Biogeography of Aegagropila linnaei (Cladophorophyceae,Chlorophyta): a widespread freshwater alga with low effective dispersal potential shows a glacial imprint in its distribution

Aim Aegagropila linnaei is a freshwater macroalga that is generally regarded as a rare species. It is apparently absent from large but seemingly suitable areas of the Northern Hemisphere, implying a limited dispersal potential and an imprint of Pleistocene glaciations in its biogeography. However, despite the popularity of its enigmatic lake ball‐form, detailed biogeographical studies of A. linnaei have never been conducted. The main means of reproduction of A. linnaei is fragmentation and akinetes are not formed, supporting the assumption of limited dispersal capacity. The aim of this study was to reconstruct the biogeography of A. linnaei, and to identify possible refugia during glaciations, as well as to evaluate dispersal potential by quantitative desiccation experiments. Location Palaearctic. Methods The current distribution of A. linnaei was inferred from herbarium specimens, literature data and recent field observations. All herbarium specimens were morphologically re‐examined. Desiccation experiments were performed with vegetative filaments of three isolates of A. linnaei, as no specialized resistant stages are known. For comparison, the widespread freshwater algae Cladophora glomerata and Rhizoclonium sp. were included. Internal transcribed spacer (ITS) ribosomal DNA sequences were generated and a ribotype network was constructed. Results Aegagropila linnaei was recorded from 283 locations in freshwater and brackish environments. The majority of locations were in central and northern Europe in previously glaciated areas. Desiccation experiments showed that A. linnaei is very susceptible to desiccation. Based on ITS sequences of 34 samples, five different ribotypes were identified. Four of these ribotypes had a restricted distribution. Aegagropila linnaei represents a single species with little genetic variation (0.1–0.5%). Main conclusions This is the most comprehensive study of this species so far, reporting many new locations and tackling several taxonomic problems. Few additional finds were made from North America, and the origin of A. linnaei is inferred to be in Asia. The highest density of its present‐day locations is in previously glaciated areas in Europe, where glacial ice‐dammed lakes might have functioned as refugia. Low effective long‐distance dispersal capacity is inferred, based on high susceptibility to desiccation and its modes of dispersal.     

Microbial toxins in plant-pathogen interactions: Biosynthesis,resistance mechanisms,and significance

In the history of phytopathology, microbial toxins have been the objects of extensive studies as possible pathogenicity or virulence factors for the producer pathogens. The recent development of molecular genetic techniques provided an experimental basis to thoroughly test the role of these secondary metabolites in pathogenesis. Some of them did prove to be highly associated with disease initiation or enhanced virulence in certain plant-pathogen interactions. In this review, we describe recent progresses in the field of plant-pathogen interactions focusing on two toxins; i.e., tabtoxin from Pseudomonas syringae and trichothecenes from Fusarium and other fungi. These microbial toxins have convincingly been shown to play causal roles in plant disease development. Studies on the biosynthesis and resistance mechanisms of these producers are outlined, and the significance of this knowledge is discussed in relation to practical applications in agriculture.     

Crystal structures of wild-type and mutant plastocyanins from a higher plant, Silene

Plastocyanin functions as an electron carrier between the cytochrome b6f complex and photosystem I. The crystal structures of the wild-type and E43K/D44K double mutant from the higher plant, Silene, have been determined at 2.0 and 1.75 A resolution, respectively. The wild-type plastocyanin comprises two monomers per asymmetric unit, one of which shows the unusually great distance between the copper ion and the Ndelta1 atom of H87 because of the hydrogen bond network formation between H87 and symmetry-related G10. The root mean square deviation for Ca atoms between the wild-type and mutant plastocyanins is 0.44 A, however, the electrostatic potential maps of their molecular surfaces are remarkably different. The low electron-transfer rate in the E43K/D44K mutant results from the hindrance of electrostatic interactions, not from the structural change due to the mutation.     

Antifouling activity of simple synthetic isocyanides against larvae of the barnacle Balanus amphitrite

Twelve simple linear isocyanides were synthesized and examined for antifouling activity and toxicity against cyprid larvae of the barnacle, Balanus amphitrite. Larval settlement was inhibited, with EC50 values of 0.046-1.90 microg ml(-1), and they were much less toxic (LD50 values ranging over 21.28 microg ml(-1)) than CuSO4 (EC50 0.30 microg ml(-1) and LD50 2.95 microg ml(-1)). The data indicate that simple linear isocyanides are promising non-toxic antifouling agents.     

Enzymatic characterization of scytalone dehydratase Val75Met variant found in melanin biosynthesis dehydratase inhibitor (MBI-D) resistant strains of the rice blast fungus

Carpropamid ((1RS,3SR)-2,2-dichloro-N-[(R)-1-(4-chlorophenyl)ethyl]-1-ethyl-3-methylcyclopropanecarboxamide) is a potent chemical against the rice blast fungus, Pyricularia oryzae. In 2001, isolates of the fungus with reduced sensitivity to this fungicide appeared in Saga Prefecture of Japan and were regarded as a potential threat to rice protection by carpropamid. The cause of the resistance has been identified genetically as a point mutation resulting in the Val75Met change in scytalone dehydratase, the primary target of the fungicide. We constructed an overexpression system of the variant enzyme and characterized the kinetics in the catalysis and the inhibition by carpropamid isomers. The variant enzyme retained a significant level of enzymatic activity. Inhibition of the variant enzyme by carpropamid was more than 200-fold reduced in comparison with that of the wild-type. Based on the results, here we propose possible mechanisms of the carpropamid-resistance of the variant enzyme in retaining the normal enzymatic activity.     

A GFP-transfected HFWT cell line,GHINK-1, as a novel target for non-RI activated natural killer cytotoxicity assay

An anchorage-dependent Wilms' tumor cell line, HFWT, has been found to be extremely susceptible to human natural killer (NK) cells. Here we established a transfectant of HFWT with the green fluorescence protein (GFP) gene, designated GHINK-1 cells, to apply to the activated NK cytotoxicity assay without radioisotope labeling. After being co-cultured with CD3 CD56+ NK cells, GHINK-1 cells released GFP into the medium. The intensity of the fluorescence from the released GFP correlated almost exactly with the radioactivity of a standard 51Cr-release assay performed with suspension-cultured K562 cells. Because it does not require separation of the remaining live target cells by centrifugation, the non-radioisotopic GFP release assay with GHINK-1 cells is a convenient alternative for monitoring human activated NK killing activity.     

Biological characteristics of cultured cells derived from various types of human brain tumors

We placed in culture brain tumors from 45 cases (7 cases of astrocytoma, 2 from oligodendrogliomas, 2 glioblastomas, 2 ependymomas, 13 meningiomas, 6 pituitary adenomas, 5 neurinomas, a malignant lymphoma, a choroid plexus papilloma, and 6 metastatic tumors) and succeeded in making a primary culture from 33, and maintained 17 in vitro over a considerable period of time (greater than three months). In the early period of the primary cultures, the astrocytoma cells had cytoplasmic processes which contacted each other, the oligodendroglioma cells were small and spindle-shaped, the glioblastoma cells were neoplastic with pleopmorphic features and possessed cytoplasmic processes, the ependymoma cells formed a rosette-like cell arrangement, the meningioma cells were spindle- or round-shaped cells and characterized as forming psammoma bodies, the pituitary adenoma cells were round- or oval-shaped cells and produced growth hormone (GH), adenocorticoid tropic hormone (ACTH), prolactin, or other hypophyseal hormones, the choroid plexus papilloma cells were round-or polygonal and showed a papillary cell arrangement, the neurinoma cells were spindle- or fibrous-shaped cells, and the malignant lymphoma cells were round and formed cell aggregates floating in the culture medium.