Fluorescent Labeling of Helminth Extracellular Vesicles Using an In Vivo Whole Organism Approach

In the last two decades, extracellular vesicles (EVs) from the three domains of life, Archaea, Bacteria and Eukaryotes, have gained increasing scientific attention. As such, the role of EVs in host-pathogen communication and immune modulation are being intensely investigated. Pivotal to EV research is the determination of how and where EVs are taken up by recipient cells and organs in vivo, which requires suitable tracking strategies including labelling. Labelling of EVs is often performed post-isolation which increases risks of non-specific labelling and the introduction of labelling artefacts. Here we exploited the inability of helminths to de novo synthesise fatty acids to enable labelling of EVs by whole organism uptake of fluorescent lipid analogues and the subsequent incorporation in EVs. We showed uptake of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (DOPE-Rho) in Anisakis spp. and Trichuris suis larvae. EVs isolated from the supernatant of Anisakis spp. labelled with DOPE-Rho were characterised to assess the effects of labelling on size, structure and fluorescence of EVs. Fluorescent EVs were successfully taken up by the human macrophage cell line THP-1. This study, therefore, presents a novel staining method that can be utilized by the EV field in parasitology and potentially across multiple species

Condensation of DNA using poly(amido amine) dendrimers: effect of salt concentration on aggregate morphology

The condensation of DNA and poly(amido amine) dendrimers of generation 1, 2, 4, 6, and 8 has been studied as a function of salt concentration in order to reveal the forces that control the aggregate size and morphology. For the lower generation dendrimers (1, 2, and 4) a dramatic increase in aggregate size occurs as a result of an increase in salt concentration. Toroidal aggregates having an outer diameter of up to several hundreds of nm are observed. For the higher generation 6 dendrimers, the size of the condensed DNA aggregates does not change, however, an alteration in morphology is seen at high salt concentration, as more rod-like aggregates are observed. The size and morphology of generation 8 dendrimers are seemingly insensitive to salt concentration. It is believed that the effective neutralisation of the dendrimer and DNA charge in the aggregate is the reason for the observed effects. It is further shown that the 2D hexagonal lattice spacing observed in toroids is close to constant irrespective of the size of the cation responsible for the DNA condensation

Morphology: An Ambiguous Indicator of Biogenicity

An integrated analysis of both airborne and field short-wave infrared hyperspectral measurements was used in conjunction with conventional field mapping techniques to map hydrothermal alteration in the central portion of the Mount Painter Inlier in the Flinders Ranges, South Australia. The airborne hyperspectral data show the spatial distribution of spectrally distinct minerals occurring as primary minerals and as weathering and alteration products. Field spectral measurements, taken with a portable infrared mineral analyzer spectrometer and supported by thin-section analyses, were used to verify the mineral maps and enhance the level of information obtainable from the airborne data. Hydrothermal alteration zones were identified and mapped separately from the background weathering signals. A main zone of alteration, coinciding with the Paralana Fault zone, was recognized, and found to contain kaolinite, muscovite, biotite, and K-feldspar. A small spectral variation associated with a ring-like feature around Mount Painter was tentatively determined to be halloysite and interpreted to represent a separate hydrothermal fluid and fluid source, and probably a separate system. The older parts of the alteration system are tentatively dated as Permo-Carboniferous. The remote sensing of alteration at Mount Painter confirms that hyperspectral imaging techniques can produce accurate mineralogical maps with significant details that can be used to identify and map hydrothermal activity. Application of hyperspectral surveys such as that conducted at Mount Painter would be likely to provide similar detail about putative hydrothermal deposits on Mars

Phase Behavior and Coassembly of DNA and Lysozyme in Dilute Aqueous Mixtures: A Model Investigation of DNA-Protein Interactions

Results from an experimental investigation of the phase behavior of an aqueous system of DNA from salmon testes and the protein lysozyme arc presented. At very low concentrations of either or both of the macromolecular components, wormlike assemblies with a width of the order of 10 nm are formed. There are strong indications that direct interactions between the protein units are instrumental both in driving the phase separation and in controlling the morphology of the formed assemblies