First report of Alternaria alternata causing leaf blight on Clausena lansium in China

Wampee (Clausena lansium) is one of the most characteristic and widely grown fruits in Guangdong Province, southern China. The fruit is suitable for processing due to its nutritional and health benefits. In April 2025, 10-15-year-old wampee plants showing leaf spots were observed in Zhanjiang City (21˚15'51''N, 110˚28'16''E). The disease incidence was greater than 20% whereas disease severity was 20% in a 1 ha field. Early symptoms on leaves appeared as small, circular, white spots (2 to 3 mm diam.) with dark margins. As the symptoms progressed, necrotic lesions expanded to produce large, dark yellow spots with irregular shapes. Ten symptomatic leaves from 10 plants were collected. Pieces of tissue (4 × 4 mm) were cut from lesion borders and surface-disinfected in 75% ethanol for 30 s then in 1% NaClO for 1 min, then rinsed three times with sterile water. Pieces were plated on potato dextrose agar (PDA) and incubated at 28°C with a 12-h photoperiod. After 5 days, ten morphologically similar fungal isolates were obtained from all 10 leaf samples, and three representative strains (HP-1, HP-2 and HP-3) were used for morphological and molecular characterization. Colonies were dark brown with cottony aerial mycelium. Conidia were solitary or in chains, dark brown, ovoid to obclavate, with 3-7 transverse and 0-3 longitudinal septa, with darker septa, usually 20-42×5-9 μm, with a beak length of 0 to 5 μm (n=50). These morphological characteristics were consistent with the description of Alternaria sp. (Simmons 2007). The rDNA internal transcribed spacer (ITS), translation elongation factor 1-alpha (tef1α), glyceraldehyde-3-phosphate dehydrogenase (gapdh), and RNA polymerase II second largest subunit (rpb2) regions were amplified with primers ITS1/ITS4, EF1-728F/EF1-986R, GDF1/GDR1 and fRPB2-5F/fRPB2-5R (Carbone and Kohn 1999). The ITS (PV915578-80), tef1α (PV929424-26), gapdh (PV929428-30) and rpb2 (PV929436-38) sequences were 100% identical to the A. alternata type strain CBS 916.96 (accession nos. AF347031, KC584634, AY278808 and JQ811951, respectively). A phylogenetic tree was generated using the concatenated sequences of ITS, tef1α, gapdh and rpb2. All three isolates clustered with A. alternata type strain CBS 916.96. Conidia suspension (105 conidia/ml) of the three isolates were inoculated onto 2-year-old wampee plants (n=10) by spraying approximately 200 μl leaf. Control leaves (n=10) were sprayed with sterile distilled water. All plants were wrapped in polyethylene bags for 24 h and incubated at 28°C in a growth chamber at 90% relative humidity. The test was conducted three times. After 7 days, all inoculated leaves showed similar symptoms to those observed in the field, whereas control leaves were asymptomatic. The fungus was consistently re-isolated from the lesions and identified as A. alternata as described above. A. alternata has been reported to cause diseases in a broad range of hosts (Sanahuja et al. 2018; Taycir et al. 2023) and Alternaria brown spot (ABS) of Rutaceae (Chung 2012; Huang et al. 2015). This is the first report of A. alternata causing leaf blight on C. lansium in China. This disease has limited fruit growth and has caused economic losses. Further studies are needed to assess the economic impact and develop management strategies for this disease.