A DNA Repair Inhibitor Isolated from an Ecuadorian Fungal Endophyte Exhibits Synthetic Lethality in PTEN-Deficient Glioblastoma.

Disruption of the tumor suppressor PTEN, either at the protein or genomic level, plays an important role in human cancer development. The high frequency of PTEN deficiency reported across several cancer subtypes positions therapeutic approaches that exploit PTEN loss-of-function with the ability to significantly impact the treatment strategies of a large patient population. Here, we report that an endophytic fungus isolated from a medicinal plant produces an inhibitor of DNA double-strand-break repair. Furthermore, the novel alkaloid product, which we have named irrepairzepine (1), demonstrated synthetic lethal targeting in PTEN-deficient glioblastoma cells. Our results uncover a new therapeutic lead for PTEN-deficient cancers and an important molecular tool toward enhancing the efficacy of current cancer treatments.

Genome of Diaporthe sp. provides insights into the potential inter-phylum transfer of a fungal sesquiterpenoid biosynthetic pathway.

Fungi have highly active secondary metabolic pathways which enable them to produce a wealth of sesquiterpenoids that are bioactive. One example is Δ6-protoilludene, the precursor to the cytotoxic illudins, which are pharmaceutically relevant as anticancer therapeutics. To date, this valuable sesquiterpene has only been identified in members of the fungal division Basidiomycota. To explore the untapped potential of fungi belonging to the division Ascomycota in producing Δ6-protoilludene, we isolated a fungal endophyte Diaporthe sp. BR109 and show that it produces a diversity of terpenoids including Δ6-protoilludene. Using a genome sequencing and mining approach 17 putative novel sesquiterpene synthases were identified in Diaporthe sp. BR109. A phylogenetic approach was used to predict which gene encodes Δ6-protoilludene synthase, which was then confirmed experimentally. These analyses reveal that the sesquiterpene synthase and its putative sesquiterpene scaffold modifying cytochrome P450(s) may have been acquired by inter-phylum horizontal gene transfer from Basidiomycota to Ascomycota. Bioinformatic analyses indicate that inter-phylum transfer of these minimal sequiterpenoid secondary metabolic pathways may have occurred in other fungi. This work provides insights into the evolution of fungal sesquiterpenoid secondary metabolic pathways in the production of pharmaceutically relevant bioactive natural products.

Stelliosphaerols A and B, Sesquiterpene-Polyol Conjugates from an Ecuadorian Fungal Endophyte.

Endophytic fungi are plant tissue-associated fungi that represent a rich resource of unexplored biological and chemical diversity. As part of an ongoing effort to characterize Amazon rainforest-derived endophytes, numerous fungi were isolated and cultured from plants collected in the Yasuní National Park in Ecuador. Of these samples, phylogenetic and morphological data revealed a previously undescribed fungus in the order Pleosporales that was cultured from the tropical tree Duroia hirsuta. Extracts from this fungal isolate displayed activity against Staphylococcus aureus and were thus subjected to detailed chemical studies. Two compounds with modest antibacterial activity were isolated, and their structures were elucidated using a combination of NMR spectroscopic analysis, LC-MS studies, and chemical degradation. These efforts led to the identification of stelliosphaerols A (1) and B (2), new sesquiterpene-polyol conjugates that are responsible, at least in part, for the S. aureus inhibitory activity of the fungal extract.

The Biological Diversity and Production of Volatile Organic Compounds by Stem-Inhabiting Endophytic Fungi of Ecuador.

Fungal endophytes colonize every major lineage of land plants without causing apparent harm to their hosts. Despite their production of interesting and potentially novel compounds, endophytes-particularly those inhabiting stem tissues-are still a vastly underexplored component of microbial diversity. In this study, we explored the diversity of over 1500 fungal endophyte isolates collected from three Ecuadorian ecosystems: lowland tropical forest, cloud forest, and coastal dry forest. We sought to determine whether Ecuador's fungal endophytes are hyperdiverse, and whether that biological diversity is reflected in the endophytes' chemical diversity. To assess this chemical diversity, we analyzed a subset of isolates for their production of volatile organic compounds (VOCs), a representative class of natural products. This study yielded a total of 1526 fungal ITS sequences comprising some 315 operational taxonomic units (OTUs), resulting in a non-asymptotic OTU accumulation curve and characterized by a Fisher's α of 120 and a Shannon Diversity score of 7.56. These figures suggest that the Ecuadorian endophytes are hyperdiverse. Furthermore, the 113 isolates screened for VOCs produced more than 140 unique compounds. These results present a mere snapshot of the remarkable biological and chemical diversity of stem-inhabiting endophytic fungi from a single neotropical country.

Multiple, novel biologically active endophytic actinomycetes isolated from upper Amazonian rainforests.

Microbial biodiversity provides an increasingly important source of medically and industrially useful compounds. We have isolated 14 actinomycete species from a collection of approximately 300 plant stem samples from the upper Amazonian rainforest in Peru. All of the cultured isolates produce substances with inhibitory activity directed at a range of potential fungal and bacterial pathogens. For some organisms, this activity is very broad in spectrum while other organisms show specific activity against a limited number of organisms. Two of these organisms preferentially inhibit bacterial test organisms over eukaryotic organisms. rDNA sequence analysis indicates that these organisms are not equivalent to any other cultured deposits in GenBank. Our results provide evidence of the untapped biodiversity in the form of biologically active microbes present within the tissues of higher plants.

Bioactive endophytes warrant intensified exploration and conservation.

BACKGROUND: A key argument in favor of conserving biodiversity is that as yet undiscovered biodiversity will yield products of great use to humans. However, the link between undiscovered biodiversity and useful products is largely conjectural. Here we provide direct evidence from bioassays of endophytes isolated from tropical plants and bioinformatic analyses that novel biology will indeed yield novel chemistry of potential value. METHODOLOGY/PRINCIPAL FINDINGS: We isolated and cultured 135 endophytic fungi and bacteria from plants collected in Peru. nrDNAs were compared to samples deposited in GenBank to ascertain the genetic novelty of cultured specimens. Ten endophytes were found to be as much as 15-30% different than any sequence in GenBank. Phylogenetic trees, using the most similar sequences in GenBank, were constructed for each endophyte to measure phylogenetic distance. Assays were also conducted on each cultured endophyte to record bioactivity, of which 65 were found to be bioactive. CONCLUSIONS/SIGNIFICANCE: The novelty of our contribution is that we have combined bioinformatic analyses that document the diversity found in environmental samples with culturing and bioassays. These results highlight the hidden hyperdiversity of endophytic fungi and the urgent need to explore and conserve hidden microbial diversity. This study also showcases how undergraduate students can obtain data of great scientific significance.

Plant endophytes as a platform for discovery-based undergraduate science education.

Project ownership is an essential but sometimes overlooked ingredient for a successful undergraduate research experience. We have embarked on an experiment in undergraduate education that targets isolation of microbes from rainforest plants and characterization of natural products as objectives for discovery-based undergraduate research.