Molecular data do not support a southern hemisphere base of Nothofagus powdery mildews.

Three powdery mildew species present on Nothofagus (viz. Erysiphe magellanica, E. nothofagi and E. patagoniaca) are endemic to South America and have unique ascomatal appendages that are not found in powdery mildews of the northern hemisphere. We determined the nucleotide sequences of the rDNA internal transcribed spacer regions and D1/D2 domains of the 28S rDNA of these three powdery mildew species to reveal their phylogenetic relationships with powdery mildews of the northern hemisphere. Although the molecular phylogenetic analyses indicated that the three Nothofagus powdery mildews are closely related to each other they did not group into one clade in either the ITS or 28S trees. Kishino-Hasegawa, Shimodaira-Hasegawa and Templeton tests could not significantly reject the constrained trees that were constructed based on the assumption that the Nothofagus powdery mildews would form a single clade. Based on this result and the evidence that all Nothofagus powdery mildews are endemic to South America and have similar morphological characteristics, it is likely that these three species diverged from a single ancestor present on Nothofagus. Calibration of evolutionary events with molecular clocks suggested that the Nothofagus powdery mildews split from the northern hemisphere relatives 22-16 million y ago (Ma) in the middle Miocene, and divergence among the Nothofagus powdery mildews occurred 17-13 Ma. These results do not support a southern hemisphere base of the Nothofagus powdery mildews.

Molecular phylogeny and evolution of the genus Neoerysiphe (Erysiphaceae, Ascomycota).

The genus Neoerysiphe belongs to the tribe Golovinomyceteae of the Erysiphaceae together with the genera Arthrocladiella and Golovinomyces. This is a relatively small genus, comprising only six species, and having ca 300 species from six plant families as hosts. To investigate the molecular phylogeny and evolution of the genus, we determined the nucleotide sequences of the rDNA ITS regions and the divergent domains D1 and D2 of the 28S rDNA. The 30 ITS sequences from Neoerysiphe are divided into three monophyletic groups that are represented by their host families. Groups 1 and 3 consist of N. galeopsidis from Lamiaceae and N. galii from Rubiaceae, respectively, and the genetic diversity within each group is extremely low. Group 2 is represented by N. cumminsiana from Asteraceae. This group also includes Oidium baccharidis, O. maquii, and Oidium spp. from Galinsoga (Asteraceae) and Aloysia (Verbenaceae), and is further divided into four subgroups. N. galeopsidis is distributed worldwide, but is especially common in western Eurasia from Central Asia to Europe. N. galii is also common in western Eurasia. In contrast, the specimens of group 2 were all collected in the New World, except for one specimen that was collected in Japan; this may indicate a close relationship of group 2 with the New World. Molecular clock calibration demonstrated that Neoerysiphe split from other genera of the Erysiphaceae ca 35-45M years ago (Mya), and that the three groups of Neoerysiphe diverged between 10 and 15Mya, in the Miocene. Aloysia citriodora is a new host for the Erysiphaceae and the fungus on this plant is described as O. aloysiae sp. nov.

Comprehensive molecular phylogenetic analysis and evolution of the genus Phyllactinia (Ascomycota: Erysiphales) and its allied genera.

Phyllactinia is a unique genus within the Erysiphales (Ascomycota) having a partly endo-parasitic nature of the mycelium within the host plant tissues. We constructed phylogenetic trees for the genus Phyllactinia and its allied genera based on a total of 120 nucleotide sequences of the 28S rDNA and ITS regions to discuss their phylogenetic relationships with special references to host plants, biogeography, evolutionary dating, and taxonomy. The analysis of the Erysiphales confirmed the monophyly of the endo-parasitic genera, i.e. Leveillula, Phyllactinia, and Pleochaeta. Phyllactinia specimens used in this study were divided into six distinctive groups and three subgroups. Interestingly, Leveillula, an obligately endo-parasitic genus of the Erysiphales, grouped together with Phyllactinia, although this was not significantly supported by the Kishino-Hasegawa and Shimodaira-Hasegawa tests. This suggests that the evolution within this group of fungi occurred from partial endo-parasitism to obligate endo-parasitism. The host range of Phyllactinia is mostly confined to woody plants, especially deciduous trees. Betulaceae, Fagaceae, Ulmaceae, Moraceae, and Rosaceae may have close connections to the divergence of the groups and subgroups of Phyllactinia concerned. Most of these plant families are known as major members of the boreotropical flora of the Tertiary, which suggests an early Tertiary origin of this genus. A comparison of the phylogenies of hosts and parasites revealed that host range expansion at higher taxonomic levels (higher than family level) is independent of the phylogeny of plants. Conversely, host range expansions in lower taxonomic levels (infrafamilial or infrageneric) tend to occur within a single family or genus. An estimation of the evolutionary timing using a molecular clock approach suggested that Phyllactinia split from Pleochaeta about 60 M years ago (Ma) in the early Tertiary and divergence of the six major clades of Phyllactinia occurred between 5 and 40 Ma during the Oligocene and Miocene. Divergence within the major clades and within Leveillula occurred maybe from more than 5 Ma onwards during the Pliocene and Quaternary. This is the first comprehensive phylogenetic study of Phyllactinia and other endo-parasitic genera of the Erysiphales.

Molecular and morphological characterization of Leveillula (Ascomycota: Erysiphales) on monocotyledonous plants.

Leveillula on monocotyledonous plants have been recorded as L. taurica by several authors, whereas the fungus on Allium has been described as an independent species, namely L. allii, by some authors. We sequenced ca 600bp of the rDNA ITS region for two Leveillula specimens from Allium and Polianthes (both from monocotyledons) and compared them with several already published sequences from Leveillula isolates from dicotyledons. Pair-wise percentages of sequence divergences were calculated for all Leveillula isolates. The ITS sequence of the Polianthes isolate was identical to L. taurica on Helianthus and Vicia. The sequence of the Allium isolate was 99.5% identical to L. taurica on Euphorbia, Haplophylum, Peganum, etc. These results suggest close relationships between monocot and dicot pathogenic Leveillula species. The identity between two monocot isolates was 98.4%. Phylogenetic analysis revealed that the two monocot isolates do not group into a clade together. This result suggests that Leveillula acquired parasitism to monocots at least twice independently.

A morphologically ill-founded powdery mildew species, Pleochaeta indica, is recognized as a phylogenetic species based on the analysis of the nuclear ribosomal DNA sequences.

Morphological characteristics of a powdery mildew fungus found on Celtis australis in the Indian Himalayas coincided with those of Pleochaeta indica, described from this tree species in India, as well with those of P. shiraiana, known to infect C. australis and other plant species in Asia. This suggested that the original description of P. indica based on morphological patterns was not well founded and this taxon could be reduced to synonymy with P. shiraiana. However, phylogenetic analyses of the rDNA 28S and ITS sequences determined in some Indian Pleochaeta specimens from C. australis showed that this fungus is closely related, but not identical to P. shiraiana infecting C. sinensis in Japan which served as the basis of the original description of P. shiraiana. Molecular clock analysis of the ITS region and that of the 28S rDNA indicated that the split between the Japanese P. shiraiana infecting C. sinensis and Pleochaeta sp. infecting C. australis in India may have occurred 2.0-8.5 million years ago in the Pliocene and may have coincided with the formation of the Himalayan mountains and the global cooling of the Earth during the late Tertiary. Thus, P. indica is recognized in this study as a distinct phylogenetic species, although our morphological study showed that its description as a morphological species was not well founded. This is a striking example of a cryptic species which is genetically different from close relatives but cannot be distinguished from them based on morphology.

Molecular phylogeny supports a Northern Hemisphere origin of Golovinomyces (Ascomycota: Erysiphales).

Golovinomyces is a strictly herb-parasitic genus in the Erysiphaceae. Host-parasite co-speciation was reported recently between the genus Golovinomyces and Asteraceae from molecular phylogenetic analyses. The Asteraceae originated in South America and latterly expanded their geographic distribution into the Northern Hemisphere. If the co-speciation between Golovinomyces and Asteraceae originated in South America, the geographic origin of Golovinomyces could be assumed to be South America. To address this question, Golovinomyces species from hosts of the tribe Mutisieae, an asteraceous tribe endemic to South America, were collected and the ITS and 28S rDNA regions sequenced. Results indicate that Oidium mutisiae and Golovinomyces leuceriae isolated from the Mutisieae do not belong at the base of the Golovinomyces tree. Instead, they are situated separately within two different clades of Golovinomyces isolates from the Northern Hemisphere. Therefore, the tribe Mutisieae is not the most early host of Golovinomyces. Present results suggest that Golovinomyces originated in the Northern Hemisphere, and not in South America. The new species Oidium reginae for the previous O. mutisiae on Mutisia decurrens is proposed.

Queirozia turbinata (Phyllactinieae, Erysiphaceae): a powdery mildew with a dematiaceous anamorph.

The powdery mildew monotypic genus Queirozia was reduced to a synonym of Pleochaeta in 1982. Now, a re-examination of the type material of Queirozia and of two other specimens showed that Queirozia turbinata has a very distinct combination of characters that do not allow its placement within Pleochaeta. It has forked special aerial hyphae, subclavate and predominantly lemon-shaped conidia, and both conidiophores and conidia varying from greyish to yellowish brown (a feature that appears to be unique for this fungus within the Erysiphales). This may represent the first dematiaceous anamorphic powdery mildew known to science. This fungus also has hemiendophytic mycelium, a character considered almost exclusive to the tribe Phyllactinieae. The outer wall surface pattern of wrinkled and turgid conidia of Q. turbinata as observed under the SEM is different from those presented in the literature for this tribe. Molecular analysis showed that Q. turbinata belongs to the tribe Phyllactinieae and is phylogenetic closer to Pleochaeta than to Leveillula or Phyllactinia. It is therefore acknowledged that Queirozia is closely related to Pleocheta but too distinct morphologically from this genus to be accepted as a synonym. Emended descriptions of Queirozia and Q. turbinata are presented.

Caespitotheca gen. nov., an ancestral genus in the Erysiphales.

The phylogenetic position of Uncinula forestalis within the Erysiphales has been inferred from 5.8S, 18S, and 28S rDNA sequences. Although the appendages of the ascomata are Uncinula-like, i.e. unbranched with curved-coiled apices, U. forestalis is situated at the very base of the large Erysiphales cluster, far from the 'Pseudoidium' clade (Erysiphe, including Microsphaera and Uncinula) and separate from the recently introduced basal genus Parauncinula. U. forestalis differs morphologically from the species of Erysiphe sect. Uncinula in having terminal, fasciculate (as in Podosphaera tridactyla), septate ascoma appendages and a Euoidium-like anamorph (conidia catenate). In Parauncinula, the appendages are also terminal but not fasciculate, the ascospores are curved, and the anamorph is lacking. Uncinula forestalis is a basal, tree-inhabiting powdery mildew with some additional ancestral characteristics, including Uncinula-like appendages and 6-8-spored asci. The new genus Caespitotheca gen. nov. is described with C. forestalis comb. nov. (syn. Uncinula forestalis) as the type species. We calculated the timing of the divergence of U. forestalis and P. septata using a molecular clock of the Erysiphales (6.5 x 10-10 changes per site per year in domains D1 and D2 of 28S rDNA) and a 28S rDNA data set. The results suggest that the divergence of U. forestalis and P. septata from other Ervsiphales occurred between 90 and 80 Myr ago; i.e. the divergence of the two ancestral species may have occurred in the late Cretaceous.