New Insights into Esca of Grapevine: The Development of Foliar Symptoms and Their Association with Xylem Discoloration.

A new study on the development of foliar symptoms of esca was carried out from 2004 to 2006 in five mature vineyards in Aquitaine, France. Symptoms were monitored for severity and changes over time. Initial foliar symptoms were characterized by the presence of drying zones or discolorations (reddening or yellowing), which are symptoms that have also been attributed to Black Dead Arm (BDA). Then, the less-severely affected leaves persisted throughout the summer and developed into typical "tiger-stripe" symptoms of esca. The most severely symptomatic leaves fell soon after symptoms appeared. Severely diseased vines showed typical apoplectic or acute forms of esca that did not differ from the severe BDA forms. The appearance of leafsymptomatic vines increased uniformly over time, reaching a maximum incidence by the end of July. A second survey in 41 European and Lebanese vineyards showed that longitudinal discolorations were visible under the bark of 95% of the vines showing foliar esca symptoms. These wood symptoms, also previously attributed to BDA, appeared as xylem orange-brown stripes. Thus, foliar symptoms of esca showed transitory phases which overlapped with some BDA descriptions. Most of these symptoms, in the west-palearctic regions that were investigated, were commonly associated with the presence of one or several xylem discolorations.

Occurrence of Grapevine Declines and First Report of Black Dead Arm Associated with Botryosphaeria obtusa in Lebanon.

Grapevine, cultivated mostly in the Bekaa Plain, is one of the most important fruit crops in Lebanon. During July 2004, a survey was made in 11 vineyards of local table or wine grapes to evaluate the sanitary status of the grapevine industry as far as wood declines are concerned. The most common grapevine decline was esca. The two forms of the disease (mild and severe) were observed. The mild form was characterized by leaf symptoms consisting of interveinal necrotic spots with yellow or red chlorotic blotches on white and red cultivars. The severe form was characterized by dieback of one or more shoots, leaf drop, shrivelling, and drying of fruit clusters. In west Bekaa, on cv. Cabernet Sauvignon, some vines showed symptoms identical to those of Eutypa dieback such as stunted chlorotic shoots with small, distorted leaves; moreover, symptoms corresponding to black dead arm (BDA) such as wine red spots on the margins of leaves and dry spots were seen as reported earlier (1). Diseased vines of various cultivars were collected: 10 Cabernet Sauvignon (7 esca, 3 BDA, and 1 Eutypa dieback), 4 Beitamouni, 3 Carignan, 2 Teifihi, 1 Zeitouni, 1 Mourverdre, 1 Caladoc, and 1 Merlot. In wood, cross sections through the trunk were made that showed mainly central necrosis, white heart rot, brown red wood, and black spotting. Wedge-shaped lesions were the least common. Particularly for BDA, peeling off the bark revealed a brown streaking of the external wood. Isolations were made on malt agar (MA) with wood chips cut from the different necroses described above. Fungal identifications were based on morphological characteristics in comparison with French isolates after subculturing at 20 to 22°C: Fomitiporia sp. (F85-1), Phaeomoniella chlamydospora (F85-2), Eutypa lata (BX1-10, 8D, and 8F), and Botryosphaeria obtusa (F99-1). The fungus most frequently isolated from central necrosis with white heart rot was the basidiomycete Fomitiporia sp. (35% of vines). Cultures of Fomitiporia sp. on MA reached 4 to 5 cm in diameter after 2 weeks and were yellowish to brownish without conidia. P. chlamydospora (associated with esca, black goo, or Petri disease) was isolated from only 9% of vines investigated. Cultures of P. chlamydospora on MA were slow growing and reached 7 to 8 mm in diameter in the dark after 8 days. Colonies were white but became light green and later became dark green. Sporulation was abundant. E. lata (causing Eutypa dieback) was isolated from the vine of cv. Cabernet Sauvignon showing typical symptoms and from two vines showing symptoms of esca only. Two strains produced characteristic pycniospores, and all strains were identified using polymerase chain reaction (Primer Scar 10A-10B) (2). Among the saprophytic fungi isolated from the different kinds of necroses, either central, wedge-shaped, or under the bark, B. obtusa associated with BDA was found most commonly (65% of vines). Cultures of B. obtusa were gray brown with dense aerial mycelium. Pycnidia started to form after 4 to 5 days and conidia (20 to 26 × 9 to 16 µm) were dark brown when mature. These results are consistent with previous descriptions. To our knowledge, this is the first report of black dead arm in Lebanon. References: (1) P. Larignon et al. Phytopathol. Mediterr. 40:S336, 2001. (2) P. Lecomte et al. Appl. Environ. Microbiol. 66:4475, 2000.

First Report of a 16SrII Group Phytoplasma Associated with Shoot Proliferation of a Cactus (Opuntia monacantha) in Lebanon.

In October 2003, during a survey to evaluate the incidence of phytoplasma diseases in Lebanon, symptoms suggestive of phytoplasma infection in Opuntia monacantha (Haworth) were observed in Saghbine, Bekaa Valley. Symptoms were excessive stem and shoot proliferation. Three symptomatic and as well as symptomless plants were collected and analyzed for the presence of phytoplasmas. Nucleic acids were extracted from 0.5 g of shoot tissue and tested using polymerase chain reaction (PCR) with universal phytoplasma primers (fU5rU3) for partial amplification of the ribosomal 16SrDNA (4). PCR resulted in amplification of an expected 881-bp rDNA fragment from the symptomatic but not from symptomless samples. For characterization, sequence of the amplified DNA was determined (Genbank Accession No. AY939815). The sequence showed a high similarity with several isolates of the 16srII group of phytoplasmas. The highest similarity has been oserved with 16S rDNA of two isolates of cactus witches'-broom phytoplasma found in China (1) and Mexico (3) (Genbank Accession Nos. AJ293216 and AF320575, respectively) (99.8%) as well as faba bean phyllody phytoplasma (Genbank Accession No. X83432) (99.7%) and "Candidatus Phytoplasma aurantifolia" (Genbank Accession No. U15442) (99.3%). The presence of phytoplasmas was confirmed using nested-PCR with primers R16mF2/R1 and R16F2n/R2 (2). The Tru9I digestion pattern of the amplified product R16F2n/F16R2 detected in O. monacantha was identical to the digestion pattern obtained from periwinkle infected by "Ca. P. aurantifolia" (subgroup 16SrII-B) and soybean phyllody phytoplasma (subgroup 16SrII-C), but different from the Tru9I digestion pattern observed for cleome phyllody phytoplasma (subgroup 16SrII-A) and tomato big bud phytoplasma (subgroup 16SrII-E). To our knowledge, this is the first report of an infection with a phytoplasma belonging to16SrII group in Lebanon. References: (1) H. Cai et al. Plant Pathol. 51:394, 2002. (2) D. E. Gundersen and I. M. Lee. Phytopathol. Mediterr. 35:144, 1996. (3) N. E. Leyva-Lopez et al. Phytopathology. (Abstr.) 89(suppl):S45, 1999. (4) B. Schneider et al. Pages 369-380 in: Molecular and Diagnostic Procedures in Mycoplasmology. Academic Press, NY, 1995.

First Report of Grapevine "Bois Noir" Disease and a New Phytoplasma Infecting Solanaceous Plants in Lebanon.

During a 2001 survey to evaluate the incidence of phytoplasma diseases in Lebanon, samples were collected from plants showing symptoms suggestive of phytoplasmal infections. Samples were also collected from symptomless plants. Sampled hosts from the Bekaa Valley included: 3 samples of tomato (Lycopersicum esculentum), 4 samples of pepper (Capsicum annuum), 10 samples of grapevine (Vitis vinifera) cvs. Chardonnay and Alicante Bouschet; 7 samples of ornamental periwinkle (Catharantus roseus) from the Tyr area; and 4 samples of weeds (Lactucca serratia). DNA was extracted from leaf midveins of diseased and symptomless plants, and from healthy periwinkle, grapevine, tomato, and pepper plants grown in a greenhouse in France. Polymerase chain reaction (PCR) with universal primers for the amplification of phytoplasma ribosomal RNA genes (3) only produced a 1.8-kbp rDNA fragment from symptomatic samples. The amplified DNAs were analyzed by restriction fragment length polymorphism (RFLP) with several restriction enzymes and sequenced. The analysis showed extracts of diseased grapevines, and two periwinkle plants had identical rDNA sequences and restriction profiles of the stolbur cluster (4). The sequences had 98% identity with two European stolbur isolates from grapevine and periwinkle (GenBank Accession Nos. X76428 and AF248959, respectively). In grapevine, the disease induced by the stolbur phytoplasma is "bois noir." Bois noir is present in Europe where its incidence is predominant in northern vineyards and has been reported in Israel (2). To our knowledge, this is the first report of the stolbur/bois noir disease in Lebanon. In tomato and pepper, the restriction profiles and sequences of the phytoplasma rDNAs were identical. Sequencing and phylogenetic analysis indicated that the phytoplasma belonged to the clover proliferation (CP) cluster, as does the eggplant little leaf phytoplasma of solanaceous plants in Asia. They differed from the stolbur phytoplasma, known to infect solanaceaous plants in Europe. Lastly, a phytoplasma belonging to the pigeon pea witches' broom (PPWB) cluster was found in L. serratia and in some periwinkle plants. A phytoplasma of the PPWB cluster was recently shown to be responsible for an emerging lethal disease of almond trees in Lebanon (1). References: (1) E. Choueiri et al. Plant Dis. 85:802, 2001. (2) X. Daire et al. Vitis 36:53, 1997. (3) B. Schneider et al. Pages 369-380 in: Molecular and Diagnostic Procedures in Mycoplasmology. Academic Press, NY, 1995. (4) E. Seemüller et al. J. Plant Pathol. 80:3, 1998.

First Report of a Phytoplasma Disease of Almond (Prunus amygdalus) in Lebanon.

During a survey conducted in October 1999 to establish the sanitary status of stone fruits in Lebanon, almond trees with symptoms of leaf yellowing, shoot proliferation, and dieback were observed in the Bekaa region. Because such symptoms are often associated with phytoplasma infections, samples were collected for analysis by PCR using universal primers for amplification of phytoplasma ribosomal RNA genes (2). DNA was extracted from the leaf midveins and/or bark phloem tissue from nine symptomatic trees and one symptomless tree in four different orchards as well as from healthy almond trees collected in France. PCR resulted in amplification of an expected 1.8 kbp rDNA fragment from all symptomatic samples but not from the healthy or symptomless samples. For characterization, the amplified DNA was analyzed by RFLP. Even though the restriction profiles were different from those published for other phytoplasmas and in particular from those infecting almond trees in Western Europe (1), sequence analysis of the amplified DNA revealed that it belongs to the pigeon pea witches' broom cluster (PPWB) (2). This is the first report of a phytoplasma infection in Lebanon and the first report for a PPWB group phytoplasma in almond trees. References: (1) W. Jarausch et al. J. Plant Pathol. 104:17-27, 1998. (2) B. Schneider et al. 1995. Molecular and diagnostic procedures in Mycoplasmology Vol. 1, 369-380, S. Razin and J. G. Tully, eds.