First Report of Spot Form of Net Blotch of Barley Caused by Pyrenophora teres f. maculata in Hungary.

A countrywide survey of fungal diseases of barley (Hordeum vulgare L.) was conducted from 2005 to 2009. Unusual leaf necrosis varying in shape from 1 × 2 mm necrotic flecks to 15 × 20 mm ovoid spots was found. Sometimes a chlorotic halo surrounding the dead area was observed. Lesions appeared on various cultivars in many commercial fields and experimental plots at a number of sampling sites. Symptomatic leaves were taken to the laboratory and incubated in a moist chamber at room temperature on the bench to induce sporulation of the pathogen. Conidiophores on the diseased tissues were single or in small groups, dark brown, and bore several hyaline-to-olive brown, almost cylindrical conidia with three to seven pseudosepta. Dimensions of conidia were 75.2 to 100.9 × 16.5 to 18.8 μm. Under a stereo microscope, single conidia were transferred aseptically from the leaves onto potato dextrose agar (PDA) with a sterile needle. Plates were kept in the dark at 20°C for 2 weeks. Cultures were gray to olive green, cottony, and did not form conidia and sexual structures. These characteristics indicated that the pathogens belonged to the genus Pyrenophora. Species identity was confirmed by PCR assays with specific primers developed for the barley pathogenic Pyrenophora spp. (3,4). Of 169 isolates, 41 were identified as P. teres Drechs. f. maculata Smed.-Pet., the spot form of net blotch pathogen (2), and two of them have been deposited at an international culture collection under accession nos. CBS 123929 and CBS 123930. The remaining isolates were either P. graminea or P. teres f. teres, the leaf stripe and net form of net blotch pathogens of barley, respectively. Pathogenicity of four P. teres f. maculata and two P. teres f. teres isolates from different regions was confirmed by Koch's postulates. Each isolate was grown on two 9-cm PDA plates at 22°C in darkness. After 10 days, aerial mycelia were scraped off, blended in 100 ml of sterile distilled water, and filtered through two layers of cheesecloth. Ten seedlings of cv. Botond were sprayed at the two-leaf stage with the mycelium suspension of each isolate and a water control until runoff. Seedlings were kept in a growth chamber at 100% relative humidity and 20°C in the dark for 24 h, then at 70% relative humidity and 24/20°C (day/night) with a 12-h photoperiod. Within 3 weeks, one to four brownish ovoid spots, typical of the spot form of net blotch symptoms, developed on the leaves inoculated with P. teres f. maculata. In contrast, the seedlings inoculated with P. teres f. teres exhibited characteristic net-like lesions, whereas the control plants sprayed with sterile water remained healthy. All strains were reisolated and identified by specific PCRs as described above. To our knowledge, this is the first report of the occurrence of P. teres f. maculata in Hungary. Resistance of barley against P. teres f. maculata and P. teres f. teres is inherited independently (1). Therefore, knowledge regarding the frequency and distribution of these pathogens is important for disease management and resistance breeding. References: (1) O. S. Afanasenko et al. J. Phytopathol. 143:501, 1995. (2) V. Smedegård-Petersen. Page 124 in: R. Vet. Agr. Univ. Yearbook. Copenhagen, 1971. (3) E. J. A. Taylor et al. Plant Pathol. 50:347, 2001. (4) K. J. Williams et al. Australas. Plant Pathol. 30:37, 2001.