The Application of Fluorescent Nanocrystals to Study the Distribution of Neuropeptides in the Aedes Aegypti Mosquito

Semiconductor nanocrystals, also known as quantum dots (QDs), have been used in studies involving mice and human tissues, but never before in research on insects. We used QDs to study the distribution of two neuropeptides in the Aedes aegypti mosquito, the vector of both dengue and yellow fever. These neuropeptides play a significant role in the production of juvenile hormone, a hormone that controls biting behavior, metamorphosis, and reproduction throughout the life of the mosquito. The two neuropeptides allatostatin-C (AS-C) and allatotropin (AT) function as inhibitory (AS-C) and stimulatory (AT) regulators of juvenile hormone synthesis in the corpus allatum gland. In other insects, they also affect heart rate, gut movement, and nutrient uptake. Conjugating these neuropeptides to quantum dots via a streptavidinlbiotin link, we were able to expose the mosquito corpus allatum and abdomen to allatostatin-C and allatotropin and then to visualize their distribution under UV light using confocal and compound light microscopy. Histological sections of the whole mosquito, incubations of tissues with conjugates (in vitro), and microinjections of conjugates into the mosquito (in vivo) were performed. The results showed that quantum dots can be used to detect neuropeptide distribution in the mosquito. The more we understand about these neuropeptides and juvenile hormone, the more we can contribute to stopping the spread of infectious diseases, such as dengue and yellow fever.

Event layer chronology from Lake Suigetsu (Japan, SG06 Project) for the last 40 kyr

Event layers in lake sediments are indicators of past extreme events, mostly the results of floods or earthquakes. Detailed characterisation of the layers allows the discrimination of the sedimentation processes involved, such as surface runoff, landslides or subaqueous slope failures. These processes can then be interpreted in terms of their triggering mechanisms. Here we present a 40 kyr event layer chronology from Lake Suigetsu, Japan. The event layers were characterised using a multi-proxy approach, employing light microscopy and µXRF for microfacies analysis. The vast majority of event layers in Lake Suigetsu was produced by flood events (362 out of 369), allowing the construction of the first long-term, quantitative (with respect to recurrence) and well dated flood chronology from the region. The flood layer frequency shows a high variability over the last 40 kyr, and it appears that extreme precipitation events were decoupled from the average long-term precipitation. For instance, the flood layer frequency is highest in the Glacial at around 25 kyr BP, at which time Japan was experiencing a generally cold and dry climate. Other cold episodes, such as Heinrich Event 1 or the Late Glacial stadial, show a low flood layer frequency. Both observations together exclude a simple, straightforward relationship with average precipitation and temperature. We argue that, especially during Glacial times, changes in typhoon genesis/typhoon tracks are the most likely control on the flood layer frequency, rather than changes in the monsoon front or snow melts. Spectral analysis of the flood chronology revealed periodic variations on centennial and millennial time scales, with 220 yr, 450 yr and a 2000 yr cyclicity most pronounced. However, the flood layer frequency appears to have not only been influenced by climate changes, but also by changes in erosion rates due to, for instance, earthquakes.

(Table 2) Relative contents of clay minerals in aeolian pelagic clays from the Pacific Ocean

For determining importance, composition, and history of aerosol material accumulation in formation of pelagic clays a study with use of light microscopy and scanning electron microscopy, X-ray diffractometry, and chemical methods has been carried out.

Global distributions of diazotrophs abundance, biomass and nitrogen fixation rates - Collection of source datasets - Contribution to the MAREDAT World Ocean Atlas of Plankton Functional Types

The MAREDAT atlas covers 11 types of plankton, ranging in size from bacteria to jellyfish. Together, these plankton groups determine the health and productivity of the global ocean and play a vital role in the global carbon cycle. Working within a uniform and consistent spatial and depth grid (map) of the global ocean, the researchers compiled thousands and tens of thousands of data points to identify regions of plankton abundance and scarcity as well as areas of data abundance and scarcity. At many of the grid points, the MAREDAT team accomplished the difficult conversion from abundance (numbers of organisms) to biomass (carbon mass of organisms). The MAREDAT atlas provides an unprecedented global data set for ecological and biochemical analysis and modeling as well as a clear mandate for compiling additional existing data and for focusing future data gathering efforts on key groups in key areas of the ocean. The present collection presents the original data sets used to compile Global distributions of diazotrophs abundance, biomass and nitrogen fixation rates

Chlorophyll-a, primary production rates, carbon concentrations and cell counts of coccolithophores, diatoms and microzooplankton measured in water samples from the North Atlantic during the M87/1 cruise in April 2012

The Deep Convection cruise repeatedly sampled two locations in the North Atlantic, sited in the Iceland and Norwegian Basins, onboard the RV Meteor (19 March - 2 May 2012). Samples were collected from multiple casts of a conductivity-temperature-depth (CTD) - Niskin rosette at each station. Water samples for primary production rates, community structure, chlorophyll a [Chl a], calcite [PIC], particulate organic carbon [POC] and biogenic silicic acid [BSi] were collected from predawn casts from six light depths (55%, 20%, 14%, 7%, 5% and 1% of incident PAR). Additional samples for community structure and ancillary parameters were collected from a second cast. Carbon fixation rates were determined using the 13C stable isotope method. Water samples for diatom and micro zooplankton counts, collected from the predawn casts, were preserved with acidic Lugol's solution (2% final solution) and counted using an inverted light microscope. Water samples for coccolithophore counts were collected onto cellulose nitrate filters and counted using polarising light microscopy. Water samples for Chl a analysis were filtered onto MF300 and polycarbonate filters and extracted in 90% acetone. PIC and BSi samples were filtered onto polycarbonate filters and analysed using an inductively coupled plasma emission optical spectrometer and a SEAL QuAAtro autoanalyser respectively.

Coccosphere geometry measurements from culture experiments on the coccolithophore species Calcidiscus leptoporus, Calcidiscus quadriperforatus and Helicosphaera carteri

Coccolithophores are a key phytoplankton group that exhibit remarkable diversity in their biology, ecology, and calcitic exoskeletons (coccospheres). An understanding of the physiological processes that underpin coccosphere architecture is essential for maximizing the information that can be retrieved from their extensive fossil record. Using culturing experiments on four modern species from three long-lived families, we investigate how coccosphere architecture responds to population shifts from rapid (exponential) to slowed (stationary) growth phases as nutrients become depleted. These experiments reveal statistical differences in cell size and the number of coccoliths per cell between these two growth phases, specifically that cells in exponential-phase growth are typically smaller with fewer coccoliths, whereas cells experiencing growth-limiting nutrient depletion have larger coccosphere sizes and greater numbers of coccoliths per cell. Although the exact numbers are species-specific, these growth-phase shifts in coccosphere geometry are common to four different coccolithophore families (Calcidiscaceae, Coccolithaceae, Isochrysidaceae, Helicosphaeraceae), demonstrating that this is a core physiological response to nutrient depletion across a representative diversity of this phytoplankton group. Polarised light microscopy was used for all coccosphere geometry measurements.

Distribution, occurrence and biotoxin composition of the main shellfish toxin producing microalgae within European waters: A comparison of methods of analysis.

Harmful algal blooms (HABs) are a natural global phenomena emerging in severity and extent. Incidents have many economic, ecological and human health impacts. Monitoring and providing early warning of toxic HABs are critical for protecting public health. Current monitoring programmes include measuring the number of toxic phytoplankton cells in the water and biotoxin levels in shellfish tissue. As these efforts are demanding and labour intensive, methods which improve the efficiency are essential. This study compares the utilisation of a multitoxin surface plasmon resonance (multitoxin SPR) biosensor with enzyme-linked immunosorbent assay (ELISA) and analytical methods such as high performance liquid chromatography with fluorescence detection (HPLC-FLD) and liquid chromatography–tandem mass spectrometry (LC–MS/MS) for toxic HAB monitoring efforts in Europe. Seawater samples (n = 256) from European waters, collected 2009–2011, were analysed for biotoxins: saxitoxin and analogues, okadaic acid and dinophysistoxins 1/2 (DTX1/DTX2) and domoic acid responsible for paralytic shellfish poisoning (PSP), diarrheic shellfish poisoning (DSP) and amnesic shellfish poisoning (ASP), respectively. Biotoxins were detected mainly in samples from Spain and Ireland. France and Norway appeared to have the lowest number of toxic samples. Both the multitoxin SPR biosensor and the RNA microarray were more sensitive at detecting toxic HABs than standard light microscopy phytoplankton monitoring. Correlations between each of the detection methods were performed with the overall agreement, based on statistical 2 × 2 comparison tables, between each testing platform ranging between 32% and 74% for all three toxin families illustrating that one individual testing method may not be an ideal solution. An efficient early warning monitoring system for the detection of toxic HABs could therefore be achieved by combining both the multitoxin SPR biosensor and RNA microarray.

Distribution, occurrence and biotoxin composition of the main shellfish toxin producing microalgae within European waters: A comparison of methods of analysis.

Harmful algal blooms (HABs) are a natural global phenomena emerging in severity and extent. Incidents have many economic, ecological and human health impacts. Monitoring and providing early warning of toxic HABs are critical for protecting public health. Current monitoring programmes include measuring the number of toxic phytoplankton cells in the water and biotoxin levels in shellfish tissue. As these efforts are demanding and labour intensive, methods which improve the efficiency are essential. This study compares the utilisation of a multitoxin surface plasmon resonance (multitoxin SPR) biosensor with enzyme-linked immunosorbent assay (ELISA) and analytical methods such as high performance liquid chromatography with fluorescence detection (HPLC-FLD) and liquid chromatography–tandem mass spectrometry (LC–MS/MS) for toxic HAB monitoring efforts in Europe. Seawater samples (n = 256) from European waters, collected 2009–2011, were analysed for biotoxins: saxitoxin and analogues, okadaic acid and dinophysistoxins 1/2 (DTX1/DTX2) and domoic acid responsible for paralytic shellfish poisoning (PSP), diarrheic shellfish poisoning (DSP) and amnesic shellfish poisoning (ASP), respectively. Biotoxins were detected mainly in samples from Spain and Ireland. France and Norway appeared to have the lowest number of toxic samples. Both the multitoxin SPR biosensor and the RNA microarray were more sensitive at detecting toxic HABs than standard light microscopy phytoplankton monitoring. Correlations between each of the detection methods were performed with the overall agreement, based on statistical 2 × 2 comparison tables, between each testing platform ranging between 32% and 74% for all three toxin families illustrating that one individual testing method may not be an ideal solution. An efficient early warning monitoring system for the detection of toxic HABs could therefore be achieved by combining both the multitoxin SPR biosensor and RNA microarray.

Oxidation and tensile behavior of ferritic/martensitic steels after exposure to lead-bismuth eutectic

Two ferritic/martensitic steels, T91 steel and newly developed SIMP steel, were subject to tensile test after being oxidized in the liquid lead-bismuth eutectic (LBE) at 873 K for 500 h, 1000 h and 2000 h. Tensile tests were also carried out on the steels only thermally aged at 873 K. The result shows that thermal aging has no effect. Exposure to LBE at 873 K leads to a slight decrease in strength, but a large decrease in elongation when tested at 873 K. When tested at 873 K after 2000 h exposure, the tensile strength of T91 decreases slightly, and elongation from 39% to 21%. For SIMP, the decreases are slightly and from 44% to 28%, for tensile strength and elongation, respectively. The room temperature strength has slightly larger percentage reductions after the LBE exposure, but the elongation changes little.

Crystallization on the Mesoscale : Self-Assembly of Iron Oxide Nanocubes into Mesocrystals

Self-assembly of nanoparticles is a promising route to form complex, nanostructured materials with functional properties. Nanoparticle assemblies characterized by a crystallographic alignment of the nanoparticles on the atomic scale, i.e. mesocrystals, are commonly found in nature with outstanding functional and mechanical properties. This thesis aims to investigate and understand the formation mechanisms of mesocrystals formed by self-assembling iron oxide nanocubes. We have used the thermal decomposition method to synthesize monodisperse, oleate-capped iron oxide nanocubes with average edge lengths between 7 nm and 12 nm and studied the evaporation-induced self-assembly in dilute toluene-based nanocube dispersions. The influence of packing constraints on the alignment of the nanocubes in nanofluidic containers has been investigated with small and wide angle X-ray scattering (SAXS and WAXS, respectively). We found that the nanocubes preferentially orient one of their {100} faces with the confining channel wall and display mesocrystalline alignment irrespective of the channel widths.  We manipulated the solvent evaporation rate of drop-cast dispersions on fluorosilane-functionalized silica substrates in a custom-designed cell. The growth stages of the assembly process were investigated using light microscopy and quartz crystal microbalance with dissipation monitoring (QCM-D). We found that particle transport phenomena, e.g. the coffee ring effect and Marangoni flow, result in complex-shaped arrays near the three-phase contact line of a drying colloidal drop when the nitrogen flow rate is high. Diffusion-driven nanoparticle assembly into large mesocrystals with a well-defined morphology dominates at much lower nitrogen flow rates. Analysis of the time-resolved video microscopy data was used to quantify the mesocrystal growth and establish a particle diffusion-based, three-dimensional growth model. The dissipation obtained from the QCM-D signal reached its maximum value when the microscopy-observed lateral growth of the mesocrystals ceased, which we address to the fluid-like behavior of the mesocrystals and their weak binding to the substrate. Analysis of electron microscopy images and diffraction patterns showed that the formed arrays display significant nanoparticle ordering, regardless of the distinctive formation process.  We followed the two-stage formation mechanism of mesocrystals in levitating colloidal drops with real-time SAXS. Modelling of the SAXS data with the square-well potential together with calculations of van der Waals interactions suggests that the nanocubes initially form disordered clusters, which quickly transform into an ordered phase.

Spliceosomal small nuclear ribonucleoprotein particle trafficking and maturation in the nuclear compartment

We show for the first time that upon injection into the cytoplasm of the oocyte, fluorescein-labeled spliceosomal snRNAs, in the context of functional snRNPs, are targeted to elongating pre-mRNAs. This finding presents us with a novel assay with which to dissect the mechanism by which snRNPs are targeted to nascent pre-mRNA transcripts. Two critical advantages offered by this system are immediately evident. First, it allows us to investigate the mechanisms employed to recruit snRNPs as it actually transpires within the realm of the cell nucleus. Second, it allows a genome-wide analysis of snRNP recruitment to nascent transcripts, and, hence, the conclusions drawn from these studies do not depend on the sequence of any particular promoter or pre-mRNA. Indeed, it is with this assay that we have stumbled upon a most unanticipated discovery: Contrary to the current paradigm, the co-transcriptional recruitment of splicing snRNPs to nascent transcripts is not contingent on their role in splicing in vivo. Based on these and other data, we have constructed a two-step recruitment-loading model wherein snRNPs are first recruited to pre-mRNA transcripts and only then loaded directly onto cis-acting sequences on nascent pre-mRNA. While conducting studies on snRNP trafficking, a new discovery was made. We found that the lampbrush chromosomes could be visualized by light microscopy in vivo, and that these chromosomes have an architecture that is identical with those in formaldehyde treated nuclear spread preparations. Importantly, we now have the first system with which we can examine the dynamic interactions of macromolecules with specific RNA polymerase II transcriptional units in the live nucleus.

Effects of silicon treatment and inoculation with Fusarium oxysporum f. sp. vasinfectum on cellular defences in root tissues of two cotton cultivars

BACKGROUND AND AIMS: Silicon has been shown to enhance the resistance of plants to fungal and bacterial pathogens. Here, the effect of potassium silicate was assessed on two cotton (Gossypium hirsutum) cultivars subsequently inoculated with Fusarium oxysporum f. sp. vasinfectum (Fov). Sicot 189 is moderately resistant whilst Sicot F-1 is the second most resistant commercial cultivar presently available in Australia. METHODS: Transmission and light microscopy were used to compare cellular modifications in root cells after these different treatments. The accumulation of phenolic compounds and lignin was measured. KEY RESULTS: Cellular alterations including the deposition of electron-dense material, degradation of fungal hyphae and occlusion of endodermal cells were more rapidly induced and more intense in endodermal and vascular regions of Sicot F-1 plants supplied with potassium silicate followed by inoculation with Fov than in similarly treated Sicot 189 plants or in silicate-treated plants of either cultivar not inoculated with Fov. Significantly more phenolic compounds were present at 7 d post-infection (dpi) in root extracts of Sicot F-1 plants treated with potassium silicate followed by inoculation with Fov compared with plants from all other treatments. The lignin concentration at 3 dpi in root material from Sicot F-1 treated with potassium silicate and inoculated with Fov was significantly higher than that from water-treated and inoculated plants. CONCLUSIONS: This study demonstrates that silicon treatment can affect cellular defence responses in cotton roots subsequently inoculated with Fov, particularly in Sicot F-1, a cultivar with greater inherent resistance to this pathogen. This suggests that silicon may interact with or initiate defence pathways faster in this cultivar than in the less resistant cultivar.

Rapid detection and quantification of the marine toxic algae, Alexandrium minutum, using a super-paramagnetic immunochromatographic strip test

The dinoflagellates of Alexandrium genus are known to be producers of paralytic shellfish toxins that regularly impact the shellfish aquaculture industry and fisheries. Accurate detection of Alexandrium including A. minutum is crucial for environmental monitoring and sanitary issues. In this study, we firstly developed a quantitative lateral flow immunoassay (LFIA) using super-paramagnetic nanobeads for A. minutum whole cells. This dipstick assay relies on two distinct monoclonal antibodies used in a sandwich format and directed against surface antigens of this organism. No sample preparation is required. Either frozen or live cells can be detected and quantified. The specificity and sensitivity are assessed by using phytoplankton culture and field samples spiked with a known amount of cultured A. minutum cells. This LFIA is shown to be highly specific for A. minutum and able to detect reproducibly 105 cells/L within 30 min. The test is applied to environmental samples already characterized by light microscopy counting. No significant difference is observed between the cell densities obtained by these two methods. This handy super-paramagnetic lateral flow immnunoassay biosensor can greatly assist water quality monitoring programs as well as ecological research.

Age estimation in common sole Solea solea larvae: validation of daily increments and evaluation of a pattern recognition technique

Close similarities have been found between the otoliths of sea-caught and laboratory-reared larvae of the common sole Solea solea (L.), given appropriate temperatures and nourishment of the latter. But from hatching to mouth formation. and during metamorphosis, sole otoliths have proven difficult to read because the increments may be less regular and low contrast. In this study, the growth increments in otoliths of larvae reared at 12 degrees C were counted by light microscopy to test the hypothesis of daily deposition, with some results verified using scanning electron microscopy (SEM), and by image analysis in order to compare the reliability of the 2 methods in age estimation. Age was first estimated (in days posthatch) from light micrographs of whole mounted otoliths. Counts were initiated from the increment formed at the time of month opening (Day 4). The average incremental deposition rate was consistent with the daily hypothesis. However, the light-micrograph readings tended to underestimate the mean ages of the larvae. Errors were probably associated with the low-contrast increments: those deposited after the mouth formation during the transition to first feeding, and those deposited from the onset of eye migration (about 20 d posthatch) during metamorphosis. SEM failed to resolve these low-contrast areas accurately because of poor etching. A method using image analysis was applied to a subsample of micrograph-counted otoliths. The image analysis was supported by an algorithm of pattern recognition (Growth Demodulation Algorithm, GDA). On each otolith, the GDA method integrated the growth pattern of these larval otoliths to averaged data from different radial profiles, in order to demodulate the exponential trend of the signal before spectral analysis (Fast Fourier Transformation, FFT). This second method both allowed more precise designation of increments, particularly for low-contrast areas, and more accurate readings but increased error in mean age estimation. The variability is probably due to a still rough perception of otolith increments by the GDA method, counting being achieved through a theoretical exponential pattern and mean estimates being given by FFT. Although this error variability was greater than expected, the method provides for improvement in both speed and accuracy in otolith readings.

Semen profiles of young men involved as bicycle taxi cyclists in Mangochi District, Malawi: A case-control study

Aim The purpose of this study was to evaluate the semen profiles of bicycle taxi cyclists and healthy controls in Mangochi district, Malawi. Methods Semen samples were collected from young bicycle taxi cyclists after two to three days of sexual abstinence. A control group, comprising young men who were not bicycle taxi operators also submitted semen samples. Samples were left to liquefy for 30 minutes before measurements were conducted of volume, concentration, total motility, and progressive motility. This was followed by preparation of morphology slides. Light microscopy was used for sperm analysis. Results Semen parameters such as volume (1.66 ± 0.18 mL vs. 3.64 ± 0.17 mL; p = 0.0001), concentration (28.31 ± 4.33 x 106/mL vs. 54.95 ± 5.93 x 106/ mL; p = 0.02) , total motility (56.98% ± 8.22% vs. 56.98% ± 8.22%; p = 0.03), progressive motility (22.57% ± 3.35% vs. 59.69% ± 4.82%; p = 0.004), and morphology (6.98% ± 3.23% vs. 19.73% ± 2.32%; p = 0.006) were significantly reduced in the bicycle taxi cyclists compared to the healthy controls. Conclusion In this case-control study, bicycle taxi operators had lower semen volume, concentration, total motility, and progressive motility, as well as a higher concentration of abnormally shaped spermatocytes, compared to healthy controls.