bluuebunny/crossref_metadata_2025 huggingface dataset:
trans-Dominant Inhibition of Geminiviral DNA Replication by Bean Golden Mosaic Geminivirus rep Gene Mutants
Authors: Hanson Stephen F. , Maxwell Douglas P.
Published: May 2007

Geminiviruses are a group of single-stranded DNA viruses that cause major losses on a number of important crops throughout the world. Bean golden mosaic virus (BGMV) is a typical bipartite, whitefly-transmitted geminivirus that causes a severe disease on beans (Phaseolus vulgaris) in the Western Hemisphere. The lack of natural resistance to geminiviruses has led to attempts to engineer resistance, particularly through the use of pathogen-derived resistance strategies. The rep gene contains several conserved domains including nucleoside triphosphate (NTP)-binding and DNA-nicking domains and is the only geminiviral gene necessary for replication. Previous analysis by our group and others has demonstrated that the NTP-binding and DNA-nicking domains are necessary for geminiviral DNA replication. The ability of the rep gene and rep gene mutants to interfere with geminiviral DNA replication, when expressed in trans, was examined using a transient assay in a tobacco suspension cell culture system. Wild-type (wt) and mutant rep genes were cloned into plasmids under the control of the cauliflower mosaic virus 35S promoter for in planta expression and coinoculated into tobacco cells with infectious clones of various geminiviruses. The wt rep gene from BGMV-GA was able to support replication of BGMV-GA DNA-B. Several different rep gene mutants, with function-abolishing mutations in the NTP-binding or DNA-nicking domains, were potent trans-dominant inhibitors of geminiviral DNA replication.
Leaf Rust Resistance Gene Lr34 Associated with Nonsuppression of Stem Rust Resistance in the Wheat Cultivar Canthatch
Authors: Kerber E. R. , Aung T.
Published: May 2007

The common wheat cultivar Thatcher and the backcross derivative Canthatch are moderately or fully susceptible to several races of stem rust because of a suppressor on chromosome 7DL that inhibits the expression of the relevant resistance gene(s). The incorporation of leaf rust resistance gene Lr34 into ‘Thatcher’ is known to enhance stem rust resistance. The effect of this gene in a ‘Canthatch’ background and its relationship with the 7DL suppressor were determined by replacing chromosome 7D of ‘Canthatch’ with 7D of ‘Chinese Spring’, which possesses Lr34 on the short arm. ‘Canthatch’ nullisomic 7D was crossed with ‘Chinese Spring’, followed by a succession of backcrosses to the nullisomic recurrent parent. Homozygous resistant disomic and monosomic substitution lines were recovered that exhibited the same resistant reaction as that of ‘Thatcher’ possessing Lr34 and as that of ‘Canthatch’ nullisomic 7D, in which the 7DL suppressor is absent. The results indicate that, in ‘Canthatch’, Lr34 permits expression of resistance genes normally inhibited by the 7DL suppressor. Furthermore, it is likely that, in ‘Thatcher’ and ‘Thatcher’ back-cross derivatives, Lr34 inactivates the 7DL suppressor.
Effect of Temperature on and Histopathology of the Interaction Between Meloidogyne incognita and Thielaviopsis basicola on Cotton
Authors: Walker N. R. , Kirkpatrick T. L. , Rothrock C. S.
Published: May 2007

Controlled environments were used to study the relationship between the root-knot nematode (Meloidogyne incognita) and Thielaviopsis basicola on cotton. Temperature treatments were continuous 20, 24, and 28°C or two cyclic linear regimes with ranges of 14 to 32 or 18 to 28°C over 24 h. Cotton seeds were planted in fumigated soil infested with T. basicola, M. incognita, or both. After 42 days, pathogen effects on plant growth and pathogen development were evaluated. Histology was conducted on roots collected 14, 28, and 42 days after planting in the continuous 24°C treatment. Reductions in plant height-to-node ratio and total fresh weight were observed for soils infested with both pathogens compared with the control or with soils infested with either pathogen, except for M. incognita-infested soil at 28°C. T. basicola reduced root galling and reproduction of the nematode at all temperatures. Vascular discoloration caused by T. basicola was greater in the presence of M. incognita compared with that by T. basicola alone. At 2 and 4 weeks, histological studies showed that plants grown in all T. basicola-infested soils contained chlamydospore chains on the root surface and in cortical cells. The fungus was not observed inside the vascular cylinder. Roots from 4-week-old plants from soils infested with T. basicola and M. incognita showed fungal sporulation in vascular tissue and localized necrosis of vascular tissue adjacent to the nematodes. At 6 weeks, plants grown in soil infested with T. basicola alone exhibited no remaining cortical tissue and no evidence of vascular colonization by the fungus. Six-week-old plants grown in T. basicola + M. incognita-infested soils exhibited extensive vascular necrosis and sporulation within vascular tissue. These studies suggest that coinfection expands the temperature ranges at which the pathogens are able to cause plant damage. Further, M. incognita greatly increases the access of T. basicola to vascular tissue.
Moisture Prediction from Simple Micrometeorological Data
Authors: Chtioui Y. , Francl L. J. , Panigrahi S.
Published: May 2007

Four linear regression methods and a generalized regression neural network (GRNN) were evaluated for estimation of moisture occurrence and duration at the flag leaf level of wheat. Moisture on a flat-plate resistance sensor was predicted by time, temperature, relative humidity, wind speed, solar radiation, and precipitation provided by an automated weather station. Dew onset was estimated by a classification regression tree model. The models were developed using micrometeorological data measured from 1993 to 1995 and tested on data from 1996 and 1997. The GRNN outperformed the linear regression methods in predicting moisture occurrence with and without dew estimation as well as in predicting duration of moisture periods. Average absolute error for prediction of moisture occurrence by GRNN was at least 31% smaller than that obtained by the linear regression methods. Moreover, the GRNN correctly predicted 92.7% of the moisture duration periods critical to disease development in the test data, while the best linear method correctly predicted only 86.6% for the same data. Temporal error distribution in prediction of moisture periods was more highly concentrated around the correct value for the GRNN than linear regression methods. Neural network technology is a promising tool for reasonably precise and accurate moisture monitoring in plant disease management.
Evaluation of Maize Streak Virus Pathogenicity in Differentially Resistant Zea mays Genotypes
Authors: Martin D. P. , Willment J. A. , Rybicki E. P.
Published: May 2007

We devised a rapid technique for the objective and precise assessment of both the pathogenicity of maize streak virus (MSV) isolates and the MSV resistance of maize genotypes. The technique involves the use of agroinoculation to infect maize seedlings and the objective symptom evaluation by quantification of infection rates, stunting, and chlorotic leaf areas. In assessing the MSV resistance of 19 maize genotypes, we describe how the use of differentially virulent virus isolates enables the analysis of MSV resistance phenotypes, ranging from extremely susceptible to completely immune. We further demonstrate how quantification of chlorotic leaf areas by image analysis permits differentiation between degrees of MSV resistance that are indistinguishable from one another using currently employed symptom assessment approaches. Using chlorotic area measurements, we quantify the virulence of a diverse group of 10 MSV isolates and, through agroinoculation of differentially susceptible maize genotypes, we demonstrate the use of our technique in evaluating the pathogenicity of these isolates.
Increase of Tospoviral Diversity in Brazil with the Identification of Two New Tospovirus Species, One from Chrysanthemum and One from Zucchini
Authors: Bezerra I. C. , de O. Resende R. , Pozzer L. , Nagata T. , Kormelink R. , De Ávila A. C.
Published: May 2007

During a survey conducted in several different regions of Brazil, two unique tospoviruses were isolated and characterized, one from chrysanthemum and the other from zucchini. The chrysanthemum virus displayed a broad host range, whereas the virus from zucchini was restricted mainly to the family Cucurbitaceae. Double-antibody sandwich-enzyme-linked immunosorbent assay and western immunoblot analyses demonstrated that both viruses were serologically distinct from all reported tospovirus species including the recently proposed peanut yellow spot virus and iris yellow spot virus (IYSV) species. The nucleotide sequences of the nucleocapsid (N) genes of both viruses contain 780 nucleotides encoding for deduced proteins of 260 amino acids. The N proteins of these two viruses displayed amino acid sequence similarities with the previously described tospovirus species ranging from 20 to 75%, but they were more closely related to each other (80%). Based on the biological and molecular features, these viruses are proposed as two new tospovirus species, designated chrysanthemum stem necrosis virus (CSNV) and zucchini lethal chlorosis virus (ZLCV). With the identification of CSNV and ZLCV, in addition to tomato spotted wilt virus, groundnut ring spot virus, tomato chlorotic spot virus, and IYSV, Brazil harbors the broadest spectrum of tospovirus species reported.
Histopathological Study of Barley Cultivars Resistant and Susceptible to Rhynchosporium secalis
Authors: Carisse K. Xi , Burnett P. A. , Tewari J. P. , Chen M. H. , Turkington T. K. , Helm J. H.
Published: May 2007

Differences in the penetration process by Rhynchosporium secalis were compared in resistant and susceptible barley cultivars at the seedling stage. Percent penetration and percent host cell wall alteration (HCWA) differed significantly among cultivars and isolates as revealed by light microscopy. Based on these two variables, the cultivars were statistically separated into two groups that corresponded to their disease reactions. The resistant cultivars, Johnston and CDC Guardian, showed 81.2 to 99.4% HCWA and 0.1 to 20.1% penetration at encounter sites, whereas the susceptible cultivars, Harrington, Argyle, and Manley, had 30.1 to 78.3% HCWA and 31.8 to 81.8% penetration. In the current study, cv. Leduc, which is susceptible at the seedling stage and resistant at the adult stage, showed the same percent HCWA and penetration as did susceptible cultivars. A significant negative correlation (P < 0.01) was found between percent penetration and percent HCWA for cultivars inoculated with two isolates of the pathogen. Isolate 1 was less virulent than isolate 2 with respect to percent penetration and induced significantly fewer HCWA. Scanning electron microscopy showed various shapes of fungal appressoria but no apparent difference in host reaction between resistant and susceptible cultivars. Transmission electron microscopy revealed interactions between the host and pathogen at various stages of penetration. The resistant cv. Johnston responded by producing appositions, as evidenced by a layer of compact osmiophilic material deposited on the inner side of the cell wall. Infection pegs produced by conidia were unable to penetrate the cuticle where an apposition had formed inside. When penetration occurred in the susceptible cv. Argyle, cytoplasmic aggregates and separation of the plasmalemma were visible from the host cell wall, but the layer of compact osmiophilic material was not always present. Data based on light microscopic observations suggested that HCWA may be one of the mechanisms responsible for resistance that is characterized as penetration prevention rather than as a slow rate of mycelial growth after successful penetration. HCWA occurred in response to attempted cuticle penetration, suggesting that HCWA may produce chemical barriers that help to prevent penetration.
Thermal Inactivation of Phytophthora nicotianae
Authors: Coelho L. , Mitchell D. J. , Chellemi D. O.
Published: May 2007

Phytophthora nicotianae was added to pasteurized soil at the rate of 500 laboratory-produced chlamydospores per gram of soil and exposed to temperatures ranging from 35 to 53°C for 20 days. The time required to reduce soil populations to residual levels (0.2 propagule per gram of soil or less) decreased with increasing temperatures. Addition of cabbage residue to the soil reduced the time required to inactivate chlamydospores. Temperature regimes were established to simulate daily temperature changes observed in the field, with a high temperature of 47°C for 3 h/day, and were good estimators of the efficacy of soil solarization for the control of P. nicotianae in soil. Cabbage amendment reduced the time required to inactivate chlamydospores of P. nicotianae and its effect was more pronounced at lower temperature regimes.
Enhanced Polymerase Chain Reaction Methods for Detecting and Quantifying Phytophthora infestans in Plants
Authors: Judelson Howard S. , Tooley Paul W.
Published: May 2007

Sensitive and specific primer sets for polymerase chain reaction (PCR) for Phytophthora infestans, the oomycete that causes late blight of potato and tomato, were developed based on families of highly repeated DNA. The performance of these primers was compared to those developed previously for the internal transcribed spacer (ITS) of ribosomal DNA. The detection limit using the new primers is 10 fg of P. infestans DNA, or 0.02 nuclei. This is about 100 times more sensitive than ITS-directed primers. Nested polymerase chain reaction (PCR) allows the measurement of down to 0.1 fg of DNA using the new primers. To enhance the reliability of diagnostic assays, an internal positive control was developed using an amplification mimic. The mimic also served as a competitor for quantitative PCR, which was used to assess the growth of P. infestans in resistant and susceptible tomato. A key dimension of this study was that two laboratories independently checked the specificity and sensitivity of each primer set; differences were noted that should be considered when PCR is adopted for diagnostic applications in any system.
Genes Governing Resistance to Puccinia hordei in Thirteen Spring Barley Accessions
Authors: Brooks W. S. , Griffey C. A. , Steffenson B. J. , Vivar H. E.
Published: May 2007

Leaf rust, caused by Puccinia hordei, is an important disease of barley in many parts of the world. In the eastern United States, this disease was effectively controlled for over 20 years through the deployment of cultivars carrying the resistance gene Rph7. Isolates of P. hordei with virulence for Rph7 appeared in this region in the early 1990s rendering barley cultivars with this gene vulnerable to leaf rust infection. From a preliminary evaluation test, 13 accessions from diverse geographic locations possessed resistance to P. hordei isolate VA90-34, which has virulence for genes Rph1, 2, 4, 6, 7, 8, and 11. Each of these 13 accessions was crossed with susceptible cvs. Moore or Larker to characterize gene number and gene action for resistance to P. hordei. Additionally, the 13 accessions were intercrossed and crossed to host differential lines possessing genes Rph3, Rph5, and Rph9 to determine allelic relationships of resistance genes. Seedlings of F1, F2, and BC1F1 populations were evaluated in the greenhouse for their reaction to P. hordei isolate VA90-34. Leaf rust resistance in six of the accessions including Collo sib, CR270.3.2, Deir Alla 105, Giza 119, Gloria, and Lenka is governed by a single dominant gene located at or near the Rph3 locus. All accessions for which the gene Rph3 was postulated to govern leaf rust resistance, except for Deir Alla 105, likely possess an allele different than Rph3.c found in Estate based on the differential reaction to isolates of P. hordei. The resistance gene in Grit and Donan is located at or near the Rph9 locus. Alleles at both the Rph3 and Rph9 loci confer resistance in Femina and Dorina. In addition to Rph3, Caroline and CR366.13.2 likely possess a second unknown recessive gene for leaf rust resistance. Resistance in Carre 180 is governed by a recessive gene that is different from all other genes considered in this study. Identification of both known and unique genes conferring leaf rust resistance in the barley germplasm included in this study provides breeding programs with the knowledge and opportunity to assess currently used sources of leaf rust resistance and to incorporate new sources of resistance into their programs.
Range of Phytoplasma Concentrations in Various Plant Hosts as Determined by Competitive Polymerase Chain Reaction
Authors: Berges R. , Rott M. , Seemüller E.
Published: May 2007

For competitive polymerase chain reaction (PCR), an internal standard DNA template was developed that consisted of a highly conserved, internally deleted 16S rDNA fragment of an aster yellows phytoplasma. The internal standard was calibrated using a quantified culture of Acholeplasma laidlawii. Serial dilutions of the internal standard and fixed amounts of target templates from infected plants were coamplified with the same primers, and the products obtained were quantified using an enzyme-linked immunosorbent assay procedure. Analysis of the data revealed that the phytoplasma concentration in the plants examined differed by a factor of about 4 × 106. Phytoplasma concentrations of 2.2 × 108 to 1.5 × 109 cells per g of tissue were identified in periwinkles infected with various phytoplasmas. High to moderate concentrations were detected in Malus domestica (apple) genotypes infected with the apple proliferation phytoplasma, Alnus glutinosa (alder) genotypes infected with the alder yellows phytoplasma, and most aster yellows-infected Populus (poplar) genotypes examined. Very low phytoplasma concentrations, ranging from 370 to 34,000 cells per g of tissue, were identified in proliferation-diseased apple trees on resistant rootstocks 4551 and 4608, yellows-diseased Quercus robur (oak) trees, and Carpinus betulus (hornbeam) trees. Such low concentrations, which corresponded to about 4 to 340 cells in the reaction mixture, could only be detected and quantified by nested PCR.
Aphid Acquisition and Cellular Transport of Potato leafroll virus-like Particles Lacking P5 Readthrough Protein
Authors: Gildow F. E. , Reavy B. , Mayo M. A. , Duncan G. H. , Woodford J. A. T. , Lamb J. W. , Hay R. T.
Published: May 2007

Lepidopteran cells (Spodoptera frugiperda) produced isometric virus-like particles (VLP) when infected with a recombinant baculovirus Ac61 that contained the Potato leafroll virus (PLRV) coat protein gene modified with an N-terminal histidine tag (P3-6H). Cells infected with AcFL, a recombinant baculovirus that expressed cDNA copies of the PLRV genome RNA, did not produce virus-like particles (VLP). In cell lines doubly infected with Ac61 and AcFL, VLP were formed that contained PLRV-RNA packaged in P3-6H coat protein (FL). Both the P3-6H and the FL particles were morphologically indistinguishable from particles of PLRV despite the fact that they lacked the P5 readthrough protein present in wild-type PLRV. When aphids (Myzus persicae) were fed on, or injected with, purified PLRV, or VLP of either type (FL or P3-6H) and examined by electron microscopy, no differences were observed among treatments for particle endocytosis, transcellular transport, or exocytosis at the aphid midgut or accessory salivary glands. Particles were observed in the salivary canals and in the salivary duct leading out of the aphid. These results suggest that P5 readthrough protein of PLRV may not be essential for cellular transport of virus through aphid vectors.
Evaluation of Seed Transmission of Turnip yellow mosaic virus and Tobacco mosaic virus in Arabidopsis thaliana
Authors: de Assis Filho F. M. , Sherwood J. L.
Published: May 2007

The mechanism of virus transmission through seed was studied in Arabidopsis thaliana infected with Turnip yellow mosaic virus (TYMV) and Tobacco mosaic virus (TMV). Serological and biological tests were conducted to identify the route by which the viruses reach the seed and subsequently are located in the seed. Both TYMV and TMV were detected in seed from infected plants, however only TYMV was seed-transmitted. This is the first report of transmission of TYMV in seed of A. thaliana. Estimating virus seed transmission by grow-out tests was more accurate than enzyme-linked immunosorbent assay due to the higher frequency of antigen in the seed coat than in the embryo. Virus in the seed coat did not lead to seedling infection. Thus, embryo invasion is necessary for seed transmission of TYMV in A. thaliana. Crosses between healthy and virus-infected plants indicated that TYMV from either the female or the male parent could invade the seed. Conversely, invasion from maternal tissue was the only route for TMV to invade the seed. Pollination of flowers on healthy A. thaliana with pollen from TYMV-infected plants did not result in systemic infection of healthy plants, despite TYMV being carried by pollen to the seed.
Characterization of a Viral EPS-Depolymerase, a Potential Tool for Control of Fire Blight
Authors: Kim Won-Sik , Geider Klaus
Published: May 2007

A 3.3-kb fragment of genomic DNA from bacteriophage Φ-Ea1h encoding an amylovoran-directed depolymerase lyase was sequenced, and three open reading frames (ORFs) were detected. The first ORF could encode a lysozyme and the second a holin that may form a pore supporting cell lysis by the lysozyme. The third ORF encodes a protein of 657 amino acids and deletion mutation in this DNA fragment abolished extracellular polysaccharide (EPS)-degrading activity. A putative promoter and a ribosome binding sequence were located in front of the gene. A polymerase chain reaction product spanning the gene was inserted into multi copy plasmids including fusions with a Histidine-tagged sequence to facilitate its purification on a nickel nitrilotriacetic acid column. Maximal activity of the purified protein was observed between pH 4 and 5 at 52°C. Visualized by staining with fluorescein isothiocyanate-labeled lectin from Abrus precatorious, the enzyme degraded the EPS-capsules of Erwina amylovora. In virulence assays, no symptoms were detected for a low inoculum of an E. amylovora strain when pear slices were soaked in a solution of depolymerase in contrast to control slices without addition of the enzyme. Furthermore, gfp- or lux-labeled E. amylovora cells were not propagated, when their amylovoran capsules were removed by the depolymerase. The enzyme could be a tool for biological control of fire blight.
Simultaneous Detection of the Three Ilarviruses Affecting Stone Fruit Trees by Nonisotopic Molecular Hybridization and Multiplex Reverse-Transcription Polymerase Chain Reaction
Authors: Saade M. , Aparicio F. , Sánchez-Navarro J. A. , Herranz M. C. , Myrta A. , Di Terlizzi B. , Pallás V.
Published: May 2007

The three most economically damaging ilarviruses affecting stone fruit trees on a worldwide scale are the related Prunus necrotic ringspot virus (PNRSV), Prune dwarf virus (PDV), and Apple mosaic virus (ApMV). Nonisotopic molecular hybridization and multiplex reverse-transcription polymerase chain reaction (RT-PCR) methodologies were developed that could detect all these viruses simultaneously. The latter technique was advantageous because it was discriminatory. For RT-PCR, a degenerate antisense primer was designed which was used in conjunction with three virus-specific sense primers. The amplification efficiencies for the detection of the three viruses in the multiplex RT-PCR reaction were identical to those obtained in the single RT-PCR reactions for individual viruses. This cocktail of primers was able to amplify sequences from all of the PNRSV, ApMV, and PDV isolates tested in five Prunus spp. hosts (almond, apricot, cherry, peach, and plum) occurring naturally in single or multiple infections. For ApMV isolates, differences in the electrophoretic mobilities of the PCR products were observed. The nucleotide sequence of the amplified products of two representative ApMV isolates was determined, and comparative analysis revealed the existence of a 28-nucleotide deletion in the sequence of isolates showing the faster electrophoretic mobility. To our knowledge, this is the first report on the simultaneous detection of three plant viruses by multiplex RT-PCR in woody hosts. This multiplex RT-PCR could be a useful time and cost saving method for indexing these three ilarviruses, which damage stone fruit tree yields, and for the analysis of mother plants in certification programs.
The Role of Water in Epiphytic Colonization and Infection of Pomaceous Flowers by Erwinia amylovora
Authors: Pusey P. L.
Published: May 2007

Detached crab apple flowers were used as an experimental model to investigate the effect of relative humidity (RH), free moisture, and water potential Ψw on the interaction between Erwinia amylovora and pomaceous flowers. Flowers were maintained at 24°C with the cut pedicel submerged in a sucrose solution. The bacterium multiplied on inoculated flower stigmas at between approximately 55 and 100% RH but not in the floral cup (hypanthium) until the RH was higher than 80%. To study the effect of free moisture, stigma-inoculated flowers were kept wet for different periods. Flowers became diseased only with wetting, and incidence was high (77%) even when water application was immediately followed by a 52-min drying period. In other experiments with hypanthium-inoculated flowers, RH or sucrose concentration in holding vials was varied to affect Ψw of flower nectar and ovary tissue. Population size of E. amylovora in the hypanthium increased with nectar Ψw following a sigmoidal curve (R2 = 0.99). Disease incidence and severity, however, were more closely related to ovary Ψw (R2 = 0.85 and 0.91, respectively) than to bacterial population size (R2 = 0.25 and 0.67, respectively) as fitted to the quadratic equation. Maximum disease incidence and severity occurred at an ovary Ψw above -2.0 MPa, and disease severity continued to increase above -1.0 MPa. These results were confirmed with detached flowers of Delicious apple and d'Anjou pear. A practical implication is that disease might be partly managed in arid climates by limiting soil irrigation water during bloom and early fruit set.
Analysis of the Association Between the Incidence of Two Spatially Aggregated Foliar Diseases of Strawberry
Authors: Turechek W. W. , Madden L. V.
Published: May 2007

Association of the incidence of leaf blight (caused by Phomopsis obscurans) and leaf spot of strawberry (caused by Mycosphaerella fragariae) was assessed at multiple scales in perennial plantings at several commercial farms over 3 years (1996 to 1998). For each field, the presence or absence of each disease was recorded from n = 15 leaflets in each of N ≈ 70 evenly spaced sampling units, and the proportion of leaflets with blight, spot, and total disease (blight or spot) was determined. Individual diseases and total disease incidence were all well described by the beta-binomial distribution but not by the binomial distribution, indicating overdispersion of disease. The Jaccard similarity index was used to measure disease co-occurrence at the leaflet, sampling-unit, and field scales. Standard errors of this index for the lower two scales were obtained using the jackknife (resampling) procedure, and data randomizations were used to determine the expected Jaccard index for an independent arrangement of the two diseases, conditioned on the incidence and spatial heterogeneity of the observed disease data. Results based on these statistics showed that only 4 of 52 data sets at the leaflet level and no data sets at the sampling-unit level had Jaccard index values significantly different from that expected under an independent rearrangement of the two diseases. Rank correlation and cross-correlation statistics were calculated to determine the degree of covariation in incidence between the two diseases. Additionally, covariation between diseases was tested using a new procedure in the Spatial Analysis by Distance IndicEs (SADIE) class of tests. Covariation was detected in 21% of the data sets using rank correlation methods and in 15% of the data sets using the SADIE-based approach. The discrepancy between these two methods may be due to the rank correlation procedure not taking into account the effects of spatial pattern of disease incidence. There was no relationship between mean disease incidence per field of spot and blight or between degree of heterogeneity of the two diseases (as measured by θ of the beta-binomial distribution), demonstrating lack of covariation at the field scale. Incidence of leaflets with either disease (total disease incidence) could be well predicted using a linear combination of the estimated probabilities of leaf blight and leaf spot incidence based on independence of the two diseases. Heterogeneity of total disease incidence, measured with the estimated θ parameter of the beta-binomial distribution, could also be well predicted using a linear combination of the weighted θ values for leaf blight and leaf spot, with weights proportional to incidence of the individual diseases.
Genetics of Host-Pathogen Relationships Between Venturia inaequalis Races 6 and 7 and Malus Species
Authors: Bénaouf G. , Parisi L.
Published: May 2007

Resistance to scab originating from Malus floribunda clone 821 is the most widely form of resistance used in apple breeding programs. A dominant gene, named Vf, was introgressed from this clone into recent cultivars, although the genetic determinants of the resistance of M. floribunda 821 are apparently more complex than a single gene. The appearance of new races overcoming the resistance of cultivars with the Vf gene, the parental clone, or both made it possible to undertake a genetic analysis of host-pathogen interactions. The segregation of resistance in progenies of crosses from ‘Golden Delicious’ × M. floribunda 821 and ‘Golden Delicious’ × ‘Idared’ into five strains of Venturia inaequalis—races 1 (strains 104, 1093, and 301), 6 (strain 302), and 7 (strain 1066)—demonstrated the existence of a second dominant gene in M. floribunda 821. This gene, independent of Vf, was named Vfh because it seemed to induce a hypersensitive reaction. The results obtained with strain 1066, virulent to M. floribunda 821, allowed identification of another dominant gene, Vg, responsible for the resistance of ‘Golden Delicious’ to this strain. Vg is also carried by ‘Florina’, which was selected for its Vf resistance. The pathogenicity of a progeny originating from a cross between V. inaequalis strains 1066 and 301, characterized in vitro on leaf disks of differential genotypes, revealed two independent avirulence genes involved in the pathogenicity toward the Vg and Vf genes, respectively. These avirulence genes were named Avr Vg and Avr Vf. The host-pathogen interactions detected are consistent with a gene-for-gene relationship.
Induced Resistance by Penicillium oxalicum Against Fusarium oxysporum f. sp. lycopersici: Histological Studies of Infected and Induced Tomato Stems
Authors: De Cal A. , Garcia-Lepe R. , Melgarejo P.
Published: May 2007

Tomato (Lycopersicon esculentum) plants of ‘Lorena’ were induced with a conidial suspension (107 conidia per ml) of Penicillium oxalicum before inoculation with Fusarium oxysporum f. sp. lycopersici, the wilt pathogen. Histological changes occurred in plants under both growth chamber and glasshouse culture conditions and there was a reduction of disease severity. In noninduced plants, the pathogen produced almost a complete loss of cambium (75 to 100% reduction), an increase in the number of bundles, and a decrease in the number of xylem vessels (20% reduction), in which the diameter also was reduced by 20 to 30% in hypocotyls and epicotyls. The percentage of vessels colonized by F. oxysporum f. sp. lycopersici was positively correlated to the area under the disease progress curve (AUDPC). However, plants induced with P. oxalicum showed less disease, did not lose the cambium, had a lower number of bundles, and had less vascular colonization by F. oxysporum f. sp. lycopersici (35 to 99%). These effects also were observed in ‘Precodor’, which is susceptible to races 1 and 2 of F. oxysporum f. sp. lycopersici, and partially in ‘Ramón’, which is resistant to both races. Renewed or prolonged cambial activity that led to the formation of additional secondary xylem could be one of the reasons for disease reduction in P. oxalicum-induced tomato plants.
Transgenic Nicotiana occidentalis Plants Expressing the 50-kDa Protein of Apple chlorotic leaf spot virus Display Increased Susceptibility to Homologous Virus, but Strong Resistance to Grapevine berry inner necrosis virus
Authors: Yoshikawa N. , Gotoh S. , Umezawa M. , Satoh N. , Satoh H. , Takahashi T. , Ito T. , Yoshida K.
Published: May 2007

The 50-kDa protein (P50) encoded by the open reading frame 2 of Apple chlorotic leaf spot virus (ACLSV), a putative movement protein, was expressed in transgenic Nicotiana occidentalis plants. P50 in transgenic plants was mainly detected in a modified form in the cell wall fraction, similar to that in infected leaves. The P50-expressing plants (P50 plants) complemented the systemic spread of the P50-defective mutants of an infectious cDNA clone of ACLSV (pCLSF), indicating that P50 in transgenic plants was functional. Severity of symptoms was greatly enhanced and accumulation of virus in upper leaves was increased in P50 plants inoculated with pCLSF or ACLSV compared with that in nontransgenic control plants (NT plants). Conversely, transgenic plants expressing the coat protein of ACLSV (CP plants) showed a significant delay in symptom development and a reduction of virus accumulation. However, most P50 plants inoculated with Grapevine berry inner necrosis virus (GINV), another species of the genus Trichovirus, neither developed obvious symptoms nor supported virus accumulation in inoculated or upper leaves. In contrast, systemic symptoms developed and virus accumulated equally in NT and CP plants inoculated with GINV. After inoculation with Apple stem grooving virus or Apple stem pitting virus, there was no difference in symptom development and virus accumulation among P50, CP, and NT plants. Our results indicate that transgenic plants expressing a functional P50 were more susceptible to homologous virus and, on the contrary, showed strong resistance to the heterologous virus GINV.
Impeded Thrips Transmission of Defective Tomato spotted wilt virus Isolates
Authors: Nagata Tatsuya , Inoue-Nagata Alice K. , Prins Marcel , Goldbach Rob , Peters Dick
Published: May 2007

Two defective RNA-containing isolates (Pe-1 and 16-2) and an envelope-deficient (env¯) isolate of Tomato spotted wilt virus (TSWV) were tested for their transmissibility by Frankliniella occidentalis. The Pe-1 isolate contained a truncated L RNA segment that barely interfered with symptom expression and replication of the wild-type (wt) L RNA segment. This isolate was transmitted with an efficiency of 51%, a value comparable to that found for wt TSWV (54%). Isolate 16-2, which contained a genuine defective interfering L RNA as concluded from its ability to suppress wt L RNA synthesis and attenuation of symptom expression, was not transmitted at all. The midguts of all larvae that ingested Pe-1 became infected, whereas limited midgut infections were found in 24% of the larvae that ingested 16-2. This difference in infection could be explained by the presence of a low number of infectious units in the inoculum ingested from plants as demonstrated in infection experiments and verified by northern blot analysis. The env¯ isolate failed to infect the midgut after ingestion and could not be transmitted by any thrips stage. This isolate also cannot infect primary thrips cell cultures. Taken together, these results suggest that the envelope of TSWV contains the determinants required for binding and subsequent infection of thrips cells.
Contamination, Error, and Nonspecific Molecular Tools
Authors: Dyer Alan T. , Leonard Kurt J.
Published: May 2007

Random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) are widely used in studies of genetic variation. Although it is recognized that contamination should be avoided in DNA samples, little is known about the potential hazards of low level bacterial contamination of samples from which DNA is extracted for RAPD or AFLP analyses. We found that contamination of Aphanomyces cochlioides cultures with a prokaryote at visibly undetectable levels markedly altered the results of RAPD and AFLP analyses. The contamination resulted in seven contaminant-specific RAPD products and in the suppression of eight products characteristic of uncontaminated A. cochlioides cultures. Prokaryote contamination resulted in 39 contaminant-specific AFLP products, but did not cause suppression of AFLP products. Comparing A. cochlioides samples with outgroup A. euteiches did not clearly indicate the presence of contaminant DNA, because uneven product suppression in RAPD analysis increased the apparent similarity between contaminated samples and A. euteiches and because a high proportion of the contaminant-specific amplified products comigrated with products from A. euteiches in both RAPD and AFLP analyses. Work with organisms that are prone to contamination should employ techniques such as restriction fragment length polymorphism or DNA sequence comparisons rather than relying solely on RAPD or AFLP analyses.
(+)-Catechin Acts as an Infection-Inhibiting Factor in Strawberry Leaf
Authors: Yamamoto M. , Nakatsuka S. , Otani H. , Kohmoto K. , Nishimura S.
Published: May 2007

An infection-inhibiting factor (IIF) was isolated from strawberry leaves and identified as (+)-catechin. This compound inhibited the formation of infection hyphae from appressoria of Alternaria alternata, but allowed both spore germination and appressorial formation. It is a normal component of strawberry leaves, but further accumulates as the major IIF in response to inoculation with nonpathogenic spores of A. alternata. The accumulation of (+)-catechin on a susceptible host was not induced, however, by inoculation with pathogenic spores of the strawberry pathotype or by inoculation with nonpathogenic spores supplemented with host-specific toxin (AF-toxin I). These results imply that (+)-catechin acts as a protective agent during induced resistance and that AF-toxin I acts as a fungal suppressor of induced resistance.
Identification and Subgrouping of Cucumber mosaic virus with Mouse Monoclonal Antibodies
Authors: Hsu H. T. , Barzuna L. , Hsu Y. H. , Bliss W. , Perry K. L.
Published: May 2007

Using a mixture of isolates of Cucumber mosaic virus (CMV) from subgroups I and II as immunogens, 20 mouse hybridoma cell lines secreting monoclonal antibodies were produced. A reliable method for efficient detection and accurate subgrouping of CMV isolates has been developed. Tests with 12 well-characterized strains of CMV and other cucumoviruses demonstrated the presence of epitopes that were virus and subgroup specific. Analyses of 109 accessions of CMV isolates collected from various parts of the world revealed 70% were subgroup I, with 20% identified as subgroup II. Seven isolates (6%) did not react with group-specific antibodies but did react with antibodies that recognized all CMV isolates. Differential reactions among isolates suggested a total of 10 epi-topes were recognized. The antigenic diversity among subgroup II CMVs was greater than for the subgroup I isolates, even though fewer subgroup II isolates were tested.
Spiroplasma citriSurface Protein P89 Implicated in Adhesion to Cells of the VectorCirculifer tenellus
Authors: Yu J. , Wayadande A. C. , Fletcher J.
Published: May 2007

Two microtiter plate assays were developed to study the adherence of the plant-pathogenic mollicute Spiroplasma citri to a monolayer of cultured cells of its leafhopper vector, Circulifer tenellus. Adherence was significantly reduced by prior treatment of the spiroplasmas with proteinase K or pronase. Electrophoresis and western blotting of spiroplasma membrane proteins, before and after exposure of intact spiroplasmas to proteases, revealed the concomitant reduction in intensity of a major membrane protein (P89) and a new polypeptide of ≈46 kDa in protease-treated preparations (P46). Triton X-114 phase partitioning demonstrated that P89 and P46 are amphiphilic, and labeling of the new polypeptide P46 with anti-P89 serum suggested that this molecule may be a breakdown product of P89. Regeneration of P89 after proteinase K treatment of spiroplasmas was directly associated with restoration of the pathogen's attachment capability. Treatment of spiroplasmas with any of several carbohydrates and glycoconjugates or with tetramethyl-urea, a compound that interferes with hydrophobic associations, had a negligible effect on attachment. These results suggest that a spiroplasma surface protein, P89, has a role in S. citri adherence to C. tenellus cells.
Influences of Cropping Practices on Verticillium dahliae Populations in Commercial Processing Tomato Fields in Ontario
Authors: Harrington M. A. , Dobinson K. F.
Published: May 2007

The abundance of Verticillium dahliae in the soil and the incidence of V. dahliae-infected plants were determined for 12 commercial processing tomato fields in Kent County, Ontario. Comparison of the data with those from a previous survey of fields in adjacent Essex County showed that soil inoculum levels and incidence of infection were generally lower in Kent County fields and that race 2 V. dahliae was not common in Kent County. From the two counties, 128 isolates were characterized by restriction fragment length polymorphism (RFLP) analysis, using the subspecies-specific repetitive DNA sequence E18. A subset of these isolates was also characterized by vegetative compatibility and DNA hybridization analysis with a second subspecies-specific DNA sequence. Isolates with E18 RFLP profiles highly similar to those of isolates previously collected from potato fields in North America were prevalent in Essex County tomato fields but not common in Kent County fields. The data are consistent with the hypothesis that the group I isolates were introduced into southwestern Ontario with potato and that the different cultural practices in Essex County and Kent County have contributed to the differences in the accumulation of these isolates in the two regions.
Characterization of Ectophosphatase Activities in Trypanosomatid Parasites of Plants
Authors: Dutra P. M. L. , Rodrigues C. O. , Romeiro A. , Grillo L. A. M. , Dias F. A. , Attias M. , De Souza W. , Lopes A. H. C. S. , Meyer-Fernandes J. R.
Published: May 2007

In the present work ectophosphatase activities of three trypanosomatid parasites of plants were characterized using intact cells. Phytomonas françai, Phytomonas mcgheei, and Herpetomonas sp. hydrolyzed p-nitro-phenylphosphate at a rate of 5.40, 7.28, and 25.58 nmol Pi/mg of protein per min, respectively. Experiments using classical inhibitors of acid phosphatases such as sodium orthovanadate (NaVO3) and sodium fluoride (NaF) showed a decrease in phosphatase activities. Lithium fluoride (LiF) and aluminum chloride (AlCl3) were also used. Although AlCl3 had no effect, LiF was able to promote a decrease in the phosphatase activities. Interestingly, the inhibition caused by LiF was enhanced by the addition of AlCl3 during the reaction, probably due to the formation of fluoroaluminate complexes. This effect was confirmed by cytochemical analysis. In this assay, electron-dense cerium phosphate deposits were visualized on the external surface of the three parasites.
Identification and Differentiation of Tilletia indica and T. walkeri Using the Polymerase Chain Reaction
Authors: Frederick Reid D. , Snyder Karen E. , Tooley Paul W. , Berthier-Schaad Yvette , Peterson Gary L. , Bonde Morris R. , Schaad Norman W. , Knorr David A.
Published: May 2007

Karnal bunt of wheat, caused by Tilletia indica, was found in regions of the southwestern United States in 1996. Yield losses due to Karnal bunt are slight, and the greatest threat of Karnal bunt to the U.S. wheat industry is the loss of its export market. Many countries either prohibit or restrict wheat imports from countries with Karnal bunt. In 1997, teliospores morphologically resembling T. indica were isolated from bunted ryegrass seeds and wheat seed washes. Previously developed PCR assays failed to differentiate T. indica from the recently discovered ryegrass pathogen, T. walkeri. The nucleotide sequence of a 2.3 kb region of mitochondrial DNA, previously amplified by PCR only from T. indica, was determined for three isolates of T. indica and three isolates of T. walkeri. There was greater than 99% identity within either the T. indica group or the T. walkeri group of isolates, whereas there was =3% divergence between isolates of these two Tilletia species. Five sets of PCR primers were made specific to T. indica, and three sets were designed specifically for T. walkeri based upon nucleotide differences within the mitochondrial DNA region. In addition, a 212 bp amplicon was developed as a target sequence in a fluorogenic 5′ nuclease PCR assay using the TaqMan system for the detection and discrimination of T. indica and T. walkeri.
Using Restriction Fragment Length Polymorphisms to Assess Temporal Variation and Estimate the Number of Ascospores that Initiate Epidemics in Field Populations of Mycosphaerella graminicola
Authors: Zhan J. , Mundt C. C. , McDonald B. A.
Published: May 2007

Restriction fragment length polymorphisms (RFLPs) and DNA fingerprints were used to assess temporal variation and estimate the effective population size of the wheat pathogen Mycosphaerella graminicola over a 6-year period. In each year, the fungal population was founded by ascospores originating from outside the sampled fields. A total of 605 fungal isolates were included in this study. Our results indicate that the genetic structure of these M. graminicola populations were stable over the 6-year period. The common alleles at each RFLP locus were present at similar frequencies each year. More than 99% of gene diversity was distributed within populations sampled from the same year and less than 1% was attributed to differences among years. The lack of population differentiation among collections taken in different years indicated that the effective size of the source population was sufficiently large that genetic drift was insignificant in this location. It also suggests that the initial colonists from ascospore founder populations were a fair reflection of the source population. We estimate that the effective sizes of these field populations ranged from 3,400 to 700,000 individuals, depending on the size of the field sampled and assumptions about mutation rates. Estimates of the number of ascospores initiating epidemics of leaf blotch disease in each field plot and factors that contribute to the large effective population size of M. graminicola are discussed.