Phytophthora Species Are Common on Nursery Stock Grown for Restoration and Revegetation Purposes in California.

Phytophthora tentaculata was detected for the first time in North America in 2012 in a nursery on sticky monkeyflower plant (Diplacus aurantiacus) and again in 2014 on outplanted native plants. At that time, this species was listed as a federally actionable and reportable pathogen by the USDA. As a result of these detections, California native plant nurseries were surveyed to determine the prevalence of Phytophthora species on native plant nursery stock. A total of 402 samples were collected from 26 different native plant nurseries in California between 2014 and 2016. Sampling focused on plants with symptoms of root and crown rot. Symptomatic tissue was collected and tested by immunoassay, culture, and molecular techniques (PCR). Identifications were made using sequences from the internal transcribed spacer (ITS) rDNA region, a portion of the trnM-trnP-trnM, or the atp9-nad9 mitochondrial regions. Phytophthora was confirmed from 149 of the 402 samples (37%), and from plants in 22 different host families. P. tentaculata was the most frequently detected species in our survey, followed by P. cactorum and members of the P. cryptogea complex. Other species include P. cambivora, P. cinnamomi, P. citricola, P. hedraiandra, P. megasperma, P. multivora, P. nicotianae, P. niederhauserii, P. parvispora, P. pini, P. plurivora, and P. riparia. A few Phytophthora sequences generated from mitochondrial regions could not be assigned to a species. Although this survey was limited to a relatively small number of California native plant nurseries, Phytophthora species were detected from three quarters of them (77%). In addition to sticky monkeyflower, P. tentaculata was detected from seven other hosts, expanding the number of associated hosts. During this survey, P. parvispora was detected for the first time in North America from symptomatic crowns and roots of the nonnative Mexican orange blossom (Choisya ternata). Pathogenicity of P. parvispora and P. nicotianae was confirmed on this host. These findings document the widespread occurrence of Phytophthora spp. in native plant nurseries and highlight the potential risks associated with outplanting infested nursery-grown stock into residential gardens and wildlands.

Phylogeny and taxonomy of the genus Tubakia s. lat.

The genus Tubakia is revised on the basis of morphological and phylogenetic data. The phylogenetic affinity of Tubakia to the family Melanconiellaceae (Diaporthales) was recently postulated, but new analyses based on sequences retrieved from material of the type species of Tubakia, T. dryina, support a family of its own, viz. Tubakiaceae fam. nov. Our phylogenetic analyses revealed the heterogeneity of Tubakia s. lat. which is divided into several genera, viz., Tubakia s. str., Apiognomonioides gen. nov. (type species: Apiognomonioides supraseptata), Involutiscutellula gen. nov. (type species: Involutiscutellula rubra), Oblongisporothyrium gen. nov. (type species: Oblongisporothyrium castanopsidis), Paratubakia gen. nov. (type species: Paratubakia subglobosa), Racheliella gen. nov. (type species: Racheliella wingfieldiana sp. nov.), Saprothyrium gen. nov. (type species: Saprothyrium thailandense) and Sphaerosporithyrium gen. nov. (type species: Sphaerosporithyrium mexicanum sp. nov.). Greeneria saprophytica is phylogenetically closely allied to Racheliella wingfieldiana and is therefore reallocated to Racheliella. Particular emphasis is laid on a revision and phylogenetic analyses of Tubakia species described from Japan and North America. Almost all North American collections of this genus were previously referred to as T. dryina s. lat., which is, however, a heterogeneous complex. Several new North American species have recently been described. The new species Sphaerosporithyrium mexicanum, Tubakia melnikiana and T. sierrafriensis, causing leaf spots on several oak species found in the North-Central Mexican state Aguascalientes and the North-Eastern Mexican state Nuevo León, are described, illustrated, and compared with similar species. Several additional new species are introduced, including Tubakia californica based on Californian collections on various species of the genera Chrysolepis, Notholithocarpus and Quercus, and T. dryinoides, T. oblongispora, T. paradryinoides, and Paratubakia subglobosoides described on the basis of Japanese collections. Tubakia suttoniana nom. nov., based on Dicarpella dryina, is a species closely allied to T. californica and currently only known from Europe. Tubakia dryina, type species of Tubakia, is epitypified, and the phylogenetic position and circumscription of Tubakia are clarified. A revised, supplemented key to the species of Tubakia and allied genera on the basis of conidiomata is provided.

Detection of Phytophthora ramorum in Nurseries and Forest Lands in California in 2004 to 2009.

From December 2004 through May 2009, samples were collected from California nurseries and wild lands to survey for Phytophthora ramorum and comply with federal regulations of nursery stock. Samples were prescreened by an enzyme-linked immunosorbent assay (ELISA) that detects Phytophthora spp. and tested by culture, P. ramorum-specific real-time polymerase chain reaction (PCR), and nested PCR. Yearly percentages of infected samples ranged from 0.6 to 2.3%. Camellia spp., Rhododendron spp., Magnolia spp., Pieris spp., and Laurus nobilis tested positive the most frequently in the nurseries and Lithocarpus densiflorus, Umbellularia californica, and Quercus agrifolia tested positive most often from wild lands. Of the 118,410 samples isolated onto PARP media, 0.8% was identified as P. ramorum. Of 115,056 samples tested by ELISA, 5.9% tested positive for Phytophthora spp. Of the 6,520 samples tested by PCR, 12.4% tested positive for P. ramorum. The false-negative, positive, and internal control failure rates of the assays are discussed. After removing the seasonal effect of sampling strategy, isolation of the pathogen into culture was found to be seasonally dependent whereas detectability by PCR and ELISA was not. To our knowledge, this is the first evaluation of a regulatory testing program for a plant pathogen on this scale using standardized assays.

Fungal Planet description sheets: 281-319.

Novel species of fungi described in the present study include the following from South Africa: Alanphillipsia aloeicola from Aloe sp., Arxiella dolichandrae from Dolichandra unguiscati, Ganoderma austroafricanum from Jacaranda mimosifolia, Phacidiella podocarpi and Phaeosphaeria podocarpi from Podocarpus latifolius, Phyllosticta mimusopisicola from Mimusops zeyheri and Sphaerulina pelargonii from Pelargonium sp. Furthermore, Barssia maroccana is described from Cedrus atlantica (Morocco), Codinaea pini from Pinus patula (Uganda), Crucellisporiopsis marquesiae from Marquesia acuminata (Zambia), Dinemasporium ipomoeae from Ipomoea pes-caprae (Vietnam), Diaporthe phragmitis from Phragmites australis (China), Marasmius vladimirii from leaf litter (India), Melanconium hedericola from Hedera helix (Spain), Pluteus albotomentosus and Pluteus extremiorientalis from a mixed forest (Russia), Rachicladosporium eucalypti from Eucalyptus globulus (Ethiopia), Sistotrema epiphyllum from dead leaves of Fagus sylvatica in a forest (The Netherlands), Stagonospora chrysopyla from Scirpus microcarpus (USA) and Trichomerium dioscoreae from Dioscorea sp. (Japan). Novel species from Australia include: Corynespora endiandrae from Endiandra introrsa, Gonatophragmium triuniae from Triunia youngiana, Penicillium coccotrypicola from Archontophoenix cunninghamiana and Phytophthora moyootj from soil. Novelties from Iran include Neocamarosporium chichastianum from soil and Seimatosporium pistaciae from Pistacia vera. Xenosonderhenia eucalypti and Zasmidium eucalyptigenum are newly described from Eucalyptus urophylla in Indonesia. Diaporthe acaciarum and Roussoella acacia are newly described from Acacia tortilis in Tanzania. New species from Italy include Comoclathris spartii from Spartium junceum and Phoma tamaricicola from Tamarix gallica. Novel genera include (Ascomycetes): Acremoniopsis from forest soil and Collarina from water sediments (Spain), Phellinocrescentia from a Phellinus sp. (French Guiana), Neobambusicola from Strelitzia nicolai (South Africa), Neocladophialophora from Quercus robur (Germany), Neophysalospora from Corymbia henryi (Mozambique) and Xenophaeosphaeria from Grewia sp. (Tanzania). Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

Leaf Spot of Arugula, Caused by Alternaria japonica, in California.

Arugula (Eruca vesicaria subsp. sativa (Mill.) Thell. is a Cruciferous plant used for culinary purposes. From 2012 to 2013, a foliar disease seriously impacted the growth and quality of about 0.1 ha of hydroponically grown arugula at a Santa Barbara County nursery. Samples of affected arugula seedlings exhibited adaxial and abaxial symptoms of mottling with circular to oval, water soaked, dark green leaf spots, each 1 to 3 mm in diameter, and some of which coalesced. Conidia of an Alternaria sp. were observed on the foliage. Symptomatic leaf pieces were disinfested with 0.6% NaOCl, blotted dry, and plated on acidified potato dextrose agar (APDA). Cultures were incubated under near-UV lights for 24 h/day. Olivaceous-grey colonies of the same Alternaria species observed on the leaves grew after 7 days. After 21 days on carrot-piece agar (3), the fungus produced beakless conidia with longitudinal and constricted transverse septa that measured 30.0 to 69.0 × 12.5 to 20.0 µm and were borne singly or in short chains of 2 to 3 conidia. In addition, knots of dark, thick-walled micro-chlamydospores were produced by the hyphae. The fungus was identified morphologically as Alternaria japonica Yoshii (2), and the species confirmed by sequence analysis. A portion of the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) was amplified using ITS1 and ITS4 primers (4). The sequence (GenBank Accession No. KJ126846) was 100% identical to the ITS rDNA sequence of an isolate of A. japonica (KC584201) using a BLASTn query. A. japonica was also detected in seeds of the lot used to grow the affected arugula crop. Pathogenicity of a single isolate was tested by inoculating four 37-day-old plants each of arugula, cabbage (Brassica oleracea L. var. capitata), and broccoli (B. oleracea L. var. botrytis L.). Inoculum was obtained from 11-day-old cultures of the isolate grown at 24°C on half-strength APDA. Half of a 2.5 cm diameter agar plug containing hyphae and conidia was ground in 2 ml of sterilized water, and the volume of water increased to 45 ml. Leaves of four plants/host species were sprayed with 3.5 to 4.0 ml of inoculum. The inoculated plants and four control plants of each species treated similarly with sterilized water were immediately incubated in a dark dew chamber at 23°C. After 72 h in the dew chamber, inoculated plants of all three hosts produced similar symptoms of wilting, water soaking, and dark green leaf spotting as the original symptomatic field plants. Conidia formed in the leaf spots on both sides of inoculated leaves. A. japonica was re-isolated from all of the inoculated plants but from none of the symptomless control plants using the method previously described. Pathogenicity tests were repeated, with similar results. Although reported in Italy in 2013 (1), to our knowledge, this is the first report of A. japonica on arugula in the United States. References: (1) G. Gilardi et al. Acta Hort. 1005:569, 2013. (2) E. G. Simmons. Page 368 in: Alternaria, An Identification Manual. CBS Fungal Biodiversity Centre, Utrecht, 2007. (3) S. Werres et al. Z. Planzenkr. Pflanzensh. 108:113, 2001. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

First Report of Stem Blight of Blueberry in California Caused by Neofusicoccum parvum.

In July 2013 in coastal (Santa Barbara County) California, commercial plantings of southern highbush blueberry (Vaccinium corymbosum) developed symptoms of a previously undiagnosed disease. Symptoms consisted of reddening and wilting of foliage, with leaves and small twigs later drying up. The bark of diseased branches was discolored and sunken; removal of this bark revealed a brown discoloration of the underlying wood. Approximately 5% of the planting was affected. When placed on acidified potato dextrose agar (A-PDA), surface disinfested pieces of symptomatic wood consistently yielded one type of fungus. On A-PDA, isolates produced extensive white aerial mycelium that turned dark gray after 4 to 5 days and formed pycnidia after 21 days. Three single-spore isolates were grown on PDA for 21 days for morphological and molecular characterization. Conidia were hyaline, smooth, and ellipsoid with round apices and truncated bases. Conidia measured 13 to 20 × 5 to 7.5 µm (n = 50; mean 16.7 × 6.1 µm), with a length/width ratio of 2.73. After 25 days, conidia became biseptate with a darker middle cell. rDNA sequences of the internal transcribed spacer (ITS) region of the isolates (GenBank KJ126847 to 49), amplified using primers ITS1 and ITS4 (5), were 99% identical to the holotype isolate of Neofusicoccum parvum Pennycook and Samuels (3) by a BLAST query (GU251125). Partial sequences of the translation elongation factor 1-alpha (EF1-α) gene (KJ126850 to 52), obtained using primers EF728Fa and EF986R (5), were 99% identical to N. parvum (GU251257). To demonstrate Koch's postulates, 14-day-old colonies of the three N. parvum isolates were grown on A-PDA. Using three blueberry cultivars (Abundance, Jewel, and Snowchaser), slits were cut beneath the epidermis of branches 1 cm diameter or less; one colonized agar plug (6 mm diameter) was placed into each cut and the epidermis was resealed with Parafilm. Ten inoculations (one inoculation per branch; two branches per plant) were made for each isolate and each cultivar; inoculated plants were maintained in a greenhouse. After 10 to 14 days, leaves on inoculated branches turned red and wilted, bark above and below the inoculation sites turned brown, and vascular tissue beneath the bark was also brown. After 21 days, diseased areas became sunken. N. parvum was recovered from all inoculated branches of all cultivars and matched the characteristics of the original isolates. Control branches, inoculated with sterile agar plugs, did not develop any symptoms and N. parvum was not isolated. This experiment was repeated with similar results. Many Botryosphaeriaceae species, including N. parvum, are associated with canker and dieback symptoms on blueberry worldwide (2). To our knowledge, this is the first documentation of stem blight caused by N. parvum on blueberry in CA. Blueberry is a rapidly expanding industry in the state, with 960 ha planted in 2005 increasing to 2,830 ha in 2012 (1). Drought stress predisposes plants to stem blight caused by Botryosphaeriacease species (4); therefore, expansion into arid areas of CA could increase the incidence and severity of N. parvum. References: (1) N. Amer. Blueberry Council. 2012 World Blueberry Acreage & Prod. Rept., 2013. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab., online publication, ARS, USDA. Retrieved February 5, 2014. (3) S. R Pennycook and G. J. Samuels. Mycotaxon 24:445, 1985. (4) W. A. Sinclair and H. H. Lyon. Diseases of Trees and Shrubs, Second Edition. Comstock Publ. Assoc. 2005. (5) B. Slippers et al. Mycologia 96:83, 2004.

First Report of Root and Stem Rot Caused by Phytophthora tentaculata on Mimulus aurantiacus in North America.

Sticky monkey flower plant, Mimulus aurantiacus (Phrymaceae), is a small, perennial shrub that is widely distributed throughout California, especially in coastal and disturbed habitats. It is also found in native plantings in parks and landscapes. In October 2012, nearly all the M. aurantiacus plants grown in a Monterey County, CA nursery for a restoration project were stunted and had dull, yellowish leaves. Roots and stem collars had necrotic, sunken lesions with few feeder roots. Thirty percent of the plants had died. Samples of diseased plants were sent to the CDFA-PPDC Lab and tested positive for Phytophthora sp. using the Agdia ELISA Phytophthora kit (Agdia, Elkhart, IN). A Phytophthora sp. was consistently isolated from the tissue on corn meal agar-PARP (CMA-PARP) (2). Sporangia were spherical to ovoid, papillate to bipapillate and 17 to 42.5 (avg. 27.5) × 12 to 35 (avg. 22.9) µm, with a length/breadth ratio of 1.2:1. Chlamydospores, which were spherical, terminal to intercalary, thin walled and 27.5 to 40 µm, and hyphal swellings formed on CMA-PARP. Spherical oospores, 25 to 36 µm, with primarily paragynous antheridia formed readily on V8 juice agar. rDNA sequences of the internal transcribed spacer (ITS) region of the isolates (GenBank KF667505), amplified using primers ITS1 and ITS4, were 100% identical to Phytophthora tentaculata (CBS 552.96, GenBank AF266775) by a BLAST query (1,3). To assess pathogenicity, exposed root crowns of three 3.78-liter potted M. aurantiacus plants were inoculated with 20 ml of zoospore suspension (2 × 104 ml-1). Plants were maintained in a 23°C growth chamber with a 12-h photoperiod and watered daily. Sterile water was applied to the exposed crowns of three control plants. At 2 weeks, all inoculated plants were wilted with chlorotic foliage. After 3 weeks, the cortical tissue of the crowns and roots was discolored and sloughing and P. tentaculata was recovered on CMA-PARP. P. tentaculata did not grow from the asymptomatic control plants. Inoculations were repeated with similar results. P. tentaculata is a homothallic species in Phytophthora clade 1 that causes crown, root, and stalk rot of nursery plants in Europe and China (1,4). A USDA PERAL analysis lists it as one of the top 5 Phytophthora species of concern to the United States (4). Genera infected with P. tentaculata include Apium, Aucklandia, Chicorium, Chrysanthemum, Delphinium, Gerbera, Lavandula, Santolina, Origanum, and Verbena (4). To our knowledge, this is the first report of P. tentaculata in North America. The source of inoculum of P. tentaculata in California remains unknown. The nursery used seed and cuttings of M. aurantiacus from nearby native areas for propagation, and P. tentaculata was not found in neighboring plant hosts or by baiting soil and water at the nursery. All infected M. aurantiacus material was destroyed. The presence of P. tentaculata in California nurseries could have serious economic impacts on the nursery industry and environmental impacts on susceptible native hosts, if spread into the wildlands. References: (1) D. E. L. Cooke et al. Fungal Genet. Biol. 30:17, 2000. (2) S. N. Jeffers and S. B. Martin. Plant Dis. 70:1038, 1986. (3) H. Krober and R. Z. Marwitz. Pflanzenkrankh. Pflanzenschutz 100:250, 1993. (4) U.S. Department of Agriculture, Animal and Plant Health Inspection Services (APHIS). Phytophthora species in the Environment and Nursery Settings New Pest Response Guidelines, USDA-APHIS-PPQ-Emergency and Domestic Programs-Emergency Management, Riverdale, MD, 2010.

First Report of Candidatus Liberibacter asiaticus Associated with Citrus Huanglongbing in California.

Huanglongbing (HLB), also known as citrus greening, is one of the most destructive citrus diseases worldwide and is seen as a major threat to the multimillion dollar citrus industry in California. The vector of the two bacterial species associated with this disease, Candidatus Liberibacter asiaticus and Ca. L. americanus, is the Asian citrus psyllid (ACP), Diaphorina citri (4). ACP was detected in California in August of 2008 and has since been detected in nine counties in southern California. As part of a long term survey and testing program for the ACP carrying the HLB associated bacteria, groups of ACP nymphs and adults were submitted to the Jerry Dimitman Citrus Research Board/Citrus Pest and Disease Prevention Program Laboratory in Riverside, CA. In March 2012, DNA extracted using the Qiagen MagAttract 96 DNA plant kit (QIAGEN Inc., 27220 Turnberry Lane, Suite 200, Valencia, CA 91355) from a group of three ACP adults tested positive for Ca. L. asiaticus with the real-time PCR assay developed by Li et al. (4). ACP adults were collected from a residential citrus tree located in the Hacienda Heights area of Los Angeles County, California. The approximately 1.8 meter tall lemon tree had 23 graft unions, primarily of lemon (Citrus × meyeri) and pomelo (Citrus maxima) varieties. The tree was unthrifty, with yellow shoots and chlorotic leaves. Symptoms on the lemon and pomelo leaves included asymmetrical blotchy mottling, yellowing, and corking of the leaf veins, with the blotchy mottle more prominent in the pomelo leaves. Pomelo leaves appeared crinkled along the thickened veins. Lemon leaves had yellow veins and a few had islands of green tissue completely surrounded by yellow tissue. The entire tree was removed, cut into sections, bagged, and transported to the CDFA Plant Pest Diagnostics Lab for analysis. Two hundred milligrams of petiole and midrib tissue from leaves apical to each graft union was collected, and DNA from each sample was extracted using the Qiagen DNeasy plant mini kit. DNA extracted from both lemon and pomelo leaves tested positive for Ca. L. asiaticus using real-time PCR (4). A 1,160-bp fragment of the 16S ribosomal RNA gene was amplified from the insect and plant DNA extracts using conventional PCR with primers Ol1 and OI2c (2). A 703-bp fragment of the ß-operon gene was amplified from the insect and plant extracts with primers A2 and J5 (1). The 16S rDNA fragments from the insect and plant respectively (GenBank Accession Nos. JX430434 and JX455745) and the ß-operon fragments (JX430435 and JX455746) showed 100% identity with the corresponding regions of Ca. L. asiaticus (CP001677) strain psy 62. Our 16S rDNA sequence showed 98% identity with Ca. L. africanus (EU921620), 97% identity with Ca. L. solanacearum (HM246509), and 96% with Ca. L. americanus (FJ036892). In response to the detection of HLB, a 241 km2 quarantine area around the detection site was established. Surveys for ACP and symptomatic host plants within the HLB quarantine area are ongoing. To date, there have been no additional positive detections. In the United States, HLB was first detected in Florida in 2005 (4) and in Texas in January of 2012 (3). To our knowledge, this is the first confirmed report of Ca. L. asiaticus associated with HLB in California. References: (1) A. Hocquellet et al. Mol. Cell. Probes 13:373, 1999. (2) S. Jagoueix et al. Mol. Cell. Probes 10:43, 1996. (3) M. Kunta et al. Phytopathology 102:S4.66, 2012. (4) W. Li et al. J. Microbiol. Methods 66:104, 2006.

First Report of Colletotrichum phormii Causing Anthracnose on New Zealand Flax in the United States.

Phormium colensoi Hook.f. (syn. P. cookianum), New Zealand flax, (family Xanthorrhoeaceae) is popular in ornamental landscapes in the United States because of its sturdy blade-like foliage available in diverse colors. In February 2012, the Oregon State University Plant Clinic received three potted plants of P. colensoi 'Black Adder' from a commercial nursery in Santa Cruz County, California. The margins and midribs of several leaves had brown lesions that were variable in size, and fusiform to ellipsoidal in shape. Embedded in the lesions were black acervuli without setae that exuded salmon-colored spore masses under moist conditions. Conidia were hyaline, cylindrical to fusiform, straight to slightly curved, and 22.4 to 35.2 × 4.0 to 6.4 (average 24.7 × 4.9) µm. Based on morphology, the fungus was confirmed by USDA-APHIS National Identification Services to be Colletotrichum phormii (Henn.) D.F. Farr & Rossman (2). In March 2012, the California Department of Food and Agriculture Plant Pest Diagnostic Lab received additional samples from the same nursery lot (25% disease incidence) from which a similar fungus was recovered. rDNA sequences of the internal transcribed spacer (ITS) region from the California isolate (GenBank KC122681), amplified using primers ITS1 and ITS4 (2), were 100% identical to multiple species of Colletotrichum, including C. phormii by a BLAST query (JQ948446 through JQ948453). ITS sequence similarity alone is not sufficient to address Colletotrichum taxonomy and must be used in combination with host range and morphology (1). Pathogenicity of C. phormii (isolate CDFA986) was tested on three 'Black Adder' plants, which were inoculated with 6-mm agar plugs from a 14-day-old culture grown on half strength potato dextrose agar (PDA). Leaves were wound-inoculated along the midrib using colonized plugs (4). Five leaves per plant were inoculated with C. phormii plugs and five leaves per plant were treated with uncolonized PDA agar plugs as controls. Plants were sprayed with water and incubated in plastic bags at 22°C with a 12-h photoperiod. After 48 h, the bags and caps were removed and plants were kept under the same conditions. Two weeks later, water-soaked lesions had developed on the inoculated leaves. Lesions expanded along the midrib and became fusiform in shape after 21 to 28 days. C. phormii was isolated from lesion margins of all the inoculated leaves, but not from control leaves. This experiment was repeated once with similar results. Another Colletotrichum species, C. gloeosporiodes, also occurs on Phormium spp., but differs from C. phormii in morphology and symptom expression. Subsequent nursery and landscape surveys showed that anthracnose caused by C. phormii occurs on several P. colensoi cultivars as well as on P. tenax in five California counties including Santa Cruz, Yolo, Sacramento, San Luis Obispo, and Solano. C. phormii is also reported to infect P. colensoi and P. tenax in New Zealand, Europe, the United Kingdom, Australia, and South Africa (2,3). To our knowledge, this is the first report of C. phormii causing anthracnose on Phormium in North America. This disease could impact the American nursery trade and New Zealand flax production due to crop loss and increased production costs for pest management. References: (1) J. Crouch et al. Mycologia 101:648, 2009. (2) D. F. Farr et al. Mycol. Res. 110:1395, 2006. (3). H. Golzar and C. Wang. Australas. Plant Pathol. 5:110, 2010. (4) L. E. Yakabe et al. Plant Dis. 93:883, 2009.

First Report of Camphor Tree (Cinnamomum camphora) as a Host of Phytophthora ramorum.

Cinnamomum camphora (Lauraceae) is an evergreen shade tree grown in many parts of the United States, including California. From 2007 to 2011, an arborist working in a residential neighborhood in Mill Valley (Marin Co.) noticed several camphor trees with branch dieback and decline. Affected trees had patchy, irregular cankers on the branches and shoot blight. Cankers were black and most had horizontal fissures. Cankers were most abundant in the inside and lower portions of the canopies. In 2011, samples sent to Bartlett Tree Laboratory tested positive for Phytophthora sp. using the Agdia ELISA Phytophthora kit (Agdia, Elkhart, IN). In February 2009 and April 2011, camphor leaf samples were collected by Sacramento Co. inspectors during an annual nursery inspection for Phytophthora ramorum and submitted to CDFA. The normally bright green leaves were reddish with small necrotic spots surrounded by green halos. Camphor samples from Marin Co. were also collected and sent to CDFA in September 2011. An organism with coralloid coenocytic hyphae, chlamydospores, and ellipsoidal semi-papillate sporangia grew on CMA-PARP (4) from both Marin and Sacramento Co. samples. Morphologically, it matched the description of P. ramorum (3). rDNA sequences of the internal transcribed spacer (ITS) region of the Marin (GenBank KC473521) and Sacramento (KC473522) isolates, amplified using primers ITS1 and ITS4 (4), were 100% identical to P. ramorum by a BLAST query (AY038058). Microsatellite loci placed the Marin isolate in the NA1 clonal lineage, while the Sacramento isolate belonged to the NA2 lineage (2). Pathogenicity of both isolates was tested on 5 trees grown in 18.93-liter pots. Three leaves on each tree were inoculated with 6-mm agar plugs taken from the margin of 7-day-old cultures grown on V8 juice agar (V8). Leaves were wounded with a sterile pushpin and two colonized plugs of each isolate were covered with a freezer tube cap filled with sterile dH2O and attached to the leaves with a pin-curl clip (4). Three branches of the same plants were wounded and inoculated with a 3-mm colonized agar plug for each isolate and secured with Parafilm. An equal number of leaves and stems were treated with uncolonized V8 plugs as controls. Plants were sprayed with dH2O, covered in large plastic bags, and placed in a growth chamber at 18°C. After 4 days, the bags, caps, and plugs were removed from the leaves. Black lesions were seen 7 days after inoculation on most leaves and 10 to 14 days on inoculated branches. After 32 days, P. ramorum was isolated from leaf lesions and canker margins onto CMA-PARP. No Phytophthora spp. grew from the controls. The experiment was repeated once with similar results. Overall, leaf and stem lesions were larger with the NA2 lineage isolate than the NA1 lineage isolate, which is consistent with previous research (1). Leaf abscission was seen in 30% of the leaves inoculated with the NA2 lineage isolate but none of the NA1 or control leaves. To our knowledge, this is the first report of P. ramorum on camphor in nursery and landscape settings. Mill Valley is known for its mild temperatures and abundant summer fog. Optimal weather conditions likely led to the spread of P. ramorum from infected neighboring forest hosts to camphor in Mill Valley, rather than from an introduction of infected nursery plants. References: (1) E. Elliott et al. For. Pathol. 41:7, 2011. (2) E. M. Goss et al. Phytopathology 101:166, 2011. (3) S. Werres et al. Mycol. Res. 105:1155, 2001. (4) L. E. Yakabe et al. Plant Dis. 93:883, 2009.

First Report of Neofabraea alba Causing Fruit Spot on Olive in North America.

Olive (Olea europaea) is a widely planted evergreen tree primarily grown for its oil, fruit for pickling, and landscape appeal in Mediterranean and temperate climates. California produces most of the olives grown in the United States; its industry was valued at $53 million in 2011 (4). In 2005 and 2008, fruit spotting occurred on coratina and picholine cultivars in two commercial orchards in Sonoma County. The spots were scattered, slightly sunken and brown, and surrounded by a green halo. Many of the spots were associated with lenticels. A slow to moderate growing, cream to rose-colored fungus was isolated from the spots onto potato dextrose agar (PDA) amended with 0.01% tetracycline hydrochloride. Sporulation was observed in vitro on PDA after 40 days under near-UV light. Macroconidia, produced from conidiomata, were hyaline, aseptate, cylindrical to fusiform-allantoid, slightly curved, and 17 to 27 × 2.5 to 3.5 µm (average 21.1 × 2.9 µm). Microconidia were aseptate, strongly curved, hyaline, and 14 to 18 × 0.75 to 1 µm (average 16.1 × 0.9 µm). rDNA sequences of the internal transcribed spacer (ITS) region of the isolate (GenBank KC751540), amplified using primers ITS1 and ITS4, were 99.8% identical to Neofabraea alba (E.J. Guthrie) Verkley (anamorph Phlyctema vagabunda) (=Gloeosporium olivae) (AF141190). Pathogenicity was tested on detached, green fruit (cv. frantoio). Olives were surface sterilized in 10% sodium hypochlorite for 5 min and air dried. Five olives were wounded with a needle and 10 µl spore suspension (105 spores/ml) was placed on each wound. An equal amount of spore suspension was placed on five unwounded olives. Water was also placed on wounded and unwounded olives to serve as a control. The olives were placed on racks in 22.5 × 30 cm crispers lined with wet paper towels and incubated at 23°C. After 21 days, the olives began to turn red. Olives wounded and inoculated with N. alba had a distinct green ring around the inoculation point where maturity was inhibited. Control olives uniformly turned red. After 35 days, wound-inoculated olives began to form a sunken, brown lesion at the inoculation point where aerial mycelium was visible. After 51 days, lesions were visibly sunken and immature conidiomata began to form in concentric rings giving a bull's eye-like appearance. Unwounded fruit exhibited uneven maturity and green spots associated with the lenticels throughout the experiment but did not develop sunken lesions. Control fruit showed no symptoms and ripened normally. After 56 days, fruit was surface sterilized in 10% sodium hypochlorite for 5 min and plated onto PDA. N. alba was isolated from the sunken and green areas of all of the wounded and unwounded fruit. No fungi grew from the control fruit. The experiment was repeated once with similar results. N. alba has been reported to cause an anthracnose disease on fruit and leaves of olives in Spain and Italy (1,2). In North America, N. alba causes a bull's eye rot on fruit of Malus and Pyrus spp. in the Pacific Northwest and coin canker of Fraxinus spp. in Michigan and Canada (3). To our knowledge, this is the first report of N. alba causing disease on olive in North America. References: (1) J. Del Maral de la Vega et al. Bol. San Veg. Plagas. 12:9. 1986. (2) S. Foschi. Annali. Sper. Agr., n.s. 9:911. 1955. (3) T. D. Gariepy et al. Can. J. Plant Pathol. 27:118. 2005. (4) United States Department of Agriculture, National Agricultural Statistics Service, California Field Office, California Agriculture Statistics, Crop Year 2011.

First Report of Powdery Mildew Caused by Podosphaera euphorbiae-hirtae on Euphorbia tithymaloides in California.

Euphorbia tithymaloides (Euphorbiaceae; known as 'Jacob's ladder,' 'Devil's Backbone') is a perennial, succulent spurge, grown primarily as a border plant in ornamental landscapes. In June 2011 and February 2012, the California Department of Food and Agriculture Plant Pest Diagnostics Lab, Sacramento, CA, received an unusual powdery mildew sample on greenhouse-grown E. tithymaloides from a Ventura County, CA nursery. Disease incidence at the nursery was 100%. White mycelial patches were present on the stems and on both sides of the leaves. Over time, heavily infected branches defoliated and brownish, roughened, scabby lesions developed on the stems. Hyphae were thin-walled, up to 8 µm wide and developed nipple-shaped appressoria. Ellipsoid-ovoid conidia measured 21.0 to 32.5 × 13 to 18 µm (avg. 26.4 × 13.9 µm, n = 20) and formed in chains. The rDNA internal transcribed spacer (ITS) region was amplified with primers PFITS-F and PF5.8-R (4). The 387-bp sequence (GenBank JX006103) was 99% similar (346/347 bp) to Podosphaera euphorbia-hirtae (AB040306) from Acalypha australis (Euphorbiaceae) (3). Based on ITS similarity and culture morphology, the fungus was identified as P. euphorbiae-hirtae U. Braun & Somani (1,3). Pathogenicity was confirmed through inoculation by gently pressing diseased leaves from the nursery onto the youngest leaves of three plants each of E. tithymaloides cultivars 'Nano' and 'Variegated.' Leaves of an equal number of control plants were pressed with healthy leaves. Plants were incubated in a dew chamber for 48 h after which they were transferred to a 22°C growth chamber with a 12-h photoperiod. The experiment was repeated once. White powdery mildew colonies formed after 7 days on 'Variegated' and 13 days on 'Nano'. Conidia measured 27.5 to 35.0 × 11 to 15 µm (avg. 30.5 × 12.6 µm, n = 30) which was within the range of P. euphorbia-hirtae. No symptoms developed on the control plants. P. euphorbiae-hirtae has been reported in Asia and the UK on E. tithymaloides and in Asia on A. australis (2). An asexual Oidium stage on Euphorbiaceae in Asia, Africa, Australia, Florida, Puerto Rico, Cuba, and the U.S. Virgin Islands may correspond to P. euphorbiae-hirtae (2). To our knowledge, this is the first report of P. euphorbiae-hirtae in California. Following the 2011 and 2012 detections, all E. tithymaloides plants in the Ventura County, CA nursery were destroyed. A regulatory trace back survey found that the plants were shipped from a Florida supplier, which was also shown to have an outbreak of P. euphorbiae-hirtae. The original source of the Florida E. tithymaloides plants was a 2010 shipment from Costa Rica. The host range of P. euphorbiae-hirtae is restricted to three landscape species in the Euphorbiaceae. References: (1) U. Braun. Beih. Nova Hedwigia 89:143, 1987. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/index.cfm May 1, 2012. (3) T. Hirata. et al. Can. J. Bot. 78:1521, 2000. (4) R. Singh et al. Plant Dis. 93:1348, 2009.

First Report of Phytophthora ramorum Causing a Leafspot on Loropetalum chinense, Chinese Fringe Flower in California.

Chinese fringe flower is a popular landscape plant in California for its red evergreen foliage and its showy red flowers in the spring. In April 2007, a sample was submitted to the California Department of Food and Agriculture diagnostic laboratory from Sacramento County as part of an inspection of a nursery for Phytophthora ramorum. A sample was taken from Loropetalum chinense because the inspector noticed very small spots and defoliation in the crop, even though P. ramorum was not detected in previous samples sent to the lab with similar symptoms. Six 5-mm2 pieces of the leaves were placed on CMA-PARP (1) medium as part of our standard nursery screening, even though no lesions were seen. An organism with coralloid coenocytic hyphae, chlamydospores, and ellipsoidal semi-papillate sporangia matching the description of P. ramorum (2) grew into a snowflake-shaped colony from two pieces. On closer inspection of the leaves, small green lesions of approximately 3 to 5 mm wide were visible, especially when the leaves were backlit. For sporangial production, a 6-mm plug was transferred from the colony margin of the isolate onto V8 juice agar (V8). Sporangia, produced on V8 plugs incubated in dH20 for 2 days, were from 41 to 61 × 23 to 32 µm (48.7 × 29.3 µm average) with a length to breadth ratio from 1.3 to 2.0 (average 1.7). Chlamydospores on CMA-PARP were 36.7 to 60.1 µm (49.1 µm diameter average). From 2008 to 2011, similar symptoms were found on L. chinense from Contra Costa, San Joaquin, and Los Angeles Counties. The same organism was isolated from these infected plants. To confirm pathogenicity on L. chinense, five nursery-grown plants in 3.78-L pots were inoculated with three isolates each. Plants were inoculated with 6-mm plugs taken from the margin of a 7- to 10-day old culture grown on V8. Plant leaves were wounded with a sterile pushpin and two colonized plugs were covered with a freezer tube cap filled with sterile dH2O and attached to the underside of the leaves with a sterile pin-curl clip (4). Inoculated plants were sprayed with water, covered with plastic bags, and incubated for 2 days, when bags and plugs were removed. Four leaves per isolate were inoculated on each plant and four leaves per plant were treated similarly with uncolonized V8 plugs as a control. Plants were incubated for 12 to 14 days at 18°C (16-h photoperiod) when lesions were visible and some of the leaves began to abscise. P. ramorum grew from each lesion produced on inoculated leaves and no Phytophthora spp. grew from the control leaves when isolated onto CMA-PARP. Inoculations were repeated with similar results. The internal transcribed spacer region (ITS) of rDNA was amplified and sequenced from the isolates using ITS1 and ITS4 primers as described by White et al. (3). BLAST analysis of the sequenced amplicons (GenBank JQ361743 through JQ361745) showed 100% identity with the ITS sequence of P. ramorum (GenBank AY594198). P. ramorum is a quarantine pathogen with many hosts (2,4). Leaf spots on L. chinense caused by P. ramorum are inconspicuous and missing this disease during nursery inspections could lead to unintended spread to neighboring host plants. References: (1) S. N. Jeffers and S. B. Martin. Plant Dis. 70:1038, 1986. (2) S. Werres et al. Mycol. Res. 105:1155, 2001. (3) T. J. White et al. Page 315 in: PCR Protocols. A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990. (4) L. E. Yakabe et al. Plant Dis. 93:883, 2009.

First Report of Neofusicoccum nonquaesitum Causing Branch Cankers on Giant Sequoia (Sequoiadendron giganteum) in North America.

Between 2001 and 2007, samples from three California native plants showing canker symptoms were submitted to the California Department of Food and Agriculture's Plant Pest Diagnostics laboratory. Giant sequoia (Sequoiadendron giganteum) and coast redwood (Sequoia sempervirens) showed branch cankers and dieback, whereas tanoak (Lithocarpus densiflora) had bleeding bole cankers. Samples were collected from mature trees in private landscapes in El Dorado, Sacramento, and Alameda counties in California. A fungus was isolated on one-half strength acidified potato dextrose agar (APDA) from the canker margins of all three hosts. Colonies were moderately fast growing, initially white, later turning olivaceous black. Pycnidia developed singly or in small groups and contained conidia that measured 18 to 29 × 6 to 8 µm (average 21.5 × 6.8 µm). Conidia were aseptate, hyaline, and fusiform, with truncate bases. rDNA sequences of the internal transcribed spacer (ITS) region of the isolates (GenBank JQ282157 through JQ282159), amplified using primers ITS1 and ITS4 (2), were 100% identical to the holotype isolate of Neofusicoccum nonquaesitum Inderb., Trouillas, Bostock & Michailides (1) by a BLAST query (GenBank GU251163). Pathogenicity of the N. nonquaesitum isolate from giant sequoia (CDFA4) was tested on five saplings using cultures grown on APDA for 14 days. A single wound was made approximately 2 cm above the soil line on the cambium of each plant using a 3-mm cork borer. One 3-mm colonized agar plug was placed on each wound and secured with Parafilm. Plugs of APDA were placed onto wounds of five plants as controls. All plants were kept in a growth chamber at 23°C with a 12-h photoperiod. After 4 days, Parafilm was removed to reveal dark brown cankers measuring 12 to 43 mm long on the inoculated plants. Fourteen days after inoculation, cankers were black, sunken, and measured 79 to 117 mm (average 91.4 mm) long. Most of the inoculated plants were wilted with chlorotic to necrotic foliage. Mature pycnidia with cirri developed in most of the cankers. N. nonquaesitum was reisolated on APDA from all of the cankers. No symptoms developed on the control plants. The experiment was repeated once with similar results. Botryosphaeria dothidea, also in the Botryosphaeriaceae, has been reported to cause similar cankers on giant sequoia and coast redwood in California (3). However, rDNA sequencing of the ITS region of this isolate obtained from the American Type Culture Collection (ATCC 60344) (GenBank JQ284384) showed it matched the type specimen of Neofusicoccum australe (GenBank GU251219), not our isolate. To our knowledge, this is the first report of N. nonquaesitum as a pathogen of giant sequoia in North America. This study expands the host range of N. nonquaesitum from almond (Prunus dulcis), California bay (Umbellularia californica), and blueberry (Vaccinium spp.) (1) to include giant sequoia, coast redwood, and tanoak, which are economically important trees in California forests and landscapes. References: (1) P. Inderbitzin et al. Mycologia 102:1350, 2010. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990. (3) J. J. Worrall et al. Plant Dis. 70:757, 1986.

First Report of Embellisia allii Causing Skin Blotch and Bulb Canker on Garlic in California.

In April 2011, commercial garlic (Allium sativum) in Monterey County, CA showed symptoms of an undocumented disease. Bulb and stem sheaths were dark, decayed, and sloughing off the plants. Dissection of diseased sheaths revealed black hyphae between layers. Lower leaves wilted, turned tan, and dried up. Disease occurred in small patches scattered in two fields. In the patches, disease incidence was as much as 50%; however, overall field incidence was less than 1%. Isolations from 80% (16 of 20 plants) of collected plants resulted in the recovery of a dark olivaceous black fungus. Conidiophores were geniculate and brown and conidia were borne singly, brown, and ellipsoidal to cylindrical. Conidia had two to five but mostly three transverse septa. Longitudinal septa were infrequent and apical cells were rounded. Conidia measured (19.0-) 26.3 to 36.6 (-42.8) × (6.7-) 9.2 to 9.9 (-12.9) µm. Dark, intercalary chlamydospores were observed as colonies aged. DNA sequencing of the internal transcribed spacer (ITS) regions of four, single-spored isolates was completed with primers ITS1 and ITS4 (3). Sequences of all isolates (GenBank Nos. JN588614 to JN588617) were identical and 100% similar to Embellisia allii (AY278840). On the basis of morphological and molecular data, the fungus was identified as E. allii (Campanile) Simmons (1). Pathogenicity of four of the sequenced E. allii isolates and one additional E. allii isolate was tested using inoculum grown on acidified potato dextrose agar and garlic (cv. California Late) planted into 15-cm pots. A transverse incision was made at a point 2 cm above the garlic bulb so that a colonized agar plug could be inserted between the second and third sheath layer. The stem was then wrapped with Parafilm. Ten plants per isolate were inoculated and kept in a greenhouse (24 to 26°C). Seven to eight days after inoculation, the tissue around the incision turned tan and dark fungal growth was observed. Fourteen days after inoculation, the inoculated area was necrotic and dark fungal growth developed between stem layers. E. allii was reisolated from all inoculated plants and matched the morphological characteristics of the original isolates. Control plants, inoculated with uncolonized agar plugs, developed no symptoms. This experiment was repeated with similar results. In addition, one isolate was used to inoculate leek (A. porrum cv. Lancelot) and onion (A. cepa cv. Evergreen). Similar symptoms developed on these two species and E. allii was reisolated from all plants. To our knowledge, this is the first documentation of skin blotch and bulb canker caused by E. allii on garlic in California. Affected plants were of poor quality and could not be harvested. Our findings that garlic isolates of E. allii can infect leek and onion provide preliminary evidence that this pathogen is not restricted to garlic; this information may be useful to growers when considering crop rotations. E. allii has been reported on garlic in a number of places in Africa, Asia, Europe, the Middle East, and North and South America (2). The sequenced E. allii isolates are deposited in the fungal collection at the CDFA Plant Pest Diagnostics Lab (CDFA798-801). References: (1) J. C. David. Mycopathologia 116:59, 1991. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , August 8, 2011, (3) B. M. Pryor and D. M. Bigelow. Mycologia 95:1141, 2003.

First Report of Fusarium Wilt Caused by Fusarium oxysporum f. sp. passiflorae on Passion Fruit in North America.

Passiflora edulis Sims f. edulis, known as purple passion fruit, is a woody, perennial vine that is grown for its attractive two-part flower and its purple, edible fruit (4). In November 2009, passion fruit vines were collected during a regulatory nursery inspection in Santa Barbara County and submitted to the California Department of Food and Agriculture Plant Pest Diagnostics Laboratory. Nearly 100% of the plants inspected, all of which were approximately 1.25 m tall, appeared stunted, defoliated, and severely wilted. Dark brown vascular discoloration was present in the roots and lower stems of the plants. A pinkish violet Fusarium oxysporum colony containing chlamydospores, multiseptate macroconidia, and microconidia formed on monophialidic conidiophores was consistently isolated from roots and stems onto half-strength acidified potato dextrose agar (aPDA). All further experiments were done with an isolate obtained from a single conidium. A portion of the translation elongation factor gene (TEF-1α) was amplified and sequenced with primers ef1 and ef2 from our isolate (GenBank No. JF332039) (3). BLAST analysis of the 615-bp amplicon with the FUSARIUM-ID database showed 99% similarity with a F. oxysporum passion fruit isolate from Australia (NRRL 38273) (3). To confirm pathogenicity, washed roots of four-leaf stage seedlings approximately 10 cm tall were submerged in a conidial spore suspension (106 spores/ml) for 15 min. The conidial suspension was prepared by flooding 10-day-old cultures grown on aPDA medium with sterile distilled water. Seven seedlings were inoculated and planted in 10-cm2 pots and kept in a 25°C growth chamber with a 12-h photoperiod. Seven seedlings were mock inoculated with sterile water. After 3 weeks, four of the seven inoculated plants had leaves with yellow veins and discolored roots and had partially defoliated. Two of the four symptomatic plants also had brown stem cankers. F. oxysporum grew from the isolated roots and stems of all the inoculated plants. F. oxysporum did not grow from root and stem pieces from the water-dipped plants and the plants remained asymptomatic. Inoculations were repeated on plants approximately 15 cm tall with F. oxysporum growing from roots and stem pieces of all inoculated plants. Symptoms of yellow veins and root necrosis were not observed until 4 weeks after inoculation. Fusarium wilt caused by F. oxysporum f. sp. passiflorae is a significant disease of P. edulis f. edulis in Australia. The disease has also been reported in South Africa, Malaysia, Brazil, Panama, and Venezuela; but it is unclear as to whether the symptoms were caused by Fusarium wilt or Haematonectria canker (1). Banana poka (P. mollissima), P. ligularis, and P. foetida are also susceptible hosts (2). To our knowledge, this is the first report of Fusarium wilt caused by F. oxysporum f. sp. passiflorae on passion fruit in North America. Passion fruit is not commercially produced for consumption in California so the economic importance of this disease appears to be limited to nursery production and ornamental landscapes. The grower of the California nursery stated that the infected passion fruit plants had been propagated on site from seed. The source of inoculum at this nursery remains unknown. References: (1) I. H. Fischer and J. A. M. Rezende. Pest Tech. 2:1, 2008 (2) D. E. Garder. Plant. Dis. 73:476, 1989. (3) D. M. Geiser et al. Eur. J. Plant Pathol. 110:473, 2004. (4) F. W. Martin et al. Econ. Bot. 24:333, 1970.

First Report of Cercospora beticola Causing a Leaf Spot Disease on Acanthus mollis in California.

The genus Acanthus (Acanthaceae) includes ~30 herbaceous, perennial species grown for their attractive foliage and flower spikes. Between June and December 2009 the CDFA Plant Pest Diagnostics Lab in Sacramento, CA received multiple leaf spot disease samples on Acanthus spinosus and A. mollis, commonly known as bear's breeches. Samples were collected four times from two nurseries in Santa Barbara County. Disease was observed in nearly 100% of the plants inspected. Leaf spots were brown, roundish to elliptical, and 1 to 4 mm in diameter. Older spots often developed grayish centers and often coalesced, leading to large necrotic areas. Conidiophores were fasciculate, amphigenous, light brown to olivaceous, multiseptate, geniculate, and had distinctive spore scars. Conidia were hyaline, straight to slightly curved with tapered tips and truncate bases. Conidia were solitary, multiseptate (1 to 10) and 48 to 160 × 2.5 to 5 µm (average 100 × 3.9 µm). Colonies obtained from single conidial isolates were established on acidified potato dextrose agar (APDA). Morphologically, the causal agent was identified as Cercospora diantherae Ellis and Kellerm (1), a species synonymous with C. apii sensu lato (2). The C. apii sensu lato complex includes three morphologically similar taxa, C. apii, C. beticola, and C. apiicola (3). Sequence analysis of the internal transcribed spacer region from the Acanthus isolate confirmed it belongs to the C. apii complex (GenBank HQ328503). Multiplex PCR to distinguish species within the complex was also performed on the isolate (3). A 176-bp fragment was only observed in the PCR reaction containing the C. beticola primers. To confirm pathogenicity, hyphal suspensions were used to inoculate healthy leaves of A. mollis plants potted in 3.7-liter containers. Hyphal suspensions were obtained by grinding 3-week-old colonies grown on APDA with distilled water using a mortar and pestle. Both sides of healthy leaves and petioles were sprayed with ~40 ml of the suspension. Five plants were inoculated with C. beticola and five plants were sprayed with sterile water. Plants were incubated in a dew chamber for 48 h and then transferred to a 25°C growth chamber with a 12-h photoperiod. The experiment was repeated. Five days after inoculation, small necrotic leaf spots developed on the leaves. After 14 days, the spots had enlarged and the leaves began to turn yellow. Over time, the spots coalesced leading to large necrotic areas, especially along the leaf margins. Petiole spots, not seen on field samples, were seen on laboratory inoculated plants. Sporulation of C. beticola occurred within most of the spots and the pathogen was successfully reisolated from all inoculated leaves. No foliar symptoms developed on any of the control plants. Worldwide, C. beticola is a destructive pathogen of sugar beet (4), and has also been reported on a number of other plant hosts (3). To our knowledge, this is the first report of C. beticola causing a leaf spot disease on a host in the Acanthaceae family. This strain has been deposited into the culture collection at Centraalbureau voor Schimmelcultures. References: (1) C. Chupp. A Monograph of the Fungus Genus Cercospora. Ithaca, N.Y., 1953. (2) P. W. Crous and U. Braun. Mycosphaerella and Its Anamorphs 1: Names Published in Cercospora and Passalora. CBS, Utrecht, the Netherlands, 2003. (3) M. Groenwald et al. Mycologia 98:275, 2006. (4) W. W. Shane and P. S. Teng. Plant Dis. 76:812, 1992.

First Report of Embellisia hyacinthi Causing a Leaf Spot and Bulb Skin Spot Disease on Scilla peruviana in California.

The genus Scilla (Hyacinthaceae) includes more than 50 species of perennial, flowering bulbs grown in landscapes worldwide. In December 2000 and May 2009, an unknown leaf spot disease on Scilla peruviana was submitted to the California Department of Food and Agriculture Plant Pest Diagnostic Lab. Samples were collected during routine phytosanitary inspections of production fields in Santa Cruz County in 2000 and Monterey County in 2009. The disease was detected before plants flowered in one field at each location each year and appeared to have a scattered distribution. Foliar spots were large, elliptical to oblong with grayish black centers and brown margins. Yellow halos surrounded many of the spots. Examination of the bulb material revealed small necrotic patches on the outer bulb scales. A rapidly growing fungus was isolated on one-half-strength acidified potato dextrose agar (APDA) from the sporulating leaf spots and necrotic patches on the bulbs. The colonies were greenish gray and became dark olivaceous with age. Dictyospores, which formed on simple to branched, geniculate conidiophores, were oblong, fusiform or obclavate and usually had a triangular apical cell. They were initially hyaline, turning olivaceous brown with age. Conidia measured 14 to 39 × 8 to 13 µm (average 24.6 × 9.9 µm) typically with two to four (but up to seven) thick, transverse septa and one to two longitudinal septa. Morphologically, the fungus matched the description of Embellisia hyacinthi de Hoog & Miller (1,3). To confirm pathogenicity, four leaves of four S. peruviana plants were inoculated by taking colonized mycelial plugs from 2-week-old cultures and placing them in a plastic screw-cap lid filled with sterile water. The water plus mycelial plug suspension in the lid was then clipped to the adaxial side of a pushpin-wounded leaf (4). Plants were placed in a dark dew chamber at 20°C for 48 h and then moved to a growth chamber at 20°C with a 12-h photoperiod. After 48 h, the clips, caps, and plugs were removed. An equal number of control plants were wounded and mock inoculated with noncolonized APDA agar plugs and the experiment was repeated. Leaf lesions were visible 3 days after clip removal and expanded to an average of 26 × 10 mm, 14 days after inoculation. Sporulation was observed in the lesions after 5 to 7 days and the fungus was isolated from all inoculated leaves. No symptoms developed on the control leaves. DNA sequencing of the internal transcribed spacer region of the isolate (GenBank Accession No. HQ425562) using primers ITS1 and ITS4 matched the identity of E. hyacinthi (2,4). E. hyacinthi has been reported as a foliar and bulb pathogen on Hyacinthus, Freesia, and Scilla in Japan and Europe including Great Britain. Bulbs infected with E. hyacinthi are generally less sound and less valuable than noninfected bulbs (1). To our knowledge, this is the first report of the disease on S. peruviana in California. References: (1) G. S. de Hoog and P. J. Muller. Neth. J. Plant Pathol. 79:85, 1973. (2) B. Pryor and D. M. Bigelow. Mycologia 95:1141, 2003. (3) E. Simmons. Mycotaxon 17:216, 1983. (4) L. E. Yakabe et al. Plant Dis. 93:883, 2009.