First Report of Leaf Spot Caused by Cochliobolus eragrostidis on Corn (Zea mays L.) in Fujian Province, China

Corn (Zea mays L.) is one of the most important crops in China and the world. Approximately 60,000 ha of corn, especially sweet and waxy corn, is planted yearly in Fujian province (Dai et al. 2016). Leaf spot symptoms were observed on corn beginning in July 2016. The symptoms began as small, water-soaked lesions on young corn leaves, and then they became larger, dark brown, necrotic lesions, 1 to 6 mm long. Lesions presented as gray-white in the center with a dark brown halo at the margin. Finally, the most affected leaves became necrotic. To determine the causal agent, a total of 30 corn leaves exhibiting leaf spots were collected from Putian, Xianyou, and Xiapu in Fujian Province of China. Small pieces (5 × 5 mm) from infected leaves were surface sterilized in 75% (v/v) alcohol for 2 min and then in 0.1% (w/v) mercuric chloride solution for 90 s. Pieces were rinsed three times in sterilized distilled water, placed on potato dextrose agar containing 50 μg/ml of rifampicin, and incubated at 28°C in the dark for 5 days. Finally, 45 pure cultures were obtained by single conidia isolation. All isolates produced gray fluffy aerial hyphae and exhibited dark green color on the reverse. The typical hyphae were hyaline, branched, septate, and 4.0 to 9.5 µm (average, 5.5 µm) in diameter. The mature conidia were ellipsoidal, straight or slightly curved, and had three transverse septa, with the central two cells approximately equal in size and brown or dark brown, whereas the cells at each end of the conidia were pale. Conidia were 8.5 to 13.0 µm (average, 11.0 µm) wide and 18.5 to 26.5 µm (average, 21.0 µm) long. Based on morphological characteristics, isolates were identified as Cochliobolus eragrostidis (anamorph: Curvularia eragrostidis [P Henn.] J. A. Meyer) (Ferreira et al. 2014; Shi et al. 2000). To confirm identification, two isolates (WB13 and PT2), which were isolated from Xianyou and Putian, were randomly selected as representatives for molecular identification. The nuclear rDNA internal transcribed spacer (ITS) regions were amplified with the universal primer ITS4/ITS5 (White et al. 1990). The resulting PCR products (GenBank accession nos. KY654327 and MF351560) from the two isolates were identical and shared a 100% identity to the ITS sequences of isolates 1172 (KT933667), 5854 (KT012655), and W612 (JN006772), which were isolated from corn in Malaysia, the United States, and China, respectively. Pathogenicity testing of the isolates was implemented utilizing a conidial suspension inoculation method on the hybrid-type corn Shangpin (Dai et al. 2016). Conidial suspensions (10⁵ conidia/ml) of each of the two isolates were sprayed on six leaves per plant on 12 corn seedlings. Control plants were sprayed with distilled water. The inoculated corn seedlings were incubated at 28°C in a greenhouse (12 h of light and relative humidity >90%). Approximately 7 days later, all inoculated plants exhibited similar symptoms as observed in the field, and the isolates of C. eragrostidis were reisolated from lesions. The specific fungus has been reported on other host plants in other areas of China (Zhu and Qiang 2004), but to our knowledge, this is the first report of C. eragrostidis causing leaf spot of corn in Fujian Province of China. The resulting disease could influence yield and quality of corn in the field, especially under temperate and moist conditions. Therefore, there is a need to determine the distribution of C. eragrostidis in Fujian Province and explore effective control measures, to improve leaf spot control.