Following basal stem rot in young oil palm plantings

The PCR primer GanET has previously been shown to be suitable for the specific amplification of DNA from Ganoderma boninense. A DNA extraction and PCR method has been developed that allows for the amplification of the G. boninense DNA from environmental samples of oil palm tissue. The GanET primer reaction was used in conjunction with a palm-sampling programme to investigate the possible infection of young palms through cut frond base surfaces. Ganoderma DNA was detected in frond base material at a greater frequency than would be expected by comparison with current infection levels. Comparisons are made between the height of the frond base infected, the number of frond bases infected, and subsequent development of basal stem rot. The preliminary results suggest that the development of basal stem rot may be more likely to occur when young lower frond bases are infected

Finding needles in haystacks: linking scientific names, reference specimens and molecular data for Fungi

DNA phylogenetic comparisons have shown that morphology-based species recognition often underestimates fungal diversity. Therefore, the need for accurate DNA sequence data, tied to both correct taxonomic names and clearly annotated specimen data, has never been greater. Furthermore, the growing number of molecular ecology and microbiome projects using high-throughput sequencing require fast and effective methods for en masse species assignments. In this article, we focus on selecting and re-annotating a set of marker reference sequences that represent each currently accepted order of Fungi. The particular focus is on sequences from the internal transcribed spacer region in the nuclear ribosomal cistron, derived from type specimens and/or ex-type cultures. Re-annotated and verified sequences were deposited in a curated public database at the National Center for Biotechnology Information (NCBI), namely the RefSeq Targeted Loci (RTL) database, and will be visible during routine sequence similarity searches with NR_prefixed accession numbers. A set of standards and protocols is proposed to improve the data quality of new sequences, and we suggest how type and other reference sequences can be used to improve identification of Fungi

Paleogene Radiation of a Plant Pathogenic Mushroom

Background: The global movement and speciation of fungal plant pathogens is important, especially because of the economic losses they cause and the ease with which they are able to spread across large areas. Understanding the biogeography and origin of these plant pathogens can provide insights regarding their dispersal and current day distribution. We tested the hypothesis of a Gondwanan origin of the plant pathogenic mushroom genus Armillaria and the currently accepted premise that vicariance accounts for the extant distribution of the species. Methods: The phylogeny of a selection of Armillaria species was reconstructed based on Maximum Parsimony (MP), Maximum Likelihood (ML) and Bayesian Inference (BI). A timeline was then placed on the divergence of lineages using a Bayesian relaxed molecular clock approach. Results: Phylogenetic analyses of sequenced data for three combined nuclear regions provided strong support for three major geographically defined clades: Holarctic, South American-Australasian and African. Molecular dating placed the initial radiation of the genus at 54 million years ago within the Early Paleogene, postdating the tectonic break-up of Gondwana. Conclusions: The distribution of extant Armillaria species is the result of ancient long-distance dispersal rather than vicariance due to continental drift. As these finding are contrary to most prior vicariance hypotheses for fungi, our result

Factors That Affect Large Subunit Ribosomal DNA Amplicon Sequencing Studies of Fungal Communities: Classification Method, Primer Choice, and Error

Nuclear large subunit ribosomal DNA is widely used in fungal phylogenetics and to an increasing extent also amplicon-based environmental sequencing. The relatively short reads produced by next-generation sequencing, however, makes primer choice and sequence error important variables for obtaining accurate taxonomic classifications. In this simulation study we tested the performance of three classification methods: 1) a similarity-based method (BLAST + Metagenomic Analyzer, MEGAN); 2) a composition-based method (Ribosomal Database Project naïve Bayesian classifier, NBC); and, 3) a phylogeny-based method (Statistical Assignment Package, SAP). We also tested the effects of sequence length, primer choice, and sequence error on classification accuracy and perceived community composition. Using a leave-one-out cross validation approach, results for classifications to the genus rank were as follows: BLAST + MEGAN had the lowest error rate and was particularly robust to sequence error; SAP accuracy was highest when long LSU query sequences were classified; and, NBC runs significantly faster than the other tested methods. All methods performed poorly with the shortest 50–100 bp sequences. Increasing simulated sequence error reduced classification accuracy. Community shifts were detected due to sequence error and primer selection even though there was no change in the underlying community composition. Short read datasets from individual primers, as well as pooled datasets, appear to only approximate the true community composition. We hope this work informs investigators of some of the factors that affect the quality and interpretation of their environmental gene surveys

Opportunistic, human-pathogenic species in the Herpotrichiellaceae are phenotypically similar to saprobic or phytopathogenic species in the Venturiaceae

Although morphologically similar, species of Cladophialophora (Herpotrichiellaceae) were shown to be phylogenetically distinct from Pseudocladosporium (Venturiaceae), which was revealed to be synonymous with the older genus, Fusicladium. Other than being associated with human disorders, species of Cladophialophora were found to also be phytopathogenic, or to occur as saprobes on organic material, or in water, fruit juices, or sports drinks, along with species of Exophiala. Caproventuria and Metacoleroa were confirmed to be synonyms of Venturia, which has Fusicladium (= Pseudocladosporium) anamorphs. Apiosporina, based on A. collinsii, clustered basal to the Venturia clade, and appears to represent a further synonym. Several species with a pseudocladosporium-like morphology in vitro represent a sister clade to the Venturia clade, and are unrelated to Polyscytalum. These taxa are newly described in Fusicladium, which is morphologically close to Anungitea, a heterogeneous genus with unknown phylogenetic affinity. In contrast to the Herpotrichiellaceae, which were shown to produce numerous synanamorphs in culture, species of the Venturiaceae were morphologically and phylogenetically more uniform. Several new species and new combinations were introduced in Cladophialophora, Cyphellophora (Herpotrichiellaceae), Exophiala, Fusicladium, Venturia (Venturiaceae), and Cylindrosympodium (incertae sedis)

Serpentine Soils Do Not Limit Mycorrhizal Fungal Diversity

Background: Physiologically stressful environments tend to host depauperate and specialized biological communities. Serpentine soils exemplify this phenomenon by imposing well-known constraints on plants; however, their effect on other organisms is still poorly understood. Methodology/Principal Findings: We used a combination of field and molecular approaches to test the hypothesis that serpentine fungal communities are species-poor and specialized. We conducted surveys of ectomycorrhizal fungal diversity from adjacent serpentine and non-serpentine sites, described fungal communities using nrDNA Internal Transcribed Spacer (ITS) fragment and sequence analyses, and compared their phylogenetic community structure. Although we detected low fungal overlap across the two habitats, we found serpentine soils to support rich fungal communities that include representatives from all major fungal lineages. We failed to detect the phylogenetic signature of endemic clades that would result from specialization and adaptive radiation within this habitat. Conclusions/Significance: Our results indicate that serpentine soils do not constitute an extreme environment for ectomycorrhizal fungi, and raise important questions about the role of symbioses in edaphic tolerance and the maintenanc

Comparing COI and ITS as DNA Barcode Markers for Mushrooms and Allies (Agaricomycotina)

DNA barcoding is an approach to rapidly identify species using short, standard genetic markers. The mitochondrial cytochrome oxidase I gene (COI) has been proposed as the universal barcode locus, but its utility for barcoding in mushrooms (ca. 20,000 species) has not been established. We succeeded in generating 167 partial COI sequences (∼450 bp) representing ∼100 morphospecies from ∼650 collections of Agaricomycotina using several sets of new primers. Large introns (∼1500 bp) at variable locations were detected in ∼5% of the sequences we obtained. We suspect that widespread presence of large introns is responsible for our low PCR success (∼30%) with this locus. We also sequenced the nuclear internal transcribed spacer rDNA regions (ITS) to compare with COI. Among the small proportion of taxa for which COI could be sequenced, COI and ITS perform similarly as a barcode. However, in a densely sampled set of closely related taxa, COI was less divergent than ITS and failed to distinguish all terminal clades. Given our results and the wealth of ITS data already available in public databases, we recommend that COI be abandoned in favor of ITS as the primary DNA barcode locus in mushrooms