R. A. Choudhury

Season-Long Dynamics of Spinach Downy Mildew Determined by Spore Trapping and Disease Incidence

Peronospora effusa is an obligate oomycete that causes downy mildew of spinach. Downy mildew threatens sustainable production of fresh market organic spinach in California, and routine fungicide sprays are often necessary for conventional production. In this study, airborne P. effusa spores were collected using rotating arm impaction spore trap samplers at four sites in the Salinas Valley between late January and early June in 2013 and 2014. Levels of P. effusa DNA were determined by a species-specific quantitative polymerase chain reaction assay. Peronospora effusa was detected prior to and during the growing season in both years. Nonlinear time series analyses on the data suggested that the within-season dynamics of P. effusa airborne inoculum are characterized by a mixture of chaotic, deterministic, and stochastic features, with successive data points somewhat predictable from the previous values in the series. Analyses of concentrations of airborne P. effusa suggest both an exponential increase in concentration over the course of the season and oscillations around the increasing average value that had season-specific periodicity around 30, 45, and 75 days, values that are close to whole multiples of the combined pathogen latent and infectious periods. Each unit increase in temperature was correlated with 1.7 to 6% increased odds of an increase in DNA copy numbers, while each unit decrease in wind speed was correlated with 4 to 12.7% increased odds of an increase in DNA copy numbers. Disease incidence was correlated with airborne P. effusa levels and weather variables, and a receiver operating characteristic curve analysis suggested that P. effusa DNA copy numbers determined from the spore traps nine days prior to disease rating could predict disease incidence.

First report of Diplodia seriata causing pear branch canker dieback in California.

California produces 26% of the United States pear crop on approximately 5,600 ha. A survey of seven northern California pear orchards (Pyrus communis cv. Bartlett) in summer 2010 revealed the presence of wedge-shaped cankers on 2- to 5-cm diameter branches, equating to 1- to 3-year-old wood. Many of the observed cankers occurred near pruning wounds, and there was decreased foliation on infected branches. Infected wood was surface disinfected with 95% ethanol and briefly flamed. After removing bark, small sections of diseased tissue were plated onto 4% potato dextrose agar (PDA) amended with 0.01% tetracycline and placed on the lab bench at 22°C until fungal growth emerged. Fungal colonies that were consistently isolated were transferred to fresh PDA using hyphal tip isolation. Fungal colonies were dark brown to gray with aerial mycelium and formed pycnidia after 15 days of incubation at 22°C. Conidia were brown, oval to oblong, and measured (16.5-) 20 to 24 (-26) × (7.5) 8.75 to 11 (-12.5) μm (n = 50). DNA from 14- to 21-day-old colonies was extracted and sequences of the rDNA internal transcribed spacer region and part of the β-tubulin gene were amplified using primers ITS4/ITS5 and Bt2a/Bt2b, respectively (2). The DNA sequences of fungal isolates from California showed 99 to 100% homology with the ex-type Diplodia seriata De Not. (1) CBS112555 deposited in GenBank. DNA sequences from three California isolates were submitted to GenBank with accession numbers KC937062, KC937065, KF481957, KF481598, KF481959, and KF481960. Pathogenicity tests were performed in March 2011 on 3-year-old Bartlett pear trees planted at an experimental farm in Davis, CA. A single, circular, 2-cm pruning wound at the top of the trunk was inoculated on each of three single-tree replications using 2-cm mycelial plugs from 14-day-old colonies growing on PDA. After inoculation, mycelial plugs were covered and sealed with Parafilm and aluminum foil for the duration of the trial. Three control trees were inoculated using sterile PDA plugs. Twelve months after inoculation, UCD103 and UCD105 were consistently re-isolated from the margin between necrotic and healthy tissue using the same methods described for the original isolation, and UCD102 was re-isolated in two out of three plants. The average lesion lengths of UCD102, UCD103, UCD105, and control plants were 12.5, 17.3, 23, and 1 mm, respectively. Control lesions were short and sterile, and seemed to be a physiological reaction from the plant. A second pathogenicity test was completed in 5 months beginning in June 2012. UCD105 was consistently re-isolated, and UCD102 and UCD103 were re-isolated in two out of three plants. The average lesion lengths for UCD102, UCD103, UCD105, and control plants were 2, 3, 5, and 1 mm, respectively. Compared to grapevine (Vitis vinifera), the pathogen grows more slowly in pear tissue under natural conditions. To our knowledge, this is the first report describing D. seriata as a causal agent of pear branch canker in California. Canker diseases can reduce the lifespan of perennial plants, ultimately leading to long term economic losses for growers (3). References: (1) A. J. L. Phillips et al. Fungal Diversity 25:141, 2007. (2) J. R. Urbez-Torres et al. Plant Dis. 90:1490, 2006. (3) J. R. Urbez-Torres and W. D. Gubler. Plant Dis. 93:584, 2009.

Spatiotemporal Patterns in the Airborne Dispersal of Spinach Downy Mildew

Downy mildew is the most devastating disease threatening sustainable spinach production, particularly in the organic sector. The disease is caused by the biotrophic oomycete pathogen Peronospora effusa, and the disease results in yellow lesions that render the crop unmarketable. In this study, the levels of DNA from airborne spores of P. effusa were assessed near a field of susceptible plants in Salinas, CA during the winter months of 2013-14 and 2014/15 using rotating-arm impaction spore-trap samplers that were assessed with a species-specific quantitative polymerase chain reaction (qPCR) assay. Low levels of P. effusa DNA were detectable from December through February in both winters but increased during January in both years, in correlation with observed disease incidence; sharp peaks in P. effusa DNA detection were associated with the onset of disease incidence. The incidence of downy mildew in the susceptible field displayed logistic-like dynamics but with considerable interseason variation. Analysis of the area under the disease progress curves suggested that the 2013-14 epidemic was significantly more severe than the 2014-15 epidemic. Spatial analyses indicated that disease incidence was dependent within an average range of 5.6 m, approximately equivalent to the width of three planted beds in a typical production field. The spatial distribution of spores captured during an active epidemic most closely fit a power-law distribution but could also be fit with an exponential distribution. These studies revealed two important results in the epidemiology of spinach downy mildew in California. First, they demonstrated the potential of impaction spore-trap samplers linked with a qPCR assay for indicating periods of high disease risk, as well as the detection of long-distance dispersal of P. effusa spores. Second, at the scale of individual crops, a high degree of spatial aggregation in disease incidence was revealed.

Multiple origins of downy mildews and mito-nuclear discordance within the paraphyletic genus Phytophthora

Phylogenetic relationships between thirteen species of downy mildew and 103 species of Phytophthora (plant-pathogenic oomycetes) were investigated with two nuclear and four mitochondrial loci, using several likelihood-based approaches. Three Phytophthora taxa and all downy mildew taxa were excluded from the previously recognized subgeneric clades of Phytophthora, though all were strongly supported within the paraphyletic genus. Downy mildews appear to be polyphyletic, with graminicolous downy mildews (GDM), brassicolous downy mildews (BDM) and downy mildews with colored conidia (DMCC) forming a clade with the previously unplaced Phytophthora taxon totara; downy mildews with pyriform haustoria (DMPH) were placed in their own clade with affinities to the obligate biotrophic P. cyperi. Results suggest the recognition of four additional clades within Phytophthora, but few relationships between clades could be resolved. Trees containing all twenty extant downy mildew genera were produced by adding partial coverage of seventeen additional downy mildew taxa; these trees supported the monophyly of the BDMs, DMCCs and DMPHs but suggested that the GDMs are paraphyletic in respect to the BDMs or polyphyletic. Incongruence between nuclear-only and mitochondrial-only trees suggests introgression may have occurred between several clades, particularly those containing biotrophs, questioning whether obligate biotrophic parasitism and other traits with polyphyletic distributions arose independently or were horizontally transferred. Phylogenetic approaches may be limited in their ability to resolve some of the complex relationships between the “subgeneric” clades of Phytophthora, which include twenty downy mildew genera and hundreds of species.

First Report of Powdery Mildew Caused by Podosphaera sp. on Triadica sebifera in California

Tweeted by 1 8/18/2017 First Report of Powdery Mildew Caused by Podosphaera sp. on Triadica sebifera in California | Plant Disease http://apsjournals.apsnet.org/doi/full/10.1094/PDIS-07-16-1001-PDN 2/2 resulting 569-bp sequences were deposited in GenBank (accession no. KX528023). There was a 99% identity match with several Podosphaera sp. sequences. Without chasmothecia or a type sequence, it is impossible to definitively identify our isolate to species. To our knowledge, this is the first report of powdery mildew caused by Podosphaera sp. on Chinese tallow tree in California. Powdery mildews of T. sebifera caused by Phyllactinia and Oidium species have been reported in Australia, Japan, and China, but not from North America (Farr and Rossman 2016).

A Framework for Optimizing Phytosanitary Thresholds in Seed Systems

Seedborne pathogens and pests limit production in many agricultural systems. Quarantine programs help prevent the introduction of exotic pathogens into a country, but few regulations directly apply to reducing the reintroduction and spread of endemic pathogens. Use of phytosanitary thresholds helps limit the movement of pathogen inoculum through seed, but the costs associated with rejected seed lots can be prohibitive for voluntary implementation of phytosanitary thresholds. In this paper, we outline a framework to optimize thresholds for seedborne pathogens, balancing the cost of rejected seed lots and benefit of reduced inoculum levels. The method requires relatively small amounts of data, and the accuracy and robustness of the analysis improves over time as data accumulate from seed testing. We demonstrate the method first and illustrate it with a case study of seedborne oospores of Peronospora effusa, the causal agent of spinach downy mildew. A seed lot threshold of 0.23 oospores per seed could reduce the overall number of oospores entering the production system by 90% while removing 8% of seed lots destined for distribution. Alternative mitigation strategies may result in lower economic losses to seed producers, but have uncertain efficacy. We discuss future challenges and prospects for implementing this approach.