bluuebunny/crossref_metadata_2025 huggingface dataset:
Antagonism of the Two-Needle Pine Stem Rust FungiCronartium flaccidumandPeridermium pinibyCladosporium tenuissimumIn Vitro and In Planta
Authors: Moricca Salvatore , Ragazzi Alessandro , Mitchelson Keith Richard , Assante Gemma
Published: May 2007

Selected isolates of Cladosporium tenuissimum were tested for their ability to inhibit in vitro aeciospore germination of the two-needle pine stem rusts Cronartium flaccidum and Peridermium pini and to suppress disease development in planta. The antagonistic fungus displayed a number of disease-suppressive mechanisms. Aeciospore germination on water agar slides was reduced at 12, 18, and 24 h when a conidial suspension (1.5 × 107conidia per ml) of the Cladosporium tenuissimum isolates was added. When the aeciospores were incubated in same-strength conidial suspensions for 1, 11, 21, and 31 days, viability was reduced at 20 and 4°C. Light and scanning electron microscopy showed that rust spores were directly parasitized by Cladosporium tenuissimum and that the antagonist had evolved several strategies to breach the spore wall and gain access to the underlying tissues. Penetration occurred with or without appressoria. The hyperparasite exerted a mechanical force to destroy the spore structures (spinules, cell wall) by direct contact, penetrated the aeciospores and subsequently proliferated within them. However, an enzymatic action could also be involved. This was shown by the dissolution of the host cell wall that comes in contact with the mycelium of the mycoparasite, by the lack of indentation in the host wall at the contact site, and by the minimal swelling at the infecting hyphal tip. Culture filtrates of the hyperparasite inhibited germination of rust propagules. A compound purified from the filtrates was characterized by chemical and spectroscopic analysis as cladosporol, a known β-1,3-glucan biosynthesis inhibitor. Conidia of Cladosporium tenuissimum reduced rust development on new infected pine seedlings over 2 years under greenhouse conditions. Because the fungus is an aggressive mycoparasite, produces fungicidal metabolites, and can survive and multiply in forest ecosystems without rusts, it seems a promising agent for the biological control of pine stem rusts in Europe.
Intrapathotype Diversity for Aggressiveness and Pathogen Evolution in Cultivar Mixtures
Authors: Lannou Christian
Published: May 2007

A model was developed and used to study the consequences of diversity for aggressiveness within pathotypes on pathogen evolution in two-component and four-component cultivar mixtures. It was assumed that, within a pathotype, a proportion of the isolates would have higher or lower spore efficacy than the average on a given host genetic background. Two situations were examined in which the pathogen can have either independent or negatively correlated values for spore efficacy on different cultivars. In the latter case, a pathogen genotype more aggressive than the average on a host genotype was always less aggressive on other host genotypes. In the simulations, isolates with greater aggressiveness relative to a host genotype were selected for and increased in frequency. However, because simple pathotypes always reproduced on the same host genotype whereas complex pathotypes were able to grow on several hosts, selection was faster for simple pathotypes. Pathotypes with two different levels of diversity for aggressiveness were compared with nondiversified pathotypes. In order to make comparisons, the effect of a 5 and 10% cost of virulence on the development of complex pathotypes was simulated. In general, increased diversity within pathotypes reduced the rate of increase of complex pathotypes in host mixtures, and this effect was stronger with greater frequencies of autodeposition of pathogen spores.
Restriction Fragment Length Polymorphism Mapping of Resistance to Two Races of Pyrenophora tritici-repentis in Adult and Seedling Wheat
Authors: Effertz R. J. , Anderson J. A. , Francl L. J.
Published: May 2007

Resistance to the chlorosis factor of tan spot of wheat, caused by the ascomycete Pyrenophora tritici-repentis, has been reported to be quantitative and a single quantitative trait loci (QTL), QTsc.ndsu-1A, explained 35% of the variation for resistance to a single isolate in seedlings of recombinant inbred (RI) lines derived from the cross W-7984/Opata 85. The objectives of this study were to determine the number and locations of genes conditioning resistance to the same isolate in adult plants of this population and three isolates in seedlings of wheat RI lines derived from the cross W-7976/Trenton. An extensive restriction fragment length polymorphism map exists for the W-7984/Opata 85 population, and markers significantly associated (P < 0.01) with resistance to tan spot were selected to analyze the W-7976/Trenton population. A multiple regression model accounted for 49% of the variation for resistance in adult plants with QTsc.ndsu-1A, explaining 26% of the variation. QTsc.ndsu-1A explained 47, 58, and 64% of the variation for resistance in seedlings to isolates Pti2, 78-62, and D308, respectively. These results showed that the QTL for tan spot resistance on chromosome 1AS was effective in both seedlings and adult plants and against isolates from different races of P. tritici-repentis.
Genetic Diversity WithinColletotrichum acutatum sensuSimmonds
Authors: Freeman Stanley , Minz Dror , Maymon Marcel , Zveibil Aida
Published: May 2007

Isolates of Colletotrichum acutatum from several hosts were characterized by various molecular methods in comparison with morphological identification. Species-specific primer analysis was reliable for grouping C. acutatum isolates to their designated species. Arbitrarily primed polymerase chain reaction and A+T-rich DNA analyses identified four subgroups within C. acutatum. Subgroup I contained U.S. isolates from almond, apple, peach, and pecan, subgroup II contained isolates from anemone, olive, and strawberry, subgroup III contained isolates from almond (Israel) and strawberry (Spain), and subgroup IV contained a single isolate from anemone (the Netherlands). Likewise, sequence analysis of the internal transcribed spacer (ITS) 2 region alone or the complete ITS (ITS 1–5.8S-ITS 2) region grouped the isolates into the same four subgroups. Percent similarity of the complete ITS region within each cluster ranged from 99.6 to 100.0, 99.8 to 100.0, and 98.6% among subgroups I, II, and III, respectively. DNA sequence analysis of the ITS 2 region alone or the entire ITS 1-2 region was more informative than that of the ITS 1 region, which could only group the isolates into two main clusters. The molecular methods employed for studying genetic variation in populations of C. acutatum determined that this species is diverse, indicating that isolates within populations of each subgroup are not host specific.
Population Structure of Botryosphaeria dothidea from Pistachio and Other Hosts in California
Authors: Ma Zhonghua , Boehm Eric W. A. , Luo Yong , Michailides Themis J.
Published: May 2007

Genetic diversity was investigated among California populations of Botryosphaeria dothidea, causal agent of Botryosphaeria panicle and shoot blight of pistachio, with random amplified polymorphic DNA (RAPD) and microsatellite-primed polymerase chain reaction (MP-PCR). We surveyed 120 isolates, 112 of which originated from the California Central Valley and included pistachio isolates (n = 52) and isolates from other plant species (n = 60). Out-group isolates (n = 8) were obtained from pistachio in Greece. There was a strong correlation (r = 0.99; P < 0.0001) between the RAPD- and MP-PCR dissimilarity data sets. Little genetic variation (haplotypic diversity [Hs] < 0.002) was detected among B. dothidea isolates collected from central and southern California pistachio plantings. We observed relatively high diversity for isolates from a northern California pistachio orchard (Hs = 0.0146), where the disease was first diagnosed, and from the Chico U.S. Department of Agriculture Germ Plasm Repository (Hs = 0.0726), where the first pistachio trees were planted in California in 1929. Isolates obtained from other hosts, especially those associated with the rare occurrence of the sexual stage of this fungus, showed the highest levels of diversity (Hs = 0.1689). Thirty-eight pistachio isolates (73.1%) had DNA fingerprints identical to 28 pycnidiospore-derived isolates (56.0%) obtained from other host species. Greenhouse inoculations demonstrated that all isolates obtained from other hosts were capable of infecting pistachio and produced characteristic disease symptomology. Thus, California populations of B. dothidea from pistachio are, for the most part, genetically uniform, with the sexual stage rare to absent. However, the rare occurrence of the sexual stage of B. dothidea on other hosts, and more importantly, the capacity of these isolates to infect pistachio, indicate that other host species may serve as sources of inoculum and genetic variation.
Suppression of Specific Apple Root Pathogens by Brassica napus Seed Meal Amendment Regardless of Glucosinolate Content
Authors: Mazzola Mark , Granatstein David M. , Elfving Donald C. , Mullinix Kent
Published: May 2007

The impact of Brassica napus seed meal on the microbial complex that incites apple replant disease was evaluated in greenhouse trials. Regardless of glucosinolate content, seed meal amendment at a rate of 0.1% (vol/vol) significantly enhanced growth of apple and suppressed apple root infection by Rhizoctonia spp. and Pratylenchus penetrans. High glucosinolate B. napus cv. Dwarf Essex seed meal amendments did not consistently suppress soil populations of Pythium spp. or apple root infection by this pathogen. Application of a low glucosinolate containing B. napus seed meal at a rate of 1.0% (vol/vol) resulted in a significant increase in recovery of Pythium spp. from apple roots, and a corresponding reduction in apple seedling root biomass. When applied at lower rates, B. napus seed meal amendments enhanced populations of fluorescent Pseudomonas spp., but these bacteria were not recovered from soils amended with seed meal at a rate of 2% (vol/vol). Seed meal amendments resulted in increased soil populations of total bacteria and actinomycetes. B. napus cv. Dwarf Essex seed meal amendments were phytotoxic to apple when applied at a rate of 2% (vol/vol), and phytotoxicity was not diminished when planting was delayed for as long as 12 weeks after application. These findings suggest that B. napus seed meal amendments can be a useful tool in the management of apple replant disease and, in the case of Rhizoctonia spp., that disease control operates through mechanisms other than production of glucosinolate hydrolysis products.
Small-Scale Isolation of Viral RNA-Dependent RNA Polymerase from Protoplasts Inoculated with In Vitro Transcripts
Authors: Adkins Scott , Lewandowski Dennis J.
Published: May 2007

Cowpea chlorotic mottle virus (CCMV) replicated in tobacco suspension cell protoplasts inoculated with in vitro transcripts of CCMV RNA1, 2, and 3. CCMV RNA-dependent RNA polymerase (RdRp) isolated from these protoplasts specifically recognized CCMV and Brome mosaic virus (BMV) subgenomic RNA promoters and directed in vitro RNA synthesis in a manner indistinguishable from CCMV RdRp more laboriously isolated from systemically infected cowpea leaves. Omission of CCMV RNA3 from the protoplast inoculum or replacement with in vitro transcripts of BMV RNA3 reduced CCMV (+)-strand RNA1 and 2 accumulation to ≈1/40 and ≈1/10, respectively, of the level attained when CCMV RNA3 was present. The absence of CCMV RNA3 did not prevent assembly and isolation of highly active, template-dependent and template-specific CCMV RdRp, which directed synthesis of products identical in size to those of RdRp isolated from protoplasts inoculated with all three CCMV genomic RNAs. These results demonstrate that CCMV RNA1 and 2 are sufficient for CCMV replication and RdRp assembly in tobacco protoplasts. This approach for isolation of functional viral RdRp will be especially useful for viruses for which large quantities of infected tissue are unavailable, such as those with specific tissue tropisms or mutants incapable of systemic movement.
The Effects of Founding Events and Agricultural Practices on the Genetic Structure of Three Metapopulations of Globodera pallida
Authors: Folkertsma Rolf T. , van Koert Paul , van der Voort Jeroen N. A. M. Rouppe , de Groot Koen E. , Kammenga Jan E. , Helder Johannes , Bakker Jaap
Published: May 2007

To assess the genetic constitution of the Globodera pallida populations in the Netherlands and the effects of agricultural practices, three geographically separated metapopulations, in total consisting of 226 local populations, were analyzed by two-dimensional gel electrophoresis (2-DGE) of total proteins. This technique allows the accurate assessment of allele frequencies in homogenates of mixtures of individuals. Based on the estimated average heterozygosity, the average proportion of polymorphic loci and the average number of alleles per locus, the genetic diversity among 226 local G. pallida populations was small. The small genetic basis of G. pallida in the Netherlands will facilitate the identification of Solanum genotypes with a broad and durable resistance to G. pallida. Instead of clusters of local populations with unique alleles, a continuous range of allele frequencies was observed. Analysis of the three metapopulations by the Shannon-Weaver index and Nei's GST revealed that the metapopulation from a region with sandy-loam soils was clearly distinguishable from the remaining two; the local populations within this metapopulation were more similar and the genetic diversity within the individual local populations was significantly higher than the local populations from the two remaining regions. These regions are characterized by wider crop rotation schemes and a very limited use of nematicides. The less intensive cultivation of potatoes in these regions with sandy-clay soils resulted in relatively little variation within and more variation between local nematode populations. To our knowledge, the effects of agricultural practices on the genetic constitution of potato cyst nematode populations have not been pinpointed before.
Reaction of Ptr ToxA-Insensitive Wheat Mutants to Pyrenophora tritici-repentis Race 1
Authors: Friesen T. L. , Rasmussen J. B. , Kwon C. Y. , Francl L. J. , Meinhardt S. W.
Published: May 2007

The host-selective toxin Ptr ToxA is produced by races 1 and 2 of Pyrenophora tritici-repentis, causal agent of tan spot of wheat. Ptr ToxA has been causally associated with pathogenicity by the race 2 phenotype in this system. However, the role of toxin in disease caused by race 1, the most prevalent form of the fungus in the central and northern Great Plains of North America, has not been rigorously investigated. Three independent wheat lines harboring mutations for insensitivity to Ptr ToxA were derived from ethylmethane sulfonate treatment of the hard red spring wheat cv. Kulm, possessing the single dominant gene for toxin sensitivity. Each of the three mutants was insensitive to Ptr ToxA in bioassays based on necrosis development and electrolyte leakage. Each mutant was crossed to each of the other mutants and to the wild-type Kulm. Segregation data indicate that each mutant line harbors a single recessive mutation for toxin insensitivity that maps to or near the same locus, possibly the toxin-sensitivity gene. Each toxin-insensitive mutant line was susceptible to two isolates of race 1 of P. tritici-repentis. F2 and F3 generations derived from crosses between Kulm and each mutant segregated for toxin reaction. However, segregation for fungal reaction was not evident, and all F3 families were tan spot susceptible regardless of toxin reaction. Host insensitivity to Ptr ToxA is not necessarily equivalent to resistance to race 1. Ptr ToxA should not be used alone as a proxy for fungal inoculations by breeding programs aimed at developing tan spot-resistant wheat.
Hyphal Anastomosis Reactions, rDNA-Internal Transcribed Spacer Sequences, and Virulence Levels Among Subsets of Rhizoctonia solani Anastomosis Group-2 (AG-2) and AG-BI
Authors: Carling D. E. , Kuninaga S. , Brainard K. A.
Published: May 2007

Hyphal anastomosis reactions, rDNA-internal transcribed spacer (ITS) sequences, and virulence of isolates representing Rhizoctonia solani AG-BI and six subsets of anastomosis group (AG)-2 (-2-1, -2-2 IIIB, -2-2 IV, -2-2 LP, -2-3, and -2-4) were compared. AG-2-4 is a subset described for the first time in this report. Anastomosis reactions within AG-BI and the listed subsets of AG-2 were generally strong but, between subsets, ranged from strong to a very weak “bridging” -type reaction. Anastomosis reaction alone generally did not provide adequate evidence for placement of an isolate into a subset of AG-2. Anastomosis reactions between AG-BI and the original subsets of AG-2 (-2-1 and -2-2) are very strong; for this reason, we propose that it be included as a subset of AG-2 (designation AG-2 BI). Subsets -2-3 and -2-4 show very weak bridging-type anastomosis reactions with all other subsets of AG-2 and thus may be candidates for independent AG status. Grouping within AG-2 based on rDNA-ITS sequences was consistent with the abovementioned subsets. However, grouping based on virulence as measured herein does not conform to established grouping patterns within AG-2 and does not seem useful as a group-defining criterion. A broad range of damage was observed among members of the most virulent subsets (-2-1, -2-2 IIIB, -2-2 IV, and -2-4), whereas other subsets (-2 BI, -2-2 LP, and -2-3) were similar to one another in causing a minimal level of damage. Group-specific primer pairs for each of the seven subsets of AG-2 were designed based on the abovementioned rDNA-ITS sequences. Primer pairs proved dependable and subset specific in polymerase chain reaction amplifications of purified genomic DNA from 109 isolates of R. solani and two isolates of binucleate Rhizoctonia. These primers will provide a simple and useful method for subset-specific characterization within AG-2 if further critical evaluations confirm their specificity.
Spatial and Temporal Distribution of Geminiviruses in Leafhoppers of the Genus Cicadulina Monitored by Conventional and Quantitative Polymerase Chain Reaction
Authors: Lett Jean-Michel , Granier Martine , Hippolyte Isabelle , Grondin Martial , Royer Monique , Blanc Stéphane , Reynaud Bernard , Peterschmitt Michel
Published: May 2007

Spatial and temporal distribution of Maize streak virus (MSV, family Geminiviridae, genus Mastrevirus) was monitored in the vector species Cicadulina mbila and the nonvector species C. chinaï using conventional and real-time quantitative polymerase chain reaction. Sustained feeding on MSV-infected plants showed that virus accumulation reaches a maximum in C. chinaï, but not in C. mbila. After a 3-day acquisition access feeding period (AAP), MSV was detected in the gut, the hemolymph, and the head of C. mbila, but only in the gut of C. chinaï. Similarly, Digitaria streak virus (genus Mastrevirus), which is not transmitted by either of the two species, was only detected in the gut. MSV was detected in the hemolymph of C. mbila 3 h after the beginning of the AAP. Although viral DNA progressively decreases in the vector and nonvector species after a 3-day AAP, MSV DNA remained stable in the salivary glands of C. mbila.
Effect of Relative Humidity and Interrupted Wetness Periods on Brown Spot Severity of Pear Caused by Stemphylium vesicarium
Authors: Llorente I. , Montesinos E.
Published: May 2007

Field observations in four pear orchards during 5 years from April to October indicated that days with uninterrupted wetness of variable length represented 83.9% of the total days studied. However, days with surface wetness interruptions and with high relative humidity (RH) (≥90%) without wetness occurred with a frequency of 7.1 and 6.2%, respectively. Accordingly, the effect of interruption of 24-h wetness periods by dry periods of high or low RH on infections caused by Stemphylium vesicarium on pear was determined. Pear plants inoculated with conidia of S. vesicarium were exposed to a 12-h wet period followed by a dry period of variable length (0, 3, 6, 12, 18, or 24 h) and a second wet period of 12 h. The dry period consisted either of low (60%) or high (96%) RH. The infection process was irreversibly stopped under low RH during dry periods between wetness, but continued at high RH. The effect of high RH on disease severity in the absence of wetness was also determined. Pear plants inoculated with S. vesicarium were exposed to periods of variable length (3 to 24 h) either at high RH (96%) in the presence of wetness or at high RH (96%) without wetness. No infections were observed on plants incubated under high RH without wetness, indicating that conidia of S. vesicarium require the presence of a water film in the plant surface to develop infections on pear.
Molecular Identification of a New Phytoplasma Associated with Alfalfa Witches'-Broom in Oman
Authors: Khan A. J. , Botti S. , Al-Subhi A. M. , Gundersen-Rindal D. E. , Bertaccini A. F.
Published: May 2007

Alfalfa (Medicago sativa) plants showing witches'-broom symptoms typical of phytoplasmas were observed from Al-Batinah, Al-Sharqiya, Al-Bureimi, and interior regions of the Sultanate of Oman. Phytoplasmas were detected from all symptomatic samples by the specific amplification of their 16S–23S rRNA gene. Polymerase chain reaction (PCR), utilizing phytoplasma-specific universal primer pairs, consistently amplified a product of expected lengths when DNA extract from symptomatic samples was used as template. Asymptomatic plant samples and the negative control yielded no amplification. Restriction fragment length polymorphism profiles of PCR-amplified 16S–23S rDNA of alfalfa using the P1/P7 primer pair identified phytoplasmas belonging to peanut witches'-broom group (16SrII or faba bean phyllody). Restriction enzyme profiles showed that the phytoplasmas detected in all 300 samples belonged to the same ribosomal group. Extensive comparative analyses on P1/P7 amplimers of 20 phytoplasmas with Tru9I, Tsp509I, HpaII, TaqI, and RsaI clearly indicated that this phytoplasma is different from all the other phytoplasmas employed belonging to subgroup 16SrII, except tomato big bud phytoplasma from Australia, and could be therefore classified in subgroup 16SrII-D. The alfalfa witches'-broom (AlfWB) phytoplasma P1/P7 PCR product was sequenced directly after cloning and yielded a 1,690-bp product. The homology search showed 99% similarity (1,667 of 1,690 base identity) with papaya yellow crinkle (PapayaYC) phytoplasma from New Zealand. A phylogenetic tree based on 16S plus spacer regions sequences of 35 phytoplasmas, mainly from the Southern Hemisphere, showed that AlfWB is a new phytoplasma species, with closest relationships to PapayaYC phytoplasmas from New Zealand and Chinese pigeon pea witches'-broom phytoplasmas from Taiwan but distinguishable from them considering the different associated plant hosts and the extreme geographical isolation.
Relationships Between Blueberry Flower Age, Pollination, and Conidial Infection byMonilinia vaccinii-corymbosi
Authors: Ngugi H. K. , Scherm H. , Lehman J. S.
Published: May 2007

Monilinia vaccinii-corymbosi infects open blueberry flowers via the gynoecial pathway, leading to mummification of the developing fruit. To determine the effect of flower age on infection, stigmata were inoculated with conidia of M. vaccinii-corymbosi between 0 and 5 days after anthesis, fungal growth rates through the stylar canal were measured in detached flowers in the laboratory, and fruit disease incidence was determined in plants grown in the greenhouse. Hyphal growth rates were greatest in flowers inoculated on the day of anthesis, declined linearly with increasing flower age at inoculation (r = 0.921; P < 0.0001; n = 12), and were unaffected by the presence or absence of pollen applied at the time of inoculation. In greenhouse-grown plants, the percentage of infected fruit decreased exponentially with increasing flower age at inoculation (R = 0.878; P = 0.0057; n = 10), with disease incidence ranging from 76.4% for flowers inoculated on the day of anthesis to 15.5% for those inoculated 4 days later. Fruit disease incidence in the greenhouse was linearly correlated with hyphal growth rates in detached flowers (r = 0.985; P < 0.0001; n = 9), justifying the use of detached flowers when investigating gynoecial infection by M. vaccinii-corymbosi. In separate experiments, the effects of timing and sequence of pollination and inoculation on hyphal growth rates through the stylar canal and on disease incidence were investigated. Application of pollen to detached flowers 1 or 2 days before inoculation reduced hyphal growth rates by between 14.0 and 42.9% compared with flowers that received pollen and conidia simultaneously. Similarly, reductions in fruit disease incidence by between 9.5 and 18.3% were observed on greenhouse-grown plants for pollination-to-inoculation intervals ranging from 1 to 4 days. These results document that newly opened flowers are most susceptible to infection by M. vaccinii-corymbosi and that fruit disease incidence is reduced if pollination occurs at least 1 day before inoculation. Strategies that lead to early pollination of newly opened flowers may be useful for managing mummy berry disease in the field.
Phytophthora infestans Populations from Tomato and Potato in North Carolina Differ in Genetic Diversity and Structure
Authors: Wangsomboondee T. , Groves C. Trout , Shoemaker P. B. , Cubeta M. A. , Ristaino J. B.
Published: May 2007

Phytophthora infestans causes a destructive disease on tomato and potato. In North Carolina (NC) potatoes are mostly grown in the east, whereas tomatoes are grown in the mountainous areas in the western part of the state. Five genotypes of P. infestans were identified from 93 and 157 isolates collected from tomato and potato over a 5 year period between 1993 and 1998. All isolates collected from potato in eastern NC were the US-8 genotype, whereas only a single isolate was the US-1 genotype. Tuber blight was found on immature daughter tubers in a single field in 1997, however infection on mature tubers was not observed. Within potato fields, a range of sensitivity to metalaxyl was observed among isolates but all were either intermediate or highly resistant to the fungicide. In contrast, isolates from tomatoes included previously reported US-7 and US-8 genotypes and two new genotypes called US-18 and US-19 (A2 mating type, allozyme genotype Gpi 100/100 and Pep 92/100). These genotypes had unique restriction fragment length polymorphism banding patterns, were sensitive to metalaxyl, and have not been reported elsewhere. All genotypes, with the exception of the US-1, were the Ia mitochondrial haplotype. Thus, isolates of P. infestans from tomato were more genetically diverse over time in NC than those from potato and include two new genotypes that are sensitive to metalaxyl.
Occurrence and Identification of Phytophthora spp. Pathogenic to Pear Fruit in Irrigation Water in the Wenatchee River Valley of Washington State
Authors: Yamak F. , Peever T. L. , Grove G. G. , Boal R. J.
Published: May 2007

Seven hundred forty-nine isolates of Phytophthora spp. were obtained from irrigation canals in eastern Washington State during the 1992 to 1995 and 1999 growing seasons. Isolates were retrieved using pear baiting techniques. All isolates were pathogenic to pear and were present in irrigation water beginning early in fruit development. Over the course of the 5 year study, 10 and 5% of isolates were identified as P. cactorum and P. citricola, respectively, using morphological criteria. The remaining isolates could not be identified using morphological criteria. Colony morphology of these isolates was characterized during all years of the study. In 1999, more detailed studies utilizing polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis of entire internal transcribed spacer (ITS) regions (ITS1, 5.8S, and ITS2) of ribosomal DNA for 180 isolates, and sequence analysis of ITS2 for 50 isolates, were used to investigate genetic variation and phylogenetic relationships among isolates. Isolates were divided into 12 groups based on their growth type on corn meal agar. Restriction digestion of the entire ITS region with three enzymes revealed 11 restriction digestion patterns among 180 isolates. PCR-RFLP and sequence data were obtained for 12 reference Phytophthora spp. (two species in each of Waterhouse's six morphological groups). Phylogenetic analysis of ITS2 regions revealed nine clades, each with strong bootstrap support. Molecular analyses revealed 23 isolates that were in the P. gonapodyides clade, 9 in the P. parasitica clade, 1 in the P. cactorum clade, 7 in the P. citricola/capsici clade, and 4 in the P. cambivora/pseudotsugae clade. The three isolates comprising clade 5 were significantly distinct from all other Phytophthora spp. in the databases and may represent a new Phytophthora sp. Colony morphology was not consistently correlated to PCR-RFLP pattern or ITS2 phylogeny, suggesting that the former criterion is insufficient for species identification. The results of this study indicate that at least nine phylogenetically distinct taxa of Phytophthora pathogenic to pear are present in irrigation water in North Central Washington.
Etiology and Population Genetics of Colletotrichum spp. Causing Crown and Fruit Rot of Strawberry
Authors: Ureña-Padilla A. R. , MacKenzie S. J. , Bowen B. W. , Legard D. E.
Published: May 2007

Isolates of Colletotrichum spp. from diseased strawberry fruit and crowns were evaluated to determine their genetic diversity and the etiology of the diseases. Isolates were identified to species using polymerase chain reaction primers for a ribosomal internal transcribed spacer region and their pathogenicity was evaluated in bioassays. Isolates were scored for variation at 40 putative genetic loci with random amplified polymorphic DNA and microsatellite markers. Only C. acutatum was recovered from diseased fruit. Nearly all isolates from crowns were C. gloeosporioides. In crown bioassays, only isolates of C. gloeosporioides from strawberry caused collapse and death of plants. A dendrogram generated from the genetic analysis identified several primary lineages. One lineage included isolates of C. acutatum from fruit and was characterized by low diversity. Another lineage included isolates of C. gloeosporioides from crowns and was highly polymorphic. The isolates from strawberry formed distinctive clusters separate from citrus isolates. Evaluation of linkage disequilibrium among polymorphic loci in isolates of C. gloeosporioides from crowns revealed a low level of disequilibrium as would be expected in sexually recombining populations. These results suggest that epidemics of crown rot are caused by Glomerella cingulata (anamorph C. gloeosporioides) and that epidemics of fruit rot are caused by C. acutatum.
Environmental Factors Affecting Pseudothecial Development and Ascospore Production of Mycosphaerella citri, the Cause of Citrus Greasy Spot
Authors: Mondal S. N. , Timmer L. W.
Published: May 2007

Mycosphaerella citri, the cause of citrus greasy spot, produces pseudothecia and ascospores in decomposing leaf litter on the grove floor. In laboratory studies, the effect of wetting and drying and temperature on the formation, maturation, and production of pseudothecia and ascospores was evaluated on mature, detached grapefruit leaves. Production of pseudothecia was most rapid when leaves were soaked five times per week for 2 h per day, but pseudothecial density and total ascospore production were greatest when leaves were soaked three times per week for 2 h per day. In duration of wetting studies, 3 h per day, 3 days per week brought about the most rapid production, but 10 to 30 min per day resulted in production of the most pseudothecia and ascospores. Pseudothecia and ascospore production were greatest at 28°C and declined rapidly at lower and higher temperatures. Maturation of pseudothecia was slow at 20 and 24°C, but production was high at 24°C; at 32°C, pseudothecia matured rapidly, but degenerated quickly. No mature pseudothecia were produced on leaves maintained continuously under wet conditions. In field studies, leaves were placed on the grove floor monthly from April 2000 to September 2001. Pseudothecia production was rapid during the summer rainy season from June to September. Pseudothecia produced on leaves placed in the grove from October to May developed and matured more slowly but were produced in much larger numbers than in summer. The number of days to first pseudothecial initials, 50% maturation, first discharge of ascospores, leaf decomposition, as well as pseudothecial density and incidence, were negatively related to average temperature. Total ascospore production was unrelated to temperature.
Population Analysis of Fusarium graminearum from Wheat Fields in Eastern China
Authors: Gale L. Rosewich , Chen L.-F. , Hernick C. A. , Takamura K. , Kistler H. C.
Published: May 2007

Wheat heads showing symptoms of Fusarium head blight were collected from four commercial fields in Zhejiang Province, China, an area where epidemics occur regularly. A total of 225 isolates were subjected to population-level analyses using restriction fragment length polymorphism (RFLP) as markers. Diagnostic RFLP markers established that all isolates belonged to Fusarium graminearum lineage 6. Nine polymorphic probes were hybridized to all isolates, resulting in 65 multilocus RFLP haplotypes (MRH). Probing with the telomeric clone pNla17, which reveals differences among isolates in the hypervariable subtelomeric region, differentiated the 65 MRH further into 144 clones. Mean gene diversity for the four field populations was similar, ranging from H = 0.306 - 0.364 over the nine RFLP loci for clone-corrected data. High levels of gene flow were inferred from a low level of population subdivision among all field populations, indicating that they were part of the same population. Pairwise linkage disequilibrium measures did not unequivocally support a random mating population, because one-third of locus pairs were significantly different from the null hypothesis of no-association between alleles. We speculate therefore that sexual recombination may not be frequent and that high levels of genotypic diversity may be maintained by relatively low selection pressure acting on a highly diverse population.
Status of Chemical Alternatives to Methyl Bromide for Pre-Plant Fumigation of Soil
Authors: Duniway J. M.
Published: May 2007

None of the chemical alternatives currently registered and available has the full spectrum of activity and versatility of methyl bromide as a pre-plant soil fumigant. Chloropicrin and 1,3-dichloropropene (Telone) can provide significant control of many plant pathogens in soil and growth stimulation in annual crops. These compounds, however, provide limited control of weeds or other residual plant materials in soil of concern in nursery production systems, and some perennial replant diseases. Methyl isothiocyanate generators such as metam sodium have broad biocidal activity in soil, but are more difficult to apply effectively. In most soil applications, the available alternatives are likely to be used in combinations, either as mixtures (e.g., 1,3-dichloropropene and chloropicrin) or sequentially (e.g., chloropicrin followed by metam sodium). They may also be supplemented with other more specific pesticides and cultural controls. Among the alternatives currently under active development but not yet available, methyl iodide and propargyl bromide probably have activity that most closely parallels that of methyl bromide in soil. However, all of the chemical alternatives to methyl bromide will be subject to continuing review and more regulation. Furthermore, we do not know the actual prospects for registration of the new fumigants currently under development and there is a risk that registered fumigants will not be available for large-scale use in soil indefinitely.
Biological Approaches for Control of Root Pathogens of Strawberry
Authors: Martin F. N. , Bull C. T.
Published: May 2007

Soil fumigation with methyl bromide plus chloropicrin is used as a preplant treatment to control a broad range of pathogens in high-value annual crop production systems. In California, fumigation is used on approximately 10,125 ha of strawberry production to control pathogens ranging from Verticillium dahliae to root pruning pathogens such as Pythium, Rhizoctonia, or Cylindrocarpon spp. In addition to pathogen control, fumigation also causes an enhanced growth response of the plant and reduces weed pressure. The development of successful, long-term cost effective biocontrol strategies most likely will require the development of an integrated systems approach that incorporates diverse aspects of the crop production system. Although application of single microbial inoculants may provide some level of control for specific production problems, it will be a challenge to provide the broad spectrum of activity needed in production fields.
Cultural Management of Microbial Community Structure to Enhance Growth of Apple in Replant Soils
Authors: Mazzola Mark , Granatstein David M. , Elfving Don C. , Mullinix Kent , Gu Yu-Huan
Published: May 2007

Apple replant disease typically is managed through pre-plant application of broad-spectrum soil fumigants including methyl bromide. The impending loss or restricted use of soil fumigants and the needs of an expanding organic tree fruit industry necessitate the development of alternative control measures. The microbial community resident in a wheat field soil was shown to suppress components of the microbial complex that incites apple replant disease. Pseudomonas putida was the primary fluorescent pseudomonad recovered from suppressive soil, whereas Pseudomonas fluorescens bv. III was dominant in a conducive soil; the latter developed within 3 years of orchard establishment at the same site. In greenhouse studies, cultivation of wheat in replant orchard soils prior to planting apple suppressed disease development. Disease suppression was induced in a wheat cultivar-specific manner. Wheat cultivars that enhanced apple seedling growth altered the dominant fluorescent pseudo-monad from Pseudomonas fluorescens bv. III to Pseudomonas putida. The microbial community resident in replant orchard soils after growing wheat also was suppressive to an introduced isolate of Rhizoctonia solani anastomosis group 5, which causes root rot of apple. Incorporation of high glucosinolate containing rapeseed (‘Dwarf Essex’) meal also enhanced growth of apple in replant soils through suppression of Rhizoc-tonia spp., Cylindrocarpon spp., and Pratylenchus penetrans. Integration of these methods will require knowledge of the impact of the biofumigant component on the wheat-induced disease-suppressive microbial community. Implementation of these control strategies for management of apple replant disease awaits confirmation from ongoing field validation trials.
The Use of Green Fluorescent Protein-Tagged Recombinant Viruses to TestLettuce mosaic virusResistance in Lettuce
Authors: Candresse T. , Le Gall O. , Maisonneuve B. , German-Retana S. , Redondo E.
Published: May 2007

Seed certification and the use of cultivars containing one of two, probably allelic, recessive genes, mo11and mo12, are the principal control methods for Lettuce mosaic virus (LMV) in lettuce. Although for a few LMV isolates, mo12confers resistance with most isolates, the genes mo11or mo12confer a tolerance, and virus accumulation is readily detected in mo1-carrying plants. This phenotype complicates evaluation of the resistance status, in particular for mo11, for which there are no viral strains against which a true resistance is expressed. Two green fluorescent protein (GFP)-tagged viruses were constructed, derived from a non-resistance breaking isolate (LMV-0) and from a resistance-breaking isolate (LMV-E). An evaluation of 101 cultivars of known status was carried out with these recombinant viruses. Using the LMV-0-derived recombinant, identification of mo1-carrying cultivars was simple because, contrary to its wild-type parent, systemic movement of LMV-0-GFP was abolished in resistant plants. This assay detected four cases of misidentification of resistance status. In all these cases, further tests confirmed that the prior resistance status information was incorrect, so that a 100% correlation was observed between LMV-0-GFP behavior and the mo1 resistance status. Similarly, the LMV-E-derived recombinant allowed the identification of mo12lettuce lines because its systemic movement was restricted in mo12lines but not in susceptible or in mo11lines. The tagged viruses were able to systemically invade another host, pea, irrespective of its resistance status against another member of the genus Potyvirus, Pea seed-borne mosaic virus. The use of these recombinant viruses could therefore greatly facilitate LMV resistance evaluation and speed up lettuce breeding programs.
Cotton Seedling Preemergence Damping-Off Incited by Rhizopus oryzae and Pythium spp. and Its Biological Control with Trichoderma spp.
Authors: Howell C. R.
Published: May 2007

Planting the cotton cv. Sure-Grow 747 in cotton seedling disease plots during the 2001 growing season resulted in high levels of preemergence damping-off among the seedlings. Four cotton pathogens, Pythium aphanidermatum, P. ultimum, an unidentified Pythium sp., and Rhizopus oryzae, were isolated from diseased seed embryos and seedlings. Disease incited by the Pythium spp. could be controlled by seed treatment with Metalaxyl, but disease incited by R. oryzae could not. Seed treatment with Metalaxyl in naturally infested field soil was only partially effective; therefore, symptoms in 47% of the diseased seedlings could be attributed to R. oryzae. Susceptibility to disease appeared to be related to release in the spermosphere, by the germinating seeds, of compounds that stimulate pathogen propagule germination, because exudates from seed of the suscept Sure-Grow 747 and extracts from wheat bran induced pathogen germination and growth, whereas exudates from resistant cv. Stoneville 213 did not. However, even Stoneville 213 became susceptible when infested soil was amended with wheat bran. Seed treatment with preparations of Trichoderma virens parent, mutant, and hybrid strains gave effective biological control of preemergence damping-off. Disease control was attributable to metabolism by the biocontrol agent of pathogen germination stimulants released by the seed, because amendment of pathogen-infested soil with the propagule germination stimulants in wheat bran negated the protective effect of the seed treatment.
Ammonia and Nitrous Acid from Nitrogenous Amendments Kill the Microsclerotia of Verticillium dahliae
Authors: Tenuta Mario , Lazarovits George
Published: May 2007

This study examined the mechanisms by which nitrogenous amendments such as meat and bone meal kill the soilborne plant pathogen Verticillium dahliae. The effect of nitrogen products from the amendments on the survival of microsclerotia of V. dahliae was examined by solution bioassay and soil microcosm experiments. Ammonia and nitrous acid but not their ionized counterparts, ammonium and nitrite, were toxic to microsclerotia in bioassays. In microcosms, addition of meat and bone meal (2.5%) to an acidic loamy sand resulted in the accumulation of ammonia and death of microsclerotia within 2 weeks. At lower concentrations (0.5 and 1%), microsclerotia were killed after 2 weeks when nitrous acid accumulated (>0.03 mM). In an alkaline loam soil, microsclerotia survived at 3% meat and bone meal and neither ammonia nor nitrous acid accumulated. The toxicity of ammonia to the pathogen was verified by increasing the concentration of meat and bone meal to 4% or addition of urea (1,600 mg of N per kg) to the loam soil resulting in the accumulation of ammonia (>35 mM) and death of microsclerotia. The toxicity of nitrous acid was verified by adding ammonium sulfate fertilizer to an acidic sand soil. Inhibiting nitrification with dicyandiamide revealed that nitrous acid was generated as a result of the accumulation of nitrite and an acidic pH. Thus, levels to which the toxins accumulated and the effective concentration of amendment were dependent upon the soil examined. Of the two mechanisms identified, accumulation of nitrous acid is the more promising strategy to control plant diseases in acidic soil because it is more toxic than ammonia and is formed at lower concentrations of amendments.
QuantifyingAphanomyces euteichesin Alfalfa with a Fluorescent Polymerase Chain Reaction Assay
Authors: Vandemark G. J. , Barker B. M. , Gritsenko M. A.
Published: May 2007

A polymerase chain reaction (PCR) assay using a set of specific primers and a dual-labeled probe (TaqMan) was developed to quantify the amount of Aphanomyces euteiches DNA in alfalfa plants exhibiting varying levels of disease severity. The study included isolates of race 1 and race 2 of A. euteiches. The assay also discriminated between alfalfa populations for resistance based on analysis of DNA extracted from bulked plant samples. Analysis of individual plants and bulked plant samples of standard check populations with both pathogen isolates resulted in Spearman rank correlations between pathogen DNA content and disease severity index ratings that were greater than 0.75 and highly significant (P < 0.0005). In experiments with a race 1 isolate, the amount of pathogen DNA present in the resistant check WAPH-1 was significantly less than in the susceptible check Saranac. In experiments with a race 2 isolate, the amount of pathogen DNA in the resistant check WAPH-5 was significantly less than in either of the susceptible checks, Saranac and WAPH-1. Discrimination between commercial cultivars based on quantitative PCR analysis of bulked plant samples was similar to classification based on visual assessment of disease severity.
Transmission by Olpidium brassicae of Mirafiori lettuce virus and Lettuce big-vein virus, and Their Roles in Lettuce Big-Vein Etiology
Authors: Lot Hervé , Campbell Robert N. , Souche Sylvie , Milne Robert G. , Roggero Piero
Published: May 2007

Big-vein disease occurs on lettuce worldwide in temperate conditions; the causal agent has been presumed to be Lettuce big-vein virus (LBVV), genus Varicosavirus, vectored by the soilborne fungus Olpidium brassicae. Recently, the role of LBVV in the etiology of big-vein disease has been questioned because a second soilborne virus, Mirafiori lettuce virus (MiLV), genus Ophiovirus, has been found frequently in big-vein-affected lettuce. LBVV and MiLV, detectable and distinguishable by enzyme-linked immunosorbent assay using specific antisera, were tested for their ability to be transmitted from lettuce to lettuce by mechanical inoculation of sap extracts, or by zoospores of O. brassicae, and to cause big-vein disease. Both viruses were mechanically transmissible from lettuce to herbaceous hosts and to lettuce, but very erratically. LBVV was transmitted by O. brassicae but lettuce infected with only this virus never showed symptoms. MiLV was transmitted in the same manner, and lettuce infected with this virus alone consistently developed big-vein symptoms regardless of the presence or absence of LBVV. With repeated mechanical transmission, isolates of both viruses appeared to lose the ability to be vectored, and MiLV appeared to lose the ability to cause big-vein symptoms. The recovery of MiLV (Mendocino isolate, from Cali-fornia) from stored O. brassicae resting spores puts the earliest directly demonstrable existence of MiLV at 1990.