bluuebunny/crossref_metadata_2025 huggingface dataset:
Novel cell segmentation and online SVM for cell cycle phase identification in automated microscopy
Authors: Wang Meng , Zhou Xiaobo , Li Fuhai , Huckins Jeremy , King Randall W. , Wong Stephen T.C.
Published: November 2007

Motivation: Automated identification of cell cycle phases captured via fluorescent microscopy is very important for understanding cell cycle and for drug discovery. In this article, we propose a novel cell detection method that utilizes both the intensity and shape information of the cell for better segmentation quality. In contrast to conventional off-line learning algorithms, an Online Support Vector Classifier (OSVC) is thus proposed, which removes support vectors from the old model and assigns new training examples weighted according to their importance to accommodate the ever-changing experimental conditions.

Results: We image three cell lines using fluorescent microscopy under different experiment conditions, including one treated with taxol. Then, we segment and classify the cell types into interphase, prophase, metaphase and anaphase. Experimental results show the effectiveness of the proposed system in image segmentation and cell phase identification.

Availability: The software and test datasets are available from the authors.

Contact:  zhou@crystal.harvard.edu

Supplementary information: Supplementary data are available at Bioinformatics online.
Expression Profiles of Genes Encoded by the Supernumerary Chromosome Controlling AM-Toxin Biosynthesis and Pathogenicity in the Apple Pathotype of Alternaria alternata
Authors: Harimoto Yoshiaki , Hatta Rieko , Kodama Motoichiro , Yamamoto Mikihiro , Otani Hiroshi , Tsuge Takashi
Published: November 2007

The apple pathotype of Alternaria alternata produces host-specific AM-toxin and causes Alternaria blotch of apple. Previously, we cloned two genes, AMT1 and AMT2, required for AM-toxin biosynthesis and found that these genes are encoded by small, supernumerary chromosomes of <1.8 Mb in the apple pathotype strains. Here, we performed expressed sequence tag analysis of the 1.4-Mb chromosome encoding AMT genes in strain IFO8984. A cDNA library was constructed using RNA from AM-toxin–producing cultures. A total of 40,980 clones were screened with the 1.4-Mb chromosome probe, and 196 clones encoded by the chromosome were isolated. Sequence analyses of these clones identified 80 unigenes, including AMT1 and AMT2, and revealed that the functions of 43 (54%) genes are unknown. The expression levels of the 80 genes in AM-toxin–producing and nonproducing cultures were analyzed by real-time quantitative polymerase chain reaction (PCR). Most of the genes were found to be expressed in both cultures at markedly lower levels than the translation elongation factor 1-α gene used as an internal control. Comparison of the expression levels of these genes between two cultures showed that 21 genes, including AMT1 and AMT2, were upregulated (>10-fold) in AM-toxin–producing cultures. Two of the upregulated genes were newly identified to be involved in AM-toxin biosynthesis by the gene disruption experiments and were named AMT3 and AMT4. Thus, the genes upregulated in AM-toxin–producing cultures contain ideal candidates for novel AM-toxin biosynthetic genes.
Regulation of a Virus-Induced Lethal Disease in Tomato Revealed by LongSAGE Analysis
Authors: Irian Saeed , Xu Ping , Dai Xinbin , Zhao Patrick X. , Roossinck Marilyn J.
Published: November 2007

Infection of Cucumber mosaic virus (CMV) and D satellite RNA (satRNA) in tomato plants induces rapid plant death, which has caused catastrophic crop losses. We conducted long serial analysis of gene expression (LongSAGE) in control and virus-infected plants to identify the genes that may be involved in the development of this lethal tomato disease. The transcriptomes were compared between mock-inoculated plants and plants infected with CMV, CMV/D satRNA, or CMV/Dm satRNA (a nonnecrogenic mutant of D satRNA with three mutated nucleotides). The analysis revealed both general and specific changes in the tomato transcriptome after infection with these viruses. A massive transcriptional difference of approximately 400 genes was found between the transcriptomes of CMV/D and CMV/Dm satRNA-infected plants. Particularly, the Long-SAGE data indicated the activation of ethylene synthesis and signaling by CMV/D satRNA infection. Results from inoculation tests with an ethylene-insensitive mutant and treatments with an ethylene action inhibitor further confirmed the role of ethylene in mediating the epinastic leaf symptoms and the secondary cell death in the stem. Results from Northern blot analysis demonstrated the partial contribution of ethylene in the induced defense responses in CMV/D satRNA-infected plants.
Suppression of Infectious TMV Genomes Expressed in Young Transgenic Tobacco Plants
Authors: Siddiqui S. A. , Sarmiento C. , Valkonen S. , Truve E. , Lehto K.
Published: November 2007

Full-length cDNAs of the wild-type (wt) Tobacco mosaic virus (TMV) and of the coat protein gene-deleted (ΔCP) derivative of wt-TMV, under control of the 35S promoter and downstream ribozyme sequence to produce accurate viral transcripts, were transformed to tobacco plants to analyze plant–virus interactions through different stages of plant development. Surprisingly, young wt-TMV transgenics accumulated only very low levels of viral RNA, remained free of symptoms, and were moderately resistant against exogenous inoculations. This early resistance caused significant stress to the plants, as indicated by reduced growth. Approximately 7 to 8 weeks after germination, the resistance was broken and plants developed typical wt-TMV symptoms, with high accumulation of the viral RNAs and proteins. The ΔCP-TMV plants likewise were initially resistant to the endogenous inoculum and were stunted, although to a lesser extent than the wt-TMV plants. The resistance was broken at the same time as in the wt-TMV plants, but the mutant replicated to much lower levels and produced much milder symptoms than the wt virus. TMV-specific small interfering RNAs as well as increased transgene methylation were detected in the plants only after the resistance break, indicating that the resistance in the young plants was not due to RNA silencing.
Duplication of a Conditionally Dispensable Chromosome Carrying Pea Pathogenicity (PEP) Gene Clusters in Nectria haematococca
Authors: Garmaroodi Hamid S. , Taga Masatoki
Published: November 2007

A supernumerary chromosome called a conditionally dispensable chromosome (CDC) is essential for pathogenicity of Nectria haematococca on pea. Among several CDCs discovered in N. haematococca, the PDA1 CDC that harbors the pisatin demethylation gene PDA1 is one of the best-studied CDCs and serves as a model for plant-pathogenic fungi. Although the presence of multiple copies is usual for supernumerary chromosomes in other eukaryotes, this possibility has not been examined well for any CDCs in N. haematococca. In this study, we produced strains with multiple copies of the PDA1 CDC by protoplast fusion and analyzed dosage effects of this chromosome. Using multiple methods, including cytological chromosome counting and fluorescence in situ hybridization, the fusion products between two transformants derived from the same strain that bears a single PDA1 CDC were shown to contain two PDA1 CDCs from both transformants and estimated to be haploid resulting from the deletion of an extra set or sets of A chromosomes in the fused nuclei. In phenotype assays, dosage effects of PDA1 CDC in the fusion products were evident as increased virulence and homoserine-utilizing ability compared with the parents. In a separate fusion experiment, PDA1 CDC accumulated up to four copies in a haploid genome.
The HC-Pro Protein of Potato Virus Y Interacts with NtMinD of Tobacco
Authors: Jin Yongsheng , Ma Dongyuan , Dong Jiangli , Li Daofeng , Deng Changwang , Jin Jingchen , Wang Tao
Published: November 2007

Potato virus Y (PVY) infections often lead to altered numbers of host plant chloroplasts, as well as changes in morphology and inhibited photosynthesis. The multifunctional protein helper component-proteinase, HC-Pro, has been identified in PVY-infected leaf chloroplasts. We used yeast two-hybrid and bimolecular fluorescence complementation assays to demonstrate that HC-Pro can interact with the chloroplast division-related factor NtMinD in yeast and tobacco cells, respectively. In addition, we confirmed that residues 271 to 314 in NtMinD are necessary for its interaction with PVY HC-Pro in a yeast two-hybrid analysis using four NtMinD deletion mutants. These residues are necessary for the dimerization of NtMinD, which plays a vital role in chloroplast division. Thus, PVY HC-Pro may affect NtMinD activity by inhibiting the formation of NtMinD homodimers, and this may interfere with chloroplast division and contribute to changes in the numbers of chloroplast per cell observed in PVY-infected plants.
Salicylic Acid and Jasmonic Acid Signaling Defense Pathways Reduce Natural Bacterial Diversity on Arabidopsis thaliana
Authors: Kniskern Joel M. , Traw M. Brian , Bergelson Joy
Published: November 2007

Terrestrial plants serve as large and diverse habitats for a wide range of pathogenic and nonpathogenic microbes, yet these communities are not well described and little is known about the effects of plant defense on microbial communities in nature. We designed a field experiment to determine how variation in two plant defense signaling pathways affects the size, diversity, and composition of the natural endophytic and epiphytic bacterial communities of Arabidopsis thaliana. To do this, we provide an initial characterization of these bacterial communities in one population in southwestern Michigan, United States, and we compare these two communities among A. thaliana mutants deficient in salicylic acid (SA) and jasmonic acid (JA) signaling defense pathways, controls, and plants with artificially elevated levels of defense. We identified 30 distinct bacterial groups on A. thaliana that differ in colony morphology and 16S rRNA sequence. We show that induction of SA-mediated defenses reduced endophytic bacterial community diversity, whereas plants deficient in JA-mediated defenses experienced greater epiphytic bacterial diversity. Furthermore, there was a positive relationship between total community size and diversity, indicating that relatively susceptible plants should, in general, harbor higher bacterial diversity. This experiment provides novel information about the ecology of bacteria on A. thaliana and demonstrates that variation in two specific plant-signaling defense pathways can influence bacterial diversity on plants.
The N-terminus of the Begomovirus Nuclear Shuttle Protein (BV1) Determines Virulence or Avirulence in Phaseolus vulgaris
Authors: Zhou Y.-C. , Garrido-Ramirez E. R. , Sudarshana M. R. , Yendluri S. , Gilbertson R. L.
Published: November 2007

The BV1 gene of the bipartite Begomovirus genome encodes a nuclear shuttle protein (NSP) that is also an avirulence determinant in common bean. The function of the NSP of two common bean-infecting bipartite begomoviruses, Bean dwarf mosaic virus (BDMV) and Bean golden yellow mosaic virus (BGYMV), was investigated using a series of hybrid DNA-B components expressing chimeric BDMV and BGYMV NSP, and genotypes of the two major common bean gene pools: Andean (cv. Topcrop) and Middle American (cvs. Alpine and UI 114). BDMV DNA-A coinoculated with HBDBG4 (BDMV DNA-B expressing the BGYMV NSP) and HBDBG9 (BDMV DNA-B expressing a chimeric NSP with the N-terminal 1 to 42 amino acids from BGYMV) overcame the BDMV resistance of UI 114. This established that the BDMV NSP is an avirulence determinant in UI 114, and mapped the domain involved in this response to the N-terminus, which is a variable surface-exposed region. BDMV DNA-A coinoculated with HBDBG10, expressing a chimeric NSP with amino acids 43 to 92 from BGYMV, was not infectious, revealing an essential virus-specific domain. In the BGYMV background, the BDMV NSP was a virulence factor in the Andean cv. Topcrop, whereas it was an avirulence factor in the Middle American cultivars, particularly in the absence of the BGYMV NSP. The capsid protein (CP) also played a gene pool–specific role in viral infectivity; it was dispensable for infectivity in the Andean cv. Topcrop, but was required for infectivity of BDMV, BGYMV, and certain hybrid viruses in the Middle American cultivars. Redundancy of the CP and NSP, which are nuclear proteins involved directly or indirectly in viral movement, provides a masking effect that may allow the virus to avoid host defense responses.
Oxalate-Degrading Bacteria Can ProtectArabidopsis thalianaand Crop Plants AgainstBotrytis cinerea
Authors: Schoonbeek Henk-jan , Jacquat-Bovet Anne-Claude , Mascher Fabio , Métraux Jean-Pierre
Published: November 2007

Botrytis cinerea and Sclerotinia sclerotiorum secrete oxalic acid as a pathogenicity factor with a broad action. Consequently, it should be possible to interfere with the infection process by degrading oxalic acid during the interaction of these pathogens with their hosts. We have evaluated the potential of oxalate-degrading bacteria to protect plants against pathogenic fungi. Such bacteria were isolated from agricultural soil and selected on agar plates with Ca-oxalate as the sole carbon source. Four strains were retained with a medium-to-strong protective activity on Arabidopsis thaliana leaves against B. cinerea and S. sclerotiorum. They can provide 30 to 70% protection against fungal infection in different pathosystems, including B. cinerea on A. thaliana, cucumber, grapevine, and tomato. The oxalate-degrading bacteria induced only some marker genes for common plant signaling pathways for defenses, but protective effects were slightly reduced in A. thaliana mutants impaired in the ethylene and jasmonic acid signaling pathways. More detailed studies on the protective mechanism were performed in ox-strain B, identified as Cupriavidus campinensis, by analysis of transposon-tagged mutants that have a reduced ability to degrade oxalic acid.
The Mungbean Yellow Mosaic Begomovirus Transcriptional Activator Protein Transactivates the Viral Promoter-Driven Transgene and Causes Toxicity in Transgenic Tobacco Plants
Authors: Rajeswaran Rajendran , Sunitha Sukumaran , Shivaprasad Padubidri V. , Pooggin Mikhail M. , Hohn Thomas , Veluthambi Karuppannan
Published: November 2007

The Begomovirus transcriptional activator protein (TrAP/AC2/C2) is a multifunctional protein which activates the viral late gene promoters, suppresses gene silencing, and determines pathogenicity. To study TrAP-mediated transactivation of a stably integrated gene, we generated transgenic tobacco plants with a Mungbean yellow mosaic virus (MYMV) AV1 late gene promoter-driven reporter gene and supertransformed them with the MYMV TrAP gene driven by a strong 35S promoter. We obtained a single supertransformed plant with an intact 35S-TrAP gene that activated the reporter gene 2.5-fold. However, 10 of the 11 supertransformed plants did not have the TrAP region of the T-DNA, suggesting the likely toxicity of TrAP in plants. Upon transformation of wild-type tobacco plants with the TrAP gene, six of the seven transgenic plants obtained had truncated T-DNAs which lacked TrAP. One plant, which had the intact TrAP gene, did not express TrAP. The apparent toxic effect of the TrAP transgene was abolished by mutations in its nuclear-localization signal or zinc-finger domain and by deletion of its activation domain. Therefore, all three domains of TrAP, which are required for transactivation and suppression of gene silencing, also are needed for its toxic effect.
A Chitin Synthase with a Myosin-Like Motor Domain Is Essential for Hyphal Growth, Appressorium Differentiation, and Pathogenicity of the Maize Anthracnose Fungus Colletotrichum graminicola
Authors: Werner Stefan , Sugui Janyce A. , Steinberg Gero , Deising Holger B.
Published: November 2007

Chitin synthesis contributes to cell wall biogenesis and is essential for invasion of solid substrata and pathogenicity of filamentous fungi. In contrast to yeasts, filamentous fungi contain up to 10 chitin synthases (CHS), which might reflect overlapping functions and indicate their complex lifestyle. Previous studies have shown that a class VI CHS of the maize anthracnose fungus Colletotrichum graminicola is essential for cell wall synthesis of conidia and vegetative hyphae. Here, we report on cloning and characterization of three additional CHS genes, CgChsI, CgChsIII, and CgChsV, encoding class I, III, and V CHS, respectively. All CHS genes are expressed during vegetative and pathogenic development. ΔCgChsI and ΔCgChsIII mutants did not differ significantly from the wild-type isolate with respect to hyphal growth and pathogenicity. In contrast, null mutants in the CgChsV gene, which encodes a CHS with an N-terminal myosin-like motor domain, are strongly impaired in vegetative growth and pathogenicity. Even in osmotically stabilized media, vegetative hyphae of ΔCgChsV mutants exhibited large balloon-like swellings, appressorial walls appeared to disintegrate during maturation, and infection cells were nonfunctional. Surprisingly, ΔCgChsV mutants were able to form dome-shaped hyphopodia that exerted force and showed host cell wall penetration rates comparable with the wild type. However, infection hyphae that formed within the plant cells exhibited severe swellings and were not able to proceed with plant colonization efficiently. Consequently, ΔCgChsV mutants did not develop macroscopically visible anthracnose disease symptoms and, thus, were nonpathogenic.
The Hypersensitive Response to Tomato leaf curl New Delhi virus Nuclear Shuttle Protein Is Inhibited by Transcriptional Activator Protein
Authors: Hussain Mazhar , Mansoor Shahid , Iram Shazia , Zafar Yusuf , Briddon Rob W.
Published: November 2007

The hypersensitive response (HR) is a common feature of plant disease resistance reactions and a type of programmed cell death (PCD). Many pathogens are able to modulate pathways involved in cell death. In contrast to animal viruses, inhibitors of PCD activity have not been identified for plant-infecting viruses. Previously, we have reported that the nuclear shuttle protein (NSP) of Tomato leaf curl New Delhi virus (ToLCNDV) induces an HR in Nicotiana tabacum and Lycopersicon esculentum plants when expressed under the control of the Cauliflower mosaic virus 35S promoter. However, HR is not evident in plants infected with ToLCNDV, suggesting that the virus encodes a factor (or factors) that counters this response. Analysis of all ToLCNDV-encoded genes pinpointed the transcriptional activator protein (TrAP) as the factor mediating the anti-HR effect. Deletion mutagenesis showed the central region of TrAP, containing a zinc finger domain and nuclear localization signal, to be important in inhibiting the HR. These results demonstrate that TrAP counters HR-induced cell death, the first such activity identified for a plant-infecting virus.
A Developmentally Linked, Dramatic, and Transient Loss of Virus from Roots of Arabidopsis thaliana Plants Infected by Either of Two RNA Viruses
Authors: Lunello Pablo , Mansilla Carmen , Sánchez Flora , Ponz Fernando
Published: November 2007

Possible effects of host developmental stage on the amount of virus present in systemically infected plant tissues hitherto have received little attention. In this study, the pattern of virus accumulation over the plant lifespan has been examined in systemically invaded tissues of Arabidopsis thaliana infected by either of two distinct (+)RNA viruses: Turnip mosaic virus, a member of Potyvirus, and Oilseed rape mosaic virus, a member of Tobamovirus. Quantitative analyses of virus coat protein and virus genomic RNA in roots versus aerial plant parts revealed generally sinusoidal temporal patterns of virus accumulation. In noninoculated leaves, a time period was found during which no virus accumulation was detected. This period was coincident with the approximately 7 days of inflorescence bud formation and differentiation. In roots, virion content reached high levels a few days after inoculation, dropping dramatically during the period of bud formation and quickly recovering after it. These results, together with electron microscopy observations, are consistent with loss of virions due to disassembly. Fluorescence observations of green fluorescent protein-tagged virus-infected root tissue also were consistent with a net loss of virus-specified proteins. Inoculations performed after the emergence of the inflorescence and on A. thaliana flowering-time mutants support the temporal link between observed changes in virus content and inflorescence bud formation. Different host-involving biochemical processes can be invoked to provide mechanistic clues, but no one of them alone seems sufficient to explain the complex patterns of tight temporal regulation of virus accumulation observed in these experiments.
Metabolism of Reactive Oxygen Species Is Attenuated in Leghemoglobin-Deficient Nodules of Lotus japonicus
Authors: Günther Catrin , Schlereth Armin , Udvardi Michael , Ott Thomas
Published: November 2007

Leghemoglobins together with high rates of respiration are believed to be major sources of reactive oxygen species (ROS) in root nodules of leguminous plants. High capacities of antioxidative systems apparently protect this organ from oxidative damage. Using leghemoglobin-RNA interference (LbRNAi) lines of Lotus japonicus, we found that loss of leghemoglobin results in significantly lower H2O2 levels in nodules. Transcript levels and catalytic activities of ascorbate-glutathione cycle enzymes involved in H2O2 detoxification as well as concentrations of reduced ascorbate were also altered in LbRNAi nodules. Thus, symbiotic leghemoglobins contribute significantly to ROS generation in functional nodules.
Dissection of the Barley 2L1.0 Region Carrying the ‘Laevigatum’ Quantitative Resistance Gene to Leaf Rust Using Near-Isogenic Lines (NIL) and subNIL
Authors: Marcel Thierry C. , Aghnoum Reza , Durand Jérôme , Varshney Rajeev K. , Niks Rients E.
Published: November 2007

Partial resistance to leaf rust (Puccinia hordei G. H. Otth) in barley is a quantitative resistance that is not based on hypersensitivity. This resistance hampers haustorium formation, resulting in a long latency period in greenhouse tests. The three most consistent quantitative trait loci (QTL) uncovered in the L94 × ‘Vada’ mapping population were introgressed by marker-assisted backcrossing into the susceptible L94 background to obtain near-isogenic lines (NIL). We also developed the reciprocal Vada-NIL for the susceptibility alleles of those QTL. The QTL Rphq2 affected latency period of P. hordei more than the QTL Rphq3 and Rphq4. The NIL confirmed the contribution of Rphq2 to partial resistance by prolonging the latency period by 28 h on L94-Rphq2 and shortening the latency period by 23 h on Vada-rphq2. On the basis of flanking restriction fragment length polymorphism-based markers, Rphq2 appeared to be located near the telomeric end of the long arm of chromosome 2H, in a physical region of high recombination, making it the target QTL for map-based cloning. Microscopic observations on the NIL confirmed the nonhypersensitive nature of the resistance conferred by Rphq2. A high-resolution genetic map of the Rphq2 region was constructed using a population of 38 subNIL with overlapping L94 introgressions in Vada background across the region. Rphq2 mapped approximately 2 centimorgans (cM) proximal from the MlLa locus. By bulked segregant analysis and use of synteny with rice, we developed additional markers and fine-mapped Rphq2 to a genetic interval of 0.11 cM that corresponds to a stretch of sequence of, at most, 70 kb in rice. Analysis of this rice sequence revealed predicted genes encoding two proteins with unknown function, retrotransposon proteins, peroxidase proteins, and a protein similar to a mitogen-activated protein kinase kinase kinase (MAP3K). Possible homologs of those peroxidases and MAP3K in barley are candidates for the gene that contributes to partial resistance to P. hordei.
The Histology of Hybrid Poplar Clones Inoculated with Septoria musiva
Authors: Weiland Jerry E. , Stanosz Glen R.
Published: November 2007

Septoria musiva causes stem cankers that severely limit production of susceptible hybrid poplars in eastern North America. A field experiment was conducted with resistant clone DN34 and susceptible clone NC11505 in order to (i) identify tissues colonized by the pathogen, (ii) describe tissue responses to S. musiva, and (iii) determine whether tissue responses to S. musiva differed between hybrid poplar clones. Branches of each clone were inoculated by removing the fourth or fifth fully expanded leaf and placing an agar plug colonized by an aggressive isolate of S. musiva over the wound. Seven weeks after inoculation, branches were harvested and prepared for histology. Data from nonwounded control, wounded control, and wounded and inoculated stems were collected and analyzed for effects of clone and treatment. In general, fungal colonization was more extensive in NC11505 and exophylactic and necrophylactic periderms (NPs) of clone DN34 were significantly thicker than those of NC11505, regardless of treatment. The number of NPs produced and the distance from the epidermis to the outermost layer of phellem were significantly affected by the pathogen. Inoculated stems of clone DN34 developed a single NP that formed closer to the wound surface than in wounded controls. In contrast, inoculated stems of NC11505 developed successive NPs and the first NP formed further from the wound surface than in wounded controls. These two host responses to inoculation, as well as measures of exophylactic and necrophylactic periderm thickness, may be useful as markers for the selection of poplar resistant to damage by S. musiva.
A Field Study on the Influence of Verticillium dahliae and Pratylenchus penetrans on Gas Exchange of Potato
Authors: Saeed Ibrahim A. M. , MacGuidwin Ann E. , Rouse Douglas I. , Malek Chris
Published: November 2007

Field experiments were conducted for three consecutive years to study the effects of low populations of Verticillium dahliae and Pratylenchus penetrans on leaf gas exchange of Russet Burbank potato. Treatments were P. penetrans, V. dahliae, the combination of the nematode with the fungus, and a no-pathogen control. Gas exchange was measured nondestructively on young, fully expanded, asymptomatic leaves one to three times per week starting the ninth week after planting. Infection with either pathogen alone had little or no effect on leaf gas exchange parameters. However, co-infection by both pathogens resulted in reduced leaf light use efficiency (mole of CO2 fixed per mole of photon), lower leaf stomatal conductance, lower leaf water use efficiency (mole of CO2 fixed per mole of water used), and increased intercellular CO2 compared with the no-pathogen control. These effects, additive relative to the impact of each pathogen alone, were first observed 9 weeks after inoculation in the first 2 years of the study and 15 weeks after inoculation in the third year.
Occurrence and Distribution of QoI-Resistant Isolates of Colletotrichum cereale from Annual Bluegrass in California
Authors: Wong Francis P. , Midland Sharon L. , de la Cerda Karla A.
Published: November 2007

Turfgrass anthracnose, caused by Colletotrichum cereale (ex. Colletotrichum graminicola), is an important disease of turf used on golf course putting greens. Recent management of the disease has become increasingly difficult, partly due to the possible development of practical resistance to the QoI fungicides. In all, 558 single-conidia isolates of C. cereale were collected from 10 California golf courses, 8 of which had been exposed to QoI fungicides and 2 where no fungicides had been used. Isolates were tested using a mycelial expansion assay on azoxystrobinamended media. For the two nonexposed populations, in vitro 50% effective dose (ED50) values ranged from 0.0060 to 0.089 μg/ml. All isolates from the exposed populations could not be fully inhibited by doses of azoxystrobin as high as 8.0 μg/ml. A subset of these isolates were tested in vitro with the QoI fungicides pyraclostrobin and trifloxystrobin and found to be similar in response, indicating that these isolates were fully cross-resistant to all three fungicides. In greenhouse pot experiments, three isolates nonresponsive to QoI fungicides in vitro were not controlled by label rates of the fungicides. Spore germination assays also were examined; for 10 isolates identified as sensitive by mycelial expansion assays, ED50 values for axoystrobin ranged from 0.0040 to 0.0047 μg/ml; for 25 isolates identified as QoI-resistant, 93 to 100% of the conidia germinated at azoxystrobin concentrations as high as 8.0 μg/ml relative to the nonamended check treatments. Mitochondrial cytochrome b genes from a subset of 15 isolates (12 resistant and 3 sensitive) were partially cloned and sequenced; all resistant isolates had an alanine substitution that corresponded to position 143 of the gene product. These results indicate that QoI resistance is present in California populations of C. cereale and is contributing to the difficulty in controlling this disease.
Sensitivity Distributions of California Populations of Colletotrichum cereale to the DMI Fungicides Propiconazole, Myclobutanil, Tebuconazole, and Triadimefon
Authors: Wong Francis P. , Midland Sharon L.
Published: November 2007

The baseline sensitivity of a California population of Colletotrichum cereale (turfgrass anthracnose) to the sterol demethylation inhibitor (DMI) fungicide propiconazole was determined using an in vitro assay with known reproducibility. The 50% effective dose (ED50) values for propiconazole for a nonexposed, baseline population ranged from 0.025 to 0.35 μg/ml with a mean of 0.14 μg/ml. Examination of two DMI-exposed populations indicated an approximate increase of 6.5× in mean ED50 values. In vivo testing of two isolates with ED50 values of propiconazole at 0.15 and 0.90 μg/ml indicated reduced control for the less sensitive isolate by propiconazole at rates ≤38 μg/ml. It was determined that single discriminatory dose testing in vitro with propiconazole at 0.50 μg/ml could differentiate sensitive and resistant isolates. Using this dose, six additional populations were tested and DMI-exposed populations were found to be three to nine times less sensitive compared with the baseline population. Two populations were assayed for sensitivity to myclobutanil, tebuconazole, and triadimefon. Mean ED50 values for a nonexposed population were 0.72, 0.082, and 5.6 μg/ml, respectively; for a DMI-exposed population, mean ED50 values were 3.8, 0.35, and 18 μg/ml, respectively. This work provides information on the development of DMI resistance in populations of C. cereale in California and methodologies for future resistance monitoring for this pathogen.
Genetic Diversity in Populations of Phytophthora citricola Associated with Horticultural Crops in California
Authors: Bhat R. G. , Browne G. T.
Published: November 2007

California populations of the plant pathogen Phytophthora citricola were examined for amplified fragment length polymorphism (AFLP), pathogenicity on almond, and sensitivity to mefenoxam. The characterizations of AFLP variation and mefenoxam sensitivity were based on 86 isolates (44 from almond, 11 from avocado, 3 from strawberry, 18 from walnut, and 10 from six other hosts). Cluster analysis of the AFLP data using the unweighted pair group method indicated a high level of genetic diversity among the isolates, and four main clusters were identified—one dominated by isolates from almond, another including all isolates from avocado, and two including isolates from several hosts other than avocado. Analysis of molecular variance revealed that 38.4 and 24.9% of the AFLP variation were associated with host and geographical factors, respectively. Of 24 isolates, including those from almond, avocado, strawberry, and walnut, 22 were aggressive on almond shoots; there was no evidence of host specificity. All but 1 of the 86 isolates grew at different rates on V8 juice medium amended with mefenoxam at 1 ppm, indicating partial tolerance to the fungicide. Isolates of P. citricola from California populations are genetically diverse, and much of the variation is associated with host and geography. These populations are all potentially pathogenic on almond and tolerant to mefenoxam.
Intergenic Spacer–RFLP Analysis and Direct Quantification of AustralianFusarium oxysporumf. sp.vasinfectumIsolates from Soil and Infected Cotton Tissues
Authors: Zambounis A. G. , Paplomatas E. , Tsaftaris A. S.
Published: November 2007

Fusarium wilt of cotton, caused by Fusarium oxysporum f. sp. vasinfectum, can have devastating effects on the vascular system of cotton plants and is a major threat to cotton production throughout the world. Accurate characterization and improved detection of these pathogenic isolates is needed for the implementation of a disease prevention program and the development of disease management strategies. Polymerase chain reaction (PCR) amplification of the ribosomal intergenic spacer (IGS) regions combined with digestion with three restriction enzymes (AluI, HaeIII, RsaI) resulted in three unique restriction profiles (IGS-restriction fragment length polymorphism [RFLP] haplotypes) for Australian F. oxysporum f. sp. vasinfectum isolates, which were capable of distinguishing them from other formae speciales of F. oxysporum. Furthermore, a portion of the IGS region flanking the 5′ end was sequenced and single nucleotide polymorphisms (SNPs) were revealed. Using these sequence data, two specific real-time PCR-based assays were developed for the absolute quantification of genomic DNA from isolates obtained from soil substrates and infected cotton tissues. Standard curves of real-time PCR-based assays showed a linear relation (R2= 0.993 to 0.994) between log values of fungal genomic DNA and real-time PCR cycle thresholds. Using these assays, it was possible to detect fungal DNA as low as 5 pg/μl. The detection sensitivity for inoculum added to sterile soils was lower than 104conidia/g soil. In plant samples, the quantified fungal DNA varied from 30 pg to 1 ng/100 ng of total plant genomic DNA.
Modifications of PARP Medium Using Fluazinam, Miconazole, and Nystatin for Detection of Pythium spp. in Soil
Authors: Morita Y. , Tojo M.
Published: November 2007

The standard Pythium selective medium PARP (pimaricin + ampicillin + rifampicin + pentachloronitrobenzene [PCNB] agar), was modified by replacing PCNB and pimaricin with other antifungal agents. Several antifungal agents such as fluazinam, miconazole, 2,4,5,6-tetrachloroisophthalonitrile (TPN), iminoctadine triacetate, tolclofos-methyl, captan, and nystatin, initially were screened for effects on Pythium growth. Based on these results, the following three media were developed: PARF (pimaricin + ampicillin + rifampicin + fluazinam agar), NARF (nystatin + ampicillin + rifampicin + fluazinam agar), and NARM (nystatin + ampicillin + rifampicin + miconazole agar). New media were comparable with PARP on yield of naturally occurring Pythium spp. from two different types of soil using the soil-dilution plating technique. PARF and NARF were significantly better than PARP on inhibition of non-pythiaceous microbes on the soil-dilution plates, but were significantly lower than PARP on the rate of mycelial growth of six of eight isolates belonging to seven species of Pythium. NARM was equivalent to PARP on inhibition of non-pythiaceous microbes except for Fusarium oxysporum, and was significantly better than PARP on rate of mycelial growth of five of eight isolates of Pythium.
Efficacy of Pre- and Postinoculation Application of Fungicides to Expanding Young Citrus Leaves for Control of Melanose, Scab, and Alternaria Brown Spot
Authors: Mondal S. N. , Vicent A. , Reis R. F. , Timmer L. W.
Published: November 2007

In greenhouse trials, copper hydroxide, pyraclostrobin, and famoxadone were applied to actively growing young citrus seedlings to determine the duration of protection of young leaves provided by these fungicides against melanose, caused by Diaporthe citri, citrus scab, caused by Elsinoe fawcettii, and Alternaria brown spot, caused by Alternaria alternata. Fungicides were applied to different sets of potted plants of grapefruit for control of melanose, of rough lemon for control of scab, and of Dancy tangerine for control of Alternaria brown spot 1 to 6 days prior to inoculation, as well as on the day of inoculation. Leaf area of treated shoots was estimated on the day of fungicide application and the day of inoculation and disease severity evaluated subsequently. In most cases, copper hydroxide and famoxadone provided at least 50% control of all three diseases for only about 2 days after application. Generally, there was little or no disease control when the products were applied 4 or more days before inoculation. In contrast, pyraclostrobin usually provided a high level of control of all three diseases when applied up to 5 days prior to inoculation. The level of disease control decreased as the interval between a fungicide application and inoculation increased and the relationship between disease control and leaf expansion best fit a quadratic equation. Effective disease control was observed with copper hydroxide and famoxadone until leaf area had increased by 100 to 200%, whereas control with pyraclostrobin was observed up to 400 to 500% increase in leaf area. In postinoculation tests with scab and melanose, pyraclostrobin provided high levels of disease control (>75%) when applied up to 2 days after inoculation, whereas copper hydroxide and famoxadone had minimal postinoculation activity. Applications of pyraclostrobin to the spring flush growth of citrus trees are much more likely to provide control of melanose, scab, and Alternaria brown spot than those of famoxadone or copper hydroxide.
Preplant Biofumigation with Sorghum or Methyl Bromide Compared for Managing Criconemoides xenoplax in a Young Peach Orchard
Authors: Nyczepir A. P. , Rodriguez-Kabana R.
Published: November 2007

Sorghum as a preplant green manure biofumigant management system of Criconemoides xenoplax was investigated from 1998 to 2003. The study was conducted on a site infested with C. xenoplax and having a previous history of peach tree short life (PTSL). Plots consisted of five preplant treatments: (i) nonfumigated soil without tarp and urea, (ii) nonfumigated soil with tarp and urea, (iii) sorghum as a green manure without tarp and urea, (iv) sorghum as a green manure with tarp and urea, and (v) preplant methyl bromide fumigation. Sorghum as a green manure with and without tarp was comparable with methyl bromide fumigation in suppressing the population of C. xenoplax in the early stages of this experiment. Nematode population densities were suppressed 11 months longer in sorghum with tarp and urea plots than in sorghum without tarp and urea plots. However, nematode population densities in sorghum with tarp and urea plots were not suppressed as long as in fumigated methyl bromide plots (19 versus 24 months, respectively).
Macroarray Detection of Solanaceous Plant Pathogens in the Fusarium solani Species Complex
Authors: Zhang Ning , Geiser David M. , Smart Christine D.
Published: November 2007

Members of the Fusarium solani species complex (FSSC), which are morphologically similar but have more than 45 distinct lineages, were chosen as targets for the development of a macroarray detection system that would be broadly adaptable. Thirty-three oligonucleotides (17 to 27 mers) were designed from the internal transcribed spacer (ITS) of the ribosomal RNA genes of 17 FSSC isolates, which belong to 12 phylogenetically closely related species. Of the 33 oligonucleotides on the array, 21 were useful in discriminating all 12 species, some of which had only a single nucleotide difference among them. The high specificity of this method was achieved by optimizing the hybridization temperature and oligo probe length, which had a more substantial effect on the array performance than the melting temperature and the DNA G+C content (G/C%) of the probes. The array was validated by testing inoculated greenhouse samples and diseased field plant samples.
Effect of Oriental Medicinal Plant Extracts on Spore Germination ofAlternaria brassicicolaand Nature of Inhibitory Substances from Speedweed
Authors: Ho Wang-Ching , Wu Tsung-Yu , Su Huey-Jen , Ko Wen-Hsiung
Published: November 2007

Among 65 species of oriental medicinal plants tested, 39 contained substances inhibitory to conidial germination of Alternaria brassicicola, with the most inhibitory extract from speedweed (Polygonum perfoliatum). The inhibitory substances in dried speedweed were insoluble in water. However, the inhibitors were readily extracted with ethanol or methanol, but not with acetone, ether, or chloroform. The ethanol extract was very effective in controlling black leaf spot of spoon cabbage, Brassica campestris subsp. chinensis, caused by A. brassicicola. The inhibitory effect of the extract was not affected by treatment with anion exchange resins, but was partially reduced by cation exchange resins, indicating the presence of two inhibitory substances in the extract, one with a positive charge and the other with no charge on its molecule. The inhibitory substances in the extract were dialyzable with molecular weight minimums of 10,000 or 1,000, but not 500 or 100, suggesting that both inhibitors have molecular weights between 500 and 1,000.
Effects of Cultivars and Fungicides on Rice Sheath Blight, Yield, and Quality
Authors: Groth D. E. , Bond J. A.
Published: November 2007

The development of sheath blight (Rhizoctonia solani)-resistant rice (Oryza sativa) cultivars will allow producers to use less fungicide and to avoid significant reductions in grain and milling yields. Among cultivars currently in cultivation in the southern United States rice-producing region, sheath blight resistance levels range from very susceptible to moderately susceptible. A study was conducted to determine the response of cultivars with different levels of susceptibility to sheath blight inoculations and fungicide applications and to determine the impact of sheath blight disease development on rice yield and quality. Sheath blight epidemics in field plots were initiated by inoculation at the panicle differentiation growth stage from 2003 through 2005. Azoxystrobin at 0.17 kg a.i. ha–1 and flutolanil at 0.56 kg a.i. ha–1 were applied in sequential applications at midboot and 50 to 70% heading. Inoculation significantly increased sheath blight severity and incidence and caused yield losses of 4% in moderately susceptible cv. Francis to 21% in very susceptible cv. Cocodrie. Milling yield was affected to a lesser extent. Fungicide treatments reduced sheath blight incidence and severity regardless of cultivar. Azoxystrobin was more effective than flutolanil in minimizing yield loss due to sheath blight in all cultivars except Francis.
Detection and Quantification of Rhizoctonia solani AG-1 IA, the Rice Sheath Blight Pathogen, in Rice Using Real-Time PCR
Authors: Sayler Ronald J. , Yang Yinong
Published: November 2007

Rhizoctonia solani Kühn is the causal organism of sheath blight, a major rice disease worldwide that severely impairs yield and quality. It is difficult to identify the pathogen in the early phase of the infection and to accurately quantify the fungal development based on visual inspection. Therefore, a rapid and reliable method is advantageous for the detection and quantification of the pathogen causing this important rice disease. In this study, a real-time, quantitative polymerase chain reaction (QPCR) assay was developed to detect and quantify R. solani AG-1 IA DNA from infected rice plants. A specific primer pair was designed based on the internal transcribed spacer region of the fungal ribosomal DNA. The specific detection of R. solani DNA was successful with quantities as low as 1 pg. The QPCR assay could be used for detecting the rice sheath blight pathogen, quantifying fungal aggressiveness, and evaluating the resistance level of rice cultivars.
A PCR-Based Assay for Detection of Puccinia striiformis f. sp. tritici in Wheat
Authors: Zhao J. , Wang X. J. , Chen C. Q. , Huang L. L. , Kang Z. S.
Published: November 2007

Monitoring the pathogenic fungus of wheat stripe rust, Puccinia striiformis f. sp. tritici, plays a key role in effective control of the disease. In the present study, we developed a specific and sensitive polymerase chain reaction (PCR) assay for detecting the pathogen in wheat (Triticum aestivum) leaves. A pair of primers (PSF and PSR) was designed based on the internal transcribed spacer (ITS) region sequence of P. striiformis f. sp. tritici. PCR products that were amplified with universal primers ITS1 and ITS4 were cloned into pGEM-T Easy vectors and sequenced. The ITS sequence was compared with those of P. striiformis f. sp. tritici, P. triticina, P. graminis f. sp. tritici, Blumeria graminis f. sp. tritici, Fusarium graminearum, Rhizoctonia cerealis, and Bipolaris sorokiniana, which are associated with early symptoms of foliar diseases on wheat. Specificity of the primers was tested in the PCR assays using DNA extracted from all tested P. striiformis f. sp. tritici isolates, other fungal species, and healthy and infected wheat leaves sampled around stripe rust foci in wheat fields, different days after inoculation with P. striiformis f. sp. tritici, as well as asymptomatic wheat leaves sampled around stripe rust foci in the fields. A PCR product of 169 bp was amplified from DNA of all P. striiformis f. sp. tritici isolates. The primers did not amplify DNA from the other tested fungal species. The pathogen was detected from asymptomatic wheat leaves inoculated with P. striiformis f. sp. tritici under greenhouse conditions, as well as leaves sampled around stripe rust foci in wheat fields. Under optimum conditions, the PCR assay was highly sensitive and required only 0.1 pg of the target DNA for a detectable and reliable amplification with the PSF and PSR primers.
Timing of Fungicide Applications for Control of Husk Spot Caused by Pseudocercospora macadamiae in Macadamia
Authors: Akinsanmi O. A. , Miles A. K. , Drenth A.
Published: November 2007

Pseudocercospora macadamiae is an important pathogen of macadamia in Australia, causing a disease known as husk spot. Growers strive to control the disease with a number of carbendazim and copper treatments. The aim of this study was to consider the macadamia fruit developmental stage at which fungicide application is most effective against husk spot, and whether application of copper-only applications at full-size fruit developmental stage toward the end of the season contributed to effective disease control. Fungicides were applied to macadamia trees at four developmental stages in three orchards in two subsequent production seasons. The effects of the treatments on disease incidence and severity were quantified using area under disease progress curve (AUDPC) and logistic regression models. Although disease incidence varied between cultivars, incidence and severity on cv. A16 showed consistent differences between the treatments. Most significant reduction in husk spot incidence occurred when spraying commenced at match-head sized-fruit developmental stage. All treatments significantly reduced husk spot incidence and severity compared with the untreated controls, and a significant positive linear relationship (R2 = 73%) between AUDPC and severity showed that timing of the first fungicide application is important for effective disease control. Application of fungicide at full-size fruit stage reduced disease incidence but had no impact on premature fruit drop.