Plant Seeds Commonly Host Bacillus spp., Potential Antagonists of Phytopathogens.

In agriculture, horticulture and plantation forestry, Bacillus species are the most commonly applied antagonists and biopesticides, targeting plant pathogens and insect pests, respectively. Bacillus isolates are also used as bacterial plant biostimulants, or BPBs. Such useful isolates of Bacillus are typically sourced from soil. Here, we show that Bacillus - and other antagonistic microbes - can be sourced from a broad range of plant seeds. We found that culturable Bacillus isolates are common in the seeds of 98 plant species representing 39 families (i.e., 87% of the commonly cultured bacteria belonged to Bacillales). We also found that 83% of the commonly cultured fungi from the seeds of the 98 plant species belonged to just three orders of fungi-Pleosporales, Hypocreales and Eurotiales-that are also associated with antagonism. Furthermore, we confirmed antagonism potential in agaro with seed isolates of Bacillus from Pinus monticola as a representative case. Eight isolates each of seed Bacillus, seed fungi, and foliar fungi, all from P. monticola, were paired in a total of 384 possible pair-wise interactions (with seed and foliar fungi as the targets). Seed Bacillus spp. were the strongest antagonists of the seed and foliar fungi, with a mean interaction strength 2.8 times greater than seed fungi (all either Eurotiales or Hypocreales) and 3.2 times greater than needle fungi. Overall, our study demonstrates that seeds host a taxonomically narrow group of culturable, antagonistic bacteria and fungi.

Phylogenetic conservatism of mycoparasitism and its contribution to pathogen antagonism.

Closely related species are expected to have similar functional traits due to shared ancestry and phylogenetic inertia. However, few tests of this hypothesis are available for plant-associated fungal symbionts. Fungal leaf endophytes occur in all land plants and can protect their host plant from disease by a variety of mechanisms, including by parasitizing pathogens (e.g., mycoparasitism). Here, we tested whether phylogenetic relatedness among species of Cladosporium, a widespread genus that includes mycoparasitic species, predicts the effect of this endophyte on the severity of leaf rust disease. First, we used congruence among different marker sequences (i.e., genealogical concordance phylogenetic species recognition criterion) to delimit species of Cladosporium. Next, in a controlled experiment, we quantified both mycoparasitism and disease modification for the selected Cladosporium species. We identified 17 species of Cladosporium; all the species reduced rust disease severity in our experiment. Cladosporium phylogeny was a significant predictor of mycoparasitism. However, we did not observe a phylogenetic effect on disease severity overall, indicating that other mechanism/s operating independently of shared ancestry also contributed to endophyte effects on disease severity. Indeed, a second experiment showed that Cladosporium endophyte exudates (no live organism) from divergent species groups equally reduced disease severity. Our results reveal that multiple mechanisms contribute to the protective effects of an endophyte against a plant pathogen, but not all traits underlying these mechanisms are phylogenetically conserved.

Systematics of the ectomycorrhizal genus Lactarius in the Rocky Mountain alpine zone.

Lactarius (Russulales) is an important component of ectomycorrhizal fungal communities in cold-dominated contiguous arctic and disjunct alpine habitats where it associates primarily with Betula, Dryas and Salix However, little is known of this genus in the central and southern Rocky Mountain alpine zone (3000-3900 m) of North America. Molecular phylogenetic analyses of nuc rDNA ITS1-5.8S-ITS2 (ITS barcode) and the second largest subunit of the RNA polymerase II gene (RPB2) partial sequences in conjunction with detailed morphological examination confirm at least six species occurring above treeline. Most have intercontinental distributions in North America and Eurasia according to molecular comparison with type material and collections from Europe, Fennoscandia, Svalbard and Alaska. Rocky Mountain collections of L. lanceolatus (subgenus Russularia), along with the type from Alaska are paraphyletic with respect to L. aurantiacus and North American taxa L. luculentus and L. luculentus v. laetus Rocky Mountain collections of L. nanus, L. glyciosmus, L. repraesentaneus and L. salicis-reticulatae (subgenus Piperites) all form clades with European material from type localities and other arctic-alpine habitats. The arctic-alpine L. pseudouvidus/L. brunneoviolaceus group appears to be a complex containing additional taxa. North American material originally described as part of this group is well-separated phylogenetically and is described here as L. pallidomarginatus sp. nov. Lactarius lanceolatus, L. nanus and L. salicis-reticulatae appear largely restricted to arctic-alpine habitats with Salix Lactarius glyciosmus and L. repraesentaneus occur in arctic-alpine, subalpine and boreal habitats with Betula and also Picea and possibly Salix for the latter. Species distributions are hypothesized to be shaped by host ranges, glaciation and long distance dispersal.

New species and new combinations in the genus Paraisaria (Hypocreales, Ophiocordycipitaceae) from the U.S.A., supported by polyphasic analysis.

Molecular phylogenetic and chemical analyses, and morphological characterization of collections of North American Paraisaria specimens support the description of two new species and two new combinations for known species. P.cascadensissp. nov. is a pathogen of Cyphoderris (Orthoptera) from the Pacific Northwest USA and P.pseudoheteropodasp. nov. is a pathogen of cicadae (Hemiptera) from the Southeast USA. New combinations are made for Ophiocordycepsinsignis and O.monticola based on morphological, ecological, and chemical study. A new cyclopeptide family proved indispensable in providing chemotaxonomic markers for resolving species in degraded herbarium specimens for which DNA sequencing is intractable. This approach enabled the critical linkage of a 142-year-old type specimen to a phylogenetic clade. The diversity of Paraisaria in North America and the utility of chemotaxonomy for the genus are discussed.

Culturable Seed Microbiota of Populus trichocarpa.

Plants harbor a diverse community of microbes, whose interactions with their host and each other can influence plant health and fitness. While microbiota in plant vegetative tissues has been extensively studied, less is known about members of the seed microbiota. We used culture-based surveys to identify bacteria and fungi found in the seeds of the model tree, Populus trichocarpa, collected from different sites. We found that individual P. trichocarpa seeds typically contained zero or one microbe, with common taxa including species of Cladosporium, Aureobasidium, Diaporthe, Alternaria, and Pseudomonas, a bacterium. Pseudomonas isolates were associated with seed mortality and were negatively associated with the occurrence of fungal isolates within Epicoccum, Alternaria, and Aureobasidium from the same seed. Next, we conducted an inoculation experiment with one of the isolated seed microbes, Pseudomonas syringae pv. syringae, and found that it reduced seed germination and increased seedling mortality for P. trichocarpa. Our findings highlight common fungi and bacteria in the seeds of P. trichocarpa, prompting further study of their functional consequences. Moreover, our study confirms that P. syringae pv. syringae is a seed pathogen of P. trichocarpa and is the first report that P. syringae pv. syringae is a lethal seedling pathogen of P. trichocarpa, allowing for future work on the pathogenicity of this bacterium in seedlings and potential antagonism with other seed microbes.