title stringlengths 0 1.13k | abstract stringlengths 1 15.7k | PMID int64 22 36.5M |
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The Complete Nucleotide Sequence and Biotype Variability of Papaya leaf distortion mosaic virus. | ABSTRACT The complete nucleotide sequence of the genome of Papaya leaf distortion mosaic virus (PLDMV) was determined. The viral RNA genome of strain LDM (leaf distortion mosaic) comprised 10,153 nucleotides, excluding the poly(A) tail, and contained one long open reading frame encoding a polyprotein of 3,269 amino acids (molecular weight 373,347). The polyprotein contained nine putative proteolytic cleavage sites and some motifs conserved in other potyviral polyproteins with 44 to 50% identities, indicating that PLDMV is a distinct species in the genus Potyvirus. Like the W biotype of Papaya ringspot virus (PRSV), the non-papaya-infecting biotype of PLDMV (PLDMV-C) was found in plants of the family Cucurbitaceae. The coat protein (CP) sequence of PLDMV-C in naturally infected-Trichosanthes bracteata was compared with those of three strains of the P biotype (PLDMV-P), LDM and two additional strains M (mosaic) and YM (yellow mosaic), which are biologically different from each other. The CP sequences of three strains of PLDMV-P share high identities of 95 to 97%, while they share lower identities of 88 to 89% with that of PLDMV-C. Significant changes in hydrophobicity and a deletion of two amino acids at the N-terminal region of the CP of PLDMV-C were observed. The finding of two biotypes of PLDMV implies the possibility that the papaya-infecting biotype evolved from the cucurbitaceae-infecting potyvirus, as has been previously suggested for PRSV. In addition, a similar evolutionary event acquiring infectivity to papaya may arise frequently in viruses in the family Cucurbitaceae. | 18,943,981 |
Different Resistance Mechanisms of Medicago truncatula Ecotypes Against the Rust Fungus Uromyces striatus. | ABSTRACT A pathosystem consisting of the model plant Medicago truncatula and the rust fungus Uromyces striatus was characterized. From a collection of 113 mostly European accessions of M. truncatula, the vast majority were found to be susceptible to U. striatus, whereas 5 accessions showed strong resistance reactions. Stomatal surface characteristics, even if partly occluded, did not interfere with the ability of U. striatus germ tubes to infect. After penetration, the resistant ecotypes reacted with various degrees of cell death during different stages of haustorial establishment. Whereas four ecotypes showed a typical hypersensitive reaction by developing necrotic lesions, one ecotype (F11.008) exhibited a prehaustorial type of defense without hypersensitive response. This ecotype may be used as a source of nonhost-type of resistance against U. striatus. | 18,943,984 |
A one-step reverse transcription-polymerase chain reaction system for the simultaneous detection and identification of multiple tospovirus infections. | ABSTRACT A one-step reverse transcription-polymerase chain reaction (RT-PCR) method has been developed for the simultaneous detection and identification of multiple tospoviruses that infect plants. The RT-PCR system is composed of six primers in a single tube: a universal degenerate primer and five virus species-specific primers. Amplifications resulted in an 848-bp PCR product for Watermelon silver mottle virus, 709-bp for Tomato spotted wilt virus, 589-bp for Impatiens necrotic spot virus, 511-bp for Melon yellow spot virus, and a 459-bp amplicon for Iris yellow spot virus. This system enables the simultaneous detection of at least three types of tospovirus infections, in addition to their species identities, from five possible tospoviruses studied, on the basis of their S RNA combinations. This multiplex RT-PCR system was applied to the detection of tospovirus in ornamental crops cultivated in fields and shows potential for epidemiological studies. | 18,943,986 |
Inducible Expression of a Phytolacca heterotepala Ribosome-Inactivating Protein Leads to Enhanced Resistance Against Major Fungal Pathogens in Tobacco. | ABSTRACT Plant genetic engineering has long been considered a valuable tool to fight fungal pathogens because it would limit the economically costly and environmentally undesirable chemical methods of disease control. Ribosome-inactivating proteins (RIPs) are potentially useful for plant defense considering their antiviral and antimicrobial activities but their use is limited by their cytotoxic activity. A new gene coding for an RIP isolated from leaves of Phytolacca heterotepala was expressed in tobacco under the control of the wound-inducible promoter of the bean polygalacturonase-inhibiting protein I gene to increase resistance against different fungal pathogens, because an individual RIP isolated from P. heterotepala showed direct antifungal toxicity. Phenotypically normal transgenic lines infected with Alternaria alternata and Botrytis cinerea showed a significant reduction of leaf damage while reverse transcription-polymerase chain reaction and western analysis indicated the expression of the RIP transgene upon wounding and pathogen attack. This work demonstrates that use of a wound-inducible promoter is useful to limit the accumulation of antimicrobial phytotoxic proteins only in infected areas and that the controlled expression of the PhRIP I gene can be very effective to control fungal pathogens with different phytopathogenic actions. | 18,943,992 |
Population Structure of Mycosphaerella graminicola: From Lesions to Continents. | ABSTRACT The genetic structure of field populations of Mycosphaerella graminicola was determined across a hierarchy of spatial scales using restriction fragment length polymorphism markers. The hierarchical gene diversity analysis included 1,098 isolates from seven field populations. Spatial scales ranged from millimeters to thousands of kilometers, including comparisons within and among lesions, within and among fields, and within and among regions and continents. At the smallest spatial scale, microtransect sampling was used to determine the spatial distribution of 15 genotypes found among 158 isolates sampled from five individual lesions. Each lesion had two to six different genotypes including both mating types in four of the five lesions, but in most cases a lesion was composed of one or two genotypes that occupied the majority of the lesion, with other rare genotypes interspersed among the common genotypes. The majority (77%) of gene diversity was distributed within plots ranging from approximately 1 to 9 m(2) in size. Genotype diversity (G / N) within fields for the Swiss, Texas, and Israeli fields was high, ranging from 79 to 100% of maximum possible values. Low population differentiation was indicated by the low G(ST) values among populations, suggesting a corresponding high degree of gene flow among these populations. At the largest spatial scale, populations from Switzerland, Israel, Oregon, and Texas were compared. Population differentiation among these populations was low (G(ST) = 0.05), and genetic identity between populations was high. A low but significant correlation between genetic and geographic distance among populations was found (r = -0.47, P = 0.012), suggesting that these populations probably have not reached an equilibrium between gene flow and genetic drift. Gene flow on a regional level can be reduced by implementing strategies, such as improved stubble management that minimize the production of ascospores. The possibility of high levels of gene flow on a regional level indicates a significant potential risk for the regional spread of mutant alleles that enable fungicide resistance or the breakdown of resistance genes. | 18,944,019 |
Internal Transcribed Spacer Regions 1 and 2 and Random Amplified Polymorphic DNA Analysis of Didymella bryoniae and Related Phoma Species Isolated from Cucurbits. | ABSTRACT Didymella bryoniae (anamorph Phoma cucurbitacearum) is the causal agent of gummy stem blight, although other Phoma species are often isolated from cucurbit plants exhibiting symptoms of the disease. The molecular and phylogenetic relationships between D. bryoniae and these Phoma species are unknown. Isolates of D. bryoniae and Phoma obtained from cucurbits grown at various geographical locations in the United States were subjected to random amplified polymorphic DNA (RAPD) analysis and internal transcribed spacer (ITS) sequence analysis (ITS-1 and ITS-2) to determine the molecular and phylogenetic relationships within and between these fungi. Using RAPD fingerprinting, 59 isolates were placed into four phylogenetic groups, designated RAPD group (RG) I, RG II, RG III, and RG IV. D. bryoniae isolates clustered in either RG I (33 isolates), RG II (12 isolates), or RG IV (one isolate), whereas all 13 Phoma isolates clustered to RG III. There was greater than 99% sequence identity in the ITS-1 and ITS-2 regions between isolates in RG I and RG II, whereas isolates in RG III, P. medicaginis ATCC 64481, and P. exigua ATCC 14728 clustered separately. On muskmelon seedlings, a subset of RG I isolates were highly virulent (mean disease severity was 71%), RG II and RG IV isolates were slightly virulent (mean disease severity was 4%), and RG III isolates were nonpathogenic (disease severity was 0% for all isolates). The ITS sequences indicate that RG I and RG II are both D. bryoniae, but RAPD fingerprints and pathogenicity indicate that they represent two different molecular and virulence subgroups. | 18,944,025 |
Risk Analysis of Brown Rot Blossom Blight of Prune Caused by Monilinia fructicola. | Experiments under controlled environmental conditions were conducted during bloom of prune (Prunus domestica, L.) in 1999 and 2000 to assess the effects of inoculum concentration (IC), wetness duration (WD), temperature, and bloom stages on development of brown rot blossom blight of prunes. Branches from trees of a prune orchard were inoculated with Monilinia fructicola at different bloom stages and incubated at different temperatures with different periods of WD. The proportion of blighted blossoms (PBB) for each inoculated branch was determined. Bloom stage, IC, temperature, and WD significantly affected blossom blight of prunes. PBB at popcorn and full bloom stages was significantly greater than PBB at later bloom stages (P </=0.05). The optimal temperatures for blossom blight development were 22 to 26 degrees C, and Gaussian functions were used to describe the relationship between PBB and temperature. PBB linearly increased with increased IC. Linear regressions of PBB on WD were obtained for each combination of bloom stage, IC, and temperature. The parameters of these regressions were used in a computer program to produce the possible maximum PBB with 90% probability (PBB(90)) using stochastic simulations. Early bloom stages with a higher IC at temperatures from 20 to 25 degrees C were associated with more severe blossom blight than late stages with a lower IC at nonoptimal temperatures. Blossom blight did not occur at <10 or >30 degrees C and less than 4-h WD. However, longer than 4-h WD linearly increased incidence of blossom blight. A risk assessment table of blossom blight was produced for different environmental conditions to guide the control of prune brown rot. | 18,944,033 |
Plant pathogen culture collections: it takes a village to preserve these resources vital to the advancement of agricultural security and plant pathology. | ABSTRACT Plant pathogen culture collections are essential resources in our fight against plant disease and for connecting discoveries of the present with established knowledge of the past. However, available infrastructure in support of culture collections is in serious need of improvement, and we continually face the risk of losing many of these collections. As novel and reemerging plant pathogens threaten agriculture, their timely identification and monitoring depends on rapid access to cultures representing the known diversity of plant pathogens along with genotypic, phenotypic, and epidemiological data associated with them. Archiving such data in a format that can be easily accessed and searched is essential for rapid assessment of potential risk and can help track the change and movement of pathogens. The underexplored pathogen diversity in nature further underscores the importance of cataloguing pathogen cultures. Realizing the potential of pathogen genomics as a foundation for developing effective disease control also hinges on how effectively we use the sequenced isolate as a reference to understand the genetic and phenotypic diversity within a pathogen species. In this letter, we propose a number of measures for improving pathogen culture collections. | 18,944,046 |
Clarification of the Etiology of Glomerella Leaf Spot and Bitter Rot of Apple Caused by Colletotrichum spp. Based on Morphology and Genetic, Molecular, and Pathogenicity Tests. | ABSTRACT Morphological characteristics and vegetative compatibility groups (VCGs) of 486 isolates of Glomerella cingulata, Colletotrichum gloeosporioides, and C. acutatum collected from apple leaves with Glomerella leaf spot (GLS) symptoms and fruit with bitter rot symptoms in the United States and Brazil were studied. From this collection, 155 isolates of G. cingulata (93 from fruit, 61 from leaves, and 1 from buds), 42 isolates of C. gloeosporioides from fruit, and 14 isolates of C. acutatum (10 from fruit and 4 from leaves) were studied using mitochondrial (mt)DNA restriction fragment length polymorphism (RFLP) haplotypes. A subset of 24 isolates was studied by examining the sequence of a 200-bp intron of the glyceraldehyde 3-phosphate dehydrogenase (GDPH) nuclear gene. In addition, 98 isolates were tested for pathogenicity on leaves of cvs. Gala and Golden Delicious in the greenhouse, and 24 isolates were tested for pathogenicity on fruit of cv. Gala in growth chambers. In total, 238 and 225 isolates of G. cingulata were separated into four distinct morphological types and six VCGs, respectively. Five morphological types and six VCGs were identified among 74 and 36 isolates of C. gloeosporioides, respectively. Three morphological types and four VCGs were identified among 74 and 23 isolates of C. acutatum, respectively. Seven different mtDNA RFLP haplotypes were observed within isolates of G. cingulata, two within isolates of C. gloeosporioides, and two within isolates of C. acutatum. Phylogenetic trees, inferred based on maximum likelihood and maximum parsimony methods using the intron sequence, produced similar topologies. Each species was separated into distinct groups. All isolates tested were pathogenic on fruit, though only isolates with specific VCGs and haplotypes were pathogenic to leaves. Vegetative compatibility was a better tool than molecular characters for distinguishing isolates of G. cingulata pathogenic on both leaves and fruit from the ones pathogenic only on fruit. Isolates of G. cingulata capable of causing both GLS and bitter rot were included in haplotypes and groups based on the sequence analysis of the 200-bp intron that also included isolates capable of causing bitter rot only. Additionally, isolates of G. cingulata from the United States and Brazil which cause GLS were included in different haplotypes and sequence analysis groups. Therefore, one hypothesis is that isolates of G. cingulata from the United States capable of causing both GLS on foliage and bitter rot on fruit may have arisen independently of Brazilian isolates of G. cingulata capable of causing both GLS and bitter rot, and the two groups of isolates may represent distinct populations. | 18,944,054 |
Activity of the Antifungal Protein from Aspergillus giganteus Against Botrytis cinerea. | ABSTRACT Botrytis blight (gray mold), caused by Botrytis cinerea, is one of the most widely distributed diseases of ornamental plants. In geranium plants, gray mold is responsible for important losses in production. The mold Aspergillus giganteus is known to produce and secrete a basic low-molecular-weight protein, the antifungal protein (AFP). Here, the antifungal properties of the Aspergillus AFP against various B. cinerea isolates obtained from naturally infected geranium plants were investigated. AFP strongly inhibited mycelial growth as well as conidial germination of B. cinerea. Microscopic observations of fungal cultures treated with AFP revealed reduced hyphal elongation and swollen hyphal tips. Washout experiments in which B. cinerea was incubated with AFP for different periods of time and then washed away revealed a fungicidal activity of AFP. Application of AFP on geranium plants protected leaves against Botrytis infection. Cecropin A also was active against this pathogen. An additive effect against the fungus was observed when AFP was combined with cecropin A. These results are discussed in relation to the potential of the afp gene to enhance crop protection against B. cinerea diseases. | 18,944,061 |
Ecological implications from a molecular analysis of phytoplasmas involved in an aster yellows epidemic in various crops in Texas. | ABSTRACT In the spring of 2000, an aster yellows (AY) epidemic occurred in carrot crops in the Winter Garden region of southwestern Texas. A survey revealed that vegetable crops, including cabbage, onion, parsley, and dill, and some weeds also were infected by AY phytoplasmas. Nested polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis of PCR-amplified phytoplasma 16S rDNA were employed for the detection and identification of phytoplasmas associated with these crops and weeds. Phytoplasmas belonging to two subgroups, 16SrI-A and 16SrI-B, in the AY group (16SrI), were predominantly detected in infected plants. Carrot, parsley, and dill were infected with both subgroups. Onion and three species of weeds (prickly lettuce, lazy daisy, and false ragweed) were predominantly or exclusively infected by subgroup 16SrI-A phytoplasma strains, while cabbage was infected by subgroup 16SrI-B phytoplasmas. Both types of phytoplasmas were detected in three leafhopper species, Macrosteles fascifrons, Scaphytopius irroratus, and Ceratagallia abrupta, commonly present in this region during the period of the epidemic. Mixed infections were very common in individual carrot, parsley, and dill plants and in individual leafhoppers. Sequence and phylogenetic analyses of 16S rDNA and ribosomal protein (rp) gene sequences indicated that phytoplasma strains within subgroup 16SrI-A or subgroup 16SrI-B, detected in various plant species and putative insect vectors, were highly homogeneous. However, based on rp sequences, two rpI subgroups were identified within the subgroup 16SrI-A strain cluster. The majority of subgroup 16SrI-A phytoplasma strains were classified as rp subgroup rpI-A, but phytoplasma strains detected in one onion sample and two leafhoppers (M. fascifrons and C. abrupta) were different and classified as a new rp subgroup, rpI-N. The degree of genetic homogeneity of the phytoplasmas involved in the epidemic suggested that the phytoplasmas came from the same pool and that all three leafhopper species may have been involved in the epidemic. The different phytoplasma population profiles present in various crops may be attributed to the ecological constraints as a result of the vector-phytoplasma-plant three-way interaction. | 18,944,064 |
Use of a Nitrate-Nonutilizing Mutant and Selective Media to Examine Population Dynamics of Fusarium oxysporum f. sp. spinaciae in Soil. | ABSTRACT Determining the population density of the spinach wilt pathogen Fusarium oxysporum f. sp. spinaciae in soil with conventional Fusarium-selective media is quite difficult because nonpathogenic strains of F. oxysporum also grow on those media and are indistinguishable from the pathogen. Therefore, a nitrate-nonutilizing (nit) mutant of the pathogen and corresponding selective media were tested in an experimental approach to determine the population density of the pathogen. Colony forming units of the pathogen were countable after soil-dilution plating onto nit mutant-selective media MMCPA, CMP, and CGMBP. Colony forming units of wild-type Fusarium spp. were countable using a wildtype Fusarium-selective medium, GMBP. By combining nit mutant- and wild-type-selective media, the population densities of pathogenic and nonpathogenic F. oxysporum in the same soil could be measured selectively. This method was useful in studying population dynamics of the pathogen after different soil treatments. Soil disinfested with hot water or chloropicrin was amended with the nit mutant pathogen, and subsequent changes in population densities of the pathogen were compared with those in nontreated field soil. The pathogen rapidly proliferated in disinfested soil and wilt developed faster than in nontreated soil. When a nonpathogenic isolate of F. oxysporum was added at high density to sterilized soil prior to the pathogen, growth of the pathogen was greatly suppressed. Nonpathogenic F. oxysporum could not, however, reduce the density of preexisting pathogen. | 18,944,103 |
Depth Distribution of Rotylenchulus reniformis Under Different Tillage and Crop Sequence Systems. | ABSTRACT The population density of the reniform nematode, Rotylenchulus reniformis, was monitored at depths of 0 to 30, 30 to 60, 60 to 90, and 90 to 120 cm in a tillage and crop sequence trial in south Texas in 2000 and 2001. Main plots were subjected to three different tillage systems: conventional tillage (moldboard plowing and disking), ridge tillage, and no-tillage. Subplots were planted with three different crop sequences: spring cotton and fall corn every year; spring cotton and fall corn in one year, followed by corn for two years; and cotton followed by corn and then grain sorghum, one spring crop per year. The population density of R. reniformis on corn and grain sorghum was low throughout the soil profile. In plots planted with spring cotton and fall corn every year, fewer nematodes were found at depths of 60 to 120 cm in the no-tillage and ridge tillage systems than in the conventional tillage system. Population densities were lower at depths of 0 to 60 cm than at 60 to 120 cm. Soil moisture and cotton root length did not affect nematode population densities in the field. When soil was placed in pots and planted with cotton in the greenhouse, lower population densities developed in soil taken from depths of 0 to 60 cm than in soil from depths of 60 to 120 cm. Final nematode populations were similar in size in soil from the different tillage systems, but reproductive factors were higher in soil from plots with reduced-tillage systems than in soil from plots with conventional tillage. Reduced-tillage practices lowered the risk of increases in R. reniformis populations and reduced population densities following 2 years of non-hosts throughout soil depths, but population densities resurged to the same high levels as in soil planted with cotton every year during one season of cotton. | 18,944,104 |
Investigating the Spatiotemporal Genetic Structure of Phytophthora capsici in Michigan. | ABSTRACT Phytophthora capsici isolates were recovered from pepper and cucurbit hosts at seven locations in Michigan from 1998 to 2000. Isolates were characterized for compatibility type (CT), mefenoxam sensitivity (MS), and amplified fragment length polymorphism (AFLP) marker profiles. In total, 94 AFLP bands were resolved. Individual populations were highly variable. Within populations, 39 to 49% of the AFLP bands were polymorphic and estimated heterozygosities ranged from 0.16 to 0.19. Of the 646 isolates fingerprinted, 70% (454) had unique AFLP profiles. No clones were recovered between years or locations. Pairwise F statistics (Phi(ST)) between populations from different locations ranged from 0.18 to 0.40. A tree based on unweighted pair-group method with arithmetic average cluster analysis indicates discrete clusters based on location. Isolates from the same location showed no clustering based on the year of sampling. Analysis of molecular variance partitioned variability among (40%) and within populations (60%). The overall estimated Phi(ST) was 0.34 (SD = 0.03). A1/A2 CT ratios were approximately 1:1, and MS frequencies were similar between years for the two locations sampled over time. These data suggest that P. capsici persists in discrete outcrossing populations and that gene flow among locations in Michigan is infrequent. | 18,944,124 |
Characterization and mapping of oat crown rust resistance genes using three assessment methods. | ABSTRACT Resistance is the primary means of control for crown rust of oat (Avena sativa L.), caused by Puccinia coronata f. sp. avenae, and better knowledge of the genetics of resistance will enhance resistance breeding. Disease data were generated in the field and greenhouse for parents and recombinant inbred lines of the Ogle/TAM O-301 (OT) oat mapping population using (i) a new quantitative assay that employs quantitative real-time polymerase chain reaction (q-PCR) to estimate fungal growth in the host, (ii) digital image analysis, and (iii) visual ratings. The objectives of this study were to evaluate each assessment method's ability to map a major gene from cv. Ogle and potential quantitative trait loci (QTL) contributed by Ogle and TAM O-301. All three assessment methods identified the major gene in Ogle, which was mapped to linkage group OT6. The resolution produced by q-PCR, however, enabled more precise mapping of the major gene. Quantitative analysis indicated that 64% of the phenotypic variation was accounted for using q-PCR, whereas 41 and 52% were accounted for using visual and digital assessments, respectively. Data generated by q-PCR permitted identification of QTL on linkage groups OT32, accounting for 6% of the phenotypic variation, and OT2, accounting for 4% of the variation. QTL on both OT32 and OT2 were conferred by TAM O-301, one of which (OT2) was indiscernible using data from the visual and digital assessments. The new method of precisely phenotyping crown rust resistance provided a more accurate and thorough means of dissecting resistance in the OT mapping population. Similar methods could be developed and applied to other important cereal rust diseases. | 18,944,171 |
Edaphic Soil Levels of Mineral Nutrients, pH, Organic Matter, and Cationic Exchange Capacity in the Geocaulosphere Associated with Potato Common Scab. | ABSTRACT In order to determine possible relationships between geocaulosphere soil properties and severity of common scab of potato caused by Streptomyces scabies, soils were collected from representative commercial potato fields in Canada: in Simcoe and Dufferin Counties, Ontario and across Prince Edward Island (PEI) in August 2004. Soils immediately adjacent to tubers were sampled and analyzed for select edaphic factors and for pathogen presence using polymerase chain reaction (PCR) tests with primers that amplify a region of the TxtA gene involved in regulating the biosynthesis of the thaxtomin toxin family. Individual tubers were assessed visually for scab severity. The relationships between soil chemical factors and disease severity were investigated for each region to detect the strongest relationships. Principal component analysis revealed a distinctive clustering of samples with respect to disease severity in PEI but not in Ontario soils. Total and percent saturation of K (%K) were the only factors found associated with high disease severity in soils from both provinces. In PEI soils, pH, Mg, Ca, Cu, and %K, %Mg, %Ca, and %Na were associated with high disease severity, whereas cation exchange capacity (CEC) and Al were correlated with low disease severity soils. In Ontario, high Mn content was strongly correlated with low disease severity soils, whereas %K and organic matter content were correlated with disease severity. Partitioning samples into presence or absence of the TxtA PCR product with corresponding high or low severity showed further significant relationships in the data. There was an excellent correlation between Streptomyces spp. presence as detected by PCR and disease severity in PEI soils; however, the relationship was not as clear in Ontario soils, where many PCR-positive soils had low disease incidence. Principal component and partial least square analysis indicated that disease severity was predicted by soil factors such as organic matter, CEC, pH, Al, %Ca, %Mg, and %K for PEI but not for Ontario soils. The data reveal that the relationship between scab severity and soil chemical components is complex and potentially soil specific. | 18,944,172 |
Using the TxtAB operon to quantify pathogenic Streptomyces in potato tubers and soil. | The phytotoxin thaxtomin, produced by plant pathogenic Streptomyces species, is the only known pathogenicity determinant for common scab diseases of potato and other root and tuber crops. Genes encoding thaxtomin synthetase (txtAB) are found on a pathogenicity island characteristic of genetically diverse plant pathogenic Streptomyces species. In this study, an SYBR Green quantitative real-time polymerase chain reaction (PCR) assay using primers designed to anneal to the txtAB operon of Streptomyces was developed to quantify pathogenic bacterial populations in potatoes and soil. The real-time PCR assay was specific for pathogenic Streptomyces strains. The detection limit of the assay was 10 fg of the target DNA, or one genome equivalent. Cycle threshold (Ct) values were linearly correlated with the concentration of the target DNA (correlation coefficient R(2) = 0.99) and were not affected by the presence of plant DNA extracts, indicating the usefulness of the assay for quantitative analyses of the pathogenic bacteria in plant tissues. The amount of pathogenic Streptomyces DNA in total DNA extracts from 1 g asymptomatic and symptomatic tubers was quantified using the assay and ranged from 10(1) to 10(6) pg. A standard curve was established to quantify pathogenic Streptomyces in soil. Using the standard curve, numbers of pathogenic Streptomyces colony forming units were extrapolated to range from 10(3) to 10(6) per gram of soil from potato fields where common scab was found. This real-time PCR assay using primers designed from the txtAB operon allows rapid, accurate, and cost effective quantification of pathogenic Streptomyces strains in potato tubers and in soil. | 18,944,188 |
Variation in Albicidin Biosynthesis Genes and in Pathogenicity of Xanthomonas albilineans, the Sugarcane Leaf Scald Pathogen. | ABSTRACT Total genomic DNA from 137 strains of Xanthomonas albilineans from worldwide locations was hybridized with two DNA probes that together harbor the entire 49-kb albicidin biosynthesis gene cluster and two additional 3-kb genomic regions required for albicidin production. Fourteen haplotypes and two major genetic groups (albicidin [ALB]-restriction fragment length polymorphism [RFLP] A and ALB-RFLP B) were identified, and strains that were isolated after recent outbreaks of leaf scald disease belonged to group ALB-RFLP B. Albicidin genetic diversity was very similar to the previously described genetic diversity of the pathogen based on the whole genome. No relationship was found between variability of albicidin biosynthesis genes and the amount of albicidin produced in vitro by X. albilineans. Leaf scald-susceptible sugarcane cv. H70-144 was inoculated with 20 strains of the pathogen belonging to different ALB-RFLP haplotypes. Among them, 10 strains from Guadeloupe belonged to the same ALB-RFLP group but differed in the amount of albicidin produced in vitro. Strains were distributed in at least three different pathogenicity groups based on symptom severity and pathogen population density in the stalk. These two pathogenicity factors varied concurrently; however, no relationship between variation in albicidin biosynthesis genes, variation in the amount of albicidin produced in vitro, and variation in pathogenicity of X. albilineans was found. Further investigation is necessary to identify other genes involved in pathogenicity of X. albilineans. | 18,944,203 |
Strawberry Anthracnose: Histopathology of Colletotrichum acutatum and C. fragariae. | ABSTRACT Ontogeny of the invasion process by Colletotrichum acutatum and C. fragariae was studied on petioles and stolons of the strawberry cultivar Chandler using light and electron microscopy. The invasion of host tissue by each fungal species was similar; however, each invasion event occurred more rapidly with C. fragariae than with C. acutatum. Following cuticular penetration via an appressorium, subsequent steps of invasion involved hyphal growth within the cuticle and within the cell walls of epidermal, subepidermal, and subtending cells. Both species of fungi began invasion with a brief biotrophic phase before entering an extended necrotrophic phase. Acervuli formed once the cortical tissue had been moderately disrupted and began with the development of a stroma just beneath the outer periclinal epidermal walls. Acervuli erupted through the cuticle and released conidia. Invasion of the vascular tissue typically occurred after acervulus maturation and remained minimal. Chitin distribution in walls of C. fragariae was visualized with gold-labeled wheat germ agglutinin. The outer layer of bilayered walls of conidia, germ tubes, and appressoria contained less chitin than unilayered hyphae in planta. | 18,944,215 |
Silicon-induced cell wall fortification of rice leaves: a possible cellular mechanism of enhanced host resistance to blast. | ABSTRACT Locations of silicon accumulation in rice leaves and its possible association with resistance to rice blast were investigated by electron microscopy and X-ray microanalysis. A blast-susceptible cultivar, Jinmi, and a partially resistant cultivar, Hwaseong, were grown under a hydroponic culture system with modified Yoshida's nutrient solution containing 0, 50, 100, and 200 ppm of silicon. Electron-dense silicon layers were frequently found beneath the cuticle in epidermal cell walls of silicon-treated plants. Increasing levels of silicon were detected in the outer regions of epidermal cell walls. Silicon was present mainly in epidermal cell walls, middle lamellae, and intercellular spaces within subepidermal tissues. Furthermore, silicon was prevalent throughout the leaf surface, with relatively small deposition on stomatal guard cells in silicon-treated plants. Silicon accumulation and epidermal cell wall thickness in leaves were greater in cv. Jinmi than in cv. Hwaseong. However, the thickness ratios of the silicon layers to epidermal cell walls were greater in cv. Hwaseong (53.25 to 93.28%) than in cv. Jinmi (36.58 to 66.54%). Leaf blast severity was lower in cv. Hwaseong than in cv. Jinmi and was significantly reduced in silicon-treated plants of both cultivars. These results suggest that silicon-induced cell wall fortification of rice leaves may be closely associated with enhanced host resistance to blast. | 18,944,220 |
Resistance to Bean pod mottle virus in Transgenic Soybean Lines Expressing the Capsid Polyprotein. | ABSTRACT Transgenic fertile soybean plants were generated from somatic embryos of soybean (Glycine max) cv. Jack transformed via particle bombardment with the capsid polyprotein (pCP) gene of Bean pod mottle virus(BPMV). The plant transformation vector (pHIG/BPMV-pCP) utilized in these experiments contained the BPMV-pCP coding sequence, an intron-containing GUS gene, and the hygromycin phosphotransferase gene. Southern blot hybridization analysis showed that 19 transgenic soybean plants selected for resistance to hygromycin contained the genes for GUS and BPMV-pCP. The progeny of five of these transgenic soybean plants (plants 137, 139, 157, 183, and 186) were characterized in detail. An additional transgenic plant (plant 200) contained the intron-GUS and hygromycin resistance genes, but lacked the BPMV-pCP gene and was used as a negative control. Southern blot hybridization analysis of the five transgenic plants showed the presence of three copies of the T-DNA in a similar banding pattern suggesting that they were derived from a single transformation event. Western and northern blot analyses showed that the expression levels of BPMV-pCP and pCP transcript were high in these five pCP plants. Infectivity assays with detached leaves demonstrated that all five pCP plants exhibited resistance to virus infection because they accumulated lower levels of BPMV compared with plant 200 and nontransformed controls. Unlike the T(2) progeny of line 183-1 that segregated with respect to the pCP gene and, consequently, to BPMV resistance, the T(2) progeny of the homozygous line 183-2 showed little or no symptoms in response to rub-inoculation with virions of a severe strain of BPMV. Although BPMV accumulation was evident in leaves on which viruliferous beetles were allowed a 72-h inoculation access period, the upper noninoculated leaves of the T(2) progeny of line 183-2 plants were symptomless and accumulated little or no virus. Because the progeny of this homozygous transgenic line exhibited systemic resistance, they could potentially be useful in generating commercial cultivars resistant to BPMV. | 18,944,228 |
Aspergillus flavus and Aflatoxin Contamination of Leguminous Trees of the Sonoran Desert in Arizona. | ABSTRACT Aspergillus spp. in section Flavi were frequently associated with desert tree legumes in uncultivated areas of the Sonoran Desert. Of 270 samples of debris and fruits of mesquite (Prosopis spp.), ironwood (Olneya tesota), acacia (Acacia spp.), and palo verde (Cercidium and Parkinsonia spp.), 87% were positive for A. flavus (S and L strains) and A. tamarii. A. flavus was the most common species (87%) among the 3,763 isolates examined. Mesquite pods were both the substrate from which A. flavus was recovered most frequently and the substrate from native habitats with the greatest aflatoxin content. In vitro, most desert legumes supported significant growth, reproduction, and aflatoxin production by A. flavus, with mesquite pods yielding 1 x 10(10) propagules/g and 5,000 mug/kg of aflatoxin B(1). Twenty percent of legume pods collected in the desert contained measurable quantities of aflatoxin, ranging from 1 to >2,500 mug/kg. Insect-damaged mesquite pods had significantly higher aflatoxin than intact pods. Legumes are apparently important reservoirs of aflatoxin-producing fungi and significant sources of aflatoxin contamination in the native Sonoran Desert habitats of Arizona. | 18,944,238 |
Maize fine streak virus, a New Leafhopper-Transmitted Rhabdovirus. | ABSTRACT A previously uncharacterized virus was isolated from fall-planted sweet corn (Zea mays L., Syngenta GSS 0966) leaves showing fine chlorotic streaks. Symptomatic plants were negative in enzyme-linked immunosorbent assay against many maize viruses, but reacted weakly with antisera to Sorghum stunt mosaic virus suggesting a distant relationship between the viruses. The virus was readily transmitted by vascular puncture inoculation (VPI), but not by leaf-rub inoculation. Symptoms on maize included dwarfing and fine chlorotic streaks along intermediate and small veins that developed 12 to 17 days post-VPI. The isolated virus was bacilliform (231 +/- 5 nm long and 71 +/- 2 nm wide), with a knobby surface, and obvious helical structure typical of rhabdovirus morphology. Nucleorhabdovirus virions were observed by transmission electron microscopy of infected maize leaf tissue sections. Proteins unique to infected plants were observed in extracts of infected leaves, and the isolated virion contained three proteins with molecular masses 82 +/- 2, 50 +/- 3, and 32 +/- 2 kDa. Preliminary sequence analysis indicated the virus had similarity to members of the family Rhabdoviridae. The virus was transmitted by Graminella nigrifrons under persistent conditions. The data indicate the virus, provisionally designated Maize fine streak virus, is a new species in the genus Nucleorhabdovirus. | 18,944,241 |
Detection of two orchid viruses using quartz crystal microbalance-based DNA biosensors. | ABSTRACT We have developed a piezoelectric DNA-sensor based on DNA-RNA hybridization for the detection of two orchid viruses, Cymbidium mosaic virus (CymMV) and Odontoglossum ringspot virus (ORSV). Specific oligonucleotide probes modified with a mercaptohexyl group at the 5'-phosphate end were directly immobilized onto 10-MHz AT-cut quartz crystal microbalance (QCM). QCMs coated with such oligonucleotide probes were exposed to test solutions containing viral RNA for hybridization. Various experimental conditions evaluated were (i) DNA probe coating concentration, (ii) sensitivity and specificity of the probes at different hybridization temperatures, and (iii) effects of incubation temperature on the hybridization time. The specific nucleotide probe-coated QCM-based DNA sensors were able to detect both CymMV and ORSV in quantities as low as approximately 1 ng in purified RNA preparations and 10 ng in the crude sap of infected orchids. This is the first application of a DNA biosensor for the detection of plant viruses. | 18,944,263 |
Protection against pathogen and salt stress by four plant growth-promoting rhizobacteria isolated from Pinus sp. on Arabidopsis thaliana. | The ability of four plant growth-promoting rhizobacteria, isolated in a previous study, to induce systemic resistance on Arabidopsis thaliana Col 0 against biotic and abiotic stress was evaluated. All the bacteria enhanced protection against the foliar pathogen Pseudomonas syringae DC3000 and increased plant tolerance to salt stress (NaCl 60 mM). Bacillus sp. strain L81 and Arthrobacter oxidans strain BB1 performed best with a decrease in the disease index of 61.2 and 52.3%, respectively, and a reduction in the mortality due to salt stress of 72.4 and 57.8%, respectively. Additionally, significant differences were found in growth and photosynthesis, again, L81 and BB1 performed best either in normal or under stress conditions. In order to elucidate the pathway elicited by these two strains to induce systemic resistance, experiments with the transgenic line of Arabidopsis thaliana NahG (defective in salicylic acid [SA]) and with the jar1 mutant (defective in jasmonic acid) were carried out. Results showed that the SA-dependent pathway was involved in the defense response induced by strains L81 and BB1. Results from quantitative reverse transcription-polymerase chain reaction analysis of the PR1 gene, related to the SA-dependent pathway and the PDF1.2 gene related to the SA-independent pathway, showed an increased expression of PR1 in BB1-treated plants, confirming involvement of the SA-dependent pathway in the defensive response. | 18,944,290 |
Capsicum Species: Symptomless Hosts and Reservoirs of Tomato yellow leaf curl virus. | ABSTRACT Five Capsicum species were tested for susceptibility to Tomato yellow leaf curl virus (TYLCV) and the mild strain of TYLCV (TYLCV-Mld). TYLCV was able to infect 30 of 55 genotypes of C. annuum, one of six genotypes of C. chinense, one of two genotypes of C. baccatum, and the only genotype of C. frutescens tested but was unable to infect the one genotype of C. pubescens tested. This is the first evidence for the susceptibility of C. baccatum, C. chinense, and C. frutescens to TYLCV. Unlike TYLCV isolates, TYLCV-Mld was unable to infect C. chinense. No host differences were observed between the Israeli and Florida isolates of TYLCV. None of the Capsicum species showed symptoms after infection with TYLCV or TYLCV-Mld. TYLCV was detected in fruits of C. annuum, but whiteflies were unable to transmit virus from fruits to plants. White-flies were able to transmit both TYLCV and TYLCV-Mld from infected pepper plants to tomato plants. Pepper plants in research plots were found infected with TYLCV at rates as much as 100%. These data demonstrate the ability of some genotypes of pepper to serve as reservoirs for the acquisition and transmission of TYLCV and TYLCV-Mld. | 18,944,303 |
Identification and Characterization of Benomyl-Resistant and -Sensitive Populations of Colletotrichum gloeosporioides from Statice (Limonium spp.). | ABSTRACT Sixty-four isolates of Colletotrichum gloeosporioides were isolated from infected Limonium spp. cultivated in 12 different locations in Israel. All isolates were identified as belonging to the C. gloeosporioides complex by species-specific primers. Of these isolates, 46 were resistant to benomyl at 10 mug/ml and 18 were sensitive to this concentration of fungicide. Based on arbitrarily primed polymerase chain reaction of all isolates and internal transcribed spacer-1 sequence analyses of 12 selected isolates, the benomyl-resistant and -sensitive populations belong to two distinct genotypes. Sequence analyses of the beta-tubulin genes, TUB1 and TUB2, of five sensitive and five resistant representative isolates of C. gloeosporioides from Limonium spp. revealed that the benomyl-resistant isolates had an alanine substitute instead of a glutamic acid at position 198 in TUB2. All data suggest that the resistant and sensitive genotypes are two independent and separate populations. Because all Limonium plant propagation material is imported from various geographic regions worldwide, and benomyl is not applied to this crop or for the control of Colletotrichum spp. in Israel, it is presumed that plants are bearing quiescent infections from the points of origin prior to arrival. | 18,944,315 |
Genotyping of Serratia marcescens Strains Associated with Cucurbit Yellow Vine Disease by Repetitive Elements-Based Polymerase Chain Reaction and DNA-DNA Hybridization. | ABSTRACT The bacterium that causes cucurbit yellow vine disease (CYVD) has been placed in the species Serratia marcescens based on 16S rDNA and groE sequence analysis. However, phenotypic comparison of the organism with S. marcescens strains isolated from a variety of ecological niches showed significant heterogeneity. In this study, we compared the genomic DNA of S. marcescens strains from different niches as well as type strains of other Serratia spp. through repetitive elements-based polymerase chain reaction (rep-PCR) and DNA-DNA hybridization. With the former, CYVD strains showed identical banding patterns despite the fact that they were from different cucurbit hosts, geographic locations, and years of isolation. In the phylogenetic trees generated from rep-PCR banding patterns, CYVD strains clearly were differentiated from other strains but formed a loosely related group with S. marcescens strains from other niches. The homogeneity of CYVD strains was supported further by the DNA relatedness study, in that labeled DNA from the cantaloupe isolate, C01-A, showed an average relative binding ratio (RBR) of 99%, and 0.33% divergence to other CYVD strains. Used as a representative strain of CYVD, the labeled C01-A had a RBR of 76%, and a 4.5% divergence to the S. marcescens type strain. These data confirm the previous placement of CYVD strains in S. marcescens. Our investigations, including rep-PCR, DNA-DNA hybridization, and previous phenotyping experiments, have demonstrated that CYVD-associated strains of S. marcescens cluster together in a group significantly different from other strains of the species. | 18,944,323 |
Interactions Between Geminiviruses in a Naturally Occurring Mixture: Pepper huasteco virus and Pepper golden mosaic virus. | ABSTRACT Pepper huasteco virus (PHV) and Pepper golden mosaic virus (PepGMV) are found in mixtures in many horticultural crops in Mexico. This combination constitutes an interesting, naturally occurring model system to study several aspects of virus-virus interactions. Possible interactions between PHV and PepGMV were studied at four levels: symptom expression, gene expression, replication, and movement. In terms of symptom expression, the interaction was shown to be host-dependent because antagonism was observed in pepper, whereas synergism was detected in tobacco and Nicotiana benthamiana. PHV and PepGMV did not generate viable pseudorecombinant viruses; however, their replication is increased during mixed infections. An asymmetric complementation in movement was observed because PHV was able to support the systemic movement of PepGMV A whereas PepGMV did not support the systemic distribution of PHV A. Heterologous transactivation of both coat protein promoters also was detected. Several conclusions can be drawn from these experiments. First, viruses coinfecting the same plant can interact at several levels (replication, movement) and in different manners (synergism, antagonism); some interactions might be host dependent; and natural mixed infections could be a potential source of geminivirus variability by generating viable tripartite combinations that could facilitate recombination events. | 18,944,336 |
Induction of Chitinase, beta-1,3-Glucanase, and Phenylalanine Ammonia Lyase in Peach Fruit by UV-C Treatment. | ABSTRACT Treatment of peach fruit with UV-C light caused a rapid induction of chitinase, beta-1,3-glucanase, and phenylalanine ammonia lyase (PAL) activities starting 6 h after treatment and reaching maximum levels at 96 h after treatment. By 96 h after UV-C treatment, chitinase, beta-1,3-glucanase, and PAL activities in UV-C-treated fruit were over twofold above the levels observed for the control. In nontreated control fruit, no apparent increase in chitinase and beta-1,3-glucanase activities was detected but a minor increase in PAL activity was seen. The transient increase in chitinase, beta-1,3-glucanase, and PAL activities in UV-C-treated fruit was preceded by a gradual activation of the corresponding genes. UV-C-treated fruit showed an increase in accumulation of beta-1,3-glucanase and chitinase mRNAs at 3 h after treatment, which peaked approximately 96 h posttreatment. A similar induction kinetic pattern was observed for PAL mRNA in response to UV-C treatment, except the induction started 6 h after UV-C treatment. These results show that the response of peach fruit to elicitor treatment is similar to that seen in other plant-elicitors interactions and suggests the involvement of peach biochemical defense responses in UV-C-mediated disease resistance. | 18,944,346 |
Evaluation of local and imported fire blight warning systems in Israel. | ABSTRACT The possibility of using local and imported warning systems for the management of fire blight (caused by the bacterium Erwinia amylovora) in pears was tested in Israel from 1997 to 2000. Three imported systems (MARYBLYT 4.3, BIS95, and Cougarblight 98C) and one local system (Fire Blight Control Advisory [FBCA]) were used. All systems were tested in simulation experiments; MARYBLYT 4.3 and FBCA were also tested in orchard experiments under natural infections. Simulation experiments included 193 orchard-plots in which the time of disease onset enabled us to determine the date of infection. Thirty-five experiments were conducted in commercial orchards; in 10 of these, fire blight developed naturally. The performance of the imported warning systems was too variable to be accurately used under Israeli conditions. In the simulation experiments, the success rate (i.e., the capacity of the systems to predict the exact date of the occurrence of infection episodes) of the imported systems was low (3 to 55%) with considerably large variability among years (CV = 30 to 67%). Similar results were obtained in the orchard experiments for MARYBLYT 4.3: in only two of five experiments where plots were managed according to that system was disease severity significantly lower than that recorded in untreated control plots. In comparison, the local system, FBCA, predicted most infection episodes in the simulation experiments with low variability (99%, CV = 1.0%). In the orchard experiments, adequate disease suppression was achieved in all eight experiments in which FBCA recommendations were followed. We concluded that it was not possible to import and successfully implement fire blight warning systems in Israel that have been developed in regions with dissimilar environmental conditions. | 18,944,347 |
The Identification of Two New Races of Pyrenophora tritici-repentis from the Host Center of Diversity Confirms a One-to-One Relationship in Tan Spot of Wheat. | ABSTRACT Pyrenophora tritici-repentis, causal agent of tan spot, induces necrosis and chlorosis in its wheat host. The tan spot system conforms to the toxin model and three host-specific toxins have been identified (Ptr ToxA, Ptr ToxB, and putative Ptr ToxC). Processing of a collection of isolates, obtained in the Fertile Crescent and Caucasus regions, yielded two new virulence patterns. Isolate Az35-5 combined the virulences of races 2 and 5 and was classified in the new race 7. Isolates TS93-71B and TS93-71F had a virulence pattern that combined those of races 2, 3, and 5 and were grouped in the new race 8. Southern analysis revealed that all three isolates possessed copies of the ToxA and ToxB genes, the first time the genes were found in a common background. The production of Ptr ToxA and Ptr ToxB by the isolates was confirmed by western blotting. Virulence patterns suggested that TS93-71B and TS93-71F may also produce Ptr ToxC, even though it was not present at detectable levels in culture filtrates. The identification of races 7 and 8 complete the theoretical maximum number of races that can be differentiated by three loci in the host (2(3) = 8), assuming a one-to-one relationship. It appears that the wheat/P. tritici-repentis system is a mirror image of the classical gene-for-gene relationship. | 18,944,352 |
Defense Gene Expression Analysis of Arabidopsis thaliana Parasitized by Orobanche ramosa. | ABSTRACT The infection of Arabidopsis thaliana roots with the obligate parasite Orobanche ramosa represents a useful model for a study of the molecular events involved in the host plant response to a parasitic plant attack. To avoid analysis problems due to the subterranean development of O. ramosa, we developed two in vitro co-culture systems: O. ramosa seedlings infesting Arabidopsis plantlet roots and callus tissues. We were then able to investigate the expression patterns of some host plant genes selected among genes known to be involved in metabolic pathways and resistance mechanisms activated during several plant-pathogen interactions including ethylene, isoprenoid, phenylpropanoid, and jasmonate biosynthesis pathways, oxidative stress responses, and pathogenesis-related proteins. Molecular analyses were carried out using polymerase chain reaction amplification methods allowing semiquantitative evaluation of transcript accumulation during early (first hours) and late (15 days) stages of infestation, in whole roots or parts close to the parasite attachment site. In A. thaliana, O. ramosa induced most of the general response signaling pathways in a transient manner even before its attachment to A. thaliana roots. However, no salicylic acid-dependent defense is observed because no activation of systemic acquired resistance markers is detectable, whereas genes, co-regulated by jasmonate and ethylene, do display enhanced expression. | 18,944,360 |
Sequence tagged site markers to rsp1, rsp2, and rsp3 genes for resistance to septoria speckled leaf blotch in barley. | ABSTRACT Five random amplified polymorphic DNA markers, two in coupling (OPAH5(545C), and OPBA12(314C)) and three in repulsion phase (UBC285(158R), OPC2(441R), and OPB17(451R)), closely linked to Rsp genes conferring resistance to Septoria speckled leaf blotch (SSLB), were identified using bulked segregant analysis in three F(2) populations, each containing a Rsp gene. These markers were converted into the sequence tagged site (STS) markers SUBC285, SOPC2, SOPAH5, and SOPBA12. Another STS marker (MWG938) linked to Rsp2 in coupling phase was also identified in an F(2) population from the cross Robust/CIho 4780. The STS markers were tested on a set of 42 resistant and susceptible barley germplasm lines and 98 landraces. The expected sizes of marker fragments associated with each allele at Rsp loci were present in resistant or susceptible accessions. Efficiency of marker-assisted selection (MAS) for Rsp1, Rsp2, and Rsp3 using STS markers were evaluated in three F(23) populations in the greenhouse and the field. Results of testing F(23) progeny demonstrated that the accuracy of MAS was, with one exception, greater than 97% in the greenhouse and in two field locations (90% in the Osnabrock, ND trial for Rsp2). The STS markers closely linked to Rsp genes also identified the SSLB resistance corresponding to Rsp1, Rsp2, or Rsp3 in gene pyramiding F(2) populations. The STS markers tightly linked to Rsp genes may be useful for M and for pyramiding with other genes in barley breeding for SSLB resistance. | 18,944,371 |
Characterization of early leaf spot suppression by strip tillage in peanut. | ABSTRACT Epidemics of early leaf spot of peanut (Arachis hypogaea), caused by Cercospora arachidicola, are less severe in strip-tilled than conventionally tilled fields. Experiments were carried out to characterize the effect of strip tillage on early leaf spot epidemics and identify the primary target of suppression using a comparative epidemiology approach. Leaf spot intensity was assessed weekly as percent incidence or with the Florida 1-to-10 severity scale in peanut plots that were conventionally or strip tilled. The logistic model, fit to disease progress data, was used to estimate initial disease (y(0)) and epidemic rate (r) parameters. Environmental variables, inoculum abundance, and field host resistance were assessed independently. For experiments combined, estimated y(0) was less in strip-tilled than conventionally tilled plots, and r was comparable. The epidemic was delayed in strip-tilled plots by an average of 5.7 and 11.7 days based on incidence and severity, respectively. Tillage did not consistently affect mean canopy temperature, relative humidity, or frequency of environmental records favorable for infection or spore dispersal. Host response to infection was not affected by tillage, but infections were detected earlier and at higher frequencies with noninoculated detached leaves from conventionally tilled plots. These data suggest that strip tillage delays early leaf spot epidemics due to fewer initial infections; most likely a consequence of less inoculum being dispersed to peanut leaves from overwintering stroma in the soil. | 18,944,374 |
Chitinases from the Plant Disease Biocontrol Agent, Stenotrophomonas maltophilia C3. | ABSTRACT Stenotrophomonas maltophilia strain C3, a biocontrol agent of Bipolaris sorokiniana in turfgrass, produced chitinases in broth media containing chitin. Chitinases were partially purified from culture fluid by ammonium sulfate precipitation and chitin affinity chromatography. The chromatography fraction with the highest specific chitinase activity was inhibitory to conidial germination and germ-tube elongation of B. sorokiniana, but it was less inhibitory than the protein fraction or the raw culture filtrate. The fraction exhibited strong exochitinase and weak endo-chitinase activity. Optimum temperature and pH for chitinase activity were 45 to 50 degrees C and 4.5 to 5.0, respectively. Chitinase activity was inhibited by Hg(2+) and Fe(3+), but not by other metal ions or enzyme inhibitors. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of the chromatography fraction revealed the presence of five protein bands of 25, 32, 48, 65, and 75 kDa. Partial amino acid sequences of the 32-, 65-, and 75-kDa proteins indicated that they are homologous to known bacterial chitinases. There was no homology found in the partial amino acid sequences of the 25- and 48-kDa proteins to any known chitinases. Five chitinase-active proteins were detected in the protein and chromatography fractions by activity gels, but when each protein was extracted and re-electrophoresed separately under denaturing conditions, only 32- or 48-kDa proteins were revealed. It was concluded that strain C3 produces at least two chitinases that are antifungal. | 18,944,395 |
Temporal and Spatial Patterns of Genetic Structure of Phytophthora infestans from Tomato and Potato in the Del Fuerte Valley. | ABSTRACT The temporal and spatial patterns of Phytophthora infestans population genetic structure were analyzed in the Del Fuerte Valley, Sinaloa, Mexico, during the crop seasons of 1994 to 1995, 1995 to 1996, and 1996 to 1997 by geographical information systems. Isolates of P. infestans were obtained from infected tissue of tomato and potato collected from two areas: (i) where both potatoes and tomatoes are grown, and (ii) where only tomatoes are grown. The isolates were characterized by mating type, allozymes at the glucose-6-phosphate isomerase and peptidase loci, restriction fragment length polymorphism (RFLP) fingerprint with probe RG57, metalaxyl sensitivity, and aggressiveness to tomato and potato. The results suggest presence of an asexual population with frequent immigrations from outside the valley. There was a shift of mating type in the population from predominantly A2 to completely A1 in this period. The co-occurrence of mating types was restricted to very few fields in the area around Los Mochis where tomato and potato crops are grown. Genotype variation based on allozyme analysis and mating type was low with only one genotype affecting both crops each year. The genotypes affecting both crops were the only genotypes highly aggressive to both tomato and potato in laboratory aggressiveness tests and the only genotypes widespread on both the tomato and potato crops in the valley each year. These predominant genotypes were highly resistant to the fungicide metalaxyl. Data on metalaxyl sensitivity indicate that allozyme analysis can discriminate between sensitive and resistant isolates in the Del Fuerte Valley. RFLP analysis with the probe RG57 gives further discrimination of genotypes within an allozyme genotype. In the 1995 to 1996 season, four different RFLP genotypes were found within an allozyme genotype. However, there were five other dilocus allozyme genotypes that could not be further split by RFLP analysis in 1995 to 1996 and 1996 to 1997 seasons. Spatial analysis of genotypes suggests that each season individual fields near Los Mochis became infected with one or more genotypes, but only a single genotype, aggressive on both potato and tomato, occurred south and east to the Guasave area. | 18,944,419 |
An Elicitor from Botrytis cinerea Induces the Hypersensitive Response in Arabidopsis thaliana and Other Plants and Promotes the Gray Mold Disease. | ABSTRACT Botrytis cinerea is a necrotrophic fungus that infects over 200 plant species. Previous studies showed that host cells collapse in advance of the hyphae, suggesting secretion of toxins or elicitors. We have partially characterized elicitor activity from intercellular fluid extracted from Arabidopsis thaliana leaves infected with B. cinerea. Treatment of intact leaves or cell cultures with either intercellular fluid from infected leaves or medium from inoculated A. thaliana cell culture induced generation of reactive oxygen species, resulting in reduced photosynthesis, electrolyte leakage, and necrotic lesions that resembled the hypersensitive response (HR). The necrosis was inhibited by diphenyleneiodonium, a specific inhibitor of NADPH oxidase, and by chelating free iron, suggesting the involvement of hydroxyl radicals. The necrosis was also suppressed in dnd1 mutants that are compromised in HR. In contrast, increased cell death was observed in acd2 mutants, indicating the involvement of the host defense signaling pathways. Treatment with the intercellular fluid from infected leaves also induced transcription of pathogenesis-related (PR) genes, such as PR-1, PR-5, HSR203J, and of senescence-associated gene SAG-13. Moreover, rapid transcription of the ethylene-dependent AtEBP gene was detected, indicating induction of ethylene production. The inter-cellular fluid from infected A. thaliana induced cell death in other plants, in line with the lack of B. cinerea specificity. In summary, the intercellular fluid mimicked a range of molecular and physiological host responses that are observed during infection with a live fungus. Moreover, it accelerated the B. cinerea infection, suggesting that the elicitor may act as a pathogenicity factor in the progression of gray mold disease. | 18,944,445 |
Necrotic Lesion Resistance Induced by Peronospora tabacina on Leaves of Nicotiana obtusifolia. | ABSTRACT Infection of Nicotiana obtusifolia plant introduction (PI) #555573 by the downy mildew pathogen Peronospora tabacina resulted in a compatible interaction, in which P. tabacina penetrated and freely colonized host leaf tissue. This interaction became incompatible 5 to 6 days later, with the appearance of necrotic lesions (NLs) and inhibition of pathogen growth and subsequent sporulation. NL development depended upon the presence of P. tabacina in host tissue, was not due to the effects of other microbes, and occurred co-incident in time with the pathogen's ability to produce asexual sporangia on a susceptible N. obtusifolia genotype. Inhibition of the necrotic response by CoCl(2) (a calcium channel blocker) and pathogen-induced transcription of a defense-related gene (PR-1a) suggested that necrosis was due to hypersensitive cell death in the host. In contrast, N. obtusifolia PI#555543 did not exhibit hypersensitivity upon infection by P. tabacina, but rather developed characteristic symptoms of tobacco blue mold disease: chlorotic lesions accompanied by abundant pathogen sporulation. Disease reactions scored on PI#555573 x PI#555543 F(2) progeny inoculated with P. tabacina sporangia indicated that the resistance phenotype was due to the action of a single gene from N. obtusifolia PI#555573, which we have named Rpt1. To date, Rpt1 is the only gene known to confer a hypersensitive response (HR) to P. tabacina infection in any species of Nicotiana. A survey of wild N. obtusifolia revealed that the HR to P. tabacina was expressed in the progeny of 7 of 21 (33%) plants collected in southern Arizona, but not in the progeny of plants originating from Death Valley National Park in California and the Big Bend National Park in west Texas. | 18,944,453 |
Formulation of Bacillus spp. for Biological Control of Plant Diseases. | ABSTRACT Maximizing the potential for successfully developing and deploying a biocontrol product begins with a carefully crafted microbial screening procedure, proceeds with developing mass production protocols that optimize product quantity and quality, and ends with devising a product formulation that preserves shelf-life, aids product delivery, and enhances bioactivity. Microbial selection procedures that require prospective bio-control agents to possess both efficacy and amenability to production in liquid culture increase the likelihood of selecting agents with enhanced commercial development potential. Scale-up of biomass production procedures must optimize product quantity without compromise of product efficacy or amenability to stabilization and formulation. Formulation of Bacillus spp. for use against plant pathogens is an enormous topic in general terms but limited in published specifics regarding formulations used in commercially available products. Types of formulations include dry products such as wettable powders, dusts, and granules, and liquid products including cell suspensions in water, oils, and emulsions. Cells can also be microencapsulated. Considerations critical to designing successful formulations of microbial biomass are many fold and include preserving biomass viability during stabilization, drying, and rehydration; aiding biomass delivery, target coverage, and target adhesion; and enhancing biomass survival and efficacy after delivery to the target. Solutions to these formulation considerations will not necessarily be compatible. Data from several biocontrol systems including the use of B. subtilis OH 131.1 (NRRL B-30212) to reduce Fusarium head blight of wheat are used to illustrate many of these issues. Using our recently described assay for efficiently evaluating biomass production and formulation protocols, we demonstrate the effectiveness, in vitro, of UV protectant compounds lignin (PC 1307) and Blankophor BBH in reducing OH 131.1 morbidity when cells were exposed to UV light from artificial sunlight. | 18,944,465 |
Analysis of Host Species Specificity of Magnaporthe grisea Toward Wheat Using a Genetic Cross Between Isolates from Wheat and Foxtail Millet. | ABSTRACT A genetic cross was performed between a Setaria isolate (pathogenic on foxtail millet) and a Triticum isolate (pathogenic on wheat) of Magnaporthe grisea to elucidate genetic mechanisms of its specific parasitism toward wheat. A total of 80 F(1) progenies were obtained from 10 mature asci containing 8 ascospores. Lesions on wheat leaves produced by the F(1) progenies were classified into four types, which segregated in a 1:1:1:1 ratio. This result suggested that the pathogenicity of the F(1) population on wheat was controlled by two genes located at different loci. This idea was supported by backcross analyses. We designated these loci as Pwt1 and Pwt2. Cytological analyses revealed that Pwt1 and Pwt2 were mainly associated with the hypersensitive reaction and papilla formation, respectively. | 18,944,467 |
Assessment of Diversity in Claviceps africana and Other Claviceps Species by RAM and AFLP Analyses. | ABSTRACT Genetic diversity among isolates of Claviceps africana, the sorghum ergot pathogen, and isolates of other Claviceps spp. causing ergot on sorghum or other hosts, was analyzed by random amplified microsatellite (RAM) and amplified fragment length polymorphism (AFLP) analyses. Of the RAM primer sets tested, one revealed polymorphism in C. africana isolates, with Australian and Indian isolates possessing a unique fragment. AFLP analysis, in addition to clearly distinguishing Claviceps spp., revealed polymorphisms in C. africana. A group of isolates from the United States, Puerto Rico, and South Africa exhibited 95 to 100% similarity with one another. Several isolates from Isabela, Puerto Rico were 100% similar to an isolate from Texas, and another isolate from Puerto Rico was identical with one from Nebraska. Australian and Indian isolates showed greater than 90% similarity with isolates from the United States., Puerto Rico, and South Africa. A number of polymorphisms existed in the United States group, indicating that the recently introduced population contains multiple genotypes. Isolates of C. sorghicola, a newly described sorghum pathogen from Japan, were very distinct from other species via RAM and AFLP analyses, as were isolates from outgroups C. purpurea and C. fusiformis. Both RAM and AFLP analysis will be useful in determining future patterns of intercontinental migration of the sorghum ergot pathogen, with the AFLP method showing greater ability to characterize levels of intraspecific variation. | 18,944,476 |
Evaluation of Infection Processes and Resulting Disease Caused by Dendryphion penicillatum and Pleospora papaveracea on Papaver somniferum. | ABSTRACT Two pathogenic fungi of opium poppy, Pleospora papaveracea and Dendryphion penicillatum, were isolated from field material in Beltsville, MD. The processes of infection by these two fungi were studied to determine the optimal environmental conditions for infection. Both fungi formed appressoria capable of penetrating directly through the plant epidermal layer. Of the two fungi, P. papaveracea was more aggressive, causing more rapid necrosis. Appressorial formation by P. papaveracea occurred as early as 4 h after application of a conidial suspension to poppy leaves. P. papaveracea formed more appressoria than did D. penicillatum, especially at cool temperatures (7 to 13 degrees C). In greenhouse studies, P. papaveracea caused more damage to opium poppy than did D. penicillatum when applied in 10% unrefined corn oil. In the field, P. papaveracea was more consistent in its effects on opium poppy from a local seed source designated Indian Grocery. P. papaveracea caused higher disease ratings, more stem lesions, and equal or greater yield losses than did D. penicillatum on Indian Grocery. The late-maturing opium poppy variety White Cloud was severely damaged by disease, regardless of formulation or fungal treatment. P. papaveracea was the predominant fungus isolated from poppy seed capsules and the only fungus reisolated from the field the following year. These studies provide a better understanding of the infection process and the differences between these two pathogenic fungi and will be beneficial for the development of the fungi as biological control agents. | 18,944,488 |
Evidence for Antibiosis and Induced Host Defense Reactions in the Interaction Between Verticillium lecanii and Penicillium digitatum, the Causal Agent of Green Mold. | ABSTRACT Chronological events of the intercellular interaction between Verticillium lecanii and the postharvest pathogen Penicillium digitatum were investigated by transmission electron microscopy and gold cytochemistry. Growth inhibition of P. oligandrum as a response to V. lecanii attack correlated with striking host changes including retraction of the plasma membrane and cytoplasm disorganization. Such changes were associated with the deposition on the inner host cell surface of a chitin- and cellulose-enriched material which appeared to be laid down as a structural defense reaction. The accumulation of chitin in the newly formed material correlated with a decrease in the amount of wallbound chitin. However, the deposition of cellulose appeared to correspond to a de novo synthesis, as evidenced by the occurrence of cellulose-containing vesicles which released their content in the space between the invaginated plasma membrane and the host cell wall. Results of the present study provide the first ultrastructural and cytochemical evidence that antagonism, triggered by V. lecanii, is a multifaceted process in which antibiosis, with alteration of the host hyphae prior to contact with the antagonist, appears to be the key process in the antagonism against P. digitatum. | 18,944,516 |
Quantitative trait Loci associated with resistance to gray leaf spot of corn. | ABSTRACT The susceptible parent FR1141, the resistant parent 061, the F1 cross, and 301 families selfed once from backcrosses to the susceptible parent were evaluated for gray leaf spot (GLS) severity for two years in Urbana, IL, and one year in Andrews, NC. Linkage between ear height and GLS severity was suspected. Therefore, plant height characteristics were noted for two years in Urbana, IL. Eighty-six polymorphic probes were used to construct a random fragment length polymorphism linkage map, and the presence, locations, effects, and interactions of quantitative trait loci (QTL) associated with GLS, plant and ear height were determined. Five QTL were significantly associated with GLS resistance across all environments and rating periods. These five regions are associated with additive effects on phenotype and account for between 51.0 and 58.7% of the phenotypic variation associated with GLS severity. Additionally, six QTL were identified with maturity-dependent associations to GLS severity. Heritability of GLS resistance was estimated to be approximately 0.73. Four QTL were identified with associations to ear height relative to total plant height. One of the four was associated with higher ear height and GLS resistance. | 18,944,528 |
Early Events During Quiescent Infection Development by Colletotrichum gloeosporioides in Unripe Avocado Fruits. | Inoculation of avocado pericarp tissue with Colletotrichum gloeospori-oides and treatment of avocado cell cultures with the cell wall elicitor of C. gloeosporioidesboth increased the production of reactive oxygen species (ROS). However, whereas the production of ROS could be detected within minutes in avocado cell suspensions, it was detected only after 2 h following inoculation of pericarp tissue. Protein kinase inhibitors such as K-252a and staurosporine and the phosphatase inhibitor microcystin-LR inhibited the release of H(2)O(2) from avocado cell suspensions. When 1 mM H(2)O(2) was exogenously applied to pericarp tissue, it enhanced ROS, phenyl-alanine ammonia lyase (PAL) activity, and epicatechin levels. But, when H(2)O(2) treatment was applied following staurosporine treatment, PAL activity was no longer induced. The uninduced ROS production in pericarp tissue of freshly harvested, unripe, resistant fruit was twice as high as in ripe, susceptible fruit. Challenge inoculation of resistant fruit further increased the ROS level; however, this increase did not occur in susceptible fruits. The current findings are consistent with the hypothesis that production of ROS is induced by fungal infection of unripe fruits and, consequently, may modulate resistance, resulting in the inhibition of fungal development and quiescence. | 18,944,563 |
Bacterial-Mediated Induced Resistance in Cucumber: Beneficial Effect of the Endophytic Bacterium Serratia plymuthica on the Protection Against Infection by Pythium ultimum. | ABSTRACT The potential of the endophytic bacterium Serratia plymuthica strain R1GC4 in stimulating defense reactions in cucumber (Cucumis sativus) seedlings inoculated with the soilborne pathogen Pythium ultimum was explored at the cellular level. Bacterial treatment prior to Pythium inoculation resulted in less seedling disease development as compared with that in nontreated control plants, in which typical root symptoms were visible by 3 days after inoculation with the pathogen. Histological investigations of root samples revealed striking differences in the extent of plant defense reactions between bacterized and nonbacterized plants. These observations were further confirmed at the ultrastructural level with the demonstration that restriction of fungal colonization to the outermost root tissues of bacterized seedlings correlated with the deposition of enlarged callose-enriched wall appositions at sites of potential pathogen penetration and the accumulation of an osmiophilic material in the colonized areas. Hyphae of the pathogen, surrounded by this electron-opaque material, exhibited considerable changes including cytoplasm disorganization and, in many cases, loss of the protoplasm. However, labeling with the beta-1,4-exoglucanase resulted in a regular labeling of Pythium cell walls, even at a time when these walls were entirely coated by the osmiophilic material. This material was also found to infiltrate into the invading hyphae to form either an internal coating of the cell wall or a network of polymorphic droplets in the area previously occupied by the cytoplasm. Cytochemical investigations revealed that callose, pectin, and cellulose appeared in the wall appositions. In addition, glucosides, lipids, and phenolics were detected in the electron-dense aggregates forming the core of most wall appositions. Finally, galactose residues were among the minor polysaccharidic compounds detected in the wall appositions. Evidence is provided in this study showing that treatment with S. plymuthica sensitizes susceptible cucumber plants to react more rapidly and more efficiently to Pythium attack through the formation of physical and chemical barriers at sites of potential fungal entry. | 18,944,571 |
The Role of Chitinase Production by Stenotrophomonas maltophilia Strain C3 in Biological Control of Bipolaris sorokiniana. | ABSTRACT The role of chitinase production by Stenotrophomonas maltophilia strain C3 in biological control of leaf spot on tall fescue (Festuca arundinacea), caused by Bipolaris sorokiniana, was investigated in vitro and in vivo. The filtrate of a broth culture of C3, with chitin as the carbon source, was separated into fractions. A high molecular-weight fraction (>8 kDa) was chitinolytic and more inhibitory than a low-molecular-weight, nonchitinolytic fraction to conidial germination and hyphal growth by B. sorokiniana and to leaf spot development. A protein fraction derived by ammonium sulfate precipitation and a chitinase fraction purified by chitin affinity chromatography also were chitinolytic and highly antifungal. The chitinolytic fractions caused swelling and vacuolation of conidia and discoloration, malformation, and degradation of germ tubes. When boiled, the chitinolytic fractions lost chitinase activity along with most of the antifungal properties. Two chitinase-deficient and two chitinase-reduced mutants of C3 were compared with the wild-type strain for inhibition of germination of B. sorokiniana conidia on tall fescue leaves and for suppression of leaf spot development in vivo. The mutants exhibited reduced antifungal activity and biocontrol efficacy, but did not lose all biocontrol activity. An aqueous extract of leaves colonized by wild-type C3 had higher chitinase activity than that of noncolonized leaves and was inhibitory to conidial germination. The addition of chitin to leaves along with the wild-type strain increased both chitinase and antifungal activity. The chitinase activity level of extracts from leaves colonized by a chitinase-deficient mutant of C3, with and without added chitin, was no higher than the background, and the extracts lacked antifungal activity. Chitinolysis appears to be one mechanism of biological control by strain C3, and it functions in concert with other mechanisms. | 18,944,588 |
Genetic Differentiation and Host Specificity Among Populations of Alternaria spp. Causing Brown Spot of Grapefruit and Tangerine x Grapefruit Hybrids in Florida. | ABSTRACT Alternaria spp. were sampled from brown spot lesions in several geographically separated citrus groves and different grapefruit and tangerine x grapefruit hybrid cultivars in Florida and screened for variation at 16 putative random amplified polymorphic DNA loci. Populations of the pathogen on two hybrids, Minneola and Orlando, in five locations throughout Florida were moderately differentiated (Nei's coefficient of gene differentiation [G(ST)] = 0.12) among locations. The hypothesis that host-specialized forms of Alternaria spp. cause brown spot on different Citrus spp. and cultivars was tested by estimating genetic differentiation among isolates sampled from different hosts and by pathogenicity assays. Isolates sampled from grapefruit and the hybrid cv. Nova were genetically distinct from isolates sampled from other hybrid cultivars including Robinson, Sunburst, Minneola, Orlando, and Murcott. No differentiation could be detected among isolates sampled from this latter group of hybrids. Quantitative pathogenicity assays on leaves using spray inoculation revealed that 'Nova' isolates were not significantly more pathogenic on 'Nova' compared with isolates from 'Minneola' and 'Orlando'. Similarly, grapefruit isolates were not significantly more pathogenic on grapefruit compared with isolates from 'Minneola'. Isolates from all hosts had similar disease rankings on each inoculated cultivar, with 'Minneola' the most susceptible, followed in decreasing order of susceptibility by 'Orlando', 'Sunburst', 'Nova', and 'Duncan' grapefruit. Rough lemon was generally immune to all isolates tested; however, occasional brown spot lesions were observed on leaves of this host with isolates from grapefruit. No evidence was found to support the hypothesis that unique genotypes of the pathogen, which are more virulent on 'Sunburst' or grapefruit, have been introduced to Florida. Populations of Alternaria spp. causing brown spot of citrus on grapefruit and 'Nova' in Florida are genetically distinct from isolates on other cultivars, and we speculate that these populations are in the early stages of adaptation to and possible speciation on these hosts. | 18,944,592 |
Early brown rot infections in sweet cherry fruit are detected by monilinia-specific DNA primers. | ABSTRACT Visible and nonvisible quiescent infections of immature and mature fruit are an integral component of the disease cycle of brown rot of sweet cherry in California. Detection of these infections is critical for developing efficient and efficacious fungicide management programs. The previously published DNA amplification primers mfs3 and NS5 for the identification of Monilinia fructicola were very specific in amplifying DNA of M. fructicola only and not M. laxa. This primer set, however, only detected DNA from some of the California isolates of M. fructicola. This genetic diversity was supported by random amplified polymorphic DNA (RAPD) analysis. Using eight 10-mer primers, seven M. fructicola isolates from California were all identified as genetically distinct. Using the same primers, only one polymorphism was detected among seven isolates of M. laxa. The multiple genotypes identified within the small population sample of M. fructicola, but not of M. laxa, using RAPD analysis could be indicative of genetic recombination within M. fructicola but not within M. laxa. To detect early brown rot infections in fruit, two primer sets that were developed from DNA sequences of either ribosomal DNA (MF5/ITS4/ITS3) or a RAPD fragment (X-09intF3/X-09R) specifically amplified DNA from isolates of M. fructicola and Monilinia species, respectively. No amplification products were present when using DNA from Botrytis cinerea or from other fungi commonly found on sweet cherry fruit. Primers X-09intF3 and X-09R were more sensitive and reliable for detecting small amounts of target DNA either extracted from conidia or from laboratory-inoculated cherry fruit with early brown rot infections that showed no visual symptoms or with visible quiescent infections. Furthermore, these primers also were effective for detecting visible quiescent infections in cherry fruit that were collected in the field. | 18,944,605 |
Molecular Variability Among Isolates of Prunus Necrotic Ringspot Virus from Different Prunus spp. | ABSTRACT Viral sequences amplified by polymerase chain reaction from 25 isolates of Prunus necrotic ringspot virus (PNRSV), varying in the symptomatology they cause in six different Prunus spp., were analyzed for restriction fragment polymorphisms. Most of the isolates could be discriminated by using a combination of three different restriction enzymes. The nucleotide sequences of the RNA 4 of 15 of these isolates were determined. Sequence comparisons and phylogenetic analyses of the RNA 4 and coat proteins (CPs) revealed that all of the isolates clustered into three different groups, represented by three previously sequenced PNRSV isolates: PV32, PE5, and PV96. The PE5-type group was characterized by a 5' untranslated region that was clearly different from that of the other two groups. The PV32-type group was characterized by an extra hexanucleotide consisting of a duplication of the six immediately preceding nucleotides. Although most of the variability was observed in the first third of the CP, the amino acid residues in this region, which were previously thought to be functionally important in the replication cycle of the virus, were strictly conserved. No clear correlation with the type of symptom or host specificity could be observed. The validity of this grouping was confirmed when other isolates recently characterized by other authors were included in these analyses. | 18,944,653 |
Chromosomal Gene Transfer by Conjugation in the Plant Pathogen Xanthomonas axonopodis pv. vesicatoria. | ABSTRACT Genes for copper resistance, located on the chromosome of strain XvP26 of Xanthomonas axonopodis pv. vesicatoria, were transferred by conjugation to a recipient strain of the bacterium. The chromosomal gene transfer was verified by analyses of the genomes of donor, recipient, and putative transconjugants for plasmid profiles, by polymorphism of DNA bands obtained by digesting total genomic DNA by a rare-cutting endonuclease and pulsed-field gel electrophoresis, and by Southern hybridization with a probe containing the copper genes. Transfer of kanamycin resistance to a recipient strain, associated with Tn5 insertion into the chromosome of another strain of the bacterial spot pathogen, was also verified. The frequency of kanamycin resistance transfer to recipient was more than 75 times greater in pepper leaves than in vitro. The transfer of chromosomal sequences containing the hypersensitive reaction and pathogenicity (hrp) genes and pigmentation (pig) genes was linked with transfer of kanamycin resistance (Tn5). Horizontal transfer in planta of the chromosomal genes (i.e., cop, pig, hrp, and Tn5 sequences) among strains of X. axonopodis pv. vesicatoria means that horizontal chromosomal gene transfer is possible in nature. This type of gene transfer may explain the presence of great diversity among strains of the bacterial spot pathogen in terms of DNA polymorphism and may also explain the apparent horizontal transfer of hrp sequences among pathovars of Xanthomonas. | 18,944,660 |
Identification of Subpopulations of Colletotrichum acutatum and Epidemiology of Almond Anthracnose in California. | ABSTRACT In recent years, almond anthracnose has developed into a major problem for the California almond industry. The identification of the causal pathogen as Colletotrichum acutatum was confirmed using species-specific primers and restriction fragment length polymorphisms of ribosomal DNA in comparative studies with isolates of C. acutatum from strawberry and C. gloeosporioides from citrus. Two distinct clonal subpopulations among the almond isolates of C. acutatum were identified. These two subpopulations differed in their colony appearance (pink versus gray cultures), conidial morphology, virulence in laboratory inoculation studies, temperature relationships for growth, and molecular fingerprints using random and simple-repeat primers in polymerase chain reactions. Both subpopulations were commonly isolated from the same orchard or even the same fruit. In other orchards, one subpopulation predominated over the other subpopulation. Using random, simple-repeat, and species-specific primers, isolates of the almond anthracnose pathogen from Israel were very similar to the California isolates that produce gray colonies. In addition to fruit, the pathogen was isolated from blighted blossoms, water-soaked or necrotic leaf lesions, symptomless peduncles, and spurs and wood from branches showing dieback symptoms, indicating that the amount of tissue that may be infected is more extensive than previously considered. Overwintering fruit mummies were identified as inoculum sources for early spring infections. Growth studies using almond kernels with different moisture contents indicated that postharvest damage of stored kernels likely originates from preharvest field infections. | 18,944,662 |
Microbial Antagonism at the Root Level Is Involved in the Suppression of Fusarium Wilt by the Combination of Nonpathogenic Fusarium oxysporum Fo47 and Pseudomonas putida WCS358. | ABSTRACT Two biological control agents, nonpathogenic Fusarium oxysporum Fo47 and Pseudomonas putida WCS358, were evaluated for suppression of Fusarium wilt of flax grown in nutrient solution and for suppression of the population density and metabolic activity of the causal organism F. oxysporum f. sp. lini strain Foln3GUS on root surfaces. Due to the presence of an introduced gusA reporter gene construct in Foln3GUS, the pathogen expressed beta-glucuronidase activity that was related to its carbon metabolism. At a Fo47 to Foln3GUS inoculum ratio of 100:1, both the population density of the pathogen and the beta-glucuronidase activity on and in flax roots were reduced by the nonpathogenic strain, and Fusarium wilt was suppressed. At a Fo47 to Foln3GUS inoculum ratio of 10:1, Fo47 decreased the severity of Fusarium wilt to a smaller extent and it also reduced beta-glucuronidase activity without reducing the density of Foln3GUS on flax roots. At a nonpathogenic to pathogenic Fusarium strains ratio of 10:1, the addition of P. putida WCS358 further suppressed Fusarium wilt and the density of the pathogen at the root level, whereas a mutant of WCS358 deficient in pseudobactin production had no significant effect. Iron availability to WCS358 on flax roots, assessed by ice-nucleation activity conferred from a transcriptional fusion (pvd-inaZ) of an ice-nucleation reporter gene to an iron-regulated promoter, was sufficiently low to allow pseudobactin production. P. putida WCS358 did not reduce the severity of Fusarium wilt of flax when inoculated without Fo47, and it did not improve disease suppression achieved by high inoculum doses of Fo47 (a Fo47 to Foln3GUS ratio of 100:1). Together, these data provide evidence that (i) suppression of Fusarium wilt of flax by Fo47 is related to reductions in the population density and metabolic activity of the pathogen on the root surface; (ii) WCS358 can enhance the biological control activity of Fo47, but this enhancement depends on the population of Fo47 relative to the pathogen; and (iii) pseudobactin contributes to suppression of Fusarium wilt by the combination of Fo47 and WCS358 on roots in which conditions are conducive to pseudobactin production by the bacterium. | 18,944,664 |
Sampling in seed health testing. | ABSTRACT Seed health tests are usually performed on a sample of a seed lot; therefore, it is crucial that the test sample be as homogeneous as possible and representative of the lot. Seed sampling procedures appropriate for seed health testing have been developed by seed testing organizations such as the International Seed Testing Association and the Association of Official Seed Analysts. Seed lot size is generally not a constraint when the distribution of contaminated/infected seed in the lot is relatively homogeneous; if the distribution is heterogeneous, increased sampling intensity is required. Sample size is determined by the damage threshold (intolerable infection level) for the pathogen and the probability of detection desired, commonly 95 or 99%. The probability of detection at a given damage threshold is greater as sample size increases, and this probability and the appropriate sample size can be determined by statistical methods. Most seed health tests utilize qualitative data based on the presence or absence of the pathogen in the test sample, with the lot being rejected if the pathogen is detected in the sample and accepted if the sample is negative. | 18,944,666 |
Effects of Temperature and Wetness Duration on Infection of Peanut Cultivars by Cercospora arachidicola. | ABSTRACT The effects of temperature and duration of wetness (relative humidity >/=95%) on infection of three peanut cultivars by Cercospora arachidicola were determined under controlled conditions. Plants of the Spanish cv. Spanco and the runner cvs. Florunner and Okrun were exposed to constant temperatures of 18 to 30 degrees C during 12-h periods of wetness each day that totaled 12 to 84 h following inoculation of leaves with conidia. Severity of disease, measured by either lesion density (number per leaf) or lesion size (diameter), was greatest for 'Spanco', intermediate for 'Florunner', and lowest for 'Okrun' in each of two experiments. Lesion density was evaluated further because it was an indicator of both the occurrence and degree of infection. Nonlinear regression analysis was employed to evaluate the combined effects of temperature (T) and wetness duration (W) on lesion density (Y). In the regression model, the Weibull function characterized the monotonic increase of Y with respect to W, while a hyperbolic function characterized the unimodal response of Y with respect to T. Parameters for the intrinsic rate of change with respect to W (b), the intrinsic rate of change with respect to T (f), the optimal value of T (g), and the upper limit (e) when T is optimum (T = g) were estimated for each cultivar and experiment. The effect of cultivar was characterized primarily by differences in the upper limit parameter e. In each experiment, e was greatest for 'Spanco', intermediate for 'Florunner', and least for 'Okrun'. The effect of cultivar on b followed a pattern similar to that for e in experiment 1, but not in experiment 2. Differences among cultivars for estimates of f and g were small and inconsistent. Estimates for g were precise for each cultivar and experiment and fell within the range of 22.3 to 23.2 degrees C. Cultivar responses to T and W were further evaluated using data pooled over the two experiments. Parameter e was estimated for each cultivar, but common values of b, f, and g were estimated. At e = 22.8 degrees C, lesion density approached an upper limit of 96, 17, and 6 lesions per leaf for the cvs. Spanco, Florunner, and Okrun, respectively. These fitted values approximated the observed values of 86, 25, and 9 lesions per leaf for the respective cultivars. Cultivars varied in their response to W at a given T. At 22.8 degrees C, one lesion per leaf was expected following 26, 30, and 36 h of wetness for 'Spanco', 'Florunner', and 'Okrun', respectively. If temperature was increased to 28 degrees C, one lesion per leaf was expected following 36, 44, and 54 h of wetness for the respective cultivars. | 18,944,677 |
Evaluation of Maize Streak Virus Pathogenicity in Differentially Resistant Zea mays Genotypes. | ABSTRACT We devised a rapid technique for the objective and precise assessment of both the pathogenicity of maize streak virus (MSV) isolates and the MSV resistance of maize genotypes. The technique involves the use of agroinoculation to infect maize seedlings and the objective symptom evaluation by quantification of infection rates, stunting, and chlorotic leaf areas. In assessing the MSV resistance of 19 maize genotypes, we describe how the use of differentially virulent virus isolates enables the analysis of MSV resistance phenotypes, ranging from extremely susceptible to completely immune. We further demonstrate how quantification of chlorotic leaf areas by image analysis permits differentiation between degrees of MSV resistance that are indistinguishable from one another using currently employed symptom assessment approaches. Using chlorotic area measurements, we quantify the virulence of a diverse group of 10 MSV isolates and, through agroinoculation of differentially susceptible maize genotypes, we demonstrate the use of our technique in evaluating the pathogenicity of these isolates. | 18,944,683 |
Cytology of Infection of Maize Seedlings by Fusarium moniliforme and Immunolocalization of the Pathogenesis-Related PRms Protein. | ABSTRACT We have investigated the histology of infection of maize seedlings by Fusarium moniliforme in association with a biochemical host defense response, the accumulation of the PRms (pathogenesis-related maize seed) protein. Light microscopy of trypan blue-stained sections and scanning electron microscopy revealed direct penetration by F. moniliforme hyphae through the epidermal cells of the seedling and colonization of the host tissue by inter- and intracellular modes of growth. Pathogen ingress into the infected tissue was associated with the induction of defense-related ultrastructural modifications, as exemplified by the formation of appositions on the outer host cell wall surface, the occlusion of intercellular spaces, and the formation of papillae. Cellular and subcellular immunolocalization studies revealed that PRms accumulated at very high levels in those cells types that represent the first barrier for fungal penetration such as the aleurone layer of germinating seeds and the scutellar epithelial cells of isolated germinating embryos. A highly localized accumulation of PRms within papillae of the inner scutellar parenchyma cells also occurred, suggesting that signaling mechanisms that lead to the accumulation of PRms in papillae of cell types that are distant from the invading pathogen must operate in the infected maize tissues. Our study also revealed the presence of a large number of fungal cells with an abnormal shape that showed PRms-specific labeling. PRms was found to accumulate in clusters over the fungal cell wall. Taken together, the occurrence of PRms in cell types that first establish contact with the pathogen, as well as in papillae, and in association with fungal cell walls suggests that PRms may have a function in the plant defense response. | 18,944,701 |
Survey Methods for Assessment of Citrus Tristeza Virus Incidence When Toxoptera citricida Is the Predominant Vector. | ABSTRACT Citrus tristeza virus (CTV) incidence may be assessed by sampling groups of citrus trees, recording the groups as CTV positive (one or more infected trees) or CTV negative (no infected trees), and then calculating disease incidence at the scale of the individual tree by means of a formula involving incidence at the group scale and the number of trees per group. This procedure works well when the CTV status of a tree can be regarded as independent of the CTV status of other trees in the same group. This is the case when the main vector species is Aphis gossypii and the groups comprise four adjacent trees, because the spatial pattern of CTV incidence at the within-group scale can be regarded as random. However, when the main vector species is Toxoptera citricida, this simple procedure is not appropriate, because the spatial pattern of CTV incidence at the within-group scale cannot be regarded as random. Using field data and computer simulation, an alternative procedure for assessment of CTV incidence when the main vector species is T. citricida was devised and tested. In the alternative procedure, the sampling scheme is operationally identical to that used when the main vector species is A. gossypii, but the calculation of CTV incidence at the scale of the individual tree is based on incidence at the group scale and an effective sample size. The analysis of CTV-incidence data collected from a number of citrus blocks in reasonable geographical and temporal proximity and the use of CTV-detection methods more sensitive than the enzyme-linked immunosorbent assay used here are also discussed. | 18,944,721 |
A Corn Trypsin Inhibitor with Antifungal Activity Inhibits Aspergillus flavus alpha-Amylase. | ABSTRACT In this study, we found that the inhibition of fungal growth in potato dextrose broth (PDB) medium by the 14-kDa corn trypsin inhibitor (TI) protein, previously found to be associated with host resistance to aflatoxin production and active against various fungi, was relieved when exogenous alpha-amylase was added along with TI. No inhibitory effect of TI on fungal growth was observed when Aspergillus flavus was grown on a medium containing either 5% glucose or 1% gelatin as a carbon source. Further investigation found that TI not only inhibited fungal production of extracellular alpha-amylase when A. flavus was grown in PDB medium containing TI at 100 mug ml(-1) but also reduced the enzymatic activity of A. flavus alpha-amylase by 27%. At a higher concentration, however, TI stimulated the production of alpha-amylase. The effect of TI on the production of amyloglucosidase, another enzyme involved in starch metabolism by the fungus, was quite different. It stimulated the production of this enzyme during the first 10 h at all concentrations studied. These studies suggest that the resistance of certain corn genotypes to A. flavus infection may be partially due to the ability of TI to reduce the production of extracellular fungal alpha-amylase and its activity, thereby limiting the availability of simple sugars for fungal growth. However, further investigation of the relationship between TI levels and fungal alpha-amylase expression in vivo is needed. | 18,944,733 |
Amy1, the alpha-Amylase Gene of Aspergillus flavus: Involvement in Aflatoxin Biosynthesis in Maize Kernels. | ABSTRACT Aspergillus flavus is the causal agent of an ear and kernel rot in maize. In this study, we characterized an alpha-amylase-deficient mutant and assessed its ability to infect and produce aflatoxin in wounded maize kernels. The alpha-amylase gene Amy1 was isolated from A. flavus, and its DNA sequence was determined to be nearly identical to Amy3 of A. oryzae. When Amy1 was disrupted in an aflatoxigenic strain of A. flavus, the mutant failed to produce extracellular alpha-amylase and grew 45% the rate of the wild-type strain on starch medium. The mutant produced aflatoxin in medium containing glucose but not in a medium containing starch. The alpha-amylase-deficient mutant produced aflatoxin in maize kernels with wounded embryos and occasionally produced aflatoxin only in embryos of kernels with wounded endosperm. The mutant strain failed to produce aflatoxin when inoculated onto degermed kernels. In contrast, the wild-type strain produced aflatoxin in both the endosperm and embryo. These results suggest that alpha-amylase facilitates aflatoxin production and growth of A. flavus from a wound in the endosperm to the embryo. A 14-kDa trypsin inhibitor associated with resistance to A. flavus and aflatoxin in maize also inhibited the alpha-amylase from A. flavus, indicating that it is a bifunctional inhibitor. The inhibitor may have a role in resistance, limiting the growth of the fungus in the endosperm tissue by inhibiting the degradation of starch. | 18,944,734 |
Evidence of Cytoplasmic Inheritance of Virulence in Cronartium ribicola to Major Gene Resistance in Sugar Pine. | ABSTRACT Tests for Mendelian segregation of virulence and avirulence in Cronartium ribicola, causal agent of white pine blister rust, to a major gene (R) for resistance in sugar pine were made using haploid basidiospore progenies from single diploid telia as inoculum on resistant genotypes. The telia were sampled from a small deme in the Siskyou Mountains of northern California, where a few mature sugar pines known to be Rr genotypes had become infected after withstanding the chronic blister rust epidemic for several decades and where intermediate frequencies of virulence in the ambient basidiospore population were subsequently measured. Infection type on inoculated seedlings with R was qualitative: all progenies of 81 single telia tested over 3 different years were either virulent (compatible) or avirulent (inducing hypersensitive necrosis), never a mixture of both reactions. The complete absence of heterozygotes in the telia population is strong evidence that virulence is not controlled by a nuclear gene. The data are consistent with earlier tests showing that basidiospore inoculum derived from aeciospores isolated from infected Rr trees produced mostly (>90%) virulent reactions on R- seedlings. The evidence indicates that transmission of virulence is uniparental via the cytoplasm of aeciospores. Exchange of spermatia between haploid thalli does not appear to be involved. | 18,944,758 |
Biotrophic Development of Sporisorium reilianum f. sp. zeae in Vegetative Shoot Apex of Maize. | ABSTRACT Sporisorium reilianum f. sp. zeae is the causal agent of maize head smut, a disease present in several regions of France. A cytological study was carried out to describe a key step of the fungal etiology, in which the mycelium invades the vegetative shoot apex. Light and transmission electron microscopy observations show that the fungus is mostly intracellular and suggest that it passes through the host cell wall by lysis and mechanical pressure. The hyphae are surrounded by an amorphous vesicle-rich layer limited by a membrane related to the host plasmalemma. The encasement can be considered as an exchange zone between the plant and the fungus. The infected host cells appear normal; therefore, the fungus seems to act like a biotrophic endophyte. | 18,944,766 |
Molecular Identification and Phylogenetic Grouping of Diaporthe phaseolorum and Phomopsis longicolla Isolates from Soybean. | ABSTRACT Diaporthe phaseolorum and Phomopsis longicolla isolates from soybean were examined using traditional mycological characteristics and molecular methods. Cultural characteristics including types of fruiting bodies and conidia were assessed for isolates collected from soybean stems and seeds. Cultures were identified as P. longicolla, D. phaseolorum var. caulivora, D. phaseolorum var. meridionalis, or D. phaseolorum var. sojae. Molecular markers for these groups were developed and analyzed using polymerase chain reaction restriction fragment length polymorphisms (PCR-RFLP) and DNA sequencing in the internal transcribed spacer (ITS) and the 5.8S ribosomal DNA. The ITS(4) and ITS(5) primers amplified PCR products for all isolates studied. Gel electrophoresis of undigested PCR products and DNA sequencing produced various fragment lengths including 604 bp for P. longicolla, 602 and 603 bp for D. phaseolorum var. caulivora, 603 bp for D. phaseolorum var. meridionalis, and from 597 to 609 bp for D. phaseolorum var. sojae. Digestion of these PCR products with enzymes AluI, HhaI, MseI, RsaI, and ScrFI resulted in distinct bands for identification of P. longicolla and the varieties of D. phaseolorum I. All P. longicolla, D. phaseolorum var. caulivora, and D. phaseolorum var. meridionalis isolates were distinguished using AluI and HhaI with RsaI or ScrFI. The banding patterns of D. phaseolorum var. sojae isolates were complex and were separated into 11 subgroups after digestion with AluI, HhaI, MseI, RsaI, and ScrFI. Phylogenetic analysis of 20 isolates of D. phaseolorum and P. longicolla based on the DNA sequence of the ITS region resolved six clades termed A, B, C, D, E, and F. Clade A included all sequenced D. phaseolorum var. caulivora isolates, two from Italy and one from the United States. Isolates in clade B were exclusively associated with D. phaseolorum var. meridionalis. Clades A and B formed a well-supported monophyletic group. Isolates in clades C, D, E, and F were morphologically defined as isolates of P. longicolla, D. phaseolorum var. sojae, and Diaporthe spp. The ITS sequences similarity of seven geographically diverse P. longi-colla isolates illustrated that P. longicolla isolates have a similar genetic background, with some affiliations to some D. phaseolorum var. sojae isolates. Morphological characteristics of the isolates along with the terminal clades of the ITS phylogeny suggest that P. longicolla is an individual species, D. phaseolorum var. caulivora and D. phaseolorum var. meridionalis are varieties of D. phaseolorum, and D. phaseolorum var. sojae is either several varieties of D. phaseolorum or possibly several distinct species. | 18,944,833 |
Phytoplasma: ecology and genomic diversity. | ABSTRACT The recent development of molecular-based probes such as mono- and polyclonal antibodies, cloned phytoplasma DNA fragments, and phytoplasma-specific primers for polymerase chain reaction (PCR) has allowed for advances in detection and identification of uncultured phytoplasmas (formerly called mycoplasma-like organisms). Comprehensive phylogenetic studies based on analysis of 16S ribosomal RNA (rRNA) or both 16S rRNA and ribosomal protein gene operon sequences established the phylogenetic position of phytoplasmas as members of the class Mollicutes, and the revealed phylogenetic interrelationships among phytoplasmas formed a basis for their classification. Based on restriction fragment length polymorphism (RFLP) analysis of PCR-amplified 16S rRNA gene sequences, phytoplasmas are currently classified into 14 groups and 38 subgroups that are consistent with groups delineated based on phylogenetic analysis using parsimony of 16S rRNA gene sequences. In the past decades, numerous phyto-plasma strains associated with plants and insect vectors have been identified using molecular-based tools. Genomic diversity of phytoplasma groups appears to be correlated with their sharing common insect vectors, host plants, or both in nature. The level of exchange of genetic information among phytoplasma strains in a given group is determined by three-way, vector-phytoplasma-plant interactions. A putative mechanism for the creation of new ecological niches and the evolution of new ecospecies is proposed. | 18,944,840 |
The importance of archival and herbarium materials in understanding the role of oospores in late blight epidemics of the past. | ABSTRACT Nineteenth and early twentieth century botanists and mycologists collected healthy and infected plant materials from many regions of the world. Some of these plant collections preserved in herbaria around the world contain samples that are of considerable significance to epidemiologists, population biologists, and botanists. The advent of the polymerase chain reaction (PCR) and the development of molecular marker technology has made DNA amplification from herbarium material a reality. In this mini-review, archival letters and herbarium samples are used to track the historical role of oospores in the biology of the potato late blight pathogen. DNA was successfully amplified by PCR with the Phytophthora infestans-specific PCR primer, PINF, and the universal primer, ITS 5, from oospores observed in a field sample of potato collected by G. P. Clinton in 1902. This experiment demonstrates the potential to utilize molecular methods to amplify DNA from historical samples of the late blight pathogen and represents the earliest definitive record of oospores of the pathogen in field samples in the United States. Studies based upon such materials and techniques, although high risk and laborious, have the potential to open a new window to epidemics of the past. | 18,944,844 |
Unusual Cytoplasmic Inclusions Induced in Tobacco by Peanut Stunt Virus Subgroup II Strains Map to RNA3. | ABSTRACT Infection of tobacco protoplasts or leaf tissues with peanut stunt virus (PSV) subgroup II strains induced the production of unusual cytoplasmic ribbon-like inclusions. The inclusion structures appeared as long, thin, densely staining sheets that were prevalent within the cytoplasm, accumulating most commonly near vacuoles. Numerous virions and ribosomes could be seen adjacent to the inclusion surfaces. The formation of these novel inclusions appeared to be subgroup specific, since infection of tobacco with PSV strains W and B (subgroup II), but not strains ER, V, and J (subgroup I), induced the inclusions. Furthermore, inclusion formation was shown to be host specific, because the inclusions were not detected in either of two leguminous host species infected with PSV subgroup II strains. Using tobacco protoplasts electroporated with various assortments of infectious RNA transcripts derived from cDNA clones of genomic RNAs of PSV-ER and PSV-W, we demonstrated that induction of the unusual ribbon-like inclusions maps to PSV-W (subgroup II) RNA3. This conclusion is consistent with the finding that PSV strain BV-15, a natural intraspecific reassortant that derives its RNA2 and RNA3 from a subgroup I strain, did not induce inclusion formation. | 18,944,853 |
Transmission of Pineapple Mealybug Wilt-Associated Virus by Two Species of Mealybug (Dysmicoccus spp.). | ABSTRACT Closterovirus-like particles associated with mealybug wilt of pineapple were acquired and transmitted by the pink pineapple mealybug, Dysmicoccus brevipes, and the gray pineapple mealybug, D. neobrevipes. Mealybugs acquired pineapple mealybug wilt-associated virus (PMWaV) from infected pineapple plants or detached leaves. The virus was detected in plants by tissue blot immunoassay and confirmed by immunosorbent electron microscopy. Plants exposed to mealybugs reared on PMWaV-free pineapple tissue remained uninfected. The presence of ants was correlated with an increased rate of virus spread when caged with D. brevipes. All stages of D. neobrevipes acquired PMWaV, although vector efficiency decreased significantly in older adult females. The probability of a single third-instar immature transmitting the virus was 0.04. Both species of mealybug acquired and transmitted PMWaV from infected pineapple material that had been clonally propagated for decades, and both species acquired PMWaV from sources previously infected with the virus by the other mealybug species. | 18,944,858 |
Phylogenetic relationships within the family potyviridae: wheat streak mosaic virus and brome streak mosaic virus are not members of the genus rymovirus. | ABSTRACT The complete nucleotide sequence of wheat streak mosaic virus (WSMV) has been determined based on complementary DNA clones derived from the 9,384-nucleotide (nt) RNA of the virus. The genome of WSMV has a 130-nt 5' leader and 149-nt 3'-untranslated region and is polyadenylated at the 3' end. WSMV RNA encodes a single polyprotein of 3,035 amino acid residues and has a deduced genome organization typical for a member of the family Potyviridae (5'-P1/HC-Pro/P3/6K1/CI/6K2/VPg-NIa/NIb/CP-3'). Because WSMV shares with ryegrass mosaic virus (RGMV) the biological property of transmission by eriophyid mites, WSMV has been assigned to the genus Rymovirus, of which RGMV is the type species. Phylogenetic analyses were conducted with complete polyprotein or NIb protein sequences of 11 members of the family Potyviridae, including viruses of monocots or dicots and viruses transmitted by aphids, whiteflies, and mites. WSMV and the monocot-infecting, mite-transmitted brome streak mosaic virus (BrSMV) are sister taxa and share a most recent common ancestor with the whitefly-transmitted sweet potato mild mottle virus, the type species of the proposed genus "Ipomovirus." In contrast, RGMV shares a most recent common ancestor with aphid-transmitted species of the genus Potyvirus. These results indicate that WSMV and BrSMV should be classified within a new genus of the family Potyviridae and should not be considered species of the genus Rymovirus. | 18,944,883 |
Immunocytochemical Evidence that Secretion of Pectin Occurs During Gel (Gum) and Tylosis Formation in Trees. | ABSTRACT During gel (gum) formation in angiosperm trees, fibrillar material accumulated in protective layers of xylem parenchyma cells before being secreted across half-bordered pit membranes into vessel elements. Immunogold labeling demonstrated that this fibrillar material was mainly composed of partially esterified pectic polysaccharides. The primary wall of expanding tyloses, an extension of the parenchyma protective layer, secreted similar pectic substances to completely block vessel elements. In most studies, these occluding structures were reported to be formed in response to causative factors such as aging processes, injuries, or infections. Current observations support the view that partial to complete embolism, which almost always accompanies these factors, might be the main cause triggering the formation of vessel occlusions. Whereas pectin seems to be the basic component of gels (gums) and of the external layer of tyloses, other substances, such as phenols, were also detected either as a part of these plugs or as accumulations beside them in vessels. Finally, it is proposed that the term 'gel' instead of 'gum' be used in future studies to describe the occluding material secreted by ray and paratracheal parenchyma cells. | 18,944,900 |
Establishment of Bacterial Antagonists of Erwinia amylovora on Pear and Apple Blossoms as Influenced by Inoculum Preparation. | The influence of inoculum preparation on the establishment of bacterial antagonists that suppress fire blight and Erwinia amylovora on blossoms was evaluated. Aqueous suspensions of Pseudomonas fluorescens A506, E. herbicola C9-1R, or E. amylovora 153N were prepared from cells harvested from the surface of an agar medium or from cells that were lyophilized after culture under similar conditions. Bacterial suspensions (1 x 10(8) CFU/ml) were sprayed on pear and apple trees at 50% bloom near midday. The incidence of recovery (proportion of blossoms containing detectable populations) and the population sizes of the bacteria on individual blossoms with detectable populations were followed over a period of several days. Fluorescent microspheres (1 mum in diameter) were added to sprays at a concentration of 1 x 10(7) microspheres per ml to mark blossoms that were open during application of bacteria. After dilution-plating, the stigmas and styles of each blossom were examined for the presence of microspheres with an epifluorescence microscope. In three of five trials, bacteria applied as suspensions of lyophilized cells were recovered from a greater proportion of blossoms than bacterial cells harvested directly from culture media. Every blossom harvested within 6 days after spraying had microspheres present on the surfaces of the styles and stigmas; thus, lack of establishment of detectable populations, rather than escape of blossoms from spray inoculation, accounted for the differences in proportion of blossoms colonized by the different preparations of bacteria. The use of lyophilized cells in field trials decreased variability in the establishment of bacteria on blossoms. | 18,944,901 |
Satellite RNA of cucumber mosaic cucumovirus spreads epidemically in natural populations of its helper virus. | ABSTRACT Three hundred thirty-eight isolates of cucumber mosaic cucumovirus (CMV), sampled from natural populations in six areas of Spain between 1989 and 1996, were screened for the presence of satellite RNA (satRNA). The frequency of CMV isolates with satRNA approached 1.00 in Valencia (east Spain) between 1990 and 1994 where a tomato necrosis epidemic induced by CMV+satRNA had started in 1986 and was smaller north and west of this area in 1992 and 1993. After 1994, satRNA almost disappeared from all CMV populations. Genetic typing of satRNA variantswas done by ribonuclease protection assay, and from these data, genetic distances were estimated for any pair of satRNA variants. CMV-satRNA populations were highly diverse, containing 0.07865 nucleotide substitutions per site on average. Data also showed that the whole compared set of 100 satRNA variants form a single population that is not structured according to place, year, host plant, or strain of helper virus (HV). This is in sharp contrast with the metapopulation structure of the Spanish CMV population. Thus, the genetic structure and dynamics of populations of CMV and its satRNA are not coupled. This shows that CMV-satRNA spreads epidemically, as a hyperparasite, in the population of its HV. This conclusion is relevant to the use of CMV-satRNA as a biocontrol agent of CMV. | 18,944,903 |
Effects of a Cover Crop on Splash Dispersal of Colletotrichum acutatum Conidia. | ABSTRACT A rain simulator, with generated rains of 11 and 30 mm/h, was used to determine the effect of a cover crop or intercrop on the splash dispersal of Colletotrichum acutatum conidia. Dispersal through sudangrass, which can be used as a 'living mulch', was tested at two planting densities (140 or 280 kg/ha) and two heights (5 and 20 cm) and compared with a control consisting of a bare soil. Dispersal of C. acutatum conidia was assessed by counting colonies formed from spore-bearing splash droplets deposited in sheltered petri plates containing a selective medium. Both a cover crop and rain intensity significantly affected splash dispersal as measured by the interpolated total number of colonies (denoted by Sigma) from 0 to 72 cm from the inoculum source and in a time span of 61 min of generated rain (P < 0.001). However, there was no significant interaction of cover crop and intensity (P > 0.90). Dispersal with a 30-mm/h rain was higher than dispersal with a 11-mm/h rain, and presence of a cover crop significantly reduced dispersal compared with bare soil (P < 0.001). Of the treatments with sudangrass, cover crop planting density did not affect dispersal overall, but there was greater spore dispersal with the taller sudangrass at the higher planting density, due in part to the higher rate of water splashing with the tall grass compared with the short grass. Spore deposition in the petri plates could be functionally related to distance and time using a diffusion-type model, and parameter estimates could be used to explain the effects of cover crop on Sigma. Although the relationship between cover crop properties and splash dispersal is complex, results show the potential beneficial effects of the cover crop on disease management. | 18,944,906 |
Survival of Fusarium moniliforme, F. proliferatum, and F. subglutinans in Maize Stalk Residue. | ABSTRACT The roles of residue size and burial depth were assessed in the survival of Fusarium moniliforme, F. proliferatum, and F. subglutinans in maize stalk residue. Stalk pieces (small or large sizes) were soaked in a spore suspension of F. moniliforme, F. proliferatum, or F. subglutinans and placed in a field on the soil surface or buried at 15- or 30-cm depths. Residue pieces were recovered periodically, cultured on a selective medium, and microscopically examined for the presence of the inoculated Fusarium species. After 630 days, the inoculated Fusarium species were recovered from 0 to 50% of the inoculated stalk pieces in a long-term, continuous maize field, from 0 to 28% of the inoculated stalk pieces placed in a maize/soybean/oat rotation field, and from 0 to 25% of the noninoculated stalk pieces at both locations. Residue size and residue depth had significant effects on survival, but there were significant interactions among strain, depth, residue size, and time. Up to 343 days after placement in the field, survival of the three Fusarium species was not consistently different between buried residues and surface residues, but after 630 days, survival was greater from surface residues. Overall, fungus survival decreased more slowly in the surface residues than in the buried residues. Linear coefficients of determination ranged from 0.35 to 0.82 for the surface residues and from 0.81 to 0.98 for the buried residues. Decline in survival over time followed a more linear pattern in buried residues than in surface residues. Vegetative compatibility tests confirmed that F. moniliforme, F. proliferatum, and F. subglutinans strains can survive at least 630 days in surface or buried maize residue. These results demonstrate that maize residue can act as a long-term source of inoculum for infection of maize plants by these three Fusarium species. | 18,944,908 |
Molecular Evidence of Distinct Introductions of the European Race of Gremmeniella abietina into North America. | ABSTRACT The presence of the European (EU) race of Gremmeniella abietina var. abietina, the causal agent of Scleroderris canker of conifers, was first reported in North America in 1975 in the northeastern United States and subsequently in southern Quebec and Newfoundland during the late 1970s, where it quickly became established. We analyzed DNA profiles in samples from a historic collection of G. abietina var. abietina that included some of the first isolates of the EU race reported in the United States to test hypotheses concerning the G. abietina var. abietina epidemic in North America. Genetic diversity was partitioned by an analysis of molecular variance with haplotype frequencies and distances. Genetic differentiation was high between populations in continental North America and Newfoundland (between region differentiation, Phi(ct) = 0.665, P < 0.001). This result was not consistent with the hypothesis of a single introduction of the pathogen into the northeastern United States followed by secondary spread into northeastern Canada. In contrast, small levels of genetic differentiation were observed among continental North American populations (Phi(ct) = 0.047, P = 0.079), suggesting gene flow among these populations. A single haplotype of G. abietina var. abietina dominated the continental populations (80% of the isolates) but was absent from Newfoundland and Europe. Five haplotypes were found in the New-foundland population, all of which were either absent or very rare on the continent. Populations from continental North America clustered together and were distinct from a second cluster composed of European and Newfoundland populations. A phylogenetic analysis of the haplotypes indicated that some of the rare haplotypes may have derived from somatic mutations, whereas others probably occurred as the result of new introductions. The results are consistent with a scenario of distinct primary introductions of this pathogen into Newfoundland and continental eastern North America followed by secondary asexual propagation. | 18,944,913 |
Colonization of Host Plants by the Fire Blight Pathogen Erwinia amylovora Marked with Genes for Bioluminescence and Fluorescence. | ABSTRACT To follow the movement of Erwinia amylovora in plant tissue without dissection, this bacterium was marked with either the lux operon from Vibrio fischeri or the gfp gene from the jellyfish Aequorea victoria, both carried on multicopy plasmids and expressed under the control of the lac promoter from Escherichia coli. Movement of the pathogen was visualized in leaves, stems, and roots of apple seedlings, and migration of E. amylovora was traced from inoculation sites in the stem to as far as the roots. Green fluorescent E. amylovora cells were observed in the xylem and later appeared to break out of the vessels into the intercellular spaces of the adjacent parenchyma. Inoculation in the intercostal region of leaves caused a zone of slow necrosis that finally resulted in bacterial invasion of the xylem vessels. Labeled bacteria could also be seen in association with the anchor sites of leaf hairs. Distortion of the epidermis adjacent to leaf hairs created openings that were observed by scanning electron microscopy. As the intercostal region, the bases of leaf hairs provided E. amylovora access to intact xylem vessels, which allowed further distribution of the pathogen in the host plant. | 18,944,920 |
Pathogenicity of Streptomyces scabies Mutants Altered in Thaxtomin A Production. | ABSTRACT To investigate the role of thaxtomin A in the pathogenicity of Streptomyces scabies, mutants altered in thaxtomin A production were obtained by N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis. Mutants of S. scabies EF-35 could be differentiated according to levels of thaxtomin production. Mutants M1, M8, and M19 produced 2 to 20 times less thaxtomin A in oat bran medium than did EF-35. M1 and M19 were deficient in tryptophan catabolism. Thaxtomin production was reduced by about 300 times in mutant M16, which was a glutamic acid auxotroph. No thaxtomin A was detected in M13 culture supernatant. This mutant had a normal growth rate, was prototrophic, and catabolized tryptophan. Pathogenicity of mutants was tested on radish and potato. Mutants M1, M8, and M19 were pathogenic but, in most cases, less virulent than EF-35. M13 and M16 were nonpathogenic. These results suggest that thaxtomin A is an important pathogenicity determinant in S. scabies. | 18,944,924 |
Isolation, Characterization, and Distribution of a Biocontrol Fungus from Cysts of Heterodera glycines. | ABSTRACT Seventy-six populations of Heterodera glycines were collected from 33 counties in 10 states of the United States along the Mississippi and Missouri Rivers in 1992 and 1993. A sterile hyphomycete fungus of an unnamed taxon, designated ARF18 and shown to be a parasite of eggs of H. glycines, was isolated from eggs and cysts of 10 of the populations from Kentucky, Louisiana, Mississippi, and Tennessee. Ten isolates of ARF18 obtained in this study and seven isolates obtained in earlier studies were characterized for cultural morphology on several growth media, the ability to produce sclerotium-like structures (SLS) on cornmeal agar, growth rates, pathogenicity to eggs of H. glycines in vitro, and mitochondrial (mt) DNA restriction fragment length polymorphisms (RFLPs). All 17 isolates of ARF18 readily grew on potato dextrose agar, cornmeal agar, and nutrient agar. Based on colony morphology and SLS appearance on cornmeal agar, the isolates could be grouped into two morphological phenotypes. Isolates that produced SLS that were composed of a compact mass of hyphae were designated ARF18-C, whereas isolates that produced SLS composed of a mass of loosely clumped hyphae were designated ARF18-L. Only minor differences in growth rates were detected among the ARF18-C and ARF18-L isolates. All 10 ARF18-C isolates, which were from Arkansas, Louisiana, Mississippi, and Tennessee, belonged to a single mtDNA RFLP haplotype. The seven ARF18-L isolates shared many comigrating mtDNA restriction fragments with one another, but belonged to three distinct mtDNA RFLP haplotypes. Ability to infect eggs of H. glycines in vitro varied considerably among the various isolates of ARF18. In particular, several of the ARF18-C isolates were consistently able to infect over 50% (mean = 70.0%, standard deviation = 16%) of the eggs of H. glycines, whereas ARF18-L infected eggs to a lesser degree (mean = 25%, standard deviation = 27%). ARF18-C was isolated only from H. glycines populations from below 37 degrees N latitude. The presence of ARF18 was associated with soils with Mg levels <314 kg/ha, cyst numbers >4.5 per 100 cm(3), and iron levels >203.5 kg/ha; or with Mg levels >314 kg/ha and Na levels <121 kg/ha. The widespread distribution of ARF18 and the ability of some isolates to aggressively colonize eggs of H. glycines are indications that it has potential as a biological control agent for H. glycines. | 18,944,928 |
Cytochemical Investigation of the Antagonistic Interaction Between a Microsphaeropsis sp. (Isolate P130A) and Venturia inaequalis. | ABSTRACT In an attempt to better understand the mode of action of the antagonistic fungus Microsphaeropsis sp., the interaction between this fungus and Venturia inaequalis was studied, using both light and electron microscopy. Cytological observations indicated that the antagonistic interaction between the two fungi likely involves a sequence of events, including (i) attachment and local penetration of Microsphaeropsis sp. into V. inaequalis hyphae; (ii) induction of host structural response at sites of potential antagonist entry; (iii) alteration of host cytoplasm; and (iv) active multiplication of antagonistic cells in pathogen hyphae, leading to host cell breakdown and release of the antagonist. The interaction was investigated further by gold cytochemistry. The use of gold-complexed beta-1,4-exoglucanase and wheat germ agglutinin/ovomucoid-gold complex to localize cellulosic beta-1,4-glucans and chitin monomers, respectively, resulted in regular labeling of V. inaequalis cell walls. This finding supports other studies refuting the classification of ascomycetes as only a glucan-chitin group. At an advanced state of parasitism, the labeling pattern of cellulose and chitin, which clearly showed that the level of integrity of these compounds was affected, suggested the production of cellulolytic and chitinolytic enzymes by Microsphaeropsis sp. Wall appositions formed in V. inaequalis in response to the antagonist's attack contained both cellulose and chitin. However, penetration of this newly formed material frequently succeeded. This study provides the first detailed picture of the cytological events associated with mycoparasitism in V. inaequalis. | 18,944,932 |
Comparison of ambisense m RNA of watermelon silver mottle virus with other tospoviruses. | ABSTRACT Double-stranded genomic RNAs (dsRNAs) extracted from Chenopodium quinoa infected with watermelon silver mottle virus (WSMV) were similar to those of tomato spotted wilt virus (TSWV, serogroup I) and impatiens necrotic spot virus (INSV, serogroup III), except that the S dsRNA of WSMV is 0.75 and 0.6 kbp longer than those of TSWV and INSV, respectively. The complete nucleotide sequence of the genomic M RNA of WSMV was determined from cDNA clones generated from separated M dsRNA. The M RNA is 4,880 nucleotides in length with two open reading frames (ORFs) in an ambisense organization. The M RNA-encoded nonstructural (NSm) ORF located on the viral strand encodes a protein of 312 amino acids (35 kDa), and the G1/G2 ORF located on the viral complementary strand encodes a protein of 1,121 amino acids (127.6 kDa). The RNA probe corresponding to the NSm or G1/G2 ORF of WSMV failed to hybridize with the M dsRNAs of TSWV and INSV. Comparison of M and S RNAs of WSMV, TSWV, INSV, and peanut bud necrosis virus (PBNV, serogroup IV) revealed a consensus sequence of eight nucleotides of 5'-AGAGCAAU...-3' at their 5' ends and 5'-...AUUGCUCU-3' at their 3' ends. The low overall nucleotide identities (56.4 to 56.9%) of the M RNA and the low amino acid identities of the NSm and G1/G2 proteins (30.5 to 40.9%) with those of TSWV and INSV indicate that WSMV belongs to the Tospovirus genus but is phylogenetically distinct from viruses in serogroups I and III. The M RNA of WSMV shares a nucleotide identity of 79.6% with that of PBNV, and the two viruses share 83.4 and 88.7% amino acid identities for their NSm and G1/G2 proteins, respectively. It is concluded that they are two related but distinct species of serogroup IV. In addition to the viral or viral complementary full-length M RNA, two putative RNA messages for the NSm gene and the G1/G2 gene, 1.0 and 3.4 kb, respectively, were detected from the total RNA extracted from WSMV-infected tissue of Nicotiana benthamiana. The 1.0- and 3.4-kb RNAs were also detected in the viral RNAs extracted from purified nucleocapsids, suggesting that the putative messages of the M RNA of WSMV can also be encapsidated by the nucleocapsid protein. | 18,944,959 |
Monoterpene and Phenolic Compound Concentrations in Water-Stressed Red Pine Inoculated with Sphaeropsis sapinea. | ABSTRACT Changes in monoterpene and phenolic compounds resulting from water stress and colonization by Sphaeropsis sapinea were examined for 9- and 11-year-old red pine trees in a plantation and 3-year-old seedlings in a growth chamber. Four treatments were assigned at random to individual trees in the field: no treatment, herbicide to kill surrounding weeds, supplemental water, and both herbicide and supplemental water. In the growth chamber, seedlings were either not watered (water stressed) or watered daily (nonstressed). Shoots were inoculated with agar plugs colonized with either S. sapinea isolates of morphotype A and B (field) or only isolates of morphotype A (growth chamber). Nine monoterpenes were detected in tissue extracts; the most common were alpha-pinene (59 to 74% of the total), beta-pinene (13 to 33% of the total), and delta-3-carene (1 to 5% of the total). Shoots inoculated with isolates of morphotype A had more severe symptoms and produced higher concentrations of monoterpenes in both experiments compared with the controls. In the growth chamber, inoculations with isolates of morphotype A caused higher concentrations of phenolics compared with the controls. In the field experiment, monoterpenes increased in quantity only in shoots of stressed trees inoculated with isolates of morphotype A. Isolates of morphotype B caused few symptoms and did not alter monoterpene concentrations. Increases in monoterpenes do not appear to be involved in the response to infection by morphotype A in nonstressed trees, and the role of phenolics is unclear. However, these results are consistent with previous observations that monoterpenes may be involved in the differences in aggressiveness between morphotypes on red pine. | 18,944,971 |
Molecular Evidence That Strain BV-15 of Peanut Stunt Cucumovirus Is a Reassortant Between Subgroup I and II Strains. | ABSTRACT In Northern hybridization assays, RNA1 of peanut stunt virus (PSV) strain BV-15 hybridized strongly with a cloned cDNA probe to RNA1 from strain PSV-W (subgroup II). Cloned probes to PSV-W RNA2 and RNA3, however, did not hybridize with the corresponding RNAs from strain BV-15. The complete nucleotide sequence of PSV-BV-15 RNA2 has been determined, and sequence comparison analysis showed that it is closely related to PSV subgroup I strains; the percent nucleotide sequence identity between PSV-BV-15 RNA2 and RNA2 sequences from PSV subgroup I and II strains were 90.5 and 75%, respectively. The possibility that PSV-BV-15 RNA2 may contain short regions derived from a subgroup II strain (i.e., represent a mosaic structure indicative of recombination) was investigated. Results indicated, however, that the entire PSV-BV-15 RNA2 sequence is derived from a subgroup I strain. PSV-BV-15 RNA3 has previously been shown to belong to subgroup I strains. These results thus establish that PSV strain BV-15 is a natural reassortant between PSV subgroups I and II strains. A reverse transcription-po-lymerase chain reaction assay is proposed for differentiating between this reassortant strain and PSV strains in subgroups I and II. | 18,944,976 |
Determinants of density- and frequency-dependent fitness in competing plant pathogens. | ABSTRACT Using mathematical models, we investigated how infection and sporulation characteristics of competing plant pathogens determine the density and frequency dependence of relative fitness. Two models, one for the infection stage and one for the sporulation stage of a pathogen's life cycle, describe reproductive output of pathogen strains in mixture as a function of the strains' population densities. Model parameters include infection and sporulation efficiencies, carrying capacities on leaves for sporulating lesions and spore production, and coefficients of interstrain competitive effects in both life cycle stages. Although the models were originally developed for rust fungi, they are generally applicable to any organism with distinct colonization (e.g., infection) and propagative (e.g., sporulation) life cycle stages. In this work, paired hypothetical strains were assigned equal baseline parameter values. Parameters were then altered one at a time for one or both strains, and relative fitness was calculated over a range of densities and strain frequencies. Except for infection efficiency, the fitness benefit conferred by an advantage in a single parameter was always density dependent. Relative fitness was frequency dependent whenever inter- and intrastrain competitive effects were not equal. These results suggest that the fitness of pathogens in nature is rarely fixed, but, rather, may typically be highly dependent on the densities and frequencies of all coexisting strains in a habitat. | 18,944,998 |
Binding of Tomato Spotted Wilt Virus to a 94-kDa Thrips Protein. | ABSTRACT Using protein blot assays, a 94-kDa thrips protein was identified that exhibited specific binding to tomato spotted wilt virus (TSWV) particles. Renaturation of the 94-kDa protein, which is conserved among the two major vector species of TSWV, Frankliniella occidentalis and Thrips tabaci, was crucial for its virus-binding properties, whereas under the same conditions no specific binding was observed with aphid (Myzus persicae) proteins. The 94-kDa protein species was present in all developmental stages of both vectoring thrips, whereas it was present mainly in the adult stage of a nonvectoring thrips species, Parthenothrips dracenae. Using antibodies against the different TSWV structural proteins, the G2 envelope glycoprotein was identified as the viral determinant involved. Because the virus-binding protein is present throughout the thrips body, but not in the gut, it may represent a receptor protein involved during circulation of the virus through its vector but probably not during viral uptake in the midgut. | 18,945,001 |
Populations of Xylella fastidiosa in Plants Required for Transmission by an Efficient Vector. | ABSTRACT Xylella fastidiosa, a xylem-limited bacterium that causes Pierce's disease (PD) of grapevine and other diseases, is transmitted efficiently by xylem-feeding leafhoppers. Acquisition of a PD strain of X. fastidiosa by the blue-green sharpshooter (BGSS) from five plant host species-grapevine (Vitis vinifera), Himalayan blackberry (Rubus discolor), California mugwort (Artemisia douglasiana), watergrass (Echinochloa crus-galli), and Bermuda grass (Cynodon dactylon)-was tested at various time intervals after vector inoculation. The minimum incubation periods in plant hosts before BGSS acquired X. fastidiosa were 4, 22, 29, and 25 days for grapevine, blackberry, mugwort, and watergrass, respectively. There were no transmissions by vectors or recoveries of X. fastidiosa by culturing from Bermuda grass in 133 attempts, including 80 attempts with the green sharpshooter, Draeculacephala minerva. The first acquisitions and subsequent transmissions by BGSS occurred after X. fastidiosa multiplied to a population of about 10(4) CFU/g of stem tissue. Higher populations of bacteria in plants resulted in higher rates of transmission. In grapevine, the rate of transmission increased over time (4.5% in the first 10 days to 55% after day 25) as the maximum number of viable CFU of X. fas-tidiosa recovered by culturing also increased (from 5 x 10(5) CFU/g during the first 10 days to 5 x 10(8) after day 25). | 18,945,018 |
Comparison of lettuce diseases and yield under subsurface drip and furrow irrigation. | ABSTRACT Subsurface drip and furrow irrigation were compared on lettuce (Lactuca sativa) cvs. Salinas and Misty Day for yield and incidence and severity of three important diseases of lettuce in the Salinas Valley, CA. Experiments were conducted between 1993 and 1995 during the spring and fall seasons. The diseases examined included lettuce drop (Sclerotinia minor), downy mildew (Bremia lactucae), and corky root (Rhizomonas suberifaciens). Replicated plots of subsurface drip and furrow irrigation were arranged in a randomized complete-block design. All plants were inoculated with S. minor at the initiation of the experiment during the 1993 spring season. Plots were not inoculated for downy mildew and corky root during any season nor were the plots reinoculated with S. minor. During each season, all plots were sprinkler irrigated until thinning, and subsequently, the irrigation treatments were begun. The furrow plots were irrigated once per week, and the drip plots received water twice per week. The distribution of soil moisture at two soil depths (0 to 5 and 6 to 15 cm) at 5, 10, and 15 cm distance on either side of the bed center in two diagonal directions was significantly lower in drip-irrigated compared with furrow-irrigated plots. Plots were evaluated for lettuce drop incidence and downy mildew incidence and severity at weekly intervals until harvest. Corky root severity and yield components were determined at maturity. Lettuce drop incidence and corky root severity were significantly lower and yields were higher in plots under subsurface drip irrigation compared with furrow irrigation, regardless of the cultivar, except during the 1994 fall season. Incidence and severity of downy mildew were not significantly different between the two irrigation methods throughout the study. The differential microclimates created by the two irrigation treatments did not affect downy mildew infection, presumably because the mesoclimate is usually favorable in the Salinas Valley. Subsurface drip irrigation is a viable, long-term strategy for soilborne disease management in lettuce in the Salinas Valley. | 18,945,057 |
Clavibacter michiganensis subsp. Sepedonicus Elicits a Hypersensitive Response in Tobacco and Secretes Hypersensitive Response-Inducing Protein(s). | ABSTRACT Strains of Clavibacter michiganensis subsp. sepedonicus, causal agent of bacterial ring rot of potato, showed marked differences in virulence on host plants. When infiltrated into tobacco leaves, virulent strains caused a rapid localized necrotic response (within 24 to 48 h) characteristic of the hypersensitive response (HR), whereas nonpathogenic strains did not. Concentrated cell-free culture supernatants (CCS) from virulent strains caused a necrotic reaction on tobacco, whereas CCS from nonpathogenic strains did not. The necrosis-inducing activity was heat stable and protease sensitive. Inhibitors of eukaryotic metabolism suppressed the necrotic reaction of tobacco to CCS. No necrotic response was observed when host plants were infiltrated with either cells or CCS from virulent strains. HR-inducing protein(s) from a virulent strain separated from the majority of other proteins on DEAE cellulose at 250 to 300 mM NaCl. Ammonium sulfate-precipitated proteins from a virulent strain produced a necrotic reaction at a total protein concentration of 18 mug/ml, whereas those from a nonpathogenic strain did not, even at a concentration of 180 mug/ml. We conclude that virulent strains of C. michiganensis subsp. sepedonicus elicit a typical HR in tobacco and secrete proteinaceous elicitor(s) of the nonhost HR. | 18,945,088 |
Molecular mapping of resistance to blight in an interspecific cross in the genus castanea. | ABSTRACT A three-generation American chestnut x Chinese chestnut pedigree was used to construct a genetic linkage map for chestnut and to investigate the control of resistance to Endothia parasitica (chestnut blight fungus). DNA genotypes for 241 polymorphic markers (eight isozymes, 17 restriction fragment length polymorphisms [RFLPs], and 216 random amplified polymorphic DNAs [RAPDs]) were assayed on an F(2) family consisting of 102 individuals. Of these markers, 196 were segregating as expected and, subsequently, used for primary linkage mapping. Two isozymes, 12 RFLPs, and 170 RAPDs were mapped to 12 linkage groups spanning a total genetic distance of 530.1 Kosambi centimorgans. F(2) plants were evaluated for a response to E. parasitica infection by directly inoculating them with two unique fungal isolates and measuring canker expansion over a period of 3.5 months. Results were compared with the marker genotype data, thereby identifying genomic regions significantly associated with a resistance response. Single-marker or nonsimultaneous analyses of variance identified seven genomic regions that appear to have an effect on host response. Multiple-marker or simultaneous models suggest that three of these regions have a significant effect on host response, together explaining as much as 42.2% of the total variation for canker size. At each of the three putative resistance loci, alleles derived from the Chinese chestnut grandparent were associated with smaller canker size, or higher levels of resistance. | 18,945,098 |
Comparison of Soil Receptivity to Thielaviopsis basicola, Aphanomyces euteiches, and Fusarium solani f. sp. pisi Causing Root Rot in Pea. | ABSTRACT Soil receptivity as a quantifiable characteristic ranging from conduciveness to suppressiveness to soilborne pea pathogens Thielaviopsis basicola and Aphanomyces euteiches was determined by analysis of differences in disease response curves obtained by artificial introduction of inoculum into natural field soil samples. Several parameters, including maximum root rot severity, the area under the health index curve, scores on the first axis of a principal component analysis (PCA) on dose responses, and Weibull model fitting were used to describe the disease responses. In all cases, the Weibull model gave satisfactory fits. PCA yielded a first axis that comprised 86% of the variance found when using Weibull predicted responses for T. basicola and 74% of the variance found for A. euteiches. This PCA axis essentially represented the average increase in disease severity due to the addition of increasing doses of inoculum to the soil. The Weibull scale parameter B, which represents the amount of inoculum necessary to increase root rot severity by 63% with respect to the level caused by pathogens naturally present in the soil, is another means of quantifying the receptivity of soils to these plant pathogens. Weibull parameter B, maximum root rot severity, the areaunder the health index curve, and the scores on the first PCA axis were strongly correlated for each of the pathogens tested individually. To compare the extent and behavior of soil receptivity responses to different pathogens, Weibull parameters B and C (slope at dose B) were chosen because of their universal definition, in contrast to PCA scores. Comparison of the average levels of Weibull parameters B and C indicated significant differences between the pathogens. Yet, no significant similarity in the ranking of the soils was found for the three pathogens, demonstrating that individual soils may interact with different pathogens in totally different ways. In general, soils were suppressive to T. basicola but conducive to A. euteiches, whereas their response to Fusarium solani f. sp. pisi ranged from conducive to suppressive. Therefore, risk assessment of soils prior to planting may require different strategies for each pathogen. Bioassays with soil samples taken before the last pea crop in 1987 and 1991 revealed a significant increase in the natural inoculum potential of soils that mainly was accounted for by A. euteiches and Pythium spp. These results strongly indicate that A. euteiches must be considered one of the most threatening pathogens to pea crops in the Netherlands. | 18,945,109 |
Characterization of the Suppressiveness of Hairy Vetch-Amended Soils to Thielaviopsis basicola. | ABSTRACT Factor(s) involved in soil suppressiveness to Thielaviopsis basicola when hairy vetch was used as a green manure were studied in a cotton production system. Soil suppressiveness was assessed in vitro at hairy vetch amendment levels of 0, 0.25, and 0.75% (wt/wt) by observing chlamydospores, using a nylon fabric technique. Chlamydospore germination in all soils was below 5%, and microscopic examination showed no germ tube lysis or visible propagule destruction. Viability (chlamydospore germination on T. basicola-carrot-etridiazol-nystatin [TB-CEN] medium) was reduced by 29% within 48 h after hairy vetch amendment. Viability also was reduced in atmospheres of amended soils, suggesting that the suppressiveness was due to a volatile factor. In a field study, chlamydospore viability in amended soils was reduced by 16%. T. basicola hyphal growth was more sensitive to ammonia than Rhizoctonia solani or Pythium ultimum, and chlamydospore mortality of T. basicola was 100% in petri dish atmospheres with 0.4 ppm of ammonia (50% lethal dose = 0.15 ppm). Soil atmospheric ammonia was 0.08 and 0.10 ppm for 0.25 and 0.75% amendment levels, respectively, both at 3 and 7 days after incorporation. In the field, 0.11 and 0.14 ppm of ammonia were detected in soil atmospheres 3 and 7 days after incorporation, respectively. The levels of ammonia detected were sufficient to account for the loss in T. basicola chlamydospore viability, indicating that ammonia is responsible for the suppressiveness observed. | 18,945,142 |
Epidemiological effect of gene deployment strategies on bacterial blight of rice. | ABSTRACT Experiments were conducted in farmers' fields at two locations of the irrigated lowlands of Laguna province in southern Luzon island, Philippines, during the wet seasons of 1993 and 1994. Nine rice populations were studied including pure stands, two-component mixtures, two-gene combinations of backcrossed lines containing varying combinations of the bacterial blight resistance genes Xa-4, xa-5, and Xa-10, and a non-isogenic cultivar containing Xa-4 and partial resistance to bacterial blight. The area under the disease progress curve (AUDPC) of both gene combinations studied was significantly less than the single most effective gene of each combination deployed singly. A mixture of a susceptible and a resistant line expressed an AUDPC significantly less than the mean of its component pure stands, but two other mixtures did not. The cultivar IR20, which contains both Xa-4 and partial resistance, reduced the AUDPC by about two-thirds as compared with IR-BB4, which contains Xa-4 and little or no partial resistance. | 18,945,155 |
Treatment with the Mycoparasite Pythium oligandrum Triggers Induction of Defense-Related Reactions in Tomato Roots When Challenged with Fusarium oxysporum f. sp. radicis-lycopersici. | ABSTRACT The influence exerted by the mycoparasite Pythium oligandrum in triggering plant defense reactions was investigated using an experimental system in which tomato plants were infected with the crown and root rot pathogen Fusarium oxysporum f. sp. radicis-lycopersici. To assess the antagonistic potential of P. oligandrum against F. oxysporum f. sp. radicis-lycopersici, the interaction between the two fungi was studied by scanning and transmission electron microscopy (SEM and TEM, respectively). SEM investigations of the interaction region between the fungi demonstrated that collapse and loss of turgor of F. oxysporum f. sp. radicis-lycopersici hyphae began soon after close contact was established with P. oligandrum. Ultrastructural observations confirmed that intimate contact between hyphae of P. oligandrum and cells of the pathogen resulted in a series of disturbances, including generalized disorganization of the host cytoplasm, retraction of the plasmalemma, and, finally, complete loss of the protoplasm. Cytochemical labeling of chitin with wheat germ agglutinin (WGA)/ovomucoid-gold complex showed that, except in the area of hyphal penetration, the chitin component of the host cell walls was structurally preserved at a time when the host cytoplasm had undergone complete disorganization. Interestingly, the same antagonistic process was observed in planta. The specific labeling patterns obtained with the exoglucanase-gold and WGA-ovomucoid-gold complexes confirmed that P. oligandrum successfully penetrated invading cells of the pathogen without causing substantial cell wall alterations, shown by the intense labeling of chitin. Cytological investigations of samples from P. oligandrum-inoculated tomato roots revealed that the fungus was able to colonize root tissues without inducing extensive cell damage. However, there was a novel finding concerning the structural alteration of the invading hyphae, evidenced by the frequent occurrence of empty fungal shells in root tissues. Pythium ingress in root tissues was associated with host metabolic changes, culminating in the elaboration of structural barriers at sites of potential fungal penetration. Striking differences in the extent of F. oxysporum f. sp. radicis-lycopersici colonization were observed between P. oligandrum-inoculated and control tomato plants. In control roots, the pathogen multiplied abundantly through much of the tissues, whereas in P. oligandrum-colonized roots pathogen growth was restricted to the outermost root tissues. This restricted pattern of pathogen colonization was accompanied by deposition of newly formed barriers beyond the infection sites. These host reactions appeared to be amplified compared to those seen in nonchallenged P. oligandrum-infected plants. Most hyphae of the pathogen that penetrated the epidermis exhibited considerable changes. Wall appositions contained large amounts of callose, in addition to be infiltrated with phenolic compounds. The labeling pattern obtained with gold-complexed laccase showed that phenolics were widely distributed in Fusarium-challenged P. oligandrum-inoculated tomato roots. Such compounds accumulated in the host cell walls and intercellular spaces. The wall-bound chitin component in Fusarium hyphae colonizing P. oligandrum-inoculated roots was preserved at a time when hyphae had undergone substantial degradation. These observations provide the first convincing evidence that P. oligandrum has the potential to induce plant defense reactions in addition to acting as a mycoparasite. | 18,945,162 |
Spatial Patterns of Microsclerotia of Verticillium dahliae in Soil and Verticillium Wilt of Cauliflower. | ABSTRACT The spatial patterns of microsclerotia of Verticillium dahliae in soil and wilt symptoms on cauliflower were determined at three sites in each of two fields in 1994 and 1995. Each site was an 8 x 8 grid divided into 64 contiguous quadrats (2 by 2 m each). Soil samples were collected to a depth of 15 cm with a probe (2.5 cm in diameter), and samples from four sites in each quadrat were bulked. Plants in each quadrat were cut transversely, and the number of plants with vascular discoloration and the number without discoloration were recorded. The soil was assayed for microsclerotia by the modified Anderson sampler technique. Lloyd's index of patchiness (LIP) was used as an indicator to evaluate the aggregation of microsclerotia in the field. Spatial autocorrelation and geostatistical analyses were also used to assess the autocorrelation of microsclerotia among quadrats. The LIP for microsclerotia was greater than 1, indicating aggregation of propagules; however, the degree of aggregation at most sites was not high. Significant autocorrelation within or across rows was detected in some spatial autocorrelograms of propagules, and anisotropic patterns were also detected in some oriented semivariograms from geostatistical analyses for microsclerotia, indicating the influence of bed preparation in the fields on pathogen distribution. The parameter estimates p and theta in the beta-binomial distribution and the index of dispersion (D) associated with the distribution were used to assess the aggregation of diseased plants at each site. A random pattern of wilt incidence was detected at 7 of 12 sites, and an aggregated pattern was detected at 5 of 12 sites. The degree of aggregation was not high. A regular pattern of wilt severity was detected at all sites. The high disease incidence (77 to 98%) observed at 11 of the 12 sites could be explained by high inoculum density. | 18,945,176 |
Ultrastructural and Cytochemical Investigations of the Antagonistic Effect of Verticillium lecanii on Cucumber Powdery Mildew. | ABSTRACT Chronological events of the intercellular interaction between Verticillium lecanii and cucumber powdery mildew, caused by Sphaerotheca fuliginea, were investigated at different times after inoculation by transmission electron microscopy. V. lecanii hyphae colonized host structures by tight binding, apparently mediated by a thin mucilaginous matrix. As early as 24 h after application of the antagonist, increased vacuolation and disorganization of the cytoplasm of the pathogen hyphae were easily detected. By 36 h after treatment, plasmalemma retraction and local cytoplasm aggregation were typical features of damage. Labeling chitin with the wheat germ agglutinin (WGA)/ovomucoid-gold complex showed that intracellular invasion of S. fuliginea by V. lecanii did not cause extensive host cell wall alterations, except in the area of hyphal penetration. By 48 h after inoculation, further cytoplasm disorganization was observed, as evidenced by the loss of cell turgor and contortion of the cell wall. Such deformation suggests that penetration of the antagonist results from mechanical pressure or localized enzymatic hydrolysis through the action of chitinases, as confirmed by the pattern of labeling obtained with the WGA/ovomucoid-gold complex. By 72 h after contact between the fungi, S. fuliginea cells were markedly collapsed, depleted of their protoplasm due to extensive multiplication of the antagonist, and totally encircled by the antagonist. Based on the current observations, the antagonism of S. fuliginea by V. lecanii appears to involve the following events: (i) attachment of the antagonist to the powdery mildew fungus; (ii) mechanical pressure and production of cell-wall degrading enzymes such as chitinases; (iii) penetration and active growth of the antagonist inside the pathogen hyphae; and (iv) digestion of host tissues and release of the antagonist from dead cells of S. fuliginea. The interaction between V. lecanii and S. fuliginea also affected the morphological and structural features of the haustorial bodies, as shown by increased vacuolation, distortion, and necrotization of the haustorial lobes. These observations provide the first experimental evidence that V. lecanii, primarily known as an entomopathogenic fungus, also has the potential to colonize mycelial structures of S. fuliginea. V. lecanii, therefore, may become a valuable alternative to current management of cucumber powdery mildew in greenhouses. | 18,945,181 |
Detection of a novel Francisella in Dermacentor reticulatus: a need for careful evaluation of PCR-based identification of Francisella tularensis in Eurasian ticks. | Francisella tularensis, the causative agent of tularemia, has been detected in ixodid ticks in some regions of North America, Europe, and Asia. In the present study, 245 Dermacentor reticulatus, 211 Ixodes ricinus, and 194 Haemaphysalis concinna adults from Hungary were tested for the presence of F. tularensis by polymerase chain reaction (PCR) assays based on 16S ribosomal RNA (16S rDNA) and T-cell epitope of a Francisella membrane protein (TUL4). No Francisella-specific amplification products were detected in I. ricinus and H. concinna ticks. Francisella DNA was identified using PCR assays based on 16S rDNA and TUL4 gene in D. reticulatus with similar prevalence (minimum 1.2%) as demonstrated in earlier European and Asian studies detecting F. tularensis in D. reticulatus. However, the 16S rDNA and TUL4 gene sequences of the Francisella-like agent occurring in D. reticulatus differed from the homologous sequences of Francisella spp. deposited in GenBank. Phylogenetic reconstructions showed that the new genotype detected in D. reticulatus was closely related to Francisella-like endosymbionts of North American Dermacentor ticks. Although further studies are needed on the relationship of this bacterium with ticks, the results highlight the need for careful evaluation of PCR-based identification in European and Asian laboratories that screen ixodid ticks for F. tularensis. | 18,945,184 |
Community participation and the emergence of late-life depressive symptoms: differences between women and men. | To understand the role of community participation in prevention of first lifetime depressive episode in older women and men. We used data from the Wisconsin Longitudinal Study to identify variables that predicted risk for the emergence of depressive symptoms and tested a hypothesis that community participation would protect women from depression more than it would protect men. The sample was drawn from Wisconsin high school graduates who were approximately 64-66 years of age in the 2003-2005 data collection period (n = 2546 with complete data meeting inclusion criteria.) The sample consisted of persons who had no evidence of current or prior lifetime depression in the 1993 data collection period. The emergence of high depressive symptoms was examined for women and men as a function of community participation and other covariates, including social support. The emergence of depressive symptoms for both sexes was predicted by poorer reported health status and higher levels of subthreshold depressive symptoms during the previous interview. For men, additional risk factors were pain and low income. For women, additional risks were widowhood, lower education, and lower community participation. Community participation, in the form of volunteering, religious attendance, and engagement in community organizations, is related to reduced risk of first-time depressive symptoms among older women. | 18,945,207 |
Reactivity of nitric oxide with the [4Fe-4S] cluster of dihydroxyacid dehydratase from Escherichia coli. | Although the NO (nitric oxide)-mediated modification of iron-sulfur proteins has been well-documented in bacteria and mammalian cells, specific reactivity of NO with iron-sulfur proteins still remains elusive. In the present study, we report the first kinetic characterization of the reaction between NO and iron-sulfur clusters in protein using the Escherichia coli IlvD (dihydroxyacid dehydratase) [4Fe-4S] cluster as an example. Combining a sensitive NO electrode with EPR (electron paramagnetic resonance) spectroscopy and an enzyme activity assay, we demonstrate that NO is rapidly consumed by the IlvD [4Fe-4S] cluster with the concomitant formation of the IlvD-bound DNIC (dinitrosyl-iron complex) and inactivation of the enzyme activity under anaerobic conditions. The rate constant for the initial reaction between NO and the IlvD [4Fe-4S] cluster is estimated to be (7.0+/-2.0)x10(6) M(-2) x s(-1) at 25 degrees C, which is approx. 2-3 times faster than that of the NO autoxidation by O2 in aqueous solution. Addition of GSH failed to prevent the NO-mediated modification of the IlvD [4Fe-4S] cluster regardless of the presence of O2 in the medium, further suggesting that NO is more reactive with the IlvD [4Fe-4S] cluster than with GSH or O2. Purified aconitase B [4Fe-4S] cluster from E. coli has an almost identical NO reactivity as the IlvD [4Fe-4S] cluster. However, the reaction between NO and the endonuclease III [4Fe-4S] cluster is relatively slow, apparently because the [4Fe-4S] cluster in endonuclease III is less accessible to solvent than those in IlvD and aconitase B. When E. coli cells containing recombinant IlvD, aconitase B or endonuclease III are exposed to NO using the Silastic tubing NO delivery system under aerobic and anaerobic conditions, the [4Fe-4S] clusters in IlvD and aconitase B, but not in endonuclease III, are efficiently modified forming the protein-bound DNICs, confirming that NO has a higher reactivity with the [4Fe-4S] clusters in IlvD and aconitase B than with O2 or GSH. The results suggest that the iron-sulfur clusters in proteins such as IlvD and aconitase B may constitute the primary targets of the NO cytotoxicity under both aerobic and anaerobic conditions. | 18,945,212 |
Oxidized and poorly glycosylated band 3 is selectively phosphorylated by Syk kinase to form large membrane clusters in normal and G6PD-deficient red blood cells. | Oxidative events involving band 3 (Anion Exchanger 1) have been associated with RBC (red blood cell) removal through binding of NAbs (naturally occurring antibodies); however, the underlying mechanism has been only partially characterized. In addition to inducing direct membrane protein oxidative modification, oxidative treatment specifically triggers the phosphorylation of band 3 tyrosine residues. The present study reports that diamide, a thiol group oxidant, induces disulfide cross-linking of poorly glycosylated band 3 and that the oligomerized band 3 fraction is selectively tyrosine phosphorylated both in G6PD (glucose-6-phosphate dehydrogenase)-deficient and control RBCs. This phenomenon is irreversible in G6PD-deficient RBCs, whereas it is temporarily limited in control RBCs. Diamide treatment caused p72 Syk phosphorylation and translocation to the membrane. Diamide also induced p72 Syk co-immunoprecipitation with aggregated band 3. Moreover, following size-exclusion separation of Triton X-100-extracted membrane proteins, Syk was found only in the high-molecular-mass fraction containing oligomerized/phosphorylated band 3. Src family inhibitors efficiently abrogated band 3 tyrosine phosphorylation, band 3 clustering and NAbs binding to the RBC surface, suggesting a causal relationship between these events. Experiments performed with the non-permeant cross-linker BS(3) (bis-sulfosuccinimidyl-suberate) showed that band 3 tyrosine phosphorylation enhances its capability to form large aggregates. The results of the present study suggest that selective tyrosine phosphorylation of oxidized band 3 by Syk may play a role in the recruitment of oxidized band 3 in large membrane aggregates that show a high affinity to NAbs, leading to RBC removal from the circulation. | 18,945,214 |
Different bispectral index values from both sides of the forehead in unilateral carotid artery stenosis. | Bispectral index (BIS) values derived from the left and right forehead are usually the same. We report on two patients with unilateral carotid artery stenosis in whom we observed differences between the BIS values obtained from sensors placed on each side of the forehead. During surgery, the BIS values of the diseased side decreased more than those of the opposite side when the mean arterial pressure decreased below 70 mmHg. BIS monitors should be used with caution in patients with unilateral carotid artery and cerebrovascular disease. | 18,945,245 |
Fluid therapy and the use of albumin in the treatment of severe traumatic brain injury. | Evidence-based guidelines for severe traumatic brain injury (TBI) do not include strategies for fluid administration. The protocol used in this study includes albumin administration to maintain normal colloid osmotic pressure and advocates a neutral to slightly negative fluid balance. The aim of this study was to analyze the occurrence of organ failure and the mortality in patients with severe TBI treated by a protocol that includes defined strategies for fluid therapy. Ninety-three patients with severe TBI and Glasgow Coma Score <or=8 were included during 1998-2001. Medical records of the first 10 days were retrieved. Organ dysfunction was evaluated with the Sequential Organ Failure Assessment (SOFA) score. Mortality was assessed after 10 and 28 days, 6 and 18 months. The total fluid balance was positive on days 1-3, and negative on days 4-10. The crystalloid balance was negative from day 2. The mean serum albumin was 38+/-6 g/l. Colloids constituted 40-60% of the total fluids given per day. Furosemide was administered to 94% of all patients. Severe organ failure defined as SOFA >or=3 was evident only for respiratory failure, which was observed in 29%. None developed renal failure. After 28 days, mortality was 11% and, after 18 months, it was 14%. A protocol including albumin administration in combination with a neutral to a slightly negative fluid balance was associated with low mortality in patients with severe TBI in spite of a relatively high frequency (29%) of respiratory failure, assessed with the SOFA score. | 18,945,246 |
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