Laser Based Mid-Infrared Spectroscopic Imaging – Exploring a Novel Method for Application in Cancer Diagnosis

A number of biomedical studies have shown that mid-infrared spectroscopic images can provide both morphological and biochemical information that can be used for the diagnosis of cancer. Whilst this technique has shown great potential it has yet to be employed by the medical profession. By replacing the conventional broadband thermal source employed in modern FTIR spectrometers with high-brightness, broadly tuneable laser based sources (QCLs and OPGs) we aim to solve one of the main obstacles to the transfer of this technology to the medical arena; namely poor signal to noise ratios at high spatial resolutions and short image acquisition times. In this thesis we take the first steps towards developing the optimum experimental configuration, the data processing algorithms and the spectroscopic image contrast and enhancement methods needed to utilise these high intensity laser based sources. We show that a QCL system is better suited to providing numerical absorbance values (biochemical information) than an OPG system primarily due to the QCL pulse stability. We also discuss practical protocols for the application of spectroscopic imaging to cancer diagnosis and present our spectroscopic imaging results from our laser based spectroscopic imaging experiments of oesophageal cancer tissue

Colony formation in Phaeocystis antarctica : connecting molecular mechanisms with iron biogeochemistry

© The Author(s), 2018. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Biogeosciences 15 (2018): 4923-4942, doi:10.5194/bg-15-4923-2018.Phaeocystis antarctica is an important phytoplankter of the Ross Sea where it dominates the early season bloom after sea ice retreat and is a major contributor to carbon export. The factors that influence Phaeocystis colony formation and the resultant Ross Sea bloom initiation have been of great scientific interest, yet there is little known about the underlying mechanisms responsible for these phenomena. Here, we present laboratory and field studies on Phaeocystis antarctica grown under multiple iron conditions using a coupled proteomic and transcriptomic approach. P. antarctica had a lower iron limitation threshold than a Ross Sea diatom Chaetoceros sp., and at increased iron nutrition (>120pM Fe') a shift from flagellate cells to a majority of colonial cells in P. antarctica was observed, implying a role for iron as a trigger for colony formation. Proteome analysis revealed an extensive and coordinated shift in proteome structure linked to iron availability and life cycle transitions with 327 and 436 proteins measured as significantly different between low and high iron in strains 1871 and 1374, respectively. The enzymes flavodoxin and plastocyanin that can functionally replace iron metalloenzymes were observed at low iron treatments consistent with cellular iron-sparing strategies, with plastocyanin having a larger dynamic range. The numerous isoforms of the putative iron-starvation-induced protein (ISIP) group (ISIP2A and ISIP3) had abundance patterns coinciding with that of either low or high iron (and coincident flagellate or the colonial cell types in strain 1871), implying that there may be specific iron acquisition systems for each life cycle type. The proteome analysis also revealed numerous structural proteins associated with each cell type: within flagellate cells actin and tubulin from flagella and haptonema structures as well as a suite of calcium-binding proteins with EF domains were observed. In the colony-dominated samples a variety of structural proteins were observed that are also often found in multicellular organisms including spondins, lectins, fibrillins, and glycoproteins with von Willebrand domains. A large number of proteins of unknown function were identified that became abundant at either high or low iron availability. These results were compared to the first metaproteomic analysis of a Ross Sea Phaeocystis bloom to connect the mechanistic information to the in situ ecology and biogeochemistry. Proteins associated with both flagellate and colonial cells were observed in the bloom sample consistent with the need for both cell types within a growing bloom. Bacterial iron storage and B12 biosynthesis proteins were also observed consistent with chemical synergies within the colony microbiome to cope with the biogeochemical conditions. Together these responses reveal a complex, highly coordinated effort by P. antarctica to regulate its phenotype at the molecular level in response to iron and provide a window into the biology, ecology, and biogeochemistry of this group.Support for this study was provided by an Investigator grant to Mak A. Saito from the Gordon and Betty Moore Foundation (GBMF3782), National Science Foundation grants NSF-PLR 0732665, OCE-1435056, OCE-1220484, and ANT-1643684, the WHOI Coastal Ocean Institute, and a CINAR Postdoctoral Scholar Fellowship provided to Sara J. Bender through the Woods Hole Oceanographic Institution. Support was provided to Andrew E. Allen through NSF awards ANT-0732822, ANT-1043671, and OCE-1136477 and Gordon and Betty Moore Foundation grant GBMF3828. Additional support was provided to GRD through NSF award OPP-0338097

Phytoplankton-Bacterial Interactions Mediate Micronutrient Colimitation at the Coastal Antarctic Sea Ice Edge

Southern Ocean primary productivity plays a key role in global ocean biogeochemistry and climate. At the Southern Ocean sea ice edge in coastal McMurdo Sound, we observed simultaneous cobalamin and iron limitation of surface water phytoplankton communities in late Austral summer. Cobalamin is produced only by bacteria and archaea, suggesting phytoplankton–bacterial interactions must play a role in this limitation. To characterize these interactions and investigate the molecular basis of multiple nutrient limitation, we examined transitions in global gene expression over short time scales, induced by shifts in micronutrient availability. Diatoms, the dominant primary producers, exhibited transcriptional patterns indicative of co-occurring iron and cobalamin deprivation. The major contributor to cobalamin biosynthesis gene expression was a gammaproteobacterial population, Oceanospirillaceae ASP10-02a. This group also contributed significantly to metagenomic cobalamin biosynthesis gene abundance throughout Southern Ocean surface waters. Oceanospirillaceae ASP10-02a displayed elevated expression of organic matter acquisition and cell surface attachment-related genes, consistent with a mutualistic relationship in which they are dependent on phytoplankton growth to fuel cobalamin production. Separate bacterial groups, including Methylophaga, appeared to rely on phytoplankton for carbon and energy sources, but displayed gene expression patterns consistent with iron and cobalamin deprivation. This suggests they also compete with phytoplankton and are important cobalamin consumers. Expression patterns of siderophore- related genes offer evidence for bacterial influences on iron availability as well. The nature and degree of this episodic colimitation appear to be mediated by a series of phytoplankton–bacterial interactions in both positive and negative feedback loops

Influence of nutrients and currents on the genomic composition of microbes across an upwelling mosaic

Metagenomic data sets were generated from samples collected along a coastal to open ocean transect between Southern California Bight and California Current waters during a seasonal upwelling event, providing an opportunity to examine the impact of episodic pulses of cold nutrient-rich water into surface ocean microbial communities. The data set consists of ∼5.8 million predicted proteins across seven sites, from three different size classes: 0.1–0.8, 0.8–3.0 and 3.0–200.0 μm. Taxonomic and metabolic analyses suggest that sequences from the 0.1–0.8 μm size class correlated with their position along the upwelling mosaic. However, taxonomic profiles of bacteria from the larger size classes (0.8–200 μm) were less constrained by habitat and characterized by an increase in Cyanobacteria, Bacteroidetes, Flavobacteria and double-stranded DNA viral sequences. Functional annotation of transmembrane proteins indicate that sites comprised of organisms with small genomes have an enrichment of transporters with substrate specificities for amino acids, iron and cadmium, whereas organisms with larger genomes have a higher percentage of transporters for ammonium and potassium. Eukaryotic-type glutamine synthetase (GS) II proteins were identified and taxonomically classified as viral, most closely related to the GSII in Mimivirus, suggesting that marine Mimivirus-like particles may have played a role in the transfer of GSII gene functions. Additionally, a Planctomycete bloom was sampled from one upwelling site providing a rare opportunity to assess the genomic composition of a marine Planctomycete population. The significant correlations observed between genomic properties, community structure and nutrient availability provide insights into habitat-driven dynamics among oligotrophic versus upwelled marine waters adjoining each other spatially

Delirium care: real-world solutions to real-world problems

Implementation research into delirium care is lacking. Exploiting known practice barriers to understand what management strategies work best in delirium is a means of prioritising care interventions. A consensus approach to determining priority interventions in delirium was derived and related to reference standards in health-care practice.A workshop of 20 experts was held at the Australasian Delirium Association conference 2016. Structured small group work, iterative ranking and a 21-member check were undertaken to (i) explore research barriers in delirium care; (ii) explore how barriers related to individual items of multicomponent interventions; and (iii) rank multicomponent interventions in relation to each statement within the newly released Australian Commission on Safety and Quality in Health Care delirium standard.Top-ranking interventions included the following: education and training, comprehensive geriatric assessment, family partnerships, individualised care and multidisciplinary engagement.Delirium experts identified a minimum standard of any care intervention for delirium

Species-specific responses of marine bacteria to environmental perturbation.

Environmental perturbations shape the structure and function of microbial communities. Oil spills are a major perturbation and resolving spills often requires active measures like dispersant application that can exacerbate the initial disturbance. Species-specific responses of microorganisms to oil and dispersant exposure during such perturbations remain largely unknown. We merged metatranscriptomic libraries with pangenomes to generate Core-Accessory Metatranscriptomes (CA-Metatranscriptomes) for two microbial hydrocarbon degraders that played important roles in the aftermath of the Deepwater Horizon oil spill. The Colwellia CA-Metatranscriptome illustrated pronounced dispersant-driven acceleration of core (~41%) and accessory gene (~59%) transcription, suggesting an opportunistic strategy. Marinobacter responded to oil exposure by expressing mainly accessory genes (~93%), suggesting an effective hydrocarbon-degrading lifestyle. The CA-Metatranscriptome approach offers a robust way to identify the underlying mechanisms of key microbial functions and highlights differences of specialist-vs-opportunistic responses to environmental disturbance

Molecular Approaches for an Operational Marine Biodiversity Observation Network

Biological health underpins ecosystem services, including the production of food from the sea. Therefore, many policy frameworks rely on biological observations to guide the sustainable use of marine resources. For example, a marine biodiversity observation network can provide evidence of ecosystem degradation, remediation success, and progress toward goals of sustainable development. The utility of biodiversity observations is limited, however, if measurements are not standardized to allow comparisons across space and time or if they are too difficult or expensive to make routinely or at large scale. Molecular approaches (e.g., metagenomics, metabarcoding, eDNA, metatranscriptomics) offer efficiencies in sample collection and processing and provide analysis across trophic levels compared to traditional methods. Increased affordability and throughput of nucleic acid sequencing allow integration of molecular biological techniques into operational monitoring with the potential benefit of expanding the spatial and temporal scale of biological observations

Κρητικό ιατροσόφιον του 19ου αιώνα / έκδοση, εισαγωγή, σημειώσεις Νικόλαος Ε. Παπαδογιαννάκης.

There is an intricate interaction between iron (Fe) and copper (Cu) physiology in diatoms. However, strategies to cope with low Cu are largely unknown. This study unveils the comprehensive restructuring of the photosynthetic apparatus in the diatom Thalassiosira oceanica (CCMP1003) in response to low Cu, at the physiological and proteomic level. The restructuring results in a shift from light harvesting for photochemistry-and ultimately for carbon fixation-to photoprotection, reducing carbon fixation and oxygen evolution. The observed decreases in the physiological parameters Fv/Fm, carbon fixation, and oxygen evolution, concomitant with increases in the antennae absorption cross section (σPSII), non-photochemical quenching (NPQ) and the conversion factor (φe:C/ηPSII) are in agreement with well documented cellular responses to low Fe. However, the underlying proteomic changes due to low Cu are very different from those elicited by low Fe. Low Cu induces a significant four-fold reduction in the Cu-containing photosynthetic electron carrier plastocyanin. The decrease in plastocyanin causes a bottleneck within the photosynthetic electron transport chain (ETC), ultimately leading to substantial stoichiometric changes. Namely, 2-fold reduction in both cytochrome b6f complex (cytb6f) and photosystem II (PSII), no change in the Fe-rich PSI and a 40- and 2-fold increase in proteins potentially involved in detoxification of reactive oxygen species (ferredoxin and ferredoxin:NADP+ reductase, respectively). Furthermore, we identify 48 light harvesting complex (LHC) proteins in the publicly available genome of T. oceanica and provide proteomic evidence for 33 of these. The change in the LHC composition within the antennae in response to low Cu underlines the shift from photochemistry to photoprotection in T. oceanica (CCMP1003). Interestingly, we also reveal very significant intra-specific strain differences. Another strain of T. oceanica (CCMP 1005) requires significantly higher Cu concentrations to sustain both its maximal and minimal growth rate compared to CCMP 1003. Under low Cu, CCMP 1005 decreases its growth rate, cell size, Chla and total protein per cell. We argue that the reduction in protein per cell is the main strategy to decrease its cellular Cu requirement, as none of the other parameters tested are affected. Differences between the two strains, as well as differences between the well documented responses to low Fe and those presented here in response to low Cu are discussed

Evolution and metabolic significance of the urea cycle in photosynthetic diatoms

Diatoms dominate the biomass of phytoplankton in nutrient-rich conditions and form the basis of some of the world’s most productive marine food webs1,2,3,4. The diatom nuclear genome contains genes with bacterial and plastid origins as well as genes of the secondary endosymbiotic host (the exosymbiont5)1,6,7,8,9,10, yet little is known about the relative contribution of each gene group to diatom metabolism. Here we show that the exosymbiont-derived ornithine-urea cycle, which is similar to that of metazoans but is absent in green algae and plants, facilitates rapid recovery from prolonged nitrogen limitation. RNA-interference-mediated knockdown of a mitochondrial carbamoyl phosphate synthase impairs the response of nitrogen-limited diatoms to nitrogen addition. Metabolomic analyses indicate that intermediates in the ornithine-urea cycle are particularly depleted and that both the tricarboxylic acid cycle and the glutamine synthetase/glutamate synthase cycles are linked directly with the ornithine-urea cycle. Several other depleted metabolites are generated from ornithine-urea cycle intermediates by the products of genes laterally acquired from bacteria. This metabolic coupling of bacterial- and exosymbiont-derived proteins seems to be fundamental to diatom physiology because the compounds affected include the major diatom osmolyte proline12 and the precursors for long-chain polyamines required for silica precipitation during cell wall formation11. So far, the ornithine-urea cycle is only known for its essential role in the removal of fixed nitrogen in metazoans. In diatoms, this cycle serves as a distribution and repackaging hub for inorganic carbon and nitrogen and contributes significantly to the metabolic response of diatoms to episodic nitrogen availability. The diatom ornithine-urea cycle therefore represents a key pathway for anaplerotic carbon fixation into nitrogenous compounds that are essential for diatom growth and for the contribution of diatoms to marine productivity