Global spread, genetic differentiation and selection of barley spot form of net blotch isolates.

Spot form of net blotch, caused by Pyrenophora teres f. maculata, is a significant necrotrophic disease of barley that spread world-wide in the 20th century. Genetic relationships were analysed to determine the diversity, survival and dispersal of a diverse collection of 346 isolates from Australia, Southern Africa, North America, Asia Minor and Europe. The results, based on genome-wide DArTseq data, indicated isolates from Turkey were the most differentiated with regional sub-structuring, together with individuals closely related to geographically distant genotypes. Elsewhere, population subdivision related to country of origin was evident, although low levels of admixturing was found that may represent rare genotypes or migration from unsampled populations. Canadian isolates were the next most diverged and Australian and South African the most closely related. With the exception of Turkish isolates, multiple independent Cyp51A mutation events (which confer insensitivity to demethylation inhibitor fungicides) between countries and within regions was evident, with strong selection for a transposable element insertion at the 3' end of the promoter and counter-selection elsewhere. Individuals from Western Australia shared genomic regions and Cyp51A haplotypes with South African isolates, suggesting a recent common origin.

Worldwide forest surveys reveal forty-three new species in Phytophthora major Clade 2 with fundamental implications for the evolution and biogeography of the genus and global plant biosecurity.

During 25 surveys of global Phytophthora diversity, conducted between 1998 and 2020, 43 new species were detected in natural ecosystems and, occasionally, in nurseries and outplantings in Europe, Southeast and East Asia and the Americas. Based on a multigene phylogeny of nine nuclear and four mitochondrial gene regions they were assigned to five of the six known subclades, 2a-c, e and f, of Phytophthora major Clade 2 and the new subclade 2g. The evolutionary history of the Clade appears to have involved the pre-Gondwanan divergence of three extant subclades, 2c, 2e and 2f, all having disjunct natural distributions on separate continents and comprising species with a soilborne and aquatic lifestyle and, in addition, a few partially aerial species in Clade 2c; and the post-Gondwanan evolution of subclades 2a and 2g in Southeast/East Asia and 2b in South America, respectively, from their common ancestor. Species in Clade 2g are soilborne whereas Clade 2b comprises both soil-inhabiting and aerial species. Clade 2a has evolved further towards an aerial lifestyle comprising only species which are predominantly or partially airborne. Based on high nuclear heterozygosity levels ca. 38 % of the taxa in Clades 2a and 2b could be some form of hybrid, and the hybridity may be favoured by an A1/A2 breeding system and an aerial life style. Circumstantial evidence suggests the now 93 described species and informally designated taxa in Clade 2 result from both allopatric non-adaptive and sympatric adaptive radiations. They represent most morphological and physiological characters, breeding systems, lifestyles and forms of host specialism found across the Phytophthora clades as a whole, demonstrating the strong biological cohesiveness of the genus. The finding of 43 previously unknown species from a single Phytophthora clade highlight a critical lack of information on the scale of the unknown pathogen threats to forests and natural ecosystems, underlining the risk of basing plant biosecurity protocols mainly on lists of named organisms. More surveys in natural ecosystems of yet unsurveyed regions in Africa, Asia, Central and South America are needed to unveil the full diversity of the clade and the factors driving diversity, speciation and adaptation in Phytophthora. Taxonomic novelties: New species: Phytophthora amamensis T. Jung, K. Kageyama, H. Masuya & S. Uematsu, Phytophthora angustata T. Jung, L. Garcia, B. Mendieta-Araica, & Y. Balci, Phytophthora balkanensis I. Milenkovic, Z. Tomic, T. Jung & M. Horta Jung, Phytophthora borneensis T. Jung, A. Durán, M. Tarigan & M. Horta Jung, Phytophthora calidophila T. Jung, Y. Balci, L. Garcia & B. Mendieta-Araica, Phytophthora catenulata T. Jung, T.-T. Chang, N.M. Chi & M. Horta Jung, Phytophthora celeris T. Jung, L. Oliveira, M. Tarigan & I. Milenkovic, Phytophthora curvata T. Jung, A. Hieno, H. Masuya & M. Horta Jung, Phytophthora distorta T. Jung, A. Durán, E. Sanfuentes von Stowasser & M. Horta Jung, Phytophthora excentrica T. Jung, S. Uematsu, K. Kageyama & C.M. Brasier, Phytophthora falcata T. Jung, K. Kageyama, S. Uematsu & M. Horta Jung, Phytophthora fansipanensis T. Jung, N.M. Chi, T. Corcobado & C.M. Brasier, Phytophthora frigidophila T. Jung, Y. Balci, K. Broders & I. Milenkovic, Phytophthora furcata T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora inclinata N.M. Chi, T. Jung, M. Horta Jung & I. Milenkovic, Phytophthora indonesiensis T. Jung, M. Tarigan, L. Oliveira & I. Milenkovic, Phytophthora japonensis T. Jung, A. Hieno, H. Masuya & J.F. Webber, Phytophthora limosa T. Corcobado, T. Majek, M. Ferreira & T. Jung, Phytophthora macroglobulosa H.-C. Zeng, H.-H. Ho, F.-C. Zheng & T. Jung, Phytophthora montana T. Jung, Y. Balci, K. Broders & M. Horta Jung, Phytophthora multipapillata T. Jung, M. Tarigan, I. Milenkovic & M. Horta Jung, Phytophthora multiplex T. Jung, Y. Balci, K. Broders & M. Horta Jung, Phytophthora nimia T. Jung, H. Masuya, A. Hieno & C.M. Brasier, Phytophthora oblonga T. Jung, S. Uematsu, K. Kageyama & C.M. Brasier, Phytophthora obovoidea T. Jung, Y. Balci, L. Garcia & B. Mendieta-Araica, Phytophthora obturata T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora penetrans T. Jung, Y. Balci, K. Broders & I. Milenkovic, Phytophthora platani T. Jung, A. Pérez-Sierra, S.O. Cacciola & M. Horta Jung, Phytophthora proliferata T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora pseudocapensis T. Jung, T.-T. Chang, I. Milenkovic & M. Horta Jung, Phytophthora pseudocitrophthora T. Jung, S.O. Cacciola, J. Bakonyi & M. Horta Jung, Phytophthora pseudofrigida T. Jung, A. Durán, M. Tarigan & M. Horta Jung, Phytophthora pseudoccultans T. Jung, T.-T. Chang, I. Milenkovic & M. Horta Jung, Phytophthora pyriformis T. Jung, Y. Balci, K.D. Boders & M. Horta Jung, Phytophthora sumatera T. Jung, M. Tarigan, M. Junaid & A. Durán, Phytophthora transposita T. Jung, K. Kageyama, C.M. Brasier & H. Masuya, Phytophthora vacuola T. Jung, H. Masuya, K. Kageyama & J.F. Webber, Phytophthora valdiviana T. Jung, E. Sanfuentes von Stowasser, A. Durán & M. Horta Jung, Phytophthora variepedicellata T. Jung, Y. Balci, K. Broders & I. Milenkovic, Phytophthora vietnamensis T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora ×australasiatica T. Jung, N.M. Chi, M. Tarigan & M. Horta Jung, Phytophthora ×lusitanica T. Jung, M. Horta Jung, C. Maia & I. Milenkovic, Phytophthora ×taiwanensis T. Jung, T.-T. Chang, H.-S. Fu & M. Horta Jung. Citation: Jung T, Milenkovic I, Balci Y, Janousek J, Kudlácek T, Nagy ZÁ, Baharuddin B, Bakonyi J, Broders KD, Cacciola SO, Chang T-T, Chi NM, Corcobado T, Cravador A, Dordevic B, Durán A, Ferreira M, Fu C-H, Garcia L, Hieno A, Ho H-H, Hong C, Junaid M, Kageyama K, Kuswinanti T, Maia C, Májek T, Masuya H, Magnano di San Lio G, Mendieta-Araica B, Nasri N, Oliveira LSS, Pane A, Pérez-Sierra A, Rosmana A, Sanfuentes von Stowasser E, Scanu B, Singh R, Stanivukovic Z, Tarigan M, Thu PQ, Tomic Z, Tomsovský M, Uematsu S, Webber JF, Zeng H-C, Zheng F-C, Brasier CM, Horta Jung M (2024). Worldwide forest surveys reveal forty-three new species in Phytophthora major Clade 2 with fundamental implications for the evolution and biogeography of the genus and global plant biosecurity. Studies in Mycology 107: 251-388. doi: 10.3114/sim.2024.107.04.

Heat shock-induced enhanced susceptibility of barley to Bipolaris sorokiniana is associated with elevated ROS production and plant defence-related gene expression.

Heat stress alters plant defence responses to pathogens. Short-term heat shock promotes infections by biotrophic pathogens. However, little is known about how heat shock affects infection by hemibiotrophic pathogens like Bipolaris sorokiniana (teleomorph: Cochliobolus sativus). We assessed the effect of heat shock in B. sorokiniana-susceptible barley (Hordeum vulgare cv. Ingrid) by monitoring leaf spot symptoms, B. sorokiniana biomass, ROS and plant defence-related gene expression following pre-exposure to heat shock. For heat shock, barley plants were kept at 49 °C for 20 s. B. sorokiniana biomass was assessed by qPCR, ROS levels determined by histochemical staining, while gene expression was assayed by RT-qPCR. Heat shock suppressed defence responses of barley to B. sorokiniana, resulting in more severe necrotic symptoms and increased fungal biomass, as compared to untreated plants. Heat shock-induced increased susceptibility was accompanied by significant increases in ROS (superoxide, H2 O2 ). Transient expression of plant defence-related antioxidant genes and a barley programmed cell death inhibitor (HvBI-1) were induced in response to heat shock. However, heat shock followed by B. sorokiniana infection caused further transient increases in expression of HvSOD and HvBI-1 correlated with enhanced susceptibility. Expression of the HvPR-1b gene encoding pathogenesis-related protein-1b increased several fold 24 h after B. sorokiniana infection, however, heat shock further increased transcript levels along with enhanced susceptibility. Heat shock induces enhanced susceptibility of barley to B. sorokiniana, associated with elevated ROS levels and expression of plant defence-related genes encoding antioxidants, a cell death inhibitor, and PR-1b. Our results may contribute to elucidating the influence of heat shock on barley defence responses to hemibiotrophic pathogens.

Six new Phytophthora species from ITS Clade 7a including two sexually functional heterothallic hybrid species detected in natural ecosystems in Taiwan.

During a survey of Phytophthora diversity in natural ecosystems in Taiwan six new species were detected. Multigene phylogeny based on the nuclear ITS, ß-tubulin and HSP90 and the mitochondrial cox1 and NADH1 gene sequences demonstrated that they belong to ITS Clade 7a with P. europaea, P. uniformis, P. rubi and P. cambivora being their closest relatives. All six new species differed from each other and from related species by a unique combination of morphological characters, the breeding system, cardinal temperatures and growth rates. Four homothallic species, P. attenuata, P. flexuosa, P. formosa and P. intricata, were isolated from rhizosphere soil of healthy forests of Fagus hayatae, Quercus glandulifera, Q. tarokoensis, Castanopsis carlesii, Chamaecyparis formosensis and Araucaria cunninghamii. Two heterothallic species, P. xheterohybrida and P. xincrassata, were exclusively detected in three forest streams. All P. xincrassata isolates belonged to the A2 mating type while isolates of P. xheterohybrida represented both mating types with oospore abortion rates according to Mendelian ratios (4-33 %). Multiple heterozygous positions in their ITS, ß-tubulin and HSP90 gene sequences indicate that P. xheterohybrida, P. xincrassata and P. cambivora are interspecific hybrids. Consequently, P. cambivora is re-described as P. xcambivora without nomenclatural act. Pathogenicity trials on seedlings of Castanea sativa, Fagus sylvatica and Q. suber indicate that all six new species might pose a potential threat to European forests.

Nothophytophthora gen. nov., a new sister genus of Phytophthora from natural and semi-natural ecosystems.

During various surveys of Phytophthora diversity in Europe, Chile and Vietnam slow growing oomycete isolates were obtained from rhizosphere soil samples and small streams in natural and planted forest stands. Phylogenetic analyses of sequences from the nuclear ITS, LSU, ß-tubulin and HSP90 loci and the mitochondrial cox1 and NADH1 genes revealed they belong to six new species of a new genus, officially described here as Nothophytophthora gen. nov., which clustered as sister group to Phytophthora. Nothophytophthora species share numerous morphological characters with Phytophthora: persistent (all Nothophytophthora spp.) and caducous (N. caduca, N. chlamydospora, N. valdiviana, N. vietnamensis) sporangia with variable shapes, internal differentiation of zoospores and internal, nested and extended (N. caduca, N. chlamydospora) and external (all Nothophytophthora spp.) sporangial proliferation; smooth-walled oogonia with amphigynous (N. amphigynosa) and paragynous (N. amphigynosa, N. intricata, N. vietnamensis) attachment of the antheridia; chlamydospores (N. chlamydospora) and hyphal swellings. Main differing features of the new genus are the presence of a conspicuous, opaque plug inside the sporangiophore close to the base of most mature sporangia in all known Nothophytophthora species and intraspecific co-occurrence of caducity and non-papillate sporangia with internal nested and extended proliferation in several Nothophytophthora species. Comparisons of morphological structures of both genera allow hypotheses about the morphology and ecology of their common ancestor which are discussed. Production of caducous sporangia by N. caduca, N. chlamydospora and N. valdiviana from Valdivian rainforests and N. vietnamensis from a mountain forest in Vietnam suggests a partially aerial lifestyle as adaptation to these humid habitats. Presence of tree dieback in all forests from which Nothophytophthora spp. were recovered and partial sporangial caducity of several Nothophytophthora species indicate a pathogenic rather than a saprophytic lifestyle. Isolation tests from symptomatic plant tissues in these forests and pathogenicity tests are urgently required to clarify the lifestyle of the six Nothophytophthora species.

First Report of Phytophthora × pelgrandis Causing Root Rot and Lower Stem Necrosis of Common Box, Lavender and Port-Orford-Cedar in Hungary.

In 2008 and 2009, necrotic bark lesions at the root collar and lower stem associated with root rot, reduced growth, and wilting were observed on container-grown common box (Buxus sempervirens L.), lavender (Lavandula angustifolia Mill. 'Hidcote'), and Port-Orford-cedar (Chamaecyparis lawsoniana (A. Murray) Parl. 'Columnaris') in three ornamental nurseries in western Hungary. Number of affected plants ranged from approximately 100 (Port-Orford-cedar) to 250 (lavender). Isolations from necrotic root collars of each host plant species yielded four Phytophthora isolates developing uniform colonies on carrot agar with a maximum growth temperature of 35 to 36°C. The isolates were homothallic with smooth-walled oogonia (32.2 ± 2.3 to 35.9 ± 3.5 µm), aplerotic oospores (27.5 ± 1.8 to 32.1 ± 3.1 µm) and both amphigynous and paragynous antheridia, and produced chlamydospores (25.8 ± 3.9 to 29.1 ± 5.2 µm) and papillate sporangia (35.2 ± 2.5 to 43.5 ± 5.6 µm long and 27.6 ± 2.2 to 32.0 ± 3.8 µm wide), mostly obpyriform to nearly spherical or rarely distorted with two or three apices. In spring water, sporangia were both caducous with short pedicel and non-caducous. Multiplex ITS-PCR assay of DNA from all isolates, using primers specific for P. nicotianae (NICF1 and NICR2.1) and P. cactorum (CACTF1 and CACTR1) (1), amplified DNA fragments of the expected size for each Phytophthora species. In addition, isoenzyme analysis revealed a characteristic banding pattern of one heterodimer and two homodimer bands at both loci of the dimeric enzyme malate dehydrogenase. These bands comigrated with those of P. × pelgrandis (Gerlach et al.) (CBS 123385) and isolate PD 93/1339 (courtesy of W. A. Man in 't Veld), two natural hybrid strains of P. nicotianae and P. cactorum (2,3), proving that our four isolates can be referred to as this interspecific hybrid. Pathogenicity was tested on 1- or 3-year-old plants of the original host species and cultivars (for common box, cv. Faulkner was used). Cultures were grown for 4 to 6 weeks at 20°C on autoclaved millet grains moistened with V8 broth. Infested and uninfested grains were mixed with autoclaved soil in a ratio of 6% (w/v), and the mixes were used as potting media for transplanting five treated and five control plants per isolate, respectively. Plants were kept in a growth chamber (20°C, 70% RH, 12-h photoperiod). Pots were flooded for 24 h on the 1st and 21st day after transplanting. All plants in infested potting mix showed symptoms of wilt associated with basal stem and root necrosis, similar to those observed on the plants from the field, within 2 and 3 months on lavender and both common box or Port-Orford-cedar, respectively. Additionally, a reduction of root weight ranging from 35 to 68% compared to the control was recorded. Growth reduction was significant at P ≤ 0.019 according to Mann Whitney test. Control plants remained healthy. The same Phytophthora hybrid was reisolated solely from inoculated plants. In Europe, hybrid isolates of P. nicotianae × P. cactorum have been reported on several ornamental plants, including lavender, in the Netherlands and Germany (2,3). However, to our knowledge, this is the first report of this hybrid in Hungary and as a pathogen of common box and Port-Orford-cedar in the world. References: (1) P. J. M. Bonants et al. Phytopathology 90:867, 2000. (2) W. A. Man in 't Veld et al. Phytopathology 88:922, 1998. (3) H. I. Nirenberg et al. Mycologia 101:220, 2009.

First Report of Phytophthora citrophthora Causing Root and Basal Stem Rot of Woody Ornamentals in Hungary.

From 2007 to 2009, necrotic bark lesions at the root collar and lower stem associated with root rot were observed on container-grown cork-bark fir (Abies lasiocarpa var. arizonica, Port-Orford-cedar (Chamaecyparis lawsoniana (A. Murray) Parl. 'Barabits Gold'), lavender (Lavandula angustifolia Mill. 'Hidcote'), flowering currant (Ribes sanguineum Pursh 'King Edward VII'), and lilac (Syringa vulgaris L. 'Belle de Nancy') in six ornamental nurseries in western Hungary. Symptoms also included reduced growth, wilting, and desiccation of branches. Mortality of affected plants ranged from a low level in flowering currant to a high frequency in cork-bark fir (1,000 plants; 50%) and lavender (2,500 plants; 50%). Isolations from necrotic tissues onto PARPB medium (1) yielded 13 isolates of a Phytophthora sp. None of the isolates produced sexual structures, sporangia, or chlamydospores but developed slightly stellate or patternless colonies with loose, aerial mycelium on nonselective carrot agar (CA) at 25°C. One isolate from each host species was further characterized and tested for pathogenicity. Growth on CA was fastest at 25°C (7.9 to 8.6 mm per day) and no growth occurred below 5°C or above 34°C. In nonsterile stream water, persistent, mono- and bipapillate, mostly ovoid, rarely distorted sporangia, measuring 40.5 to 49.4 × 29.8 to 37.3 µm were produced. In pairings on CA with A1 and A2 strains of P. cambivora and P. nicotianae, used as testers, none of our isolates produced gametangia. On the basis of these characteristics, the pathogen from ornamentals appeared to be P. citrophthora (Smith & Smith) Leon. (1). Species identity of all 13 isolates was determined in single-round PCR assays with the P. citrophthora-specific primer-pair Pc2B/Pc7 (2) and/or by sequencing the rDNA internal transcribed spacer (ITS) regions amplified with the universal ITS1/ITS4 primers. The primers Pc2B and Pc7 generated a single DNA fragment of the expected size (approximately 210 bp), and the rDNA ITS sequences (NCBI Accession Nos. GU723282, GU723284, GU723285, and GU723287) showed 99 to 100% homology with many GenBank sequences (e.g., HQ697232) of P. citrophthora as the closest match. Pathogenicity to the original host plant cultivars was tested on 2- or 3-year-old healthy plants potted in sterile soil and inoculated at the root collar (2 replicates per isolate). A 4-mm-diameter bark plug was removed and a mycelial disc of the same size from an actively growing CA culture was placed into the hole. Control plants received sterile CA plugs. Inoculation points were sealed with sterile moist cotton and Parafilm, covered with sterile soil, and then the plants were kept in a greenhouse at 24 ± 4°C. Symptoms identical to those observed on the naturally diseased hosts developed on inoculated plants within 3 months. Control plants remained healthy. P. citrophthora could be reisolated only from the infected plants. The pathogen is polyphagous, widely distributed (1), and has been associated with woody ornamentals in nurseries (3). However, to our knowledge, this is the first record of P. citrophthora in Hungary. The pathogen has to be considered as a threat to ornamental production within Hungary. References: (1) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN, 1996. (2) A. Ippolito et al. Eur. J. Plant Pathol. 108:855, 2002. (3) B. W. Schwingle et al. Plant Dis. 91:97, 2007.

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.

First Report of Phytophthora citricola on False Cypress in Hungary.

In May 2005, an estimated 10 to 15% mortality of various cultivars of false cypress (also named Lawson cypress or Port-Orford-cedar [Chamaecyparis lawsoniana]) with severe wilt was observed in field stands of an ornamental nursery in western Hungary. Wilted plants had rot-associated reduction of their root system. Root discoloration and occasional chlorosis of lower leaves commenced on potted 3-year-old plants that were held in the open air for 10 to 12 months before planting. Four species of Phytophthora (P. lateralis, P. eriugena, P. hibernalis, and P. cinnamomi) have been reported on this host (2). Direct plating of discolored roots from the most susceptible cultivar (Silver Globus) onto a selective potato dextrose agar or carrot agar medium yielded pure cultures that developed white, stellate colonies with sparse aerial mycelia. The hyphal growth was optimal at 25°C, but the growth above 32°C and below 4°C was completely inhibited. Single, terminal sporangia on simple (occasionally sympodial) sporangiophores formed abundantly in nonsterile soil filtrate but not in agar. Sporangia, 31 to 67 µm (59.1 ± 9.3 µm) long and 25 to 39 µm (31.5 ± 4.0 µm) wide, were noncaducous and semipapillate, variable in shape, mostly obpyriform, rarely obovoid, ovoid-ellipsoid and spherical or bifurcated and distorted, and the exit pore was narrow (7.2 ± 0.8 µm). No external or internal proliferation and no hyphal swellings or chlamydospores were observed. The isolates were homothallic with smooth-walled oogonia (27.3 ± 3.4 µm in diameter) and paragynous antheridia. The oospores (24.7 ± 2.1 µm in diameter) were plerotic. The morpho-physiological features suggested that our isolates belonged to Waterhouse's Group III, and in particular, represented P. citricola. This was confirmed by cellulose acetate electrophoresis of malate dehydrogenase; the isozyme pattern of false cypress isolate was identical to that of the ITS-sequenced (NCBI Accession No. AY366193) P. citricola isolate from a Hungarian alder forest (1). Pathogenicity tests on four 3-year-old potted false cypress (cv. Silver Globus) plants in the greenhouse resulted in rapidly developing (within 2 weeks) sunken, necrotic lesions at the stem base around the site of wound inoculation with a 5-mm-diameter mycelial agar plug. After 12 weeks, each inoculated plant wilted and died. The causal agent was consistently reisolated from necrotic tissues. In Hungary, P. citricola was first isolated and identified from alder forest soil (1). Nonetheless that false cypress has been listed as the host of P. citricola in Norway and Poland (3,4), to our knowledge, this report is the first definitive description of this Phytophthora sp. on this host. References: (1) J. Bakonyi et al. Plant Pathol. 52:807, 2003. (2) D. C. Erwin and O. K. Ribeiro. Pages 282-287 in: Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN, 1996. (3) V. Talgø V. and A. Stensvand. Grønn kunnskap e 7(101G):1, 2003. (4) K. Wiejacha et al. Page 45 in: Improvement and Unification of Plant Disease Diagnostics. Abstracts of International Workshop, Skierniewice, Poland, 2004.

First Report of Phytophthora Root and Collar Rot of Alder in Hungary.

In June 1999, a disease associated with mortality of Alnus glutinosa, was observed in a 12- to 18-year-old peatland plantation in northwest Hungary. The root and collar rot symptoms were similar to those caused by Phytophthora cambivora in tree species other than alders. Nine isolations were made from diseased roots and soil samples using the Rhododendron leaf baiting method. Three isolates recovered from two sites, approximately 2 km apart, exhibited similar growth and morphology in vitro and were pathogenic to 2-year-old trees of A. glutinosa following inoculation of root collars. All three isolates had amphigynous long, two-celled antheridia. The mean diameter of oogonia ranged from 39.5 to 64.6 µm. They also produced nonpapillate, ellipsoid, non-caducous sporangia 26.9 to 50.5 µm long and 19.3 to 38.5 µm wide with broad exit pores in soil filtrate. These characteristics were similar to those reported for Phytophthora on alder from elsewhere in Europe and for P. cambivora that is not a pathogen of alder (1,2). However, Hungarian isolates from alder, in contrast to P. cambivora, were homothallic like previously recorded isolates from alder, formed nonornamented oogonia and developed colonies at lower optimum (approximately 25°C) and maximum (approximately 30°C) temperatures on carrot agar. A comparison with Phytophthora from alder from other countries (courtesy of C. M. Brasier) showed that the Hungarian isolates have smooth-walled oogonia typical of Swedish isolates rather than the ornamented oogonia of U.K. isolates, but have the appressed, slightly woolly colony morphology like U.K. isolates rather than the fluffy growth found in Swedish isolates. Moreover, cellulose acetate electrophoresis of glucose-6-phosphate isomerase revealed one homodimer band in Hungarian isolates that was identical with that of the Swedish isolate from alder P876 and isolates P1010 and P1011 of P. cambivora (courtesy of C. M. Brasier). This band comigrated with the middle one of the five-banded U.K. standard isolate P772. Molecular evidence (2) indicates that the Phytophthora from alder with its unusual characteristics is not a species in the strict sense but comprises natural hybrids that may have originated in an interspecific hybridization event between a P. cambivora-like species and an unknown species similar to P. fragariae. On this basis, the Hungarian Phytophthora from alder might have evolved similarly. It remains to be determined whether the pathogen was introduced or has developed independently. References: (1) C. M. Brasier et al. Plant Pathol. 44:999, 1995. (2) C. M. Brasier et al. Proc. Natl. Acad. Sci. USA 96:5878, 1999.

First Report of the A2 Mating Type of Phytophthora infestans on Potato in Hungary.

Severe symptoms of potato late blight were observed in July 1996 on potato (Solanum tuberosum L.) cv. Desirée grown on a farm in western Hungary. Isolation was made directly from diseased leaf tissues onto selective pea-agar medium. A recovered isolate, H2a, was identified as Phytophthora infestans (Mont.) de Bary on the basis of Koch's postulates and morphological characteristics of the fungus. Pairing of H2a with isolates of known mating types, A1 and A2 from Germany, revealed that it represents the A2 mating type. After a 2-week incubation on pea-agar medium at 20°C, oospores formed in abundance in the region of contact between the colonies of H2a and the A1 mating type isolate. After extended incubation scattered formation of gametangia was observed when isolate H2a had been paired with itself or with the A2 mating type isolate. The same phenomenon of presumed self fertilization also took place when single, zoospore-derived colonies of H2a were combined with one another or with the known A2 isolate. Of an incomplete set of potato differentials, leaves of potato genotypes r, R2, R3, R4, R1.2.3.4, R7, R8, and R11 were all susceptible to infection with a zoospore suspension of the isolate H2a. The complex virulence phenotype of H2a, its tolerance to metalaxyl in agar cultures and on leaf disks (EC50 >100 mg liter-1), and its A2 mating type behavior collectively suggest that H2a represents a genotype that recently has been introduced into Hungary.

Comparison of selected species of Bipolaris, Drechslera and Exserohilum by random amplification of polymorphic DNA.

Forty-six strains representing 15 species of Drechslera, five of Bipolaris and four of Exserohilum, as well as two formae of Drechslera teres were compared by RAPD analysis. Drechslera formed a large, heterologous group, while species of Bipolaris and Exserohilum were more closely related. Strong pair-wise affinities were observed between D. graminea and D. teres, D. tritici-repentis and D. bromi, D. siccans and D. biseptata, D. fugax and D. poae, B. sorghicola and B. zeicola, as well as between E. rostratum and E. turcicum.