The in situ sinking rates of herbivore fecal pellets

The vertical flux of pigmented fecal pellets was measured forperiods of 4 h or less with closely spaced sediment traps. Thesettling of the nighttime injection resulting from macrozooplanktongrazing activity could be monitored as the pellets settled throughthe water column. Sinking rates of fecal pellets showed a rangeof 31 – 122 m d^–1 with an average of 87 m d^–1.Longer term experiments show that the vertical phaeopigmentflux is conservative, and that there is no significant lossof pigments en route to the sea floor. In Dabob Bay, coprophagyin the water column is most likely a minor consideration. Theamount of chlorophyll sinking out of the euphotic zone usuallyrepresents <1% of the standing crop, and is usually only5% or less than the phaeopigment flux. This suggests that grazingis the predominant process accounting for the major loss ofchlorophyll from the water column. ¹Contribution no. 1332 from the School of Oceanography, Universityof Washington, Seattle, WA 98195, USA. ²Present address: The Biological Laboratories, Harvard University,Cambridge, MA 02138, USA.     

Basic fibroblast growth factor and interleukins 4 and 6 stimulate the release of IFN-gamma by individual NK cells.

Both the secretory and cytotoxic activity of natural killer (NK) cells are known to be regulated by such cytokines as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma). In the present study we have used the reverse hemolytic plaque assay to investigate either the direct effects of the protein kinase activator, phorbol myristate acetate (PMA), or exposure to recombinant human interleukins 2, 4, and 6 (IL-2, IL-4, and IL-6) tumour necrosis factor alpha (TNF-alpha) and basic fibroblast growth factor (bFGF) on the release of IFN-gamma by individual, immunoidentified NK cells isolated from peripheral blood. This sensitive immunoassay was adapted and coupled with immunocytochemistry not only to immunophenotype and enumerate cells secreting IFN-gamma in a given cell population, but also to quantify the amount of this cytokine released per individual cell. These studies have confirmed mononuclear cells with the morphology of large granular lymphocytes and the immunophenotype of CD3-/CD16+ NK cells to be the predominant source of spontaneously released IFN-gamma in vitro. In contrast to this, fewer than 2% of the CD3+ T cells secreted detectable levels of this cytokine during the assay, irrespective of the stimulus applied. Whilst TNF-alpha had no significant effect on IFN-gamma release by NK cells, a 6-hr exposure to IL-2 or PMA stimulated an increase in the amount secreted per single cell. Furthermore, bFGF and interleukins 4 and 6 elicited a marked, dose-dependent stimulation of IFN-gamma secretion by this cell type. However, exposure to these cytokines did not alter the number of cells capable of releasing detectable levels of IFN-gamma during the assay. These studies demonstrate that (i) both the spontaneous and stimulated release of IFN-gamma by NK cells can be visualized and quantified at the single-cell level using this sensitive immunoassay, and (ii) bFGF and interleukins 2, 4, and 6, but not TNF-alpha, are potent stimulants of IFN-gamma secretion by CD3-/CD16+ NK cells.     

Problems of recordings of nervous activity with glass microelectrodes in the CNS of insects

1. Extracellular single unit recordings with glass microelectrodes in the central nervous system of insects display action potentials of variable amplitude, polarity and time course. This phenomenon is due to capacitive influences at the electrode in contact with the tissue. This is demonstrated by an electrical model circuit simulating extracellular recording conditions. 2. Extracellularly recorded potentials often are very similar to intracellularly recorded ones. Criteria for the decision whether the electrode is intracellularly or not are discussed. 3. Action potentials and slow potentials were recorded simultaneously in the acoustic neuropiles of Locusta. Since slow potentials may not only be distorted by capacitive properties of both the tissue and the electrode, but also are influenced by the anatomical organization of the nervous tissue, their interpretation is ambiguous.     

Biosensor Determination of the Microscale Distribution of Nitrate, Nitrate Assimilation, Nitrification, and Denitrification in a Diatom-Inhabited Freshwater Sediment

High-resolution NO₃⁻ profiles in freshwater sediment covered with benthic diatoms were obtained with a new microscale NO₃⁻ biosensor characterized by absence of interference from chemical species other than NO₂⁻ and N₂O. Analysis of the microprofiles obtained indicated no nitrification during darkness, high rates of nitrification and a tight coupling between nitrification and denitrification during illumination, and substantial rates of NO₃⁻ assimilation during illumination. Nitrification during darkness could be induced by purging the bulk water with O₂ gas, indicating that the stimulatory effect on nitrification by illumination was caused by algal production of O₂. NH₄⁺ addition did not stimulate nitrification during darkness when O₂ was restricted to the upper 1-mm layer, and there was thus a low nitrification potential in the permanently oxic top 1 mm of the sediment.     

A Rhizobium meliloti lipopolysaccharide mutant altered in competitiveness for nodulation of alfalfa.

A transposon Tn5-induced mutant of Rhizobium meliloti Rm2011, designated Rm6963, showed a rough colony morphology on rich and minimal media and an altered lipopolysaccharide (LPS). Major differences from the wild-type LPS were observed in (i) hexose and 2-keto-3-deoxyoctonate elution profiles of crude phenol extracts chromatographed in Sepharose CL-4B, (ii) silver-stained sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis patterns of crude and purified LPS fractions, and (iii) immunoreactivities otherwise present in purified LPS of the parental strain Rm2011. In addition, Rm6963 lost the ability to grow in Luria-Bertani medium containing the hydrophobic compounds sodium deoxycholate or SDS and showed a decrease in survival in TY medium supplemented with high calcium concentrations. The mutant also had altered symbiotic properties. Rm6963 formed nodules that fixed nitrogen but showed a delayed or even reduced ability to nodulate the primary root of alfalfa without showing changes in the position of nodule distribution profiles along the roots. Furthermore, 2 to 3 weeks after inoculation, plants nodulated by Rm6963 were smaller than control plants inoculated with wild-type bacteria in correlation with a transient decrease in nitrogen fixation. In most experiments, the plants recovered later by expressing a full nitrogen-fixing phenotype and developing an abnormally high number of small nodules in lateral roots after 1 month. Rm6963 was also deficient in the ability to compete for nodulation. In coinoculation experiments with equal bacterial numbers of both mutant and wild-type rhizobia, only the parent was recovered from the uppermost root nodules. A strain ratio of approximately 100 to 1 favoring the mutant was necessary to obtain an equal ratio (1:1) of nodule occupancy. These results show that alterations in Rm6963 which include LPS changes lead to an altered symbiotic phenotype during the association with alfalfa that affects the timing of nodule emergence, the progress of nitrogen fixation, and the strain competitiveness for nodulation.     

Time course study of in situ expression of antigens following DNA-vaccination against VHS in rainbow trout (Oncorhynchus mykiss Walbaum) fry

The present study was performed as a time course study of fish vaccinated with 20 microg plasmid DNA vaccine encoding either the VHSV G-protein or the VHSV N-protein. Samples of the injection site were collected sequentially over a 7-week period. The study revealed an intense positive staining by immunohistochemistry for the viral G-protein mainly in the membrane of intact myocytes, most prominent by days 10-27, and with concomitant infiltration of inflammatory cells by days 13-38 that subsequently lead to a marked reduction in the number of myocytes expressing the G-protein. By immunofluorescence, infiltrating cells positive for MHC II, IgM, and C3 were demonstrated. By contrast, in fish vaccinated with the VHSV-N construct, fewer, diffusely positive myocytes were found, most prominent by days 13-38, these having a positive reaction for the N-protein mainly in the cytoplasm and variably in the membrane. N-protein positive myocytes did not attract infiltrating cells to the same degree. Positive reaction for the N-protein almost ceased by day 48 post-vaccination.     

In silico screening of drug databases for TSE inhibitors

Today, thousands of different chemical compounds are used as drugs for a wealth of different indications. Here, we demonstrate the use of a novel conformational drug database for the search of compounds with a positive influence on Transmissible Spongiform Encephalopathies (TSEs) by using two- and three-dimensional structural similarity to compounds with known effect. Both methods are combined to deduce a list of 16 candidate drugs. The proposal of a small number of putative inhibitors out of about 2300 approved essential drugs allows testing by expensive or time-consuming methods with the advantage that all agents are well known and suitable for use in humans.     

Parallel phylogenetic analyses using the N, G or Nv gene from a fixed group of VHSV isolates reveal the same overall genetic typing

Different genetic regions representing the viral phospho- (P), nucleocapsid- (N) or glyco-protein (G) gene have been used for phylogenetic studies of viral haemorrhagic septicaemia virus (VHSV). Since these analyses were performed on different virus isolates using various genomic regions, it has been difficult to evaluate how the choice of target region affects the output of the analyses. To address this, we sequenced and performed parallel phylogenetic analysis of an N gene fragment, the entire Nv (non-structural protein) and G genes, and 4 different fragments of the G gene from a fixed virus panel. The overall genotyping of the selected isolates was identical for the 7 target regions, but separation of Genotype I sub-lineages was best when the analysis was performed on the full length G gene (1524 nucleotides, nt). Good resolution was furthermore obtained using smaller sequencing windows represented by a G gene fragment (nt 360 to 720) or the Nv gene (366 nt), although these regions had different characteristics with respect to resolution of Genotype I sublineages and resolution within Sub-lineage Ia. Phylogenetic analysis based on the deduced amino acid sequences was also performed. The phylogenetic relationship between the nucleotide and amino acid sequences of the isolates corresponded best in the case of the N gene/protein. For the 6 other genomic regions, genetically distant isolates occasionally grouped together when compared at protein levels. No clear relationship between the G gene genotyping and serotyping with neutralising (G protein specific) antibodies was observed, stressing that epidemiological analysis based on phenotypic characteristics such as serotype could be misleading.     

Genetic linkage maps of the red flour beetle, Tribolium castaneum, based on bacterial artificial chromosomes and expressed sequence tags

A genetic linkage map was constructed in a backcross family of the red flour beetle, Tribolium castaneum, based largely on sequences from bacterial artificial chromosome (BAC) ends and untranslated regions from random cDNA's. In most cases, dimorphisms were detected using heteroduplex or single-strand conformational polymorphism analysis after specific PCR amplification. The map incorporates a total of 424 markers, including 190 BACs and 165 cDNA's, as well as 69 genes, transposon insertion sites, sequence-tagged sites, microsatellites, and amplified fragment-length polymorphisms. Mapped loci are distributed along 571 cM, spanning all 10 linkage groups at an average marker separation of 1.3 cM. This genetic map provides a framework for positional cloning and a scaffold for integration of the emerging physical map and genome sequence assembly. The map and corresponding sequences can be accessed through BeetleBase (http://www.bioinformatics.ksu.edu/BeetleBase/).     

Growth and nutrient responses of Eloecharis cellulosa (Cyperaceae) to phosphate level and redox intensity

Phosphorus (P) availability limits plant growth in many ecosystems. The ability of plants to explore for soil P is often impaired by nonresource stressors. Understanding the effects of these stressors on P acquisition in oligotrophic environments is critical in predicting species dominance. Growth and nutrient responses of Eleocharis cellulosa to redox intensity and phosphate level were evaluated under three redox potentials (Eh) and three phosphate (PO(4)) levels (P). Although low Eh (-150 mV) decreased root length at low P, Eh did not affect shoot height, relative growth rate (RGR), shoot elongation, photosynthesis, or biomass of E. cellulosa. Low PO(4) (10 μg P?·?L(-1)) strongly inhibited growth. Shoot height, RGR, elongation, photosynthesis, and biomass were lower at 10 μg P?·?L(-1) than at 80 or 500 μg P?·?L(-1). None of the growth variables, except the ratio of root-supported biomass to root biomass, significantly differed between the 80 and 500 μg P?·?L(-1) treatments. At low P, plants allocated relatively more biomass to roots than to shoots, compared to the medium and high P levels. Eleocharis cellulosa is well adapted to flooded conditions that lower soil Eh, and elevated PO(4) levels further promote its growth potential.