A new destructive apple disease, causing black spots and necrotic lesions on leaves and defoliation on cvs. Gala and Golden Delicious (Malus × domestica Borkh.), was observed in August 2011 in Fengxian, Jiangsu Province, China. More than 90% of trees of those cultivars in the area were defoliated by the disease and almost no leaves were left on trees before harvest. The disease was similar to Glomerella leaf spot reported first in Brazil in 1988 (2) and in the United States in 1998 (1) on cvs. Gala and Golden Delicious. The initial symptom was small black lesions on leaves. Above 30°C, the lesions developed quickly and grew to 2 to 3 cm, with a blurred edge. Diseased leaves became dark and were shed. At lower temperatures, the black lesions stopped enlarging after 5 to 6 days and formed large necrotic spots with clear edges; these leaves gradually grew yellow and were shed. When incubated at 30°C and 100% relative humidity for 1 to 2 days, the black lesions produced a mass of saffron-yellow conidia. On fruit, the pathogen only caused circular, necrotic, sunken, red-bordered lesions 2 to 3 mm in diameter, which was different from bitter rot. Three monospored cultures were isolated from diseased leaves and new conidia were obtained from isolates. The colony, with abundant mycelium, was white but turned gray to black. Conidia were 12 to 17 × 5 to 7 μm, and were cylindrical with rounded ends. After germination, conidia formed appressoria, oval, or circular cells with black thick walls 7 to 12 × 5 to 7 um. Based on morphological characteristics, the pathogen was putatively identified as Glomerella cingulata. The conidia were inoculated in vitro on leaves of cvs. Gala and Fuji by dripping a suspension of about 104 conidia/ml of water onto upper leaf surfaces. Dark necrotic lesions were observed on all inoculated Gala leaves, which were similar to those observed in orchards, after 4 days incubation in a chamber at 30°C with 100% humidity. Only small black lesions, about 1 to 2 mm in diameter, were observed on Fuji leaves. No symptoms developed on leaves inoculated with distilled water. The internal transcribed spacer (ITS) region of ribosomal DNA and part of the 18S and 28S ribosomal RNA of the three isolates were amplified with the universal primers ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′) and ITS4 (5′-CCTCCGCTTATTGATATGC-3′). The amplified ITS sequences confirmed that the three isolates belonged to the same species, with only one base pair variation among sequences. The nucleotide sequence of isolate 1 and 2 was deposited in GenBank (JN714400 and JN714401). BLAST analysis showed that the sequence had 99% homology with the sequence of G. cingulata (EU008836), the causal agent of Glomerella leaf spot. However, the sequence of isolate 1 had 100% homology with that of G. cingulata (HQ845103.1) isolated from walnut in Shandong, China, while the sequence of isolate 2 had 100% homology with that of G. cingulata (HM015004.1) isolated from sweet pepper in Taiwan. Results suggested the disease is Glomerella leaf spot and the causal agent is G. cingulata. The disease will eliminate sensitive apple cultivars, such as Gala, from wet, warm production areas if effective control measures are not developed within a few years. To our knowledge, this was the first finding of the disease in China and will provide useful information for developing effective control strategies. References: (1) E. González and T. B. Sutton. Plant Dis. 83:1074, 1999. (2) T. B. Sutton and R. M. Sanhueza. Plant Dis. 82:267, 1998.
In China, covering developing apple (Malus × domestica) fruit with paper bags is a standard production practice. The fruit are usually covered from May to October to exclude pests and rain-dispersed pathogens and reduce pesticide residue at harvest. From 2010 to 2012, a fruit spot disease was observed on bagged fruit and caused 1 to 30% annual yield losses in most orchards in Shandong Province. Affected fruit were covered with red-brown, sunken, circular lesions 2 to 20 mm in diameter with dark violet edges often surrounded by a red halo. In many cases, the lesion cracked and pinkish mycelium was observed within the cracks. Isolations were made from bagged fruit from 12 orchards in October 2010 to 2012. Fungal isolations were made onto potato dextrose agar (PDA) medium. Two strains were consistently obtained from isolates. Strain 1 produced conidia assembled in head. Conidia were ellipsoidal to ovoid and 2.1 to 7.5 × 1.1 to 3.0 μm. Colonies were whitish with some pink and powdery on PDA. String 2 produced conidia in a long chain. Conidia were spindle-shaped with apiculate at both ends and 2.1 to 6.6 × 1.3 to 3.8 μm. Colonies were whitish at the beginning and grayish later and powdery on PDA. To further confirm the identity of the isolated fungus, the large subunit (LSU), the small subunit (SSU), and the internal transcribed spacer (ITS) sequences of ribosomal DNA, and the β-tubulin gene (β-tubulin), were amplified and sequenced with the primers V9G/LR5, NS1/NS24, ITS1/ITS4, and Bt1a/Bt1b, respectively. LSU (GenBank Accession Nos. KJ194115 and KJ194116), SSU (KJ194117 and KJ194118), ITS (KF225143 and KF225144), and β-tubulin (KF225145 and KF225146) sequences didn't have any variation between the two strains sequenced. Phylogenetic analyses of each of the examined genes indicated a high similarity (>99%) with Acremonium sclerotigenum (CBS 384.65 HQ232129). Based on the sequence data and the morphology, we identified the fungus as A. sclerotigenum (1,2). To confirm pathogenicity, a spore suspension (1 × 104 conidia per ml) was made from each of the strains isolated. Strains were subsequently inoculated on to 10 mature apple fruit by wounding them to a depth of 2 mm with an acupuncture needle. Inoculation with sterile distilled water was included as a control. Prior to inoculation, all fruit were surface-sterilized with 75% alcohol. Lesions developed on fruit inoculated with the putative pathogen 10 days after incubation in >90% humidity chamber at 25°C. The fungi that were isolated from the infected fruit were identical to the inoculated strains. No lesions developed on the control fruit. This is the first report of brown spot disease caused by A. sclerotigenum in apple and in bagged fruit production. Given that brown spot disease symptoms were usually observed in September after long periods of rain, management efforts need to focus on protecting bagged fruit before harvest. References: (1) H. Perdomo et al. J. Clin. Microbiol. 49:243, 2011. (2) R. C. Summerell et al. Stud. Mycol. 68:139, 2011.
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