High concentration of basidiolichens in a single family of agaricoid mushrooms (Basidiomycota: Agaricales: Hygrophoraceae)

The Agaricales is the largest and most diverse order of mushroom-forming Basidiomycota, with over 100 natural groups recognized in recent Fungal Tree of Life studies. Most agarics are either saprotrophic or ectomycorrhizal fungi, but the family Hygrophoraceae is in part characterized by a unique and remarkable diversity of lichenized forms. The most familiar of these is the chlorolichen genus Lichenomphalia, whose phylogenetic position in the Agaricales has been established. Recent limited evidence suggested that Hygrophoraceae also contains cyanolichens in the genus Dictyonema, which indicates a remarkable concentration and diversity of lichen-formers in a single family of agarics. To demonstrate the relationships of lichen-formers to other fungi in the family, we assembled ribosomal sequences from 52 species representing recognized groups within the Hygrophoraceae, among them new sequences representing Acantholichen and most species and forms of Dictyonema. The molecular data were evaluated using parsimony, likelihood, Bayesian, and distance analyses, including coding of ambiguous regions by means of INAASE and ARC, all of which indicate that Dictyonema and Acantholichen form a monophyletic clade derived from the primarily bryophilous genus Arrhenia and sister to the enigmatic Athelia pyriformis, a species unrelated to the Atheliales for which we are proposing a new genus name Eonema. The chlorolichen genus Lichenomphalia may be polyphyletic. Fungi in the Dictyonema-Acantholichen clade are typically tropical, entirely lichenized, and associate with cyanobacterial photobionts. Our data indicate a transition from agaricoid-omphalinoid basidiomes observed in Arrhenia to stereoid-corticioid forms in Dictyonema, and also support a previous suggestion of a connection between loss of clamp connections and lichenization. The diverse basidiome and thallus morphologies and nutritional ecologies of these fungi indicate a remarkable evolutionary flexibility that appears to have developed in part as a consequence of symbiosis.     

Distance of the reduced Brusselator from equilibrium

The new concept of a nonequilibrium parameter P is applied to a reduced Brusselator, considered as a kinetic model for cellular processes. The reduction corresponds to omitting a monomolecular reaction. The deviation from equilibrium is due to a fixed nonequilibrium value of an extracellular concentration B, responsible for energizing and determining the steady-state value of the overall chemical affinity ?. The value of ? is insensitive to different steady states possible for a given set of rate constants. In contrast, the parameter P is state dependent. In particular, it may jump together with state variables. Two limiting cases of high ? are investigated, B→ 0 and B→ 1. In the first case P grows monotonically with ?. In the second case there is always a steady state solution with P→ 0. The physical interpretation of this effect of “equilibrium far from equilibrium” reveals the real predictive power of the parameter P. Relaxation regimes are investigated for a doubly reduced Brusselator. Both P and ? are in general time dependent and have jumps in their time derivatives. The canonical form of P is compared with the noncanonical one in the context of robustness of the new concept with respect to incomplete information about the system studied. These forms of P are different in relaxation to a nonequilibrium state and coincide in relaxation to an equilibrium state.     

Two new species of Edmockfordia García Aldrete (Psocodea, ‘Psocoptera’, Epipsocidae), from Valle del Cauca,Colombia, and description of the female E. chiquibulensis García Aldrete

Two new species of Edmockfordia García Aldrete, from Valle del Cauca, Colombia, and the female of Edmockfordia chiquibulensis García Aldrete, are described and illustrated. A key to the species of Edmockfordia is included; the genus was previously known only from Belize. The genus is re-diagnosed to include female characters. The distribution of the genus is considerably widened, from Belize to northeastern South America.     

On the validity of Epeorella Ulmer, 1939 (Ephemeroptera,Heptageniidae) with general considerations on the Heptageniidae of the Sunda Islands

The type material of Epeorella borneonia Ulmer, 1939, the sole species of the genus Epeorella Ulmer, 1939 is reinvestigated and a lectotype (male imago) is designated. Based on several morphological structures, the synonymy with Epeorus Eaton, 1881 (Rhithrogeninae) is rejected. Epeorella stat. prop., known only at the winged stages, belongs to the subfamily Ecdyonurinae, and is a probable endemic of the island of Borneo. The newly erected genus Darthus Webb & McCafferty, 2007, also endemic to Borneo and only known by one species at the nymphal stage, is shown to be a junior subjective synonym of Epeorella. The new combination Epeorella vadora (Webb & McCafferty, 2007) is proposed for the species. The distribution of known heptageniid species from the Sunda Islands is discussed.     

The Inner Ear of Two Late Cretaceous Multituberculate Mammals, and Its Implications for Multituberculate Hearing

The inner ear of the Late Cretaceous multituberculates Nemegtbaatar gobiensis and Chulsan-baatar vulgaris is described from serial sections and enlarged models. The size and proportions of the inner ear as a whole are as expected for extant small mammals. The lengths of the cochlea (Nemegtbaatar gobiensis, 3.0 mm, Chulsanbaatar vulgaris, 2.0 mm) are comparable to those of other multituberculates, when ratios of length of the cochlea to skull length are calculated. The vestibule is not as expanded in the two taxa as in Lambdopsalis, ?Meniscoessus, and ?Catopsalis; the estimated volume for Nemegtbaatar gobiensis is 9 mm³. A slightly laterally curved, anteriomedially directed cochlea, relatively robust ear ossicles, and the estimations of the area of the tympanic membrane and stapedial footplate in Chulsanbaatar suggest high-frequency hearing but a relatively low sensitivity to low-decibel sounds. The semicircular canals of Nemegtbaatar and Chulsanbaatar are fully developed; the size of the anterior, posterior, and lateral canals and their angles and proportions are comparable to those of extant mammals of similar size. The anterior semicircular canal of Nemegtbaatar forms a smooth half-circle and thus is more derived than the angular canal of Ornithorhynchus. The notable differences between the ratio of the width of the lateral semicircular canal to skull length and the size of the vestibule in Nemegtbaatar and the Paleocene multituberculate Lambdopsalis bulla are probably related to different modes of life.     

Genomic structure and sequence of the pufferfish (Fugu rubripes) growth hormone-encoding gene: a comparative analysis of teleost growth hormone genes

A nested polymerase chain reaction (PCR) technique for amplifying a fragment of the gene (GH) encoding teleost growth hormone has been developed. Using this technique, a fragment of the pufferfish, Fugu rubripes and Arothron maculatus; dwarf gourami, Colisa lalia; guppy, Poecilia reticulata; and goldfish, Carassius auratus GH genes were cloned. The Fugu rubripes (Fugu) gene fragment was used to isolate the GH gene from a Fugu genomic library. The complete nucleotide sequence of a 8.5-kb SacI genomic fragment containing the Fugu GH gene has been determined. The GH gene spans 2.5 kb from the first codon to polyadenylation signal, and contains six exons and five introns similar to the GH genes of salmonids, tilapia, barramundi, flounder and yellowtail. The GH introns contain microsatellite and satellite sequences. The microsatellites found in the fifth intron of the GH gene are also present in the corresponding introns of tilapia, barramundi and flounder GH genes. Southern analysis revealed that the GH gene is a single-copy gene in the Fugu. The promoter region of the Fugu GH gene contains conserved sequences that are likely to be involved in the pituitary-specific expression of the gene. A phylogenetic tree of nucleotide (nt) sequences of all known teleost GH genes has been inferred using the distance matrix method. The topology of this tree reflects the major phylogenetic groupings of teleosts. The intron patterns and repetitive sequences of GH genes can serve as useful natural markers for the classification and phylogenetic studies of teleosts.     

Phylogenetic reassessment of Hyaloscyphaceae sensu lato (Helotiales, Leotiomycetes) based on multigene analyses

Hyaloscyphaceae is the largest family in Helotiales, Leotiomycetes. It is mainly characterized by minute apothecia with well-differentiated hairs, but its taxonomic delimitation and infrafamilial classification remain ambiguous. This study performed molecular phylogenetic analyses using multiple genes including the ITS-5.8S rDNA, the D1–D2 region of large subunit of rDNA, RNA polymerase II subunit 2, and the mitochondrial small subunit. The primary objective was to evaluate the phylogenetic utility of morphological characters traditionally used in the taxonomy of Hyaloscyphaceae through reassessment of the monophyly of this family and its genera. The phylogenetic analyses inferred Hyaloscyphaceae as being a heterogeneous assemblage of a diverse group of fungi and not supported as monophyletic. Among the three tribes of Hyaloscyphaceae only Lachneae formed a monophyletic lineage. The presence of hairs is rejected as a synapomorphy, since morphologically diversified hairs have originated independently during the evolution of Helotiales. The true- and false-subiculum in Arachnopezizeae are hypothesized to have evolved through different evolutionary processes; the true-subiculum is likely the product of a single evolutionary origin, while the false-subiculum is hypothesized to have originated multiple times. Since Hyaloscyphaceae sensu lato was not resolved as monophyletic, Hyaloscyphaceae sensu stricto is redefined and only applied to the genus Hyaloscypha.     

Diversification of the Genus Anopheles and a Neotropical Clade from the Late Cretaceous

The Anopheles genus is a member of the Culicidae family and consists of approximately 460 recognized species. The genus is composed of 7 subgenera with diverse geographical distributions. Despite its huge medical importance, a consensus has not been reached on the phylogenetic relationships among Anopheles subgenera. We assembled a comprehensive dataset comprising the COI, COII and 5.8S rRNA genes and used maximum likelihood and Bayesian inference to estimate the phylogeny and divergence times of six out of the seven Anopheles subgenera. Our analysis reveals a monophyletic group composed of the three exclusively Neotropical subgenera, Stethomyia, Kerteszia and Nyssorhynchus, which began to diversify in the Late Cretaceous, at approximately 90 Ma. The inferred age of the last common ancestor of the Anopheles genus was ca. 110 Ma. The monophyly of all Anopheles subgenera was supported, although we failed to recover a significant level of statistical support for the monophyly of the Anopheles genus. The ages of the last common ancestors of the Neotropical clade and the Anopheles and Cellia subgenera were inferred to be at the Late Cretaceous (ca. 90 Ma). Our analysis failed to statistically support the monophyly of the Anopheles genus because of an unresolved polytomy between Bironella and A. squamifemur.     

Ein neues clerodan-derivat sowie weitere inhaltsstoffe aus der gattung Macowania ag

The roots of M. corymbosa afford three new benzofuran derivatives very closely related to leysseralangelicate isolated from the related genus Leyssera. The aerial parts of M. glandulosa contain, besides labdanolic acid, a new cis-clerodane derivative, its structure being elucidated by spectroscopic methods and by chemical reactions. The roots of a further Macowania species closely related to M. hamata afford a new ester related to m-thymol.     

Brazilian species of Calliandra Benth. (tribe Ingeae) are nodulated by diverse strains of Paraburkholderia

The Chapada Diamantina in NE of Brazil is a biodiversity hotspot and a center of radiation for many Neotropical legume genera, such as Calliandra and Mimosa. The present study aimed to evaluate nodulation in Calliandra species endemic to various environments, and to characterize the diversity of their symbiotic rhizobia using housekeeping (16S rRNA, recA) and plasmid-borne, symbiosis-related (nifH and nodC) genes. The nodulation ability of selected isolates was assessed. All of the 126 bacterial isolates from 18 Calliandra species collected in six different vegetation types were identified as Paraburkholderia according to their housekeeping and symbiosis gene phylogenies. They were grouped in seven clades in relation to the dominant vegetation type in their native environments. The majority, particularly those from highland “campo rupestre” vegetation, were similar to Paraburkholderia nodosa, but had nodC genes identical to the Mimosa symbiont Paraburkholderia tuberum sv. mimosae. The other smaller groups were related to Paraburkholderia diazotrophica and Paraburkholderia sabiae, and some single strains were not close to any known species. The symbionts of Calliandra spp. in NE Brazil are Paraburkholderia strains closely-related to Mimosa symbionts from the same region. NE Brazil is a reservoir of symbiotic Paraburkholderia that have an affinity for genera in the Mimosoid clade.     

Phylogeny of the ectomycorrhizal mushroom genus Alnicola (Basidiomycota, Cortinariaceae) based on rDNA sequences with special emphasis on host specificity and morphological characters

Alnicola (=Naucoria, pro parte) is a mushroom genus of strictly temperate, obligately ectomycorrhizal species, traditionally included in the family Cortinariaceae. Most Alnicola spp. are primarily host specific on Alnus, although a few are mycobionts of Salix or other hosts. The different species of Alnicola exhibit unique morphological (cystidia, pileipellis) and cytological (dikaryotic or monokaryotic hyphae) characters. This makes the genus Alnicola of particular interest for studying the evolution of host specificity and morphological characters in ectomycorrhizal basidiomycetes. We used a combination of classical morphological and phylogenetic methods (rDNA ITS and LSU sequences) to address the following questions: (i) Is Alnicola monophyletic? And (ii) Are characters like host specificity or microscopical structures synapomorphic for certain clades? The study included nearly all currently known European Alnicola sp. Our results demonstrated that, on one hand, the genus Alnicola is polyphyletic, with sistergroup relationships to Hebeloma, Anamika or the clades /Hymenogaster I and /Hymenogaster II. On the other hand, Alnicola splits into three well-supported clades corresponding to the sections Alnicola, Submelinoides, and Salicicolae. The strict host-specificity to Alnus is a derived character and has occurred at least twice. The following morphological characters are synapomorphic for defined clades: the spindle-shaped hymenial cystidia for sect. Alnicola, the hymeniform pileipellis for sect. Submelinoides, and monocaryotic/clampless hyphae for sect. Salicicolae and its sistergroup /Hymenogaster II. As a taxonomical consequence, polyphyly of Alnicola implies that the sects. Submelinoides and Salicicolae need to be segregated from Alnicola.     

Foliar cuticular alkanes of Camarea (Malpighiaceae) and their taxonomic significance

Camarea is a South-American endemic genus comprising eight species. In the present work n-alkanes from foliar cuticular waxes of 23 specimens, representing seven species of Camarea were analyzed, aiming at establishing interspecific affinities and evaluating the usefulness of n-alkane distribution as species characteristic. The sampling included also specimens of Peixotoa reticulata and Janusia guaranitica (both Malpighiaceae). The results were used to obtain a phenogram indicating chemical affinities between species. The results are in agreement with morphological similarities among some Camarea species. Intraspecific variability was small, suggesting that n-alkane distribution may be useful for species characterization and establishment of links among Camarea species. The results support the recognition of Camarea triphylla as a synonym of Camarea axillaris and are not coherent with a hybrid condition of a population exhibiting morphological characteristics combining Camarea affinis and Camarea hirsuta, suggesting instead that the individuals analyzed belong either to Camarea hirsuta or a close species. Distribution of n-alkanes is inadequate to distinguish among Malpighiaceae genera: P. reticulata has n-alkane distribution similar to several Camarea species.     

Organization of the nar genes at the chlZ locus

The two membrane-bound respiratory nitrate reductases of Escherichia coli are encoded by distinct operons at two different loci, chlC and chlZ, on the chromosome. The chlZ locus includes a narK homologue, narU, encoding a nitrite extrusion protein, and narZYWV encoding nitrate reductase Z. No apparent homologue to the narXL operon has been found. Homology between narU and narK on the one hand and narZYWV and narGHJI on the other hand is limited to the coding regions.     

Characterization of a Theta-Type Plasmid from Lactobacillus sakei: a Potential Basis for Low-Copy-Number Vectors in Lactobacilli

The complete nucleotide sequence of the 13-kb plasmid pRV500, isolated from Lactobacillus sakei RV332, was determined. Sequence analysis enabled the identification of genes coding for a putative type I restriction-modification system, two genes coding for putative recombinases of the integrase family, and a region likely involved in replication. The structural features of this region, comprising a putative ori segment containing 11- and 22-bp repeats and a repA gene coding for a putative initiator protein, indicated that pRV500 belongs to the pUCL287 subfamily of theta-type replicons. A 3.7-kb fragment encompassing this region was fused to an Escherichia coli replicon to produce the shuttle vector pRV566 and was observed to be functional in L. sakei for plasmid replication. The L. sakei replicon alone could not support replication in E. coli. Plasmid pRV500 and its derivative pRV566 were determined to be at very low copy numbers in L. sakei. pRV566 was maintained at a reasonable rate over 20 generations in several lactobacilli, such as Lactobacillus curvatus, Lactobacillus casei, and Lactobacillus plantarum, in addition to L. sakei, making it an interesting basis for developing vectors. Sequence relationships with other plasmids are described and discussed.     

Cloning and Expression of a Chitinase Gene from Sanguibacter sp. C4

The chitinase Chi58 is an extracellular chitinase produced by Sanguibacter sp.strain C4. The gene-specific PCR primers were used to detect the presence of the chiA gene in strain C4. A chiA fragment (chiA-F) was amplified from the C4 genomic DNA and was used to blast-search the related sequences from the GenBank dadabase. By alignment and selection of the highly conserved regions of the homologous sequences, two pairs of primers were designed to amplify the open reading frame (ORF) of the chitinase from strain C4 by nested PCR. The results revealed that the Chi58 ORF consisted of 1 692 nucleotides encoding a protein of 563 amino acid residues. The molecular weight of the mature protein was predicted to be 58.544 kDa. The Chi58 ORF was a modular enzyme composed of a signal peptide sequence, a polycystic kidney disease I domain, and a glycosyl hydrolase family 18 domain. The chitinase of C4 exhibited a high level of similarity to the chitinase A of Serratia (88.9%-99.6%) at the amino acid sequence level. The Chi58 gene was cloned into the expression vector pET32a to construct the recombinant plasmid pChi58 and was expressed in E. coli BL-21 (DE3) cells with IPTG induction. The molecular weight of the Trx-Chi58 fusion protein was estimated to be 81.1 kDa by SDS-PAGE.     

Core and symbiotic genes reveal nine Mesorhizobium genospecies and three symbiotic lineages among the rhizobia nodulating Cicer canariense in its natural habitat (La Palma,Canary Islands)

Cicer canariense is a threatened perennial wild chickpea endemic to the Canary Islands. In this study, rhizobia that nodulate this species in its natural habitats on La Palma (Canary Islands) were characterised. The genetic diversity and phylogeny were estimated by RAPD profiles, 16S-RFLP analysis and sequencing of the rrs, recA, glnII and nodC genes. 16S-RFLP grouped the isolates within the Mesorhizobium genus and distinguished nine different ribotypes. Four branches included minority ribotypes (3–5 isolates), whereas another five contained the predominant ribotypes that clustered with reference strains of M. tianshanense/M. gobiense/M. metallidurans, M. caraganae, M. opportunistum, M. ciceri and M. tamadayense. The sequences confirmed the RFLP groupings but resolved additional internal divergence within the M. caraganae group and outlined several potential novel species. The RAPD profiles showed a high diversity at the infraspecific level, except in the M. ciceri group. The nodC phylogeny resolved three symbiotic lineages. A small group of isolates had sequences identical to those of symbiovar ciceri and were only detected in M. ciceri isolates. Another group of sequences represented a novel symbiotic lineage that was associated with two particular chromosomal backgrounds. However, nodC sequences closely related to symbiovar loti predominated in most isolates, and they were detected in several chromosomal backgrounds corresponding to up to nine Mesorhizobium lineages. The results indicated that C. canariense is a promiscuous legume that can be nodulated by several rhizobial species and symbiotypes, which means it will be important to determine the combination of core and symbiotic genes that produce the most effective symbiosis.