First Report of Colletotrichum siamense Causing Leaf Drop and Fruit Spot of Citrus reticulata Blanco cv. Shiyue Ju in China

Citrus reticulata Blanco cv. Shiyue Ju, which produces one of China's most popular tropical fruits, is widely planted throughout southern China. In 2008, a new citrus disease was found in Zhaoqing City in Guangdong Province on about 20,000 ha. Yield losses averaged 15% on a wide range of different aged trees of C. reticulata cv. Shiyue Ju. No yield losses were observed on C. reticulata cv. Gong gan. Symptoms first appeared on young leaves as leaf lesions, which were reddish-brown, elliptical, and 2 to 5 mm in diameter. After several weeks, 20 to 70% of leaves dropped and 10 to 50% of fruits on the trees showed brown spots (5 to 40 mm in diameter). Leaves and fruit peels adjacent to and including lesions from different trees were surface disinfested with 1% sodium hypochlorite for 1 min and rinsed three times in sterile water. Then the tissues were plated on potato dextrose agar (PDA) in alternating light and dark at 28°C for 3 days. Twenty-three similar isolates of a fast-growing fungus were recovered from all samples. For identification, single-spore cultures were grown on potato dextrose agar (PDA) at 28°C. Initially, the colonies were white, but after 5 days, they became pale gray with concentric zones and greenish black beneath. No setae, acervuli, or ascocarp were observed in the PDA culture. Conidia formed in pink conidial masses, were hyaline, fusiform, straight, obtuse at the ends, sometimes slightly curved, and 14 to 20 × 4.5 to 6 μm (x¯ = 16.2 ± 1.5 × 4.9 ± 0.5, n = 100). The cultural and morphological characteristics of these isolates matched the description of Colletotrichum siamense (3), but not that of C. gloeosporioides or C. acutatum, which cause anthracnose on citrus plants (1,2). The actin, β-tubulin, CHS I, CAL, GPDH, and ITS regions of four representative isolates (GenBank KC524462, KC524463, KC524464, KC524465, KC524466, and KC524467) were identical and with almost 100% identity to those of the type specimen of C. hymenocallidis isolate CSSN3 (C. hymenocallidis is synonymous with C. siamense) (4), except for two inconsistent nucleotide bases in the GPDH gene. Four potted plants of C. reticulata cv. Shiyue Ju were used for pathogenicity tests. On each plant, 10 randomly selected leaves and four 6-month-old fruits were wound-inoculated with 20 μl of sterile water or conidial suspensions (1 × 105 conidia per ml). Plants were then maintained at 90% relative humidity with a 12-h photoperiod at 28°C. Symptoms resembling those in the field were observed on three inoculated plants after 14 days. In another similar experiment without wounding, three of 20 inoculated plants exhibited the symptoms after 14 days. Controls remained healthy throughout this period. The tests were performed three times. C. siamense was reisolated from all diseased inoculated plants, and the culture and fungus characteristics were the same as the original isolate. Thus, C. siamense was determined to be the pathogen causing leaf drop and fruit spot on C. reticulata cv. Shiyue Ju. To our knowledge, this is the first report of leaf drop and fruit spot on C. reticulata cv. Shiyue Ju caused by C. siamense. References: (1) H. Benyahia et al. Plant Pathol. 52:798, 2003. (2) N. A. Peres et al. Plant Dis. 89:784, 2005. (3) H. Prihastuti, et al. Fungal Diversity 39:89, 2009. (4) B. Weir et al. Stud Mycol. 73:115, 2012.