Abstract:Fusarium wilt of watermelon caused by Fusarium oxysporum f. sp. niveum caused seedling losses in nurseries, as well as severe losses in many crops in Nghe An province, Vietnam, in 2008. Isolates of the fungus were shown to be pathogenic. All 20 cultivars grown in the province in 2008 were susceptible. This is the first formal report of this disease in Vietnam.
Abstract: Internal fragments of a- and b-tubulin genes were generated using reverse transcription polymerase chain reaction (RT-PCR), and the termini were isolated using 5- and 3-rapid amplification of cDNA ends. Phytophthora capsici a- and b-tubulin specific primers were then used to generate full-length cDNA by RT-PCR. The recombinant a- and b-tubulin genes were expressed in Escherichia coli BL21 (DE3), purified under denaturing conditions, and average yields were 3.38–4.5 mg of a-tubulin and 2.89–4.0 mg of b-tubulin, each from 1-l culture. Optimum conditions were obtained for formation of microtubule-like structures. A value of 0.12 mg/ml was obtained as the critical concentration of polymerization of P. capsici tubulin. Benomyl inhibited polymerization with half-maximal inhibition (IC50) =468±20 uM. Approximately 18.66±0.13 cysteine residues per tubulin dimer were accessible to 5,5'-dithiobis-(2-nitrobenzoic acid), a quantification reagent of sulfhydryl and 12.43±0.12 residues were accessible in the presence of 200 uM benomyl. The order of preference for accessibility to cysteines was benomyl > colchicine > GTP > taxol, and cysteine accessibility changes conformed that binding sites of these ligands in tubulin were folding correctly. Fluorescence resonance energy transfer technique was used for high throughput screening of chemical library in search of antimitotic agent. There was significant difference in relative fluorescence by 210-O-2 and 210-O-14 as compared to colchicine.
Abstract:Phytophthora foot rot of black pepper(Pipper nigrum L.,) caused by P.capsici is severe during the wet monsoon season. The pattern of spread was studied and analyzed using STCLASS computer software. The pattern of spread was non-random and the initially infected vines served as source and focus of secondary spread.
Abstract: Ink disease caused by Phytophthora cambivora is a major disease of sweet chestnut (Castanea sativa). In two C. sativa stands in central Italy, one (Montesanti) that is infected with P. cambivora and the trees showing symptoms of ink disease and another healthy stand (Puzzella), the ectomycorrhizal (ECM) community structure was investigated. On the roots of the surviving trees of the diseased stand, 29 different ECM species were determined compared to 23 in the healthy stand. Eleven ECM species were common to both stands; however, a number of species were unique to one of the stands. Cenococcum geophilum was dominant at both sites, but the percentage colonisation was much higher at Montesanti (40.8%) compared to Puzzella (27.2%). There was a switch in species from Russula vesca, Russula lepida and Russula azurea at Puzzella to Russula nigricans, R. lepida and Russula delica at Montesanti. Both R. vesca and R. azurea were found only at the Puzzella site. At the diseased site, the ECMs formed had a smaller root tip diameter, and the ECM at the healthy site had more abundant extramatrical hyphae.
Abstract: A severe decline of alder associated with an undescribed Phytophthora species was identified for the first time in England in 1993. No generalized decline of alder was reported in France before 1990. The first diebacks and mortalities of common alder were observed at the beginning of the 1990s, but the so-called alder Phytophthora was not isolated in France until 1996. First, a synthesis about alder declines that were known in France before 1995 is presented. Then, a survey was established in north-eastern France; 108 sites were visited and the alder Phytophthora was isolated from 57 of them. All the main rivers were found to be affected and damage levels are significant along some of them. The frequency of the alder Phytophthora and other fungi isolated from declining alders is discussed. Finally, information on other alder declines in France is presented region by region, and a map summarizes the known distribution of the disease. The alder Phytophthora is quite common and widespread in France, with western and north-eastern France being especially affected; however, the number of diseased or dead trees varies greatly from one site to another. All records are from Alnus glutinosa; other Alnus species were seldom seen in the surveys.
Abstract: Oospore viability, oospore germinability, and phenotypic variation among zoospore and hyphal tip derivatives of the standard and natural variant types of the hybrid alder Phytophthoras were investigated. Oospore viability in the standard hybrid, estimated by the tetrazolium bromide method, was low (≈ 31–36%). No germination was observed in more than 4000 oospores, although germination did occur in the Phytophthora cactorum, Phytophthora citricola and Phytophthora cambivora controls. This is consistent with the known meiotic irregularities in this hybrid. Mean oospore viabilities in the natural variants were significantly different (p < 0.001), ranging from approximately 24% in the UK variant to approximately 75% in the Dutch variant. Again, no oospore germination was observed. Zoospore and hyphal tip derivatives of the standard hybrid and of the Swedish and Dutch variants resembled the parent isolate in phenotype. The derivatives of the German and UK variants, however, often differed from the parent type. Those of the UK variant were extremely and continuously variable in colony patterns, growth rates, temperature–growth relationships and fertility levels. Although these results do not support the view that the natural variants arise as genetic breakdown products of the standard hybrid, this possibility cannot yet be excluded. The apparent inviability of the oospores suggests that the mycelium and zoospores are mainly responsible for the survival and spread of the alder Phytophthoras in the field.
Abstract:Ralstonia solanacearum, the causing agent of crop bacterial wilt, is a devastating bacterium with an unusually wide host range. To effectively integrate durable resistance to R. solanacearum into crops, comprehensive and global information on pathogenesis mechanisms employed by this pathogen is important. In this study, we report the development of an efficient Arabidopsis-based bioassay system useful for screening of large number of bacterium mutants with altered virulence. The effectiveness and uniqueness of this system was further demonstrated by the isolation of R. solanacearum mutants exhibiting differential pathogenesis and in vitro characteristics, and by the identification of mutants with the transposon inserted in known and novel loci. This highly efficient bioassay system, together with the accompanying in planta and in vitro bioassay systems described here, would facilitate comprehensive study on R. solanacearum pathogenesis mechanisms.
Abstract: Two different recombinant plasmids both containing the cyanophycin synthetase gene (cphA) of Synechocystis sp. strain PCC6308 but differing concerning the resistance marker gene were tested for their suitability to produce high amounts of cyanophycin in recombinant strains of Ralstonia eutropha. Various cultivation experiments at the 30-L scale revealed very low cyanophycin contents of the cells ranging from 4.6% to 6.2% (w/w) of cellular dry weight (CDW) only, most probably because most cells had lost the corresponding plasmid during cultivation. To establish a cost effective and high efficient system for production of cyanophycin at larger scales using recombinant strains of R. eutropha, we applied two strategies: First, we integrated cphA into the dispensable chromosomal l-lactate dehydrogenase gene (ldh) of R. eutropha. Depending on the cultivation conditions used, relatively low cyanophycin contents between 2.2% and 7.7% (w/w) of CDW were reproducibly detected, which might be due to weak expression or low gene dosage in the single cphA copy strain of R. eutropha. In a second strategy we constructed a KDPG-aldolase gene (eda)-dependent addiction system, which combined features of a multi-copy plasmid with stabilized expression of cphA. Flasks experiments revealed that the cells accumulated extraordinarily high amounts of cyanophycin between 26.9% and 40.0% (w/w) of CDW even under cultivation conditions lacking cyanophycin precursor substrates or plasmid stabilizing antibiotics. Cyanophycin contents of up to 40.0% (w/w) of CDW were also obtained at a 30-L scale or a 500-L pilot-plant scale under such non-selective conditions. This demonstrates impressively that the stabilizing effect of the constructed eda-dependent addiction system can be used for production of enhanced amounts of cyanophycin at a larger scale in recombinant strains of R. eutropha.
Abstract: Healthy and Ralstonia solanacearum-inoculated tomato genotypes susceptible or resistant to bacterial wilt including recombinant inbred lines (RILs) deriving from a cross between the resistant genotype Hawaii7996 and the susceptible Wva700 were compared for symptom and bacterial population development, and for the composition and structure of pectic polysaccharides and arabinogalactan proteins (AGPs) of xylem cell walls by immunological staining of tissue prints. Constitutive differences were observed between resistant and susceptible RILs, with a higher degree of methyl-esterification of homogalacturonan (HG) detected by antibody JIM7 in the resistant plants. After inoculation, decreased methyl-esterification of HG indicated by stronger labeling with antibody JIM5 was observed in all susceptible genotypes and in five of eleven resistant genotypes, with a clear increase in the non-blockwise de-esterification pattern of HG (LM7) only in the susceptible lines, indicating the mode of action of the pectinmethylesterase of R. solanacearum. In the susceptible lines infection generally leads to increased branching of rhamnogalacturonan I indicated by the detection of arabinan (LM6) and galactan (LM5) side chains, and of arabinogalactan protein (LM2), while only few of the resistant genotypes reacted with changes in these epitopes. All the resistant, symptomless genotypes contained relatively high pathogen populations in stems. A clear relation between cell wall composition and degree of latent infection of resistant genotypes was not found.
Abstract: A green fluorescent protein (GFP)-expressing plasmid was constructed from a filamentous bacteriophage φRSS1 that infects the phytopathogen Ralstonia solanacearum. This plasmid designated as pRSS12 (4.7 kbp in size) consists of an approximately 2248 bp region of the φRSS1 RF DNA, including ORF1-ORF3 and the intergenic region (IG), and a Km cassette in addition to the GFP gene. It was easily introduced by electroporation and stably maintained even without selective pressure in strains of R. solanacearum of different races and biovars. Strong green fluorescence emitted from pRSS12-transformed bacterial cells was easily monitored in tomato tissues (stem, petiole, and root) after infection as well as from soil samples. These results suggest that pRSS12 can serve as an easy-to-use GFP-tagging tool for any given strain of R. solanacearum in cytological as well as field studies.
Abstract