Plant compartment and genetic variation drive microbiome composition in switchgrass roots.

Switchgrass (Panicum virgatum) is a promising biofuel crop native to the United States with genotypes that are adapted to a wide range of distinct ecosystems. Various plants have been shown to undergo symbioses with plant growth-promoting bacteria and fungi, however, plant-associated microbial communities of switchgrass have not been extensively studied to date. We present 16S ribosomal RNA gene and internal transcribed spacer (ITS) data of rhizosphere and root endosphere compartments of four switchgrass genotypes to test the hypothesis that host selection of its root microbiota prevails after transfer to non-native soil. We show that differences in bacterial, archaeal and fungal community composition and diversity are strongly driven by plant compartment and switchgrass genotypes and ecotypes. Plant-associated microbiota show an enrichment in Alphaproteobacteria and Actinobacteria as well as Sordariales and Pleosporales compared with the surrounding soil. Root associated compartments display low-complexity communities dominated and enriched in Actinobacteria, in particular Streptomyces, in the lowland genotypes, and in Alphaproteobacteria, specifically Sphingobium, in the upland genotypes. Our comprehensive root analysis serves as a snapshot of host-specific bacterial and fungal associations of switchgrass in the field and confirms that host-selected microbiomes persist after transfer to non-native soil.

Detection and phylogenetic relationships of Puccinia emaculata and Uromyces graminicola (Pucciniales) on switchgrass in New York State using rDNA sequence information.

The species of rust fungi (Pucciniales) inciting disease on switchgrass (Panicum virgatum) grown in bioenergy feedstock systems across the north-central and eastern United States remain unclear. In the present study, the species number and phylogenetic relationships of rust species affecting switchgrass were examined in 2011-2013 at two sites in New York State as well as selected sites in Alabama, Iowa, Nebraska, Pennsylvania, South Dakota, and West Virginia using ribosomal RNA gene data (partial internal transcribed spacer [ITS] 1, complete 5.8 subunit [S] and ITS2, and partial 28S). Uredinial group and teliospore morphology were also utilized to delimit taxa in collection years 2012 and 2013. Maximum likelihood, maximum parsimony, and Bayesian analyses demonstrated two monophyletic clades. Clade I consisted of Puccinia emaculata and included the majority of single-sorus samples across sites, whereas, Clade II included multiple samples from Iowa, Nebraska, and South Dakota. Single-telial samples for Clade I possessed only two-celled teliospores while Clade II samples possessed only one-celled teliospores, and hence, were readily diagnosed morphologically to P. emaculata and Uromyces graminicola, respectively. No U. graminicola sequences exist in GenBank to compare with our Clade II samples; however, based on teliospore morphology, the identity of Clade II taxa is U. graminicola.

Multilocus genotyping of Amylostereum spp. associated with Sirex noctilio and other woodwasps from Europe reveal clonal lineage introduced to the US.

Sirex noctilio is a woodwasp of Eurasian origin that was inadvertently introduced to the southern hemisphere in the 1900s and to North America over a decade ago. Its larvae bore in Pinus spp. and can cause significant mortality in pine plantations. S noctilio is associated with a symbiotic white rot fungus, Amylostereum areolatum, which females inject into trees when they oviposit and which is required for survival of developing larvae. We compared the genetic diversity of A. areolatum isolated from S. noctilio and other woodwasps collected from Europe and from northeastern North America to determine the origin of introduction(s) into the United States. Multilocus genotyping of nuclear ribosomal regions and protein coding genes revealed two widespread multilocus genotypes (MLGs) among the European samples, one of which is present in the US. The other two MLGs associated with S. noctilio in the US represented unique haplotypes. These latter two haplotypes were likely from unrepresented source populations, and together with the introduced widespread haplotype reveal multiple A. areolatum MLGs introduced by S. noctilio and indicate possible multiple S. noctilio introductions to North America from Europe. Our results also showed a lack of fidelity between woodwasp hosts and Amylostereum species.

Genetic evidence for three discrete taxa of Melampsora (Pucciniales) affecting willows (Salix spp.) in New York State.

Rust fungi in the genus Melampsora (Pucciniales) are the most important pathogens of shrub willows (Salix spp.) cultivated for biomass in New York State and temperate regions worldwide. The taxonomy and species identification of these fungi historically have been problematic as they are morphologically indistinguishable on willow and often have complex life histories. Melampsora of Salix in North America, therefore, have been circumscribed to the collective species Melampsora epitea Thüm. and further delineated to formae speciales by aecial host. Ribosomal DNA (rDNA) data was obtained from 75 collections/isolates of Melampsora in NY State affecting either native and cultivated Salix spp. or suspected alternate hosts. Maximum likelihood (ML), maximum parsimony (MP), and Bayesian (BI) analyses were conducted on three data partitions (individual and concatenated): complete internal transcribed spacer (ITS) and partial large subunit (LSU) rDNA sequences for all collections. Analyses of the ITS and concatenated ITS-LSU sequences revealed that Melampsora on native and cultivated willows in NY State consisted of three phylogenetically delineable taxa (phylotaxa); monophyly for each phylotaxon was strongly supported by ML, MP, and BI credibility values. Phylotaxa were also delimited phylogenetically by aecial host: Alpine currant (Ribes alpinum), eastern larch (Larix laricina), or balsam fir (Abies balsamea).

Association of ergot alkaloids with conidiation in Aspergillus fumigatus.

Ergot alkaloids are mycotoxins that affect the nervous and reproductive systems of exposed individuals through interactions with monoamine receptors. They have been studied more widely in ergot fungi and grass endophytes but also are found in Aspergillus fumigatus, an opportunistic human pathogen that reproduces and disseminates exclusively through conidia. The ergot alkaloids festucla-vine and fumigaclavines A, B and C are present in or on conidia of A. fumigatus. Cultures of the fungus that are free of conidia are difficult to obtain, obscuring comparisons of conidia versus vegetative hyphae as sources of the ergot alkaloids. To create conidiation-deficient strains of A. fumigatus we manipulated the bristle A gene (brlA), which controls vesicle formation or budding growth necessary for conidiation in Aspergillus spp. Disruption of brlA in A. fumigatus, via homologous recombination, resulted in a nonconidiating mutant that produced bristle-like structures instead of conidiophores and conidia. Moreover the disrupted strain failed to produce ergot alkaloids as verified by HPLC analyses. Complementation with a wild-type allele restored conidiation and ergot alkaloid production. These results suggest that ergot alkaloids are not produced within the vegetative mycelium of the fungus and are associated directly with conidiation.