Amanita theophili sp. nov. (Amanitaceae) from central Mexico

Research on the molecular systematics of Cortinarius, a species-rich mushroom genus with nearly global distribution, is just beginning. The present study explores infrageneric relationships using rDNA ITS and LSU sequence data. One large dataset of 132 rDNA ITS sequences and one combined da-taset with 54 rDNA ITS and LSU sequences were generated. Hebeloma was used as outgroup. Bayesian analyses and maximum-likelihood (ML) analyses were carried out. Bayesian phylogenetic inference performed equally well or better than ML, especially in large datasets. The phylogenetic analysis of the combined dataset with species representing all currently recognized subgenera recovered seven well-supported clades (Bayesian posterior probabilities BPP > 90%). These major clades are: /Myxacium s.l., /subg. Cortinarius, the /phlegmacioid clade (including the subclades /Phlegmacium and /Delibuti), the /calochroid clade (/Calochroi, /Ochroleuci and /Allutus), the /telamonioid clade (/Telamonia, /Orellani, /Anomali), /Dermocybe s.l. and /Myxotelamonia. Our results show that Cortinarius consists of many lineages, but the relationships among these clades could not be elucidated. On one hand, the low divergence in rDNA sequences can be held responsible for this; on the other hand, taxon sampling is problematic in Cortinarius phylogeny. Because of the incredibly high diversity (~2000 Cortinarius species), our sampling included <5% of the known species. By choosing type species of subgenera and sections, our sampling is strongly biased toward Northern Hemisphere taxa. More extensive taxon sampling, especially of species from the Southern Hemisphere, is essential to resolve the phylogeny of this important genus of ectomycorrhizal fungi.

Research on the molecular systematics of Cortinarius, a species-rich mushroom genus with nearly global distribution, is just beginning. The present study explores infrageneric relationships using rDNA ITS and LSU sequence data. One large dataset of 132 rDNA ITS sequences and one combined da-taset with 54 rDNA ITS and LSU sequences were generated. Hebeloma was used as outgroup. Bayesian analyses and maximum-likelihood (ML) analyses were carried out. Bayesian phylogenetic inference performed equally well or better than ML, especially in large datasets. The phylogenetic analysis of the combined dataset with species representing all currently recognized subgenera recovered seven well-supported clades (Bayesian posterior probabilities BPP > 90%). These major clades are: /Myxacium s.l., /subg. Cortinarius, the /phlegmacioid clade (including the subclades /Phlegmacium and /Delibuti), the /calochroid clade (/Calochroi, /Ochroleuci and /Allutus), the /telamonioid clade (/Telamonia, /Orellani, /Anomali), /Dermocybe s.l. and /Myxotelamonia. Our results show that Cortinarius consists of many lineages, but the relationships among these clades could not be elucidated. On one hand, the low divergence in rDNA sequences can be held responsible for this; on the other hand, taxon sampling is problematic in Cortinarius phylogeny. Because of the incredibly high diversity (~2000 Cortinarius species), our sampling included <5% of the known species. By choosing type species of subgenera and sections, our sampling is strongly biased toward Northern Hemisphere taxa. More extensive taxon sampling, especially of species from the Southern Hemisphere, is essential to resolve the phylogeny of this important genus of ectomycorrhizal fungi.

Since the first formal description of the unicellular green algae Dunaliella salina, its presence in hypersaline environments worldwide and its physiological responses to different environmental conditions have been studied extensively. Moreover, due to massive carotenoid accumulation by some strains under specific growth conditions, its biotechnological applications have attracted a great deal of scientific interest. In this study, the phylogenetic relationship, growth, and carotenogenesis of a new strain of Dunaliella salina isolated from Maharlu salt lake in Shiraz (latitude 29.26°N, longitude 52.48°E), Iran were investigated. First, a phylogram based on neighbor-joining analysis of the nuclear rDNA ITS (ITS-1 + 5.8 rDNA + ITS-2) sequence data was constructed. The phylogenetic tree showed that the new isolate is part of a major clade containing several strains of D. salina and was designated as D. salina MSI-1. Then, the responses of the new isolate to the initial pH of the culture media and different concentrations of nitrate, NH4+, and citrate were examined. As with other strains of D. salina, growth and carotenogenesis were controlled by the levels of nitrate and NH4+ in the growth media. Low available nitrogen negatively affected growth but enhanced carotenoid accumulation. Insensitivity of carotenogenesis to citrate indicates a minor contribution of cytosolic IPP synthesis to the overall carotenoid production. Despite changes in the initial pH of the culture media over the experimental period, the initial pH had marked effects on the growth and carotenogenesis of the new isolate. These effects, together with the higher cell carotenoid content observed at pH 11.0, await further research. The results confirm that the analysis of the ITS sequences is a reliable basis for determination of the genetic relatedness among strains of the genus Dunaliella, and the search for strains with novel characteristics may have valuable biotechnological applications.

Phylogenetic relationships of Puccinia horiana and other rust pathogens of Chrysanthemum × morifolium based on rDNA ITS sequence analysis

The enigmatic Spirosphaera lignicola is revised based on recent collections from Austria. The taxon does not fit the generic circumscription of the genus Spirosphaera in its morphological features but clearly belongs to Dendroclathra. Nuclear ITS rDNA sequence data from recent collections and the type specimens indicate conspecificity of S. lignicola with Dendroclathra caeruleofusca. Spirosphaera lignicola is transferred to the genus Dendroclathra, and a recent collection with an ITS sequence identical to the type specimen is designated as epitype. Morphology is illustrated with SEM and LM pictures. Phylogenetic analyses of LSU, ITS and tef1 sequence data reveal a phylogenetic affinity of Dendroclathra to the Microascales (Hypocreomycetidae), being therefore phylogenetically distant from Spirosphaera floriformis, the generic type, which belongs to the Leotiomycetes. The recent collections are the first records of this species for Austria. The distribution and aeroaquatic ecology of D. lignicola are briefly discussed.

Fourteen new species in the Latin American Clade (LAC) of the Ceratocystis fimbriata complex recently were distinguished from C. fimbriata sensu stricto largely based on variation in ITS rDNA sequences. Among the 116 isolates representing the LAC, there were 41 ITS haplotypes. Maximum parsimony (MP) analysis of ITS sequences produced poorly resolved trees. In contrast, analyses of mating-type genes (MAT1-1-2 and MAT1-2-1) resolved a single MP tree with branches of high bootstrap and posterior probability support. Four isolates showed intragenomic variation in ITS sequences. Cloning and sequencing of PCR products from a single haploid strain identified two or more ITS sequences differing at up to 16 base positions and representing two described species. Isolates from introduced populations that appeared to be clonal based on microsatellite markers varied at up to 14 bp in ITS sequence. Strains of seven Brazilian ITS haplotypes and an isolate from Ipomoea batatas (on which the species name C. fimbriata was based) were fully interfertile in sexual crosses. These analyses support three phylogenetic species that differ in pathogenicity: C. platani, C. cacaofunesta and C. colombiana. Five ITS species (C. manginecans, C. mangicola, C. mangivora, C. acaciivora, C. eucalypticola) appear to be ITS haplotypes that have been moved from or within Brazil on nursery stock. The taxonomic status of other species delineated primarily by ITS sequences (C. diversiconidia, C. papillata, C. neglecta, C. ecuadoriana, C. fimbriatomima, C. curvata) needs further study, but they are considered doubtful species.

Fourteen new species in the Latin American Clade (LAC) of the Ceratocystis fimbriata complex recently were distinguished from C. fimbriata sensu stricto largely based on variation in ITS rDNA sequences. Among the 116 isolates representing the LAC, there were 41 ITS haplotypes. Maximum parsimony (MP) analysis of ITS sequences produced poorly resolved trees. In contrast, analyses of mating-type genes (MAT1-1-2 and MAT1-2-1) resolved a single MP tree with branches of high bootstrap and posterior probability support. Four isolates showed intragenomic variation in ITS sequences. Cloning and sequencing of PCR products from a single haploid strain identified two or more ITS sequences differing at up to 16 base positions and representing two described species. Isolates from introduced populations that appeared to be clonal based on microsatellite markers varied at up to 14 bp in ITS sequence. Strains of seven Brazilian ITS haplotypes and an isolate from Ipomoea batatas (on which the species name C. fimbriata was based) were fully interfertile in sexual crosses. These analyses support three phylogenetic species that differ in pathogenicity: C. platani, C. cacaofunesta and C. colombiana. Five ITS species (C. manginecans, C. mangicola, C. mangivora, C. acaciivora, C. eucalypticola) appear to be ITS haplotypes that have been moved from or within Brazil on nursery stock. The taxonomic status of other species delineated primarily by ITS sequences (C. diversiconidia, C. papillata, C. neglecta, C. ecuadoriana, C. fimbriatomima, C. curvata) needs further study, but they are considered doubtful species.

To study the genetic diversity of rDNA ITS sequences in different species of Bai Shouwu, utilize the molecular diversity of ITS sequences to authenticate the different species of Bai Shouwu. Firstly, total DNA was extracted from the different species of Bai Shouwu. Secondly, the ITS sequence was amplified by PCR with universal primer of ITS and sequenced after cloning and purification. From four species the complete sequence of ITS and 5.8 S rDNA, the partial sequences of 18S rDNA and 26S rDNA were obtained. The rDNA ITS sequences of Cynanchum bungei (sign in No. GU198970 and No. GU479037) were obtained. Ten variable sites among the sequences were found. ITS sequence could be used to authenticate the species. The method could be used to identify germplasm resources and authenticate.

Eleven samples of the most important pistachio rust (caused by Pileolaria terebinthi (DC.) Cast.,), which causes disease on Beneh (Pistacia atlantica Desf. subsp. mutica (Fisch. &amp; Mey.) Rech. F) and Kasoor (Pistacia khinjuk Stocks.), were collected from herbarium specimens and pistachio fields at the Pistachio Research Institute in Rafsanjan, Iran. The complete sequences of ribosomal DNA internal transcribed spacers ITS1 and ITS2 (rDNA ITS) from the samples were determined and analysed. In general, very little rDNA ITS sequence variation was observed between rDNA ITS sequences of P. terebinthi samples. The length of the PCR fragments was 621 bp (for ITS1F-ITS4) and 1177 bp (for ITS1F-rust1), and consisted of 67 bp at the 3 ́ end of 18S rDNA, 93 bp of ITS1 region, 154 bp of 5.8S rDNA, 246 bp of the ITS2 region, 57 bp (for ITS1F-ITS4) and 613 bp (for ITS1F-rust1) at the 5 ́ end of the 28S rDNA. Restriction fragment length polymorphisms (RFLPs) of the rDNA-ITS region were used to identify Pileolaria terebinthi. Three strong bands of 105, 134 and 381 bp and five bands of 105, 134, 200, 301 and 437 bp are observed for the fragment of “ITS1F-ITS4” and “ITS1F-rust1”, respectively. A PCR-RFLP diagnostic technique provided effective identification of the species by a unique pattern with the specific restriction enzyme XapI (ApoI).

Aims: To optimize the culture media of 0288 Tricholoma matsutake and to identify it according to its rDNA ITS sequence. Methods: Fruit body, collected from Heilongjiang province, is used as the material, and the mycelium is isolated and radiated under ultraviolet. Then the radiated mycelium is cultured in five different media and its morphology of fungus colony and growth rate are investigated. After that, the rDNA ITS sequences of 0288 and the natural fruit body of T. matsutake are analyzed and identified by PCR and sequencing. Results: The radiated mycelium grew faster in formula A culture medium than others and its average growth rate is up to 0.575mm/d. Results also showed that the homology of the rDNA ITS sequence is almost up to 99% between 0288 and the natural fruit body. Through this experiment, the best growth condition of 0288 is obtained and its fruit body is induced as pure cultural product. It would lay strong foundation for the artificial domestication of the wild T. matsutake.

Since 2003, Torenia fournieri plants grown for experimental purposes were repeatedly infected by powdery mildew in a laboratory in Hungary. Based on morphological characteristics, the pathogen belonged to the mitosporic genus Oidium subgen. Reticuloidium, the anamorph stage of Golovinomyces. The rDNA ITS sequence was identical to that of two other powdery mildew fungi, infecting Arabidopsis and Veronica, respectively, in different parts of the world. According to a previous phylogenetic analysis of ITS and 28S rDNA sequences, those two powdery mildews belong to a recently evolved group of Golovinomyces characterized by multiple host range expansions during their evolution. Both the ITS sequence and the morphological data indicate that the powdery mildew anamorph infecting Torenia also belongs to this group. It is likely that the powdery mildew infections of the experimental T. fournieri plants, native to south-east Asia, were the result of a very recent host range expansion of a polyphagous Golovinomyces because (i) T. fournieri is absent from our region, except as an experimental plant grown in the laboratory, (ii) the powdery mildew fungus infecting this exotic plant belongs to a group of Golovinomyces where host range expansion is a frequent evolutionary scenario, (iii) cross-inoculation tests showed that this pathogen is also able to infect other plant species, notably A. thaliana and tobacco, and (iv) no Golovinomyces species are known to infect T. fournieri anywhere in the world. Although host range expansion has often been proposed as a common evolutionary process in the Erysiphales, and also in other biotrophic plant pathogens, this has not been clearly demonstrated in any case studies so far. To our knowledge, this is the first convincing case of a host range expansion event in the Erysiphales.

This paper presents the results of phylogenetic analysis of three morphospecies in the Daedaleopsis genus (Polyporaceae, Agaricomycetes): D. confragosa, D. septentrionalis, and D. tricolor. It has been shown that the rDNA ITS sequences of D. confragosa, D. tricolor, and D. septentrionalis from the Urals, Siberia, and the Far East form a single cluster on the phylogenetic tree. The nucleotide similarity of the ITS sequences from the analyzed morphospecies is 98.95–99.29% and falls within its intragroup values range (98.65–99.44%). The nucleotide divergence (Dxy) of the ITS sequences from these morphospecies is 0.69–1.08% and does not exceed the intragroup nucleotide diversity values (π): 0.52–1.34%. Based on the nucleotide divergence and nucleotide similarity of the rDNA ITS sequences, the differences between D. confragosa, D. tricolor, and D. septentrionalis do not reach levels which would allow one to recognize them as separate species. Given this, and the sympatric nature of D. confragosa, D. tricolor, and D. septentrionalis distribution, they should be considered three morphological varieties of D. confragosa. From the ecological point of view, they can be considered three ecotypes, one of them (D. confragosa) being confined to azonal biotopes and the two other being ecotypes adapted to latitude-specific climates: north boreal (D. septentrionalis) and south boreal (D. tricolor).

Taxonomy of the genus Leveillula has long been considered as a challenge in powdery mildew systematics. The rDNA diversity has recently been used for phylogenetic analysis of several specimens of the genus Leveillula. In the present study, additional rDNA ITS sequences are provided and a new phylogenetic analysis is carried out aiming at a better understanding of the genetic diversity in the genus Leveillula. New analyses confirmed that L. taurica is unique in the genus, as it exhibits an intraspecific gene sequence diversity considerably higher than in other species. In several cases L. taurica s. lat. on a certain host plant species has a sequence different from L. taurica on other host plants. Moreover, DNA data indicated different lineages among L. taurica specimens which were hardly distinguishable by morphology. More than one genotype occurring on a single host is sometimes possible. According to these results, several races such as Leveillula on Artemisia, Acroptilon, Onobrychis, are molecularly well characterized. While there is enough molecular evidence to delimit such races as independent taxa, clear morphological delimitations between these new and already published taxa are very difficult or even impossible. However, ecological features, and above all, host specificity for biotrophic fungi such as powdery mildew, would be a good criterion to discriminate cryptic taxa along with rDNA sequences. In fact, many collections of Leveillula strains on different hosts show their own type of conidial morphology, which is usually consistent for a strain on a single host species. Hence, we have proposed to describe new species for Leveillula on some host plants such as Artemisia, Acroptilon, Echinops and Onobrychis.

Several taxonomic problems arise in the group of small, white European truffles, probably due to the over-emphasized significance of certain morphological features of ascomata. The distinction between Tuber rapaeodorum Tul. & C. Tul. and Tuber borchii Vittad. and Tuber puberulum Berk. & Broome has not been accepted in several recent studies. Furthermore, the existence of T. rapaeodorum been questioned in some recent synopses of the genus. We conducted microscopic and ITS sequence investigations of 31 herbarium specimens. Using morphological features such as peridium structure, form and size of spores and dermatocystidia and spore numbers per ascus, we could distinguish T. borchii, T. foetidum Vittad., T. maculatum Vittad., Tuber puberulum, and T. rapaeodorum. Analysis of whole ITS sequences showed sharp differences among the morphologically separated groups. Neighbour-joining and parsimony methods produced highly supported branches and confirmed the identity of these species.

Caractérisation moléculaire des populations de Diplozoidae sur cinq espèces de Cyprinidae: nouvelles données sur la spécificité parasitaire