Calcium fertilizers induce soil suppressiveness to <Emphasis Type="BoldItalic">Phytophthora cinnamomi</Emphasis> root rot of <Emphasis Type="BoldItalic">Quercus ilex</Emphasis>

Based on the observation that the root disease caused by P. cinnamomi on Q. ilex has a low incidence and severity in soils with medium-high Ca²⁺ content, we studied the ability of Ca²⁺ fertilizers to induce soil suppressiveness to the pathogen. Studies on cultures of P. cinnamomi exposed to different Ca²⁺ fertilizers in vitro showed significant inhibition of sporangial, chlamydospore and zoospore production at millimolar concentrations while mycelial growth was mainly unaffected. Experiments performed with artificially infested soil showed that some Ca²⁺ fertilizers induce a significant decrease on chlamydospore viability. Additionally, greenhouse experiments using artificially infested soils showed a significant reduction of foliar and root symptom severities in Holm oak seedlings growing in soils amended with Ca²⁺ fertilizers. We suggest that limestone amendments in oak rangelands could enhance the suppressiveness of soils to P. cinnamomi, and it is likely that the inhibition of sporangial production was the main mechanism involved.     

A quantitative PCR assay for accurate <Emphasis Type="Italic">in planta</Emphasis> quantification of the necrotrophic pathogen <Emphasis Type="Italic">Phytophthora cinnamomi</Emphasis>

A reliable method for measuring disease progression is important when evaluating susceptibility in host—pathogen interactions. We describe a sensitive quantitative polymerase chain reaction (QPCR) assay that enables quantitative measurement of in planta DNA of the necrotrophic pathogen, Phytophthora cinnamomi, that avoids problems caused by variation in DNA extraction efficiency and degradation of host DNA during host tissue necrosis. Normalization of pathogen DNA to sample fresh weight or host DNA in samples with varying degrees of necrosis led to overestimation of pathogen biomass. Purified plasmid DNA, containing the pScFvB1 mouse gene, was added during DNA extraction and pathogen biomass was normalized based on plasmid DNA rather than host DNA or sample fresh weight. This method is robust and improves the accuracy of pathogen measurement in both resistant (non-host A. thaliana–P. cinnamomi) and susceptible (host Lupinus angustifolius–P. cinnamomi) interactions to allow accurate measurement of pathogen biomass even in the presence of substantial host cell necrosis.     

<Emphasis Type="Italic">Lupinus luteus</Emphasis>, a new host of <Emphasis Type="Italic">Phytophthora cinnamomi</Emphasis> in Spanish oak-rangeland ecosystems

Phytophthora cinnamomi is an aggressive pathogen on Lupinus luteus (yellow lupin), causing root rot, wilting and death of this crop, common in oak-rangeland ecosystems ('dehesas') in south-western Spain. The oomycete, the main cause of Quercus decline in the region, was isolated from roots of wilted lupins in the field. Artificial inoculations on four cultivars of L. luteus reproduced the symptoms of the disease, both in pre- and post-emergence stages, recovering the pathogen from necrotic roots. These results suggest the potential of yellow lupin as inoculum reservoir for the infection of Quercus roots. This is the first report of P. cinnamomi as root pathogen of L. luteus.     

Population Dynamics of <Emphasis Type="Italic">Monosporascus</Emphasis><Emphasis Type="Italic">cannonballus</Emphasis> Ascospores in Marsh Soils in Eastern Spain

The population dynamics of Monosporascus cannonballus ascospores in the soils of four muskmelon fields located in a marsh area in Castellón province (eastern Spain) was studied for a 3 year period. Two of these fields were cropped to muskmelon with fallow periods between muskmelon cropping, and the others were in fallow and had extensive flooding periods. Muskmelon cultivation resulted in a progressive increase of the number of ascospores in soil, reaching a maximum 7 months after muskmelon planting (2–4 months after plant death), and a subsequent decline during fallow periods between muskmelon crops. During muskmelon cropping, in-bed and between-bed ascospore numbers were compared and, in general, there were no statistical differences between them. In the fields which were in fallow and flooded, the dynamics found was a progressive decline of the population of ascospores. Soilborne inoculum was viable and capable of infecting muskmelon at the end of the 3 year period in all fields, demonstrating that ascospores of M.␣cannonballus are able to survive for this period of time in the absence of muskmelon cultivation and also that this fungus seems to be well adapted to survive in soils which maintain a high water table during the crop or under flooding conditions.     

Pathogenicity of <Emphasis Type="Italic">Beauveria bassiana</Emphasis> and <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> against <Emphasis Type="Italic">Galleria mellonella</Emphasis>

The greater wax moth Galleria mellonella L. (Lepidoptera: Pyralidae) is occasionally found in beehives and is a major pest of stored wax. Entomopathogenic fungi have recently received attention as possible biocontrol elements for certain insect pests. In this study, 90 isolates of Beauveria bassiana and 15 isolates of Metarhizium anisopliae were screened for proteases and lipases production. The results showed significant variations in the enzymatic action between the isolates. In the bioassay, the selected isolates evinced high virulence against the 4th instar of the G. mellonella larvae. The isolates BbaAUMC3076, BbaAUMC3263 and ManAUMC3085 realized 100% mortality at concentrations of 5.5 × 10⁶ conidia ml⁻¹, 5.86 × 10⁵ conidia ml⁻¹, and 4.8 × 10⁶ conidia ml⁻¹, respectively. Strong enzymatic activities in vitro did not necessarily indicate high virulence against the tested insect pest. The cuticle of the infected larvae became dark and black-spotted, indicating direct attack of fungus on the defense system of the insects. The LC₅₀ values were 1.43 × 10³, 1.04 × 10⁵ and 5.06 × 10⁴ for Bba3263AUMC, Bba3076AUMC and Man3085AUMC, respectively, and their slopes were determined by computerized probit analysis program as 0.738 ± 0.008, 0.635 ± 0.007 and 1.120 ± 0.024, respectively.     

Biological soil disinfestation using ethanol: effect on <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">lycopersici</Emphasis> and soil microorganisms

Better soil disinfestation methods, such as biological soil disinfestation (BSD), that are environmentally safe are increasingly been developed and used because of rising concerns related to environmental risks. We evaluated the efficacy of soil disinfestation using ethanol to control the fungus Fusarium oxysporum f. sp. lycopersici, which causes fusarium wilt of tomato. Survival of bud cells and chlamydospores declined markedly in soil saturated with diluted ethanol solution in the laboratory. In field trials, artificially added nonpathogenic Fusarium oxysporum and indigenous F. oxysporum were both strongly suppressed in soil saturated with 1% ethanol solution; a wheat bran treatment was not as effective. The artificially added fungus was not detected in three of four sites treated with ethanol but was detected in three of four sites amended with wheat bran. Using ethanol in pre-autoclaved soil was not suppressive; thus native microorganisms are essential for the suppression. This ethanol-mediated biological soil disinfestation (Et-BSD) temporarily increased the number of anaerobic bacteria, but the number of fungi and aerobic bacteria was stable. Polymerase chain reaction–denaturing gradient gel electrophoresis (PCR–DGGE) analysis revealed slight but apparent differences in bacterial community structures in the soil treated with Et-BSD compared with the structure in soils after other treatments such as water irrigation and in the control soil, which received neither organic amendment nor irrigation after 15 days. Et-BSD is a potentially effective and easy soil disinfestation method, and its impact on native, beneficial microorganisms is moderate.     

Biological traits and prey consumption of <Emphasis Type="Italic">Anthocoris minki</Emphasis> fed on <Emphasis Type="Italic">Agonoscena pistaciae</Emphasis> and <Emphasis Type="Italic">Brachycaudus (Thuleaphis) amygdalinus</Emphasis>

The predatory insect Anthocoris minki Dohrn (Heteroptera: Anthocoridae) is an indigenous Anthocoris species for the biological control of pests in pistachio orchards. The pistachio psylla Agonoscena pistaciae Burckhardt and Lauterer (Homoptera: Psyllidae) is an important insect pest in pistachio trees in Turkey. Similarly, Brachycaudus (Thuleaphis) amygdalinus (Schouteden) (Hemiptera: Aphididae) is a pest of almond trees that is considered as alternative prey for A. minki when pistachio psylla are not available in early spring on pistachio trees. The development time, survival percentage of immature stages, longevity, fecundity, prey consumption, and life table parameters of A. minki fed on A. pistaciae and B. amygdalinus nymphs were determined at 25 ± 1°C, 70 ± 5% r.h., and a 16 h:8 h (L:D) photoperiod under laboratory conditions. The nymphal survival rate was significantly higher when nymphs were fed on A. pistaciae (an average of 96.7%) than on B. amygdalinus (an average of 71.4%). The development time of A. minki was significantly shorter when nymphs were fed on B. amygdalinus (10.3 days) as opposed to A. pistaciae (11.0 days). No significant differences among prey species were found for longevity and fecundity. The total female longevity and fecundity of A. minki was 38.0 days and 247.2 eggs, respectively, when nymphs were fed on A. pistaciae; and 35.4 days and 233.0 eggs, respectively, when nymphs were fed on B. amygdalinus. On average, 104.4 A. pistaciae and 77.7 B. amygdalinus nymphs were consumed during the nymphal development time for A. minki. Adults of A. minki consumed significantly more psyllids than aphids throughout their life span. The greater difference did not significantly inpact the longevity and fecundity of A. minki. Females of A. minki consumed an average of 631.0 A. pistaciae and an average of 273.3 B. amygdalinus nymphs, while female predators consumed significantly more prey than males. The intrinsic rate of increase (r _m ) of A. minki fed on A. pistaciae (0.174) was significantly greater than those fed on B. amygdalinus (0.148). The successful development and reproduction of both A. pistaciae and B. amygdalinus indicates that they are suitable prey for A. minki.     

Occurrence and distribution of <Emphasis Type="Italic">Phytophthora</Emphasis> species in European chestnut stands,and their association with Ink Disease and crown decline

The Phytophthora complex associated with Castanea sativa Mill. was investigated in five European countries in 35 regions and with respect to various domestication levels. Annual precipitation and length of drought season were the main parameters that regulated the presence of Phytophthora species in the chestnut stands. Seven species of Phytophthora were detected; three of these, P. megasperma, P. cryptogea and P. syringae had not been previously reported on sweet chestnut. P. cinnamomi. P. cambivora and P. citricola were most frequently isolated. P. cinnamomi and P. cambivora were the species significantly associated with declining trees with symptoms of Ink Disease. P. cinnamomirequired distinct ecological conditions compared to the other species. P. cinnamomi was never detected in sites characterized by minimum temperatures below 1.4 °C, maximum temperature above 28 °C, or soil pH below 5.4. The results obtained provide useful information for modeling the probability of Ink Disease, crown decline and associated Phytophthora species in chestnut groves in global climatic change scenarios.     

Susceptibility of eggs of <Emphasis Type="Italic">Tribolium confusum</Emphasis>, <Emphasis Type="Italic">Ephestia kuehniella</Emphasis> and <Emphasis Type="Italic">Plodia interpunctella</Emphasis> to four essential oil vapors

Susceptibility of eggs of Tribolium confusum du Val. (Coleoptera: Tenebrionidae), Ephestia kuehniella (Zell.) (Lepidoptera: Phycitidae) and Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae) to vapors of essential oil from garlic (Allium sativum L.), birch (Betula lenta L.), cinnamon (Cinnamonum zeylanicum (Blume)) and aniseed (Pimpinella anisum L.) was studied. Preliminary bioassay tests indicated that vapors of the essential oils had a significant effect on the eggs of tested insect species when exposed to a concentration of 20 μl l ⁻¹ air for 24 h. Generally, garlic and birch essential oils were more toxic to the eggs of tested insect species than cinnamon and aniseed essential oils (except for eggs of T. confusum). There was also a significant difference between susceptibility of eggs of T. confusum, E. kuehniella and P. interpunctella to tested essential oils. Toxicity data indicated that eggs of T. confusum were more susceptible to tested essential oils, with LC₉₀ values ranging from 3.11 to 33.49 μl l ⁻¹ air, than those of E. kuehniella and P. interpunctella; eggs of P. interpunctella were the most tolerant to the essential oils, with LC₉₀ values ranging from 22.02 to 72.42 μl l ⁻¹ air. Concentration × time (Ct) products of 0.29, 0.22, 0.13 and 1.37 mg h l ⁻¹ for garlic, birch, cinnamon and aniseed essential oil, respectively, were required to obtain 90% kill of T. confusum eggs. Although cinnamon essential oil had a much closer Ct product value to methyl bromide, garlic and birch essential oils were found to be the most promising ones since they had also high fumigant toxicity on eggs of both E. kuehniella and P. interpunctella.     

<Emphasis Type="Italic">Corythucha ciliata</Emphasis>, a new <Emphasis Type="Italic">Platanus</Emphasis> pest in Turkey

An invasive Tingidae, the platanus lace bug Corythucha ciliata (Say, 1832) (Hemiptera: Tingidae), which specializes on Platanus spp., was found for the first time in Turkey in 2007; it was recorded from a 120 km² area in the northwestern part of the country. Infestations occurred in an area between Taşkesti and Abant in Bolu Province, which is located near major cities and two main motorways. The pest species is newly spreading in Turkey, causing noticeable damage to Platanus orientalis trees.     

Pathogenicity of morphologically different isolates of <Emphasis Type="Italic">Sclerotinia sclerotiorum</Emphasis> with <Emphasis Type="Italic">Brassica napus</Emphasis> and <Emphasis Type="Italic">B. juncea</Emphasis> genotypes

Sclerotinia stem rot caused by Sclerotinia sclerotiorum is a serious threat to oilseed production in Australia. Eight isolates of S. sclerotiorum were collected from Mount Barker and Walkway regions of Western Australia in 2004. Comparisons of colony characteristics on potato dextrose agar (PDA) as well as pathogenicity studies of these isolates were conducted on selected genotypes of Brassica napus and B. juncea. Three darkly-pigmented isolates (WW-1, WW-2 and WW-4) were identified and this is the first report of the occurrence of such isolates in Australia. There was, however, no correlation between pigmentation or colony diameter on PDA with the pathogenicity of different isolates of this pathogen as measured by diameter of cotyledon lesion on the host genotypes. Significant differences were observed between different isolates (P ≤ 0.001) in two separate experiments in relation to pathogenicity. Differences were also observed between the different Brassica genotypes (P ≤ 0.001) in their responses to different isolates of S. sclerotiorum and there was also a significant host × pathogen interaction (P ≤ 0.001) in both experiments. Responses between the two experiments were significantly correlated in relation to diameter of cotyledon lesions caused by selected isolates (r = 0.79; P < 0.001, n = 48). Responses of some genotypes (e.g., cv. Charlton) were relatively consistent irrespective of the isolates of the pathogen tested, whereas highly variable responses were observed in some other genotypes (e.g., Zhongyou-ang No. 4, Purler) against the same isolates. Results indicate that, ideally, more than one S. sclerotiorum isolate should be included in any screening programme to identify host resistance. Unique genotypes which show relatively consistent resistant reactions (e.g., cv. Charlton) across different isolates are the best for commercial exploitation of this resistance in oilseed Brassica breeding programmes.     

White rust of <Emphasis Type="Italic">Ipomoea</Emphasis> caused by <Emphasis Type="Italic">Albugo ipomoeae-panduratae</Emphasis> and <Emphasis Type="Italic">A. ipomoeae-hardwickii</Emphasis> and their host specificity

In some areas of Japan, yellow spots with white pustules on leaves, stems, petioles, peduncles and calyces were found on Ipomoea nil, I. triloba, I. lacunosa and I. hederacea var. integriuscula. We demonstrated that the diseases on I. nil, I. triloba and I. lacunosa were caused by host-specific strains of Albugo ipomoeae-panduratae and defined three forma speciales of the fungus, respectively, for the three Ipomoea species: “f. sp. nile”, “f. sp. trilobae” and “f. sp. lacunosae”. Because the diseases were new to Japan, we coined the Japanese name “shirosabi-byo”, which means white rust. We also showed that the disease on I. hederacea var. integriuscula was caused by A. ipomoeae-hardwickii. We named this new disease “white rust (shirosabi-byo in Japanese)”.     

Pathogenicity and reproductive fitness of <Emphasis Type="Italic">Pratylenchus coffeae</Emphasis> and <Emphasis Type="Italic">Radopholus arabocoffeae</Emphasis> on Arabica coffee seedlings (<Emphasis Type="Italic">Coffea arabica</Emphasis> cv. Catimor) in Vietnam

The pathogenicity and reproductive fitness of Pratylenchus coffeae and Radopholus arabocoffeae from Vietnam on coffee (Coffea arabica) seedlings cv. Catimor were evaluated in greenhouse experiments. The effect of initial population densities (Pi = 0, 1, 2, 4, 8, 16, 32, 64, 128, and 256 nematodes per cm³ soil) was studied for both species at different days after inoculation (dai). The data were adjusted to the Seinhorst damage model Y = m + (1-m).z^Pi-T. Tolerance limit (T) for P. coffeae was zero for the height and the diameter of the coffee plants. For the diameter, the T-value for R. arabocoffeae was 25.6 for 30 and 60 dai and 12.8 for 90 and 120 dai. After 4 months T was zero. The low tolerance limits indicate that Arabica coffee is highly intolerant to both nematode species. At the end of the experiment (180 dai), all plants were infected and most were dead when inoculated with R. arabocoffeae at initial densities of 32, 64, 128 and 256 nematodes/cm³ soil. For P. coffeae plant death was already observed at the lowest inoculation densities. Growth of coffee was reduced at all inoculation levels for both species. Pratylenchus coffeae and R. arabocoffeae caused intense darkening of the roots, leaf chlorosis and a strong reduction of root and shoot growth. It was observed that P. coffeae mainly destroyed lateral roots rather than tap roots, whereas R. arabocoffeae reduced tap root length rather than the lateral roots. At the lowest inoculum densities, the reproduction factor of P. coffeae was 2.38 and 2.01 for R. arabocoffeae, indicating that arabica coffee is a host for both species. Plant growth as expressed by shoot height and shoot and root weight measured 60 dai was negatively correlated with nematode (both species) density as expressed by the geometric mean of nematode numbers at 30 and 60 dai.     

<Emphasis Type="Italic">Eremothecium coryli</Emphasis> and <Emphasis Type="Italic">E.</Emphasis><Emphasis Type="Italic">ashbyi</Emphasis> cause yeast spot of azuki bean

Yeast-like fungi were isolated from lesions on azuki bean (cv. Shin-Kyotodainagon) seeds that had been sucked by bean bugs in Kyoto Prefecture, Japan. On the basis of morphological and physiological characteristics and sequence data of the internal transcribed spacer (ITS) regions including the 5.8S rDNA, these yeasts were identified as Eremothecium coryli and E. ashbyi. Pathogenicity of those yeasts was confirmed by a reinoculation test. To our knowledge, this is the first report of the occurrence of yeast spot in azuki bean in Japan. The nucleotide sequence data reported are available in the GeneBank/EMBL/DDBJ database as accessions AB478291–AB478309 for E. coryli AZC1–19 and AB478310–AB478317 for E. ashbyi AZA1–8.     

First report of blueberry red ringspot disease caused by <Emphasis Type="Italic">Blueberry</Emphasis><Emphasis Type="Italic">red</Emphasis><Emphasis Type="Italic">ringspot</Emphasis><Emphasis Type="Italic">virus</Emphasis> in Japan

Virus-like symptoms—red ringspots on stems and leaves, circular blotches or pale spots on fruit—were found on commercial highbush blueberry (Vaccinium corymbosum) cultivars Blueray, Weymouth, Duke and Sierra in Japan. In PCR testing, single DNA fragments were amplified from total nucleic acid samples of the diseased blueberry bushes using primers specific to Blueberry red ringspot virus (BRRV). Sequencing analysis of the amplified products revealed 95.7–97.7% nucleotide sequence identity with the BRRV genome. This paper is the first report of blueberry red ringspot disease caused by BRRV in Japan. The nucleotide sequence data reported in this paper are available in the GenBank/EMBL/DDBJ database as accessions AB469884 to AB469893 for BRRV isolates from Japan.     

Characterization of <Emphasis Type="Italic">Pectobacterium carotovorum</Emphasis> subsp. <Emphasis Type="Italic">carotovorum</Emphasis> causing soft-rot disease on <Emphasis Type="Italic">Pinellia ternata</Emphasis> in China

Pinellia ternata is a traditional Chinese herb which has been used in China for over 1,000 years. A soft-rot disease characterized by water-soaked lesions and soft-rot symptoms with a stinking odour was commonly observed in cultivated fields of this plant, and Pectobacterium-like bacteria were consistently isolated from the infected tissues. Two typical strains (SXR1 and ZJR1), isolated from Shanxi and Zhejiang, respectively, were identified. Pathogenicity tests revealed that these strains were virulent to P. ternata and induced the same symptoms as observed in the field. Characterization involving fatty acid profile, metabolic and physiological properties, 16S rDNA sequence and PCR-RFLP identified both isolates as P. carotovorum subsp. carotovorum (Pcc). The 16S rDNA of both isolates shared 97–99% sequence similarity with that of Pcc strains. The phylogenetic trees showed that both isolates were clustered in the group of Pcc and P. carotovorum subsp. odorifera and both PCR-RFLP profiles were consistent with the pattern E produced by the minority of Pcc strains. Thus, isolates SXR1 and ZJR1 were characterized as Pcc in spite of some differences. This is the first report that Pcc has been proven as a causal agent of soft-rot disease on P. ternata.     

Survival of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> in infected tissues of <Emphasis Type="Italic">Capsicum annuum</Emphasis> and in soils of the state of Pernambuco,Brazil

The survival of Ralstonia solanacearum A1-9^Rif race 1 phylotype I was studied in ten different soil types in the absence of the host plant as well as in infected tissues of the stem and root of bell peppers buried in the soil at 0, 5, and 15 cm. The survival time of R. solanacearum A1-9^Rif in the ten soil types ranged from 42 up to 77 days. Among the chemical and physical characteristics of the soil, clay content, residual moisture, and available water were positively correlated, and pH was negatively correlated, with survival time, population size at 42 days, and area under the population curve. The pathogen survival differed significantly in relation to the plant tissues, but not with respect to the incorporation depth of the infected tissues. The root tissue of bell pepper supported a larger bacterial population at 7 and 21 days (5 × 10⁴ and 3.1 × 10⁴ CFU g⁻¹ tissue, respectively) compared with the stem tissue (0.35 × 10⁴ and 0.48 × 10⁴ CFU g⁻¹ tissue, respectively) and also had a larger area under the population curve. On the other hand, the stem tissues presented a greater decomposition rate and pH compared with the roots. In conclusion, the different types of studied soils as well as the infected bell pepper tissues were considered potential primary sources of R. solanacearum inocula, but only for a short period.     

Vascular blackening of wasabi rhizomes caused by <Emphasis Type="Italic">Pectobacterium carotovorum</Emphasis> subsp. <Emphasis Type="Italic">carotovorum</Emphasis>

Wasabi (Wasabia japonica) is grown for its highly-valued rhizome which is used as a condiment in Japanese food. Symptoms of vascular blackening in the rhizome were first observed in 2005 in plants grown in British Columbia, Canada. Microscopic observations and microbial isolation from infected tissues revealed that most of the xylem tracheid cells were blackened and bacteria were consistently associated with symptomatic plants. The bacterium most frequently recovered was identified as Pectobacterium carotovorum subsp. carotovorum (Pcc) using BioLog™ and sequencing of a specific ~510 bp IGS region. Pathogen-free plants obtained using meristem-tip micropropagation were inoculated with a wasabi isolate of Pcc. Vascular blackening symptoms developed in the rhizome after 8 weeks when the rhizome was first wounded by stabbing or cutting, or if the roots were pre-inoculated with Pythium species isolated from rhizome epidermal tissues, followed by inoculation with Pcc at 1 × 10⁸ cells ml⁻¹. Xylem tracheid cells were blackened and Pcc was reisolated from all diseased tissues. The highest frequency of rhizome vascular blackening occurred at 22°C and 27°C and these tissues occasionally succumbed to soft rot at higher temperatures, but not when inoculated tissues were incubated at 10°C. The rooting medium used by growers for vegetative propagation of wasabi was shown to contain Pcc but the pathogen was not recovered from the irrigation water. Entry of Pcc through wounds on wasabi rhizomes and the host tissue response result in symptoms of vascular blackening.