ITS rDNA Data Research Articles

Studies in Hygrocybe s.l. (Hygrocyboideae, Hygrophoraceae) in Brazil: New species of Humidicutis and Neohygrocybe

Humidicutis and Neohygrocybe are genera of wax cap mushrooms closely related to Hygrocybe. This is the first study in Humidicutis and Neohygrocybe from Brazil. We describe two new species, H. pindorama and N. fumosa, based on morphology and ITS rDNA data, and propose a new combination, N. ovinoides. Maximum Likelihood and Bayesian analyses were conducted to reconstruct the phylogenetic relationships of the new species and related taxa. The resulting trees suggest that the new species are unique and show significant similarities to existing taxa in their respective genera. We also provide a key to the species of Humidicutis and Neohygrocybe known in the world.

Characterization of bacterial-associated with Egyptian Tirmania nivea and Terfezia canariensis

Recently, various studies investigate the cultivable microbial diversity of different truffles species. However, this study is still very limited for Egyptian desert truffles. Two morphologically different kinds of wild desert truffles collected from Western Egyptian Desert were characterized using general description in combination with ITS-rDNA data. The phylogeographic analysis of the obtained sequences in comparison to Mediterranean genera from GeneBank revealed the identification of white truffle as Tirmania nivea with 100% identity of isolate from Libya. While the red truffles were Terfezia canariensis which was grouped in one clade with isolates from Canary Island Spain and Portugal. A scanning electron microscope (SEM) analysis indicated bacterial cells inhabiting the healthy gleba of the truffles. The cultivable bacterial isolates were Gram-positive related to the Firmicutes phyla identified as Bacillus boroniphilusMZ361726 (B2), Lactococcus lactisMZ361727 (B4), Bacillus licheniformisMZ361728 (B7), Bacillus haynesiiMZ361729 (B8), and Bacillus velezensisMZ361730 (B11) based on partial sequencing of 16S rRNA. The liquid chromatography-mass spectrometry (LC-MS) of the most potent identified strains (B4, B7, and B11), revealed bioactive compounds showing antimicrobial activity as docosahexaenoic acid, fucosterol, and kolaflavanone which exist in truffle literatures. In conclusion, our findings reveal new insight into the microbiota associated with T. nivea and T. canerienasis and supported that these organisms represent a rich source for the development of novel antimicrobial bioactive compounds.

First Report of Aphelenchoides Bicaudatus (Imamura, 1931) Filipjev and Schuurmans Stekhoven, 1941 Associated with Grass in South Africa.

Aphelenchoides bicaudatus associated with grass in South Africa was identified morphologically and molecularly. This population is characterized by a body length of 409 - 529 μm, a stylet length of 9.5 - 13 μm, a post-vulval uterine sac of 45 - 50 μm, and the characteristic tail bifurcated at the end with one prong longer than the other. Molecular analyses based on the 18S and ITS rDNA data confirmed the primary morphological identification of the A. bicaudatus species. The obtained phylogenetic trees revealed a close positioning of the South African population to other representatives of A. bicaudatus with the maximum (1.00) posterior probability value. Principal component analysis (PCA) also indicated a variation within the populations of A. bicaudatus. This is the first report of A. bicaudatus from South Africa.

Additional Species and ITS rDNA Data for the Lichen Mycota of the Imperial Palace Grounds, Tokyo, Japan

Additional Species and ITS rDNA Data for the Lichen Mycota of the Imperial Palace Grounds, Tokyo, Japan

Molecular Phylogeny and Taxonomy of the Genus Spumella (Chrysophyceae) Based on Morphological and Molecular Evidence.

The genus Spumella, established by Cienkowsky in 1870, is characterized by omnivory, two (rarely three) flagella, a short stick-like structure beneath the flagella, a threadlike stalk, cell division via constriction and cyst formation. Since the first phylogenetic study of Spumella-like flagellates, their paraphyly has consistently been shown, with separation into several genera. More recently, Spumella was carefully investigated using molecular and morphological data to propose seven new species. Classification of this genus and knowledge of its species diversity remain limited because Spumella-like flagellates are extremely difficult to identify based on limited morphological characters. To understand the phylogeny and taxonomy of Spumella, we analyzed molecular and morphological data from 47 strains, including 18 strains isolated from Korean ponds or swamps. Nuclear SSU, ITS and LSU rDNA data were used for maximum likelihood and Bayesian analyses. The molecular data divided the strains into 15 clades, including seven new lineages, each with unique molecular signatures for nuclear SSU rRNA from the E23-2 to E23-5 domains, the spacer between the E23-8 and E23-9 domains of the V4 region and domain 29 of the V5 region. Our results revealed increased species diversity in Spumella. In contrast to the molecular phylogeny results, the taxa showed very similar cell morphologies, suggesting morphological convergence into simple nanoflagellates to enable heterotrophy. Three new species produced stomatocysts in culture. Aspects of stomatocyst morphology, including collar structure, surface ornamentation, and cyst shape, were very useful in differentiating the three species. The general ultrastructure of Spumella bureschii strain Baekdongje012018B8 and S. benthica strain Hwarim032418A5 showed the typical chrysophyte form for the leucoplast, a vestigial chloroplast surrounded by four envelope membranes, supporting the hypothesis that Spumella evolved from a phototroph to a heterotroph via the loss of its photosynthetic ability. Seven new species are proposed: S. benthica, S. communis, S. longicolla, S. oblata, S. rotundata, S. similis, and S. sinechrysos.

Native Philippine Heterorhabditis indica isolates from banana and rice fields and preliminary results of their virulence against the larvae of super worm (Zophobas morio Fabricius Coleoptera: Tenebrionidae)

BackgroundEntomopathogenic nematodes (EPNs) of the families Heterorhabditidae and Steinernematidae are used as biological control agents of several insect pests in lieu of synthetic pesticides.ResultsUsing an insect-baiting technique, naturally occurring EPN isolates were collected from banana and rice fields in the municipality of Hagonoy, the Philippines. A total of 4 isolates were recovered from all the sites namely, HBP1, HBP2, HR3 and HR4. By combining morphometrical and ITS rDNA data, all the isolates were identified as Heterorhabditis indica. To initially assess their infectivity, virulence tests were conducted on the last instar larvae of the super worm, Zophobas morio Fabricius (Coleoptera: Tenebrionidae), the model insect. HR3 isolate had the highest percentage mortality (56%), followed by HBP1 and HBP2 (33%), which were comparable to the available nematode in the market, Heterorhabditis bacteriophora (44%), whereas the lowest was obtained in HR4 isolate (11%).ConclusionsThis study accounted 4 more H. indica isolates in addition to pre-existing isolates in the country, expanding its habitat range and geographic distribution. This also provides baseline information on the potential biocontrol utilization of native EPNs against soil-dwelling insect pests of agricultural crops. Further investigations are required to assess their efficacy against several other insect pests of economic importance for the integrated pest management (IPM) programmes in the country.

Orostachys spinosa (Crassulaceae) origin and diversification: East Asia or South Siberian Mountains? Chloroplast DNA data

Limited data are available on genetic structures of the herbaceous plant species populations inhabiting mountainous areas of Siberia and Northeastern Asian (Russian Far East). Although this area was not directly impacted by the extensive ice-sheets during the Quaternary, it experienced significant climatic fluctuations that influenced rich local flora. Orostachys spinosa (Crassulaceae) lacks any adaptations for long-distance dispersal, yet the species is characterized by an unusually wide range spanning from the Urals to the coast of the Pacific Ocean. We studied O. spinosa phylogeography and genetic diversity across its range sampling 203 individuals from 21 natural populations. Using sequences from three chloroplast DNA non-coding regions, we revealed 82 haplotypes and observed high level of population differentiation indicating presence of the phylogeographic structure (GST = 0.501 and NST = 0.822 (p < 0.01)). In concordance with the previous phylogenetic analyses based on ITS rDNA data, parsimony network revealed two distinct cpDNA haplotype lineages deferring in their structure and characteristics of genetic diversity. The split between these haplotype groups can be dated to the Pliocene (ca. 3.6 Mya). According to our estimates diversification in the Western group of populations took place ca. 1 Mya earlier than in the Eastern group (3.1 Mya and 2.26 Mya, respectively). Apart from the generally accepted notion about East Asian origin of O. spinosa, our results indicated that the species could have originated in mountains of Southern Siberia (Altai). This region was colonized independently from O. thyrsiflora which has a largely overlapping distribution in this area.

Host preference and sorus location correlate with parasite phylogeny in the smut fungal genus Microbotryum (Basidiomycota, Microbotryales)

The smut fungal genus Microbotryum (Microbotryales, Pucciniomycotina) contains species that parasitize plants from many different lineages of euasterids, with host specificity of individual parasite species in general being exceptionally high. Additionally, it has been shown that the location of spore production in some species is related to spore dispersal. In this phylogenetic study based on ITS and LSU rDNA data of 57 Microbotryum spp., host spectra and sorus location are mapped on the phylogeny of Microbotryum species in order to understand the macroevolutionary patterns of these two traits. We find that monophyletic parasite clades correspond well with monophyletic host clades and also that monophyletic parasite groups in general produce their spores in the same plant organ. Ancestral state reconstruction inferred the most probable ancestral trait for sorus location being leaves and the most probable ancestral host family for the genus Microbotryum as being the Polygonaceae. According to molecular analyses, a newly sequenced specimen of Ustilago ducellieri, a seed parasite on Arenaria leptoclados, previously treated as synonym of Microbotryum duriaeanum, belongs to a lineage distinct from specimens of M. duriaeanum. A new combination, Microbotryum ducellieri, is accordingly proposed. Taxonomic implications of the presented analyses for the genera Bauhinus and Haradaea are briefly discussed.

Insights into the Tricholomatineae (Agaricales, Agaricomycetes): a new arrangement of Biannulariaceae and Callistosporium, Callistosporiaceae fam. nov., Xerophorus stat. nov., and Pleurocollybia incorporated into Callistosporium

A new classification of the taxa formerly ascribed to Biannulariaceae (≡ Catathelasmataceae), viz. Catathelasma (type), Callistosporium, Pleurocollybia, Macrocybe, Pseudolaccaria, Guyanagarika and Anupama is here proposed. Phylogenetic inference of the Tricholomatineae based on the analysis of a combined dataset of nuclear genes including ITS, 18S and 28S rDNA, tef1 and rpb2 data supports significantly a monophyletic origin of the aforementioned genera with the exception of Catathelasma, which is significantly related with Bonomyces and Cleistocybe. Biannulariaceae is therefore emended to include the clade formed by Catathelasma, Bonomyces and Cleistocybe. Consequently, the new family Callistosporiaceae is proposed to name the clade containing Callistosporium (= Pleurocollybia) and related genera. Species of Callistosporium with distant lamellae, long hygrophoroid basidia and large amygdaliform spores are accommodated in the new genus Xerophorus. Finally, the new species Callistosporium pseudofelleum and Macrocybe sardoa are described, Clitocybe hesleri and C. fellea are combined into Callistosporium and Pseudolaccaria, respectively, and Callistosporium olivascens var. donadinii is upgraded to species rank and combined into Xerophorus.

Phylogeny and taxonomy of Golovinomyces orontii revisited

Golovinomyces orontii is a common plurivorous powdery mildew species with wide host range and worldwide distribution, usually occurring as asexual morph. Ascomata (chasmothecia) are rarely formed on most hosts. Phylogenetic analyses based on rDNA ITS and 28S rDNA data of a wide range of powdery mildew collections of G. orontii s. lat. suggested a high degree of genetic heterogeneity of this species, which is undoubtedly not monophyletic. This study revealed that sequences retrieved from numerous collections referred to as G. orontii (s. lat.) split into three distinct main clusters, previously classified as groups 1 to 3. These groups have been genetically and morphologically analyzed, circumscribed, and named. One cluster (group 2), including a sequence retrieved from powdery mildew on the type host of Erysiphe orontii, Misopates orontium, constitutes G. orontii s. str. G. tabaci comb. nov. (≡ E. tabaci) is introduced for the second cluster in group 1 that is genetically and morphologically clearly distinct from G. orontii s. str. The third assemblage of sequences (group 3) comprises powdery mildews on hosts of the composite tribe Cichorieae, including Cichorium and Lactuca spp., and a wide range of hosts belonging to various other plant families for which the name G. bolayi sp. nov. is proposed. Euoidium longipes and Euoidium lycopersici, two additional powdery mildew species on solanaceous hosts, are briefly discussed and, based on previous phylogenetic analyses, reallocated to Golovinomyces. Oidium lactucae-debilis on Ixeris japonica in Asia is tentatively reduced to synonymy with G. sonchicola, i.e., it does not pertain to the G. orontii s. lat. complex. G. orontii s. lat. on Vinca spp. (Apocynaceae), mostly known as asexual morph, represents a separate species only distantly related to G. orontii s. str., which is described herein as Golovinomyces vincae sp. nov. Some re-examined collections on Vinca major from Germany misidentified as G. orontii turned out to belong to Ramularia vincae and represent first records of this species for Germany. Golovinomyces spadiceus is a further plurivorous Golovinomyces species discussed in this work, which, however, does not belong to the G. orontii complex.

Morphological, molecular and toxigenic characteristics of Namibian Pseudo-nitzschia species – including Pseudo-nitzschia bucculenta sp. nov.

A field study was undertaken to investigate the occurrence and toxin production of species in the diatom genus Pseudo-nitzschia in Namibian waters, in the extremely productive Benguela upwelling system. From surveys conducted on the R/V Mirabilis and the R/V !Anichab, 52 strains were morphologically determined to species level, supported by nuclear ITS rDNA data. Seven species were identified; P. australis, P. decipiens, P. dolorosa, P. fraudulenta, P. plurisecta, P. pungens var. cingulata, and the new species P. bucculenta F. Gai, C. K. Hedemand, N. Lundholm & Ø. Moestrup sp. nov.Molecular and morphological diversity of the Namibian Pseudo-nitzschia species is discussed. Most importantly, P. bucculenta is both morphologically and phylogenetically most similar to P. dolorosa differing mainly in valve width and densities of striae, poroids and band striae as well as by four hemi-compensatory base changes in the ITS2. Morphological and molecular differences among the strains of P. decipiens suggest a temperate and a warm water subdivision. The geographical and toxigenic characteristics of the identified Pseudo-nitzschia species are described and compared to previous studies. Initial tests of toxin production in all seven species revealed production of domoic acid (DA) in two species: one strain of P. australis (0.074 pg DA cell−1) and two strains of P. plurisecta (0.338 pg DA cell−1 and 0.385 pg DA cell−1).

Bacidina mendax sp. nov., a new widespread species in Central Europe, together with a new combination within the genus Bacidina

AbstractBacidina mendax, described here as a new lichen species, appears to be common and widespread, at least in Central Europe. Analyses of the ITS rDNA region and the morphology of specimens showed an intraspecific variation in the new taxon. It differs from B. neosquamulosa in the lack of a subsquamulose thallus, and from B. caligans in its longer and only slightly curved to apically hooked conidia and lack of a granular (sorediate) thallus. Since ITS rDNA data support the inclusion of Bacidia pycnidiata Czarnota &amp; Coppins in the genus Bacidina, a new combination is proposed.

Anisakis Dujardin, 1845 infection (Nematoda: Anisakidae) in Pygmy Sperm Whale Kogia breviceps Blainville, 1838 from west Pacific region off the coast of Philippine archipelago.

Cetaceans are definitive hosts of anisakid nematodes known to cause human anisakidosis. Despite the reported strandings of different cetaceans in the Philippines, studies on anisakids from these definitive hosts are limited. Here, the morphologically and molecularly identified anisakid species, specifically those of the genus Anisakis Dujardin, 1845 in stranded Pygmy Sperm Whale Kogia breviceps Blainville, 1838 in the west Pacific region off Philippine waters are presented. Morphological data using SEM and LM revealed multi-infections with different Anisakis species belonging to Anisakis type I and type II groups. Molecularly, PCR-RFLP on the ITS rDNA and sequence data analyses of both ITS rDNA and mtDNA cox2 regions identified those from Anisakis type I group as A. typica (Diesing, 1860), whereas those from type II group as A. brevispiculata Dollfus, 1968, and A. paggiae Mattiucci et al. (Syst Parasitol 61:157-171, 2005). This is the first record of Anisakis infection from this host stranded in the west Pacific region off the coast of Philippine waters and new geographical record for A. paggiae.

Studies in the Bark-Dwelling Species ofHymenochaete(Hymenochaetales, Basidiomycota) Reveal Three New Species

Abstract Three new species of Hymenochaete from Russian Far East are described based on morphological and ITS rDNA data. Hymenochaete adhaerens sp. nov. is a resupinate species closely related to H. macrospora, from which it differs in having longer basidiospores and setae. The basidiocarps of H. cupulata sp. nov. are effused-reflexed, resembling those of the European H. ulmicola, from which it differs in having setae inflated at the basal part and shorter basidiospores. Both H. adhaerens and H. cupulata inhabit bark of living elm trees (Ulmus spp.). H. manshurica sp. nov. is resupinate and morphologically similar to the North American H. corticolor, from which it differs in having narrower setae and basidiospores; it has been collected from bark of living Tilia manshurica.

Refugia within refugia: in situ speciation and conservation of threatened Bertmainius (Araneae : Migidae), a new genus of relictual trapdoor spiders endemic to the mesic zone of south-western Australia

The trapdoor spider family Migidae has a classical Gondwanan distribution and is found on all southern continents except the Indian region. The Australian fauna consists of three genera including Moggridgea O. P. Cambridge from south-western Australia and Kangaroo Island, South Australia; Moggridgea is otherwise widespread throughout Africa. The sole named species of Moggridgea from Western Australia, M. tingle Main, and its unnamed relatives are the subject of the present paper, which was stimulated by concern for the long-term persistence of populations, and the discovery of deep genetic divergences between populations. A phylogeny of the Western Australian species relative to African and South Australian Moggridgea was generated using molecular COI and ITS rDNA data, and based on both molecular and morphological criteria we conclude that the Western Australian taxa should be removed from Moggridgea and transferred to a new genus, Bertmainius. The seven species are delimited using both morphological and molecular criteria: B. tingle (Main) (the type species), and six new species, B. colonus, B. monachus, B. mysticus, B. opimus, B. pandus and B. tumidus. All seven species are considered to be threatened using IUCN criteria, with the major threatening processes being inappropriate fire regimes and climate change.

Planktochlorella nurekis gen. et sp. nov. (Trebouxiophyceae, Chlorophyta), a novel coccoid green alga carrying significant biotechnological potential.

Phylogenetic position, morphology and ultrastructure were investigated for biotechnologically remarkable strain Chlorella vulgaris IFR C-111, utilized in various commercial applications. Molecular phylogenetic analyses based on the SSU and ITS rDNA data revealed that the strain IFR C-111 forms a distinct lineage within the Parachlorella-clade in Chlorellaceae. We describe this organism as a new genus and species, Planktochlorella nurekis. Vegetative cells of this newly recognized species are spherical, possessing a single pot-shaped chloroplast with starch-covered pyrenoid. Asexually it reproduces by the formation of 2-16(-32) autospores. Cell wall is composed of two layers, the outer layer containing extended microfibrillar material. The fuzzy cell wall structure improves the buoyancy resulting in low sedimentation rate of P. nurekis. To resolve the phylogenetic position of Planktochlorella and its relationship to the closely related genera, nucleotide saturation present in the ITS rDNA data was reduced by four different approaches. The resulting topologies pointed to the poor phylogenetic signal in generally utilized SSU and ITS rDNA data and the need of sequencing other molecular markers.

New Hypoxylon species from Martinique and new evidence on the molecular phylogeny of Hypoxylon based on ITS rDNA and β-tubulin data

Three new species of Hypoxylon (Xylariaceae) collected from Martinique in the French Caribbean are recognised by new combinations of morphological characters. Their status as undescribed taxa was supported by secondary metabolite profiling based on High performance liquid chromatography with diode array and mass spectrometric detection (HPLC/DAD-MS) as well as by comparison of ITS and partial s-tubulin DNA sequences with related taxa. In the course of this study, the teleomorph of Nodulisporium griseobrunneum was found, and this species could be transferred to Hypoxylon. Moreover, several names in Hypoxylon are epitypified by selecting recently collected specimens from the same geographic areas as the holotypes came from. Despite the fact that our study used the hitherto most extensive taxon sampling, the phylogenetic analyses inferred from ITS and s-tubulin sequences remain contradictory to each other, and neither genealogy was found fully in agreement with phenotype-derived traits. We conclude that the right gene (or multi-gene genealogies) to reflect the phylogeny and evolution of Hypoxylon still remains to be found. For the time being, we recommend that the application of polyphasic taxonomic concepts should be continued in taxonomic studies of Hypoxylon.

A new Cortinarius of section Calochroi (Basidiomycota, Agaricomycetes) from Mediterranean Quercus woodlands (Italy)

A new species of Cortinarius, C. flavoaurantians sp. nov., is described from Italian Quercus woods based on both morphological and ITS rDNA data. This taxon is characterized by a yellowish pileus and cortina, a white universal veil and a pileipellis that reacts yellow-orange with KOH. Illustrations of the key micromorphological features and fresh basidiomata in situ are provided. Closely related species are discussed.

Molecular phylogeny of Sydowiellaceae—resolving the position of Cainiella

Cainiella is an ascomycete genus associated with arctic alpine plants. The systematic position of Cainiella has long been unclear, with current classifications placing the genus in either Sordariales or Xylariales. Our molecular results, based on mtSSU, ITS and nLSU rDNA data, clearly show that the genus belongs in the Sydowiellaceae (Diaporthales). The study also includes new sequences of Sydowiellaceae and contributes to a better knowledge of the phylogenetic relationships of that family.

Isomucor (Mucoromycotina): a new genus from a Cerrado reserve in state of São Paulo, Brazil

During a survey of mucoralean fungi from a Cerrado reserve (Brazilian savanna) some isolates of a Mucor-like fungus were isolated from soil plates. Characterization based on morphological, physiological and molecular data from translation elongation factor (EF-1α), 28S (D1/D2) and ITS1-5.8S-ITS2 rDNA sequences was made. The isolates produce lateral branches bearing multispored sporangiola in addition to the multispored sporangia and a uniformly septate mycelium as the main differentiating characteristics. To our surprise this fungus possesses two EF-1α genes 1.4 and 1.5 kb long. Evidence from the analyzed datasets supports the delimitation of a new genus and the inclusion of Mucor fuscus based on 28S and ITS rDNA data.