bluuebunny/crossref_metadata_2025 huggingface dataset:
A statistical method for the detection of variants from next-generation resequencing of DNA pools
Authors: Bansal Vikas
Published: June 2010

Motivation: Next-generation sequencing technologies have enabled the sequencing of several human genomes in their entirety. However, the routine resequencing of complete genomes remains infeasible. The massive capacity of next-generation sequencers can be harnessed for sequencing specific genomic regions in hundreds to thousands of individuals. Sequencing-based association studies are currently limited by the low level of multiplexing offered by sequencing platforms. Pooled sequencing represents a cost-effective approach for studying rare variants in large populations. To utilize the power of DNA pooling, it is important to accurately identify sequence variants from pooled sequencing data. Detection of rare variants from pooled sequencing represents a different challenge than detection of variants from individual sequencing.

Results: We describe a novel statistical approach, CRISP [Comprehensive Read analysis for Identification of Single Nucleotide Polymorphisms (SNPs) from Pooled sequencing] that is able to identify both rare and common variants by using two approaches: (i) comparing the distribution of allele counts across multiple pools using contingency tables and (ii) evaluating the probability of observing multiple non-reference base calls due to sequencing errors alone. Information about the distribution of reads between the forward and reverse strands and the size of the pools is also incorporated within this framework to filter out false variants. Validation of CRISP on two separate pooled sequencing datasets generated using the Illumina Genome Analyzer demonstrates that it can detect 80–85% of SNPs identified using individual sequencing while achieving a low false discovery rate (3–5%). Comparison with previous methods for pooled SNP detection demonstrates the significantly lower false positive and false negative rates for CRISP.

Availability: Implementation of this method is available at http://polymorphism.scripps.edu/∼vbansal/software/CRISP/

Contact:  vbansal@scripps.edu
Efficient construction of an assembly string graph using the FM-index
Authors: Simpson Jared T. , Durbin Richard
Published: June 2010

Motivation: Sequence assembly is a difficult problem whose importance has grown again recently as the cost of sequencing has dramatically dropped. Most new sequence assembly software has started by building a de Bruijn graph, avoiding the overlap-based methods used previously because of the computational cost and complexity of these with very large numbers of short reads. Here, we show how to use suffix array-based methods that have formed the basis of recent very fast sequence mapping algorithms to find overlaps and generate assembly string graphs asymptotically faster than previously described algorithms.

Results: Standard overlap assembly methods have time complexity O(N2), where N is the sum of the lengths of the reads. We use the Ferragina–Manzini index (FM-index) derived from the Burrows–Wheeler transform to find overlaps of length at least τ among a set of reads. As well as an approach that finds all overlaps then implements transitive reduction to produce a string graph, we show how to output directly only the irreducible overlaps, significantly shrinking memory requirements and reducing compute time to O(N), independent of depth. Overlap-based assembly methods naturally handle mixed length read sets, including capillary reads or long reads promised by the third generation sequencing technologies. The algorithms we present here pave the way for overlap-based assembly approaches to be developed that scale to whole vertebrate genome de novo assembly.

Contact:  js18@sanger.ac.uk
As-rigid-as-possible mosaicking and serial section registration of large ssTEM datasets
Authors: Saalfeld Stephan , Cardona Albert , Hartenstein Volker , Tomančák Pavel
Published: June 2010

Motivation: Tiled serial section Transmission Electron Microscopy (ssTEM) is increasingly used to describe high-resolution anatomy of large biological specimens. In particular in neurobiology, TEM is indispensable for analysis of synaptic connectivity in the brain. Registration of ssTEM image mosaics has to recover the 3D continuity and geometrical properties of the specimen in presence of various distortions that are applied to the tissue during sectioning, staining and imaging. These include staining artifacts, mechanical deformation, missing sections and the fact that structures may appear dissimilar in consecutive sections.

Results: We developed a fully automatic, non-rigid but as-rigid-as-possible registration method for large tiled serial section microscopy stacks. We use the Scale Invariant Feature Transform (SIFT) to identify corresponding landmarks within and across sections and globally optimize the pose of all tiles in terms of least square displacement of these landmark correspondences. We evaluate the precision of the approach using an artificially generated dataset designed to mimic the properties of TEM data. We demonstrate the performance of our method by registering an ssTEM dataset of the first instar larval brain of Drosophila melanogaster consisting of 6885 images.

Availability: This method is implemented as part of the open source software TrakEM2 (http://www.ini.uzh.ch/∼acardona/trakem2.html) and distributed through the Fiji project (http://pacific.mpi-cbg.de).

Contact:  tomancak@mpi-cbg.de
Field Application of Calcium to Reduce Phytophthora Stem Rot of Soybean, and Calcium Distribution in Plants
Authors: Sugimoto T. , Watanabe K. , Yoshida S. , Aino M. , Furiki M. , Shiono M. , Matoh T. , Biggs A. R.
Published: June 2010

The effect of calcium compounds [Ca(HCOO)2-A and Ca(NO3)2] on the incidence of Phytophthora stem rot of soybean (Glycine max) cv. Tanbakuro was investigated in the field. Disease incidence in control plants in three fields naturally infested with Phytophthora sojae ranged from 11.7 to 52.0% at 140 days after transplanting. Independent of the pathotype diversity, 4 and 10 mM of the calcium compounds applied twice (prior to transplanting and 14 days after transplanting) significantly suppressed disease incidence and delayed onset. Ca(HCOO)2-A (Suicaru) was more effective than calcium nitrate for reducing disease incidence. In most cases, the calcium amendments increased plant height, number of nodes and pods, and seed yields, and reduced low-quality seeds. Scanning electron microscopy with fresh samples showed increased accumulation of calcium crystals around the cambium and xylem elements of soybean plants treated with 10-mM Ca(HCOO)2-A and Ca(NO3)2. Mycelial penetration was inhibited at these sites. These results indicated that calcium-rich areas may be more resistant to invasion by P. sojae, and the calcium crystals may play an important role in calcium ion storage and its availability for those tissues to maintain long-term field resistance.
Influence of Environment on Airborne Spore Concentrations and Severity of Asparagus Purple Spot
Authors: Granke L. L. , Hausbeck M. K.
Published: June 2010

Environmental conditions, airborne concentrations of Pleospora herbarum ascospores and conidia, and purple spot disease severity on spears and fern were monitored for 3 years in two no-till asparagus fields in Michigan. Purple spot lesion development on spears was correlated with low temperature and vapor pressure deficit and high rainfall. Low vapor pressure deficit and high rainfall were also correlated with airborne ascospore concentrations at both sites. Lesion development on the fern was favored by longer periods of leaf wetness and low vapor pressure deficit and rainfall. Daily airborne conidia concentrations were positively correlated with average temperature and the number of hours of leaf wetness per day and negatively correlated with vapor pressure deficit. Airborne conidia concentrations displayed a diurnal periodicity with greater concentrations between 700 and 1300 h. The results of this study indicate that a model to predict purple spot on asparagus spears should include temperature, vapor pressure deficit, and rainfall; a model for disease on the fern should include temperature, leaf wetness, vapor pressure deficit, and rainfall.
Risk Assessment Methods for the Ringspot Pathogen Mycosphaerella brassicicola in Vegetable Brassica Crops
Authors: Wakeham Alison J. , Kennedy Roy
Published: June 2010

Mycosphaerella brassicicola causes ringspot on Brussels sprouts, which can result in substantial yield loss in commercial production. Brussels sprout buttons are downgraded if this pathogen occurs on them. In this study, the effect of temperature and wetness duration was investigated on infection of Brussels sprouts using controlled environments (CE). The effect of temperature and wetness duration on inoculum production and ascospore discharge was also investigated. Infection by M. brassicicola was described using a mathematical model and was compared to estimates of ascospore availability obtained via a volumetric air sampler and immunofluorescence (IF). Infection of M. brassicicola was correlated (r = 0.92) with temperature during leaf wetness periods. The relationship between temperature and time to discharge of 5 and 50% of the cumulative total number of ascospores from ringspot lesions was r = 0.99 and 0.98, respectively (P < 0.001). In field experiments, an optimal wind run (the product of the average wind speed and the period over which that average speed was measured) of 250 to 500 km day-1 was required for the dissemination of ringspot inoculum to field bait plants. Quantification of M. brassicicola inoculum in collected field aerosols was possible using a monoclonal antibody in a plate-trapped antigen enzyme-linked immunosorbent assay. Precoating of the air sampler wells with sodium azide prevented trapped spores from germinating. Ringspot inoculum could be detected and quantified in air samples from commercial crops of Brussels sprouts in the United Kingdom. Low levels of ringspot inoculum measured within crops did not lead to disease development.
Pathogenicity ofDiatrypaceaeSpecies in Grapevines in California
Authors: Trouillas F. P. , Gubler W. D.
Published: June 2010

Diatrypaceous fungi have been isolated from cankered wood of grapevines (Vitis vinifera) in California. However, the pathogenicity of these fungi and their potential to cause diseases in grapevine was unknown. A series of pathogenicity tests were conducted to determine if these fungi were grapevine pathogens. In all experiments, species of the family Diatrypaceae were re-isolated from the margins of developing lesions, although frequency of re-isolation and average length of vascular discolorations varied among isolates. Cryptovalsa ampelina, Diatrype stigma, and Eutypa leptoplaca were capable of colonizing dormant canes and causing vascular necrosis. Cryptosphaeria pullmanensis, C. ampelina, D. stigma, Diatrype whitmanensis, and E. leptoplaca infected and caused lesions in green shoots of the new vegetative growth. Diatrype oregonensis and Diatrypella verrucaeformis isolates did not produce lesions that were significantly different from those produced in the control shoots and canes. This suggests that D. oregonensis and D. verrucaeformis fungi may be saprophytic rather than pathogenic to grapevine.
Long-Distance Spore Transport of Wheat Stripe Rust Pathogen from Sichuan, Yunnan, and Guizhou in Southwestern China
Authors: Wang Haiguang , Yang X. B. , Ma Zhanhong
Published: June 2010

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici, is the most devastating wheat disease in China. Although pathogens oversummering and overwintering in many regions of southwestern China have been studied, spore exchange among these regions is largely unknown. It is especially important to study the role of Sichuan, Yunnan, and Guizhou provinces in southwestern China in contributing to regional disease outbreaks in the major wheat-growing areas of China. Long-distance transport of P. striiformis f. sp. tritici urediniospores after oversummering and overwintering in Sichuan, Yunnan, and Guizhou was investigated using the HYSPLIT-4 model based on meteorological data. Results suggest that the pathogen spores can interchange frequently among Sichuan, Yunnan, and Guizhou in both spring and autumn. The sources of inoculum in Sichuan, Yunnan, and Guizhou have a high probability of impacting the epidemics in northern, northwestern, and southwestern China. In particular, disease epidemics in Guizhou can be affected not only by local inoculum but also by that from Yunnan in the spring.
Distribution, Pathotypes, and Metalaxyl Sensitivity of Phytophthora sojae from Heilongjiang and Fujian Provinces in China
Authors: Cui Linkai , Yin Weixiao , Tang Qinghua , Dong Suomeng , Zheng Xiaobo , Zhang Zhengguang , Wang Yuanchao
Published: June 2010

Phytophthora sojae causes root and stem rot, one of the most devastating diseases of soybean worldwide. In Heilongjiang and Fujian provinces in China, serious cases of Phytophthora stem and root rot have occurred and caused heavy losses in the past several years. To determine the current population status of this pathogen, we investigated the pathogen's distribution, pathotypes, and metalaxyl sensitivity in both provinces. P. sojae was baited and isolated from 258 soil samples in both provinces using the soybean leaf bait method. The pathotypes of all isolates were characterized on 13 differential soybean cultivars using the hypocotyl slit inoculation method, and the sensitivity of all isolates to metalaxyl was tested in vitro. In all, 75 isolates were recovered from 75 fields in 33 counties; of these, 31 counties were in Heilongjiang Province and 2 counties were in Fujian Province. Thirty-five new pathotypes were identified compared with the previously defined races. Less than 5% of the isolates were virulent to cultivars with individual Rps genes 1a, 1c, or 1k. No metalaxyl-resistant isolates were found; the half maximal effective concentration values of all isolates ranged from 0.04 to 0.22 μg ml–1. These results suggest that effective management of the disease in both provinces can be accomplished through the use of resistant cultivars with Rps genes 1a, 1c, or 1k and the fungicide metalaxyl.
Screening Common Bean for Resistance to Four Sclerotinia sclerotiorum Isolates Collected in Northern Spain
Authors: Pascual Aida , Campa Ana , Pérez-Vega Elena , Giraldez Ramón , Miklas Phillip N. , Ferreira Juan José
Published: June 2010

White mold, caused by the fungus Sclerotinia sclerotiorum, is a serious disease in common bean (Phaseolus vulgaris) causing significant yield loss. Few cultivars with high levels of physiological resistance to white mold have been described in common bean. The objectives of this study were to (i) determine variation in aggressiveness for the local S. sclerotiorum isolates and (ii) identify sources of resistance against local isolates using the greenhouse straw test. The evaluated materials included 199 accessions of a core collection established from the main bean gene bank in Spain and 29 known cultivars or lines, 5 of them described as resistant sources to white mold: G122, PC50, A195, Cornell 606, and MO162. Significant differences for aggressiveness among the four S. sclerotiorum isolates were detected. Generally, isolates 1 and 3 were more aggressive than isolates 2 and 4. In all, 19 genotypes exhibited a level of resistance equal to or significantly better than G122: 11 accessions from the core collection and 8 cultivars or lines from known materials, including the lines A195 and Cornell 606. To confirm resistance, 19 selected genotypes were tested using a more severe straw test with reactions evaluated 21 days after inoculation. Fifteen genotypes exhibited significantly less susceptibility than G122: eight accessions from the core collection and the known cultivars or lines AB136, Kaboon, BRB57, BRB130, Don Timoteo, and A195. The logical next step will be to evaluate the best genotypes for field reaction to white mold and conduct inheritance studies.
Interactive Effects of Planting Date and Cultivar on Tomato Spotted Wilt of Peanut
Authors: Culbreath A. K. , Tillman B. L. , Tubbs R. S. , Beasley J. P. , Kemerait R. C. , Brenneman T. B.
Published: June 2010

Field experiments were conducted at Gainesville and Marianna, FL in 2004 and 2005 in which severity of spotted wilt, caused by Tomato spotted wilt virus, and pod yield were compared in six peanut (Arachis hypogaea) cultivars. The six cultivars included the moderately field resistant cultivars ANorden, C-99R, and Georgia Green; the highly field resistant cultivars AP-3 and DP-1; and the susceptible cultivar SunOleic 97R. There were four trials at each location, with four planting dates that ranged from late March to early June. Tomato spotted wilt severity in moderately resistant and susceptible cultivars was lower at Gainesville than at Marianna in both years in moderately resistant and susceptible cultivars. Trends in incidence for the two locations were less evident for AP-3 and DP-1. At Gainesville, there were few differences in tomato spotted wilt severity, and severity ratings were similar for Georgia Green and SunOleic 97R in two of four trials in 2004 and across all trials in 2005. At Marianna, severity ratings were lower for Georgia Green than for SunOleic 97R in six of the eight trials, and severity of tomato spotted wilt was lower for AP-3, C-99R, and DP-1 than for Georgia Green in all eight trials. In 2004, there was a trend toward decreasing severity ratings for Georgia Green and SunOleic 97R with later planting dates, but not for AP-3 or DP-1 at Marianna. Split-plot field experiments were also conducted at Tifton, GA in 2005 through 2007 in which incidence of tomato spotted wilt and pod yield were compared for peanut cultivars AP-3 and Georgia Green across planting dates ranging from late April through late May. Incidence of tomato spotted wilt was lower for AP-3 than for Georgia Green within each planting date of all years, and planting date effects were smaller in AP-3, if observed at all, than in Georgia Green. In most planting dates of all three trials, yields were higher for AP-3 than for Georgia Green. The relationships between yield and planting date were not consistent. These results indicate that the level of field resistance in AP-3 and DP-1 cultivars is sufficient to allow planting in late April without greatly increasing the risk of losses to tomato spotted wilt.
Colonization of Potato by Colletotrichum coccodes: Effect of Soil Infestation and Seed Tuber and Foliar Inoculation
Authors: Pasche J. S. , Taylor R. J. , Gudmestad N. C.
Published: June 2010

Colonization of potato (Solanum tuberosum) tissue, including roots, stolons, and above and below ground stems, by Colletotrichum coccodes, the causal agent of black dot, was evaluated following soil infestation, inoculation of seed tubers and foliage, and every combination thereof, in field trials over two growing seasons in North Dakota and Minnesota. A total of 107,520 isolations for C. coccodes performed across four site-years allowed for an extensive comparison of fungal colonization of the host plant and disease severity. The black dot pathogen was detected in potato stems at the first sampling date in all four site-years, as early as 14 days prior to emergence. Colonization of above and below ground stems occurred at a higher frequency than in roots and stolons in all four site-years, resulting in significantly higher relative area under the colonization progress curves (RAUCPCs) (α = 0.05). Although fungal colonization and disease incidence were higher in inoculated and/or infested treatments, sufficient natural inoculum was present to result in substantial levels of disease in noninoculated and noninfested plots. However, noninoculated and noninfested plots displayed the lowest RAUCPC values across three of four site-years and those treatments with multiple inoculation events tended to have higher RAUCPC values. Isolates belonging to vegetative compatibility group (VCG)2 and -5 were recovered from plants sampled in 2004 more frequently than isolates belonging to VCG1 and -3. A significant difference in disease incidence on stems was observed only in North Dakota in 2004 and Minnesota in 2003 (α = 0.05). Noninoculated and noninfested plots displayed the lowest disease incidence, whereas those treatments with more than one inoculation and/or infestation event tended to have higher disease incidence. Results of this study, including the disease severity and yield data, provide a better understanding of colonization of potato plants by C. coccodes and its impact.
First Report of Impatiens necrotic spot virus Infecting Phalaenopsis and Dendrobium Orchids in Yunnan Province, China
Authors: Zhang Q. , Ding Y.-M. , Li M.
Published: June 2010

In November 2007, leaves of 79 Phalaenopsis and two Dendrobium orchid plants in a nursery in Yunnan Province showed large chlorotic/necrotic ringspot symptoms. Eight symptomatic leaves from Phalaenopsis and two from Dendrobium were sampled and tested for Impatiens necrotic spot virus (INSV), Tomato spotted wilt virus (TSWV), Watermelon silver mottle virus (WSMoV), Groundnut bud necrosis virus (GBNV), Tomato chlorotic spot virus (TCSV), and Groundnut ringspot virus (GRSV) with double-antibody sandwich (DAS)-ELISA kits (Agdia Inc., Elkhart, IN). All samples were positive for INSV and negative for TSWV, WSMoV, GBNV, TCSV, and GRSV. Total RNA extracts were prepared from all ELISA-positive samples with the RNeasy extraction kit (Huashun Inc., Shanghai, China). Reverse transcription (RT)-PCR was carried out with specific primers to the INSV N gene (ZI2F, 5′-GTTTAGCCTTACCAAT-3′ and ZI2R, 5′-TACCAACAACCGTGAA-3′), designed from a sequence of GenBank Accession No. AB109100. All ELISA-positive samples yielded an amplification product of the expected 539 bp as observed by gel electrophoresis in 1% agarose. Three clones from each isolate were sequenced and two N gene consensus sequences of the isolates from Phalaenopsis and Dendrobium were determined (GenBank Nos. GU289904 and GU289905, respectively). Nucleotide sequences of these two Chinese orchid isolates were 98 to 99% identical with sequences of isolates from the Netherlands, United States, Italy, and Japan (GenBank Nos. X66972, D00914, DQ425096 and AB109100, respectively). To our knowledge, this is the first report of INSV infecting Phalaenopsis and Dendrobium in Yunnan Province, although INSV has been reported in Oncidium in Yunnan Province previously (2), and the first time that INSV has been detected in Dendrobium. An investigation of the orchid nurseries looking for the thrips vector (1) of INSV was performed and a few thrips were found, suggesting that thrips may not be responsible for the observed prevalence of INSV in these nurseries. The orchids were imported from Taiwan and reproduced by tissue culture and it is possible that INSV found to be infecting orchids in these Yunnan nurseries may be from the infected source plant and was not eradicated completely through tissue culture. To reduce spread of INSV, virus-free tissue culture should be a priority for orchid production.

References: (1). S. T. Koike and D. E. Mayhew. Orchids. Mag. Am. Orchid Soc. 70:746, 2001. (2). Q. Zhang et al. Plant Quarantine. 22:348, 2008.
First Report of Seedling Blight Caused by Rhizoctonia solani on Dioscorea nipponica in China
Authors: Bai Q. , Wang N. , Gao J.
Published: June 2010

Throughhill yam (Dioscorea nipponica Makino), a perennial winding herb and a member of the Discoreaceae, is distributed principally in northeast Asia. It is used to produce medicine for treating coronary heart disease, diabetes mellitus, and inflammation. In China, this species is cultivated in many provinces such as Liaoning, Jilin, Heilongjiang, Hebei, Inner Mongolia, and Shanxi. In July 2006, seedling blight was observed on D. nipponica with disease incidence ranging from 37 to 75% in commercial fields in Antu County, China. In the early stages of disease development, water-soaked lesions appeared at the stem base and on leaves near the ground. Lesions later turned dark brown and necrotic. Leaves eventually became chlorotic, stem and petioles collapsed gradually, and plants died. Mycelium was observed to be growing on the surface of infected tissues and adjacent plants, and brown, hard sclerotia were produced on stem or petiole surfaces. A fungus with morphological characteristics of Rhizoctonia solani Kühn was consistently isolated from diseased tissues that were plated on potato dextrose agar (PDA). Mycelium was branched at right angles with a septum near the branch and a slight constriction at the branch base. Cells from hyphae grown on 2% water agar at 25°C were determined to be multinucleate when stained with 1% safranin O and 3% KOH solution (1) and examined at ×400. Anastomosis groups were determined by pairing isolates with 12 tester strains representing all subgroups of AG1 to AG5 on 2% water agar in petri plates (2). The anastomosis grouping of isolates Rs1, Rs2, and Rs5 was determined to be AG1-IB and that of isolates Rs3 and Rs6 was determined to be AG2-1. The rDNA internal transcribed spacer (ITS) gene sequence of isolates Rs1, Rs2, and Rs5 (GenBank Accession Nos. GU585667, GU596490, and GU594691) had 100, 99, and 100% nucleotide identity, respectively, with AG1-IB (GenBank Accession No. FG440191). The rDNA-ITS of isolates Rs3 and Rs6 (GenBank Accession Nos. GU596493 and GU594692) exhibited 99% homology with AG2-1 (GenBank Accession No. EU513135). Pathogenicity tests were performed on healthy, potted 2-year-old plants of D. nipponica. Twenty plants were wound inoculated by placing 0.6-cm mycelial plugs from 3-day-old PDA cultures on leaves and stems. Twenty plants were treated with PDA plugs as controls. Plants were maintained at 25°C and 95% relative humidity on a 12-h light/dark regimen. Typical symptoms of leaf and stem rotting identical to those observed in the commercial field appeared 4 days after inoculation and all inoculated plants died within 10 days. No disease symptoms were observed on control plants. Rhizoctonia solani was consistently reisolated from symptomatic tissues. To our knowledge, this is the first report of R. solani causing seedling blight on D. nipponica in the world.

References: (1) R. J. Bandoni. Mycologia 71:873, 1979. (2) C. C. Tu and J. W. Kimbrough. Mycologia 65:941, 1973.
First Report of Corynespora cassiicola Causing Leaf Spot of Cassava in China
Authors: Liu X.-B. , Shi T. , Li C.-P. , Cai J.-M. , Huang G.-X.
Published: June 2010

Cassava (Manihot esculenta) is an important economic crop in the tropical area of China. During a survey of diseases in July and September of 2009, leaf spots were observed on cassava plants at three separate plantations in Guangxi (Yunfu and Wuming) and Hainan (Baisha) provinces. Circular or irregular-shaped leaf spots were present on more than one-third of the plants. Spots were dark brown or had white papery centers delimited by dark brown rims and surrounded by a yellow halo. Usually, the main vein or small veinlets adjacent to the spots were dark. Some defoliation of plants was evident at the Wuming location. A fungus was isolated from symptomatic leaves from each of the three locations and designated CCCGX01, CCCGX02, and CCCHN01. Single-spore cultures of these isolates were incubated on potato dextrose agar (PDA) for 7 days with a 12-h light/dark cycle at a temperature of 28 ± 1°C. Conidiophores were straight to slightly curved, unbranched, and pale to light brown. Conidia were formed singly or in chains, obclavate to cylindrical, straight or curved, subhyaline-to-pale olivaceous brown, 19.6 to 150.3 μm long and 5.5 to 10.7 μm wide at the base, with 4 to 13 pseudosepta. Morphological characteristics of the specimen and their conidia were similar to the descriptions for Corynespora cassiicola (2). The isolate CCCGX01 was selected as a representative for molecular identification. Genomic DNA was extracted by the cetyltrimethylammoniumbromide protocol (3) from mycelia and used as a template for amplification of the internal transcribed spacer (ITS) region of rDNA with primer pair ITS1/ITS4. The sequence (GenBank Accession No. GU138988) exactly matched several sequences (e.g., GenBank Accession Nos. FJ852715, EF198117, and AY238606) of C. cassiicola (1). Young, healthy, and fully expanded green leaves of cassava cv. SC205 were surface sterilized. Ten leaves were inoculated with 10-μl drops of 104 ml suspension of conidia and five leaves were inoculated with the same volume of sterile water to serve as controls. After inoculation, leaves were placed in a dew and dark chamber for 36 h at 25°C and subsequently transferred to the light for 5 days. All inoculated leaves with isolates showed symptoms similar to those observed in natural conditions, whereas the controls remained symptom free. The morphological characteristics of reisolated conidia that formed on the diseased parts were identical with the nature isolates. To our knowledge, this is the first report of leaf spot caused by C. cassiicola on cassava in China.

References: (1) L. J. Dixon et al. Phytopathology 99:1015, 2009. (2) M. B. Ellis et al. Corynespora cassiicola. No. 303 in: CMI Description of Pathogenic Fungi and Bacteria. Commonwealth Mycological Institute, Kew, UK 1971. (3) J. R. Xu et al. Genetics 143:175, 1996.
First Report of Alternaria Leaf Blight on Bupleurum chinense Caused by Alternaria alternata in China
Authors: Zhang Z. , Wei J. H. , Yang C. M. , Chen H. Q. , Sui C. , Gao Z. H.
Published: June 2010

Bupleurum chinense DC. (family Umbelliferae) is an important medicinal herb in traditional Chinese medicine and is cultivated as an economically important plant in China (2). From 2006 to 2009, severe foliar disease was observed on B. chinense in Haidian, Changping, and Shunyi districts, Beijing, China. Approximately 75 to 85% of fields were affected with disease incidence ranging from 65 to 90%. Distribution of the disease in affected B. chinense fields was generally associated with high soil moisture, often corresponding to poor drainage. Initial symptoms first appeared on older leaves as irregularly shaped, minute, dark brown-to-black spots, with yellow borders on the edge of the affected leaflet blade. As the disease progressed, the lesions expanded, causing the leaflets to turn brown, shrivel, and die. Isolations performed on potato dextrose agar (PDA) initially resulted in white colonies. After 7 days of incubation at 25°C, the colonies turned gray or brown. Conidia varied in size from 10 × 6 to 40 × 12 μm, appeared brown to dark brown or olive-brown, were short beaked and borne in long chains, oval and bean-shaped with one to six transverse septa and zero to three longitudinal septa. Sequences of the rDNA from the internal transcribed spacer regions 1 and 2 and the 5.8S gene were amplified using primers ITS1 and ITS4, were obtained from three isolates, and comparisons with GenBank showed 100% similarity with A. alternata (Genbank Accession No. AB470912.1). For pathogenicity tests, three isolates were grown on PDA for 14 days. Inoculations were performed on detached, surface-sterilized, and healthy B. chinense leaflets following the method of Belisario (1). A 5-μl drop of conidial suspension containing 1 × 105 CFU/ml was placed on each leaflet and 12 leaves per isolate were used. Leaves were incubated in a growth chamber (80 to 90% relative humidity; 50 to 60 klx/m2 light intensity with a 12-h photoperiod). After 5 days, leaf spots similar to the original symptoms developed on all inoculated leaves and A. alternata was consistently reisolated from symptomatic leaf tissues on PDA. Control leaflets inoculated with sterile water remained asymptomatic. The experiment was performed three times. To our knowledge, this is the first report of A. alternata on B. chinense from China.

References: (1) A. Belisario et al. Plant Dis. 83:696, 1999. (2) C. Sui et al. Plant Dis. 93:844, 2009.
Weed Hosts of Globodera pallida from Idaho
Authors: Boydston R. A. , Mojtahedi H. , Bates C. , Zemetra R. , Brown C. R.
Published: June 2010

The potato cyst nematode, Globodera pallida (PCN), a restricted pest in the United States, was first reported in Bingham and Bonneville counties of Idaho in 2006 (1). The U.S. government and Idaho State Department of Agriculture hope to eradicate it from infested fields. Eradicating PCN will require depriving the nematodes of their hosts over a protracted time period. Functional eradication might be achieved with relatively high, proven to be efficacious dosages of soil fumigants. The presence of host weeds of PCN can play a significant role in the success of the eradication program. To determine the host status of common weeds found in potato fields of the Pacific Northwest, host suitability tests were conducted in a secured greenhouse located at the University of Idaho at Moscow. Reproduction of PCN on nine weeds including hairy nightshade (Solanum physalifolium formerly S. sarrachoides) and cutleaf nightshade (S. triflorum) (biotypes from Idaho and Washington), black nightshade (S. nigrum) (Washington biotype), bittersweet nightshade (S. dulcamara) (Idaho biotype), redroot pigweed (Amaranthus retroflexus), kochia (Kochia scoparia), and common lambsquarters (Chenopodium album) were compared with reproduction on Desiree, Russet Burbank (known hosts), and Santé (poor host) potatoes (S. tuberosum). Plants were grown in 10-cm-diameter clay pots containing sandy loam soil previously fumigated with methyl bromide and inoculated with 10 to 150 cysts that were either collected from infested fields or raised in the secured greenhouse (ample diapause period elapsed). Treatments were replicated five times and each trial lasted 3 months. Cysts were extracted from soil with a Fenwick can, and the reproductive factor (RF = final cyst count ÷ initial inoculum) was determined. While both biotypes of hairy nightshade were suitable hosts of PCN (161–668 ÷ 150; RF = >1), cutleaf biotypes, black, and bittersweet nightshades were poor hosts (1–108 ÷ 150; RF = <1). Russet Burbank (77 ÷ 40; RF = 1.9) and Desiree (21–119 ÷ 75; RF = >1) proved to be suitable hosts and Santé (1–20 ÷ 150; RF = <1) a poor host of Idaho PCN. Although some cysts were recovered from pots containing the remaining weed species, they may have been part of the original inoculum. The significance of nightshade species (whether suitable or poor hosts) in eradication of potato cyst nematode from infested fields cannot be overemphasized.

Reference: (1) A. M. Skantar et al. J. Nematol. 39:133, 2007.
First Report of Rapid Blight Caused by Labyrinthula terrestris on Poa annua in Colorado
Authors: Hyder N. , Wong F. P. , Koski A. , Tisserat N. , Stowell L.
Published: June 2010

A disease characteristic of rapid blight caused by the net slime mold, Labyrinthula terrestris (1,2), was observed on three annual bluegrass (Poa annua) putting greens from a golf course in Adams County, Colorado in April 2009. Symptoms included water-soaked lesions and browning and bronzing of leaves. With microscopic observation, the fusiform cells typically associated with Labyrinthula spp. (1) were detected inside symptomatic leaf tissue. The pathogen was isolated by placing symptomatic leaves on a selective medium modified from Bigelow et al. (1) (12 g of granulated agar [Fisher Scientific, Pittsburg, PA], 10 ml of horse serum [Hema Resource and Supply, Aurora, OR], and 250 μg of ampicillin, streptomycin sulfate, and penicillin G [Sigma, St. Louis, MO] in artificial seawater at 4.0 dS/m electrical conductivity [Instant Ocean, Atlanta, GA]). Irregular-shaped digitate colonies of fusiform cells developed within 1 to 2 days. The isolated organism was then used to fulfill Koch's postulates on 2-week-old Poa trivialis ‘Sabre III’ seedlings and 4-week-old Poa annua seedlings planted in a 90:10 peat moss/sand mixture in 6-cm-diameter pots. Plants were inoculated with a 9 × 106 cells/ml suspension of L. terrestris cells in artificial seawater (4.0 dS/m) as described by Peterson et al. (2) and irrigated daily with artificial seawater (4.0 dS/m). Negative controls consisted of either P. trivialis or P. annua plants irrigated with artificial seawater only. There were three replications for each treatment and the experiment was repeated for each grass species. Pots were maintained at 28 to 33°C in the greenhouse with ambient light. Within 8 to 10 days of inoculation, 95% of the plants showed symptoms of severe rapid blight, while noninoculated plants showed some minor salt stress symptoms but were otherwise healthy. The organism was successfully reisolated from several plants from each replication using the method described above. Results were the same for all experiments. Rapid blight is frequently associated with high soil salinity (>2.5 dS/m total dissolved salts) (1) and sodium levels above 110 mg/kg (Mehlich-3 extraction) in diagnostic samples. Soil salinity levels from the site affected by the disease were below this guideline. However, sodium levels measured an average of 184 mg/kg. The ability of this pathogen to cause disease on plants growing in soils not measuring high in salinity, and only with elevated sodium, should be considered when attempting to ascertain rapid blight as a cause of turf damage.

References: (1) D. M. Bigelow et al. Mycologia 97:185, 2005. (2) P. D. Peterson et al. Online publication. doi:10.1094/ATS-2005-0328-01-RS. Applied Turfgrass Science, 2005.
First Report of Diplodia scrobiculata Causing Canker and Branch Dieback on Strawberry Tree (Arbutus unedo) in Italy
Authors: Linaldeddu B. T. , Scanu B. , Franceschini A.
Published: June 2010

Strawberry tree, family Ericaceae, is an evergreen shrub or small tree that grows in the Mediterranean Region. In spring 2009, a survey was conducted to study the fungi associated with canker and branch dieback of strawberry tree in a natural stand located on Caprera Island (41°12′N, 9°27′E), Italy. Fungal isolates obtained from live twigs and branches showing sunken necrotic bark lesions were identified as Diplodia scrobiculata J. de Wet, Slippers & M.J. Wingf. on the basis of morphological features (1). On potato dextrose agar (PDA) at 25°C, D. scrobiculata isolates developed white colonies with appressed mycelium that became dark gray after 4 to 6 days and formed pycnidia after 2 weeks on sterile Pinus radiata needles placed on the PDA surface. The conidia were dark brown with zero to three septa, clavate with truncate base, and measured 31.1 to 41.9 (36.8) × 11.3 to 16.3 (12.6) μm, with a length/width ratio of 2.9 ± 0.1 (mean ± standard error) (n = 50). Identity was confirmed by analysis of the internal transcribed spacer region (ITS1-5.8S-ITS2) of rDNA. BLAST searches in GenBank showed 100% similarity with reference sequences of D. scrobiculata (GenBank Nos. AY160200, EU220438, EU220444, and EU392283). The representative sequence of one D. scrobiculata strain (BL5), stored in the culture collection of the Department of Plant Protection at the University of Sassari, was deposited in GenBank (Accession No. GU722102). Pathogenicity of strain BL5 was tested by stem inoculation on eight 2-year-old strawberry tree seedlings maintained in a greenhouse at 18 to 26°C. A mycelial plug (3 to 4 mm2) taken from the margin of an actively growing colony on PDA was put in a shallow wound (~3 mm) made by a scalpel on the basal part of the stem of each seedling. Four weeks after inoculation, the seedlings displayed dark brown-to-black discoloration, measuring 1.7 ± 0.6 cm (mean ± standard error) of the bark and wood tissues of the stems. The pathogen was successfully reisolated from symptomatic stem tissues, thus fulfilling Koch's postulates. Three control seedlings inoculated with sterile PDA plugs remained asymptomatic. These results demonstrate the active role played by D. scrobiculata in the aetiology of the canker and branch dieback observed on strawberry tree. D. scrobiculata is generally recognized as a weak pathogen of gymnosperms worldwide (2), however, it has recently been reported on olive in Italy (3). To our knowledge, this is the first report of D. scrobiculata on strawberry tree. Currently, further investigations are in progress to determine the possible role of biotic and abiotic factors in the development of this disease.

References: (1) J. De Wet et al. Mycol. Res. 107:557, 2003. (2) J. De Wet et al. Mol. Phylogenet. Evol. 46:116, 2008. (3) C. Lazzizera et al. Fungal Divers. 31:63, 2008.
First Report of Potato mop-top virus on Potato in Poland
Authors: Budziszewska M. , Wieczorek P. , Nowaczyk K. , Borodynko N. , Pospieszny H. , Obrêpalska-Stêplowska A.
Published: June 2010

Potato mop-top virus (PMTV) is a serious pathogen occurring in Northern Europe, North and South America, and Asia that significantly reduces potato (Solanum tuberosum) production. PMTV is transmitted by Spongospora subterranea, the casual agent of potato powdery scab, and causes the characteristic brown arcs and circles (spraing symptoms) in potato tubers, stunting of stems, shortening of internodes, and mosaic patterns (V-shaped) on leaves as well as leaf necrosis (2). S. subterranea and PMTV are mainly associated with cool, humid environments. Between 2005 and 2009, extensive surveys for PMTV were conducted in Polish potato fields with an emphasis on areas neighboring countries where the virus had previously been reported. Approximately 18,000 tubers from 39 cultivars from different regions of Poland were collected. Tubers were first visually inspected for symptoms within the flesh and then selected tubers were analyzed by double-antibody sandwich (DAS)-ELISA (3). Symptomatic samples tested by ELISA gave A405 values approximately threefold higher than negative controls and approximately two- to fivefold lower than PMTV-positive controls (supplied by J. Valkonen). Total RNA was isolated (1) from tubers testing positive for PMTV by DAS-ELISA. cDNA synthesis and subsequent PCR amplification of the CP region were carried out using primers located in RNA2: PMTV1 5′GGTTTGTTTACCACCCTTGG3′ (3) and PMTV2 5′AAAAGCCTGAGCGGTTAATTG3′ (courtesy of E. Savenkov), which amplified a 530-bp product. No PMTV was detected in Poland between 2005 and 2007. In 2008, one tuber (cv. Inwestor) from central Poland (Łódź County) tested positive for PMTV. The RT-PCR products were sequenced and the sample from 2008 was submitted to GenBank (PMTV-Pl CP, Accession No. GQ503252). In 2009, additional infected tubers were found in three Polish cultivars (Bartek, Glada, Ruta) from the same county. Sequence comparisons of PMTV-Pl revealed 99% nucleotide identity and approximately 98% amino acid identity to Czech, Swedish, and Finnish PMTV isolates. To our knowledge, this is the first report of PMTV in Poland. Poland is one of the major potato-producers in Europe with the 2008 crop around 10 million t. If PMTV spreads in Poland, the virus could threaten potato production.

References: (1) S. Chang et al. Plant Mol Biol Rep. 11:113, 1993. (2) A. Germundsson et al. J. Gen. Virol. 83:1201, 2002. (3) S. Latvala-Kilby et al. Phytopathology 99:519, 2009.
First Report of Iresine viroid 1 in Celosia plumosa in the Netherlands
Authors: Verhoeven J. Th. J. , Jansen C. C. C. , Botermans M. , Roenhorst J. W.
Published: June 2010

In 2008, in the framework of surveying for pospiviroids, nine symptomless clones of Celosia plumosa (Voss) Burv. (Amaranthaceae) from a Dutch breeding company were tested by reverse transcription (RT)-PCR with primer sets Pospi1-RE/FW and Vid-RE/FW (4). In four samples, amplicons of 227 nt were obtained with primers Pospi1-RE/FW. Sequencing of the amplicons showed identities of more than 99% to the partial sequence of Iresine viroid 1 (IrVd-1) from Alternanthera sessilis, NCBI GenBank Accession No DQ846886 (2). Subsequently, a set of primers was designed to amplify the complete viroid genome, i.e., IrVd-FW1 5′-GCG GAA GAA ACA GGA GCT CGW CT-3′ and IrVd-RE1 5′-CGC GWG GAG TTC TCC GGT CTT TA-3′ – identical to nt 168 to 190 and 145 to 167 of the complete IrVd-1 sequences in the NCBI GenBank (Nos. DQ094293, DQ094294, NC_003613, and X95734). One isolate from C. plumosa was amplified with this primer pair and amplicons were cloned into the pGEM-T Easy Vector System II. Sequencing of one individual cDNA clone (GenBank Accession No. GU911350) revealed a genome size of 370 nt and 98.1% sequence identity to the IrVd–1 isolate from Vinca major, GenBank Accession No. DQ094293 (1). Hence, the viroid was identified as IrVd-1. The isolate from C. plumosa was also mechanically inoculated to 10 healthy plants of C. plumosa, chrysanthemum (Chrysanthemum × morifolium) cv. White Delianne, potato (Solanum tuberosum) cv. Nicola, and tomato (Solanum lycopersicum) cv. Moneymaker. No symptoms were observed over a 6-week period, and RT-PCR with primers Pospi1-RE/FW on bulked samples of five plants per species only identified IrVd-1 in both samples of C. plumosa. For tomato, these results confirm those of Spieker (3). Therefore, in contrast to the other pospiviroids, it seems unlikely that IrVd-1 poses a threat to potato and tomato.

References: (1) X. Nie et al. Can. J. Plant Pathol. 27:592, 2005. (2) R. P. Singh et al. Plant Dis. 90:1457, 2006. (3) R. L. Spieker. J. Gen. Virol. 77:2631, 1996. (4) J. Th. J. Verhoeven et al. Eur. J. Plant Pathol. 110:823, 2004.
First Report of Banana bract mosaic virus in Flowering Ginger in Hawaii
Authors: Wang I.-C. , Sether D. M. , Melzer M. J. , Borth W. B. , Hu J. S.
Published: June 2010

Flowering ginger, Alpinia purpurata (Vieill.) K. Schum., is a popular cut flower and tropical landscape plant in Hawaii. In Hawaii, ginger flowers, including red and pink cultivars, are grown as field crops with an estimated annual sales of more than $1.6 million (USD) in 2006 (2). In June 2009, a commercial ginger flower grower from Waimanalo, Oahu, Hawaii reported plants with symptoms that included severe mosaic and stripes on the leaves. Flowers showed significant cupping and browning and growers report a reduction in size and shelf life. Symptomatic ginger was also identified at the Lyon Arboretum in Honolulu. Double-stranded RNAs (dsRNAs) were isolated from pooled leaf samples collected from 42 symptomatic plants at two locations on the island of Oahu to further characterize the pathogen associated with the symptomatic ginger. dsRNAs of approximately 0.7, 1.1, 1.8, 2.2, and 12 kb were present in the extractions from symptomatic plants but not in extractions from asymptomatic plants. Partial cloning and sequence analysis of the dsRNA revealed 95 to 98% nucleotide identity to sequences of P1, HC-Pro, C1, 6K2, VpG, NIb, and CP genes and the 3′ untranslated region (total approximately 6 kb) of Banana bract mosaic virus (BBrMV). Total RNAs were also isolated from the symptomatic and asymptomatic plants from the Waimanalo farm and Lyon Arboretum. These RNA isolations were used in reverse transcription (RT)-PCR with primers Bract N1: 5′-GGRACATCACCAAATTTRAATGG-3′ and Bract NR: 5′-GTGTGCYTCTCTAGCCCTGTT-3′ (1), to amplify a 279-bp conserved region of the coat protein of BBrMV. Amplicons of the appropriate size were obtained from 38 of the symptomatic plants, whereas none were obtained from asymptomatic controls. RT-PCR amplicons of arbitrarily selected samples were cloned into pGEM-T Easy, sequenced, and found to be 99% identical to corresponding sequences of BBrMV. Furthermore, using double-antibody sandwich-ELISA assay and antibodies (3), we developed a system that can specifically detect BBrMV in infected flowering ginger plants and not in healthy appearing ginger. To our knowledge, this is the first report of BBrMV in flowering ginger in Hawaii. Further research is needed to determine if BBrMV infecting ginger poses a threat to banana, edible ginger, and other closely related ornamentals in Hawaii.

References: (1) M. L. Iskra-Caruana et al. J. Virol. Methods 153:223, 2008. (2) Statistics of Hawaii Agriculture (2006). HDOA/USDA (NASS). 96, 2008. (3) J. E. Thomas et al. Phytopathology 87:698, 1997.