From 2010 to 2012, Phytophthora isolates were obtained from brownish diffusion leaf lesions usually up to 2 to 3 cm in diameter of Rhododendron caucasicum ‘Cheer,’ from withered twigs of Rhododendron sp. with blackish elongated lesions up to ~5 cm in length, and from rotten feeder roots of 2-year-old, chlorotic, wilting seedlings of Fagus sylvatica collected from ornamental and forest nurseries in three areas (central and eastern Bohemia and northern Moravia) in the Czech Republic. Isolates formed chrysanthemum-like to slightly stellate, appressed colonies with sparse aerial mycelium on V8 agar (V8A) plates at 20°C after 5 days, whereas on carrot agar (CA) plates the pattern was vague with no aerial mycelium. The cardinal growth temperatures were: min. 3°C, optimum 23 to 27°C, and max. 31°C. Radial growth was 5.7 to 6.6 mm/day at 20°C on V8A. The isolates were homothallic and produced colorless, smooth-walled, spherical oogonia with an average diameter 29.9 to 33.8 μm on CA. Oospores were aplerotic (avg. 26.4 to 29.3 μm), oospore wall was hyaline and averaged 1.3 to 1.7 μm thick, oospore wall index was 0.26 to 0.32. Paragynous or occasionally amphigynous antheridia averaged 13.4 to 15.0 × 10.9 to 12.5 μm (height × width). Sporangia were caducous, papillate, globose, spherical to ovoid, with short pedicels (avg. 2.1 to 2.6 μm) and averaged 30.9 to 41.5 × 25.5 to 30.6 μm, L:B ratio was 1.2 to 1.4. Chlamydospores were not observed. Morphological characters resembled those described for P. hedraiandra (1). The isolates were deposited in the collection of phytopathogenic oomycetes of RILOG Pruhonice and given accession nos. 450.11, 531.11, and 578.12. The isolates were sequenced for nuclear rDNA ITS region and partial Cox I gene. Obtained sequences were compared with sequences present in GenBank database using BLAST. The ITS sequences of all isolates (GenBank Accession Nos. KJ567081, 82, and 83) of overall length of 792 bp were identical to that of P. hedraiandra AY707987 (1). The Cox I sequences of overall length of 880 bp (KJ567084, 85, and 86) showed 99% homology (1 bp substitution) with AY769115 (1) and 100% identity with other Cox I sequences of P. hedraiandra, i.e., JN376067 (4) and EF174432 (3). Koch's postulates were confirmed by wound-inoculating of 3-year-old rhododendron and common beech plants (10 host plants per corresponding isolate). Rhododendron leaves were gently abraded near the midrib, whereas 5-mm-diameter bark plugs were removed from the beech collars. The inoculum (5-mm-diameter V8A plug with actively growing mycelium) was placed over wounds and sealed with Parafilm. Control plants were treated in the same manner with sterile agar plugs. Plants were maintained in a greenhouse at 22°C. All inoculated plants showed disease symptoms after 10 days of incubation; the lesions were up to 2 cm in rhododendron leaves and ~1 cm in beech collars. Control plants remained healthy. The pathogen was re-isolated from all infected plants. To our knowledge, this is the first report of P. hedraiandra in the Czech Republic. Besides it, the pathogen was found in southern and western Europe (Italy, Slovenia, Spain, the Netherlands) and in the United States (2).
References: (1) A. W. A. M. de Cock and A. Lévesque. Stud. Mycol. 50:481, 2004. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, Syst. Mycol. Microbiol. Lab., ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/, May 13, 2014. (4) E. Moralejo et al. Span. J. Agric. Res. 5:82, 2007. (2) X. Yang et al. Plant Dis. 96:915, 2012.
