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Alternaria alternata Causing Leaf Spot on Kalanchoe in Florida

Sanahuja, G., Lopez, P., Suarez, S. N., Hernández-Lauzardo, A. N., Chase, A. R.
Plant disease 2018 v.102 no.2 pp. 452
Aloe vera, Alternaria alternata, Arachis hypogaea, Aronia melanocarpa, Kalanchoe, Stevia rebaudiana, adenosinetriphosphatase, blueberries, cacti and succulents, conidia, cultivars, culture media, databases, fungi, genes, hosts, internal transcribed spacers, leaf spot, leaves, markets, microscopy, pathogens, peptide elongation factors, plant rots, plasma membrane, plastic bags, relative humidity, spraying, temperature, Africa, California, Florida, India, Korean Peninsula, Madagascar
The genus Kalanchoe has become popular in the United States in recent years with many succulent species and cultivars available. Kalanchoe (Crassulaceae family) is native to Madagascar and tropical Africa. Kalanchoe plants with many small leaf spots were submitted to the Florida Extension Plant Diagnostic Clinic in Homestead during March 2017. Individual spots were dark brown, circular to irregular, and ranged from 1 to 4 mm in diameter. Surface-disinfected tissue was plated on potato dextrose agar (PDA), and light gray to dark gray colonies grew abundantly after 7 days at 25°C under dark conditions. Microscopic observations revealed pale-brown to dark-brown conidia that were obclavate, obpyriform, or ovoid. They varied in length from 20 to 50 µm (average 32.3 ± 3.4 µm) and a width from 10 to 15 µm (10.8 ± 1.0 µm) with three to six transverse septa and zero to two longitudinal septa per conidium. The fungus was identified as an Alternaria species based on morphology (Woudenberg et al. 2013). Sequences of the complete internal transcribed spacer (ITS) region (MF174852), the gene fragment plasma membrane ATPase (MF174853), and the gene fragment elongation factor 1-alpha (EF-1a) gene (MF741188) of a representative isolate (170242) were submitted to the NCBI database. Primers used for ITS, ATPase gene, and EF-1a gene were ITS1/ITS4, ATPDF1/ATPDR1, and EF1-728F/EF1-986R, respectively. The ITS region from the representative isolate shared 99% similarity to A. alternata collected from Stevia rebaudiana in India (KJ728834); ATPase gene shared 96% similarity to A. alternata causing fruit rot of blueberry in California, U.S.A. (KJ908221; Zhu and Xiao 2015); and EF-1a gene shared 99% similarity to an isolate of A. alternata causing brown leaf spot on Aronia melanocarpa in Korea (KT355704; Kwon et al. 2016) and 99% with A. alternata ex-type on Arachis hypogaea in India (KC584634). A specimen of the A. alternata isolate 170242 fungus was deposited with the U.S. National Fungus Collections (BPI accession no. to be announced). Inoculations were performed by spraying a conidial suspension (1 × 10⁶ conidia/ml) of A. alternata isolate 170242 from 7-day-old PDA cultures in sterile water onto the adaxial leaf surface of healthy Kalanchoe cultivar Thyrsiflora plants. Inoculated and control plants were placed under 73% shade, for which temperatures ranged from 20 to 26°C and the relative humidity averaged 77%. Plastic bags were used for 24 h after inoculation to maintain high humidity for all plants. Two plants were used per treatment, and the experiment was repeated. Brown, tiny, circular leaf spots developed in 48 h after inoculation on all inoculated leaves, and spots enlarged over time. Alternaria alternata was reisolated from symptomatic leaf spots and was confirmed via morphological characteristics and molecular analysis for ITS region and EF-1a gene. No symptoms developed on the controls. Various cultivars of Kalanchoe (Purple Fendi, Thyrsiflora, Thyrsiflora Variegated, Farancacea, and Fedtschenkoiva) have been previously affected by Alternaria species causing leaf spot in south Florida (Plant Diagnostic Clinic in Homestead records) but this is the first time it has been identified to the species level. Alternaria alternata is a pathogen known to cause leaf spot and other diseases in a wide range of hosts (Rotem 1994), including other succulents such as Aloe vera (da Silva and Singh 2012). It is important to emphasize that this pathogen threatens to affect the rapidly growing succulent market.