Phenotypic and phylogenetic characterization of actinomycetes isolated from <Emphasis Type="Italic">Lasius niger</Emphasis> and <Emphasis Type="Italic">Formica cunicularia</Emphasis> ants

Multiple actinomycete strains were isolated from two ant species, Lasius niger and Formica cunicularia, and their phenotypic properties and phylogenetic position were studied. Partial sequencing of 16S rRNA assigned the greater part of them to the genus Streptomyces, but only one belonged to Nocardia. However, some isolates had significant color and morphological differences from their closest phylogenetic relatives. The abundance and biodiversity of actinomycete communities isolated from L. niger ants greatly exceeded those found for F. cunicularia. All of the actinomycetes associated with F. cunicularia ants demonstrated cellulolytic activity, but only one had such ability among the strains associated with black ants.     

Molecular taxonomy and morphological characterization reveal new species and new host records of <Emphasis Type="Italic">Torula</Emphasis> species (Torulaceae,Pleosporales)

Four new species and two new host records of Torula (Torulaceae, Pleosporales) are described and illustrated from herbaceous litter collected in Italy and Thailand. The new species possess colony, conidiophore and conidial characteristics that fit within the generic concept of Torula. Detailed morphological observations clearly demarcate four of these from extant species and are hence described as new (Torula chiangmaiensis sp. nov., Torula pluriseptata sp. nov., Torula chromolaenae sp. nov., Torula mackenziei sp. nov.). Details of asexual morphs are described, and justifications for establishing these new species are provided. The nuclear are sequenced ribosomal RNA genes as well as protein coding genes to infer phylogenetic relationships and discuss phylogenetic affinities with morphologically similar species. Our morphological distinction is further supported by phylogenetic discrimination. In particular, phylogenies depict a close relationship of Torula chiangmaiensis and T. pluriseptata to T. hollandica, while T. chromolaenae and T. mackenziei constitute an independent phylogenetic lineage basal to T. herbarum and T. ficus. Torula ficus and T. masonii are also described and their phylogeny investigated as new host records from Bidens pilosa and Iris germanica, respectively.     

Phylogenetic relationships,molecular taxonomy,biogeography of <Emphasis Type="Italic">Betula</Emphasis>, with emphasis on phylogenetic position of Iranian populations

Birch trees inhabiting the high-altitude Hyrcanian forest (the southern shores of the Caspian Sea of Iran and Azerbaijan) are classified in the EN (endangered) category of the International Union for Conservation of Nature (IUCN). Taxonomic status and phylogenetic relationship of Iranian populations, molecular taxonomy, and biogeography of the genus Betula in the world have been considered. Four remnant populations of Betula were selected from north and northwestern parts of Iran. The internal ITS and trnH-psbA intergenic spacer regions were sequenced. Based on the trnH-psbA and ITS, Iranian birch and white birch were placed in a clade, but based on trnH-psbA divided into two subclades. Phylogenetic trees based on ITS and trnH-psbA data did not completely support the morphological classification. Network analysis confirms a close relationship of B. pendula with B. platyphylla and B. papyrifera with B. humilis, and B. ermanii were in a group with the other tetraploid species of the subgenus Neurobetula. Divergence time analysis showed that about 75 Ma ago the Betula genus separated from the other genus of Betulaceae and then divided into two main clades in Oligocene. Our divergence analysis supports that two subgenera of Betulenta and Betulaster are the oldest subgenera in the genus Betula and they date back to Eocene. The ancestral reconstruction suggests that ancestors of the genus Betula originated from Southeast Asia.     

RAPD-PCR Analysis of Ground Squirrels from the Tobol-Ishim Interfluve: Evidence for Interspecific Hybridization between Ground Squirrel Species <Emphasis Type="Italic">Spermophilus major</Emphasis> and <Emphasis Type="Italic">S. erythrogenys</Emphasis>

Populations of two ground squirrel species, Spermophilus major and S. erythrogenys, from the interfluvial area of the Tobol and Ishim rivers, where their ranges overlap, have been examined using RAPD-PCR. We have identified 253 loci, which included taxon-specific markers for S. major and S. erythrogenys as well as markers for geographic populations. Estimation of genetic diversity and construction of phylogenetic relationships were performed using software programs POPGENE, TEPGA, and TREECON. In all, based on morphological traits, animals from the Tobol-Ishim interfluve were assigned to the two parental morphotypes and showed similar levels of genetic variability (H, n_a, n_e). However, the total polymorphism level proved to be higher in ground squirrels with the major morphotype (P = 40.32%,P₉₅ = 27.27%) than in animals with the erythrogenys morphotype (P = 32%,P₉₅ = 22.13%). Nevertheless, the number of rare alleles was high in both cases, constituting about 70% of the total number. Interpopulation differentiation was considerably higher in S. major δ = 0.50) than in S. erythrogenys δ = 0.41). The genetic differentiation between local samples from the Tobol-Ishim interfluvial area was lower than that between the parental species. A significant part of the genetic diversity of the species examined and animals from the zone of overlapping ranges was accounted for by intrapopulation variability. Animals from the northern and southern parts of the Tobol-Ishim interfluve were charac-terized by the core traits of S. major and S. erythrogenys, respectively, falling into two distinct clusters in the UPGMA and NJ reconstructions. In addition to three hybrid individuals, identified by the bioacoustic method, three hybrid animals were distinguished using RAPD analysis. These animals earlier were thought to be “pure” species and formed their own clusters in phylogenetic reconstructions. Thus, the RAPD-PCR results directly showed the existence of stable hybridization (20% genetic hybrids) between S. major and S. erythrogenys in the Tobol-Ishim interfluvial area, which is more extensive than inferred previously from morphological and bioacoustic data.     

Taxonomic implications of the morphological and genetic variation of cultivated and domesticated populations of the <Emphasis Type="Italic">Agave angustifolia</Emphasis> complex (Agavoideae,Asparagaceae) in Oaxaca,Mexico

The Agave angustifolia complex, distributed from Mexico to Costa Rica, comprises four species and five varieties, including three species used for mescal production. The complex is represented in the Mexican state of Oaxaca by two wild taxa, A. angustifolia var. angustifolia and A. angustifolia var. rubescens, the cultivated form A. angustifolia “Espadín” and the partially cultivated species A. rodacantha. The aims of this study were to investigate the morphological and genetic variation of the A. angustifolia complex in the state of Oaxaca and to identify traits useful for taxonomic delimitation. Four wild and three cultivated populations of A. angustifolia from Oaxaca, one population of A. tequilana from Guanajuato and one population of A. angustifolia from Sonora were sampled for morphological, genetic and cytometric analyses. We showed that cultivated populations of A. angustifolia “Espadin,” A. rhodacantha and A. tequilana could be clearly differentiated from wild populations. Furthermore, the domesticated populations of A. angustifolia, known locally as “Espadin,” had a higher ploidy level and lower genetic variation than their related wild populations. The population of A. angustifolia from Sonora could be recognized as a different entity. Populations of A. rhodacantha need to be studied throughout their entire distribution area to further evaluate their taxonomic delimitation.     

Three new <Emphasis Type="Italic">Penicillium</Emphasis> species from marine subaqueous soils

During investigation of the mycobiota in the Sea of Okhotsk, three new Penicillium species were isolated from subaqueous soils collected in the Sakhalin Gulf and in the northeastern part of the Sakhalin shelf, near the Piltun Bay. According to Raper and Thom’s classification, the newly described species P. piltunense, P. ochotense, and P. attenuatum belong to subsection Divaricata. Two analyses were performed to reveal the phylogenetic relationships of the putative new species with other Penicillium species. Phylogenetically, the new species are related to the members of the P. canescens group and share some morphological and physiological features with them. However, they differ in having a larger colony diameter that makes them similar to P. atrovenetum, P. coralligerum, and P. antarcticum. A detailed analysis based on ITS, combined β-tubulin and calmodulin datasets, and morphological features revealed that the new species formed a distinct group inside the P. atrovenetum subclade. Moreover, P. attenuatum differs from the other two species macro- and micromorphologically and may represent a distinct phylogenetic lineage.     

Evolution and systematics of Green Bush-crickets (Orthoptera: Tettigoniidae: <Emphasis Type="Italic">Tettigonia</Emphasis>) in the Western Palaearctic: testing concordance between molecular,acoustic, and morphological data

The genus Tettigonia includes 26 species distributed in the Palaearctic region. Though the Green Bush-crickets are widespread in Europe and common in a variety of habitats throughout the Palaearctic ecozone, the genus is still in need of scientific attention due to the presence of a multitude of poorly explored taxa. In the present study, we sought to clarify the evolutionary relationships of Green Bush-crickets and the composition of taxa occurring in the Western Palaearctic. Based on populations from 24 disjunct localities, the phylogeny of the group was estimated using sequences of the cytochrome oxidase subunit I (COI) and the internal transcribed spacers 1 and 2 (ITS1 and ITS2). Morphological and acoustic variation documented for the examined populations and taxa was interpreted in the context of phylogenetic relationships inferred from our genetic analyses. The trees generated in the present study supported the existence of three main lineages: “A”—composed of all sampled populations of Tettigonia viridissima and the Tettigonia vaucheriana complex, “B”—comprising Tettigonia caudata, Tettigonia uvarovi, and the Tettigonia armeniaca complex, and “C”—consisting of Tettigonia cantans. The present study provides the first phylogenetic foundation for reviewing the systematics of Tettigonia (currently classified mostly according to morphological characteristics), proposing seven new synonymies.     

Genome sizes and phylogenetic relationships suggest recent divergence of closely related species of the <Emphasis Type="Italic">Limonium vulgare</Emphasis> complex (Plumbaginaceae)

Limonium vulgare and related species form a complex group, but until now cytological and genetic studies have been based on single species and specific geographical areas. We investigated genome size, karyological and genetic diversity in samples from Western Mediterranean and evaluated the phylogenetic relationships among the species of this complex. Genome size was assessed using flow cytometry on samples from natural populations of L. vulgare, L. maritimum and L. narbonense. Chromosome counts were conducted in plants obtained from seeds collected in the field. The internal transcribed spacer ITS1 of the nuclear rDNAs was used to assess ITS polymorphisms as well as the phylogenetic relationships within the L. vulgare complex. Our analyses showed that all species were tetraploid, with the chromosome number of L. maritimum being presented here for the first time. Significant differences were observed in genome size, with L. narbonense having lower genome sizes than the other two species, and possible aneuploids being detected. Ten new ITS sequences from L. vulgare, L. narbonense and L. maritimum were provided. Most species’ populations showed unique ribotypes, and L. narbonense has the highest ribotype diversity. One of the L. maritimum populations presented a closer genetic relationship with L. vulgare, whereas the other two seemed to be more related with L. narbonense. Phylogenetic analyses confirmed that L. vulgare and L. narbonense form a monophyletic group, sister to the remaining Limonium species. Our results put into evidence that the studied species may represent a relatively early stage of divergence.     

Comparative phylogeography of capitulate <Emphasis Type="Italic">Campanula</Emphasis> species from the Balkans,with description of a new species, <Emphasis Type="Italic">C. daucoides</Emphasis>

Capitulate inflorescence is a specific, strongly adaptive and rare feature in the genus Campanula. We studied morphologically eight capitulate Campanula taxa from the Balkans (1537 individuals/52 populations) and one more species from Caucasus at the molecular level (using chloroplast markers trnG^UCC-trnS^GCU and psbA-trnH, 130 individuals/58 populations) to assess their relations and evolutionary histories. Although all studied taxa were well circumscribed at both the morphological and molecular levels (except morphologically distinct but genetically invariable C. moesiaca which acquired its single haplotype via past cytoplasmic introgression from C. cervicaria), their relations inferred from the two datasets were incongruent possibly due to the homoplasy of morphological characters frequently reported in Campanula. Interspecific hybridization and introgression affected majority of studied species and may be more common in Campanula than previously thought. These processes, along with incomplete lineage sorting and retention of ancestral polymorphisms, hampered our phylogenetic reconstructions and prevented us to fully resolve species relations, and to support monophyletic origin of capitulate Campanula species. Nonetheless, several cryptic taxa were delineated, and C. daucoides was described as a new capitulate Campanula species. Different evolutionary histories and multiple glacial refugia were inferred for all species represented by multiple samples (except C. moesiaca). According to our dating, their speciation was in most cases triggered by various geo-historic events such as the uplift of the Alpide belt, Messinian Salinity Crisis, or desiccation of the Pannonian Sea/Pliocene Lakes from the central Balkans, while their further diversification was mainly driven by the onset of the Quaternary and cycles of glacials/interglacials.     

<Emphasis Type="Italic">Cortinarius</Emphasis> sect. <Emphasis Type="Italic">Riederi</Emphasis>: taxonomy and phylogeny of the new section with European and North American distribution

Cortinarius is one of the most species-rich genera of mushroom-forming fungi. Based on phylogenetic and morphological evidence, Cortinarius, sect. Riederi, is introduced at sectional level (= subsect. Riederi sensu Brandrud & Melot). The taxonomy, phylogeny, ecology and distribution of not only mainly European but also including some North American taxa of this section are treated, which includes nine species and two varieties. Of these, three taxa are described as new (C. burlinghamiae, C. pallidoriederi and C. argenteolilacinus var. dovrensis). The sect. Riederi species possess morphological features similar to Phlegmacium group(s) and forms a phylogenetically isolated lineage, with no supported affinity to other phlegmacioid groups. Three taxa are known from both Europe and North America, two species are known only from North America and five only from Europe. Altogether, eight of the ten taxa are associated with conifers or northern (boreal-subalpine) deciduous trees (Betula spp.). Only two species occur in more temperate forests (Fagus forests), and no species have so far been found in thermophilous Quercus forests     

Variability of quantitative morphological traits of mosquito larvae in some species of the <Emphasis Type="Italic">Culex pipiens</Emphasis> complex (Diptera,Culicidae)

The variability of 13 larval morphological characteristics in the urban mosquito, Culex pipiens f. molestus and Culex torrentium, was investigated. These species, belonging to the Culex pipiens complex, are characterized by rapid ecological and evolutionary processes following the human-induced environmental changes. A comparison of 4 geographical larval populations of C. pipiens f. molestus and Culex torrentium revealed different rates of divergence of different morphological traits. In the older form in terms of evolutionary development, C. torrentium, the two local populations differed in the structures of the trophic morphological complex characterized by a higher functional trophic load, while the structures with a lower functional load did not differ between the populations. On the contrary, the populations of the younger form, C. pipiens f. molestus, which had recently colonized new urban sites, differed in the structures with a low functional trophic load (some structures of siphon and abdominal segment IX), whereas structures of the trophic morphological complex were similar in different populations. The possible mechanisms of these morphological transformations and their role in the evolution of mosquito larvae are discussed.     

Phylogenetic analysis of <Emphasis Type="Italic">Plectosphaerella</Emphasis> species based on multi-locus DNA sequences and description of <Emphasis Type="Italic">P. sinensis</Emphasis> sp. nov.

Species in Plectosphaerella are well known as pathogens of several plant species causing fruit, root and collar rot and collapse. In an investigation of endophytic fungi associated with cucurbit plants in China, we isolated 77 strains belonging to the genus Plectosphaerella. To identify the isolated strains, we collected the type or reference strains of all currently accepted species in Plectosphaerella except P. oratosquillae and conducted a phylogenetic analysis. Phylogenetic analysis of the partial 28S rDNA sequences showed that all species in Plectosphaerella were located in one clade of Plectosphaerellaceae. Based on multi-locus phylogenetic analysis of the ITS, CaM, EF1, TUB and morphological characteristics, all species in Plectosphaerella were well separated. Three endophytic strains from stems of Cucurbita moschata, Citrullus lanatus and Cucumis melo from North China were assigned to a new species described as P. sinensis in this paper. The new species differs morphologically from other Plectosphaerella species by irregular chlamydospores, and the dimensions of phialides and conidia. The other endophytic strains from several cucurbit plants were identified as P. cucumerina.     

Taxonomic re-evaluation of the genus <Emphasis Type="Italic">Lambertella</Emphasis> (Rutstroemiaceae,Helotiales) and allied stroma-forming fungi

The genus Lambertella is currently considered to be one of the largest genera in the family Rutstroemiaceae, and its major distinguishing character is the pigmentation of ascospores, besides a substratal stroma. Although Lambertella appears to be well-defined by morphological characters, its phylogenetic heterogeneity has been suggested in earlier studies. To circumscribe the genus more precisely, morphological examination and phylogenetic analysis was conducted on some Lambertella species and some allied stroma-forming fungi. In total, 58 taxa were included in the study, including 16 species with both a substratal stroma and pigmented ascospores. The polyphyly of Lambertella sensu lato was confirmed based on the phylogenetic analysis of ITS, LSU, and RPB2 regions. A highly supported clade is composed of five Lambertella species, including the type L. corni-maris. Morphologically, all species here included in Lambertella s. s. have brown ascospores prior to being discharged from asci, whereas in other species currently placed in Lambertella, but now excluded from the genus, they turn brown only after discharge. This clade was defined as Lambertella sensu stricto. Pigmentation of the ascospores was also recognized in Lanzia and Poculum, and even in the helotiaceous genus Hymenoscyphus, but always only after discharge. The convergent evolution of a dark substratal stroma in Rutstroemiaceae and Helotiaceae was also indicated.