First Report of Ilyonectria macrodidyma Associated with Black Foot Disease of Grapevine in Brazil

Cultivated grapevine (Vitis labrusca and V. vinifera) is of considerable economic importance to the Brazilian fruit industry for both fresh market consumption and for the production of wines, sparkling beverages, and juices. Black foot disease is caused by fungi of the genera Ilyonectria P. Chaverri & C. Salgado (anamorph: Cylindrocarpon Wollew.), Campylocarpon Halleen, Schroers & Crous, and Cylindrocladiella Boesew. In 2012, 4- to 40-year-old grapevines (Vitis spp.) showing reduced vigor, vascular lesions, necrotic root lesions, delayed budding, vine decline, and death were collected from seven locations at Rio Grande do Sul state, Brazil. Fungal isolations were made from root fragments and crown lesions (at least 2 cm above the bottom) on potato dextrose agar (PDA) medium added with 0.5 g L–1 streptomycin sulfate. Eight isolates were obtained and identified on the basis of morphological features and multi-gene analysis (rDNA-ITS, β-tubulin, and histone H3) as Ilyonectria macrodidyma (Halleen, Schroers & Crous) P. Chaverri & C. Salgado. One representative isolate (Cy5UFSM) was used for more detailed morphological and molecular characterization, and pathogenicity confirmation. When incubated in the dark at 20°C for 7 to 10 days, colonies of felty straw-colored mycelium (3) 4.79 cm diameter on average were observed. No sporodochia or other fruiting bodies were produced on carnation leaf agar (CLA) medium after 30 days. Microconidia that were produced after 5 weeks on spezieller nährstoffarmer agar (SNA) medium with addition of two pieces of 1 cm2 filter paper showed ovoid and ellipsoid shape (6.4 × 3.6 μm) and one-septate macroconidia (17.3 × 4.1 μm). To confirm the species, primer pairs ITS1 and ITS4 (4); Bt2a and Bt2b; and H3-1a and H3-1b (2) were used to amplify the ITS1-5.8S rRNA-ITS2, part of the β-tubulin and histone H3 genes, respectively. Sequences of these three regions showed 99, 100, and 100% of homology with I. macrodidyma, respectively. To confirm pathogenicity, 4-month-old rooted cuttings of V. labrusca cv. Bordô were inoculated by immersing them in a conidial suspension of the isolate (106 conidia ml–1) for 60 min (1). Thirty days later, inoculation was performed again by drenching the crown with 40 ml of 106 conidia ml–1 suspension to ensure infection of the roots. In the control treatment, plants were inoculated with sterile distilled water. Plants inoculated with I. macrodidyma showed necrosis of the leaf ribs, reduction in root mass, root and crown necrosis, browning of vessels, drying of shoots, and death. I. macrodidyma was re-isolated from the crown necrosis and vascular lesions, confirming Koch's postulates. To our knowledge, this is the first report of I. macrodidyma associated with black foot disease of grapevine in Brazil, which poses considerable threat to the industry unless management options are realized. References: (1) A. Cabral et al. Phytopathol. Mediterr. 51:340, 2012. (2) N. L. Glass et al. Appl. Environ. Microbiol. 61:1323, 1995. (3) R. W. Rayner. A Mycological Colour Chart. Commonwealth Mycological Institute and British Mycological Society, 1970. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of “Cylindrocarpon” pauciseptatum Associated with Black Foot Disease of Grapevine in Brazil

Plant Disease ◽

10.1094/pdis-06-13-0635-pdn ◽

2014 ◽

Vol 98 (4) ◽

pp. 567-567 ◽

Cited By ~ 1

Author(s):

R. F. dos Santos ◽

E. Blume ◽

M. F. B. Muniz ◽

S. M. Steckling ◽

G. W. Burtet ◽

...

Keyword(s):

Histone H3 ◽

Vascular Lesions ◽

Conidial Suspension ◽

First Report ◽

Rdna Its ◽

Foot Disease ◽

Histone H3 Gene ◽

Black Foot ◽

Black Foot Disease ◽

Β Tubulin

Since 1999, the decline of American grapevines (Vitis labrusca L.) has been common in Rio Grande do Sul, Brazil (1). In August 2012, V. labrusca with black foot symptoms were collected in vineyards in the Serra Gaúcha Region. Symptomatic plants had low vigor, vascular lesions, delayed budding, and decline and death of vines. Symptomatic roots had necrotic lesions and reduced biomass. Fungal isolations were made from necrotic root and crown fragments (own-rooted cultivar) on potato dextrose agar (PDA) medium amended with 0.5 g L–1 streptomycin sulfate. Putative colonies of “Cylindrocarpon” pauciseptatum Schroers & Crous were obtained from single macroconidia isolations. Two isolates were used to confirm the identity of isolated colonies: Cy12UFSM and Cy13UFSM. After incubation in the dark for 10 days at 20°C, the isolated mycelial colonies, which were cottony white to felty in texture, became dark orange to brown. Both isolates produced chlamydospores in chains at 40 days. Chlamydospores of Cy12UFSM and Cy13UFSM were 9 to 12 μm and 5 to 11.5 μm in diameter. Sporodochia formation on carnation leaf agar (CLA) medium was observed after 30 days. To encourage development of conidia, the isolates were grown on spezieller nährstoffarmer agar (SNA) medium for five weeks at 20°C with addition of two pieces of 1 cm2 filter paper. Microconidia of Cy12UFSM were 4 to 8.5 × 3.5 to 5 μm and those of Cy13UFSM were 3.5 to 7.5 × 3 to 5 μm. Macroconida were predominantly 3-septate (Cy12UFSM was 36 to 45 × 7.5 to 9 μm and Cy13UFSM was 30 to 38 × 7.5 to 8 μm), but 1-, 2- septate macroconidia were observed. The sizes of the three spore types and colony morphology for our isolates were similar to those described by Schroers et al. (3) for “C.” pauciseptatum. To further confirm the identity of Cy12UFSM and Cy13UFSM, multi-gene DNA sequence analysis (rDNA-ITS, β-tubulin, and histone H3) was conducted using primer pairs ITS1 and ITS4 (4), Bt2a and Bt2b, and H3-1a and H3-1b (2), which amplify the ITS1-5.8S rRNA-ITS2 genes, part of the β-tubulin gene, and the histone H3 gene, respectively. Sequences of these three regions had 99, 99, and 97% similarity with references sequences of “C.” pauciseptatum (isolate Cy238; accessions ITS [JF735307]; β-tubulin [JF735435], and histone H3 [JF735582], respectively). To evaluate pathogenicity, 4-month-old rooted cuttings of V. labrusca cv. Bordô were inoculated with two isolates by immersing them in a conidial suspension (106 conidia ml–1) for 60 min. Ten single-vine replicates were used for each isolate, and 10 water-inoculated vines were included as controls. Thirty days after inoculation, vines were re-inoculated with 40 ml of a 106 conidia ml–1 suspension to ensure root infection. After 4 months, the inoculated plants had reduced root mass relative to controls (39.18% for Cy12UFSM and 18.27% for Cy13UFSM). Inoculated plants also had root and crown necrosis, vascular lesions, shoot decline, and vine mortality (60 and 80% mortality for Cy12UFSM and Cy13UFSM, respectively). All water-inoculated control plants remained symptomless. The fungi Cy12UFSM and Cy13UFSM were re-isolated from infected woody tissues, confirming Koch's postulates. To our knowledge, this is the first report of “C.” pauciseptatum associated with black foot disease of grapevine in Brazil, which may potentially impact the sustainability of grapevine nurseries and vineyard productivity. References: (1) L. R. Garrido et al. Fitopatol. Brasil. 29:548, 2004. (2) N. L. Glass et al. Appl. Environ. Microbiol. 61:1323, 1995. (3) H. J. Schoers et al. Mycol. Res. 112:82, 2008. (4) T. J. White et al. Amplification Pages 315-322 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of Campylocarpon fasciculare Causing Black Foot Disease of Grapevine in Turkey

Plant Disease ◽

10.1094/pdis-03-14-0284-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1277-1277 ◽

Cited By ~ 1

Author(s):

D. S. Akgül ◽

N. G. Savaş ◽

S. Önder ◽

S. Özben ◽

S. Kaymak

Keyword(s):

Vitis Vinifera ◽

Dna Sequences ◽

Histone H3 ◽

Conidial Suspension ◽

Water Control ◽

Western Turkey ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease ◽

Β Tubulin

Soil-borne fungal diseases have become an important problem in grapevine nurseries of the Aegean region (western Turkey) in recent years. Reduced vigor, black vascular streaking in basal ends, blackish-sunken necrotic root lesions, and young vine death were observed in 15 grapevine nurseries of Manisa city in May 2011 and 2012. To determine the causal agents, symptomatic young grapevine (Vitis vinifera cv. Sultana 7) plants (grafted on 1103 Paulsen) were collected from nurseries (8 to 10 plants from each). Symptomatic basal end tissues were surface disinfested with 95% ethanol and flame sterilized. The internal tissues were plated onto potato dextrose agar amended with tetracycline (0.01%). Campylocarpon-like fungi were isolated (with 37.9% isolation frequency) from only one nursery (corresponding to 6.7% of all surveyed nurseries). Fungal colonies were incubated for 21 days in the dark to induce sporulation. Fungal colonies produced cottony aerial mycelium and turned chocolate-brown to dark brown on PDA. Abundant macroconidia were observed at branched conidiophores on long and cylindrical phialides. Microconidia were not observed. Macroconidia were generally 2 to 4 septate, cylindrical and slightly curved, with the following dimensions: 2 septate: 33.5 to 40.7 × 6.1 to 7.6 μm (mean: 35.9 × 6.8 μm), 3 septate: 36.2 to 43.4 × 6.6 to 8.3 μm (mean: 37.3 × 7.6 μm), and 4 septate: 48.9 to 53.5 × 7.6 to 8.3 μm (mean: 50.7 × 8.0 μm). Fifty macroconidia were measured. Morphologically, the isolates resembled the published description of Campylocarpon fasciculare Schroers, Halleen & Crous (2,4). For molecular identification, fungal DNA was extracted from mycelium and ribosomal DNA fragments (ITS1, 5.8S ITS2 rDNA), β-tubulin, and histone H3 genes, amplified with ITS 4-5, Bt 2a-2b, and H3 1a-1b primers (3,5), and sequenced. Sequences were compared with those deposited in GenBank. The isolate (MBAi45CL) showed 99% similarity with Campylocarpon fasciculare isolates AY677303 (ITS), AY377225 (β-tubulin), and JF735502 (histone H3). The DNA sequences were deposited into GenBank under accessions KJ573392, KJ573393, and KJ573394 for ITS, β-tubulin, and Histone H3 genes, respectively. To fulfill Koch's postulates, pathogenicity tests were conducted under greenhouse conditions on own-rooted grapevines (Vitis vinifera) cv. Sultana 7. Plants were removed from the rooting bench and the roots were slightly trimmed and submerged in a 107 ml–1 conidial suspension of the isolate for 60 min (5). After inoculation, the rooted cuttings were planted in 1-liter bags containing a mixture of soil, peat, and sand (2:1:1, v/v/v), and maintained in the greenhouse (24°C. 16/8-h day/night, 75% RH). Ten plants were inoculated with the isolate and five plants were submerged in sterile distilled water (control). After 4 months, young vines were examined for vascular discoloration, reduced root biomass, blackish lesions, and recovery of fungal isolates. The experiment was repeated twice. Blackish-brown discoloration of xylem vessels and necrosis in the basal ends was visible in the inoculated plants but not in the control plants. The pathogen was successfully re-isolated from 69.1% of the inoculated plants. This report is important for the new studies aiming at black foot disease control in Turkey viticulture. References: (1) A. Cabral et al. Phytopathol. Mediterr. 51:340, 2012. (2) P. Chaverri et al. Stud. Mycol. 68:67, 2011. (3) N. L. Glass and G. C. Donaldson. Appl. Environ. Microbiol. 61:1323, 1995. (4) F. Halleen et al. Stud. Mycol. 50:431, 2004. (5) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of Ilyonectria robusta Associated with Black Foot Disease of Grapevine in Southern Brazil

Plant Disease ◽

10.1094/pdis-09-13-0935-pdn ◽

2014 ◽

Vol 98 (6) ◽

pp. 845-845 ◽

Cited By ~ 3

Author(s):

R. F. dos Santos ◽

E. Blume ◽

G. B. P. da Silva ◽

M. Lazarotto ◽

L. E. Scheeren ◽

...

Keyword(s):

Reference Sequence ◽

Commonwealth Mycological Institute ◽

Conidial Suspension ◽

Southern Brazil ◽

Vitis Labrusca ◽

Rooted Cuttings ◽

First Report ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease

In August 2012, symptoms of black foot disease were observed on 21-year-old grapevines (Vitis labrusca cv. Bordô; own-rooted cultivar) at Nova Pádua city, Rio Grande do Sul state, Brazil. Symptomatic plants showed reduced vigor, vascular lesions, decline and death of vines, and necrotic lesions on roots. Isolation of fungi associated with necrotic root tissue was made on potato dextrose agar (PDA) medium containing 0.5 g L−1 streptomycin sulfate. Cultures were incubated at 25°C for 7 days in darkness, and single-spore cultures were obtained from the colonies emerging from the diseased tissue. For morphological characterization, cultures were transferred to PDA and spezieller nährstoffarmer agar (SNA) medium with addition of two pieces of 1 cm2 filter paper. One representative isolate (Cy9UFSM) was used for morphological and molecular characterization and pathogenicity confirmation. After 10 days growth on PDA at 20°C in the dark, colonies were umber to chestnut in color (3), appeared cottony to felty in texture, and sporulated profusely. After 5 weeks on SNA and under dark conditions at 20°C, cultures formed macroconidia predominantly on simple conidiophores, 1 to 3 septate, with both ends slightly rounded. Macroconidia varied in size depending on the number of cells as follows: one-septate (23-) 27.7 (-31) × (4.5-) 5.8 (-7) μm; two-septate (26-) 30.1 (-34) × (5-) 5.6 (-6) μm; and three-septate (24-) 31.2 (-35) × (5-) 5.8 (-6.5) μm. Microconidia were observed and did not have a visible hilum (6-) 11.2 (-17) × (3.5-) 4.2 (-5) μm (n = 30 observations per structure). Brown, thick-walled globose to subglobose chlamydospores were produced abundantly on PDA, (8.5-) 13.8 (-17) μm. To confirm the species, primer pairs H3-1a and H3-1b (2) were used to amplify a portion the histone H3 gene. Sequence of this region showed 98% similarity with a reference sequence for Ilyonectria robusta (A.A. Hildebr.) A. Cabral & Crous (GenBank Accession No. JF735530). Thus, both morphological and molecular criteria supported identification of the strain as I. robusta. This isolate was deposited in GenBank as accession KF633172. To confirm pathogenicity, 4-month-old rooted cuttings of Vitis labrusca cv. Bordô were inoculated by immersing roots in a conidial suspension (106 ml−1) for 60 min. After inoculation, the cuttings were planted in 1-L bags containing commercial substrate (MecPlant). Thirty days later, each plant was re-inoculated by applying 40 ml of a conidial suspension (106 ml−1) to the commercial substrate. Ten single-vine replicates were used for each isolate, and 10 water-inoculated vines were included as controls. After 4 months, the inoculated plants showed a 22.5% reduction of root mass, with root and crown necrosis, browning of vessels, and 20% mortality. Control plants treated with water remained symptomless. The fungus was re-isolated from blackened tissue of wood from the basal end of rooted cuttings, thereby satisfying Koch's postulates. I. robusta was first associated with black foot disease of grapevine in Portugal in 2012 (1). To our knowledge, this is the first report in southern Brazil of I. robusta associated with black foot disease of grapevine. References: (1) A. Cabral et al. Mycol. Prog. 11:655, 2012. (2) N. L. Glass et al. Appl. Environ. Microbiol. 61:1323, 1995. (3) R. W. Rayner. A mycological colour chart. Commonwealth Mycological Institute and British Mycological Society, 1970.

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First Report of Dactylonectria alcacerensis and Dactylonectria torresensis Associated with Black Foot Disease of Grapevine in Turkey

Plant Disease ◽

10.1094/pdis-02-20-0385-pdn ◽

2020 ◽

Vol 104 (7) ◽

pp. 2027

Author(s):

N. Güngör-Savaş ◽

D. S. Akgül ◽

M. Özarslandan ◽

M. Yıldız

Keyword(s):

First Report ◽

Dactylonectria Torresensis ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease

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First Report of Ilyonectria robusta Causing Black Foot Disease of Grapevine in Spain

Plant Disease ◽

10.1094/pdis-05-18-0730-pdn ◽

2018 ◽

Vol 102 (11) ◽

pp. 2381-2381 ◽

Cited By ~ 2

Author(s):

M. P. Martínez-Diz ◽

E. Díaz-Losada ◽

J. Armengol ◽

M. León ◽

C. Berlanas ◽

...

Keyword(s):

First Report ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease

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Development of a PCRRFLP method to distinguish species within the Ilyonectria macrodidyma complex

New Zealand Plant Protection ◽

10.30843/nzpp.2014.67.5746 ◽

2014 ◽

Vol 67 ◽

pp. 151-156 ◽

Cited By ~ 4

Author(s):

M.A. Outram ◽

E.E. Jones ◽

M.V. Jaspers ◽

H.J. Ridgway

Keyword(s):

In Silico ◽

Species Complex ◽

Plant Pathogens ◽

Histone H3 ◽

Individual Species ◽

Intraspecific Polymorphism ◽

Foot Disease ◽

Histone H3 Gene ◽

Black Foot ◽

Black Foot Disease

Species within the Ilyonectria macrodidyma complex are known plant pathogens and several are implicated as the causal agents of black foot disease of grapevines The seven species within the complex can be identified by DNA sequencing of the histone H3 gene In this study a PCRRFLP method to identify the species was developed In silico digestion of the 500 bp histone H3 amplimer using MnlI showed that it could identify Ilyonectria sp 1 Ilyonectria sp 2 I alcacerensis and I macrodidyma Subsequent in silico digestion with Hinf1 identified I estremocensis I novozelandica and I torresensis The PCRRFLP was validated using a collection of 40 I macrodidyma complex isolates that had been recovered from symptomatic grapevines Ilyonectria macrodidyma I novozelandica and I torresensis were detected in that collection Intraspecific polymorphism was only detected in I torresensis This method provides a rapid procedure for identifying individual species of the I macrodidyma species complex

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First Report of Ilyonectria robusta Associated with Black Foot Disease of Grapevine in the United States

Plant Disease ◽

10.1094/pdis-01-19-0042-pdn ◽

2019 ◽

Vol 103 (7) ◽

pp. 1788-1788

Author(s):

R. Guggenheim ◽

A. Oropeza ◽

S. Keith ◽

R. Maggard ◽

J. W. Woodhall

Keyword(s):

United States ◽

The United States ◽

First Report ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease

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First Report of Black Foot Disease of Grapevine Caused by Cylindrocarpon macrodidymum in Chile

Plant Disease ◽

10.1094/pdis-91-4-0470a ◽

2007 ◽

Vol 91 (4) ◽

pp. 470-470 ◽

Cited By ~ 11

Author(s):

J. Auger ◽

M. Esterio ◽

I. Pérez

Keyword(s):

Vascular Tissue ◽

Disease Incidence ◽

Phylogenetic Analyses ◽

Abnormal Development ◽

Vitis Vinifera L ◽

First Report ◽

Vascular Elements ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease

Black foot disease, caused by Cylindrocarpon macrodidymum Halleen, Schroers & Crous, is reported damaging table and wine grapes (Vitis vinifera L.) for the first time in Chile. During the summer of 2006, 2- to 5-year-old grapevines showed reduced vigor, shortened internodes, and drying and dying shoots along with abnormal development of roots with growth parallel to the soil surface, necrotic root crowns, and development of secondary roots. Internal necrosis extended from the bark to the pith in diseased parts of the plants. Other symptoms included black discoloration of the wood, gum inclusions in xylem vessels, black streaks in the vascular tissue, and reduction in root biomass, with sunken, necrotic root lesions. Eighteen Cylindrocarpon isolates were collected from roots, vascular elements, and pith tissue of grapevines cultivars (Flame Seedless, Red Globe, Thompson Seedless, Merlot, Carmenere, and Cabernet Sauvignon) from 12 locations in Chile. The isolates were identified on the basis of morphological features. All isolates produced micro- and macroconidia (one to three septa) and chlamydospores in short and intercalary chains (1,4), and by internal transcribed spacer (ITS1-5,8S-ITS4) rDNA and β -tubulin (BT1, and BT2) partial sequences, identical to those of C. macrodidymum (isolate USS074, GenBank Accession No. AY 997558 and isolate USSO150, GenBank Accession No. AY 997598) (2). Phylogenetic analyses placed these isolates in a clade closely related, but clearly distinct from other clades, to C. destructans and C. liriodendri (2,3). Pathogenicity tests were completed by drench inoculation onto 50 6-month-old rooted cuttings of ‘Red Globe’ with 25 ml of conidia suspension (106 conidia ml-1) obtained from four isolates. Ten control cuttings of ‘Red Globe’ were inoculated with an equal volume of sterile distilled water. The plants were incubated for 4 months in a controlled environment facility at 24°C. All isolates tested were pathogenic. In addition, they caused significant root rot (t-test of disease incidence, P = 0.0048) and no significant level of variation was detected between different isolates. C. macrodidymum was reisolated from the region of brown streaking in all the inoculated cuttings and was not isolated from the water-treated controls. To our knowledge, this is the first report of C. macrodidymum causing black foot disease on grapevine in Chile. References: (1) C. D. Booth. Mycol. Pap. (CMI) 104:1, 1966. (2) F. Halleen et al. Stud. Mycol. 50:431, 2004. (3) F. R. Mantiri et al. Can. J. Bot. 79:334, 2001. (4) E. Petit and W. D. Gubler. Plant Dis. 89:1051, 2005.

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Susceptibility of four grapevine rootstocks to Cylindrocladiella parva

New Zealand Plant Protection ◽

10.30843/nzpp.2013.66.5675 ◽

2013 ◽

Vol 66 ◽

pp. 249-253 ◽

Cited By ~ 4

Author(s):

D.S. Brown ◽

M.V. Jaspers ◽

H.J. Ridgway ◽

C.J. Barclay ◽

E.E. Jones

Keyword(s):

Dry Weight ◽

Conidial Suspension ◽

Sterile Water ◽

Rooted Cuttings ◽

Root Dry Weight ◽

Shoot Dry Weight ◽

Foot Disease ◽

Black Foot ◽

Black Foot Disease

The susceptibility of four common grapevine rootstocks (10114 Schwarzmann 5C and Riparia Gloire) to Cylindrocladiella parva (black foot disease) infection was assessed in a pot experiment The roots of 4monthold callused rooted cuttings were wounded in situ and inoculated with 50 ml C parva conidial suspension (106/ml) or sterile water (controls) After 6 months of growth shoot dry weight was significantly higher for control plants (242 g) than for those inoculated with C parva (165 g) but did not differ between rootstock varieties Root dry weight was not significantly affected by C parva inoculation but root dry weight of 10114 was significantly higher than other rootstocks Incidence and severity of trunk infection were significantly affected by rootstock variety being lowest in rootstock 10114 plants than other rootstocks None of the rootstocks tested was resistant to this pathogen

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First Report of Cylindrocarpon macrodidymum Associated with Black Foot Diseases of Grapevine in Turkey

Plant Disease ◽

10.1094/pdis-05-11-0393-pdn ◽

2012 ◽

Vol 96 (5) ◽

pp. 762-762 ◽

Cited By ~ 5

Author(s):

S. Özben ◽

F. Demirci ◽

K. Değirmenci ◽

S. Uzunok

Keyword(s):

Vascular Tissue ◽

Morphological Characteristics ◽

First Report ◽

Foot Diseases ◽

Vascular Elements ◽

Pathogenicity Tests ◽

Foot Disease ◽

Water Plants ◽

Black Foot ◽

Black Foot Disease

Grape (Vitis vinifera) is widely planted and is an economically important crop in Turkey for domestic consumption and export. Black foot disease, caused by Cylindrocarpon macrodidymum Halleen, Schroers & Crous, is a recently identified but worsening problem in vineyards worldwide (3,4). Symptomatic grapevines show reduced vigor, shortened internodes, small leaves with interveinal chlorosis, and necrosis frequently leading to the death of the plants (1). Roots of symptomatic grapevines exhibit black, sunken, necrotic lesions with a reduction in root biomass. Pith of affected vines is discolored (4). During the summers of 2009 and 2010, a survey was carried out in 63 vineyards (4 to 15 years old) in six locations of Ankara Province. We collected 44 samples from roots and crowns of grapevines exhibiting black foot symptoms. In cross section, extensive necrosis at the base of the trunk and brown-black spots in xylem vessels were observed, resembling those previously reported for black foot disease (2,4). Isolations were made from roots, vascular elements, and pith tissue. In this study, 26 isolates were identified as C. macrodidymum on the basis of morphological characteristics. Isolates identified as C. macrodidymum had a dark orange-brown colony color and abundant aerial mycelia when grown on potato dextrose agar. Isolates produced ellipsoid or ovoid microconidia. The macroconidia were one to three septate, straight, and cylindrical. One-septate macroconidia were 24 to 32 × 5 to 7 μm; three-septate macroconidia were 26 to 40 × 5 to 6 μm. Chlamydospores developed in short, intercalary chains. Conidiophores were simple or complex and sporodochial. Isolate identities were confirmed by sequence analysis of the ribosomal DNA internal transcribed spacer (GenBank Accession No. HM245331) with primers ITS1 and ITS4 (4). Isolates had 99% genetic identity with other isolates of C. macrodidymum present in GenBank. In pathogenicity tests, one representative isolate was used to inoculate five grapevine plants. Tests were completed by drench inoculation onto 3-month-old rooted cuttings of cv. Sultana with 25 ml of a conidia suspension (106 conidia ml–1). Controls were inoculated with an equal volume of sterile distilled water. Plants were incubated for 4 months in a controlled environment facility at 25°C. After 3 to 4 months, inoculations resulted in reduction of root mass, and C. macrodidymum was reisolated from regions of brown streaking in wood and discolored vascular tissue in all inoculated plants, fulfilling Koch's postulates. Control plants were asymptomatic and C. macrodidymum was not recovered from control plants. To our knowledge, this is the first report of the presence of C. macrodidymum causing black foot disease on grapevine in Turkey. References: (1) S. Alaniz et al. Plant Dis. 93:821, 2009. (2) F. Hallen et al. Stud. Mycol. 50:431, 2004. (3) F. Halleen et al. Phytopathol. Mediterr. 45:S55, 2006. (4) E. Petit and W. D. Gubler. Plant Dis. 89:1051, 2005.

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