First Report of Necrotic Leaf Spot Caused by Plectosphaerella cucumerina on Lamb's Lettuce in Southern Italy

During spring 2013, lamb's lettuce plants (Valerianella locusta) cv. Calarasi (Rijk Zwaan) in a commercial greenhouse in Sele Valley (Salerno Province, southern Italy) exhibited small, black-brown, irregular spots (1 mm ) that became necrotic, enlarged, and coalesced. The spots were mostly on the tips of leaves, and were surrounded by a yellow halo. The disease was severe under greenhouse conditions of 60 to 90% RH and maximum air temperature of 26°C, and affected up to 70% of the plants. The greenhouses covered an area of ~3,000 ha where many salad species are grown. Tissue fragments were excised from symptomatic leaves, sterilized by sequential dipping in 70% ethanol for 30 s and in 1% NaOCl for 30 s, rinsed in sterilized distilled water, and placed in 9-cm-diameter petri dishes containing potato dextrose agar (PDA) medium amended with streptomycin sulfate (0.1 g/liter). The plates were incubated at 24°C in the dark. A fungus was isolated consistently from infected leaf tissue after 4 days. Each colony was whitish to orange. Mycelium was hyaline, branched, septate, 3 to 4 μm wide, with numerous anastomosis-forming hyphal coils. Conidiophores were solitary, hyaline, smooth, thin-walled, unbranched or rarely irregularly branched. Conidiogenous cells were phialidic, determinate, discrete, smooth, solitary, and formed on hyphal coils. Phialides were aseptate or occasionally 1-septate near the base. Conidia (n = 100) were ellipsoidal, hyaline, smooth, septate or aseptate, and 6.6 ± 0.9 × 2.8 ± 0.4 μm. On the basis of morphological criteria (3), the fungus was ascribed to Plectosphaerella cucumerina (L.) Laterr. (anamorph Plectosporium tabacinum). An aliquot (50 ng) of genomic DNA extracted from mycelium of five cultures obtained by monosporic isolation on PDA was used as template for a PCR reaction with primers ITS5/ITS4, specific for the ITS 5.8S rDNA region of fungi (3). The 500-bp sequences amplified from the five isolates were identical, and the sequence of isolate Val-2 was submitted to GenBank (KF753234). Sequence analysis with BLASTn showed 100% identity of this sequence to the ITS-5.8S rDNA sequences of 11 isolates of P. cucumerina in GenBank. Three isolates were selected for pathogenicity tests on the lamb's lettuce cv. Calarasi. Before planting, seeds were surface-disinfected in 1% NaOCl and rinsed with sterilized distilled water. Plants (35 days old, 30 plants tested/isolate) were grown in 0.7-liter pots filled with a sterilized (autoclaved at 112°