The avocado subgroup of Phytophthora citricola constitutes a distinct species, Phytophthora mengei sp. nov.

Isolates from avocado tree cankers have been recognized as a distinct subgroup within the P. citricola complex since 1974, both morphologically and molecularly (isozyme and amplified fragment length polymorphism [AFLP] analyses). This subgroup is formally separated from P. citricola after comparative DNA fingerprinting and sequence analyses of the ITS region, as well as by morphological examinations. This new taxon is homothallic, produces plerotic oospores with paragynous antheridia and noncaducous semipapillate sporangia. Morphologically it differs from other species of Waterhouse group III by producing many large bizarre-shaped sporangia and smaller oogonia with asymmetric capitate antheridia. It belongs to clade 2 and is phylogenetically closer to P. siskiyouensis, P. capsici and P. tropicalis than to P. citricola. P. mengei can be easily differentiated from its relatives in the same clade and other species of this morpho-group by DNA fingerprints and sequence analysis. This new taxon is named Phytophthora mengei sp. nov.

Genetic Diversity in Populations of Phytophthora citricola Associated with Horticultural Crops in California.

California populations of the plant pathogen Phytophthora citricola were examined for amplified fragment length polymorphism (AFLP), pathogenicity on almond, and sensitivity to mefenoxam. The characterizations of AFLP variation and mefenoxam sensitivity were based on 86 isolates (44 from almond, 11 from avocado, 3 from strawberry, 18 from walnut, and 10 from six other hosts). Cluster analysis of the AFLP data using the unweighted pair group method indicated a high level of genetic diversity among the isolates, and four main clusters were identified-one dominated by isolates from almond, another including all isolates from avocado, and two including isolates from several hosts other than avocado. Analysis of molecular variance revealed that 38.4 and 24.9% of the AFLP variation were associated with host and geographical factors, respectively. Of 24 isolates, including those from almond, avocado, strawberry, and walnut, 22 were aggressive on almond shoots; there was no evidence of host specificity. All but 1 of the 86 isolates grew at different rates on V8 juice medium amended with mefenoxam at 1 ppm, indicating partial tolerance to the fungicide. Isolates of P. citricola from California populations are genetically diverse, and much of the variation is associated with host and geography. These populations are all potentially pathogenic on almond and tolerant to mefenoxam.

Genetic and Pathogenic Variation in Phytophthora cactorum Affecting Fruit and Nut Crops in California.

Isolates of Phytophthora cactorum and 15 other species of Phytophthora were characterized according to their genomic DNA, pathogenicity, and sensitivity to mefenoxam. Amplified fragment length polymorphism (AFLP) analysis was completed for 132 isolates of P. cactorum (30 from almond, 86 from strawberry, 5 from walnut, and 11 from other hosts) and 22 isolates of 15 other Phytophthora spp. from various hosts. All 16 Phytophthora spp. were distinguishable by unique AFLP banding patterns. Cluster analysis of the AFLP data revealed high coefficients of genetic similarity (>0.9) among all California isolates of P. cactorum. Analysis of molecular variance indicated that, among all 132 isolates of P. cactorum, 30.8 and 24.5% of the AFLP variation was associated with hosts and geographical sources of isolates, respectively, whereas 15.0% of the variation was associated with isolate niche (i.e., an aerial plant part, portion of the root system, or soil). Among the 86 isolates of P. cactorum from strawberry, characterization by source in the production system (i.e., fruiting field or plant nursery) did not account for a significant proportion of the variation (0.6%, P = 0.204). In pathogenicity tests on strawberry plants (cv. Diamante) in a greenhouse, isolates of P. cactorum from hosts other than strawberry and an isolate from a strawberry fruit caused only negligible amounts of disease, but isolates from strawberry root systems were highly aggressive. On excised shoot segments of almond (cv. Drake), all isolates of P. cactorum originally from almond were pathogenic, and 8 of 17 isolates of the pathogen from other hosts caused significantly less disease than the almond isolates. All 132 isolates of P. cactorum were sensitive to mefenoxam at 1 ppm. Populations of P. cactorum in California apparently are mefenoxam sensitive and exhibit host specificity with relatively minor variation in genomic DNA. The genetic variation observed in P. cactorum included significant geographical and host origin components, which has implications for disease management approaches.

Phytophthora Leaf Spot and Foliar Blight of Pieris japonica Caused by Phytophthora citricola in California.

During 2004, containerized nursery stock of lily-of-the-valley-bush (Pieris japonica 'Flamingo', family Ericaceae) in Santa Cruz County was affected by a foliar disease. Symptoms consisted of large leaf spots, many developing at the leaf tips that ranged in size from 1 to greater than 4 cm in diameter. Spots were dark brown to almost black, generally oval to round, visible from both sides of the leaf, and did not exhibit signs of any pathogen. Lesions typically expanded and affected the entire leaf, leaf petiole, and stems, resulting in blight-like symptoms. Severely affected leaves abscised from the plant. In advanced stages of the disease, the foliage of the plant was killed. These symptoms resembled those caused by the sudden oak death (SOD) pathogen, Phytophthora ramorum (3). A Phytophthora sp. was isolated consistently from symptomatic leaf tissue. However, the species was identified as P. citricola based on morphological traits that included the following: production of semipapillate, noncaducous sporangia that were irregular in shape and occasionally had more than one apex; presence of oospores with paragynous antheridia in single culture; and radiate to slightly petaloid colony morphology (1). P. ramorum and other fungi were not recovered. Pathogenicity of four representative isolates was confirmed by gently abrading the adaxial surfaces of attached leaves with a sterile wire brush, placing a colonized agar plug (5 mm in diameter) on the surface, misting the leaf with sterile water, and then covering the plug with a plastic cap that was secured with a wire clip. Control leaves were treated in the same manner but received sterile agar plugs. Plants were maintained in a greenhouse at 23 to 25°C. After 2 days, all leaves inoculated with the isolates exhibited dark brown lesions and by day 6, lesions measured 3 cm in diameter. P. citricola was reisolated from symptomatic lesions. Sterile plug control leaves developed no symptoms. The test was repeated and the results were similar. To our knowledge, this is the first report of P. citricola causing a foliar disease of Pieris japonica in California. P. citricola has been reported as a pathogen on Pieris spp. in Ohio (2). Our finding is important because P. ramorum causes very similar symptoms on this same host (3). The occurrence of these two foliar Phytophthora spp. on this ornamental plant may complicate P. ramorum field detection during inspections and laboratory confirmations as established by quarantine regulations. References: (1) D. C. Erwin and O. K. Ribeiro. Morphology and Identification of Phytophthora Species. Pages 96-144 in: Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN, 1996. (2) W. W. P. Gerlach et al. Phytopathology 64:1368, 1974. (3) P. W. Tooley et al. Plant Dis. 88:993, 2004.

Comparison of Media for Recovery of Verticillium dahliae from Soil.

Polygalacturonic acid (PGA) is an important constituent of Sorensen's NP-10 medium (NP-10) for estimating the population density of Verticillium dahliae in soil. Different types of PGA are available, but not all of them favor the growth of V. dahliae. Unavailability of PGA sodium salt from orange (P-1879) has created an unprecedented problem for the quantification of microscle-rotia (MS) of V. dahliae in soil. The PGA from orange (P-3889) that is now available does not support the growth of V. dahliae. Therefore, experiments were conducted to optimize the use of NP-10 prepared with P-3889 and various concentrations of NaOH. NP-10 with P-3889 amended with eight concentrations of NaOH were compared with NP-10 prepared from PGA sodium salt from orange (P-1879, now discontinued) and citrus (P-3850) along with cellophane and Na-pectate media for recovery of MS from soil and growth of V. dahliae on the media. Seven soils were assayed for MS, and eight isolates of V. dahliae were evaluated for growth and production of MS. Concentrations of NaOH >0.035N and <0.02N in NP-10 with P-3889 reduced mycelial growth, microsclerotial production, and recovery of MS from soils. Similarly, NP-10 with P-3850 alone, cellophane, and Na-pectate media had significantly reduced growth on media and recovery of V. dahliae from soils. The NP-10 with P-3889 and 0.025N NaOH consistently yielded numbers of V. dahliae MS from soil samples and supported the growth and production of MS similar to the NP-10 with P-1879. The medium developed in this study can serve as a direct replacement for the original NP-10 that was developed nearly three decades ago, an important component of which is no longer available.

Characterization of Verticillium dahliae Isolates and Wilt Epidemics of Pepper.

Epidemics of Verticillium wilt in pepper fields of the central coast of California and isolates of Verticillium dahliae associated with these epidemics were characterized. The mean incidence of wilted plants per field ranged from 6.3 to 97.8% in fields with Anaheim, jalapeno, paprika, or bell peppers. In general, incidence of wilt in jalapeno and bell pepper crops was lower than in crops of other types of pepper. Inoculum density of V. dahliae in the surveyed pepper fields ranged from 2.7 to 66.6 microsclerotia g-1 dry soil, and the correlation between disease incidence and density of microsclerotia was high (r = 0.81, P < 0.01). Distribution of Verticillium wilt was aggregated in a majority of the pepper fields surveyed, but the degree of aggregation varied. Vegetative compatibility group (VCG) characterization of 67 isolates of V. dahliae indicated that 67% belonged to VCG 2, 22% to VCG 4, and 11% to a new group, designated VCG 6. The pathogenicity of isolates of V. dahliae from bell pepper and tomato plants was tested by inoculating 1-month-old bell pepper (cv. Cal Wonder) and tomato (cv. EP 7) seedlings and incubating the inoculated plants in the greenhouse. Seedlings of bell pepper were susceptible only to the isolates of V. dahliae from pepper, whereas seedlings of tomato were susceptible to both pepper and tomato isolates. Pepper isolates belonging to VCG 2, VCG 4, and VCG 6 were highly pathogenic to bell pepper and chili pepper. Temperatures between 15 and 25°C were optimal for mycelial growth of a majority of isolates of V. dahliae. Molecular characterization of pepper isolates of V. dahliae using a polymerase chain reaction (PCR)-based random amplified polymorphic DNA (RAPD) technique revealed minor variation among these isolates, but unique polymorphic banding patterns were observed for isolates belonging to VCG 6. Verticillium wilt of pepper is a major production constraint in the central coast of California. More aggressive isolates of V. dahliae may have been selected in this region as a result of intensive cropping practices.

Reaction of Broccoli to Isolates of Verticillium dahliae from Various Hosts.

Isolates of Verticillium dahliae from 15 different hosts and V. albo-atrum from alfalfa were tested for their ability to cause wilt on broccoli using a root-dip inoculation method. None of the isolates caused vascular discoloration in broccoli except those from cabbage and cauliflower that were weakly pathogenic. Broccoli cultivars Baccus, Greenbelt, Parasol, Patriot, and Symphony showed resistance to Verticillium infection. Re-isolated strains from fresh samples of internally discolored broccoli and cauliflower root tissues were unable to cause disease symptoms on re-inoculation of broccoli, but caused severe disease on cauliflower. Inoculation of 5-, 7-, 9-, or 11-week-old plants did not alter the resistance in broccoli or the susceptibility in cauliflower against Verticillium. Immunity of broccoli against infection by isolates of V. dahliae from non-crucifer hosts, its resistance against crucifer isolates, and as previously described, attrition of V. dahliae microsclerotia in soil by broccoli residue, coupled with its importance as a commercial vegetable, make broccoli an attractive rotation crop for the management of Verticillium wilt in many cropping systems.

Mathemycin B, a new antifungal macrolactone from actinomycete species HIL Y-8620959.

A new macrocyclic lactone antibiotic mathemycin B (1) was isolated from the fermentation broth of an Actinomycete sp. culture Y-8620959. The structure of 1 was elucidated by high-resolution MS and interpretation of 2D NMR results. Mathemycin B is active against a variety of phytopathogenic organisms.

Host Range Specificity in Verticillium dahliae.

ABSTRACT Verticillium dahliae isolates from artichoke, bell pepper, cabbage, cauliflower, chili pepper, cotton, eggplant, lettuce, mint, potato, strawberry, tomato, and watermelon and V. albo-atrum from alfalfa were evaluated for their pathogenicity on all 14 hosts. One-month-old seedlings were inoculated with a spore suspension of about 10(7) conidia per ml using a root-dip technique and incubated in the greenhouse. Disease incidence and severity, plant height, and root and shoot dry weights were recorded 6 weeks after inoculation. Bell pepper, cabbage, cauliflower, cotton, eggplant, and mint isolates exhibited host specificity and differential pathogenicity on other hosts, whereas isolates from artichoke, lettuce, potato, strawberry, tomato, and watermelon did not. Bell pepper was resistant to all Verticillium isolates except isolates from bell pepper and eggplant. Thus, host specificity exists in some isolates of V. dahliae. The same isolates were characterized for vegetative compatibility groups (VCGs) through complementation of nitrate nonutilizing (nit) mutants. Cabbage and cauliflower isolates did not produce nit mutants. The isolate from cotton belonged to VCG 1; isolates from bell pepper, eggplant, potato, and tomato, to VCG 4; and the remaining isolates, to VCG 2. These isolates were also analyzed using the random amplified polymorphic DNA (RAPD) method. Forty random primers were screened, and eighteen of them amplified DNA from Verticillium. Based on RAPD banding patterns, cabbage and cauliflower isolates formed a unique group, distinct from other V. dahliae and V. albo-atrum groups. Minor genetic variations were observed among V. dahliae isolates from other hosts, regardless of whether they were host specific or not. There was no correlation among pathogenicity, VCGs, and RAPD banding patterns. Even though the isolates belonged to different VCGs, they shared similar RAPD profiles. These results suggest that management of Verticillium wilt in some crops through crop rotation is a distinct possibility.

First Report of Verticillium dahliae Causing Artichoke Wilt in California.

In mid-August 1998, artichoke (Cynara scolymus L.) plants of cultivar Imperial Star in a field in the Salinas area of the central coast of California developed wilt symptoms. The plants were stunted with chlorotic, drooping, and dried leaves near the bottom and middle of the plants as previously described in Italy (1). Diseased plants produced smaller edible buds and, in severe cases, buds were discolored with dried outer bracts. Roots exhibited the characteristic vascular discoloration of Verticillium infection. In one part of the infested field, artichoke was near harvest with 85% of plants showing wilt symptoms with vascular discoloration whereas the other part had a 60-day-old crop with 98% of plants infected. Yield in the field was reduced by as much as 50%. Verticillium dahliae was isolated from infected plant samples on NP-10 medium (2), and isolates were single spored before storing on potato dextrose agar at 4°C. Identity of the pathogen was confirmed based on colony morphology and formation of microsclerotia. In root-dip inoculation tests in the greenhouse, two V. dahliae isolates from artichoke infected 1-month-old artichoke seedlings that wilted within 6 weeks of inoculation. V. dahliae was reisolated from plants showing vascular discoloration. Cross-inoculation studies revealed that artichoke isolates caused a moderate level of disease in lettuce, but only a trace of vascular discoloration in cauliflower. Lettuce isolates caused a severe wilt in artichoke. Cauliflower isolates did not cause wilt in lettuce and caused only slight vascular discoloration in artichoke. This is the first report of a Verticillium wilt of artichoke in California. References: (1) M. Cirulli et al. Plant Dis. 78:680, 1994. (2) L. H. Sorensen et al. Phytopathology 81:1347, 1991.

Thermodynamics of monosaccharide binding to concanavalin A, pea (Pisum sativum) lectin, and lentil (Lens culinaris) lectin.

Titration calorimetry measurements of the binding of methyl alpha-D-mannopyranoside (Me alpha Man), D-mannopyranoside (Man), methyl alpha-D-glucopyranoside (Me alpha Glu), and D-glucopyranoside (Glu) to concanavalin A (Con A), pea lectin, and lentil lectin were performed at 281 and 292 K in 0.01 M dimethylglutaric acid-NaOH buffer (pH 6.9) containing 0.15 M NaCl and Mn+2 and Ca+2 ions. The site binding enthalpies, delta H, are the same at both temperatures and range from -28.4 +/- 0.9 (Me alpha Man) to -16.6 +/- 0.5 kJ mol-1 (Glu) for Con A, from -26.2 +/- 1.1 (Me alpha Man) to -12.8 +/- 0.4 kJ mol-1 (Me alpha Glu) for pea lectin, and from -16.6 +/- 0.7 (Me alpha Man) to -8.0 +/- 0.2 kJ mol-1 (Me alpha Glu) for lentil lectin. The site binding constants range from 17 +/- 1 x 10(3) M-1 (Me alpha Man to Con A at 281.2 K) to 230 +/- 20 M-1 (Glu to lentil lectin at 292.6 K) and exhibit high specificity for Con A where they are in the Me alpha Man:Man:Me alpha Glu:Glu ratio of 21:4:5:1, while the corresponding ratio is 5:2:1.5:1 for pea lectin and 4:2:2:1 for lentil lectin. The higher specificity for Con A indicates more interactions between the amino acid residues at the binding site and the carbohydrate ligand than for the pea and lentil lectin-carbohydrate complexes. The carbohydrate-lectin binding results exhibit enthalpy-entropy compensation in that delta Hb (kJ mol-1) = -1.67 +/- 0.06 x 10(4) + (1.30 +/- 0.12)T(K) delta Sb (J mol-1K-1). Differential scanning calorimetry measurements on the thermal denaturation of the lectins and their carbohydrate complexes show that the Con A tetramer dissociates into monomers, while the pea and lentil lectin dimers dissociate into two submonomer fragments. At the denaturation temperature, one carbohydrate binds to each monomer of Con A and the pea and lentil lectins. Complexation with the carbohydrate increases the denaturation temperature of the lectin and the magnitude of the increases yield binding constants in agreement with the determinations from titration calorimetry.

Aranorosinol A and aranorosinol B, two new metabolites from Pseudoarachniotus roseus: production, isolation, structure elucidation and biological properties.

Two new secondary metabolites, aranorosinol A (1) and aranorosinol B (2), were isolated from a strain of Pseudoarachniotus roseus. Their structures were elucidated on the basis of their spectral properties and chemical transformations and were found to be similar to aranorosin (3) isolated from the same strain.

Deoxymulundocandin--a new echinocandin type antifungal antibiotic.

A new echinocandin type antifungal antibiotic, deoxymulundocandin, C48H77N7O15, was isolated from the culture filtrate and mycelia of a fungal culture, Aspergillus sydowii (Bainier and Sartory) Thom and Church var. nov. mulundensis Roy (Culture No. Y-30462). The structure was established by comparative GC-MS analyses of the derivatized acid hydrolysates of deoxymulundocandin and mulundocandin as well as by the high field NMR experiments (COSY, NOESY and DEPT).