bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of Rain Distribution Alteration on Splash Dispersal of Colletotrichum acutatum
Authors: Ntahimpera N. , Madden L. V. , Wilson L. L.
Published: May 2007

Splash dispersal of Colletotrichum acutatum conidia from infected strawberry fruit was assessed using a rain simulator to determine the properties of rain (e.g., intensity [millimeters/hour] or drop size distribution) most related to dissemination. Dispersal with a simulated rain corresponding to a natural rain of about 11 mm/h was compared with dispersal of three other simulated rains that had larger and smaller drop sizes, on average, than idealized natural rains. Splash droplets were collected in sheltered petri plates with a selective medium for Colletotrichum, and colonies formed from conidia entrained in the droplets were counted and used as the measure of dispersal. Colonies were mostly confined to a 27-cm radius from the source, and density of colonies decreased exponentially with the distance squared, as indicated by the fit of a diffusion-type model to the data. Splash dispersal was more affected by drop size distribution than rain intensity or other properties of the generated rains. That is, there was a direct positive relationship between total colonies over 61 min of rain for a circular area with a 72-cm radius (Σ) and the mass (volume) median diameter of impacting drops (D0') for four rain-simulation treatments. In a separate study, strawberry fruit were exposed to the same four simulated rains at two distances from a point source and for two rain durations. Although the proportion of infected fruit (y) increased with time and decreased with distance, rain treatment did not significantly affect y, as predicted based on past work with a wide range of intensities of simulated rains.
Purification, Properties, and Diagnosis of Banana Bract Mosaic Potyvirus and Its Distinction from Abaca Mosaic Potyvirus
Authors: Thomas J. E. , Geering A. D. W. , Gambley C. F. , Kessling A. F. , White M.
Published: May 2007

Using biochemical, serological, and cytopathological evidence, we have confirmed that banana bract mosaic virus (BBrMV) is a distinct member of the family Potyviridae. Virions of a Philippine isolate of BBrMV were purified from field-infected banana cv. Cardaba. Particles were approximately 725-nm long, banded at a density equivalent to 1.29 to 1.31 g/ml in cesium chloride equilibrium gradients, and had an A260/280 of 1.17. Yields of about 4 mg/kg were obtained from fresh or frozen leaf midrib or lamina tissue. Three major protein species with sizes of 31, 37, and 39 kDa were resolved from dissociated virions, and all reacted specifically with polyclonal antibodies to BBrMV. Infected leaf cells contained typical pinwheel inclusions. Virus-specific cDNA was amplified from field samples by reverse transcription-polymerase chain reaction (RT-PCR) assay using potyvirus degenerate primers. In plate-trapped antigen-enzyme-linked immunosorbent assay (ELISA), weak serological relationships were demonstrated between BBrMV and other members of the family Potyviridae, including abaca mosaic (AbaMV), dasheen mosaic, maize dwarf mosaic, sorghum mosaic, sugarcane mosaic, and wheat streak mosaic viruses. Despite similarities in the symptoms caused by the two viruses, AbaMV was serologically distinct from BBrMV and reacted only weakly, or not at all, with BBrMV antibodies in double-antibody sandwich (DAS)-ELISA. No cross reactions were observed when RT-PCR products from the two viruses were examined by Southern blot hybridization using BBrMV- and AbaMV-specific digoxigenin-labeled DNA probes. BBrMV was consistently associated with banana bract mosaic disease, as assessed by DAS-ELISA and Southern blot hybridization using DNA probes. The known geographical distribution of BBrMV was extended to include India (Kokkan disease) and Sri Lanka.
Quantification of Fungal Antigens in Soil with a Monoclonal Antibody-Based ELISA: Analysis and Reduction of Soil-Specific Bias
Authors: Otten W. , Gilligan C. A. , Thornton C. R.
Published: May 2007

This paper describes methods to improve the use of immunoassays for quantification of soilborne fungal antigens. Calibration curves, prepared by diluting known quantities of an antigen into soil extracts and into soil, were described by a four-parameter logistic curve from which two principal criteria, the lower detection limit and the horizontal locational parameter, were used to summarize the sensitivity and bias of an immuno-assay. We identify two sources of bias, retention of the antigen in soil due to bonding and interference of soluble soil components in plate-trapped—antigen, enzyme-linked immunosorbent assays. Using a monoclonal antibody that recognizes a putative catechol oxidase secreted by hyphae of Rhizoctonia solani, we show that bias due to retention of the antigen in soil is substantially greater than bias due to interference. Three soils were compared: a sand, a clay, and a loam. The degree of retention varied with soil type, with more than a 1,000-fold reduction in sensitivity in the clay soil. Addition of CuSO4 to the extraction solution and optimizing the volume of extractant reduced the bias and increased the sensitivity of the assay for all three soils. Possible mechanisms for the effect due to Cu2+ and the implications for the design and use of calibration curves for assays involving quantification of fungal antigens in soil are discussed.
Genetic Diversity within the Population of Xanthomonas oryzae pv. oryzae in India
Authors: Yashitola J. , Krishnaveni D. , Reddy A. P. K. , Sonti R. V.
Published: May 2007

Xanthomonas oryzae pathovar oryzae causes a serious disease of rice in India and is endemic in all of the major rice-growing areas of the country. Sixty-seven X. oryzae pv. oryzae strains, collected mostly in 1994 and 1995, from 18 locations in India were analyzed by DNA fingerprinting methods using two separate repeat element probes from the X. oryzae pv. oryzae genome. These results show that strains belonging to a single pathogen lineage can be isolated from 16 of the 18 locations sampled; many of these locations are separated from each other by hundreds of kilometers and represent ecologically diverse rice-growing areas. Pathotyping analysis indicated that the strains in this lineage belong to pathotype 1b of X. oryzae pv. oryzae.
Characteristics of Beet Yellows Closterovirus Transmission to Sugar Beets by Aphis fabae
Authors: Limburg D. D. , Mauk P. A. , Godfrey L. D.
Published: May 2007

Laboratory studies were conducted on the characteristics of beet yellows closterovirus (BYV) transmission to sugar beets by the bean aphid, Aphis fabae. The relative transmission efficiency and the retention, acquisition, and inoculation thresholds were evaluated using clonal aphid colonies. Transmission efficiency was studied between different apterous aphid species (A. fabae and Myzus persicae ), between alate and apterous morphs of A. fabae and M. persicae, and among five clonal aphid colonies of A. fabae. Results indicated, on average, that apterous M. persicae transmitted twice as efficiently as apterous A. fabae (60 and 34%, respectively), apterous forms of A. fabae were equally efficient in transmitting BYV as compared with alate A. fabae (28 and 29%, respectively), and the five clonal colonies of A. fabae ranged from 28 to 40% transmission efficiency with an average of 34%. The maximum retention time recorded for BYV by A. fabae was between 24 and 48 h. After a 1-h acquisition access, a transmission efficiency of 10% was obtained, with maximum transmission efficiency (the highest average percent transmission that is statistically different from all other previous averages) of 39% reached after 6 h of acquisition access. Inoculation thresholds occurred within 1 h, with maximum transmission efficiency (42%) reached after a 6-h inoculation access. Using an electronic monitoring system, a range of 9 to 38 min (20.5 mean) was recorded for the stylets of A. fabae to reach the phloem sieve elements. This was consistent with our threshold of a 20-min acquisition access recorded through direct observation.
Genetic Diversity and Pathogenic Variation of Colletotrichum lindemuthianum in the Three Centers of Diversity of Its Host, Phaseolus vulgaris
Authors: Sicard D. , Michalakis Y. , Dron M. , Neema C.
Published: May 2007

Population subdivision of Colletotrichum lindemuthianum, the causal agent of anthracnose, was studied in three regions located in three centers of diversity of its host, Phaseolus vulgaris. Random amplified polymorphic DNA (RAPD) markers, restriction endonuclease analysis of the amplified ribosomal internal transcribed spacer region, and virulence on a set of 12 cultivars were used to assess the genetic diversity of C. lindemuthianum strains isolated in Mexican, Ecuadorian, and Argentinean wild common bean populations. The three regions were significantly differentiated for molecular markers. For these markers, Mexico was the most polymorphic and the most distant from Ecuador and Argentina. The majority of the RAPD alleles present in Ecuador and Argentina were found in Mexico, suggesting that Andean populations have been derived from the Mesoamerican center. Pathogenicity tests on a set of 12 cultivars showed that all but one of the Mexican strains were virulent exclusively on Mesoamerican cultivars. Argentinean strains were virulent preferentially on southern Andes cultivars, and the Ecuadorian strains, except for one strain, were avirulent on all cultivars. These results suggest an adaptation of strains on cultivars of the same geographic origin. Thus, based on molecular and virulence markers, C. lindemuthianum strains isolated from wild common bean populations were divided into three groups corresponding to host gene pools.
Effect of Carotenoids on Aflatoxin B1Synthesis byAspergillus flavus
Authors: Norton Robert A.
Published: May 2007

Carotenes and xanthophylls occurring in yellow corn and related terpenoids were tested for their effect on growth and aflatoxin B1production by Aspergillus flavus NRRL 3357, using the suspended disc culture method. Aflatoxin synthesis was inhibited at concentrations of β-carotene, lutein, and zeaxanthin comparable to those found in the horny endosperm of mature corn. Usually growth was not significantly affected. Inhibition of aflatoxin biosynthesis was greater for compounds with an α-ionone-type ring (α-carotene, lutein, or α-ionone) compared with compounds with a β-ionone ring. The presence of hydroxy groups on the rings tended to decrease inhibition, but did not override the effect of the ring type; lutein was similar to α-carotene and zeaxanthin was similar to β-carotene in inhibition. A mutant accumulating norsolorinic acid (NA), A. parasiticus SRRC 162, incubated with α-carotene produced reduced levels of both NA and aflatoxin, indicating that inhibition occurred before NA. Additional A. flavus strains tested against 50 μg/ml of β-carotene had 89 to 96% inhibition, which was significantly more sensitive than NRRL 3357. A. parasiticus strains were less sensitive and generally had similar or lower inhibition than NRRL 3357. The results indicate that the presence of carotenoids in endosperm may decrease the amount of aflatoxin produced by A. flavus.
Phytoalexin Production in an Apple Cultivar Resistant to Venturia inaequalis
Authors: Hrazdina Geza , Borejsza-Wysocki Wlodzimierz , Lester Cathy
Published: May 2007

Cell suspension cultures of the scab-resistant apple (Malus × domestica) cultivar Liberty were challenged with yeast extract to mimic the effect of biological stress such as fungal invasion. The cells responded to the challenge by production of novel compounds. Suspension cultures of the scab-susceptible cultivar McIntosh, when similarly challenged, showed no detectable response. The major compound produced by scab-resistant cells in response to the challenge has been identified as the 2,4-methoxy-3-hydroxy-9-O-β-D-glucosyloxydibenzofuran by UV, mass spectrometry, 1H-nuclear magnetic resonance (NMR), and 13C-NMR spectroscopy. We suggest the trivial name malusfuran for the compound. Malusfuran production was initiated approximately 24 h after being challenged. Malusfuran inhibited spore germination and growth of Venturia inaequalis at millimolar concentrations, indicating its role as a possible phytoalexin. The aglycone of malusfuran, 2,4-methoxy-3,9-hydroxy-dibenzofuran, showed higher toxicity to V. inaequalis than to the parent malusfuran. In vitro cultures of V. inaequalis produced a β-glucosidase that hydrolyzed ortho- and para-substituted nitrophenyl-β-glucosides, suggesting that the aglycone may act as the actual phytoalexin.
Pathway for Phloem-Dependent Movement of Pepper Mottle Potyvirus in the Stem of Capsicum annuum
Authors: Andrianifahanana Mahefa , Lovins Karrie , Dute Roland , Sikora Edward , Murphy John F.
Published: May 2007

Phloem-dependent movement of pepper mottle potyvirus (PepMoV) through Capsicum annuum occurs in a defined pattern through the stem and into uninoculated leaves. The route of movement of PepMoV through the stem of C. annuum ‘Early Calwonder’ was tracked using immunotissue blot analysis and immunomicroscopy. Virus was shown to move from the inoculated leaf down the stem toward the roots via the external phloem. At some location between the cotyledonary node and the roots, PepMoV entered the internal phloem through which it rapidly spread upward the length of the stem to the young tissues. Translocation of PepMoV through the stem occurred in an asymmetric fashion, i.e., virus remained on the side of the stem to which the inoculated leaf was attached as it translocated the length of the stem. Spread and accumulation of PepMoV into uninoculated leaves appeared to occur in a source-to-sink pattern similar to that described for the flow of photoassimilates and similar to other virus and viroid-host systems.
Comparison of Virulence and Isozyme Phenotypes of Pgt-QCCJ and Great Plains Races of Puccinia graminis f. sp. tritici
Authors: Roelfs A. P. , McCallum B. , McVey D. V. , Groth J. V.
Published: May 2007

Stem rust race Pgt-QCCJ was first found in the Great Plains of the United States in 1989, collected primarily from barley. This race became a major part of the Puccinia graminis f. sp. tritici population, even though it is virulent to only a few hard red winter wheat cultivars in the central Great Plains and to barley in the northern Great Plains. It threatens barley production in the northern Great Plains of the United States and Canada due to virulence to Rpg-1. Six differences in virulence and two in isozyme banding patterns from the most similar stem rust races make it unlikely that QCCJ arose as a mutant. Thus, QCCJ likely arose through sexual or parasexual recombination. Sexual recombination in the Great Plains is unlikely, as it has not been detected in many years. Avirulence to ‘McNair 70l’ is only known from the Pacific Northwest of the United States and adjacent Canada. The rust population in this area is of sexual origin, and the pattern of virulence/avirulence and isozyme banding for QCCJ occurs there. Pgt-QCCJ likely originated in the Pacific Northwest during or before 1989 and was wind-transported into the Great Plains.
Seed Transmission of Cucumber Mosaic Virus in Spinach
Authors: Yang Yanming , Kim Kyung Soo , Anderson Edwin J.
Published: May 2007

Spinach (Spinacia oleracea) seed from a commercial breeding line suspected of harboring cucumber mosaic virus (CMV) was analyzed for seed transmission of the virus. Initial seed grow-out tests and enzymelinked immunosorbent assay studies indicated that CMV was present in this seed lot at a level of nearly 15%. To verify these results and gain insight into the mechanism of seed transmission, four combinations of crosses between healthy and/or infected parent plants were conducted. None of the spinach seedlings derived from crossing healthy male and healthy female plants contained CMV, whereas a portion of seedlings derived from all of the other three crosses, i.e., healthy male and infected female, infected male and healthy female, and infected male and infected female plants, were infected with CMV. The results demonstrate that CMV is seed transmitted in spinach and indicate that both male and female parent plants can serve as infection sources. Ultrastructural studies, including immunogold labeling, revealed the presence of virus particles in the cytoplasm of ovary wall cells, ovule integuments and nucellus, anther, and seed-coat cells, as well as fine fibril-containing vesicles and electron-dense inclusions of amorphous aggregates in the central vacuoles of these cells. In addition, reverse transcription-polymerase chain reaction (RT-PCR) was used to amplify 860-bp cDNA fragments containing the CMV coat protein (CP) gene from the embryo, endosperm, and pollen tissues of CMV-infected plants. Taken together, these studies indicate that CMV occurs in virtually all spinach reproductive tissues. Analysis of several RT-PCR amplified and cloned CP genes and flanking sequences from parent and progeny plants revealed that the spinachinfecting CMV was a member of subgroup II. Furthermore, cDNA sequencing and restriction endonuclease mapping consistently revealed two sequence variants, designated SP103 and SP104, in most plants analyzed. These data suggest that there may have been mixed infections of two distinct, seed-transmitted CMV variants in spinach.
AAL-Toxin-Deficient Mutants of Alternaria alternata Tomato Pathotype by Restriction Enzyme-Mediated Integration
Authors: Akamatsu H. , Itoh Y. , Kodama M. , Otani H. , Kohmoto K.
Published: May 2007

Host-specific toxins are produced by three pathotypes of Alternaria alternata: AM-toxin, which affects apple; AK-toxin, which affects Japanese pear; and AAL-toxin, which affects tomato. Each toxin has a role in pathogenesis. To facilitate molecular genetic analysis of toxin production, isolation of toxin-deficient mutants utilizing ectopic integration of plasmid DNA has been attempted. However, the transformation frequency was low, and integration events in most transformants were complicated. Addition of a restriction enzyme during transformation has been reported to increase transformation frequencies significantly and results in simple plasmid integration events. We have, therefore, optimized this technique, known as restriction enzyme-mediated integration (REMI), for A. alternata pathotypes. Plasmid pAN7-1, conferring resistance to hygromycin B, with no detectable homology to the fungal genome was used as the transforming DNA. Among the three restriction enzymes examined, HindIII was most effective, as it increased transformation frequency two-to 10-fold depending on the pathotype, facilitating generation of several hundred transformants with a 1-day protocol. BamHI and XbaI had no significant effect on transformation frequencies in A. alternata pathotypes. Furthermore, the transforming plasmid tended to integrate as a single copy at single sites in the genome, compared with trials without addition of enzyme. Libraries of plasmid-tagged transformants obtained with and without addition of restriction enzyme were constructed for the tomato pathotype of A. alternata and were screened for toxin production. Three AAL-toxin-deficient mutants were isolated from a library of transformants obtained with addition of enzyme. These mutants did not cause symptoms on susceptible tomato, indicating that the toxin is required for pathogenicity of the fungus. Characterization of the plasmid integration sites and rescue of flanking sequences are in progress.
Variation in Pathogenicity and Genotype Among Single-Zoospore Strains of Aphanomyces euteiches
Authors: Malvick D. K. , Percich J. A.
Published: May 2007

The role of asexual reproduction in the production of pathogenic and genotypic variation in Aphanomyces euteiches was investigated. Variation was studied among three groups of 18 single-zoospore progeny of A. euteiches derived from each of three single-zoospore parental strains. Pathogenicity was assessed by evaluating disease severity (DS) on roots of five pea lines possessing different levels of resistance to Aphanomyces root rot and of a susceptible cultivar of snap bean and alfalfa. None of the single-zoospore progeny incited significantly higher DS levels than their parental strain on any of the seven hosts; however, 3 or 4 of the 18 progeny in each group incited significantly lower DS than their parental strains. The host range of the progeny either decreased or remained the same as compared with parental strains. Genotypic variation was assessed with randomly amplified polymorphic DNA (RAPD) analysis. Polymorphic RAPD markers that distinguished parental and progeny strains were detected within two of the three groups of strains with two of four RAPD primers used. Of 76 total RAPD markers that were detected among all strains in all groups, four (5%) were polymorphic. The polymorphic markers were not associated with the pathogenic variation.
Cyclic AMP Restores Appressorium Formation Inhibited by Polyamines in Magnaporthe grisea
Authors: Choi Woo-Bong , Kang Shin-Ho , Lee Yin-Won , Lee Yong-Hwan
Published: May 2007

Magnaporthe grisea, the causal agent of rice blast, forms a dome-shaped melanized infection structure, an appressorium, to infect its host. Environmental cues that induce appressorium formation in this fungus include the hydrophobicity and hardness of the contact surface and chemicals produced by the host. An elevated concentration of intracellular cyclic AMP (cAMP) has been implicated in appressorium differentiation in M. grisea. Polyamines (putrescine, spermidine, and sper-mine) are involved in cell growth and differentiation in a wide range of organisms. To understand the role of polyamines in appressorium differentiation in M. grisea, intracellular polyamines were quantified, and the effects of polyamines and polyamine biosynthesis inhibitors on conidial germination and appressorium formation were tested. High levels of polyamines were detected in freshly collected spores, but the levels decreased during conidial germination. Spermidine was found to be the major component. Polyamines and polyamine biosynthesis inhibitors did not affect conidial germination, but polyamines specifically impaired appressorium formation. Furthermore, exogenous addition of cAMP restored appressorium formation inhibited by poly-amines. These results suggest that polyamines may reduce intracellular cAMP levels in M. grisea, leading to the inhibition of appressorium formation.
Effects of Initial Epidemic Conditions, Sporulation Rate, and Spore Dispersal Gradient on the Spatio-Temporal Dynamics of Plant Disease Epidemics
Authors: Xu X.-M. , Ridout M. S.
Published: May 2007

A stochastic model that simulates the spread of disease over space and time was developed to study the effects of initial epidemic conditions (number of initial inocula and their spatial pattern), sporulation rate, and spore dispersal gradient on the spatio-temporal dynamics of plant disease epidemics. The spatial spread of disease was simulated using a half-Cauchy distribution with median dispersal distance μ (units of distance). The rate of temporal increase in disease incidence (βI, per day) was influenced jointly by μ and by the sporulation rate λ (spores per lesion per day). The relationship between βIand μ was nonlinear: the increase in βIwith increasing μ was greatest when μ was small (i.e., when the dispersal gradient was steep). The rate of temporal increase in disease severity of diseased plants (βS) was affected mainly by λ: βSincreased directly with increasing λ. Intraclass correlation (κt), the correlation of disease status of plants within quadrats, increased initially with disease incidence, reached a peak, and then declined as disease incidence approached 1.0. This relationship was well described by a power-law model that is consistent with the binary form of the variance power law. The amplitude of the model relating κtto disease incidence was affected mainly by μ: κtdecreased with increasing μ. The shape of the curve was affected mainly by initial conditions, especially the spatial pattern of the initial inocula. Generally, the relationship of spatial autocorrelation (ρt,k), the correlation of disease status of plants at various distances apart, to disease incidence and distance was well described by a four-parameter power-law model. ρt,kincreased with disease incidence to a maximum and then declined at higher values of disease incidence, in agreement with a power-law relationship. The amplitude of ρt,kwas determined mainly by initial conditions and by μ: ρt,kdecreased with increasing μ and was lower for regular patterns of initial inocula. The shape of the ρt,kcurve was affected mainly by initial conditions, especially the spatial pattern of the initial inocula. At any level of disease incidence, autocorrelation declined exponentially with spatial lag; the degree of this decline was determined mainly by μ: it was steeper with decreasing μ.
Examination of the Leaf-Drop Symptom of Virus-Infected Potato Using Anther Culture-Derived Haploids
Authors: Valkonen J. P. T. , Rokka V-M. , Watanabe K. N.
Published: May 2007

Necrotic lesions and vein necrosis characteristic of the hypersensitive response (HR) controlled by the dominant resistance gene Ny develop in potato cv. Pito after infection with potato virus Y ordinary strain (PVYº) at a low temperature (16/18°C night/day). In contrast, at high temperatures (19/24°C night/day), large coalesced lesions develop in the lower infected leaves, which wither and remain hanging from stems forming the leaf-drop symptom; mosaic symptoms with no necrosis also develop in the top leaves. The genetic basis of the leaf-drop symptom and its dependence on temperature were examined using a novel approach involving 58 haploids (2n = 24) derived from ‘Pito’ (2n = 48) through anther culture. These haploids and ‘Pito’ were graft-inoculated with PVYO at 19/24 to 25°C (night/day). Necrotic symptoms were expressed in 28 haploids, of which 18 haploids (phenotype class N) developed top necrosis, vein necrosis, or both and necrotic lesions that are characteristic of HR. Ten haploids showed leaf drop similar to ‘Pito’ (phenotype class LD). Thirty haploids were susceptible and showed only mosaic symptoms (phenotype class S). These data indicated that necrosis was induced by a single dominant gene, Ny, in the simplex condition. However, the three distinct phenotypic classes (N, LD, and S) among the haploids grown under the same environmental conditions showed that another locus (gene) was involved in modifying the HR triggered by Ny. Data suggested that this locus contains a dominant temperature-dependent modifier (Tdm) gene that alters the expression of PVY-induced HR at higher temperatures, resulting in leaf drop.
Identification of Tomato Leaf Factors that Activate Toxin Gene Expression inPseudomonas syringaepv.tomatoDC3000
Authors: Li Xiu-Zhen , Starratt Alvin N. , Cuppels Diane A.
Published: May 2007

Coronatine is a non-host-specific chlorosis-inducing phytotoxin produced by the tomato and crucifer pathogen Pseudomonas syringae pv. tomato DC3000. How the chromosomal gene cluster controlling toxin synthesis in this strain is regulated in planta is unknown. Ice nucleation-active cor:inaZ marker-exchange derivatives of strain DC3000 were used to determine coronatine gene expression in various host and nonhost plants and in a minimal medium supplemented with selected tomato plant constituents. Ice nucleation activity, which was first detected 4 h after inoculation, was highest in cabbage, tomato, and soybean and lowest in melon and cucumber. No correlation existed between bacterial population size and expression level on the various plants. Crude tomato leaf extract and intercellular fluid were strong inducers of toxin synthesis. Based on high-performance liquid chromatography analyses and bioassays, we concluded that the active components of both preparations were malic and citric acids, with minor contributions coming from shikimic and quinic acid. Although several compounds including glucose and inositol activated the toxin genes when tested at high concentrations (3 to 5 mM), shikimic and quinic acids were the only ones with activity at concentrations below 0.1 mM. Neither acid could be used as a sole carbon source by strain DC3000. The signal activity of shikimic acid was enhanced 10-fold by the addition of glucose. None of the plant phenolics that we screened affected coronatine gene expression.
Transgenic Resistance to Cucumber Mosaic Virus in Tomato: Blocking of Long-Distance Movement of the Virus in Lines Harboring a Defective Viral Replicase Gene
Authors: Gal-On Amit , Wolf Dalia , Wang Yongzeng , Faure Jean-Emmanuelle , Pilowsky Meir , Zelcer Aaron
Published: May 2007

Tomato breeding lines were transformed with a defective replicase gene from RNA 2 of cucumber mosaic virus (CMV). A total of 63 transformants from five tomato genotypes were evaluated for resistance to CMV strains. The responses of R1 transgenic offspring fit into three categories: fully susceptible lines (44%), fully resistant lines (8%), and an intermediate-type mixture of susceptible and resistant seedlings in variable proportions (48%). Further characterization of the response of two highly resistant lines was performed by mechanical inoculation, aphid transmission, or grafting experiments. No virus was detected in noninoculated leaves from these lines, although a low level of virus accumulated initially in the inoculated leaf. The homozygous R2 plants and further generations that were evaluated (up to R5) showed resistance to the Fny-CMV strain, two Israeli isolates tentatively classified as subgroup IA, and K-CMV (a representative of subgroup IB). These lines were partially resistant to LS-CMV (a representative of subgroup II) when a high-virus-titer inoculum was used. Expression of the viral transgene was verified in these lines; however, the expected translation product was not detectable. In grafting experiments, we demonstrated that CMV virions were blocked in their ability to move from infected rootstocks of nontransformed tomato or tobacco into the transgenic scions. Interestingly, virions could not move through a transgenic intersection into the upper scion. These results provide an additional indication that replicase-mediated resistance affects long-distance movement.
Relationships Between Verticillium dahliae Inoculum Density and Wilt Incidence, Severity, and Growth of Cauliflower
Authors: Xiao C. L. , Subbarao K. V.
Published: May 2007

Microplot and field experiments were conducted to evaluate the effects of inoculum density on Verticillium wilt and cauliflower growth. Soil containing Verticillium dahliae microsclerotia was mixed with various proportions of fumigated soil to establish different inoculum densities (fumigated soil was used as the noninfested control). Seven inoculum density treatments replicated four times were established, and the treatments were arranged in a randomized complete block design. Soil was collected from each microplot immediately after soil infestation for V. dahliae assay by plating onto sodium polypectate agar (NP-10) selective medium using the Anderson sampler technique. Five-week-old cauliflower was transplanted into two beds within each 1.2- by 1.2-m microplot. At the same time, several extra plants were also transplanted at the edge of each bed for destructive sampling to examine the disease onset (vascular discoloration) after planting. Cauliflower plants were monitored for Verticillium wilt development. Stomatal resistance in two visually healthy upper and two lower, diseased leaves in each microplot was measured three times at weekly intervals after initial wilt symptoms occurred. At maturity, all plants were uprooted, washed free of soil, and wilt incidence and severity, plant height, number of leaves, and dry weights of leaves and roots were determined. The higher the inoculum density, the earlier was disease onset. A density of 4 microsclerotia per g of dry soil caused 16% wilt incidence, but about 10 microsclerotia per g of soil caused 50% wilt incidence. Both wilt incidence and severity increased with increasing inoculum density up to about 20 microsclerotia per g of soil, and additional inoculum did not result in significantly higher disease incidence and severity. A negative exponential model described the disease relationships to inoculum levels under both microplot and field conditions. Stomatal resistance of diseased leaves was significantly higher at higher inoculum densities; in healthy leaves, however, no treatment differences occurred. The height, number of leaves, and dry weights of leaves and roots of plants in the fumigated control were significantly higher than in infested treatments, but the effects of inoculum density treatments were variable between years. Timing of cauliflower infection, crop physiological processes related to hydraulic conductance, and wilt intensity (incidence and severity) were thus affected by the inoculum density. Verticillium wilt management methods used in cauliflower should reduce inoculum density to less than four micro-sclerotia per g of soil to produce crops with the fewest number of infected plants.
Histochemical and Cytochemical Investigations of Phenols in Roots of Banana Infected by the Burrowing Nematode Radopholus similis
Authors: Valette C. , Andary C. , Geiger J. P. , Sarah J. L. , Nicole M.
Published: May 2007

The burrowing nematode Radopholus similis is one of the most damaging pathogens on banana plantations. The role of phenolics in plant defense responses to the nematode was histochemically and ultrastructurally investigated in susceptible and partially resistant cultivars. Histochemical observations of healthy roots revealed that high levels of lignin, flavonoids, dopamine, cafeic esters, and ferulic acids were associated with a very low rate of nematode root penetration in the resistant cultivar. The presence of lignified and suberized layers in endodermal cells contributed to limit invasion of the vascular bundle by the pathogen. After infection, flavonoids were seen to accumulate early in walls of cells close to the nematode-migrating channel in both cultivars and in all tissues of the infected resistant roots including the vascular tissues. The labeling pattern obtained with the gold-complexed laccase and with anti-pectin monoclonal antibodies showed that phenolics were distributed in a loosened pectin-rich material surrounding the nematode. This study provides indications that constitutive phenolics in banana roots are associated with the limitation of host penetration and colonization by R. similis. Accumulation of flavonoids in response to infection was detected in the vascular tissues of susceptible plants and in all root tissues in the partially resistant plants.
Environmental Factors Affecting Production, Release, and Field Populations of Conidia of Alternaria alternata, the Cause of Brown Spot of Citrus
Authors: Timmer L. W. , Solel Z. , Gottwald T. R. , Ibañez A. M. , Zitko S. E.
Published: May 2007

Alternaria brown spot, caused by Alternaria alternata pv. citri, affects many tangerines and their hybrids, causing loss of immature leaves and fruit and reducing the marketability of the remaining fruit. Conidial production of A. alternata was greatest on mature leaves moistened and maintained at near 100% relative humidity (RH) for 24 h, whereas leaves that had been soaked or maintained at moderate RH produced few conidia. Conidial release from filter paper cultures and infected leaves was studied in a computer-controlled environmental chamber. Release of large numbers of conidia was triggered from both substrates by sudden drops in RH or by simulated rainfall events. Vibration induced release of low numbers of conidia, but red/infrared irradiation had no effect. In field studies from 1994 to 1996, air sampling with a 7-day recording volumetric spore trap indicated that conidia were present throughout the year with periodic large peaks. The number of conidia captured was not closely related to rainfall amounts or average wind speed, but was weakly related to the duration of leaf wetness. Likewise, disease severity on trap plants placed in the field weekly during 1995 to 1996 was not closely related to conidial numbers or rainfall amounts, but was weakly related to leaf wetness duration. Sufficient inoculum appears to be available to allow infection to occur throughout the year whenever susceptible host tissue and moisture are available.
Nucleotide Sequence and RT-PCR Detection of a Virus Associated with Grapevine Rupestris Stem-Pitting Disease
Authors: Zhang Yun-Ping , Uyemoto Jerry K. , Golino Deborah A. , Rowhani Adib
Published: May 2007

Grapevine rupestris stem pitting (RSP) is a graft-transmissible disease of unknown etiology. We have characterized a virus associated with this disease. The entire genomic sequence (GenBank accession number AF026278) consisted of 8,725 nucleotides excluding a poly(A) tail. Six open reading frames (ORF) were found. ORF1 potentially encodes a polypeptide with a methyltransferase domain, a papain-like proteinase domain, a helicase domain, and a RNA-dependent RNA polymerase domain; ORF2, ORF3, and ORF4 compose a triple-gene block; ORF5 encodes a coat protein; and ORF6 is located near the 3′ end with unknown function. Sequence analysis indicated that the virus is most similar to apple stem-pitting virus and may be allied with the carla- and potexviruses and grouped with other viruses that infect woody hosts. A specific reverse-transcription polymerase chain reaction (RT-PCR)-based detection method was developed. Among 62 grapevine sources known to be infected with rupestris stem-pitting disease, 60 sources tested positive by RT-PCR. Among 43 healthy vines tested, all were negative. The name grapevine rupestris stem-pitting-associated virus is proposed.
Use of Degenerate Primers for Partial Sequencing and RT-PCR-Based Assays of Grapevine Leafroll-Associated Viruses 4 and 5
Authors: Routh Geoffrey , Zhang Yun-Ping , Saldarelli Pasquale , Rowhani Adib
Published: May 2007

Double-stranded RNA (dsRNA) was purified from grapevines infected with grapevine leafroll-associated viruses 4 (GLRaV-4) and 5 (GLRaV-5), two putative closteroviruses. Reverse-transcriptase polymerase chain reaction (RT-PCR) was performed on this dsRNA using degenerate oligonucleotides designed to amplify an approximately 550- to 650-nucleotide fragment from the heat shock protein 70 homolog (HSP70) of the known closteroviruses. RT-PCR products of the appropriate molecular weight were gel-isolated and cloned into the plasmid vector pGEM-T. Clones of RT-PCR products generated by using these primers on dsRNA isolated from a plant infected with GLRaV-4 were sequenced. This sequence was used to develop an immunocapture RT-PCR (IC-RT-PCR) detection protocol capable of detecting GLRaV-4. Similar clones were made from dsRNA isolated from a plant infected with GLRaV-5. These clones were also sequenced. The two sequences were compared, and RT-PCR primers were developed that were able to amplify cDNA from both. These experiments demonstrate that degenerate primers that amplify closterovirus HSP70 sequences can be used to successfully generate sequences useful for IC-RT-PCR detection of these viruses. These data also suggest that it is feasible to use HSP70 sequences to design PCR primers capable of more general PCR detection of multiple GLRaV serotypes. Lastly, the presence of closterovirus-like HSP70 sequences in these putative closteroviruses implies that they are indeed members of this taxonomic group.
Sibling Species of Cercospora Associated with Gray Leaf Spot of Maize
Authors: Wang Juan , Levy Morris , Dunkle Larry D.
Published: May 2007

Monoconidial isolates of the fungus causing gray leaf spot of maize were obtained from diseased leaves collected throughout the United States and analyzed for genetic variability at 111 amplified fragment length polymorphism (AFLP) loci. Cluster analysis revealed two very distinct groups of Cercospora zeae-maydis isolates. Both groups were found to be relatively uniform internally with an average genetic similarity among isolates of approximately 93 and 94%, respectively. The groups were separated from each other by a genetic distance of approximately 80%, a distance greater than that separating each group from the sorghum pathogen, C. sorghi (67 to 70%). Characteristics and dimensions of conidia and conid-iophores produced on infected plants or nutrient media were unreliable criteria for taxonomic differentiation of isolates composing the two groups of C. zeae-maydis. Nucleotide sequences of 5.8S ribosomal DNA (rDNA) and the internal transcribed spacer (ITS) regions were identical within each group but different between the two groups and different from C. sorghi. Restriction fragment length polymorphisms generated by digestion of the 5.8S rDNA and ITS regions with TaqI readily distinguished each group and C. sorghi. Isolates in one group were generally distributed throughout maize-producing regions of the United States; isolates in the other group were localized in the eastern third of the country. Both types were present in the same fields at some locations. The genetic distance based on AFLP profiles and different ITS nucleotide sequences between the two morphologically indistinguishable groups indicate that they are sibling species. Although it is unlikely that breeding for resistance to gray leaf spot will be confounded by local or regional variation in the pathogen, a vigilant approach is warranted, because two pathogenic species exist with unknown abilities to evolve new pathotypes.
Genetic Variation Among Vegetative Compatibility Groups of Fusarium oxysporum f. sp. cubense Analyzed by DNA Fingerprinting
Authors: Bentley S. , Pegg K. G. , Moore N. Y. , Davis R. D. , Buddenhagen I. W.
Published: May 2007

Genetic variation within a worldwide collection of 208 isolates of Fu-sarium oxysporum f. sp. cubense, representing physiological races 1, 2, 3, and 4 and the 20 reported vegetative compatibility groups (VCGs), was analyzed using modified DNA amplification fingerprinting. Also characterized were 133 isolates that did not belong to any of the reported VCGs of F. oxysporum f. sp. cubense including race 3 isolates from a Heliconia species and isolates from a symptomatic wild banana species growing in the jungle in peninsular Malaysia. The DNA fingerprint patterns were generally VCG specific, irrespective of geographic or host origin. A total of 33 different genotypes were identified within F. oxysporum f. sp. cu-bense; 19 genotypes were distinguished among the isolates that belonged to the 20 reported VCGs, and 14 new genotypes were identified among the isolates that did not belong to any of the existing VCGs. DNA fingerprinting analysis also allowed differentiation of nine clonal lineages within F. oxysporum f. sp. cubense. Five of these lineages each contained numerous closely related VCGs and genotypes, and the remaining four lineages each contained a single genotype. The genetic diversity and geographic distribution of several of these lineages of F. oxysporum f. sp. cubense suggests that they have coevolved with edible bananas and their wild diploid progenitors in Asia. DNA fingerprinting analysis of isolates from the wild pathosystem provides further evidence for the coevolution hypothesis. The genetic isolation and limited geographic distribution of four of the lineages of F. oxysporum f. sp. cubense suggests that the pathogen has also arisen independently, both within and outside of the center of origin of the host.
Molecular Identification and Phylogenetic Grouping of Diaporthe phaseolorum and Phomopsis longicolla Isolates from Soybean
Authors: Zhang A. W. , Riccioni L. , Pedersen W. L. , Kollipara K. P. , Hartman G. L.
Published: May 2007

Diaporthe phaseolorum and Phomopsis longicolla isolates from soybean were examined using traditional mycological characteristics and molecular methods. Cultural characteristics including types of fruiting bodies and conidia were assessed for isolates collected from soybean stems and seeds. Cultures were identified as P. longicolla, D. phaseolorum var. caulivora, D. phaseolorum var. meridionalis, or D. phaseolorum var. sojae. Molecular markers for these groups were developed and analyzed using polymerase chain reaction restriction fragment length polymorphisms (PCR-RFLP) and DNA sequencing in the internal transcribed spacer (ITS) and the 5.8S ribosomal DNA. The ITS4 and ITS5 primers amplified PCR products for all isolates studied. Gel electrophoresis of undigested PCR products and DNA sequencing produced various fragment lengths including 604 bp for P. longicolla, 602 and 603 bp for D. phaseolorum var. caulivora, 603 bp for D. phaseolorum var. meridionalis, and from 597 to 609 bp for D. phaseolorum var. sojae. Digestion of these PCR products with enzymes AluI, HhaI, MseI, RsaI, and ScrFI resulted in distinct bands for identification of P. longicolla and the varieties of D. phaseolorum I. All P. longicolla, D. phaseolorum var. caulivora, and D. phaseolorum var. meridionalis isolates were distinguished using AluI and HhaI with RsaI or ScrFI. The banding patterns of D. phaseolorum var. sojae isolates were complex and were separated into 11 subgroups after digestion with AluI, HhaI, MseI, RsaI, and ScrFI. Phylogenetic analysis of 20 isolates of D. phaseolorum and P. longicolla based on the DNA sequence of the ITS region resolved six clades termed A, B, C, D, E, and F. Clade A included all sequenced D. phaseolorum var. caulivora isolates, two from Italy and one from the United States. Isolates in clade B were exclusively associated with D. phaseolorum var. meridionalis. Clades A and B formed a well-supported monophyletic group. Isolates in clades C, D, E, and F were morphologically defined as isolates of P. longicolla, D. phaseolorum var. sojae, and Diaporthe spp. The ITS sequences similarity of seven geographically diverse P. longi-colla isolates illustrated that P. longicolla isolates have a similar genetic background, with some affiliations to some D. phaseolorum var. sojae isolates. Morphological characteristics of the isolates along with the terminal clades of the ITS phylogeny suggest that P. longicolla is an individual species, D. phaseolorum var. caulivora and D. phaseolorum var. meridionalis are varieties of D. phaseolorum, and D. phaseolorum var. sojae is either several varieties of D. phaseolorum or possibly several distinct species.
The Phytopathogenic Mollicute-Insect Vector Interface: A Closer Look
Authors: Fletcher Jacqueline , Wayadande Astri , Melcher Ulrich , Ye Fengchun
Published: May 2007

Spiroplasma citri, transmitted by phloem-feeding leafhoppers, moves from the gut lumen through the gut wall, hemolymph, and salivary glands and multiplies in insect tissues. Nontransmissible lines were deficient in their ability to cross these barriers. Molecular analysis revealed extensive chromosomal rearrangements between the transmissible and nontransmissible spiroplasma lines including a large chromosomal inversion and deletions of about 10 kb at each inversion border. One open reading frame of the deleted region, cloned from the transmissible strain BR3-3X, encodes an integral membrane protein of 58 kDa that shares limited sequence similarity with major adhesin proteins of two zoopathogenic mycoplasmas. Adhesion of spiroplasmas to cultured leafhopper cells was inhibited by proteases, suggesting that adherence to host cells is mediated by spiroplasma membrane protein(s). A hypothetical model for insect transmission of phytopathogenic mollicutes is presented.