Damping-off of broccoli (Brassica oleracea var. italica) and cauliflower (B. oleracea var. botrytis) seedlings occurred in several greenhouses in Fresno, CA, in 1997. Symptoms included wilting and root and stem rot. Pythium polymastum was consistently isolated from symptomatic tissues placed on corn meal agar amended with 10 ppm pimaricin, 250 ppm ampicillin, 10 ppm rifampicin, and 25 ppm pentachloronitro-benzene. On grass leaves in water, the fungus produced numerous aplerotic oospores in oogonia 43 to 50 μm in diameter (average 46 μm) with spines about 7 μm long. Spherical sporangia were only rarely observed. In the greenhouse, 4-week-old broccoli and cauliflower seedlings were transplanted into potting mix amended with a colonized vermiculite/rye/V8 juice medium to produce approximately 2,500 CFUs per gram of potting medium. Control plants were transplanted into noninfested potting mix. There were six replicate pots per treatment and three plants per pot. After 12 days, the potting mix was gently washed from the roots and the seedlings were dried and weighed. Symptoms on inoculated plants included wilting, severe root rot, black streaks on the lower stems, and death. The fungus was recovered from symptomatic tissues. There were no symptoms on the control plants. Infection by P. polymastum reduced dry weights of surviving broccoli and cauliflower seedlings by 82 and 58%, respectively. Similar results were obtained in a second experiment. This fungus was previously characterized as a pathogen of both cultivated and wild crucifers in Canada (1). This is the first report of P. polymastum in California. Reference: (1) T. C. Vanterpool. Can. J. Bot. 52:1205, 1974.

Polygonum odoratum (= Persicaria odorata), known as rau ram or sang hum, is native to southeastern Asia and is a common herb in Vietnamese cuisine (1). It has been studied most extensively for its aromatic compound content (2). In Florida, rau ram commonly is grown hydroponically in greenhouses using large, cement beds with recirculated water. The plants form dense mats from which new growth is trimmed for market. During January of 2002, a severe dieback was observed in one production house in Saint Lucie County, FL. Plants with less severe symptoms were yellowed and stunted. Roots of symptomatic plants were largely decayed with root symptoms beginning as a tip necrosis. The cortex of severely affected roots slipped off easily, leaving a stringy vascular system. Plating of symptomatic tissue from 20 randomly selected plant samples was performed with multiple general and selective media including potato dextrose agar, corn meal agar with pimaricin, ampicillin, rifampicin, and pentachloronitrobenzene (PARP) (3). All colonies produced were identified as Pythium helicoides Drechsler on the basis of sporangial, oogonial, and antheridial characteristics (4). Isolates had proliferous, obovoid, papillate sporangia, and were homothallic with smooth-walled oogonia and thick-walled, aplerotic oospores. Multiple antheridial attachments per oogonium were common with the antheridium attached along its entire length. Pathogenicity tests were conducted using P. odoratum plants grown from commercial transplants. Two tests were performed. Each test was conducted using eight inoculated and eight control plants. In the first test, plants were maintained in 10-cm pots immersed in sterilized pond water for the duration of the test. Plants were inoculated with five 7- × 70-mm sections of freshly growing mycelial culture per plant using 10-day-old cultures of Pythium helicoides grown on water agar. Chlorosis was observed at approximately 2 months after inoculation. Root necrosis was observed in inoculated plants approximately 5 months after inoculation. This test was performed in the greenhouse with temperatures ranging from 20 to 30°C. The second test was performed in growth chambers at 35 to 40°C. Plants were maintained in 10-cm pots immersed in Hoagland's solution and were inoculated with four 6-mm plugs per plant. Symptoms were observed on inoculated plants at this temperature within 1 week of inoculation. No chlorosis or root decay was observed in noninoculated, immersed plants. The pathogen was reisolated from inoculated, symptomatic tissue. To our knowledge, this is the first report of root rot of P. odoratum caused by Pythium helicoides. References: (1) R. E. Bond. Herbarist 55:34, 1989. (2) N. X. Dung et al. J. Essent. Oil Res. 7:339, 1995. (3) M. E. Kannwischer and D. J. Mitchell. Phytopathology 68:1760, 1978. (4) A. J. van der Plaats-Niterink. Monograph of the Genus Pythium. Vol. 21, Studies in Mycology. Centraalbureau voor Schimmelcutltures, Baarn, The Netherlands, 1981.

Adiaspiromycosis produced by the fungus Emmonsia crescens is a pulmonary disease afflicting primarily small wild mammals. Man, too, may become an accidental link of the saproparatrophic circulation of the agent. Humans are infected--similarly to other mammals--by inhaling the elements of the saprophytic stage of the fungus living for long time periods in the soil substrates. After the infecting cells, aleuriospores, have invaded the host lungs, they are converted into the elements of the parasitic stage--adiaspores. These are surrounded by granulomatous tissue and reach up to 700 microns in diameter. In the circulation of the agent, the infected wild mammals play a role of reservoir hosts harbouring the parasitic stage of the fungus in nature for a relatively long time. In some cases these animals also enable spread of adiaspiromycosis from exoanthropic foci into human habitations. Clinical and experimental studies show that the result of an infection may be--in addition to its liquidation--an asymptomatic form of disease or a disseminated pulmonary process. In addition to as yet insufficiently proved proper action of fungus cells, a reduction in the functional pulmonary parenchyma plays a role in the pathogenesis of the pulmonary forms. An existence of extrapulmonary forms of adiaspiromycosis is not excluded. Serological methods have not been routinely used for diagnosis as yet: immune reaction of the organism has a character of antibody response and delayed hypersensitivity. Cell mediated immunity has not been studied as yet. Treatment of human disease is primarily a surgical one. The fungistatic drugs pimaricin or amphotericin B may be employed, corticosteroids may be indicated in individual cases. The efficacy of modern antifungal substances has not been established as yet.

In May 2005, branches originating from five separate whorls below the terminal on a single California red fir (Abies magnifica) in a mixed grand fir (Abies grandis) and Douglas-fir (Pseudotsuga menziesii) Christmas tree plantation near Los Gatos, CA displayed wilting and dieback of new shoot growth. Brown dieback, delineated by needle loss, extended 6 to 8 cm into 1-year-old and sometimes 2-year-old growth. The ~7-year-old, 1-m tall tree was located near the edge of the plantation, beneath an overstory of California bay laurel (Umbellularia californica) trees that were infected with Phytophthora ramorum. Isolations from dieback margins onto corn meal agar amended with ampicillin, rifamycin, and pimaricin (CARP) yielded hyphae and large, dark brown chlamydospores that were morphologically consistent with P. ramorum (1). Microsatellite analysis confirmed that isolates were of the NA1 lineage of P. ramorum. Isolates were deposited in the Washington State University Puyallup Phytophthora Master Collection. Dormant bareroot California red fir seedlings were obtained from the USDA Forest Service Placerville Nursery (Camino, CA) in February 2006 and planted in SC-10 super cell cones (Stuewe & Sons, Inc., Tangent, OR) in a standard greenhouse potting mix. Seedlings (average height 11 cm) were then forced to initiate bud break and new shoot elongation (0.5 to 1.5 cm) in a greenhouse at 21°C. Eight unwounded seedlings were inoculated with a zoospore suspension (4.185 × 10⁵ zoospores/ml of sterile water) produced from 3- to 4-week-old V8 juice agar cultures of isolate WSU#106-0021 using an artist's airbrush powered by Badger Propel canned propellant. Eight control seedlings were sprayed with water alone. Seedlings were placed in plastic tubs with ~2.5 cm of warm water in the bottom to provide humidity. A plastic bag supported by a wire frame was used to cover each tub. Tubs were placed in a biocontainment unit at 15 to 16°C under 24 h of fluorescent light. The plastic was removed after 5 days and seedlings were left under the same conditions. Seven days after inoculation, 25 to 100% (average 68%) of the new shoots on each of the eight inoculated seedlings were wilted and 100% of these seedlings exhibited dark brown dieback into the 1-year-old stems (range of 1.0 to 2.3 cm, average 1.6 cm). Tissues from shoots and dieback edges were plated onto CARP media. All of these attempts resulted in successful isolation of P. ramorum, and cultures exhibited the same hyphal morphology and chlamydospore characteristics when compared with the isolate tested. Control plants did not develop symptoms. This trial completes Koch's postulates to establish California red fir as a host of Phytophthora ramorum. To our knowledge, this site contains the only reported infection of California red fir by P. ramorum. The potential for infection within its native range is unknown. Reference: (1) S. Werres et al. Mycol. Res. 105:1155, 2001.

In the annual Norwegian Phytophthora ramorum survey in 2009, wild bilberry samples, collected during September and October in a semimanaged park (arboretum) in the southwest coast of Norway, tested positive in a P. ramorum-specific real-time PCR test (1). Necrotic lesions were observed in shoot tips, branching points, and around leaf abscission scars. The lesions were of variable dimensions. In the samples collected in October, some lesions were confluent and completely covered some shoots. After direct detection on plant material, P. ramorum was isolated from necrotic lesions of the stems on semiselective media PARP (corn meal agar amended with pimaricin, ampicillin, rifampicin, and pentacloronitrobenzene) (2). The isolates were identified by the production of abundant chlamydospores on agar and deciduous, semipapillate sporangia that is characteristic of P. ramorum (3). Sexual structures were not observed. Three pure cultures obtained from different plant samples also tested positive for P. ramorum by the specific real-time PCR test (1). All positive samples were found in close vicinity of infected rhododendron plants. In this location, P. ramorum had already been detected on rhododendron in 2005. A pathogenicity test was performed with two isolates from bilberry and one from rhododendron. Wild asymptomatic bilberry plants were collected at the end of June in the forest around Oslo. Two shoot tips with 6 to 10 leaves each and one small part of a branch with several shoots and immature berries were used for testing each isolate. The inoculations were made by dipping the shoots in a zoospore suspension (2 to 3 × 10⁴ zoospores ml^-1) for 1 min. Inoculated material was placed in moist incubation chambers and incubated at room temperature (19 to 24°C). Controls were obtained by dipping shoots in sterile water. After 2 days, lesions were observed on leaf laminae from all the shoots inoculated with the three different isolates. After 4 days, severe petiole necroses were observed and leaves abscised easily from the stems. Symptoms on the stems were observed in the apical part or areas around the nodes. Some shoots were almost completely necrotic. Heavy sporulation was observed on the berries. P. ramorum was reisolated from leaves and stems of inoculated shoots for all the isolates. P. ramorum was not recovered from control plants. To our knowledge, this is the first report of P. ramorum on bilberry in Norway. References: (1) K. L. Hughes et al. Phytopathology 96:975, 2006. (2) M. E. Kannwischer and D. J. Mitchell. Phytopathology 68:1760, 1978. (3) S. Werres et al. Mycol. Res.105:1155, 2001.

During a 2019-2020 survey of plant pathogenic oomycetes in Nanjing, China, a cluster of five adjacent Rhododendron pulchrum plants in Xuanwuhu Park exhibited symptoms including crown and root rot and wilting. foliage blight caused due to collar and had rotting crown and root tissues resultingrot foliage blight. Diseased roots were rinsed in water, cut into 10 mm pieces, immersed in 70% ethanol for 60 sec, and plated onto clarified V8 juice agar (cV8A) containingamended with pimaricin (20 mg/liter), ampicillin (125 mg/liter), rifampicin (10 mg/liter), and pentachloronitrobenzene (20 mg/liter). After three3 days of incubation at 26°C, Ffive Pythium-like isolatescoloniesisolates were obtained using hypalhyphal-tipping after 3 days of incubation at 25°C. Ten agar plugs (2×2 mm2) of each isolate were growntransferred into 10 mLl of 10% clarified V8 juice (cV8) in a 100 -mm plate at 26°C to produce mycelial mats. After 3three days, cV8 was replaced with sterile water. To stimulate sporangial production, 3-5 drops of soil extract solution were added to each plate. Five isolates had identical morphological features. Sporangia were terminal, ovoid to globose, andmeasuring 34.2 ± 6.2 µm (24.0-42.5 µm range) in length and 30.7 ± 6.6 µm (20.9-41.1 µm range) in width. Oogonia were not observed. The following primers were used to amplify the rDNA internal transcribed spacer (ITS) region and the mitochondrial cytochrome c oxidase subunit 1 (cox1COI) and 2 (cox2COII) genes of from aA representative isolate, PH-C were amplified using the primer pairs ITS6 and ITS4 (Cooke et al. 2000), OomCoxI-Levup and OomCoxI-Levlo (Robideau et al. 2011) and Cox2-F and Cox2-RC4 (Hudspeth et al. 2000), respectivelyPhe-1. Isolate A xxx675 bp, xxx657 bp and 561xxx bp fragmentPH-C , respectively were amplified and had have identical sequences of the ITS (GenBank ACN. MT824568), and cox1 (MT834959), COI and cox2 COII genes the rDNA internal transcribed spacer (ITS) region and the mitochondrial cytochrome c oxidase subunit 1 and 2 genes (GenBank ACN. MT824568, MT834959, (MT834958, respectively) sequences identical to those of Phytopythium helicoides (MN541109, MK879709, KT595689, respectively). Based on the morphological and molecular characters, all five isolatesthe causal agent waswere identified the species represented by Phe-1 was identified as P. helicoides. One-year-old R. pulchrum plants (approx. 0.3 m in height) grown in 8×8 cm2 pots were used in to test the pathogenicity trials. Ten plants wasere carefully dug up to expose root ballsclusterballs. TenThree- days -old cultures of the isolate PH-Che-1 were used as the inoculum. Five The pplantss wereere inoculated by inserting 10 agar plugs into thee root ball of each plantcluster. For inoculatingfive control plants, sterile cV8A discsplugs were used. All inoculated plants were re-potted using original fresh potting mix and potsture .Ten 3-day-old cV8A cultural plugs (5×5 mm2) of Phe-1 were evenly insert into the root ball of each of five plants, while sterile cV8A plugs were used for five control plants. All were then planted into their original pots. Plants were maintained in a growth chamber set at 26°C with a 12/12 h light/dark cycle and irrigated as needed. After 21-25 days, the inoculated plants had symptoms identical to those in the field, while the controls remained asymptomatic. Identical outcomes were obtained from two repeated The pathogenicity trials. test was repeatedconducted twice . and the coutcome was identical. Phytopythium. helicoides (Phe-1) was reisolated from all symptomatic plants inemerging from the pathogenicity trials. Phytopythium helicoides was found causing diseases of Asian lotus (Yin et al. 2015), mandarin orange (Chen et al. 2016), and kiwifruit (Wang et al. 2015) plants in China. Phytopythium isolates with identical morphological features to those of Phe-1 were recovered from rotted crown and root tissues of all inoculated plants. In this note, P. helicoides causing crown and root rot on R. pulchrum is reported for the first time. Globally, this is the first report of P. helicoides causing crown blight and root rot of R. pulchrum. Additional surveys are being conducted forto mapping the distribution of P. helicoides in Nanjing, Province of China.

Keywords: Causal Agent; Crop Type; Epidemiology; Oomycetes; Ornamentals; Subject Areas; disease development and spread; woody ornamentals.

Mold inhibitors such as sorbates, propionates and benzoates have been used commercially for some time. Recently these and other potential inhibitors have been studied from the standpoint of their effects on growth of potentially toxic molds and mycotoxin production. In addition, other substances such as the antifungal antibiotic natamycin (pimaricin) and plant-derived products such as components of the essential oils of certain herbs and spices have recently been studied for their antifungal properties and effects on mycotoxin production. Some of these inhibitors inhibit mycotoxin production by greater than 70%, while only inhibiting growth of the mold by 25% or less. Of the organic acids, sorbic, propionic and benzoic, sorbic and its sorbate salts seems to be most effective over the widest range of conditions in preventing mold growth and mycotoxin production. Potassium sorbate is effective against toxic molds at levels of 0.10 to 0.15%. The antibiotic natamycin is very effective in preventing mold growth and toxin production at very low (0.001 to 0.005%) concentrations. A number of herbs and spices possess antifungal activity. At a level of 2.0%, in YES agar, cloves, cinnamon, mustard, allspice, garlic and oregano all completely inhibit mycotoxin production by a number of mycotoxigenic molds. Cloves, cinnamon and mustard seem to be the most effective of those tested, with complete inhibition occurring with amounts of spice less than 1%. Essential oils of orange and lemon also have antifungal properties at levels of 0.2 % and higher. Certain insecticides and fumigants also inhibit mold growth and mycotoxin production. The organophosphates naled and dichlorvos are both effective inhibitors at relatively low concentrations (0.002 to 0.01%). Phenolic antioxidants, particularly BHA, also inhibit toxic molds in concentrations of 0.025% and above. Naturally occurring methylxanthines, such as caffeine and theophylline, inhibit growth and aflatoxin production by A. parasiticus in concentrations of 0.1% and above. Chlorine, a commonly used sanitizer, will inactivate spores of toxic Aspergillus and Penicillium species at levels of residual chlorine commonly achieved with most sanitation procedures. Even though considerable information is available on inhibitory effects of a number of substances on mold growth and mycotoxin production, more work is needed to further define the conditions under which commercial antifungal agents are most effective in preventing growth of toxic molds and mycotoxin production.

In 2004, damages resembling those caused by Phytophthora spp. were observed in a 15-year-old bough plantation of noble fir (Abies procera). When removing bark upward from the roots and base of a diseased tree, a reddish brown discoloration with distinct borders to surrounding wood appeared. The discoloration extended approximately 1.5 m above ground, but only on one side of the stem. This resulted in dead basal branches (flagging) on the cankered side of the tree. Other dying trees in the same field did not show flagging symptoms but turned chlorotic to brown after being girdled by the expanding stem canker. Approximately 25% of the trees were dead or dying. Isolations were carried out from the area between healthy and diseased tissue both from roots and base of the stem of the tree with flagging symptoms. Samples were rinsed in running tap water and plated on the Phytophthora selective medium PARP (17 g of cornmeal agar, 10 mg of pimaricin, 250 mg of ampicillin, 10 mg of rifampicin, and 100 mg of pentachloronitrobenzene (PCNB) in 1 liter of water), with and without hymexazol added (50 mg/l). Morphological characters of the isolated Phytophthora sp. included nonpapillate sporangia (37 to 64 μm), internal proliferation, and characteristic hyphal swellings. The isolate was heterothallic and produced amphigynous antheridia when crossed with tester strains of P. cryptogea. The mating type was A2. The internal transcribed spacer (ITS) rDNA sequences were identical to P. cambivora (GenBank Accession No. AY880985). Thus, both morphological characters and DNA analysis supported the species identification. A pathogenesis test to fulfill Koch's postulate was carried out during 2005. Inoculation was done by placing agar with culture in the growth medium close to the roots of noble fir seedlings. Eleven weeks after inoculation, clearly visible stem canker symptoms were observed. The ITS sequences of the reisolated Phytophthora sp. were determined and found identical to P. cambivora. P. cambivora was reported to cause root rot and stem canker in a noble fir Christmas tree plantation in the United States (1). P. citricola and P. citrophthora are known to cause problems on Lawson Falsecypress/Port-Orford-cedar (Chamaecyparis lawsoniana) in Norway, but damages by Phytophthora spp. have never been reported in Abies spp. plantations or forest stands in Norway. Currently, we are also working on Phytophthora problems discovered in two different Christmas tree plantations (A. lasiocarpa and A. nordmanniana). Reference: (1) G. A. Chastagner et al. Plant Dis. 79:290, 1995.

English (Persian) walnut (Juglans regia), among the most widely cultivated species of Juglans worldwide, is cultivated primarily for fruit production but also for timber. In the last 10 years, walnut decline causing leaf yellowing, sparse foliage, overall decline, and plant death has increased in Italian commercial orchards. In Italy, Phytophthora cactorum, P. cambivora, P. cinnamomi, and P. cryptogea are associated with this disease (1,4). Over the last 5 years, P. cinnamomi was the most widely isolated and destructive species (1). Recently, a different species of Phytophthora was isolated from diseased roots and soil from around lateral roots of 10 declining trees in two orchards in the Veneto Region of northern Italy. Another species of Phytophthora was isolated consistently from rotted roots of declining walnut trees in two orchards in the Campania Region of southern Italy. Phytophthora spp. were isolated directly from plant material or Rhododendron spp. leaf baiting on soil samples with PARBhy selective medium (10 mg of pimaricin, 250 mg of ampicillin [sodium salt], 10 mg of rifampicin, 50 mg of hymexazol, 15 mg of benomyl, 15 g of malt extract, 20 g of agar in 1,000 ml of H₂O). Two species of Phytophthora were identified based on morphological and cultural characteristics (2). The species from trees in the Veneto Region was identified as P. nicotianae. All isolates produced papillate, spherical to obturbinate, occasionally caducous sporangia with short pedicels, terminal and intercalary chlamydospores, and were mating type A2. The species isolated from trees in the Campania Region was identified as P.citricola. Isolates were homothallic, produced semipapillate, persistent, obclavate to obpyriform sporangia, occasionally with two apices, and antheridia paragynous. Identifications were confirmed by comparing restriction fragment length polymorphism patterns of the internal transcribed spacer region of rDNA with those obtained from previously identified species of Phytophthora. Pathogenicity of two isolates each of P. citricola and P. nicotianae was tested on 2-year-old potted walnut seedlings. Inocula were prepared by inoculating sterilized millet seeds moistened with V8 broth with plugs of mycelium and incubated for 4 weeks at 20°C in the dark. Infested seeds were added to potting soil at a rate of 3% (wt/vol). One day later, pots were flooded for 48 h to promote sporulation. Ten noninoculated seedlings were used as the control. Symptoms were assessed 2 months after inoculation. Seedlings inoculated with P. nicotianae developed necrosis of feeder and lateral roots, but only limited infection of taproots. Seedlings inoculated with P. citricola developed necroses at the insertion points of lateral roots. All four isolates produced visible damage to lateral roots on inoculated plants. P. nicotianae and P. citricola were reisolated from respectively infected roots. Results from these inoculations confirmed P. nicotianae and P. citricola as root pathogens of English walnut. Both species were associated with walnut decline as reported in the United States (3). To our knowledge, this is the first report of P. nicotianae and P. citricola on J. regia in Europe. References: (1) A. Belisario et al. Petria 11:149. (2) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN, 1996. (3) M. E. Matheron and S. M. Mircetich. Phytopathology 75:977, 1985. (4) A. M. Vettraino et al. Plant Dis. 86:328, 2002.

Phytophthora ramorum causes shoot and foliar blight on Rhododendron spp., Viburnum spp. (4), Pieris spp. (2), Kalmia latifolia, and Camellia spp. in several European countries (1-4). In December 2002, we received diseased C. japonica growing in containers from several nurseries in Galicia (northwestern Spain). These young camellia plants had leaves with brown-to-black, water-soaked lesions with diffuse borders that expanded into larger blotches resulting in dead leaves and necrotic lesions on the petioles. Eventually the entire plant wilted and died. Tissue from the leading edge of the lesions was transferred to a selective medium (V8 agar supplemented with pimaricin (10 μg/ml), rifampicin (25 μg/ml), hymexazol (5 μg/ml), and benomyl (5 μg/ml)) and incubated for 3 to 4 days at 20°C in the dark. A Phytophthora sp. was isolated, transferred to carrot piece agar (CPA) (4), and incubated in alternating light. Isolates exhibited coralloid mycelium with concentric rings and a radial growth of 2.5 to 3 mm per day at 20°C. The hyaline-to-yellowish chlamydospores were terminal and intercalary, occasionally lateral, and 24 to 74 μm in diameter. The caducous, sympodial, semipapillate sporangia had a length/breadth ratio of 1.8 to 2.1 and a short pedicel (<5 μm) or no pedicel. Oogonia, antheridia, and oospores were produced by pairing the isolates with P. cryptogea A2 tester BBA 63651 (3,4) provided by S. Werres. Oogonia were subspherical and smooth-walled, antheridia were amphyginous, and oospores were plerotic. The internal transcribed spacer-rDNA polymerase chain reaction (PCR) product obtained by using DNA extracted from mycelium and nested PCR with P. ramorum-specific primers was the size reported for P. ramorum (1). Pathogenicity of the isolates was confirmed by inoculating detached leaves of C. japonica. Five isolates were tested on leaves from 15 young plants growing in containers. Three leaves of each plant were detached and inoculated with each isolate. Leaves were dipped for 5 min into a suspension of sporangia and mycelial fragments and maintained at 20°C and 95% humidity. After 15 days, lesions that developed from the petiole base exceeded 25 mm, and the pathogen was consistently reisolated from the lesions. Leaves inoculated with water from sterile CPA plates did not develop symptoms. A C. japonica isolate has been deposited in the Spanish Type Culture Collection (CECT 20519). To our knowledge, this is the first report of P. ramorum on C. japonica in Spain, though the pathogen has been isolated from Rhododendron spp. and Viburnum tinus growing in several nurseries in Galicia. References: (1) J. M. Davidson et al. On-line publication doi:10.1094. Plant Health Progress, PHP, 2003-0707-01-DG, Plant Management Network. (2) A. J. Inman et al. First Report of Ramorum Dieback (Phytophthora ramorum) on Pieris in England. On-line publication. New Dis. Rep. Vol. 7, British Society for Plant Pathology, 2003. (3) E. Moralejo et al. Plant Dis. 86, 9:1052, 2002. (4) S. Werres et al. Mycol.Res.105:1155, 2001.

European beech (Fagus sylvatica) is an important forest tree species common in northern and central Europe. In Italy, this species is typical in mountain areas over 1,000 m above sea level. In the last decade, decline and death was reported on European beech caused by several Phytophthora species (2), and P. pseudosyringae was recently reported in Italy (3). During 2004 and 2005, seven declining and dying F. sylvatica trees, older than 20 years, were observed in the Veneto Region of Italy with symptoms of bleeding cankers at the base of trunks and on branches. Cankers on the collar showed tongue-shaped necroses of the inner bark and cambium tissues. Four trees were in a public park of Mestre and three were in a forest stand in the province of Belluno. Samples were taken from declining trees, one in the park and two in the forest. Inner bark and cambium tissue pieces were cut from the canker margins, cultured on CARPBHy-agar (corn meal agar amended with 250 μg ml^-1 ampicillin, 10 μg ml^-1 rifampicin, 10 μg ml^-1 pimaricin, 15 μg ml^-1 benomyl, 50 μg ml^-1 hymexazol), and incubated at 20°C. Ten morphologically similar isolates were subcultured as single hyphal tips and characterized. These isolates produced nonpapillate sporangia that were ovoid, obpyriform or ellipsoid, and exhibited predominately internal proliferation in soil extract. Hyphal swellings with outgrowths were present under those conditions. The morphological characteristics were consistent with those of P. cambivora (1). Base sequences of the ITS region of rDNA were determined for six of the isolates, and an 832-bp fragment was amplified for each isolate and that sequence was 100% homologous with sequences DQ396418 and AY880985 of P. cambivora in the NCBI database ( http://www.ncbi.nlm.nih.gov/BLAST/ ). The sequence of one isolate, ISPaVe 1950, was deposited in GenBank (Accession No. AM269752). Pathogenicity tests were conducted with 2-year-old potted beech seedlings. Inoculum of representative isolates was grown for 4 weeks on sterilized millet seeds moistened with V8 broth and added to soil at 3% (wt/vol). Control plants received sterilized inoculum only. The soil was flooded for 48 h. Inoculations were performed during May 2005 at 15 to 35°C with six replicates for the inoculated and control plants. The plants were maintained outdoors and assessed after 3 months. Wilt, root rot, and dark brown lesions at the collar developed on inoculated plants, but not on the controls. Symptoms were similar to those on naturally infected trees. The pathogen reisolated from the inoculated plants was morphologically identical to the original isolates, which confirmed P. cambivora as the causal agent. To our knowledge, this is the first report of P. cambivora on beech in Italy. References: (1) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN, 1996. (2) T. Jung et al. Mycologist, 19:159, 2005. (3) E. Motta et al. Plant Dis. 87:1005, 2003.

Zucchini (Cucurbita pepo) is intensively cropped in approximately 4,500 ha of plastic houses in southern Spain. In 2008 to 2009, Consul, Cronos, and Tosca zucchini plants showed symptoms of leaf wilting, basal stem necrosis, and plant death. Incidences of dead plants were 20 to 30% and these plants were distributed in clusters. Phytophthora capsici Leonian was isolated from the basal stems of symptomatic plants, using PDA and cornmeal agar amended with a pimaricin, ampicillin, and rifampicin. Five resultant isolates (PCl-211, PCl-221, PCl-611, PCl-612, and PCl-811) on lima beans agar (LBA) produced white mycelia with lemon-shaped and papillate sporangia borne on long pedicels, but no oospores or chlamydospores. These isolates had an identical ribosomal DNA ITS sequence, matching with that of P. capsici in GenBank. The sequences of two representative isolates, PCl-211 and PCl-811, were deposited in GenBank with accession nos. KC662328 and KC688317, respectively. The pathogenicity of these five isolates was tested on zucchini cv. Consul in 1-liter containers filled with vermiculite in May and September of 2009. Plants were inoculated at the 2 to 3 true-leaf stage. Plates with LBA fully covered with colony of each isolate were separately blended and homogenized with 300 ml of sterile distilled water. Inocula were poured around stem at 50 ml per plant. Each experiment had three replicates and four plants per replicate. Treatments with different isolates were arranged in a randomized complete block design. In both experiments, 12 uninoculated plants served as controls. Test plants were maintained for a month following inoculation in a greenhouse with mean temperatures ranging from 21.9 to 27.9°C and from 20.7 to 24.6°C for the May and September experiments, respectively. The first wilting occurred 5 days after inoculation. At the end of the May experiment, all control plants and those inoculated with PCl-221 remained asymptomatic while 83.3% of those inoculated with PCl-211 and 100% of those with the other isolates were dead. Inoculated plants exhibited crown and root rots, excluding the secondary roots. In the September experiment, 83.3% and 33.3% of plants inoculated with PCl-211 and PCl-221, respectively, were symptomatic, while all plants inoculated with the other isolates were dead. The control plants remained healthy. The pathogen was consistently recovered from symptomatic plants in both experiments. Although P. capsici was reported in peppers (Capsicum annuum) in several provinces of Spain (1), to our knowledge, this is the first report of P. capsici as the causal agent of crown rot in zucchini plants in plastic houses in the Almería Province of Spain, one of the world's largest concentrations of greenhouses. Reference: (1) J. L. Andrés et al. Span J Agric Res 3:326, 2005.

During 2006, spears, roots, and crowns of asparagus (Asparagus officinalis) exhibiting brown necrotic lesions with water soaking were collected from several sites across Peru (Ica, Lima, and Trujillo). Small infected tissue sections were washed thoroughly with tap and sterile distilled water and transferred to corn meal agar plates (CMA) amended with PARP (100 ppm of pimaricin, 100 ppm of ampicillin, 30 ppm of rifampicin, and 100 ppm of pentachloronitrobenzene) and incubated for five days at 25°C. Hyphal tips were subcultured from actively expanding mycelium. Sporangia produced on CMA were papillate and averaged 38 μm long × 29 μm wide. Chlamydospores were terminal or intercalary and averaged 35 μm in diameter. Isolates incubated in the dark for more than 3 weeks did not produce oospores in single culture. Mating with Phytophthora capsici tester isolates CBS 121656 = A1 and CBS 121657 = A2 indicate that all five isolates were A2. For pathogenicity tests, inoculum was generated by incubating 300 g of autoclaved wheat seeds with four agar plugs (7 mm) of expanding mycelium in polyethylene bags for 1 month at 25°C. Nine-week-old asparagus plants (UC151 F1) were transferred into pots containing autoclaved substrate (1 part sand, 1 part potting soil, and 1 part peat). Inoculum was added as 1 g of inoculum per kilogram of substrate. Plants were maintained in the greenhouse at 23°C and watered daily. Decline symptoms as well as root and spear rot were observed after 7 days and a Phytophthora sp. was reisolated from infected tissue. No symptoms were observed on asparagus plants inoculated with sterile inoculum. DNA was isolated from two representative isolates, and the nuclear ribosomal internal transcribed spacer (ITS) region was amplified with ITS4 and ITS6 primers and sequenced. ITS sequence was submitted for a BLAST search in the NCBI database, showing Phytophthora nicotianae strain UQ848 Accession No AF266776 as the closest match with 99% sequence similarity (1). The consensus ITS sequence was deposited in NCBI (Accession No. EU433396). These results, together with the morphological characteristics, indicate that the Phytophthora sp. isolated from asparagus in Peru is P. nicotianae (Breda de Haan) (2). To our knowledge, this is the first report of P. nicotianae infecting asparagus and represents a new threat for asparagus growers in Peru. Control methods such as moderate watering and metalaxyl application are being applied to reduce Phytophthora outbreaks. References: (1) D. E. Cooke et al. Fungal Genet. Biol. 30:17, 2000. (2) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society. St Paul, MN, 1996.

The S-adenosylmethionine: delta 24-sterol methyltransferase (24 SMT) primarily considered as a mitochondrial enzyme, was recently mainly detected in lipid particles of yeasts. It catalyses the methylation of zymosterol which is an essential reaction for the synthesis of ergosterol. We have investigated in cellular extracts of two Kluyveromyces lactis strains the action of polyenic antifungal agents on the activity of this enzyme. Low concentrations of amphotericin B, candicidin and pimaricin strongly stimulate this activity, while high concentrations inhibit it or have no effect. Whatever the doses used, nystatin and filipin had no significant influence on this activity. According to the molar ratio amphotericin B/total sterols of the enzyme preparation, the interference of amphotericin B on the 24 SMT activity may result of two mechanisms.

In spring 2012, maize farmers in southeast and south central Iowa reported stand losses due to pre- and post-emergence damping-off, and many of the fields had to be replanted. Symptoms of the disease included rotted seed, or brown, rotted, water-soaked mesocotyls and root tips. Maize seedlings with severe root and mesocotyl symptoms were yellow and wilted, stunted, or dead. The disease occurred approximately 2 weeks after cool, wet conditions. Symptomatic mesocotyls and roots were washed for 30 min, rinsed with sterile distilled water, and blotted dry on sterile paper towels. Isolation of the pathogen was performed by aseptically cutting 2- to 3-mm sections of tissue from the edge of a lesion, placing the segments under corn meal agar (CMA) containing pimaricin, ampicillin, rifampicin, and pentachloronitrobenzene (PARP), and incubating at 22°C in the dark. Colonies that developed were putatively identified as Pythium species based on morphological characteristics and cultural features when compared to published descriptions (2,3). Characteristics of isolate IAC12F21-3 included spherical and smooth-walled oogonia 18 to 26 μm in diameter, monoclinous or usually diclinous antheridia 10 to 22 μm long and 5 to 10 μm wide with one or occasionally two per oogonium, and plerotic oospores 15 to 25 μm in diameter. Sporangia were globose to ellipsoidal, 22 to 41 μm in diameter, and zoospores were 7 to 10 μm long. Primers ITS1 and ITS4 were used to amplify the ITS region within clade E1 of 88 isolates. The resultant amplicons were sequenced and a BLAST search in GenBank confirmed isolate IAC12F21-3 as Pythium schmitthenneri based on 100% similarity with GenBank accession numbers JF836869 and JF836870. Pathogenicity testing was conducted using seed and seedling assays (1,4). Koch's postulates was performed by sampling pieces of symptomatic mesocotyl and root tissue from the inoculated pots, placing segments under CMA + PARP, and incubating at 22°C. Symptoms were similar to those observed in the field and P. schmitthenneri was re-isolated successfully. Non-inoculated control plants showed no symptoms. This is the first report of P. schmitthenneri causing seedling blight on maize in Iowa. Previously, P. schmitthenneri was reported as a pathogen on maize in Ohio (2). References: (1) K. Broders et al. Plant Dis. 91:727, 2007. (2) M. Ellis et al. Mycologia, 104:477, 2012. (3) J. Middleton. Memoirs of the Torrey Botanical Club 20:171, 1943. (4) A. Rojas et al. Phytopathology, 102(Suppl):S5.8, 2012.

In connection with a case report, the mycological diagnosis and treatment of orbital aspergillosis are reviewed. In a 59-year-old male patient an exophthalmus on the right side could be diagnosed as aspergillosis spreading from the corresponding sinus maxillaris into retro- and peri-orbicular regions, after other causes had been excluded by differential diagnosis. In a biopsy specimen from the retrobulbar tumor, a fungal granuloma was found which, histologically and by isolation of the fungus, could be identified as an infection by A. fumigatus. Because of the good visus of the right eye, no exenteratio orbitae but an enoral revision of the sinus maxillaris, ethmoidal bone, and bottom of orbita was performed.--By infusions of amphotericin B and local washings with pimaricin (natamycin) in the sinus maxillaris and the region of the operation, a complete healing of the mycotic process was achieved.--Since 1 1/2 years, the patient has not had a relaps. This success of therapy was possible by exemplary cooperation of otorhinolaryngologists, ophthalmologists, pathologists and microbiologists.--In view of the prophylaxis of such infections, recent contributions to the epidemiology of aspergillosis are discussed.

During a survey of pathogenic oomycetes in Nanjing, China from June 2019 to October 2020, at least ten adjacent Rhododendron pulchrum plants at a Jiangjun Mountain scenic spot showed symptoms of blight, and crown and root discoloration . Symptomatic root tissues collected from three 6-year-old plants were rinsed with water, cut into 10-mm pieces, surface sterilized with 70% ethanol for 1 min, and plated onto 10% clarified V8 PARP agar (cV8A-PARP) containing pimaricin (20 mg/liter), ampicillin (125 mg/liter), rifampicin (10 mg/liter), and pentachloronitrobenzene (20 mg/liter). Four Pythium-like isolates were recovered after three days of incubation at 26°C, and purified using hyphal-tipping. Ten agar plugs (2×2 mm2) of each isolate were grown in 10 mL of 10% clarified V8 juice (cV8) in a 10 cm plate at 26°C for 3 days to produce mycelial mats, and then the cV8 was replaced with sterile water. To stimulate sporangial production, three to five drops of soil extract solution were added to each plate. Sporangia were terminal, ovoid to globose, and the size is 24 to 45.6 (mean 34.7) (n=10.8) in length x 23.6 to 36.0 (mean 29.8) (n=6.2) in width. Gametangia were not observed in cV8A or liquid media after 30 days. For colony morphology, the isolates were sub-cultured onto three solid microbial media (cV8A-PARP, potato dextrose agar, corn meal agar) . All isolates had identical morphological features in the three media. Complete ITS and partial LSU and cox2 gene regions were amplified using primer pairs ITS1/ITS4, NL1/NL4, and FM58/FM66 , respectively. The ITS, LSU, and cox2 sequences of isolate PC-dj1 (GenBank Acc. No. MW205746, MW208002, MW208003) were 100.00% (936/936 nt), 100.00% (772/772 nt), and 99.64% (554/556 nt) identical to those of JX985743, MT042003, and GU133521, respectively. We built a maximum-likelihood tree of Phytopythium species using the concatenated dataset (ITS, LSU, cox2) to observe interspecific differences. Based on the morphological characters and sequences, isolate PC-djl was identified as Phytopythium litorale . As the four isolates (PC-dj1, PC-dj2, PC-dj3 and PC-dj4) tested had identical morphological characters and molecular marker sequences, the pathogenicity of the representative isolate, PC-dj1, was tested using two inoculation methods on ten one-year-old R. pulchrum plants. For the first inoculation method, plants were removed from the pot, and their roots were rinsed with tap water to remove the soil. Each of these plants was placed in a glass flask containing 250 mL of sterile water and 10 blocks (10 x 10 mm2) of mycelial mats harvested from a three-day-old culture of P. litorale, while the other plant was placed in sterile water as a control, and incubated at 26°C. After three days, symptoms including crown rot, root rot and blight was observed on the inoculated plants whereas the control remained asymptomatic. For the second inoculation method, ten plants were dug up to expose the root ball. Ten three-day-old cV8A plugs (5×5 mm2) from a PC-dj1 culture or sterile cV8A plugs were evenly insert into the root ball of a plant before it was planted back into the original pots. Both plants were maintained in a growth chamber set at 26°C with a 12/12 h light/dark cycle and irrigated as needed. After 14 to 21 days, the inoculated plant had symptoms resembling those in the field , while the control plant remained asymptomatic. Each inoculation method was repeated at triplicate and the outcomes were identical. Phytopythium isolates with morphological features and sequences identical to those of PC-dj1 were recovered from rotted crown and root tissues of all inoculated plants. Previously, P. litorale was found causing diseases of apple and Platanus orientalis in Turkey, fruit rot and seedling damping-off of yellow squash in southern Georgia, USA. This is the first report of this species causing crown and root rot on R. pulchrum, an important ornamental plant species in China. Additional surveys are ongoing to determine the distribution of P. litorale in the city of Nanjing.

Keywords: Causal Agent; Crop Type; Epidemiology; Oomycetes; Subject Areas; Trees; disease development and spread; forest.

Antitumor activity of bleomycin against Ehrlich ascites carcinoma in mice was examined in the presence and absence of a polyene antibiotic, pimaricin. A single intraperitoneal (ip) injection of bleomycin and pimaricin in combination into tumor-bearing mice on day 1 produced a synergistic life-prolongation effect as compared with that of each agent alone. Accumulation of abdominal ascites was almost completely blocked by bleomycin in combination with the tetraene antibiotic. Frequent ip injection of small doses of bleomycin and pimaricin on day 1 produced a much greater increase in life span than the single ip injection. To obtain effective synergism under these treatment schedules, it appeared that the dose of bleomycin should be reciprocally proportional to the dose of pimaricin.

Sugar beet (Beta vulgaris L.) is a globally important crop for sugar. In May 2019, sugar beet seedlings were observed with wilting, lodging and a few were dead in Billings (45.7833° N, 108.5007° W), Montana. Symptoms appeared near the soil line as the stem (hypocotyl) turned dark brown to black with characteristic thread-like infections which resembled Pythium root rot. It affected approximately 10% of the growing seedlings. Diseased sugar beet root tissues were excised with a sterile scalpel and small pieces (10 mm²) were surface sterilized with 70 % ethanol for 30 seconds, rinsed twice with autoclaved water, air-dried and transferred to potato dextrose agar (PDA) media amended with pimaricin-vancomycin-PCNB (Conway, 1985). Four plates were incubated at 25° C in the dark (Masago et al., 1977) and two weeks later white, dense colony was observed (Zhang et al., 2018). The terminal smooth, globose oogonia (average 18.5 µm in diameter) and antheridia (average 14.5 × 9.5 µm) extended below the oogonium were observed via VWR N. A. 0.30 microscope. The morphological features of the four isolates were consistent with Pythium ultimum Trow (Watanabe, 2002). Genomic DNAs (NORGEN BIOTEK CORP, Fungi DNA Isolation Kit #26200) of four isolates were used for polymerase chain reaction (PCR) with the ITS6-ITS7 primers (Taheri et al., 2017). Subsequently, PCR products were flushed by E.Z.N.A ®Cycle Pure Kit, OMEGA and four samples were sent for Sanger sequencing to GenScript (GenScript, Piscataway, NJ). The sequences were identical and submitted to GenBank, NCBI (accession no. MN398593). The NCBI Blast analysis showed 100% sequence homology to Pythium ultimum with the following GenBank accessions; KF181451.1, KF181449.1 and AY598657.2. Pathogenicity test was done on sugar beet with the same isolates in the greenhouse. Two week old, pythium culture was mixed with vermiculite and perlite mixer (PRO-MIX FLX) in the plastic trays (24´´ x 15´´× 3˝), (22 °C, 75% Relative Humidity). Sterile water (500 ml/each tray) was added in the mixer to keep the sufficient moisture. Twenty seeds of cv. Hilleshog 4302 were sown in the tray, and the trays were replicated thrice with inoculated and mock treatments. Plants were watered as needed to maintain adequate soil moisture conducive for plant growth and disease development. Seven days of sowing, 50% and 100% germination was observed in the inoculated and control treatments, respectively. At the beginning after the second week, 30% post-emergence damping-off was observed in the inoculated treatments. Diseased seedlings were gently pulled out from the pots where similar infection signs were observed in the sugar beet seedlings as described previously. No incidence of disease was observed in mock-treated seedlings. Consistent reisolation of Pythium ultimum was morphologically and molecularly confirmed from the diseased seedlings, thus fulfilling Koch's postulates. Pythium spp identification is prerequisite to develop effective management of pre and post-emergence damping-off. Pythium ultimum was previously reported in Nebraska to cause sugar beet seed rot and pre-emergence damping-off (Harvenson 2006). To our knowledge, this is the first report of Pythium ultimum causing damping-off on sugar beet in Montana.

Keywords: Pythium ultimum; Sugar beet; damping-off.