First Report of Leaf Spot Disease on Spathiphyllum sp Caused by Alternaria alternata in Serbia

Abstract

Peace lily (Spathiphyllum spp.) are considered to be among the most popular ornamental plants grown commercially all over the world (Stamps and Evans 1999). A leaf spot disease of Spathiphyllum sp. was observed in commercial greenhouses (near Belgrade) in June 2015. The symptoms first appeared on lower, older leaves and the infection subsequently progressed to the younger leaves. The infection symptoms were numerous circular, small, dark brown spots on the leaf surface surrounded by intensive chlorotic areas measuring 0.2 to 1.0 cm in diameter. A small piece of leaf tissue was taken from the margin of the spot and the surface was disinfected with 3% sodium hypochlorite (NaOCl) for 30 s, rinsed with sterile distilled water, and placed onto potato dextrose agar (PDA, Difco, Detroit, MI). The PDA plates were incubated at 24°C in the dark for 5 days. To study morphological characteristics, the mycelium was transferred on V8 juice agar (Simmons 2007) at 24°C for 7 days, under cool white fluorescent bulbs in 10/14-h light/dark photoperiod. The six isolated strains developed cottony, brown olivaceous mycelium with white margins, rounded at the circumference. Sporulation of colonies was intensive and formed clear concentric zonation on the surface of the mycelium. Conidiophores were straight, septate, green to brown, and 43.2 to 58.0 × 4.0 to 4.1 µm in size. The conidia were light olivaceous to dark brown, long ellipsoid to obclavate, and measured 24.3 to 36.4 × 8 to 9.7 µm in size, with 1 to 4 transverse and 0 to 3 longitudinal septa for all isolates. Comparing all above mentioned characteristics with data of Simmons (2007), all isolates obtained from Spathiphyllum sp. were identified as Alternaria alternata. To confirm the pathogen’s identity, DNA was extracted and major allergen precursor (Alt a 1) and glyceraldehyde-3-phosphate dehydrogenase (gpd) gene regions were amplified by using primers Alt-for/Alt-rev (Hong et al. 2005) and gpd1/gpd2 (Berbee et al. 1999), respectively. The resulting amplicons were sequenced by Macrogen Inc. (Seoul, South Korea) and deposited in GenBank (accession nos. KP851750 to KP851761). BLAST analysis revealed 100% homology for Alt a 1 and 99% homology for gpd with gene sequences of A. alternata strain EGS 34-016 deposited in GenBank (AY563301 and AY278808). The pathogenicity assays of six isolates were performed by spraying healthy leaves of 2-year-old Spathiphyllum sp. plant with a spore suspension containing 1 × 105 conidia ml–1 prepared from the cultures grown on V8 agar at 24°C for 7 days. Three plants per isolate were used for pathogenicity test. The leaves sprayed with sterile distilled water were used as negative controls. Treated plants were placed in plastic boxes and incubated at 22°C for 30 days under natural daylight conditions. Koch’s postulates were conducted by reisolations from the symptomatic leaves onto V8 agar and the reisolated strains showed the same colony morphology as A. alternata. To our knowledge, this is the first report of leaf spot disease on Spathiphyllum plants caused by A. alternata in the Serbia. Leaf spots caused by A. alternata rarely kill plants, but reduce their aesthetic quality and commercial value. Therefore, further investigation of management practices is required to control this new disease.