Attenuation of phytofungal pathogenicity of Ascomycota by autophagy modulators.

Autophagy in eukaryotes functions to maintain homeostasis by degradation and recycling of long-lived and unwanted cellular materials. Autophagy plays important roles in pathogenicity of various fungal pathogens, suggesting that autophagy is a novel target for development of antifungal compounds. Here, we describe bioluminescence resonance energy transfer (BRET)-based high-throughput screening (HTS) strategy to identify compounds that inhibit fungal ATG4 cysteine protease-mediated cleavage of ATG8 that is critical for autophagosome formation. We identified ebselen (EB) and its analogs ebselen oxide (EO) and 2-(4-methylphenyl)-1,2-benzisothiazol-3(2H)-one (PT) as inhibitors of fungal pathogens Botrytis cinerea and Magnaporthe oryzae ATG4-mediated ATG8 processing. The EB and its analogs inhibit spore germination, hyphal development, and appressorium formation in Ascomycota pathogens, B. cinerea, M. oryzae, Sclerotinia sclerotiorum and Monilinia fructicola. Treatment with EB and its analogs significantly reduced fungal pathogenicity. Our findings provide molecular insights to develop the next generation of antifungal compounds by targeting autophagy in important fungal pathogens.

Draft Genome Sequences of 28 Actinobacteria of the Family Microbacteriaceae Associated with Nematode-Infected Plants.

Draft genome sequences of 28 strains of Microbacteriaceae from plants infested by plant-parasitic nematodes were obtained using Illumina technology. The sequence data will provide useful baseline information for the development of comparative genomics and systematics of Microbacteriaceae and facilitate understanding of molecular mechanisms involved in interactions between plants and nematode-associated bacterial complexes.

The Effect of Applied Salinity and Water Stress on Chemical Suppression of Phytophthora ramorum from Soilborne Inoculum in Rhododendron.

A serious concern for nurseries is the potential for Phytophthora ramorum and other Phytophthora species to colonize roots without inducing aboveground symptoms in plants that then serve as cryptic reservoirs of inoculum. Episodic abiotic stresses that reduce plant water potential can compromise the host resistance to trigger disease development from root and crown infections during many Phytophthora-plant interactions. We conducted a series of experiments with root-inoculated Rhododendron plants in a potting soil mix to assess the influence of excess salt or water deficit on ramorum blight development and the potential for these abiotic stresses to affect the efficacy of oomycete-suppressive chemical soil treatments. During growth chamber trials, P. ramorum colonized roots of both nonsalt-treated and salt-treated plants. However, salt treatment offset the benefit realized from soil treatment with mefanoxam (Subdue Maxx) and mandipropamid (Micora), as evidenced by the enhanced pathogen colonization of roots. A 3-week episode of water stress imposed after chemical treatment but before inoculation eliminated protection against P. ramorum root colonization conferred by fosetyl-Al (Aliette). At an outdoor experimental nursery, foliar symptoms were apparent in 23% of root-inoculated plants during two trials and absent during one trial. However, the majority of inoculated plants in all trials had colonized roots with little or no aboveground symptoms. A single application of Subdue Maxx or Aliette reduced root colonization by P. ramorum in Rhododendron plants. Although salt stress did not enhance ramorum blight symptom expression at the nursery, salt partially offset protection from P. ramorum root colonization obtained by Subdue Maxx.

Analysis of Volatile Profiles for Tracking Asymptomatic Infections of Phytophthora ramorum and Other Pathogens in Rhododendron.

Phytophthora ramorum is an invasive, broad host range pathogen that causes ramorum blight and sudden oak death in forest landscapes of western North America. In commercial nurseries, asymptomatic infections of nursery stock by P. ramorum and other Phytophthora species create unacceptable risk and complicate inspection and certification programs designed to prevent introduction and spread of these pathogens. In this study, we continue development of a volatile organic compound (VOC)-based test for detecting asymptomatic infections of P. ramorum in Rhododendron sp. We confirmed detection of P. ramorum from volatiles collected from asymptomatic root-inoculated Rhododendron plants in a nursery setting, finding that the VOC profile of infected plants is detectably different from that of healthy plants, when measured from both ambient VOC emissions and VOCs extracted from leaf material. Predicting infection status was successful from ambient volatiles, which had a mean area under the curve (AUC) value of 0.71 ± 0.17, derived from corresponding receiver operating characteristic curves from an extreme gradient boosting discriminant analysis. This finding compares with that of extracted leaf volatiles, which resulted in a lower AUC value of 0.51 ± 0.21. In a growth chamber, we contrasted volatile profiles of asymptomatic Rhododendron plants having roots infected with one of three pathogens: P. ramorum, P. cactorum, and Rhizoctonia solani. Each pathogen induced unique and measurable changes, but generally the infections reduced volatile emissions until 17 weeks after inoculation, when emissions trended upward relative to those of mock-inoculated controls. Forty-five compounds had significant differences compared with mock-inoculated controls in at least one host-pathogen combination.

First report of the bent seed gall nematode, Anguina agrostis (Steinbuch, 1799) Filipjev, 1936 from Poa palustris L. in Wyoming, USA.

In September 2020, several plants of fowl bluegrass, Poa palustris with seed galls were collected on a bank of river in Teton County, Wyoming, USA. Isolated nematodes were identified by both morphological and molecular methods as Anguina agrostis. This is a first report of A. agrostis in Wyoming and its report on fowl bluegrass.

Effects of Phytophthora ramorum on volatile organic compound emissions of Rhododendron using gas chromatography-mass spectrometry.

Phytophthora ramorum is an invasive and devastating plant pathogen that causes sudden oak death in coastal forests in the western United States and ramorum blight in nursery ornamentals and native plants in various landscapes. As a broad host-range quarantine pest that can be asymptomatic in some hosts, P. ramorum presents significant challenges for regulatory efforts to detect and contain it, particularly in commercial nurseries. As part of a program to develop new detection methods for cryptic infections in nursery stock, we compared volatile emissions of P. ramorum-inoculated and noninoculated Rhododendron plants using three gas chromatography-mass spectrometry methods. The first used a branch enclosure combined with headspace sorptive extraction to measure plant volatiles in situ. Seventy-eight compounds were found in the general Rhododendron profile. The volatile profile of inoculated but asymptomatic plants (121 days post-inoculation) was distinguishable from the profile of the noninoculated controls. Three compounds were less abundant in inoculated Rhododendron plants relative to noninoculated and mock-inoculated control plants. A second method employed stir bar sorptive extraction to measure volatiles in vitro from leaf extractions in methanol; 114 volatiles were found in the overall profile with 30 compounds less abundant and one compound more abundant in inoculated Rhododendron plants relative to mock-inoculated plants. At 128 days post-inoculation, plants were asymptomatic and similar in appearance to the noninoculated controls, but their chemical profiles were different. In a third technique, volatiles from water runoff from the soil of potted healthy and inoculated Rhododendron plants were compared. Runoff from the inoculated plants contained four unique volatile compounds that never appeared in the runoff from mock-inoculated plants. These three volatile detection techniques could lead to innovative approaches that augment detection and diagnosis of P. ramorum and oomycete pathogens in nurseries and other settings. Graphical abstract Detection of volatile signatures may aid in discriminating healthy vs. infected but asymptomatic plants in nursery and greenhouse facilities.

Predisposition in plant disease: exploiting the nexus in abiotic and biotic stress perception and response.

Predisposition results from abiotic stresses occurring prior to infection that affect susceptibility of plants to disease. The environment is seldom optimal for plant growth, and even mild, episodic stresses can predispose plants to inoculum levels they would otherwise resist. Plant responses that are adaptive in the short term may conflict with those for resisting pathogens. Abiotic and biotic stress responses are coordinated by complex signaling networks involving phytohormones and reactive oxygen species (ROS). Abscisic acid (ABA) is a global regulator in stress response networks and an important phytohormone in plant-microbe interactions with systemic effects on resistance and susceptibility. However, extensive cross talk occurs among all the phytohormones during stress events, and the challenge is discerning those interactions that most influence disease. Identifying convergent points in the stress response circuitry is critically important in terms of understanding the fundamental biology that underscores the disease phenotype as well as translating research to improve stress tolerance and disease management in production systems.

Abscisic acid in salt stress predisposition to phytophthora root and crown rot in tomato and chrysanthemum.

Plants respond to changes in the environment with complex signaling networks, often under control of phytohormones that generate positive and negative crosstalk among downstream effectors of the response. Accordingly, brief dehydration stresses such as salinity and water deficit, which induce a rapid and transient systemic increase in levels of abscisic acid (ABA), can influence disease response pathways. ABA has been associated with susceptibility of plants to bacteria, fungi, and oomycetes but relatively little attention has been directed at its role in abiotic stress predisposition to root pathogens. This study examines the impact of brief salinity stress on infection of tomato and chrysanthemum roots by Phytophthora spp. Roots of plants in hydroponic culture exposed to a brief episode of salt (sodium chloride) stress prior to or after inoculation were severely diseased relative to nonstressed plants. Tomato roots remained in a predisposed state up to 24 h following removal from the stress. An increase in root ABA levels in tomato preceded or temporally paralleled the onset of stress-induced susceptibility, with levels declining in roots prior to recovery from the predisposed state. Exogenous ABA could substitute for salt stress and significantly enhanced pathogen colonization and disease development. ABA-deficient tomato mutants lacked the predisposition response, which could be restored by complementation of the mutant with exogenous ABA. In contrast, ethylene, which exacerbates disease symptoms in some host-parasite interactions, did not appear to contribute to the predisposition response. Thus, several lines of evidence support ABA as a critical and dominant factor in the salinity-induced predisposition to Phytophthora spp. infection.

Episodic Abiotic Stress as a Potential Contributing Factor to Onset and Severity of Disease Caused by Phytophthora ramorum in Rhododendron and Viburnum.

Phytophthora ramorum attacks many forest and nursery species, primarily causing trunk or stem cankers, foliar blight, and dieback, and in some species root infection has been demonstrated. However, the abiotic and edaphic factors that influence infection and disease development are unresolved. Root infection by P. ramorum and the potential for mild abiotic stress in disease predisposition was examined with Rhododendron hybrid Cunningham's White and Viburnum tinus cv. Spring Bouquet. To impose water stress in a uniform and synchronous manner, osmotic stress induced with 0.2 M NaCl was selected. Roots were exposed to NaCl for 16 to 24 h in modified hydroponic culture or standard potting soil, removed from the NaCl, and then inoculated with zoospores. In the hydroponic regime, disease symptoms developed in Rhododendron and V. tinus plants within 1 week after inoculation of salt-stressed roots, whereas symptom development was delayed in nonstressed, inoculated plants. Microscopic examination of roots from both species revealed that their apices were covered with sporangia of P. ramorum. On potted Rhododendron plants inoculated by applying zoospores directly to the soil, stem lesions developed rapidly in salt-stressed plants, with death of the plant occurring within 4 weeks after inoculation. Nonstressed plants survived for 6 to 8 weeks before succumbing to disease, and symptom development in these plants was delayed by 1 to 2 weeks relative to the inoculated, salt-stressed plants. A postinfection episode of salt stress to inoculated roots in the hydroponic regime resulted in significantly faster development of stem lesions in Rhododendron relative to nonstressed, inoculated plants.