Detection of perchlorate using Ag/DMAH(+) SERS-active capture matrices.

In this communication, the fabrication of SERS-active capture matrices for the detection of perchlorate is described. The amine groups of amine-modified magnetic microparticles were used to immobilize silver colloidal particles. Once immobilized, the silver was reacted with dimethylaminoethanethiol hydrochloride (DMAH(+)Cl(-)) to form a self-assembled monolayer (SAM). The DMAH(+) SAM exhibits reasonably good selectivity for perchlorate. It was shown that calibration curves could be generated by ratioing the perchlorate peak with a DMAH(+) peak that did not change upon interaction with the perchlorate ion. Flow experiments, using Ag/DMAH(+) capture matrices held in place by a magnet, showed instantaneous response to changes in perchlorate concentration. The use of solid phase extraction (SPE) to eliminate chloride ion interference was explored.

Detection of hexavalent chromium using gold/4-(2-mercaptoethyl) pyridinium surface enhanced Raman scattering-active capture matrices.

In this communication, the fabrication of SERS-active capture matrices for the detection of hexavalent chromium is described. The amine groups of amine-modified magnetic microparticles were used to immobilize gold colloidal particles. Once immobilized, the gold was reacted with 4-(2-mercaptoethyl) pyridinium (MEP) hydrochloride to form a self-assembled monolayer (SAM). The MEP SAM exhibits great selectivity for hexavalent chromium. It was shown that calibration curves could be generated by ratioing MEP peaks that increased in intensity upon complexation with chromate with a peak that did not change. Flow experiments, using Au/MEP capture matrices held in place by a magnet, showed instantaneous response to changes in chromate concentration.

First Report of Bacterial Gall on Loropetalum chinense Caused by Pseudomonas savastanoi in the United States.

Bacterial gall symptoms were observed on Loropetalum chinense (R. Br.) Oliv. in two separate commercial nurseries in South Alabama during the spring of 2012. Limb dieback and plant death was first reported by the growers. Plants with dieback symptoms had galling and irregular dark callus formation on the lower stem and lower branches. Galls were small, 0.2 to 1 cm, inconspicuous, and in some cases girdled the stem causing breakage of the main stem. In both locations, 30 to 40% of the crop was affected. Similar symptoms have been observed on L. chinense in nursery and landscape plantings in central Alabama, North Carolina, and Georgia in previous years. Bacterial colonies were isolated from four plants representing two different locations. Isolates were recovered from surface sterilized symptomatic tissue on nutrient agar and King's medium B (KMB). All isolates were gram-negative and fluoresced blue-green under UV light after 48 h of growth at 28°C on KMB. One representative isolate from each site was identified as Pseudomonas savastanoi based on their fatty acid profiles (similarity index of 0.776; MIS-TSBA, version 4.0, MIDI Inc., Newark, DE) and LOPAT tests (2). The identity was confirmed by sequencing a 900-bp portion of the 16S rDNA gene, which revealed 98% similarity to the P. savastanoi type strain in NCBI (Accession No. AB021402). In greenhouse pathogenicity tests, eight Loropetalum liners were inoculated with a bacterial suspension (107 CFU/ml) of each of the two isolates. Plants were inoculated by injecting the suspension into the lower stem after wounding by puncturing with needles or slicing sections of the bark. Controls were inoculated with water. All plants inoculated with the bacteria developed gall symptoms in 8 weeks under 90% relative humidity at 30°C. The bacteria were reisolated from five inoculated plants. DNA was extracted from each isolate, amplified using primer pair 27F/1492R targeting the 16S rDNA gene (1), and sequenced. Sequences (900 bp) from all isolates shared 98 to 99% similarity to P. savastanoi type strain in GenBank (Accession No. AB021402). Nucleotide sequence data reported are available in GenBank under accessions JX915832 to 37. To our knowledge, this is the first report of bacterial gall of L. chinense caused by P. savastanoi in the United States. Given the increasing prevalence of this disease in South Alabama, its confirmation is a significant step toward management recommendations for growers. References: (1) D. J. Lane. 16S/23S rRNA sequencing. Page 115-175 in: Nucleic Acid Techniques in Bacterial Systematics. E. Stackebrandt and M. Goodfellow, eds. John Wiley and Sons, New York, 1991. (2) N. W. Schaad et al. Laboratory Guide for Identification of Plant Pathogenic Bacteria. 3rd ed. The American Phytopathological Society, St. Paul, MN, 2001.

Loop-Mediated Isothermal Amplification and Polymerase Chain Reaction Methods for Specific and Rapid Detection of Rhodococcus fascians.

Rhodococcus fascians is a phytopathogenic actinobacterium which causes leafy galls and other plant distortions that result in economically significant losses to nurseries producing ornamental plants. Traditional assays for detection and identification are time-consuming and laborious. We developed a rapid polymerase chain reaction (PCR) diagnostic assay based on two primer pairs, p450 and fas, which target the fasA and fasD genes, respectively, that are essential for pathogenicity. We also developed a faster, more convenient, loop-mediated isothermal amplification (LAMP) assay targeting the fasR gene, which regulates expression of virulence genes. Both assays were evaluated for sensitivity and specificity in vitro and in planta. The p450 and fas primers amplified DNA only from pure cultures of pathogenic reference isolates of R. fascians. Nonpathogenic isolates and 51 other plant-associated bacteria were not amplified. The PCR primers correctly detected pathogenic R. fascians from 73 of 75 (97%) bacterial strains isolated from naturally infected plants. The PCR assay correctly discriminated between pathogenic R. fascians and other bacteria in 132 of 139 (95%) naturally infected plants, and in 34 of 34 (100%) artificially inoculated plants. The fas primers were slightly more accurate than the p450 primers. The LAMP assay accurately detected pathogenic R. fascians in 26 of 28 (93%) naturally infected plants and did not react with 23 asymptomatic plants. The LAMP primers also amplified product for DNA extracts of 40 of 41 bacterial strains isolated from plants with leafy galls. The detection limit of both the PCR and LAMP assays was approximately 103 CFU/30-µl reaction. These new tools allow fast, reliable, and accurate detection of R. fascians in vitro and in planta. The LAMP assay in particular is a significant advancement in rapid R. fascians diagnostics, and enables those with limited laboratory facilities to confirm the presence of this pathogen in infected plants.

Longitudinal study comparing the dynamics of Clostridium difficile in conventional and antimicrobial free pigs at farm and slaughter.

Clostridium difficile is the leading cause of nosocomial diarrhea in humans and a major cause of enteritis in neonatal piglets, foals and calves. The aim of this longitudinal study was to determine and compare the prevalence, antimicrobial susceptibility, and toxinotype profiles of C. difficile isolated from pigs and their environment in the indoor conventional and outdoor antimicrobial free (ABF) production systems. Ten conventional and eight ABF cohorts of 35 pigs each and their environment were sampled at different stages of production at farm and slaughter. C. difficile prevalence in pigs was highest at the farrowing stage in both conventional (34%, 120/350) and ABF (23%, 56/244) systems, and decreased with age. This reduction in C. difficile prevalence in pigs at later stages of production mirrored the decreased prevalence in the farm environment. At slaughter, C. difficile was isolated at a low frequency from the carcasses and processing environment in both production systems. All but three isolates were resistant to ciprofloxacin (99%, 505/508), while 1.0% (5/508) and 6.0% (23/508) of isolates exhibited resistance to tetracycline and erythromycin, respectively. Toxinotype V (tcdA(+)tcdB(+)) was the predominant strain identified in both systems (conventional: 94%, 376/401; ABF: 82%, 88/107), while the rest were toxinotype XIII (tcdA(+)tcdB(+)). To conclude, we isolated antimicrobial resistant C. difficile regardless of antimicrobial use on the farm. Based on the phenotypic and genotypic similarity of C. difficile isolated in this study, we conclude that the unique production practices employed in conventional and ABF production systems have no impact on the pathogen population.

Sclerotinia Wilt of Hop (Humulus lupulus) Caused by Sclerotinia sclerotiorum in the Pacific Northwest United States.

In June 2009, wilted hop bines were observed in a yard in Marion County, OR. The wilt was associated with a stem rot that occurred ~1 m from the ground near the point where bines are tied together for horticultural purposes. Samples of affected stems were submitted to the Oregon State University Plant Clinic. White hyphae and large, black sclerotia were present on the stems, with a clear delineation between healthy and diseased tissue. The pathogen was identified as Sclerotinia sclerotiorum based on morphological characters. In June 2011, bine wilting was observed on the same farm but in a different hop yard (cv. Nugget) ~10 km from the 2009 occurrence. Affected plants had upward curled leaves with necrotic margins or wilted bines that were severed at the soil line. Wilted bines tended to have smaller diameters than bines with foliar symptoms only. Of 100 plants examined, 75% displayed some foliar symptoms and 66% had at least one bine that was wilted. Yield loss was estimated at 10 to 20% due to bine wilting before cone development. Unlike the 2009 occurrence, wilted bines did not display aerial signs of S. sclerotiorum. Rather, water-soaked lesions covered in white, cottony mycelium were apparent on affected stems 2.5 to 5 cm below the soil surface, some bearing large, irregularly shaped sclerotia. Isolations made onto potato dextrose agar yielded isolates with rapid growth rates and morphological characters consistent with S. sclerotiorum (1). DNA was extracted (2) and pathogen identity was confirmed by PCR amplification and sequencing of the internal transcribed spacer regions from isolates SS001 and SS002 as described before (4). The amplicons were sequenced bidirectionally and consensus sequences were 100% similar to S. sclerotiorum (GenBank No. AAGT01000678.1). Two nucleotide polymorphisms were present that differentiated the sequences from those of 12 S. trifoliorum accessions in GenBank that could be aligned (2). Greenhouse assays utilizing a toothpick inoculation procedure (3) were conducted to fulfill Koch's postulates. Stems of five 4-week-old hop plants of cv. Agate were pierced with a toothpick colonized with S. sclerotiorum. Five control plants were similarly inoculated with toothpicks without the fungus. Inoculated plants developed symptoms similar to those observed in the field within 11 days; four of five plants inoculated with isolate SS001 and two of five plants inoculated with isolate SS002 completely wilted. S. sclerotiorum was reisolated from all inoculated plants but not the control plants. To our knowledge, this is the first report of Sclerotinia wilt on hop in Oregon or the Pacific Northwest (1), where nearly all commercial hop production occurs in the United States. The disease appears to be localized to a limited number of yards, although given the widespread distribution and host range of S. sclerotiorum, it is plausible that the disease may occur in other yards. Recurrent outbreaks and spread of the disease among yards on the affected farm suggests that Sclerotinia wilt has the potential to become a perennial problem on hop and efforts to limit the introduction of S. sclerotiorum into other yards are warranted. References: (1) D. H. Gent. Page 32 in: Compendium of Hop Diseases and Pests. The American Phytopathological Society, St. Paul, MN, 2009. (2) E. N. Njambere et al. Plant Dis. 92:917, 2008. (3) M. L. Putnam. Plant Pathol. 53:252, 2004. (4) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.

Molecular characterization of a new tymovirus from Diascia ornamental plants.

Two tymoviruses were identified in plants of Diascia x hybrida 'Sun Chimes Coral' that exhibited chlorotic mottling and reduced growth. A strain of Nemesia ring necrosis virus (NeRNV) designated NeRNV-WA was detected in symptomatic plants; the deduced amino acid sequence is virtually identical to that of the previously reported NeRNV-Nf from Nemesia fruticosa. Sequence analysis also revealed the presence of a new tymovirus, and the entire genomic sequence of this virus was determined. The genome of 6,290 nucleotides was organized into three potential open reading frames (ORFs) typical of viruses in the genus Tymovirus. Based on sequence identity to tymovirus sequences, ORFs I to III encoded the replicase, movement protein and coat protein, respectively. Amino acid sequence identities to those of NeRNV-Nf were 84.8, 50.3 and 94.8%, respectively. The 5'-untranslated region could potentially form four hairpin structures. Secondary structure analysis of the 3'-terminus showed that the RNA can form a transfer-RNA-like structure that has an anticodon specific for histidine. Only 77.9% nucleotide identity was found when complete genomic sequences of this tymovirus from diascia and NeRNV-Nf were compared. The name Diascia yellow mottle virus (DiaYMV) is proposed for this new tymovirus.

First Report of a New Potyvirus, Tricyrtis virus Y, and Lily virus X, a Potexvirus, in Tricyrtis formosana in the United States.

Tricyrtis formosana (toad lily) is an herbaceous perennial in the family Liliaceae. Native to Asia, T. formosana is now used in the United States as an ornamental border plant in woodland and shade gardens. A T. formosana var. stolonifera plant showing chlorosis and mild mosaic symptoms obtained from a commercial grower in Columbia County, Oregon tested positive for potyvirus by ELISA using our genus Potyvirus broad spectrum reacting PTY-1 Mab (3). Electron microscopic examination of negatively stained leaf-dip preparations from symptomatic leaves showed a mixture of two sizes of flexuous rod-shaped particles, approximately 700 nm long (resembling potyviruses) and 470 nm long (resembling potexviruses). Total RNA extracts from symptomatic leaves were used in reverse transcription (RT)-PCR assays with potyvirus- or potexvirus-specific primers. The degenerate primers for the genus Potyvirus (2) direct the amplification of approximately 1,600-bp fragments from the 3' terminus of most potyviruses. Overlapping potexvirus cDNA clones were generated using degenerate genus Potexvirus replicase primers, and later, virus-specific primers in 3' RACE (4). The RT-PCR amplified fragments were cloned and sequenced. Analysis of the 1,688 nt potyvirus sequence (GenBank Accession No. AY864850) using BLAST showed highest identity with members of the Bean common mosaic virus (BCMV) subgroup of potyviruses. Pairwise amino acid comparisons of the CP region of the new potyvirus showed 78% identity to strains of Bean common mosaic necrosis virus, 77% identity with Soybean mosaic virus and Ceratobium mosaic virus, 72 to 76% identity to strains of BCMV, and only 50 to 64% identity with 54 other potyviruses. Additionally, similar pairwise analysis of the CP nucleotide sequence and 3'NCR of the new potyvirus generally revealed the same identity trend as described for the CP amino acid sequences, albeit with the highest nucleotide identities at less than 73% for CP and less than 66% for the 3'NCR. These results suggest that this virus is a new species in the genus Potyvirus (1), which we have tentatively named Tricyrtis virus Y (TrVY). BLAST analysis of the 3' terminal 3,010 nt potexvirus sequence (GenBank Accession No. AY864849) showed 89% nucleotide identity with Lily virus X (LVX). Pairwise amino acid comparisons of the putative gene products revealed 98, 95, 94 and 99% identity with LVX TGBp1, TGBp2, TGBp3-like, and CP, respectively, and 97% identity with the 108 nt 3'NCR. Homology with other members of the genus Potexvirus was less than 50% for these corresponding genes and gene products. ELISA and RT-PCR analysis for these two viruses in toad lily plants obtained from a grower in Illinois also revealed the presence of TrVY in three of seven cultivars and LVX coinfecting only one of the plants. The standard propagation method for T. formosana is plant division, which along with mechanical contact, provides efficient means for spread of both viruses. To our knowledge, this is the first description of this potyvirus and the first report of any potyvirus in T. formosana. LVX has been reported in Lilium formosanum, but to our knowledge, this is also the first report of LVX in T. formosana. References: (1) P. H. Berger et al. Potyviridae. Page 819 in: Virus Taxonomy: 8th Rep. ICTV, 2005. (2) M. A. Guaragna et al. Acta. Hortic. 722:209, 2006. (3) R. L. Jordan and J. Hammond. J. Gen. Virol. 72:1531, 1991. (4) C. J. Maroon-Lango et al. Arch. Virol. 150:1187, 2005.

Pathogenic Isolates of Rhodococcus fascians from New Hosts in the United States.

Rhodococcus fascians is important to the nursery industry due to its broad host range (68 genera) (2) and potential for horizontal gene transfer of plasmid-borne virulence genes. Since 2001, many herbaceous ornamental plants with symptoms of leafy galls or basal or axillary shoot proliferation suggestive of infection by R. fascians have been submitted to the Oregon State University Plant Clinic for diagnosis. R. fascians was isolated from symptomatic plants by placing affected tissues into saline or liquid D2 medium (1) for 30 to 120 min and then dilution plating onto D2 agar. Orange colonies were purified by dilution streaking and identified as R. fascians by substrate utilization (Biolog, Hayward, CA) and fatty acid analysis (L. Barnes, Texas A & M University, College Station, TX). Pathogenicity was confirmed by inoculation of 10 newly germinated Pisum sativum 'Laxton Progress' and 'Sugar Pod' seedlings with bacteria from 2-day-old cultures (107 CFU/ml) or water (controls). Our isolates produced shoot proliferations typical of R. fascians infection of peas, confirming pathogenicity. Control plants remained healthy. Pathogenic R. fascians isolates were associated with and isolated from eight species not previously reported as hosts: Acanthus mollis, Campanula sarastro, Heliopsis helianthoides 'Loraine Sunshine', Nemesia × 'Natalie', Hosta × 'Blue Umbrella', Verbascum 'Sierra Sunset', Veronica spicata 'Minuet' and Viola × 'Purple Showers'. References: (1) N. W. Schaad et al. Laboratory Guide to Plant Pathogenic Bacteria. The American Phytopathological Society, 2001. (2) D. Vereecke et al. Mol. Plant-Microbe Interact. 6:53, 2003.

Phlyctema vagabunda Causes Coin Canker of Ash (Fraxinus spp.) in North America.

During the spring of 2001, nursery-grown ash trees in Michigan and Ontario, Canada displayed coin cankers that were previously described (1). Cankered cultivars included Cimmaron®, ChampTree®, and Urbanite® (Fraxinus pennsylvanica), and Autumn Purple® (F. americana). Tissue from surface-sterilized cankers (0.06% sodium hypochlorite or 70% ethanol, 3 min) was placed onto one-half strength potato dextrose agar amended with 200 ppm of streptomycin sulfate (½ SPDA). Plates were incubated at 20 and 4°C in the dark. Phlyctema vagabunda (anamorph of Neofabraea alba (E.J. Guthrie) Verkley, (1999)) was isolated on the 4°C plates from most tissue pieces from all cultivars. P. vagabunda causes stem cankers of apple (Malus spp.) (1) but its pathogenicity to ash has never been demonstrated (2). Inoculations with P. vagabunda were made to green and white ash to fulfill Koch's postulates. Apple trees were also inoculated with the ash isolate. In Corvallis, OR, 2-year-old, bare-root ash trees and 15 1-year-old apple seedlings were inoculated in January, 2003. In Michigan, 10 trees each of cvs. Cimmaron® and Autumn Purple® were inoculated in December, 2002. The trees were inoculated with a wound-freezing method (3). Wounded sites received a 5-mm-diameter plug of ½ SPDA on which a 30-day-old culture derived from a single conidium of P. vagabunda AR3664 was actively growing, with sterile ½ SPDA used as a negative control. The plugs were held in place with Parafilm. Each ash in Oregon received three inoculations per stem. Six trees each of cvs. Autumn Purple®, ChampTree®, and Urbanite® and four trees of cv. Cimmeron® were inoculated with the fungus. Four additional trees of each cultivar were treated with sterile ½ SPDA (three sites per tree). Ten seedling apple trees were also inoculated with the fungus (three sites per tree) and five additional trees (five sites per tree) were inoculated with negative control plugs. In Michigan, each tree received two plugs of inoculum and two negative control plugs. All trees were evaluated 4 to 6 months after inoculation. Cankers similar to the originals formed on all ash cultivars in Oregon and Michigan; 97 of 106 inoculated wounds developed cankers on ash, and 29 of 30 inoculated wounds on Malus spp. developed cankers. P. vagabunda was recovered from cankers on each of the inoculated ash and apple trees. Five of 88 controls developed necrotic areas, but Phlyctema spp. were not recovered from any of these wounds. To our knowledge, this is the first report that P. vagabunda causes cankers on Fraxinus spp. and the first to show that Malus spp. can be infected with an ash isolate. References: (1) T. D. Gariépy et al. Mycol. Res. 107:528, 2003. (2) A. Y. Rossman et al. Plant Dis. 86:442, 2002. (3) R. Scorza and P. L. Pusey. Phytopathology 74:569, 1984.

First Report of Ash Anthracnose Caused by Discula fraxinea in Oregon.

Anthracnose on ash trees has been observed on landscape trees in western Oregon, yet there has been no formal report of the disease or its causal fungus. Anthracnose symptoms are observed annually in May and become severe by July when defoliation starts to occur. From 1989 to now, samples have been received from Benton, Josephine, and Marion counties, suggesting that ash anthracnose has been present throughout western Oregon for some time. To identify the causal agent, a fungus was isolated from acervuli on necrotic lesions on leaves of cultivated white ash (Fraxinus americana L.) trees in Benton County in July 2003. The acervuli produced hyaline, nonseptate, ellipsoid conidia 5 to 11 × 3.5 to 6 µm in diameter. The fungus was identified as Discula fraxinea (Peck) Redlin & Stack (teleomorph Gnomoniella fraxini Redlin & Stack) (2). The sequences of the internal transcribed spacer (ITS) region (GenBank Accession No. AY455814) and large subunit (GenBank Accession No. AY455818) nrDNA agreed with those of D. fraxinea from Maryland, except for three single-base substitutions and three insertions/deletions in ITS1. Ash anthracnose has been reported from the central and eastern United States and California, the prairie provinces in Canada, and recently, from British Columbia (1). A specimen (U.S. National Fungus Collections BPI 843391) and culture (Centraal Bureau voor Schimmelcultures CBS 114053) of D. fraxinea from Oregon were deposited. References: (1) D. F. Farr et al. Fungal Databases. Systematic Botany and Mycology Laboratory. On-line publication. ARS USDA, 2003. (2) S. C. Redlin and R. W. Stack. Mycotaxon 32:175, 1988.

First Report of Bean yellow mosaic virus (Pea Mosaic Strain) in Verbena × hybrida.

Verbena × hybrida is an ornamental annual used in rock gardens as an edging plant and hanging baskets. It comes in a variety of colors and grows approximately 1.5 to 2.5 cm (6 to 10 inches) high. In the spring of 2002, verbena cv. Lavender Shades plants from California showing leaf mosaic symptoms tested positive for potyvirus using an antigen-coated plate enzyme-linked immunosorbent assay with our genus Potyvirus broad spectrum reacting PTY-1 monoclonal as the detecting antibody (3). The virus was transmitted mechanically to Nicotiana benthamiana by sap inoculation from infected verbena plants. Infected tobacco showed systemic mild mosaic symptoms. Total RNA extractions from infected verbena and tobacco leaves were used in reverse transcription-polymerase chain reaction (RT-PCR) assays with generic potyvirus-specific primers that amplify highly conserved 700-bp or 1,600-bp fragments from the 3' terminus of most potyviruses. This region includes the 3' noncoding region (3'NCR) and the potyviral coat protein (CP). The PCR-amplified fragments were cloned by using standard TA cloning procedures and sequenced using dye-terminator chemistry. The cloned nucleotide and putative coat protein amino acid sequences from the infected verbena and tobacco plants were compared with the corresponding regions of other potyviruses. Amino acid comparison of the CP region of the verbena po-tyvirus showed 95 to 96% identity to four pea mosaic strains (PMV) of Bean yellow mosaic virus (BYMV), 85 to 89% identity to 20 other strains of BYMV, 74 to 76% identity with six strains of Clover yellow vein virus (CYVV), and only 50 to 64% identity with 28 other potyviruses. Pairwise comparisons among and between the CP sequences of PMV, BYMV, CYVV, and other potyviruses revealed identities of 92 to 99% for BYMV∷ BYMV, PMV∷PMV, and CYVV∷CYVV; 84 to 89% for BYMV∷ PMV, 69 to 78% for BYMV∷CYVV and PMV∷CYVV, and 50 to 64% for all other potyvirus combinations. Additionally, similar pairwise analysis of the 3'NCR of the verbena potyvirus revealed 98 to 99% identity to PMV strains, 81 to 94% to other BYMVs, 68 to 75% to CYVVs, and 52 to 64% with other potyviruses. Other 3'NCR pairwise comparisons generally revealed the same identity trend as described for the CP. Further serological analysis with our panel of BYMV-specific, BYMV-subgroup, and potyvirus cross-reactive monoclonal antibodies (3) confirmed the designation of the verbena potyvirus isolate as a pea mosaic strain of BYMV. To our knowledge this is the first confirmed report of BYMV-pea mosaic strain in Verbena (1,2). References: (1) Agdia, Inc. Positive Ornamental Plant Samples. Agdia On-line Publication, 2003. (2) A. A. Brunt et al. Verbena hybrida. Plant Viruses Online: Descriptions and Lists from the VIDE Database. Version 20. On-line publication, August 1996. (3) R. L. Jordan, and J. Hammond. J. Gen. Virol. 72:1531, 1991.

First Report of Sirococcus conigenus on Deodar Cedar in Oregon.

Cedrus deodara is a highly valued conifer widely grown as an ornamental in the Pacific Northwest and southern United States. C. deodara in the Pacific Northwest is normally problem free but occasionally is damaged by dieback of shoot tips, which has been associated with a fungus resembling Sirococcus conigenus. In February 2002, bleeding cankers were observed on 2- to 4-year-old stems of potted nursery stock of C. deodara cv. Karl Fuchs from Clackamas County, OR. Cankers were dark with indistinct margins, shallow, and up to 30 cm long. Infection appeared to have originated with small twigs that had died. Cultures isolated from discolored bark on streptomycin-amended potato dextrose agar (PDA) produced conidiomata with hyaline, fusiform, two-celled conidia typical of S. conigenus (1,3). Inter-simple sequence repeat-polymerase chain reaction fingerprints of an isolate from one of these trees were consistent with the P group of S. conigenus (mostly from hosts in Picea and Pinus spp.) (2). This isolate (02-04, ATCC MYA-2969) was used to inoculate two shoots on each of 12 3-year-old potted deodar cedars in each of two trials. Removing a needle wounded each shoot, and an agar plug colonized with mycelium was placed over the wound and held in place for 2 weeks with Parafilm. Sterile agar plugs were applied to two wounded control shoots on each tree in each trial. After 10 weeks, 25 of 48 inoculated shoots were blighted and drooped with yellow to brown needles that eventually dropped. The pathogen was reisolated from 24 of 25 symptomatic shoots but not from asymptomatic or control shoots. To our knowledge, this is the first confirmed report of S. conigenus as a pathogen of C. deodara. References: (1) P. F. Cannon and D. W. Minter. Taxon 32:572, 1983. (2) D. R. Smith et al. For. Pathol. 33:141, 2003. (3) B. Sutton. The Coelomycetes. Commonw. Mycol. Inst., Kew, Surrey, England, 1980.

Phlyctema vagabunda Isolated from Coin Canker of Ash Trees in Michigan.

The coelomycetous fungus Phlyctema vagabunda Desm. (teleomorph Neofabraea alba (E.J. Guthrie) Verkley, synonym Pezicula alba E.J. Guthrie) is associated with a serious canker disease of cultivated ash trees in Michigan. Four- to five-year-old trees of Fraxinus americana cv. Autumn Purple and F. pennsylvanica cvs. Champ Tree, Cimmaron, and Urbanite had cankers that were smooth, round, brownish yellow, approximately 2 to 4 cm in diameter with distinct reddish, cracked margins. Immersed, eventually erumpent, unilocular acervuli developed in the central portions of these cankers. The same fungus was isolated both from the conidia as well as from the margin of the canker. The internal transcribed spacer (ITS) (AY064704) and beta-tubulin (AY064702) sequences were identical to sequences identified in GenBank as Pezicula alba from apple (1), and the morphology was consistent with Phlyctema vagabunda as well (2). P. vagabunda has been studied primarily as the cause of Bull's eye canker of apple (1). P. vagabunda under its numerous synonyms has been reported on various hardwood and herbaceous hosts from temperate regions around the world, including the United States. However, it has not been reported previously on species of Fraxinus. A specimen and culture from the ash cankers in Michigan have been deposited (BPI 841384 and CBS 109875). References: (1) S. N. De Jong et al. Mycol. Res. 105:658, 2001. (2) B. C. Sutton. The Coelomycetes. CMI, Kew, Surrey, UK, 1980.

Wrist position affects loading of the dorsal scaphoid: possible effect on extrinsic scaphoid blood flow.

Cadaver studies using radial artery injection techniques were used to study the vascular supply along the dorsal ridge of the scaphoid. These revealed an intraarticular membrane between the wrist capsule and the dorsal ridge of the scaphoid through which arteriolar vessels (25-100 microm internal diameter) passed. Biomechanical tests revealed that the extensor carpi radialis brevis may apply significant pressure to the dorsal ridge of the scaphoid when the wrist is flexed. The highest pressures occurred with the wrist flexed at 60 degrees or 90 degrees and in slight (15 degrees ) ulnar deviation. The authors suggest that these vascular and biomechanical factors may contribute to the aetiology for idiopathic osteonecrosis of the scaphoid.

First Report of Garlic Rust Caused by Puccinia allii in Oregon.

In June of 2000, garlic rust caused by Puccinia allii F. Rudolphi was detected in a single commercial garlic field (Allium sativum L.) in the Willamette Valley of western Oregon. To our knowledge, this is the first report of P. allii in Oregon. The pathogen was present in low levels throughout the majority of the garlic field with little apparent effect; however, several 10 to 20 m wide lenses of severely damaged plants were observed. In the lenses, the plants were stunted and the outer leaves were prematurely senescent. Uredia were abundant and irregularly scattered on the leaves and gave the heavily infected plants an orange cast. The field was resurveyed in August and telia and uredia were collected. Urediospores were ellipsoid and measured 29 to 35 × 20 to 26 µm. Urediospore walls were generally hyaline, 1 to 2 µm thick and echinulate. Telia were black, oval to elongate, and non-erumpent to slightly erumpent. Teliospores were predominantly two-celled and measured 40 to 62 × 19 to 30 µm. The cap cells were angular to acuminate and irregular. Pedicels were generally shorter than the length of the spore. These characteristics are consistent with published descriptions of P. allii (1). P. allii was not detected in any of the 103 garlic or 113 onion seed fields enrolled in the state's phytosanitary certification programs during 2000. References: (1) G. F. Laundon and J. M. Waterson. CMI Descr. Pathog. Fungi Bact. No. 52, 1965.