scholarly journals First report of leaf blight caused by Phytophthora ramorum on cherry laurel (Prunus laurocerasus) in Washington State, USA.

Plant Disease ◽  
2020 ◽  
Author(s):  
Marianne Elliott ◽  
Lucy Rollins ◽  
Tyler Bourret ◽  
Gary Chastagner
Keyword(s):  
The United States ◽  
Washington State ◽  
Phytophthora Ramorum ◽  
Sterile Water ◽  
Long Distance ◽  
First Report ◽  
Plant Nursery ◽  
Cherry Laurel ◽  
Zoospore Suspension ◽  
Prunus Laurocerasus

In April 2014, Phytophthora ramorum (Werres, De Cock & Man in't Veld) was recovered from symptomatic foliage of cherry laurel (Prunus laurocerasus) at an ornamental plant nursery in Washington State. Cherry laurel, also known as English laurel, is widely propagated in WA because it is commonly used in landscaping. It is invasive in forests near the urban/wildland interface in the western US and in Europe (Rusterholz et al. 2018). Given its popularity as an ornamental species, the potential of this host to spread P. ramorum is of regulatory concern due to possible long distance spread to other states via nursery stock. Foliar symptoms consisted of dark brown lesions near wounds or around leaf margins where water collected. Shot-hole symptoms characterized by abscission zones and dropping of infected tissues were also observed. Lesions expanded beyond the margin of the shot-hole in some cases (Figure S1A). Phytophthora was isolated from symptomatic foliage by surface-sterilizing leaf pieces in 0.6% sodium hypochlorite and 2 rinses in sterile water. They were plated on PARP medium (Ferguson and Jeffers 1999). After 2-3 days, a slow-growing dense colony with coralloid hyphae was isolated onto V8 agar. P. ramorum was identified by observing morphological features (Figure S1B). Colony and spore morphology matched that of P. ramorum (Werres et al. 2001). The isolate was confirmed as P. ramorum by PCR and sequencing of ITS and COX1 regions using primers ITS1/ITS4 (White et al. 1990) and COX1F1/COX1R1 (Van Poucke et al. 2012). Sequences were submitted to GenBank (accession nos. ITS MT031969, COX1 MT031968). BLAST results showed at least 99% similarity with sequences of P. ramorum (ITS, KJ755124 [100%]; COX1, EU124926 [99%]). Multilocus genotyping with microsatellite markers placed the isolate in the EU1 clonal lineage. Pathogenicity of P. ramorum on cherry laurel was confirmed by completing Koch's Postulates using the isolate taken from this host. Two trials were done in a biocontainment chamber (USDA-APHIS permit # 65857) since P. ramorum is a quarantine pathogen and greenhouse trials could not be conducted, using detached stems from mature, visibly healthy cherry laurel plants growing in a landscape. Phytophthora ramorum inoculum was grown on V8A plates at 20®C for 2 weeks until sporangia were abundant. A zoospore suspension was produced by flooding plates with 7 ml sterile water, incubating for 2 hours at 5®C, then 1 hour at 24®C. Zoospores were observed with light microscopy, quantified with a hemocytometer and diluted to 1 x 104 zoospores/ml. A 10 µl droplet was placed at 3 wounded and 3 unwounded sites on 4 leaves per branch. In addition, a set of samples was inoculated by dipping foliage into the zoospore suspension for 30 seconds. A set of controls was mock inoculated using sterile water. Four branches per inoculation treatment were used and the trial was repeated once. Inoculated plant materials were incubated in moist chambers for 3-5 days at 20®C. Free moisture was present on foliage upon removal. Symptom development was assessed after incubation in the biocontainment chamber at 20®C for 7 days (Figure S1C). Phytophthora ramorum was reisolated from symptomatic tissue and the recovered culture was verified morphologically and by PCR and sequencing. It was isolated more often from foliage dipped in zoospore suspension than droplet inoculated, and more from wounded than unwounded sites. None of the water-inoculated controls were positive for P. ramorum. The presence of P. ramorum was also confirmed with DNA extraction from surface-sterilized symptomatic foliage followed by PCR and sequencing of the COX1 gene (EU124926, 100%) (Figure S2). To our knowledge, this is the first report of P. ramorum naturally infecting cherry laurel in the United States. Acknowledgements This work was supported by the USDA National Institute of Food and Agriculture, McIntire-Stennis project 1019284 and USDA APHIS Cooperative Agreement AP17PPQS&T00C070 Literature cited Ferguson and Jeffers, 1999. Plant Disease 83:1129-1136 Van Poucke, K. et al. 2012. Fungal Biology 116: 1178-1191. http://dx.doi.org/10.1016/j.funbio.2012.09.003 Werres, S. et al. 2001. Mycol. Res. 105:1155-1165. White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA.

scholarly journals First report of leaf blight caused by Phytophthora ramorum on periwinkle (Vinca minor) in Washington State, USA

Plant Disease ◽  
2021 ◽  
Author(s):  
Marianne Elliott ◽  
Lucy Rollins ◽  
Tyler Bourret ◽  
Gary Chastagner
Keyword(s):  
Ground Cover ◽  
Botanical Garden ◽  
Phytophthora Ramorum ◽  
Colony Morphology ◽  
Cox1 Gene ◽  
Sterile Water ◽  
Long Distance ◽  
Plant Materials ◽  
Food And Agriculture ◽  
Zoospore Suspension

Phytophthora ramorum (Werres, De Cock & Man in't Veld) was recovered from symptomatic foliage of periwinkle at a botanical garden in WA in March 2015. Symptoms were tan colored lesions with a dark brown margin visible on both surfaces of the leaf and were found on wounds or around leaf margins. Periwinkle is native to Europe and is commonly used for ground cover in ornamental landscapes. It is known to be invasive in US forests near the urban/wildland interface. Potential spread of P. ramorum into WA forests is of regulatory concern, as well as long distance spread to other states via nursery stock (7 CFR §301.92-2). Phytophthora ramorum was isolated from symptomatic foliage by excising leaf pieces 4-6 mm in diameter and surface-sterilizing in 0.6% sodium hypochlorite followed by two rinses in sterile water. Leaf pieces were plated on PARP medium (Ferguson and Jeffers 1999) and after 2-3 days at 20°C, slow-growing dense colonies with coralloid hyphae were isolated onto V8 agar. Colony morphology and chlamydospore production were consistent with descriptions of P. ramorum (Werres et al. 2001), except that the isolate was slower growing and had irregular, non-wildtype morphology (Elliott et al. 2018) compared to other isolates of P. ramorum. ITS and COX1 regions of mycelial DNA was amplified and sequenced to confirm the identity of P. ramorum using primers ITS1/ITS4 (White et al. 1990) and COX1F1/COX1R1 (Van Poucke et al. 2012). Sequences were submitted to GenBank (accession nos. ITS MT031975, COX1 MT031974). BLAST results showed at least 98% similarity with sequences of P. ramorum (ITS, MN540640 [98%]; COX1, EU124920 [100%]), and belonged to the NA1 clonal lineage. Pathogenicity of P. ramorum to periwinkle was confirmed by completing Koch's Postulates. Inoculum was grown on V8 agar plates at 20°C for two weeks until sporangia were abundant. A zoospore suspension was produced by flooding plates with 7 ml sterile water, incubating for 2 hours at 5°C, then for an additional hour at 24°C. Zoospores were observed under the microscope and quantified with a hemocytometer, then diluted to 2 x 105 zoospores/ml. A 10 µl droplet of inoculum was placed at one wounded and one unwounded site on six leaves on each of four plants. In addition, a set of four plants was inoculated by dipping foliage on one branch per plant into the zoospore suspension for 30 seconds. A set of four control plants were mock inoculated in the same manner using sterile water. The trial was repeated once. Inoculated plant materials were incubated in a moist chamber for 3-5 days and free moisture was present on foliage upon removal. Plants were held in a biocontainment chamber (USDA-APHIS permit # 65857) at 20C and symptom development assessed after 7 days (Figure S1). . Symptoms developed on foliage inoculated using both methods in both trials. Phytophthora ramorum was isolated once from droplet inoculated foliage at a wounded site on one plant. Reisolation onto PARP and then V8 agar was conducted from surface-sterilized symptomatic tissue and the presence of P. ramorum confirmed by observation of colony morphology and chlamydospore production. The presence of P. ramorum was also confirmed with DNA extraction from symptomatic foliage from plants from each of the two trials followed by PCR and sequencing of the COX1 gene (EU124920, 100%) (Figure S2). None of the water-inoculated controls were positive for P. ramorum. Low isolation success could be attributed to reduced pathogenicity due to being a non-wildtype isolate. Acknowledgements This work was supported by the USDA National Institute of Food and Agriculture, McIntire-Stennis project 1019284 and USDA APHIS Cooperative Agreement AP17PPQS&T00C070

scholarly journals First Report of Subanguina radicicola, the Root-Gall Nematode Infecting Poa annua Putting Greens in Washington State

Plant Disease ◽  
2007 ◽  
Vol 91 (7) ◽  
pp. 905-905 ◽  
Author(s):  
N. A. Mitkowski
Keyword(s):  
United States ◽  
New York ◽  
The United States ◽  
Washington State ◽  
Poa Annua ◽  
Golf Course ◽  
Severe Damage ◽  
Surrounding Water ◽  
Putting Greens ◽  
First Report

In the fall of 2006, a golf course in Snoqualmie, WA renovated five putting greens with commercially produced Poa annua L. sod from British Columbia, Canada. Prior to the renovation, the greens had been planted with Agrostis stolonifera L. cv. Providence, which was removed during the renovation. In February of 2007, chlorotic patches were observed on the newly established P. annua greens. When the roots were examined, extensive galling was observed throughout plant roots. Galls were slender and twisted in appearance and less than one millimeter long. Upon dissection of washed galls, hundreds of eggs were exuded into the surrounding water droplet and both mature male and female nematodes were observed. Further morphometric examination of males, females, and juvenile nematodes demonstrated that they were Subanguina radicicola (Greef 1872) Paramanov 1967 (1). Amplification of nematode 18S, ITS1, and 5.8S regions, using previously published primers (2), resulted in a 100% sequence match with the publicly available sequence for S. radicicola, GenBank Accession No. AF396366. Each P. annua plant had an average of six galls (with a range of 1 to 8), primarily located within the top 2 cm of the soil. All five new P. annua putting greens at the golf course were infested with the nematode. Additionally, P. annua from two A. stolonifera cv. Providence greens that had not been renovated was infected, suggesting that the population occurred onsite and was not imported from the Canadian sod. S. radicicola has been identified as causing severe damage in New Brunswick, Canada on P. annua putting greens and in wild P. annua in the northwestern United States, but to our knowledge, this is the first report of the nematode affecting P. annua on a golf course in the United States. References: (1) E. L. Krall. Wheat and grass nematodes: Anguina, Subanguina, and related genera. Pages 721–760 in: Manual of Agricultural Nematology. Marcel Dekker, New York, 1991. (2) N. A. Mitkowski et al. Plant Dis. 86:840, 2002.

scholarly journals First Report of Grapevine leafroll associated virus-3 in American Vitis spp. Grapevines in Washington State

Plant Disease ◽  
2006 ◽  
Vol 90 (11) ◽  
pp. 1461-1461 ◽  
Author(s):  
M. J. Soule ◽  
K. C. Eastwell ◽  
R. A. Naidu
Keyword(s):  
New York ◽  
Economic Impact ◽  
Virus Disease ◽  
The United States ◽  
Washington State ◽  
The State ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
First Report ◽  
Table Grapes

Washington State is the largest producer of juice grapes (Vitis labruscana ‘Concord’ and Vitis labrusca ‘Niagara’) and ranks second in wine grape production in the United States. Grapevine leafroll disease (GLD) is the most wide spread and economically significant virus disease in wine grapes in the state. Previous studies (2) have shown that Grapevine leafroll associated virus-3 (GLRaV-3) is the predominant virus associated with GLD. However, little is known about the incidence and economic impact of GLD on juice and table grapes. Because typical GLD symptoms may not be obvious among these cultivars, the prevalence and economic impact of GLD in Concord and Niagara, the most widely planted cultivars in Washington State, has received little attention from the grape and nursery industries. During the 2005 growing season, 32 samples from three vineyards and one nursery of ‘Concord’ and three samples from one nursery of ‘Niagara’ were collected randomly. Petiole extracts were tested by single-tube reverse transcription-polymerase chain reaction (RT-PCR; 3) with primers LC 1 (5′-CGC TAG GGC TGT GGA AGT ATT-3′) and LC 2 (5′-GTT GTC CCG GGT ACC AGA TAT-3′), specific for the heat shock protein 70 homologue (Hsp70h gene) of GLRaV-3 (GenBank Accession No. AF037268). One ‘Niagara’ nursery sample and eleven ‘Concord’ samples from the three vineyards tested positive for GLRaV-3, producing a single band of the expected size of 546 bp. The ‘Niagara’ and six of the ‘Concord’ RT-PCR products were cloned in pCR2.1 (Invitrogen Corp, Carlsbad, CA) and the sequences (GenBank Accession Nos. DQ780885, DQ780886, DQ780887, DQ780888, DQ780889, DQ780890, and DQ780891) compared with the respective sequence of a New York isolate of GLRaV-3 (GenBank Accession No. AF037268). The analysis revealed that GLRaV-3 isolates from ‘Concord’ and ‘Niagara’ share nucleotide identities of 94 to 98% and amino acid identities and similarities of 97 to 98% with the Hsp70h gene homologue of the New York isolate of GLRaV-3. Additional testing by double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) using antibodies specific to GLRaV-3 (BIOREBA AG, Reinach, Switzerland) further confirmed these results in the ‘Niagara’ and two of the ‘Concord’ isolates. GLRaV-3 has previously been reported in labrusca cvs. Concord and Niagara in western New York (4) and Canada (1), but to our knowledge, this is the first report of GLRaV-3 in American grapevine species in the Pacific Northwest. Because wine and juice grapes are widely grown in proximity to each other in Washington State and grape mealybug (Pseudococcus maritimus), the putative vector of GLRaV-3, is present in the state vineyards, further studies will focus on the role of American grapevine species in the epidemiology of GLD. References: (1) D. J. MacKenzie et al. Plant Dis. 80:955, 1996. (2) R. R. Martin et al. Plant Dis. 89:763, 2005. (3) A. Rowhani et al. ICGV, Extended Abstracts, 13:148, 2000. (4) W. F. Wilcox et al. Plant Dis. 82:1062, 1998.

scholarly journals First Report of Phytophthora hydropathica Causing Wilting and Shoot Dieback on Viburnum in Italy

Plant Disease ◽  
2014 ◽  
Vol 98 (11) ◽  
pp. 1582-1582 ◽  
Author(s):  
S. Vitale ◽  
L. Luongo ◽  
M. Galli ◽  
A. Belisario
Keyword(s):  
Natural Infection ◽  
Morphological Characteristics ◽  
Leaf Tissue ◽  
The United States ◽  
Selective Medium ◽  
Phytophthora Species ◽  
Phytophthora Ramorum ◽  
Width Ratio ◽  
Morphological Identification ◽  
First Report

The genus Viburnum comprises over 150 species of shrubs and small trees such as Laurustinus (Viburnum tinus L.), which is one of the most widely used ornamental plants in private and public gardens. Furthermore, it commonly forms stands of natural woodland in the Mediterranean area. In autumn 2012, a survey was conducted to determine the presence of Phytophthora ramorum on Viburnum in commercial nurseries in the Latium region where wilting, dieback, and death of twigs were observed on 30% of the Laurustinus plants. A Phytophthora species was consistently recovered from soil rich in feeder roots from potted Laurustinus plants showing symptoms. Soil samples were baited with rhododendron leaves. Small pieces of leaf tissue cut from the margin of lesions were plated on P5ARPH selective medium (4). Pure cultures, obtained by single-hypha transfers on potato dextrose agar (PDA), were petaloid. Sporangia formation was induced on pepper seeds (3). Sporangia were almost spherical, ovoid or obpyriform, non-papillate and non-caducous, measuring 36.6 to 71.4 × 33.4 to 48.3 μm (average 53.3 × 37.4 μm) with a length/width ratio of 1.4. Chlamydospores were terminal and 25.2 to 37.9 μm in diameter. Isolates were considered heterothallic because they did not produce gametangia in culture or on the host. All isolates examined had 30 to 35°C as optimum temperatures. Based on these morphological characteristics, the isolates were identified as Phytophthora hydropathica (2). Morphological identification was confirmed by internal transcribed spacer (ITS), and mitochondrial partial cytochrome oxidase subunit 2 (CoxII) with BLAST analysis in the NCBI database revealing 99% identity with ITS and 100% identity with CoxII. The sequences of the three isolates AB234, AB235, and AB236 were deposited in European Nucleotide Archive (ENA) with the accession nos. HG934148, HG934149, and HG934150 for ITS and HG934151, HG934152, and HG934153 for CoxII, respectively. Pathogenicity tests were conducted in the greenhouse on a total of six 1-year-old shoots cut from V. tinus plants with two inoculation points each. Mycelial plugs cut from the margins of actively growing 8-day-old cultures on PDA were inserted through the epidermis into the phloem. Controls were treated as described above except that sterile PDA plugs replaced the inoculum. Shoots were incubated in test tubes with sterile water in the dark at 24 ± 2°C. After 2 weeks, lesions were evident at the inoculation points and symptoms were similar to those caused by natural infection. P. hydropathica was consistently re-isolated from the margin of lesions, while controls remained symptomless. In the United States in 2008, P. hydropathica was described as spreading from irrigation water to Rhododendron catawbiense and Kalmia latifolia (2). This pathogen can also attack several other horticultural crops (1), but to our knowledge, this is the first report of P. hydropathica causing wilting and shoot dieback on V. tinus. References: (1) C. X. Hong et al. Plant Dis. 92:1201, 2008. (2) C. X. Hong et al. Plant Pathol. 59:913, 2010. (3) E. Ilieva et al. Eur. J. Plant Path. 101:623, 1995. (4) S. N. Jeffers and S. B. Martin. Plant Dis. 70:1038, 1986.

Estimated Economic Losses Associated with the Destruction of Plants Due to Phytophthora ramorum Quarantine Efforts in Washington State

2007 ◽  
Vol 8 (1) ◽  
pp. 20 ◽  
Author(s):  
Norman L. Dart ◽  
Gary A. Chastagner
Keyword(s):  
United States ◽  
The United States ◽  
Washington State ◽  
Phytophthora Ramorum ◽  
Economic Losses ◽  
United States Department ◽  
Department Of Agriculture ◽  
States Department ◽  
The Mean ◽  
Inspection Service

The number and retail value of plants destroyed in Washington State nurseries due to Phytophthora ramorum quarantine efforts was estimated using Emergency Action Notification forms (EANs) issued by the United States Department of Agriculture Animal and Plant Health Inspection Service between 2004 and 2005. Data collected from EANs indicate that during this period 17,266 containerized nursery plants were destroyed at 32 nurseries, worth an estimated $423,043. The mean loss per nursery was estimated at $11,188 in 2004, $11,798 in 2005, and at $13,220 per nursery over the 2-year period. Accepted for publication 26 January 2007. Published 8 May 2007.

scholarly journals First Report of Persian Buttercup Downy Mildew Caused by Peronospora sp. in Korea

Plant Disease ◽  
2013 ◽  
Vol 97 (3) ◽  
pp. 422-422
Author(s):  
Y. J. Choi ◽  
K. S. Han ◽  
J. H. Park ◽  
H. D. Shin
Keyword(s):  
Downy Mildew ◽  
The United States ◽  
Commercial Production ◽  
Ornamental Plant ◽  
Width Ratio ◽  
Sterile Water ◽  
First Report ◽  
Ranunculus Asiaticus ◽  
Length Width ◽  
Ficaria Verna

Persian buttercup (Ranunculus asiaticus L.) is an ornamental plant cultivated mainly in the countries surrounding the Mediterranean Sea, and has recently become popular in Korea. During March and April 2012, Persian buttercups ‘Elegance’ showing symptoms of downy mildew were found in plastic greenhouses in Hwaseong City of Korea. Infection resulted in chlorotic leaves with a dark greyish and dense fungal-like growth on the lower surfaces, and finally led to necrosis of the lesions. A sample was deposited in the Korea University herbarium (KUS-F26431). Conidiophores emerging from stomata were hyaline, 250 to 550 × 7 to 15 μm, straight, and dichotomously branched in 6 to 8 orders. Ultimate branchlets were mostly in pairs, slightly curved, 5 to 15 μm long, and had obtuse tips. Conidia were brown, broadly ellipsoidal to subglobose or ellipsoidal, often pedicellated, and measured 24 to 33 × 20 to 27 μm with a length/width ratio of 1.15 to 1.30. Fourteen species of Peronospora have previously been described on the genus Ranunculus (2), of which P. ficariae was mostly considered the causal agent of downy mildew on Persian buttercup (1,3). The present Korean accession is morphologically distinct from P. ficariae on R. ficaria (a synonym of Ficaria verna) by somewhat larger conidia with often pedicel-like ends. The nuclear ribosomal LSU and ITS regions were PCR-amplified and sequenced as described in Göker et al. (4), and the resulting sequences deposited in GenBank (Accession Nos. KC111207 and JX465737, respectively). A comparison with the GenBank sequences revealed that the present Korean pathogen differed from P. ficariae on R. ficaria at 10 of 688 characters (about 1.5%) in LSU (AF119600) and 11 of 802 characters (about 1.4%) in ITS sequences (unpublished sequence). In addition, the ITS sequence exhibits a dissimilarity of 1.5 to 2.0% from three species of Peronospora parasitic on Ranunculus; P. alpicola on R. aconitifolius (AY198271), P. illyrica on R. illyricus (AY198268), and P. ranunculi on R. acris (AY198267) and R. recurvatus (AY198269). Based on morphological and molecular distinction between P. ficariae and the Korean pathogen, we provisionally indicate this pathogen as an undetermined species of Peronospora. Pathogenicity was demonstrated by shaking diseased leaves onto the leaves of healthy Persian buttercup ‘Elegance’, incubating the plants in a dew chamber at 20°C for 24 h, and then maintaining them in a greenhouse (20 to 24°C and relative humidity 60 to 80%). After 3 to 4 days, inoculated plants developed downy mildew symptoms, from which an identical fungus was observed, thus fulfilling Koch's postulates. Control plants treated with sterile water did not develop any symptoms of downy mildew. To our knowledge, this is the first report of a downy mildew on Persian buttercup in Asia, although this disease has been found in other continental countries, such as Italy (1), New Zealand, South Africa, and the United States (3). The presence of a downy mildew on Persian buttercup in Asia can be considered as a potentially new and serious threat to commercial production of this ornamental plant. References: (1) E. Buonocore and R. Areddia. Informatore Fitopatologico 49:25, 1999. (2) O. Constantinescu. Thunbergia 15:1, 1991. (3) D. F. Farr and A. Y. Rossman. Fungal Databases, Syst. Mycol. Microbiol. Lab., Online publication, ARS, USDA, Retrieved August 4, 2012. (4) M. Göker et al. Mycol. Res. 113:308, 2009.

scholarly journals First Report of the Occurrence of Grapevine fanleaf virus in Washington State Vineyards

Plant Disease ◽  
2008 ◽  
Vol 92 (8) ◽  
pp. 1250-1250 ◽  
Author(s):  
T. Mekuria ◽  
R. R. Martin ◽  
R. A. Naidu
Keyword(s):  
Pacific Northwest ◽  
Mixed Infection ◽  
Consensus Sequence ◽  
Amino Acid Level ◽  
Pairwise Comparison ◽  
The United States ◽  
Washington State ◽  
Rt Pcr ◽  
Grapevine Fanleaf Virus ◽  
First Report

Grapevine fanleaf virus (GFLV; genus Nepovirus, family Comoviridae), responsible for fanleaf degeneration disease, is one of the most important viruses of grapevines worldwide (1). During our reconnaissance studies during 2007, dormant wood cuttings from individual grapevines of wine grape cv. Chardonnay were collected randomly from two geographically separate vineyards in eastern Washington State. Extracts made from cambial scrapings of these cuttings were tested separately for different viruses by single-tube reverse transcription (RT)-PCR using virus-specific primers. Two of the thirty-one grapevines in one vineyard tested positive for GLFV as mixed infection with Grapevine leafroll-associated virus (GLRaV)-3. In another vineyard, six of the twenty-six grapevines tested positive for GFLV as mixed infection with GLRaV-1, GLRaV-3, and Grapevine virus A (GVA) A forward primer (5′-ACCGGATTGACGTGGGTGAT, corresponding to nucleotides [nt] 2231–2250) and reverse primer (5′-CCAAAGTTGGTTTCCCAAGA, complementary to nt 2533–2552) specific to RNA-2 of GFLV-F13 isolate (GenBank Accession No. X16907) were used in RT-PCR assays for the detection of GFLV (4). Primers used for RT-PCR detection of GLRaV-1, GLRaV-2, and GVA were described in Martin et al (2) and Minafra et al (3). The RT-PCR results indicated mixed infection of GFLV with GLRaV-1, GLRaV-3, and GVA. To confirm the presence of GFLV, the 322-bp sequence representing a portion of the coat protein encoded by RNA-2 genomic segment was cloned into pCR2.1 (Invitrogen Corp., Carlsbad, CA). Amplicons obtained from six individual grapevines in the two vineyards were used for cloning. Three independent clones per amplicon were sequenced from both orientations. Pairwise comparison of these sequences showed 99 to 100% nucleotide sequence identity among themselves, indicating that GFLV isolates from the two vineyards may be identical. A comparison of the consensus sequence (GenBank Accession No. EU573307) with corresponding sequences of other GFLVs deposited in GenBank showed 89 to 91% identity at the nucleotide level and 95 to 99% identity at the amino acid level. However, mixed infection of GFLV with different viruses in the two vineyards suggests separate introduction of the planting material. ELISA with GFLV-specific antibodies further confirmed the presence of the virus in samples that were positive in RT-PCR. To our knowledge, this is the first report of GFLV in grapevines grown in the Pacific Northwest states of the United States. Further investigations are being carried out on the distribution, symptoms, molecular variability, and nematode vector transmission of GFLV. References: (1) P. Andret-Link et al. J. Plant Pathol. 86:183, 2004. (2) R. R. Martin et al. Plant Dis. 89:763, 2005. (3) A. Minafra et al. Arch. Virol. 142:417, 1997 (4) A. Rowhani et al. Phytopathology 83:749, 1993.

scholarly journals First Report of Leaf Spot of Sesame Caused by Xanthomonas sp. In the United States

Plant Disease ◽  
2012 ◽  
Vol 96 (8) ◽  
pp. 1222-1222 ◽  
Author(s):  
T. Isakeit ◽  
B. T. Hassett ◽  
K. L. Ong
Keyword(s):  
United States ◽  
Leaf Area ◽  
Its Region ◽  
The United States ◽  
Pcr Analysis ◽  
Sterile Water ◽  
First Report ◽  
Genomic Dna Isolation ◽  
Leaf Spots ◽  
Mist Chamber

In July 2010 in Texas, extensive leaf spots (10 to 30% leaf area affected) occurred on a commercial planting of sesame (Sesamum indicum L.) in Hidalgo County and to a lesser extent (1 to 5% leaf area) on leaves of several varieties in experimental trials in Colorado and Victoria Counties. The leaf spots were light to dark brown, somewhat circular, and 1 to 3 mm in diameter. A symptomatic leaf from each of three to five plants per county was sampled for isolations. Leaves were sprayed with 70% ethanol and immediately blotted dry with a paper towel. The margins of spots (2 mm2) were excised with a scalpel and placed in a drop of sterile water for 5 min. Drops were streaked on nutrient agar (NA) and incubated at 30°C. The 12 isolations consistently yielded gram-negative, rod-shaped bacteria with yellow, translucent colonies that were visible after 2 days of incubation. The DNA of 11 isolates was extracted with the Norgen (Thorold, ON) Bacterial genomic DNA isolation kit (Cat. #17900) and the ITS region was amplified by 16S uni 1330 and 23S uni 322 anti primers (1). PCR products were treated with the ZymoResearch (Irvine, CA) DNA clean & concentrator kit (Cat. #D4003) and sequenced. With the NCBI database, a BLAST search of the 1,100 bp amplicons showed 93 to 99% identity with pathovars of either Xanthomonas oryzae or X. axonopodis (GenBank Accession Nos. CP003057.1 and CP002914.1, respectively). Amplicon sequences of the sesame isolates were deposited in GenBank as Accession Nos. JQ975037 through JQ975047. The reported species on sesame is X. campestris pv. sesami (2). To fulfill Koch's postulates, potted sesame plants (var. Sesaco 25), 15 to 20 cm tall, were sprayed until runoff with a suspension of bacteria (106 to 107 CFU/ml) from a 2-day-old NA culture. All 12 isolates were evaluated, with five to seven plants per isolate. Plants were maintained in a mist chamber in a greenhouse at 27 to 30°C and 100% relative humidity. The pathogenicity trial was repeated once. Leaf spots were first seen 7 days after inoculation and were prevalent 14 days after inoculation. All 12 isolates were pathogenic. There were no symptoms on leaves sprayed with sterile water. Bacteria that produced colonies consistent with Xanthomonas were reisolated on NA from symptomatic leaves but not from controls. The identities of three isolates were reconfirmed with PCR analysis and sequencing. In 2007, more than 2,000 ha of sesame were grown in the continental United States, with 80% of that in Texas. Currently, acreage of shatter-free varieties of sesame is increasing in arid areas of Texas, Oklahoma, and Kansas. In such areas, the yield impact of this disease is likely to be minimal, except in years with above-average rainfall. To our knowledge, this is the first report of this disease in the United States. References: (1) E. R. Gonçalves and Y. B. Rosato. Int. J. Syst. Evol. Microbiol. 52:355, 2002. (2) J. M. Young et al., New Zealand J. Agric. Res. 21:153, 1978.

scholarly journals First Report of Dieback and Leaf Lesions on Rhododendron sp. Caused by Phytophthora hedraiandra in the United States

Plant Disease ◽  
2006 ◽  
Vol 90 (1) ◽  
pp. 109-109 ◽  
Author(s):  
B. W. Schwingle ◽  
J. A. Smith ◽  
R. A. Blanchette ◽  
S. Gould ◽  
B. L. Blanchette ◽  
...  
Keyword(s):  
United States ◽  
Average Length ◽  
The United States ◽  
Phytophthora Species ◽  
Phytophthora Ramorum ◽  
Width Ratio ◽  
Its Sequence ◽  
Voucher Specimen ◽  
First Report ◽  
Quercus Species

Surveys for Phytophthora ramorum in Minnesota nurseries revealed the presence of P. hedraiandra de Cock & Man in't Veld and several other Phytophthora species but not P. ramorum. Symptomatic leaf and stem tissues from diseased Rhododendron and Quercus species were cultured on PARP, a selective growth medium for Phytophthora (3). The Phytophthora isolates obtained were later identified by sequencing the internal transcribed spacer (ITS) region of the rDNA and comparing the sequences with those in GenBank using BLAST searches (1). The ITS sequences of six cultures (GenBank Accession Nos. DQ139804-DQ139809), isolated during 2003 from various Rhododendron cultivars exhibiting leaf lesions and shoot dieback, showed 100% identity with the ITS sequence of P. hedraiandra (GenBank Accession No. AY707987) (2). This is a recently described pathogenic species from the Netherlands responsible for causing leaf spots on Viburnum spp. Since the ITS sequence of P. hedraiandra differs little from that of P. cactorum (2), we verified one isolate to be P. hedraiandra by sequencing the mitochondrial cytochrome c oxidase subunit I gene (cox1) (GenBank Accession No. DQ139810). Comparison of this sequence with the P. hedraiandra voucher specimen in GenBank (Accession No. AY769115) showed 99% identity, which was the closest match. Reproductive structures were measured on V8 juice agar. The average oogonium diameter for three isolates was 29 μm with a range of 26 to 32 μm, while the average antheridium length was 13 μm (11 to 15 μm). Sporangium length and width averages on crushed hemp seeds were 32 μm (28 to 36 μm) and 26 μm (21 to 30 μm), respectively, with the average length to width ratio of 1.25 (1.23 to 1.29). Pathogenicity tests on Rhododendron cv. Mikkeli were carried out using three of our P. hedraiandra isolates. Spore suspensions of 2 × 104 zoospores per ml were used to mist-spray shoots of five, 3-year-old plants for each isolate. Five controls were mist sprayed with water. After inoculation, plants were placed in plastic bags in a dark growth chamber (22°C) for 7 days and then moved to a greenhouse. Leaf blotches and shoot dieback were apparent 5 days after inoculation, and P. hedraiandra was reisolated from those symptomatic tissues and identified by an exact match of the ITS sequence. Necrotic areas lengthened from the shoot tips to the main stems of the plants while expanding into petioles and leaves. No symptoms were observed on control plants. To our knowledge, this is the first report of P. hedraiandra in the United States as well as the first report of Koch's postulates performed with P. hedraiandra on Rhododendron cv. Mikkeli. The significance of this disease to other woody plants in nurseries or the landscape is unknown, and further study is needed to determine the host range and extent of the disease that may occur from this introduction. References: (1) S. F. Altschul et al. J. Mol. Biol. 215:403, 1990. (2) A. W. A.M de Cock and C. A. Lévesque. Stud Mycol 50:481, 2004. (3) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN, 1996.

scholarly journals First Report in Hawai'i of Xanthomonas citri pv. mangiferaeindicae Causing Bacterial Black Spot on Mangifera indica

Plant Disease ◽  
2013 ◽  
Vol 97 (9) ◽  
pp. 1244-1244 ◽  
Author(s):  
J. Yasuhara-Bell ◽  
A. S. de Silva ◽  
A. M. Alvarez ◽  
R. Shimabuku ◽  
M. Ko
Keyword(s):  
Mangifera Indica ◽  
Infected Plant ◽  
Black Spot ◽  
The United States ◽  
Negative Control ◽  
Sterile Water ◽  
Xanthomonas Citri ◽  
Mango Fruit ◽  
First Report ◽  
16S Rdna Pcr

Bacterial black spot of mango (Mangifera indica) caused by Xanthomonas citri pv. mangiferaeindicae (Xcm), is an economically important disease in tropical and subtropical areas (3). Xcm can infect a wide range of mango cultivars and induces raised, angular, black leaf lesions, sometimes with a chlorotic halo. Fruit symptoms appear as small, water-soaked spots on the lenticels that become star-shaped, erumpent, and exude an infectious gum (3). The bacterium can also cause latent infections (2). Immature mango fruit with black spots on the epidermis were collected in August 2012 from mango trees of the cvs. Raposa and Pirie at a residence in Pukalani, Hawai'i, on the island of Maui. Similar symptoms were seen on a tree of the mango cv. Common (also known as ‘Spanish’ or ‘Lahaina’) at a nearby golf course. Mango fruit with black lesions, and leaves showing black lesions with yellow halos, were collected in August 2012 from mango trees of the cv. Haden at a residence in Kaimuki, Hawai'i, on the island of O'ahu. Xanthomonas-like bacterial colonies were isolated on TZC agar. Suspect colonies were non-pigmented on YDC agar. A fruit strain of the bacterium from Maui (A6081A) and a strain from each of a fruit (A6081B) and a leaf (A6082) from O'ahu were each gram-negative, oxidative, positive for both starch and esculin hydrolysis, and negative for nitrate reduction, resulting in presumptive identification as a Xanthomonas sp. The three strains were further characterized by Microlog (Biolog, Inc. Hayward, CA), which showed the closest match with X. campestris. In addition, 16S rDNA PCR assays showed the closest match (99% similarity) with X. citri strains, and RIF marker analysis of dnaA (4) grouped the three strains with Xcm strain LMG 941 (Accession No. CAHO01000002.1). Hypersensitivity responses typical of xanthomonads were observed when these strains were infiltrated into tobacco leaves, whereas no response was observed using sterile water. Leaves of 3-week-old mango seedlings were infiltrated using 10 μl (~108 CFU/ml) of each strain suspended in sterilized water (six to eight inoculations per leaf, four leaves per plant, and three replicate plants per strain). The negative control treatments consisted of inoculation with sterile water, as well as an incompatible pathogen, X. hortorum pv. vitians (A6076), isolated from lettuce. Typical symptoms of bacterial black spot were observed for all strains assayed approximately 2 weeks after inoculation. No lesions were observed on the negative control plants. Koch's postulates were satisfied following reisolation and identification of the Xanthomonas strains from the infected plant tissues, using the biochemical and PCR methods described above. Results for strains from the two islands confirmed published descriptions of the pathogen, indicating that the pathogen causing symptoms on these mango trees is Xcm (1). Cultures and infected plant samples were sent to USDA APHIS and CPHST NPGLB facilities where this identification was confirmed. To our knowledge, this is the first report of bacterial black spot of mango in Hawai'i or anywhere in the United States. It is unknown whether this disease is a new occurrence or has not been reported previously. The origin of the primary inoculum is unknown. References: (1) B. Manicom and F. Wallis. Int. J. Syst. Bacteriol. 34:77, 1984. (2) O. Pruvost et al. Microbial Ecol. 58:928. (3) O. Pruvost et al. Plant Dis. 95:774, 2011. (4) K. Schneider et al. PLoS 6:e18496, 2011.

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