Source: UNIVERSITY OF ARIZONA submitted to NRP
MOLECULAR ECOLOGY, EVOLUTION AND SYSTEMATICS OF ENDOPHYTIC FUNGI: CLUES TO THE EVOLUTION OF MAJOR TROPHIC MODES IN THE ASCOMYCOTA
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0203733
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
Oct 1, 2009
Project End Date
Dec 31, 2012
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
UNIVERSITY OF ARIZONA
888 N EUCLID AVE
TUCSON,AZ 85719-4824
Performing Department
Plant Science
Non Technical Summary
Fungal symbionts of plants are a fundamental aspect of all human-made and natural plant communities. Fungal endophytes -- fungi that live within healthy plant tissues such as leaves and stems -- are an understudied but ubiquitous group of symbionts whose diversity, host affinity, ecological roles, and evolutionary history are very poorly known. Because these fungi represent a trove of previously undiscovered diversity, they are a key component in addressing broad questions regarding fungal evolution, including the evolution of factors such as virulence. Moreover, endophytes represent a wealth of novel metabolic products of use in agriculture, medicine, and industry. This study aims to survey previously unknown endophytic fungi, to examine their diversity and host associations, to infer their evolutionary relationships with disease-causing and decay-causing fungi, and to examine these fungi for biological products (metabolites) of use in human endeavors. Methods will range from field surveys to culturing, DNA sequencing, computational bioinformatics, and in vivo and in vitro assessments of fungal products in a wide array of assays. Anticipated benefits of this work include an enhanced understanding of earth's fungal diversity and a new perspective on the factors that shape the ecological roles of fungi, as well as the discovery of potentially novel and important metabolites with diverse applications.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2062420107050%
2122499110250%
Knowledge Area
212 - Pathogens and Nematodes Affecting Plants; 206 - Basic Plant Biology;

Subject Of Investigation
2420 - Noncrop plant research; 2499 - Plant research, general;

Field Of Science
1070 - Ecology; 1102 - Mycology;
Goals / Objectives
The goals of this project are (1) to expand our knowledge of the diversity and host affinities of endophytic fungi, which are ubiquitous associates of agricultural and non-domesticated plants; (2) to use these previously unknown fungi to enrich current perspectives on the phylogenetic relationships of fungi, as a means to infer the evolution of major ecological modes such as virulence and mutualism; and (3) to seek novel uses of these fungi in agricultural, pharmaceutical, and industrial applications. These goals will be achieved through a multidisciplinary research program combined with mentorship of graduate and undergraduate students and ongoing outreach activities on the Navajo Nation.
Project Methods
Fungal endophytes will be isolated from fresh, asymptomatic plant material gathered in field surveys of diverse environments, chosen to satisfy a series of criteria (biogeographic distinctiveness, ecological variation, phylogenetic novelty of plant communities). Leaves and stems will be surface-sterilized and fragments corresponding to the infection domain of individual fungi will be plated on standard media under sterile conditions. Isolates obtained in axenic culture will be vouchered and accessioned in a culture library and retained for further work. Total genomic DNA will be extracted from all samples and used to amplify both fast-evolving and phylogenetically informative loci (nuclear internal transcribed spacer and the nuclear large subunit, respectively) for preliminary identification. Fungi representing distinctive genotypes will be further sequenced for loci with known phylogenetic utility. Resulting sequences will be used to infer phylogenetic relationships using Bayesian, maximum parsimony, and maximum likelihood methods, and trophic mode transitions will be estimated using ancestral state reconstruction. Resulting trees will provide the basis for estimating host breadth, geographic structure, and the directionality of evolutionary trajectories in the species-rich Ascomycota, with this perspective being especially rich due to the inclusion of a wealth of previously unstudied fungi. Clades of particular interest will be screened for antimicrobial, anti-cancer, and cellulase activity using established protocols. Through collaboration the active metabolites will be isolated and identified. Data will be published in peer-reviewed journals and disseminated through presentations at national and international conferences and symposia, as well as through the PI's web presence.

Progress 10/01/09 to 12/31/12

Outputs
OUTPUTS: Our proposal targeted six biogeographically distinctive provinces of special interest for recovering diverse and potentially novel endophytic and endolichenic fungi. As of the conclusion of this project, we have completed fieldwork at five study sites, including the biotically rich Chiricahua Mountains of southeastern Arizona; the southern Appalachian Mountains of western North Carolina; subtropical scrub forest of south-central Florida; and coastal and continental tundra in Alaska. We have limited our study to five sites because of the extreme diversity and abundance of fungi recovered in each site: our intensive sampling has yielded a higher than expected richness of symbiotrophic fungi in each site. In each site, we identified three microsites along a ca. 100-200m transect and collected living, symptomless material from replicate sets of 10 lichen species and 10 vascular plant species (total of 20 representative species in each of 3 microsites/site). Our sampling maximized the representation of all major lineages of lichens and plants and was designed to permit comparisons among the fungal communities associated with congeneric hosts in disparate sites. Plant- and lichen tissue was surface-sterilized, cut into small (ca. 2mm2) fragments, and placed onto a general medium under sterile conditions. We plated 2880 fragments/site of healthy lichen thalli and leaves (144 fragments/species/site) using portable laminar flow hoods, which allowed us to process fresh material very rapidly. After observing that a small number of fungi emerge very slowly in culture, and recognizing that many of these late-emerging fungi are phylogenetically interesting, we incubated tissue segments for up to one year, rather than for 1-2 months (typical for most endophyte research). Concurrent with the present award, Jana URen, a Ph.D. candidate in my laboratory, initiated a complementary project examining the diversity and taxonomic composition of fungi associated with dead leaves before soil contact (i.e., in tree canopies), and in the leaf litter, in each of our focal sites (four focal species/site, replicated in each of three microsites/site). This was supported with supplementary funds from the Clark Rogerson Travel Award and a Graduate Fellowship award from the Mycological Society of America We isolated and vouchered fungi, sequencing the nuclear ribosomal internal transcribed spacer of each isolate, and conducting genotype comparisons to identify distinctive fungi for subsequent multilocus phylogenetic analyses in the Lutzoni lab. To date, we have isolated 6784 strains. All have been deposited as living vouchers at the Robert L. Gilbertson Mycological Herbarium at the University of Arizona, where they are publicly available. We have successfully sequenced the ca. 600 bp ITS region (ITS 1, 5.8S, and ITS 2) and, as part of the same fragment, the first 600 bp of the nuclear ribosomal large subunit (LSUrDNA), for 5988 isolates. PARTICIPANTS: Anne Elizabeth Arnold, Jana URen, Mariana del Olmo Ruiz, Rowena Dolino, Steve Uyeda, Alex Delgado, Frankie Orozco, 12 additional undergraduates TARGET AUDIENCES: Public, stakeholders, collaborating researchers, students PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
This award supported research in the Arnold lab by two female Ph.D. students (one Latina); one female technician who has now enrolled as a MS student (Latina), with a focus on endophytic fungi; 12 undergraduates (six female; five minority); two community college students (one female); two teachers (one female and Latina, based on the Navajo Nation); one community college faculty member (female, and based on the Navajo Nation); and one high school student (Latino). Both received supplemental funding from extramural sources based on this award. I hosted three summer interns associated with this award, one of whom later returned for a second internship (for a total of four summer internships overall): Alex Delgado, a first-year student at Pima Community College; Steven Uyeda, a teacher at Sunnyside High School in Tucson, AZ, and an instructor at Pima Community College; and Rowena Dolino, a teacher on the Navajo Nation. All were supported by supplemental funds. I have worked with a high school student, Frankie Orozco, who is a gifted young scientist whose outstanding project has led to numerous awards. We are actively recruiting Frankie to enroll as an undergraduate at the University of Arizona. This award resulted in 50 presentations by Arnold and project participants in the Arnold lab in local, regional, and international venues. Of these, 25 were invited presentations. A total of 12 intramural presentations also were supported by this award. Three high school students presented their work at science fairs. The award resulted in 27 peer-reviewed papers, with several to be submitted after the awards conclusion.

Publications

  • 1. Bascom-Slack, C., A.E. Arnold, S.A. Strobel. 2012. Student-directed discovery of the plant microbiome and its products. Science 338: 485-486.
  • 2. Tedersoo, L., A.E. Arnold, K. Hansen. 2013. Novel aspects in the life cycle and biotrophic interactions in the Pezizomycetes. Molecular Ecology, in press.
  • 3. Devan, M.M.N., S.H. Furr, A.E. Arnold. 201x. Persistent effects of wildfire on fungal endophyte communities in Quercus hypoleucoides and Juniperus deppeana. Microbial Ecology, accepted.
  • 4. Hoffman, M.T., M. Gunatilaka, E.M.K. Wijeratne, A.A.L. Gunatilaka, and A.E. Arnold. 2013. IAA production by endophytic Pestalotiopsis is enhanced by an endohyphal bacterium. PLoS ONE, accepted.
  • 5. Milani, N.A., D.P. Lawrence, A.E. Arnold, and H.D. vanEtten. 2012. Origin of pistatin demethylase (PDA) in the genus Fusarium. Fungal Genetics and Biology 49: 933-942.
  • 6. Martinson, E.O., E.A. Herre, C.A. Machado, A.E. Arnold. 2012. Culture-free survey reveals diverse fungal communities associated with figs (Ficus spp.) in Panama. Microbial Ecology 64: 1073-1084.
  • 7. Gazis, R., J. Miadlikowska, F. Lutzoni, A.E. Arnold, P. Chaverri. 2012. Culture-based study of endophytes associated with rubber trees in Peru reveals a new class of Pezizomycotina: Xylonomycetes. Molecular Phylogenetics and Evolution 65: 294-304.
  • 8. URen, J.M., F. Lutzoni, J. Miadlikowska, A. Laetsch, A.E. Arnold. 2012. Host and geographic structure of endophytic and endolichenic fungi at a continental scale. American Journal of Botany 99: 898-914.
  • 9. Higgins, K.L., A.E. Arnold, P. Coley, T. Kursar. 2012. Communities of fungal endophytes in tropical forest grasses: highly diverse host- and habitat generalists characterized by strong spatial structure. Fungal Ecology, in press.
  • 10. Varughese, T., N. Riosa, S. Higginbotham, A.E. Arnold, and 4 others. Antifungal depsidone metabolites from Cordyceps dipterigena, an endophytic fungus antagonistic to the phytopathogen Giberella fujikoroi. Tetrahedron Letters 28: 1624-1626.
  • 11. Molinar, E., N. Rios, C. Spadafora, A.E. Arnold, and 4 others. 2012. Coibanoles, a new class of meroterpenoids produced by Pycnoporus sanguineus. Tetrahedron Letters 53: 919-922.
  • 12. Lawrence, D.P., B.M. Pryor, S.B. Kroken, A.E. Arnold. 2011. Interkingdom horizontal gene transfer of a hybrid NRPS/PKS from bacteria to filamentous Ascomycota. PLoS ONE 6: e28231.
  • 13. Wijeratne, E.M.K., 7 others, A.E. Arnold, L. Whitesell, A.A.L. Gunatilaka. 2012. Geopyxins A-E, ent-kaurane diterpenoids from endolichenic fungi, Geopyxis aff. majalis and Geopyxis sp. AZ0066: structure-activity relationships of geopyxins and their analogues. Journal of Natural Products 75: 361-369.
  • 14. Wang, X.N., B.P. Bashyal, E.M.K. Wijeratne, J.M. URen, M. Gunatilaka, A.E. Arnold, A.A.L. Guna-tilaka. 2011. Smardaesidins A-G, new isopimarane and 20-Nor-isopimarane diterpenoids isolated from Smardaea sp., endophyte of Ceratodon purpureus. Journal of Natural Products 74: 2052-2061.
  • 15. Moreno, E., T. Varughese, C. Spadafora, A.E. Arnold, and 4 others. 2011. Chemical constituents of the new endophytic fungus Mycosphaerella sp. nov. and their antiparasitic activity. Natural Products Communications 6: 835-840.
  • 16. Dalling, J.W., A.S. Davis, B.J. Schutte, A.E. Arnold. 2011. Seed survival in soil: integrating effects of predation, dormancy, and the soil microbial community. Journal of Ecology 99: 89-95.
  • 17. Higgins K.L., P.D. Coley, T.A. Kursar, A.E. Arnold. 2011. Culturing and direct PCR suggest prevalent host-generalism among fungal endophytes of tropical grasses. Mycologia 103: 247-260.
  • 18. URen, J.M., F. Lutzoni, J. Miadlikowska, A.E. Arnold. 2010. Intensive sampling reveals ecological distinctiveness and continua among culturable symbiotrophic and saprotrophic Ascomycota in a montane forest. Microbial Ecology 60: 340-353.
  • 19. Epps, M.J. and A.E. Arnold. 2010. Diversity, abundance, and community network structure in sporocarp-associated beetle communities in the central Appalachian Mountains. Mycologia 102: 785-802.
  • 20. Hoffman, M.T. and A.E. Arnold. 2010. Diverse bacteria inhabit living hyphae of phylogenetically diverse foliar endophytes. Applied and Environmental Microbiology 76: 4063-4075.
  • 21. Arnold, A.E., L.J. Lamit, M. Bidartondo, C. Gehring, H.S. Callahan. 2010. Interwoven branches of the plant and fungal trees of life. New Phytologist 185: 874-878.
  • 22. Peay, K., M. Bidartondo, A.E. Arnold. 2010. Not every fungus is everywhere: scaling to the biogeography of fungal-plant interactions across roots, shoots, and ecosystems. New Phytologist 185: 878-882.
  • 23. Parrent, J.L., K. Peay, A.E. Arnold, L. Comas, P. Avis, A. Tuininga. 2010. Moving from pattern to process in fungal symbioses: linking functional traits, community ecology, and phylogenetics. New Phytologist 185: 882-886.
  • 24. Maddison, D.R. and A.E. Arnold. 2009. A review of the Bembidion (Odontium) aenulum subgroup (Coleoptera: Carabidae), with description of a new species. Zootaxa 2214: 45-61.
  • 25. Arnold, A.E., J. Miadlikowska, K.L. Higgins, S.D. Sarvate, P. Gugger, A. Way, V. Hofstetter, F. Kauff, F. Lutzoni. 2009. Hyperdiverse fungal endophytes and endolichenic fungi elucidate the evolution of major ecological modes in the Ascomycota. Systematic Biology 58: 283-297.
  • 26. URen, J.M., J.W. Dalling, R.E. Gallery, D.R. Maddison, E.C. Davis, C.M. Gibson, A.E. Arnold. 2009. Diversity and evolutionary origins of fungi associated with seeds of a neotropical pioneer tree: a case study for analyzing fungal environmental samples. Mycological Research 113: 432-449.
  • 27. Rodriguez, R., J. White, A.E. Arnold, R. Redman. 2009. Fungal endophytes: diversity and ecological roles. New Phytologist 182: 314-330.


Progress 01/01/11 to 12/31/11

Outputs
OUTPUTS: OUTPUTS: Outputs from this project in 2011 included activities in teaching, research, and service. In 2011 I convened four courses that were thematically tied to ARZT-136823-H-25-195. I taught two one-semester discussions focusing on fungal ecology and systematics for graduate students and advanced undergraduates (Topics in Fungal Ecology and Evolution) and one undergraduate course in both spring and fall, serving approximately 125 students from diverse majors (Microbial Diversity). Together these efforts served more than 140 students and benefited greatly from links with my research program. I also gave guest lectures in five courses. In 2011 I continued to mentor my current group of graduate students (4 Ph.D. students), one of whom graduated in late 2011. I accepted a new MS student into the lab in fall 2011. I supervised a postdoctoral associate based in Panama and supported the work of a new postdoc whom I recruited to the lab in November 2011. I supported research and training for five high school students, 18 undergraduates, and two K-12 educators. I developed and led a two-day workshop on teaching in fungal biology for graduate students, postdocs, and young faculty, held through a National Science Foundation Research Coordination Network. My research program grew significantly with newly garnered funding from NSF and NIH. In addition to publications listed below, my group and I presented our work in numerous international and national venues (7 invited talks; 4 additional contributed presentations). I led two days of intensive outreach/research activities with citizen scientists at the 2011 National Geographic BioBlitz and received news coverage for these activities. I submitted grant proposals to the National Science Foundation, National Institutes of Health, and numerous smaller agencies/sources. Finally, I continued a strong tradition of service by participating in several departmental committees (graduate program committee, curriculum committee, and others), as Curator of the Robert L. Gilbertson Mycological Herbarium, and in nationally elected or appointed roles in societies in my discipline. I received a college-level award for outstanding teaching and was recognized by the International Mycological Society as the outstanding young mycologist in North America. PARTICIPANTS: PARTICIPANTS: Participants in 2011 included four Ph.D. students whom I advise (Mariana del Olmo, Mary Jane Epps, Ellen Suurmeyer, and Jana U'Ren), one MS student (Dustin Sandberg), two additional Ph.D. students for whom I have provided significant training (Demetra Kandelepas, LSU; Simon Stump, UA), five high school student interns (Frankie Orozco, Ochana Otto, Wes MacDonald, Alexia Avey, Amber Ross), 18 undergraduate assistants and young researchers (Kayla Arendt, Lauren Bautista, Brett Baxter, Duan Copeland, MM Nandi Devan, Lauren Dominick, Chan Jung, Nick Massimo, Thaddeus Metz, Christie Moss, Jamie Moy, Frankie Orozco, Brittany Pena, Ethan Posey, Syed Ali Raza, Cole Steen, Adrian Ramirez, Jakob Riddle), researchers, and volunteers, and two K-12 educators (Furr, Wilch). TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Outcomes and impacts of this Hatch award in 2011 included the establishment of new collaborations, the furthering of existing research partnerships with nationally and internationally recognized groups, advancement of my students toward degree completion, garnering of numerous awards by students whom I mentor, and reception of local and international awards.

Publications

  • Seven papers were published in 2011. An additional seven papers are currently in review (not listed below). URen, J.M., F. Lutzoni, J. Miadlikowska, A. Laetsch, and A.E. Arnold. Host and geographic structure of endophytic and endolichenic fungi at a continental scale. American Journal of Botany, in press. Lawrence, D.P., B.M. Pryor, S.B. Kroken, and A.E. Arnold. 2011. Interkingdom horizontal gene transfer of a hybrid NRPS/PKS from bacteria to filamentous Ascomycota. PLoS ONE, in press. Higgins, K.L., A.E. Arnold, P. Coley, and T. Kursar. 2012. Communities of fungal endophytes in tropical forest grasses: highly diverse host- and habitat generalists characterized by strong spatial structure. Fungal Ecology, in press. Molinar, E., N. Rios, C. Spadafora, A.E. Arnold, P.D. Coley, T.A. Kursar, W.H. Gerwick, and L. Cubilla-Rios. 2012. Coibanoles, a new class of meroterpenoids produced by Pycnoporus sanguineus. Tetrahedron Letters, in press. Wijeratne, E.M.K., B.P. Bashyal, M.X. Liu, D.D. Rocha, G.M. Kamal, B. Gunaherath, J.M. URen, M.K. Gunatilaka, A.E. Arnold, L. Whitesell, and A.A.L. Gunatilaka. 2012. Geopyxins A-E, ent-kaurane diterpenoids from endolichenic fungal strains, Geopyxis aff. majalis and Geopyxis sp. AZ0066: structure-activity relationships of geopyxins and their analogues. Journal of Natural Products, in press. Wang, X.-N., B.P. Bashyal, E.M.K. Wijeratne, J.M. URen, M. Gunatilaka, A.E. Arnold, and A.A. Leslie Gunatilaka. 2011. Smardaesidins A-G, new isopimarane and 20-Nor-isopimarane diterpenoids isolated from Smardaea sp., endophyte of the moss Ceratodon purpureus. Journal of Natural Products, in press. Moreno, E., T. Varughese, C. Spadafora, A.E. Arnold, P.D. Coley, T.A. Kursar, W.H. Gerwick, and L. Cubilla-Rios. 2011. Chemical constituents of the new endophytic fungus Mycosphaerella sp. nov. and their antiparasitic activity. Natural Products Communications 6: 835-840.


Progress 01/01/09 to 12/31/09

Outputs
OUTPUTS: Outputs from this project in 2009 included activities in teaching, research, and service. In 2009 I convened four courses that were thematically tied to ARZT-136823-H-25-195. I taught two one-semester discussions focusing on fungal ecology and systematics for graduate students and advanced undergraduates (Topics in Fungal Ecology and Evolution) and one undergraduate course serving approximately 50 students from diverse majors (Microbial Diversity), as well as co-taught Advanced Mycology for graduate students. Together these efforts served more than 65 students and benefited greatly from links with my research program. In 2009 I continued to mentor my current group of graduate students (5 Ph.D. students), two of whom passed prelims and one of whom will defend in early 2010. I supervised a postdoctoral associate based in Panama and supported research and training for a large number of young researchers, including high school students, undergraduates, graduate students conducting rotations or seeking training in our areas of expertise, and K-12 educators (including one teacher from the Navajo Nation). I also developed and led a week-long bilingual workshop on fungal systematics in Panama, in partnership with INDICASAT. My research program grew significantly in 2009, with progress on all major projects. In addition to publications listed below, my group and I presented our work in numerous venues. My invited presentations included seminars presented at the University of California, Berkeley, Arizona State University, University of Indiana, and symposium presentations at the NIH International Cooperative Biodiversity Group, Panama, Botanical Society of America, and the International Symbiosis Society. I was a coauthor on seven additional contributed presentations at national and international meetings. I submitted grant proposals to the National Science Foundation, National Institutes of Health, and numerous smaller agencies/sources. Finally, I continued a strong tradition of service by participating in several departmental committees (for example: chair, graduate recruitment committee; Member, graduate program committee), as Curator of the Robert L. Gilbertson Mycological Herbarium, and in nationally elected or appointed roles in societies in my discipline. PARTICIPANTS: Participants in 2009 included five Ph.D. students whom I advise (Mariana del Olmo, Mary Jane Epps, Michele Hoffman, Ellen Suurmeyer, and Jana U'Ren), additional Ph.D. students for whom I have provided significant training (Demetra Kandelepas, LSU; Simon Stump, UA), a high school student intern (Frankie Orozco), numerous undergraduate assistants, researchers, and volunteers (Douglas Mahana, Jamal Alafifi, Alex Lovinger, Dustin Sandberg, Barbara Beauchamp, Darren Stensrud, Andrea Woodard, Patrick Campbell, Patricia Espiritu, Brittany Wohl), a community college student (Sheri Steidl), an instructor from a tribal college (Barb Klein), and and a middle school teacher from the Navajo Nation (Rowena Dolino). TARGET AUDIENCES: Efforts include the teaching, mentorship, and outreach activities listed above, as well as a number of guest lectures and interactions with the public. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
In 2009 the outcomes and impacts of this Hatch award included the establishment of new collaborations, the furthering of existing research partnerships with nationally and internationally recognized groups, advancement of my students toward degree completion, garnering of numerous awards by students whom I mentor, my reception of the Tsujimoto Award from UC Berkeley, apreparation of new research proposals (currently in review), and submission of my tenure and promotion package (currently in review).

Publications

  • Arnold, A.E., L.J. Lamit, M. Bidartondo, C. Gehring, and H.S. Callahan. 2010. Interwoven branches of the plant and fungal trees of life. New Phytologist 185: 847-878.
  • Peay, K., M. Bidartondo, and A.E. Arnold. 2010. Not every fungus is everywhere: scaling to the biogeography of fungal-plant interactions across roots, shoots, and ecosystems. New Phytologist 185: 878-882.
  • U'Ren, J.M., F. Lutzoni, J. Miadlikowska, and A.E. Arnold. 2010. Community analysis reveals close affinities between endophytic and endolichenic fungi from mosses and lichens. Microbial Ecology, in press.
  • Epps, M.J. and A.E. Arnold. 2010. Diversity, abundance, and community network structure in sporocarp-associated beetle communities in the central Appalachian Mountains. Mycologia, in press.
  • Hoffman, M. and A.E. Arnold. 2010. Diverse bacteria inhabit living hyphae of phylogenetically diverse fungal endophytes. Applied and Environmental Microbiology, in press.
  • Parrent, J.L., K. Peay, A.E. Arnold, L. Comas, P. Avis, and A. Tuininga. 2010. Moving from pattern to process in fungal symbioses: linking functional traits, community ecology, and phylogenetics. New Phytologist 185: 882-886.
  • Arnold, A.E., J. Miadlikowska, K.L. Higgins, S.D. Sarvate, P. Gugger, A. Way, V. Hofstetter, F. Kauff, and F. Lutzoni. 2009. Hyperdiverse fungal endophytes and endolichenic fungi elucidate the evolution of major ecological modes in the Ascomycota. Systematic Biology 58: 283-297.
  • Rodriguez, R., J. White, A.E. Arnold, and R. Redman. 2009. Fungal endophytes: diversity and ecological roles. New Phytologist 182: 314-330.
  • U'Ren, J.M., J.W. Dalling, R.E. Gallery, D.R. Maddison, E.C. Davis, C.M. Gibson, and A.E. Arnold. 2009. Diversity and evolutionary origins of fungi associated with seeds of a neotropical pioneer tree: a case study for analyzing fungal environmental samples. Mycological Research 113: 432-449.


Progress 01/01/07 to 12/31/07

Outputs
OUTPUTS: The goal of my research program is to elucidate the ecological roles and evolutionary history of little-known plant-symbiotic microfungi, with a focus on highly diverse fungal endophytes inhabiting asymptomatic foliage of plants. My work addresses four questions: (1) What factors influence the abundance, diversity, and distributions of fungal symbionts of plants? (2) How do unculturable species influence our understanding of microfungal diversity? (3) Do phylogenetic relationships among endophytes reflect coevolution with plant hosts? (4) How can these diverse, avirulent symbionts illuminate the evolutionary pathways to virulence in fungal pathogens? In 2007, significant progress was made in addressing each question. Over 6000 novel fungi were isolated and integrated into phylogenetically referenced collections; together, these fungi provide the basis for a diversity of studies integrating systematics, fungal ecology, plant pathology, evolutionary biology, and bioprospecting. For example, in a study assessing the evolution of endophytism, my colleagues and I showed a high degree of evolutionary concordance between endophytic and pathogenic fungi. In a study of moth dispersal of economically important plant pathogens, we found that moths also transmit a mycoparasite that limits the fitness of the pathogen, as well as endophytic fungi that shape plant responses to disease. In a study of seed-associated fungi in tropical forests, undergraduate and graduate students working with me found strong evidence of surprising host-generalism, but also extreme spatial heterogeneity over small spatial scales. Through an international collaboration my colleagues and I demonstrated the efficacy of endophytic fungi as biological control agents against three major pathogens of the chocolate tree, Theobroma cacao. A related collaboration led to the description of a new species of endophytic fungus from Panama, and a third collaboration provided first evidence of the tremendous diversity of endophytes associated with a major commodity crop (coffee) in Colombia, Hawaii, and Mexico. Finally, two studies highlighted the novel secondary metabolites produced by these previously unknown fungi, and describe for the first time the anticancer potential of these newly discovered natural products. In addition, over the past year I have (1) received grant support from the National Science Foundation (2) submitted additional proposals to the National Institutes of Health and the National Science Foundation (currently in review); (3) presented research in 10 invited symposia or seminars and two contributed papers at national conferences; (4) mentored four Ph.D. students in my lab (all female; one from a traditionally under-represented ethnic group); (5) served on the graduate committees of nine additional Ph.D. students, seven of whom are female; (6) supported research by 13 undergraduates, including six Native American students; (7) expanded new collaborations regarding bioprospecting at the University of Arizona (Arizona fungi) and University of Utah (tropical fungi), and cellulolytic activity of Arizona fungi. PARTICIPANTS: Primary participants at the graduate level included Mariana del Olmo, Mary Jane Epps, Michele Hoffman, Jana U'Ren; short-term contributions were also made by Amritha Wickramage through a research rotation. Major participants at the undergraduate level included Christine Coursodon, Dylan Grippi, Linh Huynh, Judy Kwan, Alex Laetsch, Chan Maketon, Rebecca Porter (University of Arizona), and Faith Baldwin, Carmelita Gray, Chaudelle Begaye, Ratasha McCabe, Jacky Thomas, and Kathy Yazzie (Dine College, Navajo Nation). Technical support was provided by Malkanthi Gunatilaka.

Impacts
My research program has led to new insights regarding the scale of fungal diversity, the evolution of major ecological modes in fungi, and the application of previously unknown fungi to medicine and industry. As a result of our large-scale surveys and molecular analyses of fungi, we have developed a predictive framework for efficient bioprospecting, a suite of methods for rapidly identifying novel fungi, tools for conducting experimental manipulations of fungal symbionts, and the infrastructure necessary to expanding this body of research over the coming years. Concurrently, I have sought to create a research environment conducive to mentoring students at the undergraduate and graduate levels.

Publications

  • Rojas, E.I., E.A. Herre, L.C. Mejia, A.E. Arnold, P. Chaverri, and G.J. Samuels. 2008. Endomelanconium endophyticum, a new Botryosphaeria leaf endophyte from Panama. Mycologia. In revision.
  • Vega, F.E., A. Simpkins, M.C. Aime, F. Posada, S.W. Peterson, S.A. Rehner, F. Infante, A. Castillo, and A.E. Arnold. 2008. Fungal endophyte diversity in coffee plants from Colombia, Hawaii, Mexico and Puerto Rico. Canadian Journal of Botany. In Review.
  • Bruns, T., A.E. Arnold, and K. Hughes. 2008. Fungal networks made of humans: UNITE, FESIN, and frontiers in fungal ecology. 2008. New Phytologist 177: 586-588.
  • Hoffman, M. and A.E. Arnold. 2008. Geography and host identity interact to shape communities of endophytic fungi in cupressaceous trees. Mycological Research 112: 331-344.
  • Kithsiri Wijeratne, E.M., P.A. Paranagama, M.T. Marron, M.K. Gunatilaka, A.E. Arnold, and A.A.L. Gunatilaka. 2008. Sesquiterpene quinines and related metabolites from Phyllosticta spinarum, a fungal strain endophytic in Platycladus orientalis in the Sonoran Desert. Journal of Natural Products 71: 218-222.
  • Paranagama, P.A., E.M.K. Wijeratne, M.K. Gunatilaka, A.E. Arnold, and A.A.L. Gunatilaka. 2007. Bioactive and other naphthopyrans from Corynespora sp. occurring in Usneacavernosa: first report of metabolites of an endolichenic fungus. Journal of Natural Products 70: 1700-1705.
  • Gallery, R., J.W. Dalling, and A.E. Arnold. 2007. Diversity, host affinity, and distribution of seed-infecting fungi: A case study with neotropical Cecropia. Ecology 88: 582-588.
  • Arnold, A.E., J. Miadlikowska, K.L. Higgins, S.D. Sarvate, P. Gugger, A. Way, V. Hofstetter, F. Kauff, and F. Lutzoni. 2008. Hyperdiverse fungal endophytes and endolichenic fungi elucidate the evolution of major ecological modes in the Ascomycota. Systematic Biology. In revision.
  • Feldman, T.S., H. O,Brien, and A.E. Arnold. 2008. Moth dispersal of mycoparasites and endophytes associated with Claviceps paspali and the grass Paspalum (Poaceae). Microbial Ecology. In press.
  • Kluger, C., C. Weeks-Galindo, R. Gallery, E. Sanchez, J.W. Dalling, and A.E. Arnold. 2008. Prevalent host-generalism among fungi associated with seeds of four neotropical pioneer species. Journal of Tropical Ecology. In press.
  • Mejia, L.C., E.I. Rojas, Z. Maynard, A.E. Arnold, S. Van Bael, G.J. Samuels, N. Robbins, and E. A. Herre. 2008. Endophytic fungi as biocontrol agents of Theobroma cacao pathogens. Biological Control. In press.
  • Rodriguez, R.J., J.F. White, Jr., A.E. Arnold, and R.S. Redman. 2008. Fungal endophytes: diversity and functional roles in plant eco-physiology. New Phytologist. In press.


Progress 01/01/05 to 12/31/05

Outputs
Research goals for 2005 focused on (1) elucidating the evolutionary origins and ecological lability of endophytism (the ability to colonize and persist in foliage without causing disease); (2) developing new approaches to rapidly integrate unknown fungi into rigorous phylogenetic analyses; (3) gaining a comprehensive view of the latitudinal, geographic, and host-specific structure underlying fungal symbiont communities; and (4) exploring the secondary metabolites of these previously unknown fungi for pharmaceutical applications, and developing a predictive framework for more efficient bioprospecting. Major progress in 2005 includes a series of contributions to the fields of evolutionary biology, conservation biology, ecology, systematics, and bioprospecting. First, this project has led to a new understanding of the evolution of major ecological modes among the most economically important group of Fungi, providing new insight into the evolution of virulence and symbiosis in the Ascomycota. Comparative ecological studies have demonstrated the potential for non-native plants to harbor distinct symbiont communities relative to closely related, native hosts, highlighting the potential for symbionts to cross borders during ornamental and agricultural plant introductions. Large-scale surveys from the Arctic to tropical forest show that endophyte diversity, abundance, and host-generalism increase from the poles to the equator, and fundamentally different groups of fungi dominate in different bioclimatic zones. Collaborative work has elucidated the role of insects in transmitting endophytic fungi, and the potential for those endophytes to interact with parasitic fungal species in a model plant pathogen-grass system. We have developed new strategies for rapidly identifying novel symbiotic fungi using both fast-evolving and phylogenetically informative markers, and are using previously unknown fungi to examine the structure of the fungal tree of life. Through a new collaboration, work in my lab has led to the recovery of novel secondary metabolites from a previously unknown endophytic fungus, with potential pharmaceutical applications. In addition, progress was made toward the stated goal of obtaining extramural funding. Small grants were awarded to the PI from the Center for Tropical Forest Studies (comparative studies of endophytic fungi on three continents) and the Arizona Department of Agriculture. Four additional extramural proposals were submitted. Collaborations developed during this year have led to several new avenues of research, and to the submission of grant proposals to the USDA Tribal Colleges program, the National Science Foundation, the National Institutes of Health, and other agencies. A second proposal to NIH, and additional proposals to NSF and the Environmental Protection Agency, are anticipated during 2006. The research described here contributed to the training of four Ph.D. students (three female), three undergraduate researchers (all female; two from minority groups), two high school students (one female), and one female faculty member at a tribal college.

Impacts
This project has led to new insights regarding the scale of fungal diversity, the evolution of major ecological modes (including pathogenicity and symbiosis), the structure of the fungal tree of life, and the ecological roles of cryptic leaf-inhabiting fungi. As a result of our large-scale surveys, ecological studies, and work in phylogenetic systematics, we have developed a predictive framework for efficient bioprospecting, a robust series of methods for quickly identifying novel fungal symbionts, and an array of testable hypotheses that, when fully investigated, will contribute to our knowledge of the potential applications and cryptic ecological roles of fungal endophytes.

Publications

  • Arnold, A.E., J. Miadlikowska, K.L. Higgins, S.D. Sarvate, P. Gugger, A. Way, V. Hofstetter, F. Kauff, and F. Lutzoni. 2006. In review. Leaves and lichens are the cradles of fungal diversification. Science.
  • Hoffman, M. and A.E. Arnold. 2006. In review. Fungal endophytes in native vs. non-native Cupressaceae: community structure in mesic and xeric sites. Mycological Research.
  • Arnold, A.E. and L.C. Lewis. 2005. Evolution of fungal endophytes, and their roles against insects. Ecological and Evolutionary Advances in Insect-Fungus Associations (F. Vega and M. Blackwell, eds.). Oxford University Press, Oxford. pp. 74-96.
  • Feldman, T.S., H. O'Brien, and A.E. Arnold. 2006. In review. Moth dispersal of mycoparasites and endophytes associated with Claviceps paspali and the grass Paspalum (Poaceae). Mycological Research.
  • Gallery, R.E., J.W. Dalling, B. Wolfe, and A.E. Arnold. 2006. In review. Role of seed-infecting fungi in the recruitment limitation of neotropical pioneer species. Chapter 23 in Seed dispersal: Theory and its application in a changing world. Eds. A. Dennis, R. Green, E. Schupp and D. Westcott. CABI Press.
  • Higgins, K.L., A.E. Arnold, J. Miadlikowska, S.D. Sarvate, and F. Lutzoni. 2006. In press. Phylogenetic relationships, host affinity, and geographic structure of boreal and arctic endophytes from three major plant lineages. Molecular Phylogenetics and Evolution.
  • Arnold, A.E., D.A. Henk, R.L. Eells, F. Lutzoni, and R. Vilgalys. 2006. In press. Diversity and phylogenetic affinities of foliar fungal endophytes in loblolly pine inferred by culturing and environmental PCR. Mycologia.
  • Arnold, A.E. and F. Lutzoni. 2006. In press. Diversity and host range of foliar fungal endophytes: Are tropical leaves biodiversity hotspots? Ecology.
  • Gallery, R., J.W. Dalling, and A.E. Arnold. 2006. In press. Diversity, host affinity, and distribution of seed-infecting fungi: A case study with neotropical Cecropia. Ecology.
  • Arnold, A.E. Nonpathogenic, cryptic fungi shape plant communities in tropical forests. 2006. In press. Tropical Forest Community Ecology. (S. Schnitzer and W. Carson, eds.) Blackwell Scientific, Inc.