Zoospore Cysts Research Articles

Effects of temperature, zoospore concentration, infection period, and fruit maturity on Phytophthora palmivora infection of figs

White powdery rot in figs caused by Phytophthora palmivora is an important disease resulting in severe fruit rot, but is not currently effectively controlled in Japan due to a lack of understanding of its epidemiology. Therefore, the effects of temperature, zoospore concentration, infection period, and fruit maturity on infection of figs were examined by inoculating the fruit with a suspension of P. palmivora zoospores. The zoospores germinated at temperatures from 5 to 35 °C, with the optimum temperature range being 20–35 °C. Germ tube length in zoospore cysts was greatest at 20–30 °C. The disease developed in green figs at temperatures from 20 to 30 °C. Figs inoculated with as few as 10 zoospores per fruit developed severe symptoms at the optimum temperature (25 °C). The minimum infection period required for infection was 2 h at 20–28 °C. All of the figs developed symptoms within an 8 h infection period at 25 or 28 °C, and with a 6 h infection period at 25 °C. All fruit at different stages of development (immature fruit, yellow fruit, and mature fruit) developed symptoms. These results indicate that P. palmivora is capable of infecting figs over a wide range of temperatures, within a short infection period, at a low concentration of zoospores, and at any stage of development. These data could be used to construct forecasting models and develop effective control systems for white powdery rot.

Baseline Sensitivities of New Fungicides and Their Toxicity to Selected Life Stages of Phytophthora Species from Citrus in California.

Phytophthora citrophthora, P. syringae, P. nicotianae, and P. hibernalis are important pathogens of citrus in California but few chemical treatments are currently available. In vitro toxicities of four new fungicides to isolates of Phytophthora spp. from California were determined. Mean effective concentration values to inhibit mycelial growth by 50% for ethaboxam, fluopicolide, mandipropamid, oxathiapiprolin, and mefenoxam were 0.068, 0.04, 0.004, 0.0003, and 0.039 µg/ml, respectively, for 62 isolates of P. citrophthora; 0.005, 0.045, 0.003, 0.0001, and 0.008 µg/ml, respectively, for 71 isolates of P. syringae; 0.016, 0.057, 0.005, 0.0005, and 0.183 µg/ml, respectively, for 31 isolates of P. nicotianae; and 0.030, 0.018, 0.005, <0.0003, and ≤0.001 µg/ml, respectively, for two isolates of P. hibernalis. Mean values for ≥90% inhibition of sporangia formation of four isolates of P. citrophthora were 0.1, 0.28, 0.026, 0.005, and 55 µg/ml for ethaboxam, fluopicolide, mandipropamid, oxathiapiprolin, and mefenoxam, respectively. Zoospore cyst germination of P. citrophthora was most inhibited by oxathiapiprolin and mandipropamid. Chlamydospore formation of P. nicotianae was most sensitive to oxathiapiprolin, with a mean ≥90% reduction (EC>90) of 0.002 µg/ml, moderately sensitive to mandipropamid (EC>90 = 0.2 µg/ml) and mefenoxam (EC>90 = 0.6 µg/ml), and least sensitive to ethaboxam and fluopicolide (EC>90 = 1 µg/ml). Oospore formation of P. nicotianae was inhibited by ≥90% using oxathiapiprolin at 0.0004 µg/ml, mandipropamid at 0.02 µg/ml, ethaboxam at 0.1 µg/ml, or fluopicolide at 0.4 µg/ml, whereas 62% inhibition was obtained by mefenoxam at 40 µg/ml.

Influence of pH and Etridiazole on Pythium Species

Pythium root rot (Pythium sp.) is ubiquitous in Michigan greenhouses that produce herbaceous ornamentals, an industry worth $393 million in the state. Disease symptoms include stunting, flowering delay, root rot, and death. Fungicides that are highly effective against pythium root rot are limited, and pathogen resistance has been documented. The objectives of this study were to determine the sensitivity of Pythium irregulare, Pythium ultimum, and Pythium aphanidermatum isolates from symptomatic herbaceous greenhouse ornamentals to the fungicide etridiazole and to determine the influence of pH and etridiazole on Pythium mycelial growth and asexual reproduction. Isolates were tested in vitro for sensitivity to etridiazole by growing the pathogen on amended V8-agar plates sealed in plastic containers to minimize fungicide loss from the vapor phase. The majority of isolates of all three species were sensitive to the fungicide with EC50 (effective concentration resulting in 50% inhibition of linear growth) values ranging from 0.10 to 5.03 μg·mL−1. Two isolates of P. irregulare had an EC90 (effective concentration resulting in 90% inhibition of linear growth) value &gt;80 μg·mL−1. The acidity of the medium influenced the ability of etridiazole to inhibit Pythium mycelial growth and asexual reproduction. Agar plates amended with 1 μg·mL−1 etridiazole and adjusted to pH 4.5 limited the mycelial growth of two P. aphanidermatum isolates and two P. irregulare isolates by 90% and 56%, respectively, compared with amended agar at pH 6.5. Sporangial formation by P. aphanidermatum was less frequent on mycelial disks incubated in etridiazole-amended sterile distilled water (SDW) at pH 4.5 than pH 6.5 (P &lt; 0.05). P. aphanidermatum zoospore cyst germination was less sensitive to etridiazole than sporangia or mycelial growth; however, the influence of pH and fungicide on cyst germination was significant (P &lt; 0.01). At 250 μg·mL−1 etridiazole and solution pH 4.5, zoospore cyst germination was inhibited 99.9% compared with 94.2% at pH 6.5. In a greenhouse experiment, disease symptoms were observed on ‘Pinto White’ geranium (Pelargonium ×hortorum) in a potting medium infested with P. aphanidermatum and adjusted to pH 4.5 or 6.5; however, plant health and fresh weight were greater in low pH potting medium. Etridiazole, applied as a drench at transplant, did not improve control of root rot for plants grown at low pH (P &gt; 0.05). Fresh weight of plants grown in infested potting medium adjusted to pH 4.5 and amended with a single drench of etridiazole (100 μg·mL−1) was reduced 20%, statistically similar to the untreated control. Adjusting the acidity of irrigation water at the time of etridiazole application in ebb and flow and flood floor production systems could be beneficial in pythium root rot management of certain ornamental crops if plants have tolerance to low pH.

Detection of damping-off of Cape gooseberry caused by Pythium aphanidermatum and its suppression with phosphonate

Cape gooseberry cultivated in commercial greenhouses developed wilt and root rot during the summer growing season of 2013 in Aichi Prefecture, Japan. A Pythium species was isolated from affected roots and identified as P. aphanidermatum on the basis of morphological characteristics and sequence homologies in the rDNA internal transcribed spacer regions. In pathogenicity tests, this isolate caused severe wilt and root rot. P. aphanidermatum, which previously was not known to cause damping-off in Cape gooseberry, was isolated by a bait method from soil samples collected from within and outside of the greenhouses. Pre- and post-inoculation activity of phosphonate and two fungicides, cyazofamid and metalaxyl, was evaluated with in vivo experiments. The three compounds were applied 3 days before inoculation, and phosphonate and cyazofamid also were applied at 24 h after inoculation. All three substances provided similar protectant activity. Phosphonate and cyazofamid also exhibited curative activity. In vitro activity of phosphonate on mycelial growth, sporulation, and zoospore cyst germination was evaluated; the EC50 concentration for the inhibition of mycelial growth was low. Zoospore formation was sensitive to phosphonate, while the germination of encysted zoospores was not.

Penetration of suberized periderm of a woody host by Phytophthora cinnamomi

The mechanisms by which Phytophthora cinnamomi zoospores infect inundated, above‐ground woody stem tissue are described. Using 4–6‐ and 18‐month‐old jarrah seedlings, the infection courts were identified and the invasion of the stems at sites of zoospore cyst binding were described. Stems were inoculated with a suspension of motile zoospores on the green stem/young periderm region. Light microscopy was used to examine penetration at sites of taxis, and fluorescent microscopy was used to examine penetration sites of seedlings with intact periderm. Two main infection courts were identified on stems: the emerging axillary shoot and the region of stem immediately surrounding an axillary shoot, where the periderm was thin or discontinuous. Invasion also occurred at sites where the developing shoot had not yet emerged but was at the stem surface. At these sites the pathogen also directly invaded through the thin‐walled phellem of the periderm surrounding the shoot. Zoospores of P. cinnamomi were not attracted to stomata on mature leaves or green stems. Penetration of the epidermal cell layer of the axillary bud leaf primordia was inter‐ and intra‐cellular; growth of hyphae in the periderm surrounding the shoot was intercellular; while in collenchyma it was inter‐ and intra‐cellular, being intercellular between polyphenolic‐rich cells. Exposed stem collenchyma was also directly invaded immediately adjacent to the young axillary shoot. Zoospores demonstrated taxis to sites of discontinuous periderm, similar to wounded areas where the outer protective layers of the plant are breached. This study presents the first evidence that P. cinnamomi is capable of intercellular penetration of suberized periderm.

BZIP Transcription Factors in the Oomycete Phytophthora infestans with Novel DNA-Binding Domains Are Involved in Defense against Oxidative Stress

Transcription factors of the basic leucine zipper (bZIP) family control development and stress responses in eukaryotes. To date, only one bZIP has been described in any oomycete; oomycetes are members of the stramenopile kingdom. In this study, we describe the identification of 38 bZIPs from the Phytophthora infestans genome. Half contain novel substitutions in the DNA-binding domain at a site that in other eukaryotes is reported to always be Asn. Interspecific comparisons indicated that the novel substitutions (usually Cys, but also Val and Tyr) arose after oomycetes diverged from other stramenopiles. About two-thirds of P. infestans bZIPs show dynamic changes in mRNA levels during the life cycle, with many of the genes being upregulated in sporangia, zoospores, or germinated zoospore cysts. One bZIP with the novel Cys substitution was shown to reside in the nucleus throughout growth and development. Using stable gene silencing, the functions of eight bZIPs with the Cys substitution were tested. All but one were found to play roles in protecting P. infestans from hydrogen peroxide-induced injury, and it is proposed that the novel Cys substitution serves as a redox sensor. A ninth bZIP lacking the novel Asn-to-Cys substitution, but having Cys nearby, was also shown through silencing to contribute to defense against peroxide. Little effect on asexual development, plant pathogenesis, or resistance to osmotic stress was observed in transformants silenced for any of the nine bZIPs.

Histopathological Investigations of the Infection Process and Propagule Development of &lt;em&gt;Phytophthora ramorum&lt;/em&gt; on Rhododendron Leaves

In Europe, cultivated rhododendron is one of the most important hosts for Phytophthora ramorum . To better understand leaf infection and leaf capacity for sporulation, infection studies were carried out. Detached leaves of Rhododendron ‘Catawbiense Grandiflorum ́ and `Brigitte ́ were inoculated with zoospore suspensions of P. ramorum isolates of mating type A1 and A2. ‘Catawbiense Grandiflorum’ developed much more leaf necrosis than ‘Brigitte’ (average necrotic leaf area after 55 days 106.5 mm 2 versus 0.12 mm 2 ). The trichomes on the ‘Brigitte’ leaves seemed to prevent the germ tubes from detecting the stomata. P. ramorum germ tubes from encysted zoospores invaded the leaf tissue via the stomata. Appressoria-like structures were observed. On the infected leaves new hyphae grew out of the stomata. Sporangia and chlamydospores developed on the mycelium from germinated zoospore cysts growing on the leaf surface as well as on the hyphae growing out of the stomata after infection. They could be observed mainly on the necrotic leaf areas. Single healthy-looking oospores developed within 12 days on the ‘Brigitte’ leaves. Observation of the infection process on leaves of Rhododendron ‘Cunningham’s White’ with the scanning electron microscope confirmed the various stages of the infection process.

Galtierella biscalithecae nov. gen. et sp., a Late Pennsylvanian endophytic water mold (Peronosporomycetes) from France

A new fossil water mold (Peronosporomycetes), Galtierella biscalithecae nov. gen. et sp., consisting of coenocytic hyphae occurs as an intracellular endophyte in a partially degraded specimen of the reproductive organ Biscalitheca cf. musata (Zygopteridales) from the Upper Pennsylvanian Grand-Croix cherts (Saint-Étienne Basin, France). Some hyphal tips produce small spheres that subsequently develop into ornamented, opaque-walled oogonia; amphigynous antheridia encircle the necks of several immature oogonia. Also present are ovoid structures, which may represent differently shaped oogonia, hyphal swellings, or zoosporangia. Small dome-shaped structures, probably zoospore cysts, are attached to numerous host cell walls. This discovery sheds new light on the morphology and biology of water molds in a Carboniferous ecosystem.

Use of Cross-Flow Membrane Filtration in a Recirculating Hydroponic System to Suppress Root Disease in Pepper Caused by Pythium myriotylum

Zoosporic pathogens in the genera Pythium and Phytophthora cause extensive root disease epiphytotics in recirculating hydroponic vegetable-production greenhouses. Zoospore cysts of Pythium myriotylum Drechsler were used to evaluate the effectiveness of cross-flow membrane filters to control pythiaceous pathogens in recirculating hydroponic systems. Four membrane filter brands (Honeycomb, Polypure, Polymate, and Absolife) were tested alone or in combination to determine which filters would effectively remove infective propagules of P. myriotylum from solutions and reduce disease incidence and severity. Zoospore cysts of P. myriotylum generally measured 8 to 10 microm, and it was hypothesized that filters with pore-sizes<5 microm would be effective at removing 100% of the infective propagules and protect pepper plants from root infection. Single-filter assays with Honeycomb and Polypure brands removed 85 to 95% of zoospore cysts when pore sizes were rated at 1, 5, 10, 20, or 30 microm. Single-filter assays of Polymate and Absolife brands were more effective, exhibiting apparently 100% removal of zoospore cysts from nutrient solutions on filters rated at 1 to 10 microm. However, plant bioassays with Honeycomb and Polymate single filters failed to give long-term protection of pepper plants. Double-filter assays with 1- and 0.5-microm Polymate filters significantly increased the protection of pepper plants grown in nutrient film technique systems but, eventually, root disease and plant wilt could be observed. Insect transmissions by shore flies were not factors in disease development. Scanning electron microscopy images of zoospore cysts entrapped on Polymate filters revealed zoospore cysts that were either fully encysted, partially encysted, or of unusually small size (3 microm in diameter). It was concluded that either the atypically small or pliable pleomorphic zoospore cysts were able to penetrate filter membranes that theoretically should have captured them.

Assessment of the control of Phytophthora root rot disease spread by Spin Out ®-treated fabrics in container-grown hardy nursery-stock

A non-woven capillary matting fabric (Tex-R ® Pro), coated with a latex polymer-based formulation of cupric hydroxide (Spin Out ®), developed to cover standing-out areas for weed control and to prevent rooting through, was assessed for its potential to reduce the spread of Phytophthora root rot in container-grown hardy nursery-stock. As well as production bed covers, the fabric was cut into discs and (a) used to cover the tops of plant containers (pot toppers) and (b) inserted to cover the holes in the bottoms of plant containers (disc inserts). These were all tested as barriers to the passage of infective zoospores of Phytophthora cryptogea in enclosed re-circulating irrigation systems growing test plants of Chamaecyparis lawsoniana. Bed covers and disc inserts significantly reduced disease spread, as indicated by the incidence of symptoms and infection, and by bait and colony-forming unit tests of re-circulating water, both in overhead- and trickle-irrigated systems. Pot toppers were not effective. This may be explained by the mode of spread of Phytophthora in such irrigation systems, which appears to be largely via capillary water and therefore not intercepted by toppers. These results were supported by in vitro studies on the passage of zoospores through fresh and used fabric samples which showed that all Tex-R Pro fabric materials significantly reduced the survival of zoospores and zoospore cysts (to between 0% and 4.8%) compared with untreated fabrics (between 32.3% and 42.8%). Analysis of Cu 2+ concentrations in re-circulating irrigation water and in matting samples at the end of the field experiments showed that the majority of the active ingredient stayed bound to the fabric. These results demonstrate that it is possible to deploy Spin Out-treated fabrics in nurseries to help prevent the spread of Phytophthora propagules in addition to their primary use for controlling weeds and rooting through.

Sensitivity of Populations of Phytophthora capsici from South Carolina to Mefenoxam, Dimethomorph, Zoxamide, and Cymoxanil.

In summer and fall 2003, Phytophthora blight and crown rot, caused by Phytophthora capsici, was found in three fields each of summer squash and pepper on three farms in two counties in South Carolina. Although this disease had been confirmed previously in the state, five of these outbreaks were in fields thought to be free of P. capsici. The objectives of this study were to determine whether isolates of P. capsici in South Carolina were sensitive to mefenoxam and to determine baseline sensitivities to dimethomorph, zoxamide, and cymoxanil, fungicides recently registered to control Phytophthora blight. Of 120 isolates tested for sensitivity to mefenoxam at 100 mg/liter, 8 isolates were resistant (relative colony diameter [RCD] > 90% of nonamended control), 60 isolates were sensitive (RCD < 30%), and 52 isolates were intermediately sensitive. Only sensitive isolates were found in two fields in which no mefenoxam-containing fungicides had ever been used. Intermediately sensitive or resistant isolates were found in the four fields in which mefenoxam had been applied previously. In all, 15 to 61 isolates were tested for sensitivity to dimethomorph, zoxamide, and cymoxanil. The concentrations at which RCD, percent cyst germination, and relative zoospore production were reduced to 50% (EC50 values) for mycelial growth were 0.19 ± 0.02 (± standard deviation) mg/liter for dimethomorph, 0.50 ± 0.50 mg/liter for zoxamide, and mostly >50 mg/liter for cymoxanil. EC50 values for zoospore cyst germination were 0.07 ± 0.02 mg/liter for dimethomorph and >50 mg/liter for cymoxanil. EC50 values for zoospore production were 0.63 ± 0.42 mg/liter for dimethomorph, 0.47 ± 0.51 mg/liter for zoxamide, and <50 mg/liter for cymoxanil. Sensitivity values obtained in this South Carolina study can be used as a comparative baseline to monitor shifts in sensitivity to the fungicides mefenoxam, dimethomorph, zoxamide, and cymoxanil in populations of P. capsici.

Transgene-Induced Silencing of the Zoosporogenesis-Specific NIFC Gene Cluster of Phytophthora infestans Involves Chromatin Alterations

Clustered within the genome of the oomycete phytopathogen Phytophthora infestans are four genes encoding spore-specific nuclear LIM interactor-interacting factors (NIF proteins, a type of transcriptional regulator) that are moderately conserved in DNA sequence. NIFC1, NIFC2, and NIFC3 are zoosporogenesis-induced and grouped within 4 kb, and 20 kb away resides a sporulation-induced form, NIFS. To test the function of the NIFC family, plasmids expressing full-length hairpin constructs of NIFC1 or NIFC2 were stably transformed into P. infestans. This triggered silencing of the cognate gene in about one-third of transformants, and all three NIFC genes were usually cosilenced. However, NIFS escaped silencing despite its high sequence similarity to the NIFC genes. Silencing of the three NIFC genes impaired zoospore cyst germination by 60% but did not affect other aspects of the life cycle. Silencing was transcriptional based on nuclear run-on assays and associated with tighter chromatin packing based on nuclease accessibility experiments. The chromatin alterations extended a few hundred nucleotides beyond the boundaries of the transcribed region of the NIFC cluster and were not associated with increased DNA methylation. A plasmid expressing a short hairpin RNA having sequence similarity only to NIFC1 silenced both that gene and an adjacent member of the gene cluster, likely due to the expansion of a heterochromatic domain from the targeted locus. These data help illuminate the mechanism of silencing in Phytophthora and suggest that caution should be used when interpreting silencing experiments involving closely spaced genes.

Effect of 2,4-Diacetylphloroglucinol on Pythium: Cellular Responses and Variation in Sensitivity Among Propagules and Species

ABSTRACT The antibiotic 2,4-diacetylphloroglucinol (2,4-DAPG) plays an important role in the suppression of plant pathogens by several strains of Pseudomonas spp. Based on the results of this study, there is variation within and among Pythium spp. to 2,4-DAPG. Also, various propagules of Pythium ultimum var. sporangiiferum, that are part of the asexual stage of the life cycle, differ considerably in their sensitivity to 2,4-DAPG. Mycelium was the most resistant structure, followed by zoosporangia, zoospore cysts, and zoospores. Additionally, we report for the first time that pH has a significant effect on the activity of 2,4-DAPG, with a higher activity at low pH. Furthermore, the level of acetylation of phloroglucinols is also a major determinant of their activity. Transmission electron microscopy studies revealed that 2,4-DAPG causes different stages of disorganization in hyphal tips of Pythium ultimum var. sporangiiferum, including alteration (proliferation, retraction, and disruption) of the plasma membrane, vacuolization, and cell content disintegration. The implications of these results for the efficacy and consistency of biological control of plant-pathogenic Pythium spp. by 2,4-DAPG-producing Pseudomonas spp. are discussed.

Laboratory studies to assess the risk of development of resistance to zoxamide.

Laboratory studies were conducted to evaluate the risk of developing field resistance to zoxamide, a new Oomycete fungicide which acts on microtubules. Zoxamide, metalaxyl and dimethomorph were compared with respect to the ease with which fungicide-resistant mutants could be isolated and their level of resistance. Attempts to generate mutants of Phytophthora capsici and P infestans with resistance to zoxamide by mycelial adaptation on fungicide-amended medium were unsuccessful. Similarly, changes in sensitivity to zoxamide were small (resistance factors < or = 2.2) in mutants of P capsici isolated by chemical mutagenesis of zoospore cysts. In parallel experiments with metalaxyl, highly resistant mutants were obtained using both adaptation (P capsici or P infestans) and chemical mutagenesis (P capsici). For dimethomorph, chemical mutagenesis (P capsici) yielded moderately resistant mutants (maximum resistance factor = 20.9), and adaptation (P capsici or P infestans) did not induce resistance. It is proposed that failure to isolate mutants resistant to zoxamide results from the diploid nature of Oomycete fungi and the likelihood that target-site mutations would produce a recessive phenotype. Our studies suggest that the risk of a highly resistant pathogen population developing rapidly in the field is much lower for zoxamide than for metalaxyl. However, as with any site-specific fungicide, appropriate precautions against resistance development should be taken.

Impact of Azoxystrobin, Dimethomorph, Fluazinam, Fosetyl-Al, and Metalaxyl on Growth, Sporulation, and Zoospore Cyst Germination of Three Phytophthora spp.

In vitro activity of azoxystrobin, dimethomorph, and fluazinam on growth, sporulation, and zoospore cyst germination of Phytophthora capsici, P. citrophthora, and P. parasitica was compared to that of fosetyl-Al and metalaxyl. The 50% effective concentration (EC50) values for)inhibition of mycelial growth of the three pathogens usually were lowest for dimethomorph and (metalaxyl, ranging from <0.1 to 0.38 μg/ml. However, the 90% effective concentration (EC90) levels for dimethomorph always were lower than the other four tested compounds, with values ranging from 0.32 to 1.6 μg/ml. Mycelial growth of P. capsici, P. citrophthora, and P. parasitica was least affected by azoxystrobin and fluazinam, with estimated (EC90) values >3,000 μg/ml. Reduction of sporangium formation by P. capsici, P. citrophthora, and P. parasitica in the presence of dimethomorph at 1 μg/ml was significantly greater than that recorded for the same concentration of azoxystrobin, fluazinam, and fosetyl-Al. For the three species of Phytophthora, zoospore motility was most sensitive to fluazinam (EC50 and EC90 values of <0.001 μg/ml) and (least sensitive to fosetyl-Al, with (EC50 and EC90 values ranging from 299 to 334 and 518 to 680 μg/ml, respectively). Germination of encysted zoospores of P. capsici, P. citrophthora, and P. parasitica was most sensitive to dimethomorph (EC50 and EC90 values ranging from 3.3 to 7.2 and 5.6 to 21 μg/ml, respectively), intermediate in sensitivity to fluazinam (EC50 and EC90 from 18 to 108 and 67 to >1,000 μg/ml, respectively) and metalaxyl (EC50 and EC90 from 32 to 280 and 49 to 529 μg/ml, respectively), and lowest in sensitivity to azoxystrobin and fosetyl-Al (EC50 and EC90 from 256 to >1,000 μg/ml). The activity of azoxystrobin, dimethomorph, and fluazinam on one or more stages of the life cycle of P. capsici, P. citrophthora, and P. parasitica suggests that these compounds potentially could provide Phytophthora spp. disease control comparable to that of the established fungicides fosetyl-Al and metalaxyl.

Ultrastructural analysis of Saprolegnia secondary zoospore cyst ornamentation from infected wild brown trout, Salmo trutta L., and river water indicates two distinct morphotypes amongst long‐spined isolates

Transmission electron microscopy was used to study the ornamentation of secondary cysts of 52 long‐spined Saprolegnia isolates, with and without oogonia, obtained from wild brown trout, Salmo trutta L., and water samples from various rivers in León (NW Spain). All the isolates had secondary cysts with long hooked hairs grouped in bundles, although significant individual differences were observed. There was a direct relationship between the number of bundles per cyst, the number of hairs per bundle and hair length. The isolates were classified into two morphological groups. The isolates with a higher number of bundles per cyst and bundles with a greater number and length of hairs, were included in cyst morphological Group I. The isolates with a smaller number and shorter length of these bundles and hairs were included in cyst morphological Group II. There was a relationship between the type of ornamentation of the secondary cysts and the origin of the isolates. The isolates from salmonids with saprolegniosis belonged to Group II, whereas most isolates from mucus and water belonged to Group I.

Two new species of Haptoglossa, H. erumpens and H. dickii, infecting nematodes in cow manure

Two species of Haptoglossa isolated from nematodes from cow manure from N.E. England are described. H. erumpens sp. nov., infecting Bunonema nematodes, is aplanosporic, the cysts being released after rupture of the nematode cuticle in the absence of an evacuation tube. H. dickii sp. nov. is zoosporic, infecting several species of rhabditid nematodes. Aplanospores and zoospore cysts germinated to produce secondary spores (gun cells) with which to infect further nematode hosts. Some key ultrastructural features in both species have been included.

Germination triggers of zoospore cysts of Aphanomyces euteiches and Phytophthora parasitica

Zoospore cysts of four isolates of Aphanomyces euteiches with different host-specific pathogenicity germinated in response to peptone, pea root extract, CaCl2 or gum arabic but, in tests on one isolate, did not respond to several individual sugars, amino acids or polysaccharides. A. euteiches showed differential responses to substances applied at different times in the encystment process. Gum arabic triggered germination only when added to motile zoospores, which it caused to encyst. CaCl2 triggered germination when added to motile zoospores or during vortex-encystment of zoospores, but seldom immediately ( Zoospore taxis, encystment and germination of A. euteiches and P. parasitica cysts were recorded on pea roots by video microscopy. The speed of germination and pattern of sporeling development on roots were most closely matched by treatment of motile spores with gum arabic in pure culture. The behaviour of A. euteiches conformed to a previous model for Pythium and Phytophthora spp., in which zoospores encyst by recognition of root surface components (simulated by gum arabic), germinate autonomously by a calcium-mediated process (simulated by calcium treatment during vortexing), and the germ-tube produces an assimilative hypha (simulated by peptone or pea root extract).

Calcium Interference with Zoospore Biology and Infectivity of Phytophthora parasitica in Nutrient Irrigation Solutions

ABSTRACT Calcium, applied as either CaCl2 or Ca(NO3)2 to water or calcium-free soluble fertilizer solution (Peters 20-10-20 Peat Lite Special), affected several important stages of Phytophthora parasitica zoospore behavior relevant to infection and disease spread. Release of zoospores from sporangia was suppressed by Ca(2+) concentrations in the range of 10 to 50 meq. These concentrations also curtailed zoospore motility; 20 meq of Ca(2+) in fertilizer solution caused all zoospores to encyst within 4 h, whereas 94% of zoospores remained motile in unamended solution. In addition, Ca(2+) in the range of 10 to 30 meq stimulated zoospore cysts to germinate in the absence of an organic nutrient trigger, while suppressing the release of a single zoospore (diplanetism) from cysts that did not germinate. In growth chamber experiments, the amendment of the fertilizer solution with 10 or 20 mM Ca(NO3)2 greatly suppressed infection of flood-irrigated, containerized vinca seedlings in a peat-based mix by motile or encysted zoospores of P. parasitica. These results demonstrate that Ca(2+) amendments interfere with P. parasitica zoospore biology at multiple stages, with compounding effects on epidemiology, and suggest that manipulation of Ca(2+) levels in irrigation water or fertilizer solutions could contribute to management of Phytophthora in recirculating irrigation systems.

Cues for Zoospore Release by Marine Oomycotes in Naturally Decaying Submerged Leaves

Two common potential environmental impacts upon leaf-decomposers in the marine intertidal zone (mild drying, dilution of salinity/lowering of temperature due to rainfall) were simulated and tested as enhancers of zoospore release by species of Halophytophthora. Senescent leaves of mangroves and oak were used as bait in flow-through chambers to quantitate zoospore passage from disks of naturally decaying leaves. Older-decaying leaves of red and black mangrove had low rates of zoospore release by H. vesicula, the predominant oomycote encountered (< 15% of bait samples occupied in 24 h), and environmental cues had no detectable effect. Younger-decaying leaves of red mangrove had considerably higher rates of release, and rates were stimulated 2-fold by dilution (to 50% of bait samples positive). Rates for oak leaves were doubled by both dilution and drying (to 86–92% positive). We speculate that: i) halophytophthoras complete their occupation of submerged leaves (from attachment of zoospore cysts to release of zoospores from sporangia) early in the decomposition of leaves, before substantial entry into the leaves of labyrinthulids (predators); ii) environmental cuing results in high concentrations of halo- phytophthoran zoospores in shallow sheets or pools of low-tidal waters, enhancing probability of rapid contact with new-fallen leaves.