First Report of a Leaf Spot on Pepper Caused by Cladosporium oxysporum in China

Abstract

Pepper (Capsicum annuum L.) is an important vegetable crop grown in Liaoning Province of China. In June 2009, leaf spotting was observed on hot pepper cv. 37-74 in Wafangdian County of Liaoning Province. By August 2011, the disease had spread to Pulandian and Donggang Counties. Symptoms initially appeared on both sides of leaves as pinpoint chlorotic spots that enlarged and developed into irregular, brown lesions, 1 to 8 mm in diameter. To identify the pathogen, leaf pieces (3 to 5 mm) taken at the edge of lesions with both infected and healthy portions were surface disinfected by placing them in 75% ethanol for 5 s, then transferred to a 0.1% aqueous mercuric chloride solution for 30 s and rinsed with sterilized water three times. The sections were placed on potato dextrose agar (PDA) at 25°C in the dark. Ten pure fungal cultures were obtained from single spores. For growth rate determination and morphological description of colonies, single conidial isolates were inoculated on PDA, malt extract agar (MEA), and oatmeal agar (OA), and incubated at 25°C for 14 days in darkness. For studies of microscopic morphology, isolates were grown on synthetic nutrient agar (SNA) in slide cultures. Colonies grew up to 77 to 80 mm in diameter on PDA, 62 to 66 mm on MEA, and 58 to 60 mm on OA after 14 days. Conidiophores were straight to slightly flexuous. Conidia ranged in shape from subglobose or ovoid to subcylindrical. Macronematous conidiophores measured 40 to 670 × 3 to 5.5 μm with swellings, micronematous conidiophores 15 to 137 × 1.5 to 3.5 μm, terminal conidia 2 to 4 × 2 to 3 μm, and intercalary conidia 4 to 10 × 2.5 to 4 μm. Ramoconidia were rarely observed. Secondary ramoconidia were zero- to one-septate and measured 5.2 to 14.8 × 2.4 to 4 μm. On the basis of these characteristics, the isolates were identified as Cladosporium oxysporum (1). The internal transcribed spacer (ITS) region from isolate W10-02 was amplified using primers ITS1 and ITS4 and sequenced (GenBank Accession No. JQ775499). The 525-bp amplicons had 99% identity to C. oxysporum (GenBank Accession No. EF029816). On the basis of morphological characteristics and nucleotide homology, the isolate was identified as C. oxysporum. Koch's postulates were fulfilled in the laboratory on pepper leaves inoculated with C. oxysporum conidial suspensions (107 conidia ml-1). Eight inoculated 4-week-old seedlings were incubated under moist conditions for 8 to 10 days at 25°C. All leaf spots that developed on inoculated leaves were similar in appearance to those observed on diseased pepper in the field. C. oxysporum was reisolated from lesions and its identity was confirmed by morphological characteristics. C. oxysporum was first reported as a pathogen of pepper in the Sultanate of Oman (2). To our knowledge, this is the first report of C. oxysporum causing leaf spot on pepper in China. The outbreak and spread of this disease may decrease the yield of pepper in the northern regions of China. More studies are needed on the management strategy including the resistance of pepper cultivars against C. oxysporum.