bluuebunny/crossref_metadata_2025 huggingface dataset:
A Putative Role for Fusaric Acid in Biocontrol of the Parasitic Angiosperm Orobanche ramosa
Authors: Bouizgarne Brahim , El-Maarouf-Bouteau Hayat , Madiona Karine , Biligui Bernadette , Monestiez Michèle , Pennarun Anne Marie , Amiar Zahia , Rona Jean Pierre , Ouhdouch Yedir , El Hadrami Ismaïl , Bouteau François
Published: May 2007

Fusarium spp. are ubiquitous fungi found in soil worldwide as both pathogenic and nonpathogenic strains. The signals leading to disease or the absence of disease are poorly understood. We recently showed that fusaric acid (FA), a nonspecific toxin produced by most Fusarium spp., could elicit various plant defense responses at 100 nM without toxic effect. In this study, we checked for the effect of FA on root and root hairs, probable first site of contact between the fungi and the host. Large FA concentrations reduce root and root-hair growth and induce a rapid transient membrane hyperpolarization, followed by a large depolarization, due to the inhibition of H+-ATPase currents. Nanomolar concentrations of FA induced only an early transient membrane hyperpolarization of root hairs compatible with the induction of a signal transduction pathway. FA at 10-7 M failed to induce salicylic acid- and jasmonic acid/ethylene-dependent defense-related genes but inhibited the germination of the angiosperm parasite Orobanche ramosa in contact of FA-pretreated Arabidopsis thaliana seedlings. These data suggest that FA at nontoxic concentrations could activate signal transduction components necessary for plant-defense responses that could contribute to biocontrol activity of Fusarium spp.
A Fatty Acid Synthase Gene in Cochliobolus carbonum Required for Production of HC-Toxin, Cyclo(d-Prolyl-l-Alanyl- d-Alanyl- l-2-Amino-9,10-Epoxi-8-Oxodecanoyl)
Authors: Ahn Joong-Hoon , Walton Jonathan D.
Published: May 2007

The fungal maize pathogen Cochliobolus carbonum produces a phytotoxic and cytostatic cyclic peptide, HC-toxin, of structure cyclo(D-prolyl-L-alanyl-D-alanyl-L-Aeo), in which Aeo stands for 2-amino-9,10-epoxi-8-oxodecanoic acid. Here we report the isolation of a gene, TOXC, that is present only in HC-toxin-producing (Tox2+) fungal strains. TOXC is present in most Tox2+ strains in three functional copies, all of which are on the same chromosome as the gene encoding HC-toxin synthetase. When all copies of TOXC are mutated by targeted gene disruption, the fungus grows and sporulates normally in vitro but no longer makes HC-toxin and is not pathogenic, indicating that TOXC has a specific role in HC-toxin production and hence virulence. The TOXC mRNA is 6.5 kb and the predicted product has 2,080 amino acids and a molecular weight of 233,000. The primary amino acid sequence is highly similar (45 to 47% identity) to the β subunit of fatty acid synthase from several lower eukaryotes, and contains, in the same order as in other β subunits, domains predicted to encode acetyl transferase, enoyl reductase, dehydratase, and malonyl-palmityl transferase. The most plausible function of TOXC is to contribute to the synthesis of the decanoic acid backbone of Aeo.
Apoplastic Peroxidase Generates Superoxide Anions in Cells of Cotton Cotyledons Undergoing the Hypersensitive Reaction to Xanthomonas campestris pv. malvacearum Race 18
Authors: Martinez C. , Montillet J. L. , Bresson E. , Agnel J. P. , Dai G. H. , Daniel J. F. , Geiger J. P. , Nicole M.
Published: May 2007

Cotton cotyledons displayed a hypersensitive reaction (HR) in the resistant cultivar Reba B50 after infiltration with the avirulent race 18 of Xanthomonas campestris pv. malvacearum (Xcm). Generation of active oxygen species during the HR was studied biochemically and cytochemically. O2·¯ was detected in cotyledon disks by the cytochrome c reduction assay 3 h after inoculation. This activity was inhibited by superoxide dismutase (SOD) and by the peroxidase inhibitors salicylhydroxamic acid (SHAM) and KCN but not by the NADPH oxidase inhibitor diphenyleneiodonium chloride (DPI). Strong NADH oxidation activity also was found 3 h after inoculation in crude extracts or in apoplastic washing fluid and was dramatically decreased after treatment with SHAM or KCN. NADH oxidation was activated by 2,4- dichlorophenol and MnCl2, indicating the involvement of a peroxidase. Activity of cationic peroxidase isoforms (pI 9 to 9.5) constitutively expressed in cotyledons was found to be enhanced 3 h after inoculation in the resistant cultivar. Activities of apoplastic peroxidase(s) and H2O2 accumulation were observed cytochemically, 3 and 4 h post inoculation, respectively. When digitonin, a O2·- elicitor, was infiltrated into cotyledons of resistant and susceptible cultivars, generation of O2·¯ radicals was shown to be reduced by SOD and inhibited by SHAM and KCN as observed after infection, and also by DPI. Our results strongly suggest that cotton cotyledons contain two O2·¯- generating systems and that cells undergoing the HR in response to an avirulent race of Xcm produce O2·¯ through the activation of an apoplastic peroxidase.
Ectopic Expression of the Rhizobium etli amtB Gene Affects the Symbiosome Differentiation Process and Nodule Development
Authors: Taté Rosarita , Cermola Michele , Riccio Anna , Iaccarino Maurizio , Merrick Mike , Favre Reneé , Patriarca Eduardo J.
Published: May 2007

Under conditions of nitrogen limitation, soil bacteria of the genus Rhizobium are able to induce the development of symbiotic nodules on the roots of leguminous plants. During nodule organogenesis, bacteria are released endocytotically inside the invaded plant cells where they differentiate into their endosymbiotic form called bacteroids. Bacteroids surrounded by a plant-derived peribacteroid membrane are nondividing, organelle-like structures, called symbiosomes, that use nitrogenase to reduce N2 to ammonia. Experiments performed in vitro with isolated symbiosomes have previously led to the suggestion that the NH3 produced by the bacteroids is released as NH4+ into the plant cytosol. Furthermore, it was observed that the bacterial amtB (ammonium/methylammonium transport B) gene is switched off very early during symbiosis, just when bacteria are released into the host cells. We report here that the ectopic expression of amtB in bacteroids alters the ability of bacteria to invade the host cells and the symbiosome differentiation process. Both the NtrC protein, which controls the expression of the bacterial genes involved in NH4+ assimilation, and the nitrogenase activity are essential to observe the amtB-mediated effect. Our results support the idea that in vivo bacteroids do not take up NH4+ and demonstrate that the transcriptional down-regulation of the amtB gene is essential for an effective symbiotic interaction.
Use of Green Fluorescent Protein Color Variants Expressed on Stable Broad-Host-Range Vectors to Visualize Rhizobia Interacting with Plants
Authors: Stuurman Nico , Bras Cristina Pacios , Schlaman Helmi R. M. , Wijfjes André H. M. , Bloemberg Guido , Spaink Herman P.
Published: May 2007

We developed two sets of broad-host-range vectors that drive expression of the green fluorescent protein (GFP) or color variants thereof (henceforth collectively called autofluorescent proteins [AFPs]) from the lac promoter. These two sets are based on different replicons that are maintained in a stable fashion in Escherichia coli and rhizobia. Using specific filter sets or a dedicated confocal laser scanning microscope setup in which emitted light is split into its color components through a prism, we were able to unambiguously identify bacteria expressing enhanced cyan fluorescent protein (ECFP) or enhanced yellow fluorescent protein (EYFP) in mixtures of the two. Clearly, these vectors will be valuable tools for competition, cohabitation, and rescue studies and will also allow the visualization of interactions between genetically marked bacteria in vivo. Here, we used these vectors to visualize the interaction between rhizobia and plants. Specifically, we found that progeny from different rhizobia can be found in the same nodule or even in the same infection thread. We also visualized movements of bacteroids within plant nodule cells.
Exploring Infection of Wheat and Carbohydrate Metabolism in Mycosphaerella graminicola Transformants with Differentially Regulated Green Fluorescent Protein Expression
Authors: Rohel Eric A. , Payne Andrew C. , Fraaije Bart A. , Hollomon Derek W.
Published: May 2007

A Mycosphaerella graminicola strain transformed with the green fluorescent protein (GFP) downstream of either a carbon source-repressed promoter or a constitutive promoter was used to investigate in situ carbohydrate uptake during penetration of the fungus in wheat leaves. The promoter region of the acu-3 gene from Neurospora crassa encoding isocitrate lyase was used as a carbon source-repressed promoter. The promoter region of the Aspergillus nidulans gpdA gene encoding glyceraldehyde-3-phosphate dehydrogenase was used as a constitutive promoter. Fluorometric measurement of GFP gene expression in liquid cultures of acu-3-regulated transformants indicated that the N. crassa acu-3 promoter functions in M. graminicola as it does in N. crassa, i.e., acetate induced and carbon source repressed. Glucose, fructose, and saccharose triggered the repression, whereas mannitol, xylose, and cell wall polysaccharides did not. Monitoring the GFP level during fungal infection of wheat leaves revealed that acu-3 promoter repression occurred after penetration until sporulation, when newly differentiated pycnidiospores fluoresced. The use of GFP transformants also allowed clear visualization of M. graminicola pathogenesis. No appressoria were formed, but penetration at cell junctions was observed. These results give new insight into the biotrophic status of M. graminicola.
Arabidopsis thaliana Genes Expressed in the Early Compatible Interaction with Root-Knot Nematodes
Authors: Vercauteren Isabel , Van Der Schueren Els , Van Montagu Marc , Gheysen Godelieve
Published: May 2007

In the compatible interaction between Arabidopsis thaliana and the endoparasitic nematode Meloidogyne incognita, galls are formed on the roots of the host plant. Differential display was used to identify alterations of gene expression in young A. thaliana root galls caused by M. incognita. Six genes were confirmed as plant genes by DNA gel blot hybridizations. Significant homology was found with a trypsin inhibitor, peroxidase, mitochondrial uncoupling protein, endomembrane protein, 20S proteasome α-subunit, and diaminopimelate decarboxylase. The cellular and temporal expression of each of the six genes was analyzed by mRNA in situ hybridizations.
Rhizobium leguminosarum bv. trifolii PssP Protein Is Required for Exopolysaccharide Biosynthesis and Polymerization
Authors: Mazur Andrzej , Król Jarosław E. , Wielbo Jerzy , Urbanik-Sypniewska Teresa , Skorupska Anna
Published: May 2007

Rhizobium leguminosarum bv. trifolii produces an acidic exopolysaccharide (EPS) that is important for the induction of nitrogen-fixing nodules on clover. Recently, three genes, pssN, pssO, and pssP, possibly involved in EPS biosynthesis and polymerization were identified. The predicted protein product of the pssP gene shows a significant sequence similarity to other proteins belonging to the PCP2a family that are involved in the synthesis of high-molecular-weight EPS. An R. leguminosarum bv. trifolii TA1 mutant with the entire coding region of pssP deleted did not produce the EPS. A pssP mutant with the 5′ end of the gene disrupted produced exclusively low-molecular-weight EPS. A mutant that synthesized a functional N-terminal periplasmic domain but lacked the C-terminal part of PssP produced significantly reduced amounts of EPS with a slightly changed low to high molecular form ratio. Mutants affected in the PssP protein carrying a stable plasmid with a constitutively expressed gusA gene induced nodules on red clover that were not fully occupied by bacteria. A mutant with the entire pssP gene deleted infected only a few plant cells in the nodule. The pssP promoter-gusA reporter fusion was active in bacteroids during nodule development.
BacS: An Abundant Bacteroid Protein in Rhizobium etli Whose Expression Ex Planta Requires nifA
Authors: Jahn Olivia J. , Davila Guillermo , Romero David , Noel K. Dale
Published: May 2007

Rhizobium etli CFN42 bacteroids from bean nodules possessed an abundant 16-kDa protein (BacS) that was found in the membrane pellet after cell disruption. This protein was not detected in bacteria cultured in tryptone-yeast extract. In minimal media, it was produced at low oxygen concentration but not in a mutant whose nifA was disrupted. N-terminal sequencing of the protein led to isolation of a bacS DNA fragment. DNA hybridization and nucleotide sequencing revealed three copies of the bacS gene, all residing on the main symbiotic plasmid of strain CFN42. A stretch of 304 nucleotides, exactly conserved upstream of all three bacS open reading frames, had very close matches with the NifA and sigma 54 consensus binding sequences. The only bacS homology in the genetic sequence databases was to three hypothetical proteins of unknown function, all from rhizobial species. Mutation and genetic complementation indicated that each of the bacS genes gives rise to a BacS polypeptide. Mutants disrupted or deleted in all three genes did not produce the BacS polypeptide but were Nod+ and Fix+ on Phaseolus vulgaris.
Nonspecific Lipid-Transfer Protein Genes Expression in Grape (Vitissp.) Cells in Response to Fungal Elicitor Treatments
Authors: Gomès Eric , Sagot Emeric , Gaillard Cécile , Laquitaine Laurent , Poinssot Benoit , Sanejouand Yves-Henri , Delrot Serge , Coutos-Thévenot Pierre
Published: May 2007

Nonspecific lipid transfer proteins (nsLTPs) are small, basic cystein-rich proteins believed to be involved in plant defense mechanisms. Three cDNAs coding nsLTPs from grape (Vitis vinifera sp.) were cloned by reverse-transcriptase-polymerase chain reaction (RT-PCR) and PCR. The expression of nsLTP genes was investigated in 41B-rootstock grape cell suspension, in response to various defense-related signal molecules. Ergosterol (a fungi-specific sterol) and a proteinaceous elicitor purified from Botrytis cinerea strongly and rapidly induced the accumulation of nsLTP mRNAs. Jasmonic acid, cholesterol, and sitosterol also promoted nsLTPs mRNA accumulation, although to a lesser extent, whereas salicylic acid had no effect. High performance liquid chromatography analysis indicated that the amounts of three LTP isoforms (previously named P1, P2, and P4) were increased by ergosterol. None of the four isoforms displayed any significant antifungal properties, with the exception of the P4 isoform, which reduced Botrytis mycelium growth in vitro, but only in calcium-free medium. The results are discussed in the context of plant-pathogen interactions.
Expression of the Baculovirus p35 Protein in Tobacco Affects Cell Death Progression and Compromises N Gene-Mediated Disease Resistance Response to Tobacco mosaic virus
Authors: del Pozo Olga , Lam Eric
Published: May 2007

The p35 protein from baculovirus is a broad-range caspase inhibitor and suppresses programmed cell death in animals. We report here the effects of transgenic expression in tobacco of the p35 protein during the hypersensitive response (HR). Expression of p35 causes partial inhibition of nonhost HR triggered by bacteria and gene-for-gene HR triggered by virus. Infection of p35-expressing tobacco plants with Tobacco mosaic virus (TMV) disrupts N-mediated disease resistance, causing systemic spreading of the virus within a resistant background. Mutant variants altered in aspartate residues within the loop region of p35 are inefficient substrates for caspases in vitro, and they do not suppress caspase proteolytic activity in animal systems. Tobacco plants expressing these mutant variants of the p35 protein do not show inhibition of HR cell death or enhanced virus systemic movement. Thus, HR inhibition and TMV systemic spreading phenotype in p35-expressing plants correlate with the ability of the p35 protein to suppress caspase activity in animal systems. In addition, a C-terminal truncated variant of p35 is unable to suppress cell death in animals as well as HR cell death in transgenic tobacco. Our results provide evidence for the participation of caspase-like proteases during the HR. In addition, they suggest that timely activation of cell death is necessary for effective TMV containment within the primary infection site.
The White Barley Mutant Albostrians Shows a Supersusceptible but Symptomless Interaction Phenotype with the Hemibiotrophic Fungus Bipolaris sorokiniana
Authors: Schäfer Patrick , Hückelhoven Ralph , Kogel Karl-Heinz
Published: May 2007

Bipolaris sorokiniana (teleomorph: Cochliobolus sativus) is a cereal pathogen of increasing global concern, with most significance in Asiatic cropping systems. In order to gain insight into the mechanism of host resistance, we studied fungal development on the supersusceptible barley mutant albostrians and its parent cv. Haisa. A microscopic dissection of early fungal growth on Haisa and green albostrians leaves revealed a distinct epidermis-localized biotrophic and a mesophyll-based necrotrophic phase. White, green, and striped white-green albostrians leaves showed extreme differences in disease development. When comparing cellular defense responses, we found restriction of fungal spreading after successful infection of host mesophyll tissue to be the most important mecha nism limiting outbreak of the disease. Colonization of susceptible green leaves, but not extreme colonization of supersusceptible white albostrians leaves, was associated with macroscopically visible lesion formation and mesophyll accumulation of hydrogen peroxide (H2O2), implying a symptomless growth of the pathogen in supersusceptible host tissue. In contrast, early epidermal papilla-based resistance was closely linked to H2O2 accumulation in all leaf types. In white leaves, ascorbate peroxidase (APX), glutathione-S-transferase (GST), and the cell death regulator Bax-inhibitor-1 (BI-1) showed a stronger constitutive or pathogen responsive activation, whereas glycolate oxidase (GLOX) and catalase (CAT2) expression was stronger in green leaves. We discuss super-susceptibility and symptomless growth on the basis of the histochemical and the gene expression data.
Fusarium Wilt of Bitterroot (Lewisia cotyledon) in Italy Caused by Fusarium oxysporum
Authors: Garibaldi A. , Gilardi G. , Gullino M. L.
Published: May 2007

During the spring of 2004, bitterroot (Lewisia cotyledon (S. Wats.) B.L. Robins), a flowering perennial of interest for rock gardens, showed symptoms of a wilt disease in several commercial nurseries located in Piedmont (northern Italy). Ten to thirty percent of plants from several different nurseries were affected. Wilted plants were first observed approximately 20 days after being transplanted into containers when outside temperatures ranged from 15 to 22°C. Symptomatic plants were stunted with yellowed leaves and brown-to-black veins. The vascular browning extended into the crown and was continuous with a brown discoloration in the vascular system of the crown and upper taproot. Fusarium oxysporum, identified on the basis of colony and conidia morphology (1), was consistently and readily isolated from symptomatic vascular tissue onto a Fusarium-selective medium (2). Three isolates were transferred to casein hydrolysate and grown for 10 days to produce conidial inoculum. Healthy 30-day-old plants were inoculated by dipping roots into a (1 × 106 CFU/ml) conidial suspension. Plants were then transplanted into pots filled with steam-sterilized soil. Noninoculated plants served as a control. Plants (12 per treatment) were placed in a climatic chamber at 25°C. Wilt symptoms and vascular discoloration in the roots, crown, and veins developed within 20 days on each inoculated plant, while noninoculated plants remained healthy. F. oxysporum was consistently reisolated from symptomatic plants. The pathogenicity test was conducted twice with the same result. One isolate of the pathogen has been deposited in the ATCC culture collection. To our knowledge, this is the first report of F. oxysporum on L. cotyledon in Italy and elsewhere in the world. Currently, this disease is present in several nurseries in the Piedmont Region of Italy.

References: (1) C. Booth. Fusarium. CMI, Kew, UK, 1977. (2) H. Komada. Rev. Plant Prot. Res. 8:114, 1975.
First Report of White Mold Caused by Sclerotinia sclerotiorum on Rosmarinus officinalis ‘Prostratus’ in Italy
Authors: Garibaldi A. , Minuto A. , Gullino M. L.
Published: May 2007

Rosmarinus officinalis L. ‘Prostratus’ is an evergreen shrub that is native to the Mediterranean Region in southern Europe and grown as a potted plant in Italy. This cultivar is widely used in gardens and landscapes. During the winter of 2002, extensive chlorosis was observed on 8-month-old potted plants of R. officinalis L. ‘Prostratus’ grown outdoors in commercial farms near Albenga in northern Italy. Initial symptoms included stem necrosis at the soil level and darkening of leaves. As stem necrosis progressed, infected plants wilted and died. Wilt, characterized by the presence of soft and watery tissues, occurred within a few days on young plants. The disease infected 15% of the plants. Necrotic tissues became covered with a whitish mycelium that produced dark sclerotia. The diseased stem tissue was surface sterilized for 1 min in 1% NaOCl and plated on potato dextrose agar (PDA) amended with 100 ppm of streptomycin sulfate. Sclerotinia sclerotiorum (1) was consistently recovered from infected stem pieces. Sclerotia observed on infected plants measured 0.30 to 3.33 × 1.00 to 4.23 mm (average 1.31 × 1.88 mm). Sclerotia produced on PDA measured 0.09 to 3.08 × 0.38 to 4.05 mm (average 1.94 × 2.43 mm). Pathogenicity of three isolates obtained from infected plants and used in mixture was confirmed by inoculating 60-day-old plants grown in 14-cm-diameter pots in a glasshouse. Inoculum (wheat kernels infested with mycelium and sclerotia) for each isolate was placed on the soil surface around the base of each plant. Pathogenicity tests included three inoculated plants grown in separate pots per isolate. Three noninoculated plants grown in three pots served as controls. The inoculation trial was conducted twice. All plants were kept at temperatures ranging between 8 and 34°C (average 18°C) and watered as needed. Plants were covered with plastic for 96 h after inoculation to increase the moisture level. All inoculated plants developed symptoms of leaf yellowing within 30 days, soon followed by the appearance of white mycelium and sclerotia, and eventual wilt. Control plants remained symptomless. S. sclerotiorum was reisolated from the stems of inoculated plants. To our knowledge, this is the first report of white mold of R. officinalis L. ‘Prostratus’ in Italy and in Europe. S. sclerotiorum has been previously reported on R. officinalis in India (2) and the United States (3). The economic importance of this disease for the crop in Italy can be considered low at the moment.

References: (1) N. F. Buchwald. Den. Kgl. Veterin.er-og Landbohojskoles Aarsskrift, 32:75, 1949. (2) L. Mohan. Indian Phytopathol. 47:443, 1994. (3) M. L. Putnam. Plant Pathol. 53:252, 2004.
Avenacin Production in Creeping Bentgrass (Agrostis stolonifera) and Its Influence on the Host Range of Gaeumannomyces graminis
Authors: Thomas S. L. , Bonello P. , Lipps P. E. , Boehm M. J.
Published: May 2007

Avenacinase activity has been shown to be a key factor determining the host range of Gaeumannomyces graminis on oats (Avena sativa). G. graminis var. avenae produces avenacinase, which detoxifies the oat root saponin avenacin, enabling it to infect oats. G. graminis var. tritici does not produce avenacinase and is unable to infect oats. G. graminis var. avenae is also reported to incite take-all patch on creeping bentgrass (Agrostis stolonifera). It is unknown whether creeping bentgrass produces avenacin and if the avenacin-avenacinase interaction influences G. graminis pathogenicity on creeping bentgrass. The root extracts of six creeping bentgrass cultivars were analyzed by fluorimetry, thin-layer chromatography, and high performance liquid chromatography for avenacin content. Avenacin was not detected in any creeping bentgrass cultivars, and pathogenicity assays confirmed that both G. graminis var. avenae and G. graminis var. tritici can infect creeping bentgrass and wheat (Triticum aestivum), but only G. graminis var. avenae incited disease on oats. These results are consistent with the root analyses and confirm that avenacinase activity is not required for creeping bentgrass infection by G. graminis.
First Report of Asian Soybean Rust Caused by Phakopsora pachyrhizi on Soybean in Alabama
Authors: Mullen J. M. , Sikora E. J. , McKemy J. M. , Palm M. E. , Levy L. , DeVries-Paterson R.
Published: May 2007

On November 4, 2004, soybean leaves (Glycine max (L.) Merr) were submitted to the Auburn University Plant Diagnostic Lab by a State Department of Agriculture and Industries Inspector. Samples were collected from an 80-ha field of soybean plants in a late-reproductive-growth stage in Mobile County, Alabama. Under microscopic examination, leaves showed rust pustules in advanced stages of development with urediniospores and sori characteristic of Phakopsora spp. Uredinia were ostiolate in small, brown, angular leaf spots (2 to 3 mm) on lower leaf surfaces. Urediniospores were pale yellow-to-white, globose or ovate, 20 to 40 × 15 to 25 μm. In a subsequent visit to the field, symptoms and signs of the rust disease were observed on plants bordering the edge of the field since the majority of plants were senescent. Tan lesions on lower leaf surfaces contained small pustules surrounded by a small zone of slightly discolored necrotic tissue. Masses of tan spores covered the lower leaf surface pustules. Leaves were mailed overnight to the USDA National Identification Services (Mycology) Laboratory in Beltsville, MD. The fungal structures were confirmed to be a Phakopsora sp., and the sample was forwarded to the USDA National Plant Germplasm and Biotechnology Laboratory in Beltsville, MD. DNA was extracted from leaf pieces containing sori using the Qiagen DNeasy Plant Mini kit (Qiagen, Valencia, CA). Phakopsora pachyrhizi was detected using a real-time polymerase chain reaction (PCR) protocol (1) performed in a Cepheid SmartCycler (Sunnyvale, CA). The PCR master mix was modified to include OmniMix beads (Cepheid). The field and microscopic suspect diagnosis of P. pachyrhizi was confirmed officially by APHIS on November 18, 2004. This was the fourth USDA official confirmation of Asian soybean rust in the continental United States during 2004, and to our knowledge, this is the first report of the disease in Alabama. This report helps confirm that early occurrences of Asian soybean rust in the United States were present in other areas in addition to the first reported finding in Louisiana (2).

References: (1) R. D. Frederick et al. Phytopathology 92:217, 2002. (2) R. W. Schneider et al. Plant Dis. 89:774, 2005.
Geraldton Wax as a New Host for Agrobacterium tumefaciens in Argentina
Authors: Alippi A. M. , López A. C. , Wolcan S. M.
Published: May 2007

During 2004, Geraldton wax plants (Chamaelaucium uncinatum cv. BM Violet) from commercial greenhouses in La Plata, Argentina showed gall-like structures on collars and roots similar to those reported by Carsten et al. (2). No pathogenic fungi were associated with lesions. Bacteria isolated from galls grown on medium 1A and D1M agar yielded colonies typical of Agrobacterium sp. (4). Four isolates were selected for further study. All isolates were aerobic, gram-negative rods and produced 3-ketolactose, but did not produce alkali from l-tartrate or accumulate poly-β-hydroxybutyrate inclusions. The isolates were able to grow with 2% NaCl, at 35°C, and also on potato dextrose agar medium containing CaCO3 (4) but without acid clarification. Polymerase chain reaction analysis with primers VCF/VCR that amplified the expected 730-bp product of virC operon confirmed that all strains harboured a Ti plasmid (3,4). In addition, strains were screened for extrachromosomal DNA by the in-well lysis and electrophoresis procedure of Eckhardt with minor modifications as reported by Albiach and Lopez (1) and compared with strain Agrobacterium tumefaciens LBA 958, all Argentinean strains tested harbored a plasmid similar in size. Pathogenicity was verified on Geraldton wax and tobacco plants. Bacterial suspensions of each isolate (108 CFU/ml) were pricked into the stems. Control plants were pricked with sterile distilled water. Plants were maintained at 23 ± 3°C and symptoms were recorded after 45 days. Development of almost spherical, white-to-flesh-colored, rough, spongy and wart-like galls at the inoculation sites of the inoculated collars, stems, and trunks were registered. In Geraldton wax, as galls aged, they become dark brown to black, rough, and woody. Bacteria were reisolated from these galls fulfilling Koch's postulates. No lesions were observed on the controls. To our knowledge, this is the first report of A. tumefaciens on C. uncinatum in Argentina.

References: (1) M. R. Albiach and M. M. López. Appl. Environ. Microbiol. 58:2683, 1992. (2) E. Carstens et al. Plant Dis. 83:783, 1999. (3) H. Sawada et al. Appl. Environ. Microbiol. 61:828, 1995. (4) N. W. Schaad et al. Laboratory Guide for Identification of Plant Pathogenic Bacteria. 3rd ed. The American Phytopathological Society. St. Paul, MN, 2001.
First Report of Alfalfa mosaic virus on Rhamnus alaternus in France
Authors: Cardin L. , Moury B.
Published: May 2007

Rhamnus alaternus L. (evergreen buckthorn), family Rhamnaceae, is a small, hardy shrub from Mediterranean regions grown for its ornamental persistent green or variegated foliage. Chlorotic oak leaf or ringspot symptoms on R. alaternus leaves have been observed in southern France (Bellegarde, Gard department in 1998; Fanjaux, Aude department in 2000; and Saint-Jean-Cap-Ferrat, Alpes-Maritimes department in 2002). Samples from these three localities revealed the presence of Alfalfa mosaic virus (AMV) due to (i) the symptoms observed in an inoculated diagnostic host range previously described (1), (ii) observation of typical bullet-shaped virion particles of different sizes with transmission electron microscopy, (iii) nonpersistent transmission to Nicotiana tabacum cv. Xanthi-nc by Myzus persicae, and (iv) positive reaction in double-antibody sandwich-enzyme-linked immunosorbent assays (DAS-ELISA) to antibodies raised against AMV (gift of G. Marchoux). In addition, in Fanjaux, Viburnum tinus L. plants located close to the infected R. alaternus plants were also infected by AMV and exhibited typical intense calico mosaics (3). The close species, R. frangula L., was previously identified as a natural host for AMV in Italy (2). Following isolation from local lesions on Vigna unguiculata, the Fanjaux isolate was grown in cv. Xanthi-nc plants, where it induced a severe mosaic and stunting of the plants, and inoculated to 2-year-old virus-free R. alaternus plants either mechanically or with M. persicae (10 plants each). Plants were subsequently kept in an insect-proof greenhouse. At 8 and 12 months postinoculation, only one aphid-inoculated plant showed symptoms on young leaves and was AMV-positive in DAS-ELISA, while no mechanically infected plants were infected. This low infection level together with the rare observation of symptoms in natural conditions suggest that R. alaternus is not frequently infected by AMV.

References: (1) L. Cardin and B. Moury. Plant Dis. 84:594, 2000. (2) F. Marani and L. Giunchedi. Acta Hortic. 59:97, 1976. (3) N. Plese and D. Milicic. Phytopathol. Z. 72:219, 1971.
First Report of Asian Soybean Rust Caused by Phakopsora pachyrhizi from Mexico
Authors: Cárcamo Rodríguez A. , Rios J. Aguilar , Hernández J. R.
Published: May 2007

Leaves of soybean (Glycine max (L.) Merr.; Fabaceae) cv. Huasteca 400, with conspicuous chlorotic spots and associated hypophyllous cinnamon-brown sori, were collected in commercial soybean plantings in Ébano and Tamuín in the state of San Luis de Potosí, Mexico on 26 October 2005. Uredinia, Malupa-type, are mostly hypophyllous, minute, pulverulent, cinnamon-brown, scattered or in groups, subepidermal becoming erumpent, cone like, surrounded by paraphyses; paraphyses are cylindric to clavate, 25 to 50 × 6 to 14 μm, colorless to yellow brownish with wall thickened at the apex. Urediniospores are obovoid to broadly ellipsoidal, measuring 18 to 37 × 15 to 24 μm, and have a minutely echinulate thin wall, hyaline to pale yellowish brown. This morphology is typical of Phakopsora pachyrhizi Syd. & P. Syd. and P. meibomiae (Arthur) Arthur. DNA was extracted from leaves containing sori with the PureLink Plant DNA Reagent (Invitrogen, Carlsbad, CA), and the identity of P. pachyrhizi was confirmed by the polymerase chain reaction protocol (1) with Ppa1/Ppa2 primers at the National Phytosanitary Reference Center of Mexico. The morphological and molecular diagnosis and presence of P. pachyrhizi in Mexico was officially communicated by the North American Plant Protection Organization (NAPPO) on 16 February 2006. Asian soybean rust was reported for the first time in North America in 2004 (2). To our knowledge, this the first report of P. pachyrhizi in Mexico. Voucher specimens have been placed in the USDA National Fungus Collection as BPI 871130, BPI 871131, and BPI 871132. Images and a complete description of Asian soybean rust can be viewed at http://nt.arsgrin.gov/taxadescriptions/factsheets/index.cfm?thisapp=Phakopsorapachyr hizi .

References: (1) R. D. Frederick et al. Phytopathology 92:217, 2002. (2) R. W. Schneider et al. Plant Dis. 89:774, 2005.
New Diversity of Ralstonia solanacearum Strains Associated with Vegetable and Ornamental Crops in Florida
Authors: Ji Pingsheng , Allen Caitilyn , Sanchez-Perez Amilcar , Yao Jian , Elphinstone John G. , Jones Jeffrey B. , Momol M. Timur
Published: May 2007

In 2003 and 2004, 15 isolates of Ralstonia solanacearum were obtained from wilting plants of field-grown pepper (Capsicum annuum) in south Florida and from pot-grown hydrangea (Hydrangea paniculata and H. macrophylla) and geranium (Pelargonium × hortorum) in commercial nurseries and retention ponds in north Florida. Diagnostic immunoassays and polymerase chain reaction (PCR) analyses identified all the isolates as R. solanacearum but not race 3 biovar 2. Pathogenicity studies on tomato, pepper, and tobacco revealed that all 15 strains had similar high virulence on tomato and all caused wilting of tobacco, although there were significant differences among the strains in aggressiveness on tobacco. An indigenous Florida tomato strain, race 1 biovar 1 (Rs5), caused no disease on tobacco and little or none on pepper. The three pepper strains were more aggressive than Rs5 or two hydrangea strains on all three pepper cultivars studied. Phylogenetic analysis based on an endoglucanase gene sequence indicated that these strains had three distinct origins. The three pepper strains belonged to phylotype I biovar 3 and clustered with strains from diverse hosts in Asia belonging to sequevar 13. The six geranium strains and four of the hydrangea strains were closely related to strains in sequevar 5, a distinct subcluster of phylotype II biovar 1 strains isolated from the French West Indies and Brazil. Two other biovar 1 strains from hydrangea and strains K60, AW, and Rs5 belonged to sequevar 7 in phylotype II and probably are native to North America. None of the Florida isolates belong to the highly regulated Select Agent race 3 biovar 2 subgroup, according to both the DNA sequence analysis and the biovar phenotypic test results. However, the race 3 biovar 2-specific B2 primers weakly amplified a product from some race 1 biovar 1 strains in real-time PCR, indicating that this assay may give false positives under some conditions. Given the high cost of a misdiagnosis, it seems advisable to use at least two independent diagnostic methods to confirm that a suspect isolate is R. solanacearum R3B2. This is the first report of the presence of R. solanacearum race 1 biovar 3 or phylotype I strains in North America, and the first report confirming R. solanacearum causing natural infection of hydrangea in Florida. Thus, R. solanacearum strains that are quite distinct from presumably indigenous strains are present and can infect diverse hosts in Florida.
Curvularia Blight on Lolium perenne on Turfgrasses in Argentina
Authors: Goldring L. , Lacasa M. , Wright E. R.
Published: May 2007

Lolium perenne L. is commonly used alone or in association with blue-grass and fescues in sport fields, parks, and gardens. During 2003, symptoms of an unknown disease were observed on L. perenne turfgrass in western Buenos Aires. Initial symptoms were indefinite yellow and green dappled spots that extended downward from the leaf tip, turned brown and finally gray, causing leaf death. Segments of symptomatic leaf tissues were surface sterilized and placed on 2% potato dextrose agar in petri dishes. After 4 days at room temperature, blackish brown colonies developed with dark brown septate conidiophores. Conidia were 21 to 29 × 9 to 13 μm, 3-septa, curved at the third cell from the base that is longer and darker than the others. Cells at each end are subhyaline and intermediate cells are medium brown. These characteristics are consistent with Curvularia lunata (Wakker) Boedijng (1). Pathogenicity tests were performed in five plastic trays with substrate of natural soil and sand (1:1 [v/v]) where the turfgrass (L. perenne cv. El Cencerro) was seeded. Plants were inoculated by spraying a suspension of 2 × 106 conidia per ml of sterile distilled water. Controls were sprayed with sterile distilled water. The trays were covered with transparent plastic bags, sprayed periodically with water, and incubated at 25°C in a greenhouse for 20 days. The first symptoms were observed 3 days later. After 9 days, 24% of the grass surface area showed blight lesions. C. lunata was consistently reisolated from affected tissues. Control plants remained symptomless. To our knowledge, this is the first report of C. lunata affecting L. perenne in Argentina.

Reference: (1) M. B. Ellis and I. A. S. Gibson Cochliobolus lunatus (conidial state: Curvularia lunata). Page 474 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK, 1975.
First Report of Late Blight Caused by Phytophthora infestans on Cape Gooseberry (Physalis peruviana) in Colombia
Authors: Vargas A. M. , Correa A. , Lozano D. C. , González A. , Bernal A. J. , Restrepo S. , Jiménez P.
Published: May 2007

Late blight caused by Phytophthora infestans is the most limiting disease for several species of the Solanaceae family in Colombia. A potential host for P. infestans is Cape gooseberry (Physalis peruviana), a species belonging to the Solanaceae family. Its center of origin is the highlands of Peru and it is grown at approximately 1,500 to 3,000 m above sea level. Cape gooseberry has become an important export fruit in Colombia. Consequently, in the last few years, the area cultivated with Physalis peruviana has increased dramatically. P. infestans was isolated from this crop in the province of Cundinamarca, Colombia. Symptoms caused by this oomycete appeared initially on the leaf margins as small, irregular, necrotic spots that expanded and merged, increasing the necrotic area. These spots had a soft texture resulting from the degradation of plant tissue by the pathogen. On old lesions, white mycelia and sporangia were observed. Affected plants were rarely killed, but under favorable conditions, severe symptoms were observed in leaves and yield was reduced. Ten isolates were obtained from infected tissue by placing a lesion directly on a potato slice in a moist chamber (2). Mycelia grown on the potato slice were then transferred to rye agar. Identification of the pathogen was performed based on morphological characteristics, specifically, sporangiophores of P. infestans are compoundly branched and develop sympodially, with swellings at the points where sporangia were attached (1). Further confirmation was obtained by sequencing the internal transcribed spacer (ITS) regions (GenBank Accession Nos. EF173467-EF173476). Koch's postulates were completed in the laboratory by spray inoculating detached leaves of Cape gooseberry with a zoospore suspension obtained from each of the 10 isolates. Inoculum was prepared by flooding 10-day-old cultures with sterile distilled water to obtain a 104/ml sporangial suspension followed by zoospore induction at 4°C. Leaves were sprayed with this suspension, placed in moist chambers, and incubated at 20°C in the dark. Control leaves were sprayed with sterile distilled water. Two separate leaves were inoculated with each isolate. The pathogen was reisolated from leaf lesions in all cases. The period between infection and the appearance of symptoms ranged from 5 to 7 days. To our knowledge, this is the first report of P. infestans causing damage on Cape gooseberry in Colombia. Chemical control measures are to some extent successfully applied in most regions where solanaceous crops are grown in Colombia. Nevertheless, suitable disease management for Physalis peruviana has not been achieved and further studies on the epidemiology of the disease on this new host are needed.

References: (1) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society. St. Paul, MN, 1996. (2) G. A. Forbes et al. Phytopathology 87:375, 1997.
First Report of Blumeriella jaapii Teleomorph on Wild Cherry in Italy
Authors: Annesi T. , Motta E. , Forti E.
Published: May 2007

Leaf spot disease on wild cherry (Prunus avium L.) was frequently observed in plantations for timber production in central Italy. Acervuli and conidia of Phloeosporella (= Cylindrosporium) padi (Lib.) Arx were detected. Since the presence in Italy of its teleomorph, Blumeriella jaapii (Rehm) Arx (1), has never been reported, the life cycle of the pathogen was investigated. In the autumn of 1996, in some diseased leaves in different locations, microconidia were observed in the same acervuli that produced macroconidia. Also, in a plantation near Rome, fallen leaves with dark, stromatic spots were present. Leaf samples were put into plastic net bags and laid onto the ground under the trees, to be observed after overwintering in the field. In March, fully developed apothecia were present on the lower leaf surface, containing asci and ascospores that matched the description of B. jaapii. To verify the capability of the ascospores produced in nature to infect the host, overwintered leaf fragments with mature apothecia were fixed to the underside of the lids of petri dishes, above healthy cherry leaves laid upside down on moistened filter paper. Uninoculated, healthy leaves kept in petri dishes served as control. Dishes were incubated at room light and temperature conditions. The ascospores were readily discharged and dark spots with hypophyllous or amphigenous acervuli of P. padi developed on infected leaves 20 days later. No symptoms were present in the control leaves. In conclusion, B. jaapii can complete its life cycle in wild cherries in Italy. The possibility of spring infection by ascospores should be considered in disease management programs.

Reference: (1) S. Kaszonyi. Acta Phytopathol. Acad. Sci. Hung. 1:93, 1966.
First Report of the Cymbidium Mosaic Potexvirus (CymMV) Infecting the Terrestrial Orchid Phaius tankervilliae in Costa Rica
Authors: Moreira L. , Villalobos W. , Hsu H. T. , Rodríguez-Cerezo E. , Rivera C.
Published: May 2007

In 1996, plants of the terrestrial orchid Phaius tankervilliae from a nursery in the Central Valley of Costa Rica were observed with mild to severe foliar symptoms of chlorotic streak. No differences were observed in growth, bulb production, flowers, or flowering time between symptomatic and asymptomatic plants, except the symptomatic plants had earlier senescence. Occasionally, the flowers displayed symptoms of chlorosis and white rings in the sepals. Extracts from symptomatic leaves were concentrated by differential centrifugation and analyzed after sucrose gradients. Negative staining of fractions from gradients from symptomatic plants showed the presence of filamentous viral particles 500 by 17 nm. Purified particles contained a single major protein of about 28 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and a single RNA of about 7 kb, which is greater than the 6.2 kb reported (GenBank). These data suggest the presence of a potexvirus in symptomatic plants (1,2). In enzyme-linked immunosorbent assays, symptomatic plants reacted strongly with antiserum specific for Cymbidium mosaic potexvirus (CymMV). This is the first report of CymMV in P. tankervilliae in Costa Rica.

References: (1) J. A. Frowd and J. H. Tremaine. Phytopathology 67:43, 1977. (2) H. T. Hsu et al. Phytopathology 82:491, 1992.
Recent Outbreak of “Mal de Rio Cuarto” Virus on Corn in Argentina
Authors: Lenardon S. L. , March G. J. , Nome S. F. , Ornaghi J. A.
Published: May 2007

“Mal de Río Cuarto” (MRC) is the most important viral disease affecting corn in Argentina. Reovirus-like particles were observed in diseased plants (1,4) and were later serologically related to an isolate of maize rough dwarf virus (3), though this relationship was recently questioned (2). Based on estimates of the prevalence and severity of MRC and yield losses, government agencies, corn hybrid seed companies, and growers agreed that the worst epidemic in the country occurred during the 1996 to 1997 agricultural year. Approximately 300,000 ha of corn were affected by the disease and yield losses were estimated at $120 million. Affected areas included the central and southern Santa Fe, the central, northern, southeastern, and western Buenos Aires, and the eastern and southern (originally the endemic center of MRC in Río Cuarto County) parts of Córdoba. Virus infections were confirmed by double-antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) in root samples from each surveyed location, using an antiserum to MRC virus. The occurrence of MRC in non-endemic areas suggests an unusual phenological coincidence of high vector populations, abundant natural virus reservoirs, and susceptible stages in the crop. Most commercial hybrids surveyed were apparently susceptible to the virus, although some were tolerant.

References: (1) O. E. Bradfute et al. Phytopathology 71:205, 1981. (2) C. Marzachi et al. Sem. Virol. 6:103, 1995. (3) R. G. Milne et al. Phytopathology 73:1290, 1983. (4) S. F. Nome et al. Phytopathol. Z. 101:7, 1981.
Expansion of Potato Late Blight Forecasting Models for the Columbia Basin of Washington and Oregon
Authors: Johnson Dennis A. , Alldredge J. Richard , Hamm Philip B.
Published: May 2007

A regional potato late blight forecasting system for irrigated potatoes in the semiarid environment of the Columbia Basin was expanded by developing specific forecasting models for four vicinities throughout the Basin. Relationships between weather and outbreaks of late blight at the locations over a 27-year period were examined using logistic regression analysis. The response variable was a year either with or without a late blight outbreak. An indicator variable representing the occurrence of an outbreak during the preceding year (Yp) and number of days of rain during April and May (Ram) correctly classified the disease status (presence or absence of late blight) of 89, 82, 78, and 78% of the years at Prosser, Washington, Hermiston, Oregon, and Hanford and Othello, Washington, respectively. The percentage of years with disease outbreaks correctly classified was 93, 85, 79, and 79% at the four respective locations. All years with late blight outbreaks and 96% of the total years were correctly classified using data from at least one of the four locations. These predictors are particularly important early in the growing season and can be used to make area forecasts. A second set of predictors, Yp and number of days of rain in July and August (Rja), for Hermiston and Hanford, and a third set, Yp, Ram, and Rja, for Prosser and Othello were found effective for making additional late blight forecasts later in the growing season.