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.

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Verticillium dahliae is an important soil-borne fungal pathogen that causes vascular wilt diseases in a large variety of important crop plants. Due to its persistence in the soil, control of Verticillium wilt relies heavily on soil fumigation. The global ban on methyl bromide, a highly effective soil fumigant, poses an urgent need to develop alternative control measures against Verticillium wilt; and these might be more forthcoming with a better understanding of the molecular and cellular mechanisms that underpin the pathogenicity of V. dahliae. In this study, we assessed the role in growth, development, and pathogenicity of VMK1, a gene encoding a mitogen-activated protein (MAP) kinase (hence, Verticillium MAP Kinase 1). Disruption of VMK1 via Agrobacterium tumefaciens-mediated transformation, in two V. dahliae isolates, one from lettuce and the other from tomato, resulted in severely reduced virulence in diverse host plants, suggesting that VMK1 is essential for pathogenicity and that the MAP kinase-mediated signaling pathway has a conserved role in fungal pathogenicity. The vmk1 mutants also exhibited reduced conidiation and microsclerotia formation, suggesting that the gene is important for multiple cellular processes.

Bhat, R. G., Smith, R. F., Koike, S. T., Wu, B. M., and Subbarao, K. V. 2003. Characterization of Verticillium dahliae isolates and wilt epidemics of pepper. Plant Dis. 87:789-797.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.

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 microsclerotia (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 Napectate 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.

The seed transmission of Verticillium dahliae was evaluated in lettuce (Lactuca sativa). Seed collected from lettuce plants infected with V. dahliae were plated with or without surface sterilization on Sorenson's modified NP10 medium. Of the seed plated with or without surface sterilization, 90 and 66%, respectively, yielded colonies of V. dahliae. The incidence of Verticillium wilt ranged from 55 to 80% among lettuce plants grown from seed harvested from infected plants. All evaluated isolates of V. dahliae were capable of seed transmission in lettuce. A V. tricorpus isolate failed to cause significant disease in lettuce or to become seedborne. Storage of contaminated seed at seven temperatures ranging from -20 to 15°C for up to 72 weeks did not reduce the incidence of V. dahliae in seed, whereas storage at room temperature (23 ± 2°C) for 20 to 52 weeks reduced the incidence of V. dahliae without affecting seed viability. Of the 11 weed species collected from fields with a known history of Verticillium wilt of lettuce, four yielded V. dahliae. Pathogenicity tests demonstrated that isolates of V. dahliae from Sonchus oleraceus, Capsella bursa-pastoris, and Solanum sarrachoides were as virulent as or more virulent than an isolate of V. dahliae from lettuce. These results demonstrate the potential of seedborne and weedborne inoculum to disseminate V. dahliae.

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.

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