
Ebony and algal green: Characterizing fungal-algal interactions in a biological soil crust
- E. Carr
- W. Riekhof
- J. Herr
- S. Harris
Biological soil crusts (BSCs) are a type of biofilm that can withstand a wide range of extreme conditions, including: heat, freezing, desiccation, osmolarity, UV, and heavy metals. Some of the most common fungi in BSCs are the polyextremotolerant black yeasts in the Chaetothyriales, such as those in the genus Exophaila. Unique features of these organisms make them intriguing to study alone, but may also provide insight into the evolution of fungal-algal interactions. These polyextremotolerant fungi share similar resistance features with the lichens, therefore black yeasts may enable study of the lichen lifestyle without the challenges of directly working with lichens. In addition to sharing similar traits, black yeasts are also almost always found in the same location as algae, even in desert and polar ecosystems. We hypothesize that black yeasts and algae indeed interact in a mutualistic fashion that resembles lichens, and that observing these interactions will reveal clues about the nature of the fungal-algal communication that underlies the formation of BSCs. To test this hypothesis we have isolated black yeasts and algae from BSCs and rock surfaces, identified isolates using their ITS sequences, and subjected isolates to multiple stressors and nutritional conditions to investigate their phenotypic diversity. Culturing efforts and Sanger sequencing resulted in the identification of 24 black yeasts and 43 algal isolates from eight locations sampled within BSCs found in a semi-arid sand dune ecosystem (Jackman Flats Provincial Park, BC). In addition to these culturing efforts, we sequenced both ITS1 and 16S amplicons using Illumina sequencing to elucidate the microbial composition of the BSCs. Additionally, we have co-cultured isolated fungi and algae together in a pair-wise manner to observe phenotypes of their interactions. Current efforts are focused on integrating phylogenetic, phenotypic, and microbiome data to understand the functional interactions that enable the formation and maintenance of BSCs.
Ultrastructural study of the algae within the globule of Multiclavula mucida
- H. Masumoto
- Y. Degawa
Multiclavula mucida (Pers.) R. H. Petersen is a basidiomycete forming tiny white fruit bodies on rotten wood and always exists together with algae. Although the ecological association between M. mucida and algae is not elucidated, M. mucida is often regarded as one of the basidiolichens. There are many green globules on rotten wood where M. mucida produces fruiting bodies. In the light microscopic observation, Geitler (1955) and Oberwinkler (1970) reported that these globules consisted of pseudoparenchymatous hyphae of M. mucida and several algal cells. Oberwinkler (1984) also observed these globules by transmission electron microscopy (TEM), but it was not mentioned much about intracellular structures of both fungi and algae. In this study, we observed these globules by TEM, mainly focused on algae within the globules. TEM observation showed that the outer surface of the globules was consisted of hyphae. The cytoplasm of the hyphae was often filled with osmiophilic granules with a diameter of about 1 μm. Such granules were also observed in the other basidiolichen Omphalina ericetorum (Botrydina vulgaris globules) and treated as glycogen particles (Honegger and Brunner, 1981). Hyphae were also observed within the globule, but no haustorium was found. Each globule contains several to 15 algal cells. Each algal cell was occupied by several electron-dense storage bodies and a single chloroplast. The chloroplast is biased towards the cell wall, thylakoids overlap in many layers, filling the whole chloroplast. At the center of the chloroplast several electron-dense granules were gathered. These granules were similar to the pyrenoglobuli which are found within the chloroplast of Trebouxia and always associated with a pyrenoid. If these granules are pyrenoglobuli, this alga is considered to have a pyrenoid. The osmiphilic granules with in the hyphae and pyrenoglobuli-like granules within the chloroplast can be also recognized in the TEM photographs by Oberwinkler (1984) (but not enough explanation was given). Geitler (1955) reported that the algae within the globule belong to the genus Coccomyxa, but Komárek & Fott (1983) and Tschermak-Woess (1988) pointed out that the alga had a pyrenoid and could not belong to Coccomyxa, a pyrenoid-lacking genus. We established 24 algal cultures from M. mucida globules. As a result of molecular identification using the sequence of ITS, they were divided into at least three species (two Coccomyxa spp. and Elliptochloris subsphaerica). The chloroplast of E. subsphaerica is filled with thylakoids that overlap in many layers, and pyrenoglobuli-like granules exist in the center. These characters are consistent with those of algae within the globule of M. mucida, whereas both were absent from the two Coccomyxa spp. According to Ettl and Gärtner (2014), E. subsphaerica has a pyrenoid. In M. mucida-E. subsphaerica co-cultivation, algal cells were closely surrounded by the hyphae under a certain condition, which is similar to the early stage of the globule development in the field as reported by Geitler (1955). Based on the above, E. subsphaerica is eligible to be considered as photobiont of M. mucida.
Exploring foliar fungal endophyte assemblage, diversity, and host specialization in pine
- J. Sarver
- R. Oono
- R. Salas-Lizana
- D. S. Gernandt
Host specialization of foliar fungal endophytes (FFE’s) remains a cryptic and uncommon phenomena. Since patterns of host specificity are sensitive to host taxonomic and spatial scales, a field study that investigates the fungal endophytic community of conifers, a taxonomically well-defined and diverse group, across a wide geographic range (spanning much of North America) was conducted. Conifer trees have a high incidence of FFE infection, likely due to the longevity of their evergreen foliage as well as their dominance in some ecosystems. Furthermore, Lophodermium (Rhytismataceae), a well-studied FFE genus that seems to be common within needles of the Pinaceae family, indicates high phylogenetic host specificity that is rarely documented for other FFE’s. In total, 51 species of conifers were sampled from 69 localities across the United States and Mexico. Illumina MiSeq sequences were collected on the community assemblage of FFE’s with a closer investigation of Lophodermium OTUs. Host specificity of common OTUs were analyzed and compared across geographic and taxonomic groups of conifers. These findings will be crucial for furthering our understanding of the evolutionary and ecological nature of these mysterious microfungi on their hosts.
A new species of Lophodermium (Chevall.) described using a combination of morphological data, ITS-LSU rDNA and ddRAD loci
- R. Salas-Lizana
- R. Oono
Lophodermium is a common endophyte of pine needles with ca. 30 putative species. The species within this genus have large-within species genetic variation with the possible occurrence of cryptic species. The relatively poor sampling isolates with molecular and morphological characterization, together with the lack of genetic markers to resolve its phylogeny with good support, are problems that need to be addressed before attempting further studies or experiments to better understand their ecological role, beyond their ‘endophyte’ status. We performed a non-exhaustive survey in five-needle pines of Norther California and the Pacific Northwest obtaining cultures from both monosporic ascocarp isolates and surface-sterilized green needles. Ascocarps were identified using morphological characters, and DNA was extracted from cultures to sequence the internal transcribed spacer and a partial sequence of large rDNA subunit (ITS-LSU) using Sanger technology. Furthermore, we used a subsample of isolates to recover putative homologous loci using a novel restriction site-associated DNA sequencing (RADseq) method, developed by our research group. In total, we recovered 250 isolates from both green needles and ascospores. Most isolates belong to L. nitens Darker and a new species, L. fissumilis sp. nov. Salas-Lizana and Oono. The new described species resembled L. nitens morphology because both present dark subcuticular ascocarps without lips. However, the upper wall of both ascocarps is very different, as the new species forms an inward v-shape folding, not present in L. nitens. A phylogeny using ITS-LSU confirmed that L. fissumilis represents a divergent species, although the support for deeper branches was low. After several experiments using different combinations of similarity clustering and quality filtering, a final concatenated matrix of ddRAD loci at 70% similarity and up to 30 ambiguous sites per locus was used to infer a phylogeny of a subsample of six putative species, including the new species. The topology of this phylogeny is highly similar to that of ITS-LSU but with high support for all branches. This fine-resolved phylogeny also revealed that the populations of the new species are highly structured as well as the potential occurrence of cryptic species within L. nitens. This work shows the potential of combining classical morphological and cutting-edge technologies to describe accurately the diversity of understudied groups, such as Lophodermium.
Local habitat, leaf biochemistry and seasonality drive phyllosphere fungal communities of European beech
- A. Siddique
- M. Unterseher
Comparative studies targeting the drivers of endophytic fungal biodiversity are rare and identified multiple effectors, such as plant chemistry, climate and seasonal attributes. Our project aimed to study the pattern of the leaf-associated mycobiomes of European beech (Fagus sylvatica) at different altitudes to reveal diversity, composition and seasonal dynamics of fungal endophytes by a combination of metabarcoding, cultivation and subsequent ecological analyses. An experimental field site consisting of 100 (2-years old beech) trees was established called ‘beech phytometer system’ at two altitudes (517 and 975 m a.s.l.) in a German mountain forest. Ten trees from each site were chosen and 10 leaves per tree were sampled. The following workflow included leaf biochemistry measurement and fungal DNA metabarcoding with 97% OTU threshold and biodiversity analysis for two continuous years: five trees from beech phytometer and five trees from surrounding beech trees. Metabarcoding resulted in a total of 15,703,599 demultiplexed and quality filtered ITS1 reads from 165 samples. Clustering at 97% similarity resulted in 1199 OTUs. In addition, 438 isolates from an autumn sampling event were cultivated via the dilution-to-extinction method; endophytes were identified via barcoding based on ribosomal ITS (internal transcribed spacer) markers and Sanger sequencing. A significant correlation of community composition with elevation was observed. The mycobiome was little affected by the physiological state of the leaves; only a partial shift of taxonomic composition was observed from vital towards senescent leaves. Mycobiome diversity and composition correlated significantly with the origin of the trees, pointing to local habitat condition as the main driver. Under natural conditions, the mycobiome was more diverse at the lower elevation. Additionally, leaf chlorophyll and flavonoid contents showed negative correlations with fungal richness in natural stands. Metabarcoding and cultivation approach resulted in non-overlapping community compositions and pronounced differences in taxonomic classification and trophic stages. However, both methods revealed similar correlations of the fungal communities with local environmental conditions. Our results indicate undeniable advantages of metabarcoding over cultivation in terms of representation of the major functional guilds, rare taxa and diversity signals of leaf-inhabiting fungi. We observed a strong seasonal turnover in phyllosphere fungi in both habitats over the two years of investigation, suggesting that the plant- fungal system not only responds to cyclic climatic conditions but depends as well on various parameters, e.g., geographic position, substrates age and surrounding vegetation. In general, the altitudinal difference is the most important explaining factor for community differences, which shapes many dependent abiotic and biotic habitat factors. Regarding cost and time per sequence, metabarcoding is superior to cultivation approaches and offers surprisingly profound insights by yielding much more data, allowing to test at once multiple hypotheses in fungal ecology.
Habitat modeling of foliar endophytic fungi across the Hawaiian archipelago
- J. Darcy
- S. Swift
- G. Cobian
- A. Amend
A phylogenetically diverse array of fungi live within leaf tissue, and in many cases, these foliar endophytic fungi (FEF) live mutualistically with the plant host by producing secondary metabolites or conferring disease-resistance. Many studies have examined FEF across single plant species, or focused on individual FEF taxa, but the broad-scale distributions of these fungi are not well understood, either across geographic space, across climactic conditions, or in the context of host plant phylogeny. The Hawaiian archipelago represents a uniquely tractable system for studying the habitat preferences of FEF, because the islands each have a wide range of climactic conditions, and host species can be found across islands, but dispersal barriers are significant between islands and between the archipelago and other land-masses. Therefore, the Hawaiian archipelago represents an ideal system to apply island biogeographic principles to the study of FEF. Here, we use high-throughput DNA sequencing of the ITS1 region to characterize the FEF communities of over 1000 individual native plants across 5 islands in the Hawaiian archipelago. Using climactic information, geographic locations, a host-plant phylogeny, and a FEF phylogeny, we characterize the dominant forces structuring FEF communities, and model the environmental preferences of the most cosmopolitan endophytic fungi. We found that host phylogeny and precipitation were strong determinants of community structure, but individual FEF species exhibited a wide array of habitat preferences, suggesting that aggregate beta-diversity analyses represent a narrow view of the factors structuring FEF community assembly. These results are the culmination of a four-year effort to characterize FEF communities across the Hawaiian archipelago, and although our findings are limited to Hawaii, the patterns we show here for communities and FEF taxa may serve as hypotheses for future studies in other locations. Within Hawaii, our results answer important questions about host- and habitat-specificity of FEF in native plants.
How to encompass epiphytic lichen diversity in a protected area? A case-study in a natural park in NE Iberian Peninsula
- E. Llop
Since 2012 a survey has been carried on in order to assess lichen diversity in the Cadí-Moixeró National Park (NE Iberian Peninsula). Like in several protected areas in the region, lichen biota has showed up as one of the underrepresented components of biodiversity. The main goal of the survey was to disclose epihytic lichen diversity from the natural park. To achieve this aim, and with the idea to cover as many habitats as possible, a sampling protocol was designed. The protocol was first tested in three coniferous forests, and results were compared with previous studies from other areas within the Iberian Peninsula. The obtained results prompted the application of the protocol in representative habitats of the Cadí-Moixeró National Park. During five campaigns, epiphytic lichen diversity was sampled from ten forestal and arbustive communities. Wood communities included broadleaved woods; such as beechwood, oak grove, and holm oak forests; and coniferous forests, as fir wood, Scots pine and Pinus uncinata forests. Additinally, some shrub dominated communities and fluvial forests were also examined. The sampling campaigns yielded an increase in the number of species mentioned in the natural park. In 2012, the lichen catalogue compiled 105 taxa. Five years later, the list of lichen has risen to 319 taxa. Among them, six taxa were newly quoted for the Iberian Peninsula. The epiphytic lichen diversity was represented by 272 species. The study on the species composition from the habitats showed that there was a clear difference between coniferous and broadleaved forests (including shrub and fluvial forests). However, the comparison of ecological traits such thallus growth or photobiont did not present the same pattern. No significant differences were observed between both main sorts of forests. The species-area curve suggested that the potential epiphytic lichen diversity ranges between 345 and 359 taxa, depending on the applied model (jackniffe or Chao2). Although the sampling protocol has provided a good insight in the lichen diversity in the Cadí-Moixeró Natural Park, the complete knowledge is still far of being achieved. The selection of representative habitats has resulted in a good way to increase the knowledge on lichen diversity; however, more especific habitats, like meadows or rocky places, should be included in order to gain a complete view on lichen biota.
Host specificity of endophytic fungi isolated from roots of Calanthe discolor and Cephalanthera longibracteata in Korea
- L. Bong-Hyung
- A. Eom
Orchidaceae is the most diverse and widespread family of flowering plants, with 25,000 species currently recognized. In Korea, more than 100 species have been reported in the wild. However, currently, many orchids are in danger of extinction due to over-collection and habitat destruction. Orchidaceous plants have symbiotic relationships with endophytic fungi, including mycorrhizal fungi, which play important roles in seed germination and growth of the host plants. In this study, the endophytic fungal communities were isolated from the roots of Calanthe discolor Lindl. and Cephalanthera longibracteata Blume that were collected from two different sites in Korea. The fungal isolates were identified by sequence analysis of the internal transcribed spacer regions of rDNA. In total, 35 species of endophytic fungi, including two species of mycorrhizal fungi belonging to the genus Tulasnella and were identified in C. discolor. Furthermore, 29 species of endophytic fungi were identified in C. longibracteata. The species diversity and richness were not significantly different among the two sites. However, the endophytic fungal community was highly specific to the host, suggesting that the host characteristics affected the community composition of the endophytic fungi that colonized the roots of the orchids. Our findings will help in developing methods that use symbiotic fungi for orchid conservation and restoration of native habitats.
Towards deciphering the fungal diversity associated with Malus domestica in Germany using high-throughput amplicon sequencing
- F. Witfeld
- D. Begerow
- M. Kemler
The holobiont concept and hologenome theory highlight the influence that microbial communities can have on the ecology and evolution of their associated macro-organismal hosts. The outcome of those associations thereby not only depends on interactions between the microbiome and the host, but also on the interactions between members of the microbiome, including mainly bacteria and fungi, but also viruses, nematodes and protozoa. Fungi form an important part of the plant-associated microbiomes and the composition of fungal communities and the resulting interactions are highly relevant for the ecology of plants, affecting them in beneficial and/or harmful ways. Therefore, processes within natural ecosystems (e.g., nutrient cycling), as well as in agricultural systems (e.g., plant health) are strongly influenced by the mycobiome. Apple (Malus domestica Borkh) is one of the most widely cultivated fruit crops in temperate regions and in Germany by far the most important fruit tree. A better knowledge about the apple tree associated mycobiome and its functioning could enable a targeted application, resulting in better crop yields or reduction of fungicide usage. In this study, fungal communities associated to apple trees in Germany were assessed by high-throughput sequencing of amplicon barcodes. To characterize the fungal diversity associated with apple trees and the factors controlling the community assembly, we sampled 85 apple trees at seventeen locations along two transects (North-South and West-East) throughout Germany. Sampling locations were consistently selected for lack of pesticides, herbicides and fungicides. From each host, leaves, twigs and surrounding soil were collected. DNA was extracted from three independent replicates, the fungal barcode ITS1 amplified and sequenced by Illumina MiSeq. We obtained 11,311,311 quality filtered ITS1 sequences for leaf and twig samples, which were clustered into 575 OTUs. Our results revealed that the host organ and location were major factors shaping the detected fungal communities, while the distance between locations did not show a major effect. Leaf samples were more equally dominated by Ascomycetes and Basidiomycetes, while Ascomycetes prevailed by far in twig samples. A slight similarity decay along the North-South transect was also observed. We will provide insights into community assembly and further details on the fungal communities, their structure, the taxonomic distribution according to organ and geographical location.
The structure of fungal communities on clonal Eucalyptus shows effects of host genotype and environment
- M. Kemler
- M. Vivas
- B. Slippers
Fungi and plants often form intimate relationships that have influenced the ecology and evolution of both groups in a significant way. The phyllosphere (i.e. the aboveground parts of plants) is one of the interfaces of plant-fungus interaction that has gained recent attention, as it has been hypothesized that the fungi inhabiting it can have beneficial effects on their host. Additionally, high-throughput amplicon based diversity assessments have shown that this habitat harbors an enormous species diversity of non-symptomatic fungi. In this study, we applied NGS amplicon sequencing to understand the influence of geography and host genotype on the structure of fungal communities in replicates of three different clonal Eucalyptus lineages in two geographic locations of South Africa. Additionally, we studied the formation of fungal communities in seedlings from these Eucalyptus clones, grown in an incubator and subsequently in a nursery. Our analyses on the plantation trees show that the structure of fungal communities is influenced by the different geographic locations. The fungal communities of seedlings were unique to their environment and showed a high turnover when moved between locations.
Effect of ectomycorrhizosphere bacteria on growth, ectomycorrhizal formation and sporocarps occurrence of Laccaria laccata
- K. Obase
Ectomycorrhizal (EcM) fungi colonize plant roots and form symbiotic associations with the host plants. It has been shown that a variety of bacteria inhabits surrounding the symbiosis and several of them affect either the mycelial growth of EcM fungi and/or establishment of EcM symbiosis from positively to negatively. However, we have still limited views of functional meanings of the ectomycorrhiza-associated bacteria on the ecology of EcM fungi, especially how they affect on EcM sporocarps occurrence. In this study, I targeted on Laccaria laccata that forms sporocarps in vitro with symbiosis with host plants. First, one soil core was collected from 5 locations where sporocarps of L. laccata occurred in a chestnut (Castanea crenata) plantation in Yamanashi prefecture, Japan, to understand bacterial community on EcM roots of L. laccata. Ten and 50 EcM root tips from each soil core were subjected to bacterial isolation and cloning, respectively. Two-hundred-forty-eight isolates were obtained in total and they were divided into 34 MOTUs (99% similarity threshold). Most of MOTUs were infrequent, while one MOTU of Rhizobium sp. and Bradyrhizobium sp. were commonly and frequently obtained across sampling locations. In cloning method, 83 MOTUs were detected. Most of MOTUs were infrequently detected, while one MOTU of Bradyrhizobium sp. that were also commonly found in isolation-based method, were commonly detected across sampling locations. The results indicate that Rhizobium and Bradyrhizobium are common bacteria on EcM roots of L. laccata in the chestnut plantation. Next, I examined the effect of the ectomycorrhizal-associated bacteria on the hyphal growth of L. laccata using dual-culture method. One mycelial disc of L. laccata was placed on the center of a plastic dish (9 cm diam) with 15 mL of modified Melin Norkrans (MMN) agar media with ten-fold dilution of glucose, and each bacterial strain was incubated 2 cm apart from the mycelial disc. Areas of hyphal growth were calculated after 1-month incubation. Effects of bacteria on hyphal growth largely differed according to the combination of isolates of bacteria and L. laccata but most of bacteria tended to inhibit hyphal growth of L. laccata. Bradyrhizobium and members of Rhizobium and Burkholdeliaceae did not inhibit or rather promote hyphal growth of L. laccata. Last, bacterial isolates of Bradyrhizobium and Rhizobium spp. were introduced into L. laccata—Pinus densiflora symbiotic system in vitro, and EcM formation and frequency of sporocarps production of L. laccata were examined. Introduction of bacteria did not significantly increase host plant growth, EcM formations and frequency of sporocarps occurrence but addition of one taxon of Rhizobium tended to increase host plant growth and frequency of sporocarps occurrence. The results indicated that several members of ectomycorrhizosphere bacteria would support sporocarps occurrence of the EcM fungi.
The importance of declining mammalian fungal specialists for ectomycorrhizal fungal dispersal in Australia
- S. Nuske
- K. Vernes
- T. May
- A. Claridge
- S. Anslan
- L. Tedersoo
- B. Congdon
- A. Krockenberger
- S. Abell
Hypogeous sequestrate (truffle) fungi rely primarily on consumption by mammals for dispersal. Most truffle fungi are ectomycorrhizal (ECM), making mammalian dispersers essential to the maintenance of plant-fungal relationships, soil fungal diversity and ecosystem functioning. Australia has the highest current global rate of mammalian extinctions, including important specialist mycophagists within the family Potoroidae. Knowing the functional redundancy of different mammal species as dispersers helps us to understand how this loss in mammal diversity could affect plant-fungi interactions and fungal diversity. We present data from a meta-analysis of Australian mammalian diets and a field study including data on an endangered specialist mycophagist. Data from the literature support our hypothesis that specialist mycophagists within the family Potoroidae consumed and potentially dispersed a significantly higher abundance and diversity of fungi than other mycophagous mammals with generalist diets. This finding was further corroborated in the field study; the specialist mycophagist, Bettongia tropica, consumed a higher diversity and more unique species of ECM truffle fungi than nine co-occurring mycophagous mammal species. This implies that between specialist and generalist mycophagous mammals in Australia, there is little functional redundancy with respect to fungal dispersal. Taken together, the results suggest changes to mammalian communities, particularly the loss of specialist mycophagists, could, over time, induce significant changes to truffle diversity, shifting ECM communities with unknown consequences for plant health and nutrient cycling.
Coupling effects of climate and tree growth on the productivity of the prized Tricholoma matsutake mushroom
- L. Guo
- Z. Fan
- J. Li
- X. Yang
- Z. Yu
- B. Yingfeng
The on-site observation research on wild edible mycorrhizal mushrooms is rare, which hampers the understanding of the ecological process related to the production of the mushroom. Taking the ecotomycohhrizal fungal, Tricholoma matsutake as a study system, we attempted to understand how the production of this prized mushroom is influenced by climate variation and the growth status of their host trees. A 15-year long term monitoring data set about T. matsutake fruiting in Southwest China were analyzed against climatic and tree ring data (Pinus yunnanensis and P. armandi) with a synopsis framework established to characterize the parameters related to T. matustake productivity. The Partial Least Squares was used to identify the most relevant climate data and key period of time, and the Path Analysis was used to determine the indirect and direct causes. Our results showed that both climate and tree ring width influence the production of T. matsutake, with tree ring width and precipitation in early spring as the most significant, positive and direct effects. The temperature in early spring and in summer also plays significant direct cause to the T. matsutake production. These result suggest that production of mycorrhizal fungal is determined by the coupling effects of climate and the growth status of the host trees. These findings may have major implications for improvement of mycorrhizal mushroom productivity by considering the interplay between climate, mushroom and forest.
Keywords: Tricoloma matsutake, Productivity, Tree-ring, PLS, Path analysis
Communication in Tricholoma vaccinum-spruce ectomycorrhizosphere
- K. Krause
- K. Wagner
- W. Boland
- E. Kothe
The ectomycorrhizospheric habitat contains a diverse pool of organisms, including the host plant, mycorrhizal fungi, and other rhizosphere microorganisms. Different signaling molecules may influence the ectomycorrhizal symbiosis.
Here, we show that the mutual symbiosis between the basidiomycete Tricholoma vaccinum and Norway spruce (Picea abies) shapes the surrounding mycorrhizosphere. In a forest biotope, this was characterized by a high diversity in basidiomycetes and a rich bacterial community. This consisted of mainly bacteria plant growth promoting abilities dominated by Rhizobiales, with Nitrobacter winogradski being most abundant (3.9 %). Other taxa mainly were pseudomonads and bacilli. The bacterial isolates showed symbiosis-relevant traits with 74 % producing the phytohormone indole-3-acetic acid, 23 % producing siderophores, and 23 % mobilizing phosphate.
The mycorrhizal fungus T. vaccinum was able to excrete plant hormones into the medium upon axenic cultivation. These include auxins, salicylic and abscisic acid, and jasmonates. The spruce roots exudated auxins and salicylic acid. With these compounds present in soil of a natural ectomycorrhizospheric habitat, a communication network with a response of T. vaccinum to the environmentally available salicylic and abscisic acids, which led to altered hyphal branching relevant for mycorrhization. In addition, the fungus protected the mycorrhizal tree against the spruce pathogens Botrytis cinerea and Heterobasidion annosum. Thus, the finely tuned phytohormone interactions in the mycorrhizosphere represent a specifically rich system to study microbial communication.
Allocation of nonstructural carbohydrates by Pinus strobus colonized by three ectomycorrhizal symbionts: Suillus, Rhizopogon, and Cenoccocum geophillum
- C. Victoroff
- G. Potter
- J. Castracane
- T. Horton
The NANAPHID is a newly developed nanobiosensor, which mimics an aphid to enable monitoring of three non-structural carbohydrates (glucose, sucrose, and fructose). By measuring these carbohydrates in situ, the NANAPHID will allow researchers to quickly measure plant carbohydrates in tissues under varied environmental conditions. So far, the NANAPHID has been utilized successfully to measure glucose levels (g/ml) over a wide concentration range in a phosphate buffered solution. The NANAPHID is also being beta tested to measure sucrose and fructose. The NANAPHID sensor is integrated with a laptop-based, fully automated data acquisition, processing and display system to enable field use by researchers. My research will utilize NANAPHID technology to measure the allocation of carbohydrates to mycorrhizal roots as a proxy for examining the allocation of carbohydrates by Pinus strobus to mycorrhizal fungi with varying carbon demands. I am focusing on mycorrhizae formed by three different fungal partners; Suillus, Rhizopogon, and Cenoccocum geophillum. Ectomycorrhizal associations with plants have been shown to range from more to less mutualistic. This spectrum is perhaps an outcome of the varied levels of carbon needed to support different symbionts. All three mycorrhizal fungi were collected from the Rome Sand Plains in Rome, New York. The fungi have been cultured in the lab to inoculate white pine seedlings. These root systems will be examined for mycorrhizal structures and the root tips compared for varied levels of glucose, sucrose, and fructose using the NANAPHID. Suillus and Rhizopogon are both thought to allocate large amounts of carbon to sexual reproductive structures and the growth of extensive mycelial networks. Cenoccocum, alternatively, does not produce any known sexual structures or extensive mycelial networks. Therefore, it is predicted that Suillus and Rhizopogon demand more carbohydrates be delivered to the roots than Cenococcum. Thus, using the NANAPHID technology, higher measurements of nonstructural carbohydrates may be observed in the root systems of trees colonized with Suillus and Rhizopogon than those colonized with Cenoccocum. Data collected using the NANAPHID will be analyzed using ANOVA to compare the variation of nonstructural carbohydrate allocation between trees with different fungal partners.
Using MiSeq on DNA from in-growth bags to observe ectomycorrhizal fungi with N, P and N+P additions in mature forest plots in Bartlett Experimental Forest, NH
- T. Horton
- J. Lamit
- P. Kennedy
- R. Yanai
Forests growing on young, recently glaciated soils are predicted to be N limited in early succession and then become more P limited as they age. Experimental tests of N vs. P limitation in such forest systems are few, and those few have been short-term with very high rates of fertilization. Further, while there has been work on the response of mycorrhizal fungi to N additions, little work been conducted with P. We investigated ectomycorrhizal (EM) fungi in mature forest soils exposed to nutrient additions in the Bartlett Experimental Forest, NH. N (30 kg N/ha/yr as NH4NO3), P (10 kg P/ha/yr as NaH2PO4) and N+P were added annually starting in 2011 in an experiment to investigate multiple element limitation in northern hardwood ecosystems. We used hyphal ingrowth bags to study the response of soil fungi to nutrient additions in three stands. After two growing seasons, the bags were harvested and DNA extracted from the soil. We generated community profiles from these extracts based on the fungal ITS1 region, using the Illumina MiSeq platform. Sequence data were processed using the BBMap package, VSearch 2.5.1 and Qiime 1.9, while FUNGuild was used to identify EMF taxa. As expected, N additions reduced the richness of OTUs identified as EM taxa. P additions did not change the richness of EM taxa compared to the controls, and plots with N+P additions had intermediate levels of richness. Treatment effects were observed in the assemblages of dominant genera, dominance being measured as the total number of sequences recovered. In control plots the three most dominant genera were Tomentella, Tuber and Tomentellopsis. In N plots the three most dominant genera were Sebacina, Tuber and Tomentella. In P plots the three most dominant genera were Genea, Pseudotomentella and Pachyphleous. In N+P plots the three most dominant genera were Paxillus, Tomentella and an unidentified member of the Ceratobasidiaceae. The majority of sequences recovered for a genus in each nutrient treatment were assigned to one OTU in the genus, the exception being Tomentella in control plots with three high abundance OTUs. These data provide a picture of EM fungi in soils as active mycelial networks under the various nutrient treatments. Further, these are some of the first such data from P addition plots and identify some EM taxa that may be functioning in forest P cycling.
Ectomycorrhizal fungal community with conifereous trees in Mt. Seorak
- H. Son
- A. Eom
Ectomycorrhizal fungi (ECM) mainly form a symbiotic relationship with conifererous trees and play a crucial role in nutrient cycling in the forest ecosystem. In this study, we investigated ECM communities of Abies nephrolepis, Abies holophylla, and Pinus koraiensis. Roots of host plants and rhizosphere soil were collected at two sites of different altitudes, higher and lower, of Mt. Seorak, in Korea. ECM were identified using morphological characteristics and sequence analysis of internal transcribed spacer regions of rDNA from ECM root tips. In A. nephrolepis, 9 species from higher altitudes and 6 species from lower altitudes were identified. In A. holophylla, 15 species from higher altitudes and 14 species from lower altitudes were identified. In P. koraiensis, 10 species from higher altitudes and 17 species from lower altitudes were identified. Shannon’s index, species evenness, and number of species of ECM communties were significantly higher for samples collected at the higher altitutes than lower altitudes, and the ECM communities were also correlated with soil characteristics. The results showed that the ECM communities of conifer roots showed differences according to the altitudes and host plants.
Macrofungi communities are shaped by local topography in tropical rainforests of French Guiana
- H. Schimann
- G. Jaouen
- E. Louisanna
- S. Manzi
- A. Sagne
- C. Baraloto
- M. Roy
Current knowledge of tropical Fungi distribution remains patchy, biased towards accessible sites, and extremely limited in the dense and hyper-diverse Amazonian forests. Despite an increasing number of checklists, recent evidence confirms that unknown lineages remain to be described and highlights the need to systematically collect fungal specimens from undersampled areas with standardized inventories. French Guiana remains largely an uncharted territory, with existing collections of Fungi confined to a few coastal localities. Regional geomorphological gradients, as well as local soil types and topographies, are correlated with contrasting floristic composition, providing a unique framework to investigate the diversity and the structure of fungal communities at different scales. We hypothesized that differences in topography and soil found in contrasting forest habitats would shape fungal communities, especially fungal guilds with a high specificity toward their host plant. We evaluated the taxonomic and functional diversity of macrofungi assemblages in three forests habitats distinguished by their local topography - hilltops, seasonally-flooded forests and slope forests; and two types of soils - clay-rich (terra-firme) and white-sand soils. We sampled Basidiomycetes fructifications in 36 plots distributed in four sites across French Guiana. Results showed significant differences between fungal communities growing on clay-rich and white-sand soils and, to a lesser extent, between habitats: seasonally flooded forests harbored slightly different communities than plateau and slope forests. These results suggest that fungal communities show preferences for soils and that the distribution of biotrophic fungi appears to be particularly linked with the topography and appears to be shaped by the distribution of putative host plants. The dataset constitutes an unprecedented and original collection of Fungi for the region, showing an extraordinarily diversity of Basidiomycetes, with more than 253 Genera belonging to 76 Families broadly distributed across French Guianian forest habitats. These data will contribute significantly to record and describe Fungi in the Guiana Shield known to differ from the Amazonia basin by a high level of endemism, unusual tree species composition and different geomorphological landscapes.
Isolation of microfungi from urea-treated litter in ex-situ model
- Q. Ho
- T. Ngo
- D. Vu
- M. Luong
- L. Tran
- A. Suzuki
- H. Pham
Ammonia fungi are a fungal group that forms a fungal community sequentially at the restricted sites of animal materials (e.g., decomposing carcasses and animal waste) or artificial disturbance of nitrogen compounds. Researches about ammonia fungi are fragmentary at global scale, especially sparse in tropical areas as Southeast Asia. In Vietnam, ammonia fungal study was firstly conducted by urea application in Pinus kesiya forest, DaLat, Lam Dong Province from 2010 with a new record of Hebeloma vinosophyllum at Southeast Asia. In 2012, 2 urea treated plots in forest of Pinus dalatensis and Quercus spp. at BiDoup - Nui Ba National Park, Lam Dong Province were observed in 7 months. Amblyosporium botrytis, Ascobolus denudatus, Lyophyllum tylicolor, and Coprinopsis sp. appeared in early phase, followed by Hebeloma lactariolens, Hebeloma sp., Laccaria sp. in late phase of ammonia fungi. For investigating the composition of ammonia microfungi, litter from Pinus and Quercus forests at BiDoup - Nui Ba National Park was applied urea in ex-situ model. The aqueous urea solution was added to litter for setting up the concentration of 40mgN/g dry litter, 20mgN/g dry litter. Based on morphological characteristics and ITS analyses, 14 isolates were identified to genus as Talaromyces, Fusarium and Pseudallescheria. These preliminary results contributed to ammonia fungi data in Vietnam, especially ammonia microfungi data. In further experiments, physiological characteristics of isolated microfungi will be studied.
Physiological characters support to distributional patterns of pinecone fungi Strobilurus spp. in Japan
- H. Pham
- Y. Terashima
There are 5 or more species of Strobilurus recorded from Honshu Island to southernmost of Japan. From the different substrates and climates, Strobilurus spp. in Japan can be divided into 3 groups: group A with basidiomata growing from pinecones in temperate areas such as S. stephanocystis, S. esculentus and S. tenacellus; group B with those growing from pinecones in sub-tropical areas such as the new species S. luchuensis nom. prov. in the Yaeyama area and group C with those not growing from pinecones in temperate areas as S. ohshimae. We aim to know the effects of substrates and temperatures on the growth of S. stephanocystis, S. luchuensis and S. ohshimae as the representatives of the 3 groups above . Four powder substrates from pinecone of P. densiflora, dominate in the Japanese main islands; P. kesiya dominate in Southeast Asia; P. luchuensis dominate in Okinawa; and beech sawdust (Fagus spp.) were used to test the substrate adaptation. Temperature effects were tested on a range of temperature from 5 – 35oC, 5oC difference in each step. Survival tests were applied on all non-growth experiments. In results, S. stephanocystis grew in all substrate but not all experiments. However, S. ohshimae grew well in all substrates except P. densiflora while S. luchuensis did not grow in all substrates except P. luchuensis. On the other hand, all fungal strains died at 35oC and grew weakly at 5oC. S. ohshimae was not able to grow in 30oC and grew weakly in 25oC while S. luchuensis and S. stephanocystis grew well at those temperatures. These results could be explain in that S. ohshimae does not appear in pine forests of main islands and that new species S. luchuensis nom. prov. was adapted to Okinawan pine and climate.
Fungal foraging behaviour and space exploration in a pristine, micro-engineered habitat
- K. Aleklett
- P. Mafla Endara
- C. Arellano
- M. Bengtsson
- P. Ohlsson
- E. Hammer
The soil space is a maze-like habitat for saprotrophic fungi, with walls made of quartz grains and food sources heterogeneously scattered in dead-ends. Because of the micro scale heterogeneity and opaque nature of soils, the hyphal-scale foraging behaviours of these fungi have been intrinsically hard to study and remained largely unknown. However, increasing our knowledge of what their limitations and abilities are could allow us to better understand fungal growth patterns, how fungi contribute to carbon sequestration in soils, and what role soil micro structures play in these processes. In this study, we developed transparent microfluidic chips in which we manipulated the fungal habitat by introducing microstructures. We further examined how fungal traits and growth patterns were affected by this structural heterogeneity at the micro-scale in real time. Our results show proof of fungi’s ability to colonise nutrient free environments, bridge dry pore spaces, and navigate through complex micro-confinements. We also show that even fungal species traditionally classified as having the same nutritional strategy - litter decomposition - express very different growth strategies when colonising pristine habitats, and we present results of how six different species explore restricted spaces and overcome structural heterogeneity (tight channels, nutrient free spaces and obstacles) at the micrometre scale inside the microfluidic chips. When challenged with channels forcing the fungi to turn in angles steeper than 90 degrees, and U-turns, we found that some species cope better than others with re-finding their growth direction or getting out of a dead-end corner. Ecologically, this could mean that in the soil, some litter decomposers would be better at accessing remote soil spaces and resources in cores of complex aggregates, potentially also making a larger contribution towards building a stable carbon pool of fungal necromass in remote soil spaces, protected from other decomposers. To conclude, we believe that the microfluidic devices developed in this study open up for new possibilities to study fungal foraging behaviour and decision making at the resolution of single hyphae, something that could further allow us to decipher the role different fungal species play in shaping the soil environment and whether they all contribute equally to carbon sequestration in the soil.
Which criteria should be considered when appraising ectomycorrhizal communities for forest research?
- C. Perini
- D. Barbato
- S. Maccherini
- P. Leonardi
- A. Zambolnelli
- M. Iotti
- E. Salerni
Forests host a large part of terrestrial biodiversity and provide a wide range of ecosystem services; they regulate local, regional and global climate, store carbon, purify air and freshwater. Plants and their networks with associated ectomycorrhizal fungi play a crucial role in biogeochemical cycles, biodiversity, climate stability and economic growth. Furthermore, ectomycorrhizal fungi are themselves important drivers towards sustainable innovation in many research fields such as food industry, biotechnology, biomedicine and agroforestry. Due to complexity of natural environment, the evaluation of any type of change is often very difficult, since it may not be clear which environmental component will be affected by the stressor, what type of change will occur and what the exposure will be. Before–After Control-Impact (BACI) design overcomes the problem of attributing changes to an impact rather than natural variability. In this context, SelpiBioLife project was established to evaluate the effects of an innovative silvicultural treatment on different biological groups (flora, fungi, bacteria, carabids, nematods and microarthropods) in Pinus nigra plantations. In order to analyze management effects, we adopted a BACI design applied to two study areas, one located in Pratomagno and one in Monte Amiata (Appenines, Italy). Here, our main aim was to demonstrate whether the sampling criteria were appropriate to describe exhaustively the composition of ectomycorrhizal communities, presenting the results of sampling activity before any type of silvicultural treatment. Diversity and abundance of ectomycorrhizal fruiting bodies (EMFb) were determined using mycocoenological analysis. Soil sampling was set up to test ectomycorrhizal root tips (EMRt) community. Morphological structure of each morphotype was examined and molecularly identified by means of a direct PCR approach. Analysis of species richness and composition as well as the effect of spatial scale on EMRt and EMFb communities were assessed using rarefaction technique, permutation tests for multivariate analysis of variance (PERMANOVA), Non-metric multidimensional scaling (NMDS), Mantel’s tests and similarity Percentage Analysis (SIMPER).Rarefaction curves concerning variation in species richness among fungal communities supported the lowest species richness in EMRt respect to EMFb community. PERMANOVA revealed that spatial-topographic factors significantly affected community composition. The Pair-wise t-test showed significant differences between EMRt and EMFb. NMDS confirmed this trend, showing a clear separation between fungal communities in terms of species composition. Furthermore, Mantel’s test resulted in no correlation in distance matrices of community structure. SIMPER analysis indicated that the average dissimilarity between EMRt and EMFb communities was relevant. In conclusion, these results suggest that the adopted sampling criteria are appropriate to exhaustively and quickly appraise the composition of ectomycorrhizal communities for forest research.
Response of microbial community to forest succession in Odaesan National Park, South Korea
- H. Lee
- Y. Jang
- S. Oh
- Y. Lee
- Y. Lim
- C. Kim
- J. Kim
Succession is variation of ecological communities involved by environmental changes. There are many types of succession caused by extreme stress factors such as volcanic eruptions, landslides, and floods, whereas a succession that occurs slowly over time. Odaesan National Park is well-preserved forest located in the Taebaek mountain range in South Korea. The forest succession of the national park is progressing from a mixed-wood forest to a hardwood forest. In this study, the microbial community composition was investigated using 454 sequencing of the soil samples collected from 13 different locations in Odaesan National Park. We assessed whether the communities are affected by environmental factors such as water contetnt (WC), nutrient availability (total carbon and nitrogen) and pH, which were caused by forest succession. As results, water content, total carbon (TC), total nitrogen (TN) and pH were statistically different according to the succession stages of the forest. WC, TC and TN of forest soils tended to increase as succession progressed, while pH tended to decrease. In both succession stages, the bacterial genus Pseudolabrys was most abundant, followed by Afipia and Bradyrhizobium. In addition, the fungal genus Saitozyma showed the highest abundance in the forest soils. The beta diversity of microbial communities was determined by NMDS analysis, which showed a clear discrimination of microbial communities according to forest succession stages, and soil properties (WC, TC, TN, and pH). Furthermore, a network analysis of both bacterial and fungal taxa showed the strong relationship of the microbial community depending on the soil properties caused by forest succession. Further study about functional profiling of each microbial components will help to understand the succession of forest ecology.
Soil depth matters: Communities of bacteria, fungi and micro-eukaryotes structured along soil stratification
- S. Mundra
- L. Morgado
- O. Kjønaas
- H. Kauserud
The boreal forest is a key ecosystem for global C sequestration and storage. Microorganisms in soil have crucial functions in regulating these processes. Fungi are typically sharply structured with soil depth, but we largely lack such information for other microorganism, including bacteria and other micro-eukaryotes. To improve our knowledge of how different microorganisms are structured vertically and how they might interact, we investigated the community of bacteria, fungi and micro-eukaryotes in four different soil horizons in natural birch forests in Western Norway. The communities of all three organismal groups were strongly structured along the vertical depth. Our results support the hypothesis that natural decrease in nutrient availability and pH differences between organic and mineral horizons affect the distribution of soil microorganisms. Proteobacteria, Actinobacteria and Planctomycetes dominated in the uppermost organic layer while Acidobacteria and Firmicutes in mineral layers. Proportionally, fungi dominated in mineral layers whereas other micro-eukaryotes (Metazoa, Apicomplexa, Conosa, Ochrophyta and Chlorophyta) in organic layers. Ascomycota were relatively more abundant in mineral layers compared to Basidiomycota and Cryptomycota. Nematoda, Annelida and Arthropoda showed decreasing trends with depth. Furthermore, separation in the optima of different ectomycorrhizal and saprotrophic genera was observed, supporting the view that different genera are adapted to different niches along the soil depth gradient. Network analyses are used to infer tentative biotic interactions between the microbial groups.
Impact of deadwood on fungal communities in underlying soils
- C. Sadlowski
- C. Minnich
- W. Borken
- D. Peršoh
Fungal communities in soil are crucial to ecosystem functioning by decomposing dead organic matter and supplying plants with nutrients. Composition of the communities is predominantly shaped by edaphic parameters and the local plant community. However, recent studies show that application of deadwood locally alters composition of the fungal communities in underlying soil. While the underlying mechanisms have not been studied so far, it is conceivable that nutrient transfer by fungal hyphae connecting deadwood and soil entails changes in the soil community. However, wood and soil are usually inhabited by different fungi and only few species are known to colonize both habitats. In contrast to this perception, we hypothesized that a considerable proportion of wood fungi may invade the underlying soil. To test this hypothesis, we sampled soil directly under 9-years-old deadwood logs of 13 different tree species at 29 plots in three regions of Germany. Since wood inhabiting fungi are often selective for wood of one or few different tree species, occurrence of such fungi in the corresponding underlying wood would support our hypothesis. Composition of the fungal communities was assessed by Next Generation Sequencing (Illumina MiSeq) of the ITS rRNA gene region amplified from DNA extracts. Analyses of the data confirmed that soil fungal communities are predominantly shaped by geographic location, soil type, and plant community. Our hypothesis was rejected due to the wide absence of wood-selective fungi in the underlying soil. Effects on soil fungal communities are therefore more likely to result from soil fungi invading the deadwood. However, species richness was higher in soils beneath deadwood than in the uncovered control soils. Further implications on soil fungal community structure and potentially underlying mechanisms will be discussed.
Coprinoid coprophilous fungi in Vietnam
- T. Fukiharu
- T. Nguyen
- K. Shimizu
- D. Nguyen
- D. Nguyen
About 125 species have been recognized and recorded for coprinoid agaricales, consisting of Coprinus, Coprinopsis, Coprinellus and Parasola, (Basidiomycota). Among them, about 40 species are known to have coprophilous habit, which appear on animal excrement (2005 Uljé). In Vietnam, 28 species of coprinoid agaricales have been recorded (Patouillad 1910; Trinh Tam Kiet 1998, 2011, 2013) and 6 species of them were coprophilous. We have studied on the coprophilous fungi in Vietnam (around South area). We and our colleagues have collected dungs of wild animals such as elephant etc. in 2015-2016. By the moist chamber method, the emergence of coprophilous fungi was induced, and the fruiting bodies appeared on the substrates were isolated. The collected samples were morphologically and phylogenetically determined. So far, we have identified 8 coprinoid coprophilous species, including 3 new records in Vietnam and 3 new species from Vietnam. These 3 new taxa, designated as Coprinopsis sp. 1, Coprinopsis sp. 2 and Coprinellus sp. 3, are characterized as follows: Coprinopsis sp. 1: This species is similer to C. nivea and C. igarashii, that all share the mealy powdery veil of globose element but differs in the size of veils and basidiospores, collected from a dung of elephant (Elephas maximus); Coprinopsis sp. 2: this species is similar to C. clastophylla in phylogenetically but differs in having globose type veil element; Coprinellus sp. 3: this species is similer to C. marculentus but differs in lacking pileo-, pleuro- cystidia, collected from a elephant dung. The phylogenetic analysis of ITS data also supports these new taxa. In the session we will show the morphological details and phylogenic positions of newly recognized coprinoid coprophilous species from Vietnam.
The genus Coprinellus in Pakistan with the description of three new species
- S. Hussain
Mushrooms with thin-fleshed pileus that becoming plicate on opening, deliquescent gills and dark brown to blackish basidiospores are commonly called coprinoid mushrooms. The genus Coprinellus is one of the important lineage of coprinoid mushroom in Psathyrellaceae. Species-level taxonomy in Coprinellus is based on veil structure and basidiospore morphology. In this study three new species of Coprinellus (C. campanulatum, C. dissminatus-similis and C. tenuis) are described from Pakistan. Species description are based on morphological and molecular data. Phylogeny based on nuc rDNA region encompassing the internal transcribed spacers 1 and 2 along with the 5.8S rDNA (ITS) show that the new species C. campanulatum sp. nov. and C. dissminatus-similis sp. nov. are clustered in a clade formed by the members of section Micacei; C. tenius sp. nov. falls in section Domestici of genus Coprinellus. Morpho-anatomical descriptions of the new species and comparison with closely allied taxa are provided. With this study the number of known species of Coprinellus in Pakistan reached to 7.
Taxonomy and Phylogeny of genus Lycoperdon from Pakistan
- N. Yousaf
- A. Khalid
- A. Niazi
Genus Lycoperdon was first established by Persoon in 1796 by describing L. perlatum as a type species. It is characterized by sub globose to pyriform basidomata with conspicuous sterile base and dehiscence by an apical pore. It is represented by 54 species worldwide. Recent molecular phylogenetic studies have widened the concept of the genus and new limits of Lycoperdon have been established by proposing many subgenera within the genus, i.e. Apioperdon, Bovistella, Lycoperdon, Morganella, Utraria and Vascellum. In Pakistan, twenty one (21) Lycoperdon spp. have been reported so far. During this investigation, eight (8) Lycoperdon species have been identified and described using morphological and molecular methods based on ITS-nrDNA region. Among these, three (3) belong to subgenus Vascellum, two (2) each to subgenera Bovistella and Lycoperdon and one (1) to subgenus Apioperdon. Out of these, three (3) species have been found previously undescribed viz., L. lahorense, L. olivoflavum, L. parvisporum, and reported here as new species. L. utriforme is a new record for Pakistan and three (3) species have been reported from new localities of the country. One taxon, previously published as Bovistella japonica on morphological basis has been shifted to subgenus Bovistella of genus Lycoperdon based on molecular analysis in this study.
Diversity and phylogeny of Leucoagaricus and Leucocoprinus (Agaricales, Basidiomycota) in the Neotropics
- A. Justo
- C. Angelini
- A. Bizzi
Among the lepiotaceous fungi in the Agaricaceae the Leucoagaricus/Leucocoprinus clade is particularly diverse in the tropics, with a lot of species awaiting discovery and formal description. The major mycofloristic studies of this group in the American tropical region date back to before the molecular era, and most of the molecular data currently available for this area come from the studies on ant-associated fungi and their free-living relatives, that do not focus on taxonomy. Over the past ten years, one of the authors (C.A.) has collected and studied fungi in the Dominican Republic. More than 300 species of macrofungi have been recorded, and almost all voucher specimens are deposited in the herbarium of the Jardin Botánico Nacional Dr. Rafael Ma. Moscoso (Santo Domingo, Dominican Republic). Approximately 20% of the collections represent lepiotaceous fungi of different genera (Chlorophyllum, Cystolepiota, Lepiota, Leucoagaricus, Leucocoprinus) and are currently being studied and sequenced. In this contribution we studied the species of Leucoagaricus and Leucocoprinus present in the Dominican Republic, using morphological and molecular methods, carefully comparing all our collections to the previously described Neotropical taxa. Seven species of Leucoagaricus (La. bulbiger, La. margaritifer, La. peglerii, La. roseovertens, La silvestris, La. stillatus, La. turgipes) and three of Leucocoprinus (Lc. antillarum, Lc. fuligineopunctatus, Lc. microlepis) are proposed as new. Additional records of previously described taxa are also discussed, including the first confirmed occurrence of La. rubroconfusus in its putative natural habitats.
The genus Agaricus in the Caribbean II. Phylogenetic placement of collections from the eastern Greater Antilles using multigene sequences analyses
- B. Ortiz Santana
- D. Lodge
- R. Kerrigan
- L. Parra
- P. Callac
- C. Angelini
- J. Chen
The taxonomy of the genus Agaricus was recently evaluated and reconstructed using multigene (ITS, LSU, TEF, RBP2) analyses of collections from temperate zones (mostly Europe and United States) plus tropical areas of Africa, Asia and the Americas, including some material from the Caribbean region. Six subgenera and 23 sections were proposed under this new taxonomic arrangement of infrageneric taxa in Agaricus. Following the same approach, we conducted phylogenetic analyses using ITS, LSU and TEF data to determine the placement of additional Caribbean collections within this new arrangement. This research was initiated when only ITS sequences were available for comparison, but now with tropical data available for ITS, LSU and TEF we were able to compare 29 collections from Puerto Rico, 11 from the Dominican Republic and 7 from the Virgin Islands in a multigene analysis. Among the studied collections, we found representatives of all 6 subgenera and 13 sections, including sections recently described from material collected in the Dominican Republic. Most of the collections studied fell within A. subg. Minores, A. subg. Spissicaules and A. subg. Pseudochitonia, but since there are not multigene data available for several species of the sections in these subgenera, further studies including more temperate species are needed to define their closest relationships. We also placed collections in sections Agaricus, Leucocarpi, Minores, Rarolentes, Subrutilescentes and Xanthodermatei; these collections may represent new species, which we will investigate further using morphological comparison. In the present study, morphological and multigene data have confirmed the presence of A. bisporus, A. californicus, A. endoxanthus, A. martinicensis, A. pocillator, A. subrufescens and the two recently described A. lodgeae and A. porphyropos in the Caribbean region.
New taxa of Agaricomycotina from Brazil
- T. Gibertoni
- C. Sousa De Lira
- R. Chikowski
- R. Alvarenga
- A. Soares
- A. Ottoni
- P. Alves
Agaricomycotina is a large group of mostly macroscopic Basidiomycota, usually known as mushrooms, jelly fungi, boletes, earth-stars, corticioid fungi, polypores, clavarioid fungi among others. During the last 20 years, efforts of collecting and identifying them in Brazil have been undertaken and were reinforced in the past 5-10 years. The field trips were mostly conducted in the Atlantic Rain Forest, Amazonia and Cerrado in North and Northeast Brazil and several new taxa are being described using morphological and molecular characteristics. In the corticioid fungi, we present one new species of Amyloathelia, three of Botryodontia, two of Byssomerulius, two of Gloeocystidiellum, 13 of Hyphodontia, two of Luteoporia, one of Lyomyces, one of Meruliopsis, four of Phlebiopsis, one of Resinicium, one of Sistotremastrum, one of Subulicystidium, five of Trechispora, two of Vararia, two of Xylobolus, one of Xylodon, and two new genera. Among the jelly fungi, two new species of Calocera, two of Dacryopinax, two of Tremella and two new genera are introduced. In the poroid fungi, we present one new species of Henningsia and one of Ceriporia and three new genera; additionally, two synonyms are solved, one confirmed and one old name is recovered. Among the clavarioid fungi, one new species of Clavulinopsis and one of Ramariopsis are introduced. The results indicate the high, but still undiscovered mycodiversity in the Brazilian forests and also that the addition of Brazilian specimens in the phylogenies improves the delimitation of taxa.
Phylogeny overview of Hygrophoraceae from China
- C. Wang
- T. Li
Hygrophoraceae, a large, attractive and divers family in Basidiomycetes, includes mainly the agaricoid white-spored mushrooms with waxy pileus and gills, some basidiolichens and some corticioid fungi. Some important morphological characters of its members are often very susceptible to environments, so it is challenging to do morphological recognition, especially for the Chinese taxa since the knowledge of Chinese Hygrophoraceae is still very limited and there is no monograph about them. Therefore, it is significant to do a systematic study based on Chinese materials. In the past seven years, with over 200 Hygrophoraceae fresh samples collected from China, three gene fragments (ITS, LSU, RPB2) were used to reconstruct the phylogenetic relationships of the family Hygrophoraceae based on both newly generated and downloaded sequences. In the results, phylogenetic framework of worldwide Hygrophoraceae shows it as a monophyly family with three monophyly subfamilies, conforming to the subfamily concepts of the previous studies without Chinese data. Under subfam. Hygrophoroideae, genera Hygrophorus and Haasiella are strongly supported as sister groups, while Chrysomphalina is the basal group. The division of subfam. Hygrocyboideae into three tribes is accepted: tribe Hygrocybeae is made by the largest genus Hygrocybe and the rough-spored genus Hygroaster; under tribe Humidicuteae, Humidicutis and Gliophorus are confirmed as sister groups with high support value, Porpolomopsis is close to them, while Neohygrocybe is located at the base of the tribe; and tribe Chromosereae is a basal group of the subfamily, including two sister genera Chromosera and Gloioxanthomyces and a new genus from China. Multigene phylogenetic analysis shows that China is rich in Hygrophoraceae resources: 13 known genera and a new genus are present, over 60 species can be recognized and at least 30 of which are new to science. Although the recognized taxa can constitute the basic categories of a monograph, more samples and sequences are still needed in order to make a more comprehensive monographic study on Chinese Hygrophoraceae. This study was financed by the National Natural Science Foundation of China (Nos. 31170026, 31370071, 31493011).
Taxonomy and molecular phylogeny of bioluminescent mushrooms in Taiwan
- C. Chang
- C. Chen
- H. Kao
Bioluminescent mushrooms are capable of emitting light from their basidiocarp, mycelia or both. Eighty-nine species of bioluminescent mushrooms have been recorded worldwide. Twelve of them were found in Taiwan. In this study, specimens were recently collected from mountainous areas in Taiwan, and were identified by both morphological characters and sequences of internal transcribed spacers (ITS). ITS-based phylogenetic tree was inferred by maximum likelihood (ML) and Bayesian inference (BI) algorithms. In this study, 15 species of bioluminescent mushrooms were collected, including three new species of genus Mycena and four new records (M. deeptha , M. stellaris, Omphalotus japonicus and Panellus luxfilamentus). The phylogenetic analyses showed that the three new species formed a monophyletic group respectively. Key to bioluminescent mushrooms from Taiwan and morphological characters of the three new species were provided. The report of this study brings the total numbers of bioluminescent mushrooms worldwide to 92 species.
The white elephant in the black box: characterizing ITS copy number variation and its influence on fungal metabarcoding studies
- L. Lofgren
- J. Uehling
- F. Martin
- P. Kennedy
High-throughput (HTS) amplicon-based ‘metabarcoding’ studies of fungi frequently target the Internal Transcribed Spacer (ITS) region, producing millions of reads from a single sample. These reads are frequently used for both species identification and to quantify relative abundance by using read abundance as a proxy for species abundance. The ITS region is multi-copy and the number of ITS copies may vary between species, altering the amount of template DNA available for amplification. The vast differences between observed and expected read abundance in fungal mock communities have been attributed, in part, to potential differences in ITS CNV. However, the actual CNV between fungal species, and its relative influence on HTS read abundance is largely unknown due to the difficulty in discerning ITS copy number using molecular techniques. Here, we investigate ITS CNV using a genome-based in silico read depth approach, applied to 21 species of ectomycorrhizal fungi, and characterize the functional consequences of ITS CNV in HTS datasets using mock community analysis over both the ITS1 and ITS2 regions. We report that ITS CNV varies over an order of magnitude between closely related species with no indication of phylogenetic conservation. However, HTS read abundance shows no clear correlation with ITS CNV. Instead, the primary source of variation in sequence abundance in HTS datasets appears to be associated with the process of PCR, with inhibition or preferential amplification of certain lineages in samples containing mixed communities.
International Collection of Microorganisms from Plants (ICMP)
- B. Weir
- M. Petterson
The International Collection of Microorganisms from Plants (ICMP) is New Zealand’s national culture collection of living Bacteria, Fungi, and Chromists. The collection and associated databases considered ‘Nationally Significant’ by the government and is in part publicly funded.The ICMP holds 20,000 cultures predominantly from plant, soil, and water in the natural environment, as well as important reference and type cultures of the world’s plant pathogenic fungi and bacteria. All cultures are databased and available online at cultures are available for a fee to cover retrieval costs. New accessions into the collection are welcome, and recommended when publishing papers on microbes to provide a stable permanent resource for future researchers. The cultures are preserved under liquid nitrogen or in freeze dried ampoules. The ICMP containment and transitional facility conforms to enhanced PC2 Containment criteria, with generic permits to import quarantine and unwanted organisms into New Zealand.
Describing and communicating species
- U. Kõljalg
Describing and communicating species are fundamentally different approaches and can be referred to as a “The two faces of taxonomy”. Here they stand for Karl Popper and Carl Linnaeus.
According to Popper, scientific theories or hypotheses must be falsifiable. It means all data and criteria used for species descriptions must be presented and available for scrutiny – they must, in short, be reproducible. Otherwise we can’t falsify species as hypotheses, which according to Popper, would leave them unscientific. In digital era this means that all data (anatomical, genetical, etc.) of all specimens or individuals of the described species must be available as open data. Even better would be if these data are available in machine readable format, which makes reanalyses quick and comparable over time. There are already repositories that provide services to publish species datasets in such formats. Prior to the digital era, the “face of the taxonomy” was secured by listing all studied specimens lodged in public collection(s). In order to falsify described species, taxonomists reanalysed those specimens through loans or by visiting collections. In conclusion, high quality species descriptions were and are scientific when they make data open.
Once a species has been described, the second “face of the taxonomy” - communication - will enter the stage. Communication of species is backed by the Code, and Carl Linnaeus is the face indeed. The Code is built for communication of species only and does not govern their description. It is a brilliant example how communication must be built. However, the digital era and the availability of DNA sequences combine to change the way we describe species. They are also changing the way species are used by the research community in metabarcoding and other studies. It is obvious that current Code should follow these advances in order to serve us today and in future. But what has to be changed in the species communication system?
There is a proposal to allow DNA sequences to be served as a type of the species. Proponents argue that this change in the Code will allow us to describe and communicate species based on DNA sequences only - and that this will solve the major problems were are facing in this field.
In this talk I will demonstrate that this change will not solve current challenges in taxonomy. An alternative way how we need to develope Code in order to serve modern taxonomy will be presented.
Updating the status of global fungal red-listing and a call for participation
- A. Dahlberg
- G. Mueller
Species of fungi are not immune to the threats that put animal and plant species at risk, i.e., habitat loss, loss of symbiotic hosts, pollution, over exploitation, and climate change. Yet, fungal conservation is only now receiving significant attention. So, it is not surprising that fungi have rarely been included in broader conservation discussions, policy decisions, or land management plans. A critical way to help politicians and citizens be more aware of the importance of fungi and the need to conserve them is to have fungal species included in the IUCN (International Union for Conservation of Nature) Global Red List. The Red List is a compilation of rigorous assessments of the extinction risk of individual species made using strict universal criteria and categories (www.iucnredlist.org). Until recently, some thought that it was not possible to rigorously assess the conservation status of fungal species using IUCN criteria because of the unique biology of fungi and insufficient information on their taxonomy, distribution and ecology. However, much progress has been made to address these challenges.
The voluntary Global Fungal Red List Initiative aims to facilitate and coordinate efforts by the global mycological community to get species of threatened fungi assessed and included in the global IUCN Red List. The goal of the initiative is to raise awareness of fungal conservation among mycologists, the conservation community, policy makers and the general public. By the end of 2018, more than 100 fungi will be in. A broader engagement by the mycological community is needed to keep this initiative moving forward with the goal of having a sufficient number of species assessed to provide an indication of the conservation status of fungi in relation to other groups of organisms. The initiative needs additional contributors with knowledge of distributions, ecologies and population trends of individual species as well as help with checking facts and suggesting species to assess. Gaps in our knowledge of fungal diversity, distributions, phenology, and responses to threats will continue to pose significant challenges to fungal conservation initiatives for the foreseeable future. However, we have sufficient knowledge on an increasing number of species to enable fungi and mycologists to play a larger role in regional, national, and global fungal conservation activities. Contribute by contacting the Global Fungal Red List Initiative or directly through their web-page.
A standard protocol for selecting rare or threatened lichens for IUCN red listing: Results from trial implementation in North America
- J. Lendemer
- J. Allen
- T. McMullin
Lichens are ecologically important and threatened by diverse forces worldwide. Yet their conservation has long been hampered by a widespread perception that 1) species are poorly known, 2) cannot be readily identified by non-experts, 3) direct threats are poorly documented/not identifiable, and 4) immediate actionable conservation items are unclear or impossible to implement. We will present a standard protocol that outlines how species can be selected for IUCN Red Listing and specific criteria that should be used to rank species as priorities for IUCN assessment (vs. those requiring further study). This protocol and associated criteria will be placed in the context of the first IUCN Red List assessment workshop for North American lichens that was held in New York in 2016.
An examination into the efficacies of various genomic analyses using Ophiocordyceps camponoti-floridani
- H. Christensen
- I. Will
- A. Forsman
- A. Brachmann
- R. Ohm
- C. De Bekker
Entomopathogenic fungi infect insects of several orders. Some of these fungi have the ability to manipulate the behavior of their insect hosts. Such behavior manipulating fungi can be found in the family Ophiocordycipitaceae, and more specifically the genus Ophiocordyceps. This genus contains many species that induce behavioral manipulation in ants, however many species are still undescribed. In the Southeastern United States, a recently discovered endemic species, Ophiocordyceps camponoti-floridani, infects and manipulates the native, abundant Florida Carpenter ant. This manipulation occurs with a light-induced biting mechanism that allows the fungus to complete its life cycle. Gaining a better understanding of this species, such as identifying potential genes associated with secreted bioactive compounds, is integral to our understanding of how this parasite is able to control the behavior of its host. To this end, we have isolated the fungus from an infected Florida Carpenter ant collected from Central Florida, and sequenced the genome of O. camponoti-floridani using two sequencing platforms: Illumina MiSeq and Nanopore MinIon. Due to the relatively new field of genomics, there are many different approaches to assemble, annotate and analyze the raw data. A stepwise pipeline that leads to an annotated genome can thus combine a variety of approaches, with one approach being more efficient and/or effective than the other. For each of these steps, we have examined several de novo assemby and annotation methods and parameters. These results were compared to data of previously produced Ophiocordyceps reference genomes. The various assembly and annotation outcomes are then verified through a completeness analysis. This eventually helps us create the most efficient pipeline for obtaining future de novo assembled and annotated genomes of related, novel Ophiocordyceps species. The most complete O. camponoti-floridani genome is subsequently further analyzed by the identification and quantification of various annotated gene functions, focusing on identifying genes that could be contributing towards neurological and immunological regulation/domination on the host species. Our work thus contributes to a better understanding of parasitic manipulation of host behavior by a fungal species, while also generating a robust next-generation sequencing pipeline.
Botryosphaeria spp. and Phomopsis spp. Causing Leaf Blight and Necrotic Spots on Rambutan (Nephelium lappaceum L.) in Puerto Rico
- A. Aviles Noriega
- L. Serrato-Diaz
- L. Rivera Vargas
- P. Bayman-Gupta
Rambutan (Nephelium lappaceum L.) is a tropical fruit tree that has been cultivated since 1998 on commercial orchards in Mayaguez, Puerto Rico. From 2003 to 2015, during a disease survey, leaf blight and necrotic spots were observed at commercial and experimental orchards throughout the island. Diseased leaves were disinfected and plated onto potato dextrose agar (PDA). Four isolates, A1 and A2, of Botryosphaeria spp. and A3 and A4, of Phomopsis spp. were purified and identified using taxonomic keys and DNA GeneBank sequence comparison. PCR amplifications of the ITS1-5.8S-ITS2 region and partial sequence of elongation factor 1-alpha (EF1-α) genes were used to support the identification. PCR products were sequenced and compared using BLASTn with other sequences of Botryosphaeria spp. and Phomopsis spp. submitted to the NCBI GenBank. Pathogenicity test was conducted on eight Rambutan trees, using three healthy non-detached leaves per isolate. Trees were inoculated with two separate or combined isolates for each leaf with 5mm mycelial disks from pure cultures grown on PDA. Leaves were kept in a humid chamber using plastic bags for 8 days under greenhouse conditions. Two of eight trees were untreated, inoculated with PDA disks only, and were used as controls. The test was repeated twice. Eight and 14 days after inoculations (DAI) isolates of Botryosphaeria spp. caused leaf blotch and leaf blight, respectively. For all isolates, diseased leaves turned from light brown to dark brown starting from the apex and spreading through the lamina with necrotic tissues ranging from 10mm to 40mm in leaf length. Only one isolate of Phomopsis spp. caused necrotic spots on the leaves of 5mm in diameter. To prove the pathogenicity test, Botryosphaeria spp. and Phomopsis spp. were re-isolated from diseased leaf, fulfilling Koch's postulates. Untreated controls showed no symptoms and no fungi were re-isolated from tissue. By having identified the fungal pathogens involved in leaf blight, a more specific management approach can be implemented against Rambutan tree diseases in Puerto Rico.
Inhibition of seedling establishment of soybean by Phomopsis longicolla Hobbs
- K. Tomioka
- A. Kawaguchi
- A. Masunaka
- K. Nomiyama
- H. Sekiguchi
- S. Mori
- R. Yamazaki
- Y. Kawasaki
- K. Katayama
Severe wilt with stem blight of soybean has been occasionally come across in our trial fields located in Hiroshima Prefecture in western Japan throughout the months of June until July since 2012. Leaves initially droop down to fade in color. The whole plants gradually wilt as turning brownish, and then wither entirely. Subglobose, dark brown to black pycnidia of a fungus frequently appear on lower parts of stems of the diseased plants. White to pale beige masses of conidia come out from pycnidia on the stems under moist conditions. Four representative isolates of the fungus obtained by single-conidium isolation (isolates A3, Fuk, Sac and Sta) produced pycnidia on cuts out of stems of soybean or common bean on potato dextrose agar (PDA) at 25ºC under irradiation with black light. The isolates grew on PDA in the dark at 5-35ºC with maximum growth of 2.9-6.1 mm/day at 25-28ºC. No sclerotia, chlamydospores or teleomorphs were found with the isolates. The isolates were identified as Phomopsis longicolla Hobbs (Hobbs et al., 1985) from the morphological and cultural characters. For isolates A3, Fuk and Sac, the identification was supported sequences of rDNA-ITS amplified with primer pair ITS4 and ITS5 (White et al., 1990). No sequence data of rDNA-ITS could be obtained for isolate Sta. Mycelia of each isolate cultured on PDA at 25ºC in the dark for 6 days were suspended in distillated water, and incubated with seeds of soybean (cv. Sachiyutaka) on petri dishes at 24-26ºC in the laboratory. Seeds incubated with distillated water were served as controls. Seed decay, budding delay and/or sprout decay occurred in the incubation with respective isolates. Control seeds healthily germinated and grew, showing compatibility of the isolates with soybean. When seeds dipped in the same suspension of isolate Sac were planted in pots with normal nursery soil, growth delay of seedlings without stem blight was recognized. Part of them recovered. Seed decay of soybean by P. longicolla has been reported (Hobbs et al., 1985). This has also been recorded in Japan (Sato et al., 1989), and compatibility of the fungus with stems of soybean has been noted in the report. After that, stem blight of the plant by P. longicolla has been reported from China (Chen et al., 2013; Cui et al., 2009). The present symptom in Japan is likely to be due to P. longicolla, though pod and stem blight or bean sprout rot of soybean by Phomopsis phaseoli (Desm.) Sacc. [Syn. Diaporthe phaseolorum var. caulivora Athow & Caldwell, Diaporthe phaseolorum var. sojae (Lehman) Wehm., Phomopsis phaseoli var. sojae (Lehman) Sacc.] have been recorded in Japan (Goto, 1925; Sato et al., 2014). We have not yet led to clear-cut reproduction of the present symptom in Japan by inoculation with the isolates, and will retry it as considering conditions of inoculation tests. The present isolates were deposited to Genetic Resources Center of National Agriculture and Food Research Organization, Japan, with accessions MAFF150072-150075, and DNA sequence data obtained here were registered in the DDBJ/EMBL/GenBank databases as accessions LC333208-333210.
Identification of Gliocephalotrichum spp. associated with post-harvest rot of fruits in Brazil
- C. Almeida
- A. Reis
- D. Pinho
The brazilian Cerrado has a large quantity of endemics plants and until now more than 150 new species of fungi have been described in association with hosts belonging to several botanical families. Although few studies have been carried out to identify species causing post-harvest rots, several diseased fruits have been observed in the plants of this biome. Thus, this study aimed to identify the fungal species associated with post-harvest rot of different hosts in brazilian Cerrado. The isolates were obtained from fruits rots of Caryocar brasiliense (Pequi), Syzygium jambos (Jambo amarelo), Syzygium cumini (Jamelão), Spondias purpurea (Seriguela), Spondias mombin (Cajazinho), Dypsis madagascariensis (Areca de locuba), and Roystonea sp. Only Gliocephalotrichum-like isolates were selected for identification. The morphological characteristics are conidiophores septate, hyaline, erect, consisting of a stipe and stipe extensions subtending a penicillate conidiogenous. The conidiogenesis is phialidic, with phialides hyaline and producing conidia cylindrical, hyaline and aseptate, accumulating in a mucilaginous mass. Total genomic DNA was extracted from cultures grown on 2% malt extract agar (MEA) for 7 d, using the Wizard® Genomic DNA Purification Kit. The partial region of the translation elongation factor 1-alpha (TEF) gene was amplified and sequenced using the primers EF1F and EF2R. The nucleotide sequences obtained were compared with sequences of type species and specimens available from GenBank. From the phylogenetic analysis by Bayesian inference, it was possible to identify five different species of Gliocephalotrichum. A total of 39 isolates were obtained, and from these, 22 specimens from C. brasiliense show absent stipe extensions and grouped in a well-supported clade that is phylogenetically distant from the other species while 3 isolates grouped with specimens of G. longibrachium. The isolates from S. cumini and S. jambos grouped in a phylogenetically distinct clade, representing a possible new species too. G. simplex was found associated to D. madagascariensis while Gliocephalotrichum bulbilium was obtained associated to Roystonea sp. (n=1), S. cumini (n=2) and S. jambos (n=5). This study reveals new hosts for G. longibrachium, G. bulbilium and G. simplex and the discovery of two possible new species, which will be described according the current International Code of Nomenclature for algae, fungi, and plants. Financial support: FAPDF, CNPq and UnB.
Phytopathogenic fungi affecting Mentha nemorosa in Puerto Rico
- J. Santiago
- L. Rivera Vargas
- L. Simbaña
Mentha nemorosa, an aromatic herb known in the Caribbean as “yerbabuena”, belongs to the Lamiaceae family. This species is valuable for its aromatic qualities and culinary use in dishes and tropical drinks. For the fiscal year 2014-2015, the cultivation of aromatic plants in general, contributed $334,000 to the revenue of Puerto Rico. Although this plant is produced under controlled conditions in greenhouses, it is affected by pathogenic microorganisms that cause yield losses. The most common symptoms observed were necrosis of leaves and stems; and chlorotic spots on leaves. Root and delayed plant growth were also observed. Therefore, the objective of this study was to identify phytopathogenic microorganisms of M. nemorosa. Symptomatic plants were collected, and microorganisms were isolated on PDA. Koch postulates were completed using healthy plants in a humid chamber. After a week, most of the isolates produced chlorosis on inoculated leaves. Interveinal necrosis was observed after two weeks. Using morphological keys, two pathogenic fungi were identified: Colletotrichum spp. and one isolate belonging to the Botryosphaeriaceae’s family. Colletotrichum sp. colonies were grey with scant aerial mycelium towards the center of the plate and scattered ooze of orange conidial masses. Conidia measured 4.9 x 17μm. The species was molecularly identified as C. queenslandicum after DNA analysis of rDNA ITS region, actin, alfa-elongation factor, β-tubulin, and glyceraldehyde-3-phosphate dehydrogenase (GADPH) genes. Our isolate showed 95 to 98% of homology with C. queenslandicum sequences from Genbank. The isolate belonging to the Botryosphaeriaceae’s family showed 98 to 99% homology with Lasiodiplodia parva using rDNA ITS region, β-tubulin and elongation factor genes. On PDA, Lasiodiplodia parva colonies had initial greenish grey mycelia that turn white in the margins of the means, but sporulation was not achieved. This information is very valuable to farmers when considering effective disease management practices under greenhouse conditions.
Characterization of secondary metabolite production among Cercospora species associated with Cercospora Leaf Blight
- J. Searight
- V. Doyle
- J. Solorzano
Cercospora is a species-rich genus of phytopathogenic fungi with a cosmopolitan distribution and causes disease on a wide range of host species, including many of economic importance. Among the diseases caused by Cercospora spp. with a major economic impact in North and South America is Cercospora Leaf Blight (CLB) and Purple Seed Stain (PSS) of soybean (Glycine max). Recent work has shown several species are associated with CLB and PSS, but typically a single species dominates in a given geographic region. For example, C. kikuchii, C. cf. sigesbeckiae, and C. cf. flagellaris have been associated with CLB and PSS in Louisiana, but a single species, C. cf. flagellaris, dominates in field populations. Cercosporin, a mycotoxin produced by several species of Cercospora, is an important virulence factor capable of increasing disease extent and severity. However, there is variation in the quantity of cercosporin produced among strains within a species and among species and high cercosporin-producing species and isolates may have a competitive advantage under certain conditions. The aim of this study is to quantify cercosporin production in several strains of Cercospora and assess variation among and within species. Several isolates from each of three species associated with CLB and PSS in Louisiana were characterized for cercosporin production corrected for differences in growth rate. The cultures were monitored for fifteen days with growth measurements taken every three days. Media variants included potato dextrose agar (PDA), minimal medium with ground soybean leaf, and complete medium. Cercosporin production was quantified by placing mycelial plugs from the margin of actively growing cultures in four milliliters of five molar potassium hydroxide and measuring absorbance at 480 nm. The three species responsible for CLB and PSS show variation in cercosporin production across media. All species produced minute amounts of cercosporin (1-2µM) on soybean leaf minimal medium and complete medium. Cercosporin production on PDA was 5.5 to 29.9 times greater than that produced on other media. Cercospora flagellaris produced the highest average cercosporin concentration (25.45 µM), C. sigesbeckiae produced 16.65 µM, and C. kikuchii produced 7.27 µM. While there is significant variation in cercosporin production within species, an inverse correlation was observed between cercosporin production and relative standard deviation. The relative standard deviation in Cercospora flagellaris was 61%, while C. sigesbeckiae and C. kikuchii were 73% and 78%, respectively. We will characterize the range in cercosporin production within the genus using ex-type strains from an additional 24 species. Our results suggest there is significant variation in cercosporin production both within and among species and characterizing Cercospora isolates for cercosporin content is sensitive to the conditions under which the phenotyping is being done.
Phytophthora nicotianae causing heart rot of pineapple (Ananas comosus) in Puerto Rico
- L. Simbaña Carrera
- Y. Vélez Negrón
- L. Rivera Vargas
Pineapple is an important crop in Puerto Rico, representing 5 million dollars of annual revenues for the economy of the island. Phytophthora spp. strikes pineapple production and is one of the most important pathogens for local farmers. Worldwide, ‘Heart rot’ is an important disease caused by P. nicotianae and has increased concern among producers who have unsuccessfully applied chemical fungicides for its control. The excessive use of the only chemical fungicides registered in Puerto Rico (i.e. Metalaxyl and Fosetyl Aluminium) has increased pathogen resistance, redounding in an ineffective control. Infected fields were evaluated every month for a year to characterize oomycetes and fungal species from five pineapple production areas of the island. During the rainy season from February to May, 2017, a 40% disease incidence of ‘heart root’ was observed at the southwestern region of Lajas. In Guanica, 75% disease incidence were observe in all plots sampled. Symptoms were chlorosis, necrotic leaf tips and death of many young plantlets. Diseased tissue sections were transferred to V8 PARPH agar. Two P. nicotianae isolates were identified using taxonomic keys from tissue samples collected at Guanica, Puerto Rico. Sharpy papillated, ovoid sporangium measuring 32 x 29µm with abundant chlamydospore production were observed. Sequences of rDNA ITS region and COX gene showed 99% of homology with data of P. nicotianae obtained from Gen Bank, confirming our morphological identification. Pathogenicity tests were conducted in vitro using tissue culture pineapple plantlets. Fourteen days after inoculation, a heart soft rot was clearly observed. We speculate that the expression of symptoms caused by this pathogen were directly related to high precipitation occurring during April, 2017 with 24.43mm. In seasons of low rainfall, from September to December 2017, P. nicotianea was not expressed. This study allowed us to assess the distribution and expression of pineapple heart rot disease during the production cycle to improve disease control. Providing a better idea about the disease management for the pineapple productors for the season.
Root-invading fungus of Vicia sativa at different growth stages
- L. Ma
To explore Root-invading fungus of Vicia sativa at different stages, providing scientific basis for Vicia sativa disease prevention and control work , Use Lanjian1､2､3 and 333 / A four kings of Vicia sativa varieties as materials, at Vicia sativa seedling, branch, flowering and mature period sampling in 2017.Through isolating root invasion fungi by conventional tissue separation method, and identificating by morphological and ITS sequence analysis. The results showed that: (1) A total of 23kinds of root invading fungi were distributed from four kings of Vicia sativa varieties throughout the growing season, and eight 8 kinds of root invasion fungi were 4 kinds of Vicia sativa varieties shared, including: Microdochium tabacinum, Fusarium acuminhtum, F. tricinctum, F. acuminatum, F. solani, F. trichothecioides, Rhizoctonia solani and Phoma multirostrata. Both R.solani and F. acuminatum can be isolated during the whole growth period of four kinds of Vicia sativa varieties. (2)The dominant population of root invasing fungus are different , but with little difference at different growth period.(3) The carrier rate of root segment With the growth period is "low - high - high - low" trend. (5) Pathogenicity testin doors by found that: Scytalidium thermophilum, Mortierella alpine and Oidiodendron cerealis have no pathogenicity.The five most pathogenic fungus are: F. avenaceum, F. oxysporum, F. acuminhtum, Myrothecium roridum, and Clonostachys rosea. Followed by R. Solani and F.tricinctum.
Keyword: Vicia sativa; Different growth period; Percentage of root fungi; Pathogenic
Diversity of powdery mildew fungi (Ascomycota, Erysiphales) in Azerbaijan
- L. Abasova
- D. Aghayeva
- S. Takamatsu
Powdery mildews are an economically important group of fungi. They cause serious damage on about 10.000 angiosperms including many crops, vegetables, fruits, cereals, and ornamental plants worldwide (Braun, Cook 2012; Amano 1986). Biodiversity of this fungal group has been explored well in temperate regions of the Northern Hemisphere, such as Europe, North America and East Asia. Taxonomy of this fungal group has significantly changed during the last 20 years. Investigations of powdery mildews in Azerbaijan began in middle of the last century during exploring of mycobiota of the country, when comprehensive numbers of samples were collected and numerous taxa from Erysiphales were recorded. However, the exploration of powdery mildews in Azerbaijan was largely neglected in the past 30 years, because other fungal groups were in the focus of interest. The purpose of the present study was to conduct comprehensive investigations on powdery mildews, in order to identity and clarify taxonomy of the preserved herbarium collections in the Mycological Herbarium of the Institute of Botany, ANAS, and newly collected samples by application of modern morphological and molecular approaches. Samples were examined by means of molecular and morphological methods. Molecular analyses were done by using chelex method according to Meeboon and Takamatsu (2015). Morphological examinations were conducted under the optical microscope, using the lactic acid method (Shin, La 1993) and 3% NaOH for observations of asexual and sexual stages, respectively. Re-examination of herbarium samples and identification of new collections revealed 120 taxa from nine genera (Erysiphe, Podosphaera, Sawadaea, Phyllactinia, Leveillula, Golovinomyces, Neoërysiphe, Arthrocladiella, Blumeria) distributed on more than 400 plant species in Azerbaijan. Erysiphe is the largest genus consisting of 43 taxa, followed by genus Podosphaera with 22, and Golovinomyces with 21 species respectively. The genera Phyllactinia and Leveillula, each compile 12 species. Remaining genera include one or two species. During our study one new species – Erysiphe azerbaijanica Abasova, Aghayeva & Takamatsu was described from the Microsphaera lineage, and five species (Erysiphe arcuata U. Braun, E. corylacearum U. Braun & S. Takam., E. quercicola S. Takam. & U. Braun, E. syringae-japonicae (U. Braun) U. Braun & S. Takam., E. viciae-unijugae (Homma) U. Braun) were new powdery mildew records for Azerbaijan. The list of host plant species was amended by including new host species, such as Castanea sativa Mill. for E. quercicola, Lathyrus odorathus L. for E. viciae-unijugae, Carpinus orientalis Mill. for E. arcuata, Polygonum alpinum All. for Golovinomyces spadiceus (Berk. & M.A. Curtis) U. Braun, and Alcea rosea L. for G. magnicellulatus (U. Braun) Heluta.
Herpomyces (Laboulbeniomycetes): a new order and a new species spread through pet trade
- D. Haelewaters
- D. Pfister
The class Laboulbeniomycetes comprises fungi that are obligately associated with arthropods for dispersal or as biotrophs. Two orders are currently recognized, Laboulbeniales and Pyxidiophorales. The class is severely understudied and the available class-wide phylogeny is still provisional, because it is based on a single gene (SSU) and excludes a majority of the currently recognized taxa. Herpomyces is a morphologically and phylogenetically isolated genus with 25 species that exclusively parasitize cockroaches (Blattodea). Presenting the highest level of taxon sampling across Laboulbeniomycetes to date, we evaluate a three-gene phylogeny (nrSSU, ITS, nrLSU) and propose a new order in the class Laboulbeniomycetes. We describe Herpomycetales to accommodate the single genus Herpomyces. In addition, building on the six-gene dataset from the Ascomycota Tree of Life monumental paper by Conrad Schoch and colleagues (2009), we confirm that Laboulbeniomycetes and Sordariomycetes are sister orders and we apply ‘Laboulbeniomyceta’ as a rankless taxon for the now well-resolved node that describes the most recent common ancestor of both classes. A molecular clock analysis of this six-gene dataset, using five fossil calibration points, revealed that Laboulbeniomycetes and Sordariomycetes diverged around the Triassic-Jurassic boundary (206 Mya). Within Laboulbeniomycetes, the earliest split (divergence of Pyxidiophorales) occurred around 160 Mya. Finally, Herpomycetales and Laboulbeniales diverged around 143 Mya. With this contribution, we add a robust molecular phylogenetic component to a group of fungi that has been almost exclusively defined by morphology. Our analysis of the ITS phylogeny of the genus Herpomyces brought to light an undescribed species on Shelfordella lateralis. Study of its morphology supports separation from other species in the genus. This new species has been discovered in colonies of S. lateralis in Hungary, Poland, and Massachusetts, USA. Interestingly, these colonies were all retrieved from pet stores, suggesting a role of international pet trade in the global distribution of these biotrophic fungi.
Metabarcoding of the pathogenic funus, Ophidiomyces ophiodiicola
- S. Gentry
Onygenales is a medically important order of fungi because it houses various fungal pathogens of humans and animals. Within recent years, emerging infectious diseases perpetuated by fungal pathogens have increased in wildlife, many resulting in currently uncontrolled epidemics such as chytridiomycosis among amphibians and white nose syndrome seen among bats. Ophidiomyces ophiodiicola is the primary agent of an infectious disease affecting various species of snakes across midwestern and eastern states. The pathogen colonizes the skin of its host, causing skin lesions, facial swelling, and hardened scales that, depending on the species of snake, can be fatal. There is a large variation in species susceptibility and aspects surrounding the fungus remain unclear; primarily its diversity. The objective of my experiment is to implement high throughput sequencing to understand pathogen diversity among its geographical distribution from cultured samples obtained from the National Wildlife Health Center in Madison, Wisconsin. Targeting and amplifying the internal transcriber spacer (ITS) region, which is universal among fungi, and BLASTing the collected sequences across databases such as the National Center for Biotechnology Information and GenBank will help compare and confirm all documented strains. Understanding diversity and distribution are integral in grasping mechanisms of infectious diseases. Using high throughput sequence analysis will aid in making connections behind fungal ecology and diversity among a relatively new pathogen that is threatening species such as the Eastern Massasauga Rattlesnake and Timber Rattlesnake. Furthermore, while extensive surveying has driven higher awareness, there is a lack of vigilance along western states. Extending this method of analysis to acquired swab samples from western facilities such as the Oregon Department of Fisheries and Wildlife and from universities with invested interests in this disease such as UC Davis, a more thorough and complete geographical distribution can be built. In conclusion, diversity and ecology are two sides of the same coin when attempting to understand infectious diseases of which not only elucidates climate tolerance but also the extent of diversity.
Fungal skin assemblages on hibernating bat species that are susceptible and resistant to white-nose syndrome in North America
- K. Vanderwolf
- J. Lorch
- D. Blehert
North American bats have experienced catastrophic population declines from white-nose syndrome (WNS), an emergent disease caused by the fungus Pseudogymnoascus destructans (Pd). Although Pd has a broad host range on hibernating bats in eastern North America, population-level impacts of WNS vary by host species. For example, little brown (Myotis lucifugus), northern long-eared (M. septentrionalis), and tricolored (Perimyotis subflavus) bats have experienced precipitous declines due to WNS, whereas species such as big brown (Eptesicus fuscus), gray (M. grisescens), and Virginia big-eared (Corynorhinus townsendii virginianus) bats appear to exhibit some degree of resistance to the disease and have not suffered such devastating losses. Mechanisms of WNS-resistance have not been fully elucidated, but are likely multifactorial. Microbial skin communities can influence host resistance to infectious diseases by inhibiting or competing with pathogens. We compared fungal assemblages on hibernating bat species with low or no WNS-related mortality (n=262) to WNS-susceptible bat species (n=163) using culture-dependent methods. The diversity and abundance of skin fungi differed between many of the bat species. Known WNS-resistant bat species had a mean of 2.5 ± 1.8 fungal genera per bat (mean Shannon index 0.48 ± 0.55), which was significantly higher than WNS-susceptible bats (1.1 ± 1.0 with a Shannon index of 0.12 ± 0.30; χ2=103.54, p < 2.2e-16). Of particular note was an abundance of certain yeast species (namely Debaryomyces spp.) on the skin of resistant bat species; these yeasts were absent or rare on the skin of bat species susceptible to WNS. Analyses to investigate potential relationships between fungal assemblages and resistance to WNS are underway.
Speciation by host-switching in two related cutaneous fungal pathogens of pet animals
- A. Čmoková
- V. Hubka
- T. Větrovský
- M. Kolarik
We selected two closely related skin pathogens transmitted from pets to human (mainly children) with increasing tendency to infect human during the last few years. In Europe, species Trichophyton benhamiae and T. erinacei are transmitted to human mainly from guinea pigs and hedgehogs, respectively. A considerable genetic and phenotypic variability has been revealed in these emerging pathogens. To substantiate the initial finding, we assembled strains isolated from various hosts in different European countries, Japan and USA. We conducted several analyses to elucidate whether the detected level of variability reflects undescribed species diversity or a high infraspecific variability. A total number of 326 and 146 strains from T. benhamiae and T. erinacei complexes, respectively, associated with human and animal dermatophytoses were analysed using two sequenced loci, 10 and 7 microsatellites loci, respectively, and morphological and physiological methods. Among T. benhamiae isolates, we revealed four very distinct population clusters. Three of them were present in Europe and the last only in America. Among T. erinacei isolates, only two poorly differentiated clusters were found. The two T. erinacei populations seem to be strongly carrier-specific. First, most frequently human-associated population of T. erinacei (89% of human isolates) was specific for favourite African pet hedgehog Atelerix albiventris, second population was specific for wildlife European hedgehog Erinaceus europaeus. In T. benhamiae, we observed not so strong carrier-specific population pattern. As main carrier of American population of T. benhamiae we identified a dog and for European–Japan population rabbit. However, guinea pig was also recorded as frequent carrier in case of both populations. Another two populations, including most common population (78 % of all strains), are transmitted exclusively from guinea pig. The most common population of T. benhamiae, which is responsible for the current epidemic of dermatophytosis in Europe, was identified as closely related to American population. Based on genetic relatedness of strains of both populations, we suggest that virulent population of T. benhamiae was introduced from North America to Europe. There, it began to clonally spread among different host (guinea pig) even though original host (dog) also occurred here. High genetic divergence and phenotypic differences between all four populations of T. benhamiae indicate that they can be considered as independent species. In contrast, we found small genetic and phenotypic differences between populations of T. erinacei, although they seem to be more carrier-specific.
The architecture of mucoralean genomes: Lessons from a ploidy point of view
- K. Voigt
- V. Schwartze
Mucoralean fungi represent one of the most ancient terrestrial fungi. While most species are soil-inhabiting saprotrophic organisms, some species can also cause life-threatening infections in humans, called mucormycosis. While mucormycoses are uncommon infections, they have been increasingly recognized in immunocompromised patients during the last decades. More than 80% of these infections are caused by members of the genera Rhizopus, Mucor and Lichtheimia. Besides their clinical importance, mucoralean fungi also represent an interesting model to study evolution of basal fungi. However, research on this fungal group is still limited and only little is known about genome structure and genome evolution of these fungi. To get insights into the transition from saprobe to pathogen, a tripartite -omics based approach was applied using genomics, transcriptomics and lipidomics approaches of Lichtheimia species and specimens. The genomes of a total of six isolates from from environmental and clinical background were sequenced and compared to other fully sequenced genomes of mucoralean and non-mucoralean fungi. All genomes were characterised by the presence of extensive gene duplications, which seems to be a universal feature of mucoralean genomes. The increased amount of duplicated genes is believed to be a result of a combination of an ancient whole genome duplication event in the early evolution of mucoralean fungi and additional lineage-specific single gene duplications. However, the exact mechanisms are unknown since all current knowledge relies on phylogenetic reconstruction of rather distantly related species. Our analyses revealed that strains of Lichtheimia species are generally haploid with an average genome size between 30 – 35 Mb and are well conserved between isolates. In contrast to these results, we identified a single strain which shows a genome size of about 60 Mb and appears to possess a diploid genome. Results of the analysis of single-spore isolates indicate that the nuclei of the strain are not identical, resulting in strong phenotypic differences between the colonies which are associated with defects in central signalling pathways and virulence. Virulence-associated traits appear to be well-conserved also in non-pathogenic species. However, differences were found in thermal adaptation, which was associated with metabolic changes. Interestingly, the sensitivity of L. hyalospora to thermal stress could be reduced by the simultaneous increase of osmotic pressure as a measure for the flexibility of the stress response. The data of this study give first insights into the variability of the genome architecture, the genomic, transcriptomic and lipidomic transition to stress resistance and pathogenicity of a mucormycotic agent. and the early consequences of genome duplication in basal mucoralean fungi. In addition, the availability and analysis of isolates with reduced virulence will contribute to the understanding of virulence-associated traits in these human pathogens.
Fifty strains of black: Resequencing of the melanised polyextremotolerant yeast Aureobasidium pullulans
- C. Gostinčar
- M. Turk
- J. Zajc
- N. Gunde-Cimerman
Aureobasidium pullulans (de Bary) G. Arnaud is a melanised yeast-like fungus of considerable interest due to its ubiquitous distribution, polyextremotolerant physiology and large biotechnological potential. It is best known as an epiphyte on various plant surfaces, but it is also frequently found in domestic environments and in food, in hypersaline water, in certain types of glacial ice, and a number of other habitats. It is used for the production of pullulan and aureobasidin A and is commercially available as a biocontrol agent for limiting the damage caused by several plant pathogens (both bacterial and fungal) in agriculture. In 2014 we published a de novo genome sequence of A. pullulans and three closely related species, revealing a redundancy in several gene families that could be linked to the nutritional versatility of these species and their particular stress tolerance. To build upon these initial genomic discoveries and investigate the population genomics of the species, we recently re-sequenced fifty additional A. pullulans strains from our culture collection, which includes isolates from various habitats and with a wide geographic distribution. Initial analyses of the data showed that the genomes share a high degree of similarity in their size and the number of predicted genes. Single nucleotide polymorphisms (relative to the reference genome) cover approximately 2% of the sequence. No clear population structure was detected with several methods and these results could possibly be explained by a fair amount of recombination within the species. The genome of A. pullulans contains a well-defined mating locus, but the teleomorph of the fungus has never been described. In addition to population genomics and evidence for recombination, other presented results will be focused on genomic traits linked to the polyextremotolerant character and biotechnological use of this interesting and useful black yeast.
Recent advances on taxonomy and systematics Boletaceae: the contribution of species from Thailand
- O. Raspé
- S. Vadthanarat
- S. Lumyong
The taxonomy and systematics of the Boletaceae have long been based on morphological species and genera concepts from Europe and North America. From about 2010 on, molecular (DNA) analyses including tropical boletes have started to profoundly change the systematics this group, with many new genera being published. This revolution is still ongoing, and the purpose of this paper is to present recent advances on Boletaceae systematics based on specimens collected in Thailand. We used both multi-gene phylogenetic analyses and morphological descriptions to better understand the systematics of Thai boletes. The most recent findings are two new genera, Chromatophyllum and Cacaoporus, and the identification of phylogenetic affinities of the genus Rhodactina. Chromatophyllum is a new phylloporoid genus forming a clade within the Pulveroboletus group, distant from the Phylloporus clade, which belong in the Xerocomoideae. Morphologically, Chromatophyllum can easily be distinguished from Phylloporus by the ovoid spores, lack of bacillate spore ornamentation, and deeply yellowish-orange to red lamellae. We described two new species of Chromatophyllum from Thailand, and recombined two Phylloporus species from the Americas in this new genus. Cacaoporus, which also belongs in the Pulveroboletus group, is unusual among Boletaceae in having a completely dark chocolate-brown hymenophore. Finally, with the discovery of a new Rhodactina species, R. rostratispora, we revealed the phylogenetic affinities of this truffle-like genus. It belongs in the Leccinoideae, and forms a well-supported clade with the other genera showing a purple color change of the spores when in contact with aqueous KOH solution, namely Borofutus and Spongiforma. The project is ongoing, and the impressive diversity of Boletaceae from Thailand might still bring interesting insights on the systematics and evolution of this family.
Mating genes evolution and population genetics in Suillus brevipes
- Y. Ke
- S. Branco
- A. Rojas
- K. Hameed
- I. Grigoriev
- A. Kuo
- K. Barry
- H. Liao
- N. Nguyen
- P. Kennedy
- T. Bruns
- R. Vilgalys
Mating system is a crucial feature in population genetics. Sexual reproduction in fungi is controlled by MAT loci that define different mating types. MAT loci of basidiomycetes are characterized by a high level of polymorphism. The high diversity of mating alleles has attracted much attention, and is explained by strong negative frequency dependent selection in which rare alleles have a selective advantage over popular alleles which have lower fitness. Under this scenario, multiple alleles of MAT loci are predicted to have an extended coalescence time relative to other neutral loci. Previously, mating systems and allele number in basidiomycetes are inferred by pairing of single-basidiospore isolates. In this study, we employed aTRAM (automated target restricted assembly method) to recover haplotypes of the HD MAT locus from genomic shotgun sequencing data in populations of the bolete mushroom Suillus brevipes across North America. The HD MAT locus of S. brevipes only contains a pair of homeodomain encoding HD protein. De novo assembly of shotgun sequence data shows 56 distinct alleles among 55 dikaryotic samples, where heterozygotes in HD MAT are mostly found. Comparison of different haplotypes shows that the same HD1 allele always coexists with the same HD2 in our samples; multipartite linkages are not observed. Population genetic analysis suggests a distinct mating allele composition for each population, consistent with restricted gene flow among these populations in former studies. When compared to selectively neutral loci, we find HD MAT locus has weaker geographic differentiation among populations as theory predicted. Our results confirm that illumina short reads can be used to recover “idiomorphic” alleles of mating types from dikaryotic individuals, the high mating type diversity under balancing selection, and restricted recombination within the MAT region. However, phylogenetic incongruence between HD1 and HD2 phylogenies and analysis of recombination surprisingly implies that deep recombination also contributes to the generation of MAT allele diversity. More samples and analysis are required to fully reveal the evolutionary process of HD MAT alleles.
What do we know about the associated mycobiota of the built environment?
- U. Thrane
- B. Andersen
The list of fungal species reported to be associated with the built environment is long, even when broken down to comparable building characteristics. The reason is that surveys are very diverse in terms of sampling methods, identification protocols and environmental conditions. Moreover, the rapid development in taxonomic schemes is a major challenge. The phylogenetic species concept splits ‘old’ indoor related species of e.g. Aspergillus, Penicillium, and Cladosporium into numerous new species, often delimitated by differences in gene sequences only. The methods used for detection and identification are frequently updated and fine-tuned, but there will always be a lag-time before implementation is completed. Consequently, many reports and scientific papers on the mycobiota are using yesterday’s species concept, which may be beneficial as there is a substantial body of knowledge describing the functionality and ecology of yesterday’s species. However, an updated sequenced based identification will generate results based on today’s species concept where no or limited information on physiology, toxicology, ecology etc. is present. The modern molecular tools are getting faster and faster, but the functional characterization of taxonomic novelties cannot match the pace of the taxonomic development. The result is a crucial loss of knowledge of important indoor related species of Aspergillus, Penicillium and Cladosporium and several examples will demonstrate this unfortunate situation. The outcome is that many descriptive scientific papers contain long lists of fungal species detected; but what do these results tell us about the mycobiota of built environment or health impact? In most cases, nothing or only speculations that reflect the limited body of knowledge of today’s species. The knowledge gaps represent an overwhelming amount of different types of very important data, far too much for a single research unit to cope with. It is proposed to develop and launch an international infrastructure that supports compilation of data on functionality of the fungal species. Ideally, in an open and easy accessible set-up e.g. like GenBank, and with the possibility to link between databanks to support a much better and deeper understanding of the mycobiota of the built environment and its impact on human health.
Improving precision in the study of the built mycobiome: recent changes and additions in the UNITE database for molecular identification of fungi
- R. Nilsson
- U. Kõljalg
- A. Taylor
- K. Abarenkov
Recent molecular ecology studies have found the built environment to contain a surprisingly large number of transient and resident fungi. Some of them are associated with medical conditions in humans, notably asthma and eczema development, rendering the taxonomic affiliation of these fungi important from medical and many other points of view. Molecular (DNA-based) identification of fungi is, however, fraught with technical and biological complications, including lack of reference DNA sequences, incorrectly annotated reference sequences, chimeric or otherwise low-quality molecular data, missing metadata on, e.g., country and substrate of collection, and failure to follow relevant metadata standards. This presentation details the steps that the UNITE database for molecular identification of fungi has taken to facilitate molecular identification of the built mycobiome. The measures include organizing two sequence annotation workshops, sequencing >500 previously unsequenced fungal type specimens, implementing support for the MIxS-BE metadata standard, and devising several new software tools, including a search engine designed to find truly unknown fungi in the corpus of public fungal DNA sequences. This project, funded by the Alfred P. Sloan foundation, resulted in more than 75,000 improvements (such as name changes and metadata additions) to public DNA sequences relevant to the built mycobiome. These improvements were shared with a range of other databases and resources, including GenBank and the ISHAM database.
Fungal diversity of airborne samples is uncovered by DNA metabarcoding
- L. Muggia
- E. Banchi
- C. Ametrano
- D. Stankovic
- A. Pallavicini
Fungal spores and mycelium fragments become and remain airborne and have been subjects of aerobiological studies. The presence and the abundance of certain taxa in aerobiological samples can be very variable and impaired by changeable weather conditions. In particular, it is of key importance monitoring the presence of those fungi which produce mycotoxins, and both their mycelium fragments and spores are regarded as potential allergens. So far the traditional, morphology-based methodologies used to analyze fungi in aerobiological samples have mainly assessed the few, most abundant and easily identifiable taxa and have focused only on certain environments. This research presents a first, comprehensive assessment of fungal diversity from airborne samples using a DNA metabarcoding analysis. The region ITS2 was selected as fungal barcode to catch fungal diversity in mixed airborne samples gathered for one year in five sites of North-Eastern and Central Italy. Molecular data of fungal diversity within and among the sampled sites was assessed and compared with the identifications performed by traditional microscopy. The molecular analyses find fully correspondence with the morphological inspections and provide an almost ten-fold more accurate determination of the fungal taxa. The results prove that the metabarcoding analysis is a promising approach to increases quality and sensitivity of the aerobiological monitoring and pave the way for an automated fungal identification in airborne samples to be applied in routine aerobiological monitoring. To this aim, laboratory and bioinformatics workflows have been ad hoc implemented and are here presented.
Fungal communities associated with a condemned public housing project in Richmond, CA.
- I. Sylvain
- R. Adams
- J. Taylor
Water-damaged housing has been associated with a number of negative health outcomes, principally respiratory disease and asthma. Much of what we know about fungi associated with water-damaged buildings has been gleaned from culture-based and immunochemical methods. A limited number of studies have used high-throughput sequencing technologies to assess the impact of water-damage on microbial communities in residential buildings. In this study we used amplicon sequencing and quantitative-PCR to evaluate fungal communities in a condemned public housing building in Richmond, CA, before it’s residents were relocated. We recruited 21 households to participate in this study and characterized their apartments as either a unit with visible mold or no visible mold. We collected settled dust from bathrooms, kitchens, bedrooms and living rooms from units with and without visible mold, and from the outdoors. We recovered 5,333 OTUs from 92 samples. We found that fungal biomass was greater outdoors compared to indoors, yet there was no significant difference in fungal biomass in units with visible mold and no visible mold. Fungal richness was significantly reduced in units with visible mold compared to units with no visible mold and the outdoors. We also found that units with visible mold harbored fungal communities distinct from units with no visible mold or from the outdoors. Units with visible mold were dominated by taxa within the classes Eurotiomycetes, Microbotryomycetes, Saccharomycetes, and Wallemiomycetes. A number of the OTUs recovered in significantly greater abundance from units with visible mold, such as Alternaria alternata, Cladosporium sphaerospermum, Rhodotorula mucilaginosa, and Wallemia muriae, have previously been reported with water-damaged building materials. This study demonstrates that long-term negligence and poor building maintenance in low-income public housing impacts not only the human inhabitants, but also the fungi.
Carbon for phosphorus trade in AM fungi in response to host adaptation
- Z. Alazzawi
The majority of land plants establish mutualistic symbioses with arbuscular mycorrhizal (AM) fungi. An association that benefits both partners by mediating uptake of essential plant nutrients, particularly P and N fueled by recently fixed carbon from host plants. AM fungi develop extraradical hyphal networks that link different plant species. The ability of the same fungal mycelium can colonize different plants at the same time involves an extraordinary level of compatibility and ability to adapt to different hosts by the fungus, though the adaptation to a single host might increase fitness of fungi at the expense of the other host. Our question is how plant and fungal fitness are linked and the genetics behind that. It has been suggested that AM fungi are heterokaryotic, contain genetically distinct haploid nuclei, and in response to a new plant species (changing environment), the developing fungal hyphae could temporarily segregate nucleotypes in the emerging spores to produce adapted offsprings that have different traits than the parents. We are conducting a greenhouse experiment to test if the symbiotic performance of host-adapted fungus spores (generation 1) propagated in a two plant species system, when inoculated onto their same plant host species (generation 2). Symbiotic performance will be estimated using C to P trade in both generations. We will test for differences in C for P exchange between generations and if the one host-adapted generation will grow better at the expense of plant growth/fitness on the other host. Spores of AM fungus Claroideoglomus candidum (NC268, INVAM) were inoculated onto an outer soil between two mesh bags filled with soil and one plant seedling. The developed fungal mycelium has access to two different plants; Petunia grandiflora and Allium ampeloprasum (leek) split by the mesh bags that allow only the fungal hyphae to pass through and colonize the roots. The mesh bags were filled with low P soil; P in the outer soil is doubled. Four treatments were set up as follows: Treatment 1- two leek seedlings, Treatment 2- two petunia seedlings, Treatment 3- one leek and one petunia seedling, Treatment 4- one leek and one petunia seedling (the mesh bags will be split physically in new pots around month 4). The effect of the fungus will be assessed by comparison with control treatments in which fungus was not added. To investigate preferential plant C allocation and AM fungi P uptake, we will use radioactive isotope probing before the first harvest occasion (week 12) when mycorrhiza association is still physiologically active for all the treatments. 33P isotope will be applied to the soil two weeks before 14C labeling. C to P ratio will be estimated using liquid scintillation counting. At harvest, plant fitness will be assessed by biomass and fungal fitness spore counts. The roots will be stained to estimate the colonization rate. If we observe fungal fitness changes in response to host adaptation the genomic basis for this pattern will be analyzed using single cell genomics technique.
Host specificity of mycorrhizal fungi in Australian Cryptostylis orchids
- A. Arifin
- A. Weinstein
- R. Peakall
- R. Phillips
- C. Linde
The genus Cryptostylis is unique among Australian sexually deceptive orchids in that all five species are pollinated by the same wasp species - Lissopimpla excelsa. Cryptostylis erecta, C. leptochila, C. hunteriana and C. subulata occur sympatrically in eastern Australia, while C. ovata is restricted to Western Australia. Despite their sympatry and pollinator sharing, eastern Australian Cryptostylis do not hybridise. We investigated the mycorrhizal diversity associated with Australian Cryptostylis to determine whether an inter-species difference in mycorrhizal association could be contributing to the failed establishment of hybrids. We examined a minimum of 25 plants from across five populations of each species, except for the rare C. hunteriana where we analysed only two populations. Results of both fungal isolations and direct sequencing of the ITS locus from peloton rich orchid tissue show that all the Cryptostylis species are associated with closely related Tulasnella fungi. We discovered four previously unidentified Tulasnella operational taxonomic units (OTUs). Specifically, Tulasnella OTU A is shared by four Cryptostylis species, C. erecta, C. subulata, C. leptochila, and C. ovata. Tulasnella OTU B is only associated with C. ovata, while Tulasnella OTU C has associations both with C. erecta and C. subulata. Tulasnella OTU D is was only found in association with the vulnerable species, C. hunteriana. In addition to these four new OTUs, Cryptostylis were found to associate with T. prima and T. sphagneti, both of which are known to also associate with multiple species of sexually-deceptive Chiloglottis orchids. Tulasnella sphagneti was found to only associate with C. subulata, whereas T. prima also associates with C. leptochila and C. ovata. The association with C. ovata extends the known distribution of T. prima from eastern Australia to Western Australia. The Tulasnella symbionts of Cryptostylis all belong to a closely related group of species that are found with other sexually deceptive orchids in Australia such as Drakaea, Caleana and Arthrochilus. Due to the shared association with OTU A, mycorrhizal specificity is unlikely to explain the absence of hybrids in Australian Cryptostylis.
Mycorrhizal specificity and dependence of the leafless Ghost Orchid, Dendrophylax lindenii
- L. Johnson
- D. Taylor
- G. Mueller
Dendrophylax lindenii, the Ghost Orchid, is an endangered species that is native to far western Cuba and southern Florida. The leafless morphology of D. lindenii suggests that it has a high dependency for orchid mycorrhizal fungi (MF) when growing in its natural habitat because of the reduced photosynthetic capacity compared to orchids with leaves. We investigated the root mycobiota of D. lindenii individuals using amplicon sequencing and the potential dependence of the orchid on fungi for carbon resources through stable isotope analyses. We hypothesized that the root mycobiota of D. lindenii would be distinct from those of co-occurring epiphytic orchids and that δ13C, δ15N, and δ2H of D. lindenii root samples would be consistent with fungus to plant transfer. We collected root samples from D. lindenii individuals and several co-occurring epiphytic orchids from the Florida Panther National Wildlife Refuge. We also collected bark samples from several host trees of D. lindenii, i.e., Fraxinus caroliniana and Annona glabra, to investigate fungal composition. In total, we recovered 526 OTUs (i.e. 95% sequence similarity) from root samples. Dendrophylax lindenii samples were dominated by a narrow clade of Ceratobasidium OTUs, suggesting a high specificity for these fungi. Sequences of this Ceratobasidum clade were also obtained from bark samples of host trees. D. lindenii samples were highly enriched for 13C, δ15N, and 2H. We hypothesize that orchid MF of D. lindenii may potentially be drivers of their fine scale distribution and rarity, but this needs to be tested.
The ectomycorrhizal relationship between Tricholoma vaccinum and its plant host, Picea abies and geosmin biosynthesis in the ectomycorrhizal fungus
- O. Abdulsalam
- M. Kunert
- K. Krause
- W. Boland
- E. Kothe
The relationship between ectomycorrhiza fungi and plant roots has been understood to be mutualistic. It is a relationship where the interacting fungus provides water, inorganic and auxotrophic nutrients and many other benefits to the plant host while the plant provide carbon for the fungus in return. Recent preliminary findings in Picea abies-Tricholoma vaccinum ectomycorrhiza interactions, however, have shown plant defense responses while interacting with the “supposed” mutual partner, the ectomycorrhizal fungus T. vaccinum. Metabolomics studies and volatile analyses showed a very high upregulation of alkaloids and methyl-salicylates in the plant when interacting with the fungus. The effect of such plant defense responses was also evident for the fungus. The fungus showed lower fitness and reduced mycelial growth when grown in co-culture with P. abies. The plant, on the other hand, shows a significantly higher fitness when grown with the mycorrhizal partner than without it. To test if this is a species-specific scenario, ectomycorrhiza formation between Paxillus involutus and Picea abies was investigated, the reduced fitness of the fungus was also evident in P. involutus-P. abies co-culture. To study the molecular basis of the plant response to the ectomycorrhizal fungus interactions, gene expression analysis of different conifer defense genes while interacting with the ectomycorrhizal fungus will be studied. T. vaccinum produces geosmin, a decalol known to contribute to the earth smell observed post rain after a long spell of dryness. A gene in T. vaccinum has been shown, using mRNA transcript analyses, to be involved in geosmin biosynthesis in the fungus. Stable isotope labelling using deuterium labelled substrates was used to elucidate the pathway utilized in geosmin biosynthesis in this fungus. Concluding results showed that the basidiomycete fungus produces geosmin using the classical mevalonate pathway and it was shown that the alternative MEP pathway was not used for the biosynthesis of this chemical. Recent efforts are now put into the characterization of geosmin as a communication chemical in ectomycorrhiza interaction and the function of geosmin as a communication signal.
Cophylogeny of the lichenicolous Tremella species and their hosts: disentangling a species complex mainly based on cospeciation
- S. Freire Rallo
- M. Wedin
- P. Diederich
- A. Millanes
Cospeciation between parasites and their hosts is difficult to demonstrate. One of the signs of cospeciation is that hosts and parasites share a congruent phylogenetic history and their tree topologies mirror each other. However, in the majority of host-parasite systems, other events such as host switching, lineage sorting or extinction are more frequent. High host specificity is well acknowledged within the lichenicolous species in the Tremellales (Basidiomycota, Fungi). Previous studies of Biatoropsis and their Usnea and Protousnea hosts revealed that host-switching is the most probable cophylogenetic event explaining host specificity in this system. With this work we extend these studies to another putative tremellalean lichenicolous species complex (Tremella caloplacae s. lat.) and its hosts, to evaluate if their joint evolutionary histories can be explained by parallel reciprocal speciation. We base this on a multi-loci matrix with data from Blunt-End Illumina® libraries. For species delineation we combine morphological, ecological and molecular data, as well as different molecular-based species delimitation methods. For the coevolution study we use a combination of event-cost methods, distance methods and dependence between phylogenies testing. Our results suggest that Tremella caloplacae is a species complex formed by at least six independent lineages, here interpreted as species. Each of these lineages grows on a distinct host clade, demonstrating high host specificity. Cospeciation is the most plausible cophylogenetic event to explain the joint evolutionary histories of T. caloplacae s. lat. and its hosts since it can explain the origin of five of the six independent lineages.
A molecular assessment of ectomycorrhizal diversity of cottonwoods
- B. Looney
- A. Rojas
- G. Bonito
- P. Busby
- P. Meidl
- J. Chen
- J. Labbé
- R. Vilgalys
Poplars (Populus ssp.) are an economically important wood and bioenergy crop that are nutritionally supported by both arbuscular mycorrhizal and ectomycorrhizal fungi. While aspens (P. tremula and P. tremuloides) are known to host a diverse array of ectomycorrhizal fungi, cottonwoods (P. trichocarpa, P. deltoides, and P. nigra) are considered to have a more restricted set of ectomycorrhizal associates. Here we provide a comparative overview of the reported ectomycorrhizal associates of cottonwoods informed by metagenomics and sporocarp sampling from natural stands of P. trichocarpa in Washington and Oregon and P. nigra in France. Based on our sampling we have identified a well-defined community of cottonwood associates, including specific members of Cortinarius, Hebeloma, Inocybe, Laccaria, Lactarius, and others. It is our objective to phylogenetically place these species within their generic/species complex context and investigate patterns of their host association in relation to their distribution. We hypothesize that host-restricted species are common associates of cottonwoods as they are co-evolved and selected by their host, whereas ECM invaders, or generalist ECM species, are diverse on cottonwoods but rare.
Insights on the phylogeny of Phyllachora-like fungi infecting Myrtaceae
- D. Guterres
- M. Santos
- S. Elias
- D. Pinho
- J. Dianese
Phyllachora is a biotrophic fungus found on many Myrtaceae in Brazil; worldwide with ca 50 species recorded on myrtaceous hosts. Molecular phylogeny clearly showed that Phyllachora is a polyphyletic genus with species recently transferred to Telimena (Telimenaceae, Phyllachorales) and Neophyllachora (Phyllachoraceae), the latter considered as a genus with species infecting exclusively members of the Myrtaceae. However, both genera have species on myrtaceous host. To elucidate the relationship among these fungi within the Phyllachorales, six fungal species on Myrcia, Psidium, Eugenia and Campomanesia from Brazil had the rDNA (nuclear small subunit 18S, nuclear large subunit 28S and nuclear internal transcribed spacer ITS), and the translation elongation factor 1 (TEF1) nuclear loci partially sequenced and were used in multilocus phylogenetic analyses. Taxonomic identity of the fungal specimens were confirmed by morphological examination. The results suggest that the phyllachora-like fungi on myrtaceous host are not a monophyletic group, with species grouping in distinct families within Phyllachorales. Therefore the circumscription of Neophyllachora might be revised. As an example, Phyllachora furnasensis became part of a strongly supported clade within Telimena and will eventually be recombined. Besides that, the phylogenetic analyses that led to the establishment of Neophyllachora need revision considering the new rules guiding the GenBank in terms of the size of acceptable sequences, and the fact that some sequences of mitochondrial DNA were used as if they were rDNA sequences.
Diversity and biogeography of Botryosphaeriaceae associated with Myrtales trees in Southern Africa and Southern China
- D. Pavlic-Zupanc
- A. Maduke
- Y. Zhang
- M. Wingfield
- B. Slippers
- S. Chen
- W. He
- J. Roux
Botryosphaeriaceae species are important latent pathogens causing diseases on many woody plants, including trees in the Myrtales. The diversity and biogeography of Botryosphaeriaceae on trees in the Myrtales in regions such as Southern Africa and Southern China is poorly documented. In this study, we identified Botryosphaeriaceae species on more than 30 native and non-native tree and shrub species in the Myrtales in five African countries (Malawi, Mozambique, South Africa, Tanzania, Zimbabwe) and the GuangDong and HaiNan Provinces of the Southern China. Isolates were identified based on phylogenetic analyses of ITS rDNA, β-tubulin, TEF-1α, LSU and RPB2 sequences. The isolates originating from African countries resided in the genera Diplodia, Neofusicoccum and Lasiodiplodia and representing 10 species, of which four are previously unknown. The isolates originating from Southern China resided in four genera namely Cophinoforma, Lasiodiplodia. Neofusicoccum and Botryosphaeria, of which the latter two were absent in the HaiNan Province. In this region, fourteen species were identified of which two represented novel taxa. The Botryosphaeriaceae species composition differed significantly between the regions where they were collected. Lasiodiplodia species were dominant in Southern China representing 80% of all isolates obtained in this study. Lasiodiplodia theobromae was the most common species from the latter region. In contrast, L. theobromae was absent from the studied hosts in Southern Africa, while N. parvum was the most abundant species collected. Only L. gonubiensis and N. parvum overlapped between Southern China and the Southern African countries. Other than Caphinoforma mamane, which was sporadically identified in both provinces in Southern China, there was no overlap of species identified on Myrtales in the two provinces. It appears that location rather than host association shapes Botryosphaeriaceae species composition on Myrtales hosts considered in this study.