Differences in resource storage pattern between Laminaria longissima and Laminaria diabolica (Laminariaceae; Phaeophyta) reflecting their morphological characteristics

The kelps Laminaria longissima and L. diabolica, belonging to the groups of L. angustata and L. japonica, respectively, differ greatly in their morphological characteristics although their geographical distributions overlap widely along the eastern coast of Hokkaido. To clarify the interaction between the morphological and physiological characteristics of the two species, and their link with environmental variables, hatchery-raised young sporophytes of L. longissima and L. diabolica collected from Hokkaido were cultivated simultaneously under similar conditions in Matsushima Bay, Miyagi, from January to July 2004. Seasonal morphological characteristics, gross photosynthetic rate, nutrient uptake rates, and resource contents were examined. The blade lengths of L. longissima and L. diabolica reached a maximum of 329.9 cm and 256.7 cm, respectively, in April to May, and decreased to 284.4 cm and 68.6 cm, respectively, in July. The total elongation length of L. longissima (412.5 cm) was similar to that of L. diabolica (373.8 cm). However, the total erosion length of L. longissima (145.9 cm) was approximately half that of L. diabolica (302.9 cm). The gross photosynthetic rate and uptake rates of NH₄-N, NO₃-N, and PO₄-P of the two species were similar. However, the carbon, nitrogen, and phosphorus contents were transferred and stored in the whole blade tissues in the case of L. longissima, but in the meristem of L. diabolica from May to June. These results suggest that morphological differences are a response to different resource storage patterns. The storage patterns of L. longissima and L. diabolica are likely to be genetically fixed characteristics, which have evolved in adaptation to the specific habitat environments of the groups of L. angustata and L. japonica. The low water temperature and rich nutrients provided by the Oyashio Current are conducive to storage of resources in the whole blade tissues and a large surface area retained for photosynthesis and nutrient uptake in the L. angustata group. Conversely, high temperature and poor nutrients, or large fluctuations in these parameters, provided by the Tsushima Warm Current are more conducive to intensive storage of resources in the meristem for maturation and further growth in the L. japonica group. L. diabolica retains the storage pattern of the L. japonica group but grows in regions affected by the Oyashio Current, allowing it to become the widest distributed Laminaria species.     

Study on Cortinarius subgenus Telamonia section Hydrocybe in Europe, with especial emphasis on Mediterranean taxa

In this paper we have attempted to clarify the taxonomy and nomenclature of thirteen taxa of the genus Cortinarius subgenus Telamonia (sections Hydrocybe, Fraternii) well represented in the southwestern Mediterranean area of Europe (C. atrocoeruleus, C. bombycinus, C casimiri, C. contrarius, C. decipiens, C. fraternus, C. gallurae, C. hoffmannii, C. petroselineus, C. sertipes, C. subturibulosus, C. urdaibaiensis and C. vernus). To this end we have performed a combined study of morphological and molecular data (rDNA ITS sequences). The morphological analysis was carried out on 114 collections and the molecular analysis involved 31 of the 114 collections, including 11 type collections (types for C. casimiri and C. fraternus were not available). In addition, a study of spores under field emission scanning electron microscopy (FESEM) was conducted. The results of the combined analysis allowed us to asign the studied material to five species (C. casimiri s.l., C. decipiens s.l., C. gallurae, C. subturibulosus s.l. and C. vernus s.l.). Thus, all collections from more continental areas, which were originally identified as six different taxa (C. atrocoeruleus, C. contrarius, C. decipiens, C. fraternus, C. sertipes, C. flexipes fo. sertipes) corresponded to C. decipiens sensu lato, a widely distributed, genetically and morphologically variable species. Cortinarius casimiri is also found in such habitats, but it is confirmed as distinct taxon. Collections from Mediterranean sclerophyllous communities correspond to C. gallurae, C. vernus sensu lato and C. subturibulosus sensu lato. Due to close phylogenetic relationships we propose the new combinations C. casimiri var. hoffmannii (=C. decipiens var. hoffmannii non C. hoffmannii) and C. subturibulosus var. bombycinus (=C. bombycinus), and the new variety C. vernus var. nevadavernus (=C. vernus H. Lindstr. & Melot sensu auct.).     

The development and in vitro characterisation of an intracellular nanosensor responsive to reactive oxygen species

Advances in sensor technologies have enhanced our understanding of the roles played by reactive oxygen species (ROS) in a number of physiological and pathological processes. However, high inter-reactivity and short life spans has made real-time monitoring of ROS in cellular systems challenging. Fluorescent dyes capable of intracellular ROS measurements have been reported. However, these dyes are known to be intrinsically cytotoxic and thus can potentially significantly alter cellular metabolism and adversely influence in vitro data. Reported here is the development and in vitro application of a novel ROS responsive nanosensor, based on PEBBLE (Probes Encapsulated By Biologically Localised Embedding) technology. The ROS sensitive fluorescent probe dihydrorhodamine 123 (DHR 123) was employed as the sensing element of the PEBBLE through entrapment within a porous, bio-inert polyacrylamide nanostructure enabling passive monitoring of free radical flux within the intracellular environment. Successful delivery of the nanosensors into NR8383 rat alveolar macrophage cells via phagocytosis was achieved. Stimulation of PEBBLE loaded NR8383 cells with phorbol-12-myristate-13-acetate (PMA) enabled real time monitoring of ROS generation within the cell without affecting cellular viability. These data suggest that PEBBLE nanosensors could offer significant advantages over existing technologies used in monitoring the intracellular environment.     

Potential errors in the quantitative evaluation of biogas production in anaerobic digestion processes

Errors that are commonly made in the quantification of biogas from anaerobic digestion experiments were investigated.For liquid displacement gasometers where a barrier solution separates the biogas and the atmosphere, inaccuracy due to gas diffusion was examined experimentally. Acidified saturated saline solution was the most suitable barrier solution, as biogas characteristics changed least with time. Using acidified or tap water caused considerable biogas losses and should therefore be avoided where biogas is stored before measurement.Errors associated with volume calculation from three common liquid displacement gasometer types were investigated theoretically. Corrections that must be made to obtain gas volumes at standard temperature and pressure when using this equipment are discussed. Regarding experimental errors, gasometer designs where displaced liquid is weighed to determine the volume are the most versatile since errors depend mainly upon balance sensitivity. Using liquid heights to calculate volume requires appropriate sizing of the gasometer relative to the volume of gas measured.The calibration of a low flow gas meter was investigated and an approximately linear variation with flow rate was found; hence in situ calibration is advised for this type of instrument. Correction for atmospheric conditions should be performed in real time to reduce errors.     

Circular permutation of ligand‐binding module improves dynamic range of genetically encoded FRET‐based nanosensor

Quantitative measurement of small molecules with high spatiotemporal resolution provides a solid basis for correct understanding and accurate modeling of metabolic regulation. A promising approach toward this goal is the FLIP (fluorescent indicator protein) nanosensor based on bacterial periplasmic binding proteins (PBPs) and fluorescence resonance energy transfer (FRET) between the yellow and cyan variants of green fluorescent protein (GFP). Each FLIP has a PBP module that specifically binds its ligand to induce a conformation change, leading to a change in FRET between the two GFP variant modules attached to the N‐ and C‐termini of the PBP. The larger is the dynamic range the more reliable is the measurement. Thus, we attempted to expand the dynamic range of FLIP by introducing a circular permutation with a hinge loop deletion to the PBP module. All the six circularly permutated PBPs tested, including structurally distinct Type I and Type II PBPs, showed larger dynamic ranges than their respective native forms when used for FLIP. Notably, the circular permutation made three PBPs, which totally failed to show FRET change when used as their native forms, fully capable of functioning as a ligand binding module of FLIP. These FLIPs were successfully used for the determination of amino acid concentration in complex solutions as well as real‐time measurement of amino acid influx in living yeast cells. Thus, the circular permutation strategy would not only improve the performance of each nanosensor but also expand the repertoire of metabolites that can be measured by the FLIP nanosensor technology.     

Building Developmental Integration into Functional Systems: Function-Induced Integration of Mandibular Shape

The mammalian mandible is a developmentally modular but functionally integrated system. Whether morphological integration can evolve to match the optimal pattern of functional integration may depend on the developmental origin of integration, specifically, on the role that direct epigenetic interactions play in shaping integration. These interactions are hypothesized to integrate modules and also to be highly conservative, potentially constraining the evolution of integration. Using the fox squirrel (Sciurus niger) mandible as a model system, we test five a priori developmental hypotheses that predict mandibular integration and we also explore for correlations between shapes of mandibular regions not anticipated by any of the developmental models. To determine whether direct epigenetic interactions are highly conserved in rodents, we examine the correlation structure of fluctuating asymmetry, and compare integration patterns between fox squirrels and prairie deer mice (Peromyscus maniculatus bairdii). In fox squirrels, we find a correlation structure unanticipated by all a priori developmental models: adjacent parts along the proximodistal jaw axis are correlated whereas more distant ones are not. The most notable exception is that the shape of the anterior incisor alveolus is correlated with the shape of the ramus (FA component) or coronoid (symmetric component). Those exceptions differ between species; in prairie deer mice, the molar alveolus is connected to more parts, and the incisor alveolus to fewer, than in fox squirrels. The structure of integration suggests that the mandible cannot be decomposed into parts but rather is a single connected unit, a result consistent with its functional integration. That match between functional and developmental integration may arise, at least in part, from function-induced growth, building developmental integration into the functional system and enabling direct epigenetic interactions to evolve when function does.     

Single-fluorophore monitoring of DNA hybridization for investigating the effect of secondary structure on the nucleation step

Nucleic acid hybridization is one of the essential biological processes involved in storage and transmission of genetic information. Here we quantitatively determined the effect of secondary structure on the hybridization activation energy using structurally defined oligonucleotides. It turned out that activation energy is linearly proportional to the length of a single-stranded region flanking a nucleation site, generating a 0.18 kcal/mol energy barrier per nucleotide. Based on this result, we propose that the presence of single-stranded segments available for non-productive base pairing with a nucleation counterpart extends the searching process for nucleation sites to find a perfect match. This result may provide insights into rational selection of a target mRNA site for siRNA and antisense gene silencing.     

Selection of improved Beauveria bassiana (Bals.) Vuill. strains based on 2-deoxy-d-glucose resistance and physiological analysis

A series of 2-deoxy-d-glucose resistant mutants was obtained from wild type Beauveria bassiana 88 (Bb 88) by UV irradiation. Five mutants were characterized on Sabouraud Dextrose Agar and Chitin Agar for both radial extension rate (V_r) and specific growth rate (μ). These values were obtained after adjusting morphometric data to a mathematical model used for filamentous fungi. Additionally, the protease and lipase potency index, conidial size, viability, and production levels were analyzed. The highest values for those physiological measurements were obtained by mutant 882.5 which, relative to Bb 88, showed a 30% reduction in half-life (LT₅₀) on Sphenarium purpurascens, 70% on Acheta domesticus, and 71% on Tenebrio molitor larvae and adults. The half lethal concentration (LC₅₀) on T. molitor larvae was 2.8 × 10⁵ conidia/mL (con/mL) and 1.5 × 10⁶ con/mL, respectively, for mutant 882.5 and Bb 88. This demonstrates that mutant 882.5 is more virulent, with up to an 80% reduction in LC₅₀. This work provides a convenient method for improving strains to be used in biocontrol as a suitable alternative to transgenic constructs.     

Recommendations for Improved Assessment of Noise Impacts on Wildlife

ABSTRACT Research to determine noise impacts on animals benefits from methodology that adequately describes the acoustical stimulus as well as the resulting biological responses. We present acoustical considerations and research techniques that we have found to be useful. These include acoustical definitions and noise measurement techniques that conform to standardized acoustical practice and advice for controlled experimentation to supplement behavioral observation. Specific considerations include characteristics of noise stimulus, selection of noise metrics, use of frequency-weighting algorithms tailored to a specific animal species, selection and placement of noise measurement equipment, and methods for documenting animal responses. We also present arguments for measuring the noise stimulus at the location and time of each response observation. Our purpose is to recommend some baseline terminology, metrics, and techniques prerequisite to effective assessment of noise impacts on terrestrial wildlife whenever and wherever potential conflicts arise.     

Measuring tendon properties in mdx mice: Cell viability and viscoelastic characteristics

Muscular dystrophy is a genetic disorder of skeletal muscle characterized by progressive muscle weakness. Here we assessed whether muscle wasting affects cell viability and mechanical properties of extensor digitorum longus (EDL) and of tibialis anterior (TA) tendons from mdx dystrophic mice compared to wild type (WT) mice. mdx mice represent the classical animal model for human Duchenne muscular dystrophy, and show several signs of the pathology, including a decrease in specific force and an increase of fibrotic index. Cell viability of tendons was evaluated by histological analysis, and viscoelastic properties have been assessed by a rapid measurement protocol that allowed us to compute, at the same time, tissue complex compliance for all the frequencies of interest. Confocal microscopy and mechanical properties measurements revealed that mdx tendons, compared to WT ones, have an increase in the number of dead cells and a significant reduction in tissue elasticity for all the frequencies that were tested. These findings indicate a reduced quality of the tissue. Moreover, mdx tendons have an increase in the viscous response, indicating that during dynamic loading, they dissipate more energy compared to WT. Our results demonstrate that muscular dystrophy involves not only muscle wasting, but also alteration in the viscoelastic properties of tendons, suggesting a paracrine effect of altered skeletal muscle on tendinous tissue.