First Report of Alternaria alternata Causing Black Spot on Tomato (Solarium lycopersicum) in Tongzhou, China

A leaf spot disease of tomato (Solanum lycopersicum L.) was observed during 2014–15 in the Tongzhou district of China. The disease was extensive, with 65 to 85% of fields inspected in the region being affected. Disease incidence of 70 to 90% was recorded in affected fields where severe losses in yield occurred. The symptoms began as small, circular, brown necrotic spots all over the foliage. The spots gradually enlarged in size and later became irregular in shape or remained circular with concentric rings or zones. In later stages of infection, the spots coalesced, and the leaves withered, dried, and fell from the plants (Zhang 2003). A fungus was consistently isolated on potato dextrose agar (PDA) amended with streptomycin sulfate. The fungus was transferred on potato carrot agar (PCA) medium, and six isolates were maintained. After 4 to 6 days of incubation at 24 to 25°C with a 12-h photoperiod in a growth chamber, the fungal colonies were regular and flat. The growth of the fungus was smooth. Conidiophores were 16 × 7 to 45 × 13 µm, mid to dark brown or olive brown in color. Spores were short-beaked, borne in long chains, oval and bean shaped, with 1 to 6 transverse and 0 to 2 longitudinal septa (Zhang 2003; Zhou et al. 2014). The rDNA of the internal transcribed spacer regions 1 and 2 and the 5.8S gene in the six isolates were amplified using primers ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′) and ITS4 (5′-TCCTCCGCTTATTGATATGC-3′). The nucleotide sequences of all six isolates were the same and the sequences were deposited in GenBank (accession no. KM501062). A BLAST search showed 100% identity with the strain Alternaria alternaria ALTL2 (KT900857). In addition, PCR identification for the six isolates was confirmed with the following primers (C_for/C_rev) (Gat et al. 2012), (Alt-4for/Alt-4rev) (Lawrence et al. 2012), (ACTDF1/ACTDR1), (ATPDF1/ATPDR1), (CALDF1/CALDR1) (Lawrence et al. 2013), and (gpd1/gpd2) (Berbee 1996). For pathogenicity tests, the six isolates were grown on PDA for 2 weeks and the spores were harvested in sterile distilled water. Inoculations were done using detached, surface-sterilized, and healthy tomato leaflets. A drop (5 μl each) of spore suspension (1 × 105 spores/ml) was placed on each leaflet, and 10 leaves were used for each isolate. Leaves were incubated in a growth chamber (80 to 90% relative humidity, 50 to 60 klx/m2 light intensity, with a photoperiod of 12 h). After 5 days, black spots similar to the original symptoms developed on all the inoculated points for all the six isolates and A. alternata was consistently reisolated. Control leaflets inoculated with sterile distilled water remained symptomless. The experiment was performed three times. To our knowledge, this is the first report of A. alternata causing black spot on tomato in Tongzhou, China. © 2017, American Phytopathological Society. All rights reserved.