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First Report of Cladosporium cladosporioides Causing Leaf Spot on Alstroemeria aurea in Brazil

Meneses, P. R., Dorneles, K. R., Belle, C., Moreira-Nuñez, V. L., Gonçalves, V., de Farias, C. R. J.
Plant disease 2018 v.102 no.9 pp. 1849
Alstroemeria aurea, Antirrhinum majus, Carya illinoinensis, Cladosporium cladosporioides, DNA, Dahlia pinnata, Lilium, Solanum lycopersicum var. lycopersicum, actin, conidia, conidiophores, control methods, culture media, flowers, fungal diseases of plants, genes, greenhouses, herbaria, hosts, internal transcribed spacers, leaf spot, leaves, oligodeoxyribonucleotides, pathogenicity, peptide elongation factors, plant pathogenic fungi, plant pathology, plastic bags, sequence analysis, sodium hypochlorite, solar radiation, spraying, texture, translation (genetics), Brazil
Alstroemeria aurea is a flowering plant that belongs to the family Alstroemeriaceae. It is commonly grown and commercialized as a flower-of-court and is appreciated for its beautiful flowers. In September 2017, symptoms of leaf spot were observed on A. aurea (cv. Puccini) at a commercial flower-production plant in Pelotas, Rio Grande do Sul state, Brazil. Initially, the symptoms appeared as small dark elliptical necrotic lesions. Some leaf spots coalesced and in severe cases covered the entire leaf surface. Symptomatic leaves were surface sterilized using1% NaClO for 1 min, placed on potato dextrose agar (PDA), and incubated at 25°C for 5 days. Colonies on PDA were olive-green to brown and showed a velvety texture. Conidiophores were pale and were smooth or sometimes verruculose (141.3 μm [60 to 250 μm] × 3.8 μm [3.2 to 4 μm]) (n = 30). Ramoconidia were 0- to 1-septate and were smooth or sometimes minutely verruculose (16 μm [12 to 30 μm] × 2.7 μm [2 to 4 μm]) (n = 30). Conidia formed in long, branched chains that were readily disarticulate, single celled, and elliptical to limoniform (4.6 μm [2 to 9 μm] × 2.2 μm [2 to 3 μm]) (n = 100). Conidia were pale- to olive-brown and smooth to verruculose. The specimens were deposited in the herbarium at Manoel Alves Oliveira Museum of Plant Pathology, Federal University of Pelotas, Brazil. To further confirm the identity, the internal transcribed spacer (ITS) regions as well as the translation elongation factor (TEF-1α) and partial actin (ACT) genes were sequenced, using primer pairs ITS1-5.8s and ITS2 for ITS, EF1-728F and EF-2 for TEF-1α, ACT512F and ACT783R for ACT, following the method described by Bensch et al. (2010). Sequences of the studied DNA regions were submitted to GenBank (accession nos.: ITS, MG775703; TEF-1α, MG775038; and ACT, MG775039). BLAST searches showed 99 to 100% identity with the existing sequences of Cladosporium cladosporioides (Fresen.) G.A. de Vries (1968) (ex-type CBS 11238) (ITS, HM148003; TEF-1α, HM148244; and ACT, HM148490). To confirm Koch’s postulates, three plants of A. aurea (cv. Puccini) were inoculated by spraying on the adaxial surface of the leaves with a spore suspension (10⁵ conidia/ml) of C. cladosporioides from a 5-day-old culture grown on PDA; three plants, with control treatments, were sprayed with sterile water. All plants were covered with plastic bags. Two days after inoculation, the bags were removed. Pathogenicity tests were conducted in a greenhouse at 25°C under natural daylight conditions. Seven days after inoculation, inoculated plants exhibited typical symptoms similar to those previously observed in the field, whereas control plants remained healthy. The experiment was performed two times. Cultures reisolated from symptomatic leaves showed the same morphological characteristics and molecular traits as those initially isolated from infected leaves in the field. On the basis of morphological characteristics and sequence analysis, the fungus was identified as C. cladosporioides (Bensch et al. 2010; Schoch 1999). Several hosts, including Carya illinoinensis, Lycopersicon esculentum, Antirrhinum majus, Dahlia pinnata, and Lilium sp., have been associated with C. cladosporioides (Farr and Rossman 2018). This is the first report of the infection of C. cladosporioides on A. aurea worldwide. This discovery is of great importance for Brazilian flower growers because the fungus is well established in a certain geographic area, and effective control measures are necessary to manage the disease.