bluuebunny/crossref_metadata_2025 huggingface dataset:
Correlations Between Most Probable Number and Activity of Nematode-Trapping Fungi
Authors: Jaffee B. A.
Published: May 2007

Soil cages were used to determine whether nematode-trapping fungi population density, as measured by most probable number (MPN) procedures, was correlated with the trapping of nematodes. Fungi studied (and trap type) were Arthrobotrys oligospora (adhesive networks), A. eudermata (adhesive networks), A. dactyloides (constricting rings), Dactylellina ellipsospora (adhesive knobs), and D. haptotyla (adhesive knobs). The fungi were formulated as assimilative hyphae in dried alginate pellets. Pellets were added to field soil, the soil was packed into 80-cm3 cages (PVC pipe, 3.0 cm long and 3.9 cm in diameter), and the cages were buried in vineyards. After 14 to 61 days, the cages were recovered, and MPN data and trapping activity were determined. For all five fungi, MPN data were correlated with the number of pellets added. Regardless of fungus population density, A. oligospora and A. eudermata trapped few if any nematodes in soil, and consequently, trapping and fungus population density were not correlated. The correlation between population density and trapping was weak for A. dactyloides but relatively strong for D. ellipsospora and D. haptotyla. High levels of trapping by the latter two fungi required more than 102 fungus propagules per gram of soil.
Inheritance of Resistance to Bacterial Blight in 21 Cultivars of Rice
Authors: Lee K. S. , Rasabandith S. , Angeles E. R. , Khush G. S.
Published: May 2007

Genetic analysis for resistance to bacterial blight (Xanthomonas oryzae pv. oryzae) of 21 rice (Oryza sativa L.) cultivars was carried out. These cultivars were divided into two groups based on their reactions to Philippine races of bacterial blight. Cultivars of group 1 were resistant to race 1 and those of group 2 were susceptible to race 1 but resistant to race 2. All the cultivars were crossed with TN1, which is susceptible to all the Philippine races of X. oryzae pv. oryzae. F1 and F2 populations of hybrids of group 1 cultivars were evaluated using race 1 and F1 and F2 populations of hybrids of group 2 cultivars were evaluated using race 2. All the cultivars showed monogenic inheritance of resistance. Allelic relationships of the genes were investigated by crossing these cultivars with different testers having single genes for resistance. Three cultivars have Xa4, another three have xa5, one has xa8, two have Xa3, eight have Xa10, and one has Xa4 as well as Xa10. Three cultivars have new, as yet undescribed, genes. Nep Bha Bong To has a new recessive gene for moderate resistance to races 1, 2, and 3 and resistance to race 5. This gene is designated xa26(t). Arai Raj has a dominant gene for resistance to race 2 which segregates independently of Xa10. This gene is designated as Xa27(t). Lota Sail has a recessive gene for resistance to race 2 which segregates independently of Xa10. This gene is designated as xa28(t).
Multiplex Polymerase Chain Reaction Identification of Single Individuals of the Longidorid Nematodes Xiphinema index, X. diversicaudatum, X. vuittenezi, and X. italiae Using Specific Primers from Ribosomal Genes
Authors: Wang Xinrong , Bosselut Nathalie , Castagnone Chantal , Voisin Roger , Abad Pierre , Esmenjaud Daniel
Published: May 2007

The species X. index, X. diversicaudatum, X. vuittenezi, and X. italiae are established (E) or putative (P) vectors of Grapevine fanleaf virus (GFLV) (E), Arabis mosaic virus (E), Grapevine chrome mosaic virus (P), and GFLV (P) nepoviruses of grapevine, respectively. All four species are very closely related taxonomically and their low field densities make them difficult to identify from morphological and morphometrical diagnostic characters when only single or few individuals are detected. To improve diagnostic accuracy, a simple method was developed. The internal transcribed spacer 1 (ITS1) region spanning the 18S and 5.8S ribosomal genes was sequenced in one population of each species using two conserved primers from these genes. The ITS1 fragments were 1,132 bp (X. vuittenezi), 1,153 bp (X. index), 1,175 bp (X. diversicaudatum), and 1,190 bp (X. italiae), i.e., a difference of over 5% between the extremes. The sequence variability made it possible to design species-specific internal sense primers that amplified, in combination with the same antisense ITS1 primer, a single signature fragment (340 bp for X. index, 414 bp for X. italiae, 591 bp for X. vuittenezi, and 813 bp for X. diversicaudatum). Tests with DNA from a single adult or juvenile nematode confirmed the specificity of the primers from diverse isolates or populations. The primers were successfully used in a multiplex test for the reliable detection of two to four mixed species, each represented by a single individual. This multiplex-based diagnostic tool will be particularly useful for successful nematode management practices in vineyards.
Use of Inter-Simple Sequence Repeats and Amplified Fragment Length Polymorphisms to Analyze Genetic Relationships Among Small Grain-Infecting Species of Ustilago
Authors: Menzies J. G. , Bakkeren G. , Matheson F. , Procunier J. D. , Woods S.
Published: May 2007

In the smut fungi, few features are available for use as taxonomic criteria (spore size, shape, morphology, germination type, and host range). DNA-based molecular techniques are useful in expanding the traits considered in determining relationships among these fungi. We examined the phylogenetic relationships among seven species of Ustilago (U. avenae, U. bullata, U. hordei, U. kolleri, U. nigra, U. nuda, and U. tritici) using inter-simple sequence repeats (ISSRs) and amplified fragment length polymorphisms (AFLPs) to compare their DNA profiles. Fifty-four isolates of different Ustilago spp. were analyzed using ISSR primers, and 16 isolates of Ustilago were studied using AFLP primers. The variability among isolates within species was low for all species except U. bullata. The isolates of U. bullata, U. nuda, and U. tritici were well separated and our data supports their speciation. U. avenae and U. kolleri isolates did not separate from each other and there was little variability between these species. U. hordei and U. nigra isolates also showed little variability between species, but the isolates from each species grouped together. Our data suggest that U. avenae and U. kolleri are monophyletic and should be considered one species, as should U. hordei and U. nigra.
Quantification of Fusarium solani f. sp. phaseoli in Mycorrhizal Bean Plants and Surrounding Mycorrhizosphere Soil Using Real-Time Polymerase Chain Reaction and Direct Isolations on Selective Media
Authors: Filion M. , St-Arnaud M. , Jabaji-Hare S. H.
Published: May 2007

The capacity of the arbuscular mycorrhizal fungus Glomus intraradices in reducing the presence of Fusarium solani f. sp. phaseoli in bean plants and the surrounding mycorrhizosphere soil was evaluated in a compartmentalized experimental system. Quantification of the pathogen and the symbiont in plant tissues, the soil regions of the mycorrhizosphere (rhizosphere and mycosphere), and the bulk soil was accomplished using specific polymerase chain reaction (PCR) primers in real-time PCR assays, culture-dependant methods, and microscopic determination techniques. Nonmycorrhizal bean plants infected with the pathogen had distinctive Fusarium root rot symptoms, while infected plants previously colonized by G. intraradices remained healthy. The amount of F. solani f. sp. phaseoli genomic DNA was significantly reduced in mycorrhizal bean plants and in each mycorrhizosphere soil compartment. The presence of G. intraradices in the mycorrhizosphere was not significantly modified, although the mycorrhizal colonization of roots was slightly increased in the presence of the pathogen. The results suggest that the reduced presence of Fusarium as well as root rot symptoms are caused by biotic and/or abiotic modifications of the mycorrhizosphere as a result of colonization with G. intraradices.
Biological Control of Pathogens Causing Root Rot Complex in Field Pea Using Clonostachys rosea Strain ACM941
Authors: Xue Allen G.
Published: May 2007

Pea root rot complex (PRRC), caused by Alternaria alternata, Aphanomyces euteiches, Fusarium oxysporum f. sp. pisi, F. solani f. sp. pisi, Mycosphaerella pinodes, Pythium spp., Rhizoctonia solani, and Sclerotinia sclerotiorum, is a major yield-limiting factor for field pea production in Canada. A strain of Clonostachys rosea (syn. Gliocladium roseum), ACM941 (ATCC 74447), was identified as a mycoparasite against these pathogens. When grown near the pathogen, ACM941 often was stimulated to produce lateral branches that grew directly toward the pathogen mycelium, typically entwining around the pathogen mycelium. When applied to the seed, ACM941 propagated in the rhizosphere and colonized the seed coat, hypocotyl, and roots as the plant developed and grew. ACM941 significantly reduced the recovery of all fungal pathogens from infected seed, increased in vitro seed germination by 44% and seedling emergence by 22%, and reduced root rot severity by 76%. The effects were similar to those of thiram fungicide, which increased germination and emergence by 33 and 29%, respectively, and reduced root rot severity by 65%. When soil was inoculated with selected PRRC pathogens in a controlled environment, seed treatment with ACM941 significantly increased emergence by 26, 38, 28, 13, and 21% for F. oxysporum f. sp. pisi, F. solani f. sp. pisi, M. pinodes, R. solani, and S. sclerotiorum, respectively. Under field conditions from 1995 to 1997, ACM941 increased emergence by 17, 23, 22, 13, and 18% and yield by 15, 6, 28, 6, and 19% for the five respective pathogens. The seed treatment effects of ACM941 on these PRRC pathogens were greater or statistically equivalent to those achieved with thiram. Results of this study suggest that ACM941 is an effective bioagent in controlling PRRC and is an alternative to existing chemical products.
The Population Structure of Phytophthora infestans from the Toluca Valley of Central Mexico Suggests Genetic Differentiation Between Populations from Cultivated Potato and Wild Solanum spp.
Authors: Flier Wilbert G. , Grünwald Niklaus J. , Kroon Laurens P. N. M. , Sturbaum Anne K. , van den Bosch Trudy B. M. , Garay-Serrano Edith , Lozoya-Saldaña Hector , Fry William E. , Turkensteen Lod J.
Published: May 2007

The population structure of Phytophthora infestans in the Toluca Valley of central Mexico was assessed using 170 isolates collected from cultivated potatoes and the native wild Solanum spp., S. demissum and S. xendinense. All isolates were analyzed for mitochondrial DNA (mtDNA) haplotype and amplified fragment length polymorphism (AFLP) multi-locus fingerprint genotype. Isolate samples were monomorphic for mtDNA haplotype because all isolates tested were of the Ia haplotype. A total of 158 multilocus AFLP genotypes were identified among the 170 P. infestans isolates included in this study. P. infestans populations sampled in the Toluca Valley in 1997 were highly variable and almost every single isolate represented a unique genotype based on the analysis of 165 AFLP marker loci. Populations of P. infestans collected from the commercial potato-growing region in the valley, the subsistence potato production area along the slopes of the Nevado de Toluca, and the native Solanum spp. on the forested slopes of the volcano showed a high degree of genetic diversity. The number of polymorphic loci varied from 20.0 to 62.4% for isolates collected from the field station and wild Solanum spp. On average, 81.8% (135) of the AFLP loci were polymorphic. Hetero-zygosity varied between 7.7 and 19.4%. Significant differentiation was found at the population level between strains originating from cultivated potatoes and wild Solanum spp. (P = 0.001 to 0.022). Private alleles were observed in individual isolates collected from all three populations, with numbers of unique dominant alleles varying from 9 to 16 for isolates collected from commercial potato crops and native Solanum spp., respectively. Four AFLP markers were exclusively found present in isolates collected from S. demissum. Indirect estimation of gene flow between populations indicated restricted gene flow between both P. infestans populations from cultivated potatoes and wild Solanum hosts. There was no evidence found for the presence of substructuring at the subpopulation (field) level. We hypothesize that population differentiation and genetic isolation of P. infestans in the Toluca Valley is driven by host-specific factors (i.e., R-genes) widely distributed in wild Solanum spp. and random genetic drift.
A Generalized Linear Modeling Approach for Characterizing Disease Incidence in a Spatial Hierarchy
Authors: Turechek W. W. , Madden L. V.
Published: May 2007

Several statistical models are introduced to quantify the effect of heterogeneity on disease incidence relationships in a three-scale spatial hierarchy: the sampling unit level (highest), the leaf scale (intermediate), and the leaflet scale (lowest). The models are an extension of the theory previously developed for a two-scale hierarchy and were tested using data collected from strawberry leaf blight epidemics. Disease incidence at the sampling-unit scale (proportion of sampling units with one or more diseased leaflets) increased as a saturation-type curve with increasing leaflet or leaf disease incidence (proportion of leaflets or leaves diseased) as predicted by the good fit of the beta-binomial distribution to the leaflet and leaf data. The relationship could be accurately described, without curve fitting, by several simple nonlinear models, in which the aggregation of disease was represented by a modified binomial function incorporating an effective sample size that was either constant or dependent on mean incidence. The relationship between incidence at the leaflet and leaf scales could be modeled based on the combined sampling-unit models for leaflets and leaves. By taking the complementary log-log (CLL) transformation of incidence, the equations could be expressed as generalized linear models, and curve fitting used to estimate the parameters. Generally, curve fitting gave slight to no improvement in the accuracy of the predictions of incidence. These models have broad applicability in sampling for disease incidence, and results can be used to interpret how diseased individuals at the lowest level in a hierarchy are arranged within sampling units.
Infection of Commercial Hybrid Primula Seed by Botrytis cinerea and Latent Disease Spread Through the Plants
Authors: Barnes S. E. , Shaw M. W.
Published: May 2007

Botrytis cinerea occurred commonly on cultivated Primula ×polyantha seed. The fungus was mostly on the outside of the seed but sometimes was present within the seed. The fungus frequently caused disease at maturity in plants grown from the seed, demonstrated by growing plants in a filtered airflow, isolated from other possible sources of infection. Young, commercially produced P. ×polyantha plants frequently had symptomless B. cinerea infections spread throughout the plants for up to 3 months, with symptoms appearing only at flowering. Single genetic individuals of B. cinerea, as determined by DNA fingerprinting, often were dispersed widely throughout an apparently healthy plant. Plants could, however, contain more than one isolate.
Coevolution in Natural Pathosystems: Effects of Dominance on Host-Pathogen Interactions
Authors: Ye Terrance Z. , Yang Rong-Cai , Yeh Francis C.
Published: May 2007

We have developed a new complementary model of gene interaction between diploid host and haploid pathogen by allowing for arbitrary levels of dominance in the host. This model enables us to assess the effects of overdominance, incomplete dominance, and underdominance on the equilibrium frequencies of resistance and virulence genes and on the stability of equilibria. Our model reduces to a gene-for-gene model when complete dominance of resistance is assumed. Computer simulations show that our model has two new features. First, when there is overdominance or underdominance of resistance, the internal equilibrium points exist even when there is no cost of unnecessary virulence or when there is a cost of necessary virulence at the balance between cost of unnecessary virulence and effectiveness of resistance. Second, the occurrence of stable resistance and virulence polymorphism is strongly dependent on the level of dominance. These two features suggest the need for caution when using the gene-for-gene model, especially in the presence of overdominance or underdominance. Our model is particularly suitable for studying the coevolutionary dynamics between hybrid populations and their pathogens in natural pathosystems.
Population Genetic Structure of Tapesia acuformis in Washington State
Authors: Douhan G. W. , Murray T. D. , Dyer P. S.
Published: May 2007

Eyespot of wheat is caused by Tapesia yallundae and T. acuformis. Historically, T. yallundae has been considered the more important causal agent of the disease in Washington state and consists of a large homogeneous population with a genetic structure consistent with both sexual and asexual reproduction. T. acuformis has increased significantly in Washington in the past 10 years and apothecia were found recently under natural field conditions, indicating that T. acuformis may have a more important role in eyespot of wheat than previously was thought. To determine the genetic structure of T. acuformis in Washington, 141 single conidial isolates were sampled from four subpopulations in the eastern wheat-growing region of the state. Isolates were scored for mating type and six amplified fragment length polymorphism markers. All markers segregated in a 1:1 ratio and were determined to be unlinked based on genetic analysis of 24 progeny from an in vitro cross. No significant differences in allele frequencies (0.127 < P < 0.809) were found among individual loci across the four subpopulations and over all loci based on contingency table analysis of the log-likelihood ratio statistic G2. Likewise, no overall differences between subpopulations were detected using the population differentiation statistic θ (θ = -0.004, P = 0.537). Random mating could not be rejected within each subpopulation or for the combined data using clone-corrected data sets based on (i) 1:1 ratio of mating-type, (ii) multilocus gametic disequilibrium analyses (index of association), (iii) phylogenetic analyses (parsimony tree length permutation test), and (iv) genotypic diversity analyses. T. acuformis has a genetic structure similar to that of sympatric populations of T. yallundae in Washington, with both sexual and asexual reproduction contributing to the structuring of this species.
Origin and Distribution of Cr2, a Gene for Resistance to White Pine Blister Rust in Natural Populations of Western White Pine
Authors: Kinloch Bohun B. , Sniezko Richard A. , Dupper Gayle E.
Published: May 2007

The distribution and frequency of the Cr2 gene for resistance to white pine blister rust (Cronartium ribicola) in western white pine (Pinus monticola) was surveyed in natural populations of the host by inoculation of open-pollinated seedlings from 687 individual seed parents from throughout most of the species' range. Because Cr2 is dominant and results in a conspicuous hypersensitive reaction (HR) in pine needles, the phenotype can readily be detected in offspring of susceptible seed parents fertilized by unknown Cr2 donors in the ambient pollen cloud. Gametic frequencies of Cr2 were thus determined as the proportion of total challenged seedlings that were pollen receptors exhibiting the Cr2 phenotype. Zygotic frequencies, the proportion of seed parents with progeny that segregated in Mendelian ratios for the Cr2 phenotype to the total number of parents, were a complementary, though less precise, measure. Cr2 frequency was rare overall, ranging from 0.004 to 0.008 in the Sierra Nevada to about 0.001 in the central Cascade Range; it was undetectable further north in the Cascades, as well as in the Rocky Mountains and Coast Mountains of the United States and Canada. The diminishing frequency of Cr2 from the southern and central Sierra Nevada northward mirrors that of Cr1 in sugar pine (P. lambertiana) and points to this region as the origin of both genes. We rationalize that this coincidence may have resulted from protection that these genes may have conferred on both species to an endemic pine stem rust congeneric with C. ribicola (C. occidentale) in recent geologic epochs.
Temporal Dynamics of the Biocontrol Agent Pseudomonas fluorescens Strain A506 in Flowers in Inoculated Pear Trees
Authors: Lindow Steven E. , Suslow Trevor V.
Published: May 2007

The colonization of individual flowers in mature pear orchards by Pseudomonas fluorescens strain A506 applied at different times during bloom was measured to determine the receptivity of flowers to colonization and the extent of intra-tree movement over time. Strain A506 populations in flowers open at inoculation were initially about 104 cells per flower and increased to approximately 106 cells per flower in flowers that were inoculated within about 5 days of opening. However, eventual populations decreased with further increases in flower age at inoculation to as few as about 103 cells per flower when inoculated flowers were more than 10 days old. Populations of strain A506 on flowers that opened after inoculation was initially very low at the time of petal expansion (<100 cells per flower) but increased rapidly with time after flower opening. The maximum population of strain A506 that developed on such flowers decreased with increasing time between inoculation and petal expansion; 104 to 105 cells of strain A506 eventually colonized flowers that opened within 7 days of inoculation, whereas fewer than 100 cells colonized flowers that opened 24 days or more after inoculation. Large total bacterial populations on A506-treated trees were associated with significant reductions in populations of Erwinia amylovora and reduced incidence of fire blight and severity of fruit russet.
A Species-Specific Polymerase Chain Reaction Assay for Rapid Detection of Phytophthora nicotianae in Irrigation Water
Authors: Kong Ping , Hong Chuanxue , Jeffers Steven N. , Richardson Patricia A.
Published: May 2007

Phytophthora nicotianae is a common and destructive pathogen of numerous ornamental, agronomic, and horticultural crops such as tobacco, tomato, and citrus. We have developed a species-specific polymerase chain reaction (PCR) assay for rapid and accurate detection of this pathogen in irrigation water, a primary source of inoculum and an efficient means of propagule dissemination. This PCR assay consists of a pair of species-specific primers (PN), customization of a commercial soil DNA extraction kit for purification of DNA from propagules in irrigation water, and efficient PCR protocols for primer tests and sample detection. The PN primers proved adequately specific for P. nicotianae in evaluations with 131 isolates of P. nicotianae, 102 isolates from 15 other species of Phytophthora, and 64 isolates from a variety of other oomycetes, true fungi, and bacteria. These isolates originated from a wide range of host plants, three substrates (plant tissue, soil, and irrigation water), and numerous geographic locations. The detection sensitivity is between 80 and 800 fg DNA/μl. The assay detected the pathogen in naturally infested water samples from Virginia and South Carolina nurseries more rapidly and accurately than standard isolation methods. Use of this PCR assay can assist growers in making timely disease management decisions with confidence.
Origin of Race 3 of Fusarium oxysporum f. sp. lycopersici at a Single Site in California
Authors: Cai G. , Gale L. Rosewich , Schneider R. W. , Kistler H. C. , Davis R. M. , Elias K. S. , Miyao E. M.
Published: May 2007

Thirty-nine isolates of Fusarium oxysporum were collected from tomato plants displaying wilt symptoms in a field in California 2 years after F. oxysporum f. sp. lycopersici race 3 was first observed at that location. These and other isolates of F. oxysporum f. sp. lycopersici were characterized by pathogenicity, race, and vegetative compatibility group (VCG). Of the 39 California isolates, 22 were in VCG 0030, 11 in VCG 0031, and six in the newly described VCG 0035. Among the isolates in VCG 0030, 13 were race 3, and nine were race 2. Of the isolates in VCG 0031, seven were race 2, one was race 1, and three were nonpathogenic to tomato. All six isolates in VCG 0035 were race 2. Restriction fragment length polymorphisms (RFLPs) and sequencing of the intergenic spacer (IGS) region of rDNA identified five IGS RFLP haplotypes, which coincided with VCGs, among 60 isolates of F. oxysporum from tomato. Five race 3 isolates from California were of the same genomic DNA RFLP haplotype as a race 2 isolate from the same location, and all 13 race 3 isolates clustered together into a subgroup in the neighbor joining tree. Collective evidence suggests that race 3 in California originated from the local race 2 population.
Reliability and Accuracy of Visual Estimation of Phomopsis Leaf Blight of Strawberry
Authors: Nita M. , Ellis M. A. , Madden L. V.
Published: May 2007

Six different individuals (raters) assessed the severity of Phomopsis leaf blight on strawberry leaflets in five experimental repetitions over 2 years by making a direct visual estimation of the percentage of diseased area of each leaflet or by using the Horsfall-Barratt (H-B) disease scale. Intra-rater and inter-rater reliability and accuracy were determined, and then the relationship between visually estimated severity values and actual severity values was evaluated. Agreement in estimated disease severity values between assessment times by the same raters (i.e., intra-rater reliability), and agreement in disease severity values among raters at a single assessment time (i.e., inter-rater reliability), were both high, with most correlation coefficients being greater than 0.85. The intra-class correlation for overall agreement among raters ranged from 0.80 to 0.96 for the five repetitions. Based on the concordance coefficient calculated for each rater in each repetition, agreement between estimated and actual severity (i.e., accuracy) was somewhat lower than reliability. The relationship between estimated and actual severity was linear, and there was a slight trend to overestimate disease severity. The H-B scale was not more reliable or accurate than direct estimation of severity, and the linear relationship between estimated and actual severity did not support the principles underling the H-B scale. Both size of leaflets and number of lesions per leaflet slightly affected the error in estimate of disease severity.
Molecular Diversity of Ralstonia solanacearum Isolated from Ginger in Hawaii
Authors: Yu Q. , Alvarez A. M. , Moore P. H. , Zee F. , Kim M. S. , de Silva A. , Hepperly P. R. , Ming R.
Published: May 2007

The genetic diversity of Ralstonia solanacearum strains isolated from ginger (Zingiber officinale) growing on the island of Hawaii was determined by analysis of amplified fragment length polymorphisms (AFLPs). Initially 28 strains of R. solanacearum collected from five host plant species worldwide were analyzed by AFLP. A second analysis was conducted on 55 R. solanacearum strains collected from three ginger farms along the Hamakua Coast of Hawaii, the principle area of ginger cultivation in the state. From the initial analysis, R. solanacearum strains from ginger in Hawaii showed a high degree of similarity at 0.853. In contrast, the average genetic similarity between R. solanacearum strains from heliconia and ginger was only 0.165, and strains from ginger showed little similarity with strains from all other hosts. The second analysis of 55 strains from ginger on different Hawaiian farms confirmed that they were distinct from race 1 strains from tomato. Strains from ginger also showed greater diversity among themselves in the second analysis, and the greatest diversity occurred among strains from a farm where ginger is frequently imported and maintained. Our results provide evidence that R. solanacearum strains from ginger in Hawaii are genetically distinct from local strains from tomato (race 1) and heliconia (race 2).
Detection of Begomoviruses in Clarified Plant Extracts: A Comparison of Standard, Direct-Binding, and Immunocapture Polymerase Chain Reaction Techniques
Authors: Rampersad S. N. , Umaharan P.
Published: May 2007

Three polymerase chain reaction (PCR) techniques—standard PCR (Std-PCR), direct-binding PCR (DB-PCR), and immunocapture PCR (IC-PCR)—using degenerate primers were optimized and evaluated for the detection of begomoviruses. Tomato leaf samples were ground in three different extraction buffers and subjected to Std-PCR. The effect of the buffers on the detection limits of amplification of the virus (detection of the initial and end points of dilution) was determined. With the optimal extraction buffer determined by the first experiment, the antibody concentration and incubation conditions for IC-PCR were evaluated to determine the requirements for maximum capture of antigens during the capture phase of the technique. The incubation conditions of DB-PCR were also investigated to determine the most favorable conditions for adsorption of the viral template. The reproducibility of all assays was evaluated. With the results of the optimization experiments, the applicability of the three techniques to different plant species was assessed. Extracts of plant species belonging to three families were subjected to the optimized Std-, DB-, and IC-PCR protocols. Std- and IC-PCR both achieved reproducible detection of begomoviruses, but the detection limits and amplified band intensity for all plant species tested were superior for the latter. DB-PCR was an unreliable method of detection, because of poor reproducibility and low intensity of amplified bands. These results indicate that the optimized IC-PCR detection system using degenerate primers is the most effective for the detection of begomoviruses in clarified plant extracts.
Effect of the Host and Temperature on the Formation of Defective RNAs Associated with Broad bean mottle virus Infection
Authors: Llamas Susana , Sandoval Claudio , Babin Mar , Pogany Judy , Bujarski Jozef J. , Romero Javier
Published: May 2007

Previously, we demonstrated that Broad bean mottle virus (BBMV), a member of the genus Bromovirus, could accumulate RNA 2-derived defective interfering (DI) RNAs during infection. In this work, we study how host and environmental factors affect the accumulation of DI RNAs. Serial passages of BBMV through selected plant species reveal that, with low-multiplicity inocula, some systemic hosts (Vicia faba, Nicotiana clevelandii, and N. tabacum cv. Samsum) support DI RNA accumulation after the first passage cycle but other hosts (Phaseolus vulgaris, Pisum sativum, and Glycine max) do not. However, several passages with the high-multiplicity inocula can generate DI RNAs in pea plants. Local lesion hosts (Chenopodium quinoa, C. amaranticolor, and C. murale) remain free of the DI RNA components. The size of the de novo-formed DI RNAs depends on the host and on environmental conditions. For instance, broad bean plants cultivated in a greenhouse or in a growth chamber at 20°C accumulated DI RNAs of 2.4 or 1.9 kb in size, respectively. A reverse trend was observed in pea plants. Lower temperatures greatly facilitated the formation of DI RNAs in broad bean and pea hosts after the first passage. The importance of these findings for the studies on DI RNAs are discussed.
Control of Plant Diseases by Extracts of Inula viscosa
Authors: Wang Wenqiao , Ben-Daniel B. H. , Cohen Yigal
Published: May 2007

Leaves of Inula viscosa were collected from the field, dried, and extracted with a mixture of acetone and n-hexane. The oily, water-insoluble pastes obtained after evaporation of the solvents were used for the control of foliar diseases in growth chambers. The pastes, either dissolved in acetone or emulsified in water, effectively controlled downy mildew of cucumber, late blight of potato or tomato, powdery mildew of wheat, and rust of sunflower. Mean effective dose (concentration) required for 90% inhibition of disease values for acetone solutions and water emulsions ranged from 0.68 to 1.02% and 0.65 to 1.00% (wt/vol), respectively. Dry matter content in fresh leaves, paste-extract yield in dry leaves, and disease control efficacy of paste extracts were similar in leaves of I. viscosa collected during May to October, suggesting that, for practical use, harvests can be conducted during most of the growing season. The results show that I. viscosa may be used as an herbal source for fungicidal preparations against foliar diseases caused by pathogens belonging to the families Oomycetes, Ascomycetes, and Basidiomycetes.
Cotton leaf crumple virus Is a Distinct Western Hemisphere Begomovirus Species with Complex Evolutionary Relationships Indicative of Recombination and Reassortment
Authors: Idris A. M. , Brown J. K.
Published: May 2007

The bipartite DNA genome of Cotton leaf crumple virus (CLCrV), a whitefly-transmitted begomovirus from the Sonoran Desert, was cloned and completely sequenced. The cloned CLCrV genome was infectious when biolistically delivered to cotton or bean seedlings and progeny virus was whitefly-transmissible. Koch's postulates were completed by the reproduction of characteristic leaf crumple symptoms in cotton seedlings infected with cloned CLCrV DNA, thereby verifying the etiology of leaf crumple disease, which has been known in the southwestern United States since the 1950s. Sequence comparisons confirmed that CLCrV has a genome organization typical of yet sufficiently divergent from all other bipartite begomoviruses to justify recognition as a distinct species. Phylogenetic analyses indicated that CLCrV has a complex evolutionary history probably involving both recombination and reassortment. The relatively low nucleotide sequence identity (77%) of the common region shared by the CLCrV DNA-A and DNA-B components and the distinct phylogenetic relationships of each component are consistent with component reassortment. Sequence analyses indicated that the CLCrV DNA-A component was likely derived by recombination among ancestors of two divergent clades (e.g., the Squash leaf curl virus [SLCV] clade and the Abutilon mosaic virus clade) of Western Hemisphere begomoviruses. The CLCrV DNA-B component also may have originated by recombination among an ancestor of the SLCV clade and another distantly related but unknown Western Hemisphere begomovirus.
The Bacillus Spore Coat
Authors: Driks Adam
Published: May 2007

Bacilli, which are abundant in the soil, form highly resistant dormant cell types, called spores, in response to starvation. The spore is organized into a series of concentrically arranged structures, each of which contribute in a different way to resistance against environmental stress. In certain bacteria, including Bacillus subtilis, the outermost of these structures is a multilayered protein shell, called the coat. The coat is both an armor plating and, almost certainly, possesses enzymatic activities, allowing it to have active roles as well. Assembly of the proteins comprising the coat is carefully controlled during spore assembly, resulting in a distinct pattern of layers, seen in cross section, and a discreet pattern of ridges on the surface. Although our understanding of spore coat composition and assembly is deepening, we still know little about the roles of the coat in interactions between spores and other organisms, particularly in the soil. Critical future directions for spore coat research include continued identification of the proteins that comprise the coat surface, characterization of the global chemical characteristics of this surface, and elucidation of how these features impact on other organisms in the soil.
Bioavailability of Iron to Pseudomonas fluorescens Strain A506 on Flowers of Pear and Apple
Authors: Temple Todd N. , Stockwell Virginia O. , Loper Joyce E. , Johnson Kenneth B.
Published: May 2007

The addition of 0.1 mM FeCl3 to a defined culture medium induces the bacterial epiphyte Pseudomonas fluorescens strain A506 (A506) to produce an antibiotic toxic to the fire blight pathogen, Erwinia amylovora. Consequently, because A506 is registered and applied as a commercial product to suppress E. amylovora before floral infection of pear and apple, the relative availability of iron to A506 on surfaces of pear and apple flowers is of potential significance. An ‘iron biosensor’ construct of A506 was developed by transformation with an iron-regulated promoter (pvd) fused to a promoterless ice nucleation reporter gene (inaZ). This construct, A506 (pvd-inaZ), established high populations on pear and apple flowers, ranging from 104 to 106 CFU/flower. In seven trials on pear and apple trees, A506 (pvd-inaZ) expressed high ice nucleation activity (INA) on flowers, indicating limited iron bioavailability or a low-iron environment unlikely to induce antibiotic production by A506. A506 (pvd-inaZ) also colonized flowers when mixed with chemicals containing iron: FeSO4 or the iron chelates ferric ethylenediaminedi-(o-hydroxyphenyl-acetic) acid (FeEDDHA) and ferric diethylenetriamine pentaacetate (FeDTPA). These compounds represent an array of commercial iron formulations applied to foliage to avert iron chlorosis. Treatment of flowers with a mixture of A506 (pvd-inaZ) and 3 mM FeEDDHA or FeDTPA significantly decreased INA compared with flowers treated with A506 (pvd-inaZ) in water. Lower concentrations (0.3 mM) of FeEDDHA, however, did not consistently suppress INA. These results indicate that apple and pear flowers represent an iron-limited environment to A506 and that treatment with 3 mM FeEDDHA is needed to increase significantly the level of iron available to this bacterium.
Effect of Timing of Application of the Biological Control Agent Microsphaeropsis ochracea on the Production and Ejection Pattern of Ascospores by Venturia inaequalis
Authors: Carisse O. , Rolland D.
Published: May 2007

Field and in vitro trials were conducted to establish the influence of the biological control agent Microsphaeropsis ochracea on the ejection pattern of ascospores by Venturia inaequalis and on apple scab development, and to establish the best timing of application. The ejection pattern of ascospores was similar on leaves sprayed with M. ochracea and on untreated leaves. Fall application of M. ochracea combined with a delayed-fungicide program was evaluated in orchards with intermediate and high scab risk. For both orchards, it was possible to delay the first three and two infection periods in 1998 and 1999, respectively, without causing significant increase or unacceptable leaf and fruit scab incidence. To evaluate the best timing of application, sterile leaf disks were inoculated with V. inaequalis and then with M. ochracea 0, 2, 4, 6, 8, 10, 12, 14, and 16 weeks later. After incubation under optimal conditions for pseudothecia development, the number of ascospores was counted. Similarly, M. ochracea was sprayed on scabbed leaves on seven occasions from August to November 1999 and 2000. Leaves were overwintered on the orchard floor and ascospore production was evaluated the following spring. Ascospore production was reduced by 97 to 100% on leaf disks inoculated with M. ochracea less than 6 weeks after inoculation with V. inaequalis, but ascospore production increased with increasing period of time when M. ochracea was applied 8 to 16 weeks after the inoculation with V. inaequalis. In the orchard, the greatest reduction in production of ascospores (94 to 96% in 2000 and 99% in 2001) occurred on leaves sprayed with M. ochracea in August. The production of ascospores was reduced by 61 to 84% in 2000 and 93% in 2001 on leaves sprayed with M. ochracea in September, reduced by 64 to 86% in 2000 and 74 to 89% in 2001 on leaves sprayed in October, and reduced by 54 and 67% in 2000 and 2001, respectively, on leaves sprayed in November. It was concluded that M. ochracea should be applied in August or September and that ascospore maturation models and delayed-fungicide program could be used in orchards treated with this biological control agent.
Dual Activity of a Viral Lysozyme with High Efficiency for Growth Inhibition of Erwinia amylovora
Authors: Salm Heike , Geider Klaus
Published: May 2007

The lysozyme from Erwinia amylovora phage ΦEa1h was investigated for its ability to inhibit growth of bacteria and compared with the lysozyme from Escherichia coli phage T4. The assays to measure lysozyme activity included cell lysis and growth inhibition of bacteria. Bacterial strains with kanamycin resistance were not affected by lysates containing the ΦEa1h-enzyme. The titer of Micrococcus luteus but not of Erwinia amylovora was diminished by cell extracts containing T4 lysozyme. In contrast, ΦEa1h lysozyme preferentially inhibited E. amylovora, exceeding the T4 lysozyme activity at least one million-fold. Spherical cells were formed after application to E. amylovora similar to lyz-gene expression in Escherichia coli. Heating of cell extracts destroyed the murami-dase activity, but retained an antibacterial activity. Other plant-associated bacteria related to Erwinia amylovora also were inhibited for growth when cell extracts with ΦEa1h lysozyme were applied to soak pear slices and potato slices. Ooze formation and soft rot caused by E. amylovora or E. carotovora subsp. atroseptica, respectively, were strongly reduced and the ΦEa1h lysozyme was more efficient compared with extracts containing T4 lysozyme.
Polyphasic Characterization of Xanthomonas Strains from Onion
Authors: Gent David H. , Schwartz Howard F. , Ishimaru Carol A. , Louws Frank J. , Cramer Robert A. , Lawrence Christopher B.
Published: May 2007

Xanthomonas leaf blight has become an increasingly important disease of onion, but the diversity among Xanthomonas strains isolated from onion is unknown, as is their relationship to other species and pathovars of Xanthomonas. Forty-nine Xanthomonas strains isolated from onion over 27 years from 10 diverse geographic regions were characterized by pathogenicity to onion and dry bean, fatty acid profiles, substrate utilization patterns (Biolog), bactericide resistance, repetitive sequence-based polymerase chain reaction fingerprinting, rDNA internally transcribed spacer (ITS) region, and hrp b6 gene sequencing. Multiplication of onion Xanthomonas strain R-O177 was not different from X. axonopodis pv. phaseoli in dry bean, but typical common bacterial blight disease symptoms were absent in dry bean. Populations from each geographical region were uniformly sensitive to 100 μg of CuSO4, 100 μg of ZnSO4, and 100 μg of streptomycin sulfate per ml. Biolog substrate utilization and fatty acid profiles revealed close phenoltypic relatedness between onion strains of Xanthomonas and X. axonopodis pv. dieffenbachiae (57% of strains) and X. arboricola pv. poinsettiicola (37% of strains), respectively. A logistic regression model based on fatty acid composition and substrate utilization classified 69% of strains into their geographical region of origin. Sequencing of a portion of the hrp B6 gene from 24 strains and ITS region from 25 strains revealed greater than 97% sequence similarity among strains. DNA fingerprinting revealed five genotype groups within onion strains of Xanthomonas and a high degree of genetic diversity among geographical regions of origin. Based on pathogenicity to onion, carbon substrate utilization, fatty acid profiles, rDNA genetic diversity, and genomic fingerprints, we conclude that the strains examined in this study are pathovar X. axonopodis pv. allii. Implications of genetic and phenotypic diversity within X. axonopodis pv. allii are discussed in relation to an integrated pest management program.
Recovery in Apple Trees Infected with the Apple Proliferation Phytoplasma: An Ultrastructural and Biochemical Study
Authors: Musetti R. , di Toppi L. Sanità , Ermacora P. , Favali M. A.
Published: May 2007

Localization of hydrogen peroxide (H2O2) and the roles of peroxidases, malondialdehyde, and reduced glutathione in three apple cultivars were compared in healthy trees, trees infected with apple proliferation phytoplasma (APP), and trees that had recovered from the infection. In recovered apple trees, symptoms of the disease and the pathogen had disappeared from the canopy, but phytoplasmas remained in the roots. H2O2 was detected cytochemically by its reaction with cerium chloride to produce electron-dense deposits of cerium perhydroxides.H2O2 occurred in the plasmalemma of the phloem of leaves of recovered apple trees, but not in healthy or APP-infected leaves. In all cultivars, the peroxidase activity detected in tissue from APP-diseased trees was greater than or equal to that of tissue from recovered trees, which equaled or exceeded that of tissue from healthy trees, at two sampling times (May and September). In contrast, the glutathione content of leaves decreased in the reverse order. More malondialdehyde was observed in leaves from recovered trees than in leaves from healthy or APP-infected trees in three of six cultivar-date combinations; in the other three combinations, the malondialdehyde contents of leaves from healthy, infected, and recovered trees were not significantly different from one another. The results suggest that some components of the oxidant-scavenging system in recovered leaves are not very active, leading to an overproduction of H2O2 and, possibly, to a membrane lipid peroxidation.The production of H2O2 appears to be involved in counteracting pathogen virulence.
Canopy Size and Induced Resistance in Stylosanthes scabra Determine Anthracnose Severity at High CO2
Authors: Pangga I. B. , Chakraborty S. , Yates D.
Published: May 2007

This study examines the relative importance of canopy size and induced resistance to Colletotrichum gloeosporioides at 350- and 700-ppm atmospheric CO2 concentrations on susceptible Stylosanthes scabra ‘Fitzroy’ from two studies in a controlled environment facility (CEF) and in the field. Plants were grown at the two CO2 concentrations in a repeated experiment in the CEF and inoculated at 6, 9, or 12 weeks of age. Although the physiological maturity of plants was at a similar stage for all three ages, the number of lesions per plant increased with increasing plant age at both CO2 concentrations. At 350 ppm, the increase was associated with canopy size and increasing infection efficiency of the pathogen, but at 700 ppm, it was associated only with canopy size, because infection efficiency did not change with increasing age. A level of resistance was induced in plants at 700 ppm CO2. In a second study, plants were raised for 12 to 14 weeks at the two CO2 concentrations in the CEF and exposed to C. gloeosporioides inoculum in replicated field plots under ambient CO2 over three successive years. Fitzroy developed a dense and enlarged canopy, with 28 to 46% more nodes, leaf area, and aboveground biomass at high CO2 than at low CO2. Up to twice as many lesions per plant were produced in the high CO2 plants, because the enlarged canopy trapped many more pathogen spores. The transient induced resistance in high CO2 plants failed to operate when exposed to pathogen inoculum under ambient CO2 in the field. These results highlight the need to consider both canopy size and host resistance in assessing the influence of elevated CO2 on plant disease.