Discovery of Botryosphaeria eucalypti sp. nov. from blighted Eucalyptus leaves in India.

Eucalyptus spp. are undoubtedly one of the most favored plantation trees globally. Accurately identifying Eucalyptus pathogens is therefore crucial for timely disease prevention and control. Recently, symptoms of a leaf blight disease were observed on Eucalyptus trees in plantations at Jhajjar and Karnal in the state of Haryana, northern India. Asexual morphs resembling the features of the Botryosphaeriaceae were consistently isolated from the symptomatic leaves. Morphological features coupled with DNA sequence analysis confirmed a novel species, which is described and illustrated here as Botryosphaeria eucalypti sp. nov. Conidia of the new taxon are longer and wider than those of its phylogenetic neighbors. A distinct phylogenetic position for the new taxon was established through combined analysis of the internal transcribed spacer (ITS), partial translation elongation factor-1α (tef1) and partial ß-tubulin (tub2) regions. Recombination analysis provided additional support for the new species hypothesis. The pathogenicity of the novel species was proved on Eucalyptus leaves, and Koch's postulates were fulfilled. The discovery of new Botryosphaeria species is important because it will help in understanding the species diversity, host range, possible threats and disease control in the long run.

Effects of microclimate on disease prevalence across an urbanization gradient.

Increased temperatures associated with urbanization (the "urban heat island" effect) have been shown to impact a wide range of traits across diverse taxa. At the same time, climatic conditions vary at fine spatial scales within habitats due to factors including shade from shrubs, trees, and built structures. Patches of shade may function as microclimate refugia that allow species to occur in habitats where high temperatures and/or exposure to ultraviolet radiation would otherwise be prohibitive. However, the importance of shaded microhabitats for interactions between species across urbanized landscapes remains poorly understood. Weedy plants and their foliar pathogens are a tractable system for studying how multiple scales of climatic variation influence infection prevalence. Powdery mildew pathogens are particularly well suited to this work, as these fungi can be visibly diagnosed on leaf surfaces. We studied the effects of shaded microclimates on rates of powdery mildew infection on Plantago host species in (1) "pandemic pivot" surveys in which undergraduate students recorded shade and infection status of thousands of plants along road verges in urban and suburban residential neighborhoods, (2) monthly surveys of plant populations in 22 parks along an urbanization gradient, and (3) a manipulative field experiment directly testing the effects of shade on the growth and transmission of powdery mildew. Together, our field survey results show strong positive effects of shade on mildew infection in wild Plantago populations across urban, suburban, and rural habitats. Our experiment suggests that this relationship is causal, where microclimate conditions associated with shade promote pathogen growth. Overall, infection prevalence increased with urbanization despite a negative association between urbanization and tree cover at the landscape scale. These findings highlight the importance of taking microclimate heterogeneity into account when establishing links between macroclimate or land use context and prevalence of disease.

Distribution of a Foliage Disease Fungus Within Canopies of Mature Douglas-Fir in Western Oregon.

Nothophaeocryptopus gaeumannii is a common native, endophytic fungus of Douglas-fir foliage, which causes Swiss needle cast, an important foliage disease that is considered a threat to Douglas-fir plantations in Oregon. Disease expression is influenced by fungal fruiting bodies (pseudothecia), which plug the stomata and inhibit gas exchange. Trees are impacted when pseudothecia plug stomates on 1-year-old and older needles resulting in early needle abscission. Mature (100 years+) trees appear to be less impacted from disease, and we hypothesize this is due to the greater emergence of pseudothecia on older than younger needles, which allows for more needle retention. We measured the density of pseudothecia occluding stomates across 2- to 5-year-old needles from upper, middle, and lower canopy positions of mature trees at three sites in the Oregon Coast Range and two sites in the western Oregon Cascade Mountains. Binomial generalized linear mixed model (GLMM) was used to test for the effects of canopy position (upper, middle, and lower), sites, needle age (2-5 years old), and years (2016 and 2017), and their interactions on the pseudothecia density. Pseudothecia density varied annually depending on sites, needle age and canopy positions. Pseudothecia density peaked on 3-, and 4-year-old needles, however, needles emerging from the same year, like 2-year-old needles in 2016 and 3-year-old needles in 2017 both emerged in 2014, had consistently similar patterns of pseudothecia density for both years, across site and canopy positions. Canopy position was important for 3-, and 4-year-old needles, showing less pseudothecia in the lower canopy. This research confirms that N. gaeumannii pseudothecia density is greatest in 3- and 4-year old needles in mature trees in contrast to plantations where pseudothecia density usually peaks on 2-year-old needles, and that pseudothecia density (disease severity) is generally lower in mature trees. Something about mature forest canopies and foliage appears to increase the time it takes for pseudothecia to emerge from the needles, in contrast to younger plantations, thus allowing the mature trees to have greater needle retention.

Tea Plants With Gray Blight Have Altered Root Exudates That Recruit a Beneficial Rhizosphere Microbiome to Prime Immunity Against Aboveground Pathogen Infection.

Tea gray blight disease and its existing control measures have had a negative impact on the sustainable development of tea gardens. However, our knowledge of safe and effective biological control measures is limited. It is critical to explore beneficial microbial communities in the tea rhizosphere for the control of tea gray blight. In this study, we prepared conditioned soil by inoculating Pseudopestalotiopsis camelliae-sinensis on tea seedling leaves. Thereafter, we examined the growth performance and disease resistance of fresh tea seedlings grown in conditioned and control soils. Next, the rhizosphere microbial community and root exudates of tea seedlings infected by the pathogen were analyzed. In addition, we also evaluated the effects of the rhizosphere microbial community and root exudates induced by pathogens on the performance of tea seedlings. The results showed that tea seedlings grown in conditioned soil had lower disease index values and higher growth vigor. Soil microbiome analysis revealed that the fungal and bacterial communities of the rhizosphere were altered upon infection with Ps. camelliae-sinensis. Genus-level analysis showed that the abundance of the fungi Trichoderma, Penicillium, and Gliocladiopsis and the bacteria Pseudomonas, Streptomyces, Bacillus, and Burkholderia were significantly (p < 0.05) increased in the conditioned soil. Through isolation, culture, and inoculation tests, we found that most isolates from the induced microbial genera could inhibit the infection of tea gray blight pathogen and promote tea seedling growth. The results of root exudate analysis showed that infected tea seedlings exhibited significantly higher exudate levels of phenolic acids and flavonoids and lower exudate levels of amino acids and organic acids. Exogenously applied phenolic acids and flavonoids suppressed gray blight disease by regulating the rhizosphere microbial community. In summary, our findings suggest that tea plants with gray blight can recruit beneficial rhizosphere microorganisms by altering their root exudates, thereby improving the disease resistance of tea plants growing in the same soil.

Molecular detection of Phytophthora pluvialis, the causal agent of red needle cast in Pinus radiata.

BACKGROUND: Phytophthora pluvialis was first described in 2013 and is the causal agent of red needle cast (RNC) in Pinus radiata as well as infection in Douglas fir (Pseudotsuga menziesii). A species-specific PCR is necessary for detection of this pathogen and diagnosis of RNC. OBJECTIVE: To design and validate a species-specific molecular assay for P. pluvialis using isolates from infected pine needles. METHODS: Species-specific PCR primers were generated from the ras-related GTP-binding protein 1 gene (ypt1) gene sequence, concentrating on DNA regions unique to P. pluvialis, and real-time and quantitative polymerase chain reaction (qPCR) were used to detect P. pluvialis from both artificially inoculated and naturally infected samples. RESULTS: The species-specific PCR assay was generated following P. pluvialis DNA sequence analysis. In vitro tests of the specificity of the probe-based, quantitative, polymerase chain reaction (qPCR) assay showed that no amplification was observed with other Phytophthora species including other closely-related clade 3 species, or with fungal species associated with pine or with pine DNA. The limit of detection of the qPCR assay was 2 pg/µl. When the qPCR assay was used to detect P. pluvialis in artificially-inoculated and naturally infected P. radiata needles, a PCR product was detected in all inoculated samples; the mean concentration ranges of P. pluvialis DNA in the inoculated and naturally infected samples tested were 5.9-124.5 pg/µl and 8.1-340.2 pg/µl, respectively. The assays described herein were used with serological diagnostic strips, providing the ability to identify to species level. CONCLUSIONS: The assay described herein detects P. pluvialis with high specificity and sensitivity from a range of DNA samples, including those extracted from infected plant material and serological diagnostic strips. The ability to detect and identify P. pluvialis, from infected tissues directly, provides value and practicality to diagnostics, biosecurity and research.

Foliar pathogens of eucalypts.

Species of eucalypts are commonly cultivated for solid wood and pulp products. The expansion of commercially managed eucalypt plantations has chiefly been driven by their rapid growth and suitability for propagation across a very wide variety of sites and climatic conditions. Infection of foliar fungal pathogens of eucalypts is resulting in increasingly negative impacts on commercial forest industries globally. To assist in evaluating this threat, the present study provides a global perspective on foliar pathogens of eucalypts. We treat 110 different genera including species associated with foliar disease symptoms of these hosts. The vast majority of these fungi have been grown in axenic culture, and subjected to DNA sequence analysis, resolving their phylogeny. During the course of this study several new genera and species were encountered, and these are described. New genera include: Lembosiniella (L. eucalyptorum on E. dunnii, Australia), Neosonderhenia (N. eucalypti on E. costata, Australia), Neothyriopsis (N. sphaerospora on E. camaldulensis, South Africa), Neotrichosphaeria (N. eucalypticola on E. deglupta, Australia), Nothotrimmatostroma (N. bifarium on E. dalrympleana, Australia), Nowamyces (incl. Nowamycetaceae fam. nov., N. globulus on E. globulus, Australia), and Walkaminomyces (W. medusae on E. alba, Australia). New species include (all from Australia): Disculoides fraxinoides on E. fraxinoides, Elsinoe piperitae on E. piperita, Fusculina regnans on E. regnans, Marthamyces johnstonii on E. dunnii, Neofusicoccum corticosae on E. corticosa, Neotrimmatostroma dalrympleanae on E. dalrympleana, Nowamyces piperitae on E. piperita, Phaeothyriolum dunnii on E. dunnii, Pseudophloeospora eucalyptigena on E. obliqua, Pseudophloeospora jollyi on Eucalyptus sp., Quambalaria tasmaniae on Eucalyptus sp., Q. rugosae on E. rugosa, Sonderhenia radiata on E. radiata, Teratosphaeria pseudonubilosa on E. globulus and Thyrinula dunnii on E. dunnii. A new name is also proposed for Heteroconium eucalypti as Thyrinula uruguayensis on E. dunnii, Uruguay. Although many of these genera and species are commonly associated with disease problems, several appear to be opportunists developing on stressed or dying tissues. For the majority of these fungi, pathogenicity remains to be determined. This represents an important goal for forest pathologists and biologists in the future. Consequently, this study will promote renewed interest in foliar pathogens of eucalypts, leading to investigations that will provide an improved understanding of the biology of these fungi.

Root exudates drive the soil-borne legacy of aboveground pathogen infection.

BACKGROUND: Plants are capable of building up beneficial rhizosphere communities as is evidenced by disease-suppressive soils. However, it is not known how and why soil bacterial communities are impacted by plant exposure to foliar pathogens and if such responses might improve plant performance in the presence of the pathogen. Here, we conditioned soil by growing multiple generations (five) of Arabidopsis thaliana inoculated aboveground with Pseudomonas syringae pv tomato (Pst) in the same soil. We then examined rhizosphere communities and plant performance in a subsequent generation (sixth) grown in pathogen-conditioned versus control-conditioned soil. Moreover, we assessed the role of altered root exudation profiles in shaping the root microbiome of infected plants. RESULTS: Plants grown in conditioned soil showed increased levels of jasmonic acid and improved disease resistance. Illumina Miseq 16S rRNA gene tag sequencing revealed that both rhizosphere and bulk soil bacterial communities were altered by Pst infection. Infected plants exhibited significantly higher exudation of amino acids, nucleotides, and long-chain organic acids (LCOAs) (C > 6) and lower exudation levels for sugars, alcohols, and short-chain organic acids (SCOAs) (C ≤ 6). Interestingly, addition of exogenous amino acids and LCOA also elicited a disease-suppressive response. CONCLUSION: Collectively, our data suggest that plants can recruit beneficial rhizosphere communities via modification of plant exudation patterns in response to exposure to aboveground pathogens to the benefit of subsequent plant generations.

Effect of Climatic Variables on Abundance and Dispersal of Lecanosticta acicola Spores and Their Impact on Defoliation on Eastern White Pine.

The disease complex white pine needle damage (WPND), first reported in 2006, has now escalated to an epidemic state across the northeastern United States. Although this complex is composed of several fungal species, Lecanosticta acicola is considered to be the primary causal agent. Knowledge regarding the epidemiology, specific climatic factors that affect the spread of L. acicola on eastern white pine (Pinus strobus) in natural forest settings, and potential risks repeated defoliation may have on tree health is limited. Therefore, this study examined how climatic variables affect the abundance and distance of spore dispersal of L. acicola and compared litterfall caused by defoliation versus natural needle abscission. Conidia were observed on spore traps from May through August, with a peak in abundance occurring in June, corresponding to the defoliation of second- and third-year foliage measured in litter traps. During peak spore production, relative humidity and the occurrence of rainfall was found to have the greatest influence on spore abundance. Our results will aid managers in determining how far from infected trees natural regeneration will likely be affected and predicting future disease severity based on climatic conditions.

Selection of alfalfa genotypes for resistance to the foliar pathogen Curvularia geniculata

ABSTRACT Foliar diseases impose severe restrictions on the persistence and productivity of Medicago sativa, both of which may be increased by developing disease resistant and more competitive genotypes that can improve pasture quality. We found Curvularia geniculata as the principal alfalfa foliar pathogen in the Brazilian state of Rio Grande do Sul (RS). Growth chamber experiments evaluated the resistance of alfalfa genotypes 'E1C4', 'CPPSul', 'ABT 805' and 'CUF-101' to C. geniculata as compared the control 'Crioula' genotype. These genotypes were also evaluated in field trials at a sea level site in Eldorado do Sul in central RS and at two sites £200 m above sea level, one in Bagé municipality in south west RS and the other at a farm near the town of Alto Feliz in north east RS. Plants were spray-inoculated with 1.6 x 106 ml-1 of C. geniculata spores and visually evaluated for leaf damage 14 days later. The C. geniculata infection rates varied from zero to 100%. Alfalfa persistence and forage mean dry mass (DM) production at the Eldorado site were measured during different seasons from November 2013 to January 2015 by calculating the incidence of invasive plants and morphologically separating leaves from stems and calculating both leaf and stem DM. Data were analysed using mixed statistical models. The best results for persistence and forage DM were shown by the 'CPPSul' genotypes (DM = 16,600 kg ha-1) and 'Crioula' (DM = 15,750 kg ha-1). These two genotypes will be used for subsequent investigations and selection cycles.

Pseudodidymellaceae fam. nov.: Phylogenetic affiliations of mycopappus-like genera in Dothideomycetes.

The familial placement of four genera, Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina, was taxonomically revised based on morphological observations and phylogenetic analyses of nuclear rDNA SSU, LSU, tef1, and rpb2 sequences. ITS sequences were also provided as barcode markers. A total of 130 sequences were newly obtained from 28 isolates which are phylogenetically related to Melanommataceae (Pleosporales, Dothideomycetes) and its relatives. Phylogenetic analyses and morphological observation of sexual and asexual morphs led to the conclusion that Melanommataceae should be restricted to its type genus Melanomma, which is characterised by ascomata composed of a well-developed, carbonaceous peridium, and an aposphaeria-like coelomycetous asexual morph. Although Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina are phylogenetically related to Melanommataceae, these genera are characterised by epiphyllous, lenticular ascomata with well-developed basal stroma in their sexual morphs, and mycopappus-like propagules in their asexual morphs, which are clearly different from those of Melanomma. Pseudodidymellaceae is proposed to accommodate these four genera. Although Mycodidymella and Xenostigmina have been considered synonyms of Petrakia based on sexual morphology, we show that they are distinct genera. Based on morphological observations, these genera in Pseudodidymellaceae are easily distinguished by their synasexual morphs: sigmoid, multi-septate, thin-walled, hyaline conidia (Mycodidymella); globose to ovoid, dictyosporus, thick-walled, brown conidia with cellular appendages (Petrakia); and clavate with a short rostrum, dictyosporus, thick-walled, brown conidia (Xenostigmina). A synasexual morph of Pseudodidymella was not observed. Although Alpinaria was treated as member of Melanommataceae in a previous study, it has hyaline cells at the base of ascomata and pseudopycnidial, confluent conidiomata which is atypical features in Melanommataceae, and is treated as incertae sedis.

Reassessing Vermisporium (Amphisphaeriaceae), a genus of foliar pathogens of eucalypts.

The genus Vermisporium presently accommodates 13 species, 11 of which are associated with leaf spots of eucalypts in the Southern Hemisphere. Vermisporium is chiefly distinguished from Seimatosporium (Amphisphaeriaceae) on the basis of a short exogenous basal appendage, and the absence of a recognisable apical appendage. Due to the increasing importance of these species in native forests, and confusion pertaining to their taxonomy, a revision of the genus was undertaken based on fresh collections and dried herbarium specimens. Results from DNA sequence data analyses of the nrDNA-ITS and 28S nrRNA genes for species of Vermisporium indicated the genus to be a synonym of Seimatosporium. New combinations are introduced in Seimatosporium for several species: S. acutum, S. biseptatum, S. brevicentrum, S. obtusum, S. orbiculare, S. verrucisporum and S. walkeri. An updated key to all species occurring on eucalypts is also provided.