Margery L. Daughtrey

Etiology of Phytophthora drechsleri and P. nicotianae (=P. parasitica) diseases affecting floriculture crops

Phytophthora nicotianae and P. drechsleri isolates (n = 413) recovered from eight floricultural hosts at 11 different production sites were described according to compatibility type, resistance to mefenoxam, and amplified fragment length polymorphism (AFLP) profiles. Sample sizes ranged from 2 to 120. In all cases, isolates recovered from a single facility had the same compatibility type and resistance to mefenoxam. AFLP analysis indicated that six clonal lineages of P. nicotianae and two clonal lineages of P. drechsleri were responsible for the 11 epidemics and that isolates recovered from the same facility were identical. A single clone of P. nicotianae was recovered from snapdragons at two field production sites in the southeastern United States receiving seedlings from the same source. This clone persisted at one site from 2000 to 2001. Another clone was recovered from verbena at three separate greenhouse facilities where one facility was supplying verbena to the other two. These results suggest that asexual reproduction of these pathogens plays an important role in epidemics and spread may occur between distant facilities via movement of plants.

Epidemiology and Management of Petunia and Tomato Late Blight in the Greenhouse

Factors affecting the management of petunia and tomato late blight, caused by Phytophthora infestans, under greenhouse conditions were investigated. Late blight-infected petunias (Petunia × hybrida) and tomatoes (Lycopersicon esculentum) each produced sporangia that were dispersed throughout the greenhouse via air currents. Infected petunias produced and released fewer sporangia than infected tomatoes, but infected petunias released sporangia two times longer. Surface-directed irrigation reduced disease incidence compared with overhead irrigation that wetted the foliage. The fungicides dimethomorph-mancozeb, fosetyl-Al, azoxystrobin, and dipo-tassium phosphonate/phosphate suppressed late blight development, as did the plant defense activator acibenzolar-S-methyl. All products were applied twice at 7-day intervals. The other plant defense activator (harpin protein) and the bioantagonists (Trichoderma harzianum, Glio-cladium virens, and Bacillus subtilis) were ineffective at the rates tested.

Temperature and Leaf Wetness Requirements for Pathogen Establishment, Incubation Period, and Sporulation of Phytophthora infestans on Petunia × hybrida

The temperature and leaf wetness requirements for pathogen establishment (germination, infection, and colonization) and the temperature effects on incubation period and sporulation of Phytophthora infestans on petunia were compared with those on tomato. The responses to environmental parameters were found to be similar on petunia and tomato and agreed with those previously reported for late blight development on tomato and potato. In the current study, temperatures ranging from 13 to 23°C generally were conducive to establishment. Very little establishment occurred at 28°C. The minimum leaf wetness period that enabled pathogen establishment was 2 h, whereas most establishment occurred within 6 h of inoculation. The incubation period (time period from inoculation to lesion development) and the time required for development of sporangia after lesions were formed were shortest at 23 and 28°C, respectively. Production of sporangia was greatest (per square centimeter) at 18°C and was nearly absent at 28°C on both petunia and tomato. The sporulation density at 18°C was only slightly less on petunia compared with tomato (20,000 and 24,000 sporangia/cm2, respectively); however, the total lesion area on petunia was only 20% of that on tomato.

Oidium longipes, A New Powdery Mildew Fungus on Petunia in the USA: A Potential Threat to Ornamental and Vegetable Solanaceous Crops

This is the first North American report of Oidium longipes, an anamorphic powdery mildew species described recently in Europe. It was found on vegetatively propagated petunia grown in a commercial greenhouse in New Jersey, USA, where it caused a rapidly spreading disease. The pathogen might have originated offshore and may have already been distributed in the United States through horticultural trade. During field surveys in Europe, it was found on petunia in Hungary and Austria as well; this is the first report of O. longipes from these two countries. A detailed light microscopy study of American and European specimens of O. longipes, including freshly collected samples and authentic herbarium specimens, revealed that its conidiophore morphology is more variable than illustrated in the original species description or in subsequent works. Microcycle conidiation, a process not yet known to occur in powdery mildews, was repeatedly observed in O. longipes. The rDNA internal transcribed spacer (ITS) sequences were identical in colonies containing different conidiophore types as well as in a total of five specimens collected from petunia in the United States, Austria, Hungary, Germany, and Switzerland. A phylogenetic analysis of the ITS sequences revealed that the closest known relative of O. longipes is O. lycopersici, known to infect tomato only in Australia. Cross-inoculation tests showed that O. longipes from petunia heavily infected tobacco cv. Xanthi, while the tomato and eggplant cultivars tested were moderately susceptible to this pathogen. These results indicate that its spread represents a potential danger to a number of solanaceous crops. Our ad hoc field surveys conducted in 2006 and 2007 did not detect it outside New Jersey in the United States; all the other powdery mildew-infected petunias, collected in New York and Indiana, were infected by Podosphaera xanthii. In Europe, most of the powdery mildew-infected petunias examined in this study were infected by P. xanthii or Golovinomyces orontii. Our multiple inoculation tests revealed that the same petunia plants and even the same leaves can be infected concomitantly by O. longipes, O. neolycopersici, G. orontii, and P. xanthii. Thus, it is at present unclear to what extent O. longipes contributes to the powdery mildew epidemics that develop year after year on solanaceous plants in many parts of the world.

First Report of Boxwood Blight Caused by Calonectria pseudonaviculata in Delaware, Maryland, New Jersey, and New York

Boxwood (Buxus spp.) are commercially important evergreen ornamental plants with an annual market value of over

First report of downy mildew on greenhouse and landscape coleus caused by a Peronospora sp. in Louisiana and New York.

103 million in the United States. The recent U.S. incursion of boxwood blight disease caused by the fungus Calonectria pseudonaviculata (syn. Cylindrocladium pseudonaviculatum, Cy. buxicola) threatens the health and productivity of boxwood in both landscape plantings and nurseries. The first confirmed U.S. reports of the disease were made from Connecticut and North Carolina in November 2011 (2,4), followed by diagnoses in 10 additional states during 2012 and 2013. By August 2013, symptoms consistent with boxwood blight had been observed from B. sempervirens in Delaware, Maryland, New Jersey, and southeastern New York. Affected plants showed rapid onset of disease symptoms: dark brown to black spots or diffuse dark areas on leaves, followed by defoliation. Narrow, elongate black cankers also formed on current season shoots. Symptomatic stems and leaves were placed in petri dishes with moistened filter paper at 22°C for 3 days under continuous light. Conidiophores were excised, then placed on potato dextrose agar amended with streptomycin and neomycin (0.3 g/l). Resultant colonies showed dark brown pigmentation at the colony center surrounded by tan to reddish brown rings with white mycelia at the advancing edge. Conidia (n = 30 per isolate) were hyaline, cylindrical, rounded at both ends, with a single septum (45 to 76 × 4 to 6 μm; avg. 63 × 5 μm). Conidiophores (n = 20 per isolate) comprised a stipe, a hyaline septate stipe extension (length 119 to 192 μm; avg. 150 μm) and a terminal ellipsoidal vesicle (diameter 4 to 10 μm; avg. 7 μm). Based on morphological characteristics, the causal agent was identified as C. pseudonaviculata (1,4). Voucher specimens were deposited in the U.S. National Fungus Collections (BPI 892698 to 701). To verify morphological diagnosis, genomic DNA was extracted from fungal biomass grown in liquid cultures of yeast extract peptone dextrose media. A portion of the β-tubulin gene (TUB2) was PCR amplified and sequenced bi-directionally using primers Bta/Bt2b (3). BLASTn searches of NCBI GenBank databases using the TUB2 sequences (Accession Nos. KF785808 to 11) demonstrated 96 to 100% sequence identity with other C. pseudonaviculata isolates. To confirm pathogenicity, 5-month-old B. sempervirens and B. microphylla seedlings were spray-inoculated with a spore suspension of 1 × 104 conidia/ml. One isolate from each state was independently tested with four replicates each. Non-inoculated water-sprayed plants served as negative controls. Plants were maintained in growth chambers at 22°C under constant light. Blight symptoms developed 4 to 5 days post inoculation. C. pseudonaviculata was re-isolated from inoculated plants; no symptoms or signs were observed from control plants. To our knowledge, this is the first report of C. pseudonaviculata in the states of Delaware, Maryland, New Jersey, and New York. This report demonstrates that C. pseudonaviculata is now widespread across the United States eastern seaboard, and represents a substantial threat to boxwood plants in North American landscapes and nurseries. References: (1) P. Crous et al. Sydowia 54:23, 2002. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, USDA-ARS. Retrieved from http://nt.ars-grin.gov/fungaldatabases , 30 August 2013. (3) N. L. Glass and G. C. Donaldson. Appl. Environ. Microbiol. 61:1323, 1995. (4) K. L. Ivors et al. Plant Dis. 96:1070, 2012.

First Report of Erysiphe sedi on Sedum spectabile in North America

In May 2005, two commercial greenhouse flower growers, one in Louisiana (LA) and one in New York (NY), submitted coleus, Solenostemon scutellarioides (L.) Codd, plants for diagnosis after observing stunted growth, inward curling and twisting of leaves, and leaf abscission on multiple cultivars. Downy mildew-like growth was observable with hand lens or a microscope on the abaxial leaf surfaces of affected plants. Irregular necrotic spotting was present on some, but not all, plants on which sporulation was evident. Microscopic examination of LA material led to tentative identification of the pathogen as Peronospora lamii A. Braun (2). The pale brown conidia ranged from 17 to 26 × 15 to 26 μm (average 23 × 19 μm). Conidiophores ranged from 345 to 561 × 9 to 15 μm. No oospores were found. Additional coleus plants with downy mildew were subsequently found in three retail nurseries in LA in early summer. In NY, infected coleus plants were observed in landscapes in Farmington, Rochester, Ithaca, and in two commercial greenhouses between August and October 2005. NY samples sent to the USDA/APHIS in Beltsville, MD were examined, and the fungus was found to have morphology consistent with P. lamii. Two pathogenicity trials were conducted in NY. Conidia were rubbed from an infected coleus leaf onto the leaves of six healthy potted coleus plants of five cultivars and two basil plants that were placed in a shaded plastic tent in the greenhouse where temperatures ranged from 17 to 22°C. A household humidifier was used to supply mist inside the tent for 5 h per day. Six noninoculated plants of each coleus cultivar and two basil plants, placed in the same environment, served as controls. Downy mildew sporulation and some curling and twisting of leaves were observed 14 days after inoculation on all inoculated plants for three of the five cultivars (Florida Rustic Orange, Aurora Peach, and Aurora Mocha). Cvs. Florida Sun Rose and Lava showed no symptoms or signs of downy mildew. An irregularly shaped brown lesion developed on one inoculated basil leaf, and downy mildew sporulation was evident on the abaxial surface 35 days after inoculation. All noninoculated control plants remained disease free. In a second trial, conidia were rinsed from infected coleus leaves and sprayed onto the abaxial leaf surfaces of three coleus cv. Aurora Mocha plants. Three noninoculated plants served as controls and all were placed in a humidity tent. Leaf twisting and downy mildew sporulation were observed 13 days later on all inoculated plants, and control plants showed no sporulation or symptoms. A downy mildew causing disease of greenhouse-grown basil in Europe, originally identified as P. lamii on the basis of morphology, has recently been reported to be taxonomically distinguishable from P. lamii when tested by molecular methods (1). ITS sequences of coleus downy mildew from NY and LA were nearly identical (99% homology) to those of basil downy mildew from Switzerland and Italy (1). To our knowledge, this is the first report of downy mildew occurrence on coleus. References: (1) L. Belbahri et al. Mycol. Res. 109:1276, 2005. (2) S. M. Francis. Peronospora lamii. Descriptions of Pathogenic Fungi and Bacteria. No. 688. CMI, Kew, England, 1981.

First Report of Powdery Mildew (Oidium sp.) on Pincushion Flower (Scabiosa columbaria) in New York

Since 1997, powdery mildew infections have been repeatedly observed on Sedum spectabile plants, cv. Autumn Joy, grown as ornamentals in commercial greenhouses in New York. Circular patches of gray mycelia appeared and spread on upper and occasionally on lower leaf surfaces followed by necrosis of the leaf tissues and defoliation. The new disease reduced the market value of the infected ornamentals and required chemical control. The pathogen produced conidia singly on 2- to 3-celled conidiophores occurring on the ectophytic hyphae. Conidia were subcylindrical, measured 27 to 36 μm × 13 to 17 μm, and contained no fibrosin bodies. Germinating conidia produced a short germ tube, 5 to 30 μm, terminating in a lobed appressorium. Hyphal appressoria were lobed to multi-lobed, opposite or spread along the hyphae. Cleistothecia were not found. Based on conidial characteristics, the pathogen was identified as Erysiphe sedi Braun. To confirm pathogenicity, potted healthy S. spectabile plants were placed near infected plants in the greenhouse. In addition, detached S. spectabile leaves were inoculated with the pathogen by touching them to powdery mildew colonies and then placed in plates filled with one layer of polystyrene balls floated in water. Plates were covered and kept in the laboratory. Uninfected potted plants kept in another greenhouse and noninoculated detached leaves served as controls. After 1 week, powdery mildew appeared on all infected plants and leaves exposed to or inoculated with the pathogen. The pathogen was morphologically identical to the original fungus. No symptoms were observed on the controls. E. sedi is a common Asiatic powdery mildew species infecting many crassulaceous plants (1,2) and was introduced to Eastern Europe from Asia (2). To our knowledge, this is the first report of E. sedi in North America. References: (1) U. Braun. Beih. Nova Hedwigia 89:1, 1987. (2) U. Braun. The Powdery Mildews (Erysiphales) of Europe. Gustav Fisher Verlag, Jena, 1995.

Tospoviruses strike the greenhouse industry: INSV has become a major pathogen on flower crops.

Scabiosa columbaria (Dipsacaceae) is a popular perennial ornamental in the United States. It is native to Europe and was introduced to North America by nursery trade only recently. In the spring of 2006, symptoms of powdery mildew infection were observed on overwintered plants of S. columbaria cv. Butterfly Blue in a nursery in Cutchogue, NY. White powdery mildew mycelia with abundant sporulation were observed on upper and lower leaf surfaces. The portions of leaves with powdery mildew colonies often showed purplish discoloration. Conidia were cylindric to doliiform, measured 20 to 33 × 10 to 15 μm, and were produced singly on 60 to 130 μm long conidiophores consisting of a foot-cell measuring 20 to 50 × 6 to 10 μm, followed by one to three, 12 to 40 μm long cells. Hyphal appressoria were lobed or multilobed. The teleomorph stage was not found. On the basis of these characteristics, the pathogen was identified as an Oidium sp. belonging to the subgenus Pseudoidium. Recently, an anamorphic powdery mildew fungus with similar morphological characteristics, identified as Erysiphe knautiae, was reported on S. columbaria cv. Butterfly Blue in Washington (2). E. knautiae is a common powdery mildew species of dipsacaceous plants such as Scabiosa spp. and Knautia spp. in Europe and Asia (1). To determine whether the fungus reported here was E. knautiae, DNA was extracted from its mycelium, and the internal transcribed spacer (ITS) region of the ribosomal DNA was amplified and sequenced as described earlier (4). No ITS sequences are available in public DNA databases for E. knautiae, thus, we determined this sequence in a specimen of E. knautiae collected from Knautia arvensis in The Netherlands. Herbarium specimens of the Oidium sp. infecting S. columbaria in New York and E. knautiae from the Netherlands were deposited at the U.S. National Fungus Collections under accession numbers BPI 878259 and BPI 878258, respectively. The ITS sequence from Oidium sp. infecting S. columbaria in New York (GenBank Accession No. EU377474) differed in two nucleotides from that of E. knautiae infecting K. arvensis in the Netherlands (GenBank Accession No. EU377475). These two ITS sequences were also more than 99% similar to those of some newly emerged anamorphic powdery mildew fungi: Oidium neolycopersici and other Oidium spp. infecting Chelidonium majus, Passiflora caerulea, and some crassulaceous plants (3,4). Thus, it is unclear whether the fungus reported here was E. knautiae known from Eurasia or an Oidium sp. that has acquired pathogenicity to S. columbaria. To our knowledge, this is the first report of powdery mildew on S. columbaria in New York. References: (1) U. Braun. Beih. Nova Hedwigia 89:1, 1987. (2) D. A. Glawe and G. G. Grove. Online publication. doi:10.1094/PHP-2005-1024-01-BR. Plant Health Progress, 2005. (3) B. Henricot. Plant Pathol. 57:779, 2008. (4) T. Jankovics et al. Phytopathology 98:529, 2008.