Publications by authors named "Andrew S Mount"

12 Publications

  • Page 1 of 1

Chitin is a functional component of the larval adhesive of barnacles.

Commun Biol 2020 01 17;3(1):31. Epub 2020 Jan 17.

Biological Sciences, Clemson University, Clemson, SC, 29634, USA.

Barnacles are the only sessile crustaceans, and their larva, the cyprid, is supremely adapted for attachment to surfaces. Barnacles have a universal requirement for strong adhesion at the point of larval attachment. Selective pressure on the cyprid adhesive has been intense and led to evolution of a tenacious and versatile natural glue. Here we provide evidence that carbohydrate polymers in the form of chitin provide stability to the cyprid adhesive of Balanus amphitrite. Chitin was identified surrounding lipid-rich vesicles in the cyprid cement glands. The functional role of chitin was demonstrated via removal of freshly attached cyprids from surfaces using a chitinase. Proteomic analysis identified a single cement gland-specific protein via its association with chitin and localized this protein to the same vesicles. The role of chitin in cyprid adhesion raises intriguing questions about the evolution of barnacle adhesion, as well as providing a new target for antifouling technologies.
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http://dx.doi.org/10.1038/s42003-020-0751-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6969031PMC
January 2020

Characterization of Calcification Events Using Live Optical and Electron Microscopy Techniques in a Marine Tubeworm.

J Vis Exp 2017 02 28(120). Epub 2017 Feb 28.

Department of Biological Sciences, Clemson University;

Characterizing the first event of biological production of calcium carbonate requires a combination of microscopy approaches. First, intracellular pH distribution and calcium ions can be observed using live microscopy over time. This allows identification of the life stage and the tissue with the feature of interest for further electron microscopy studies. Life stage and tissues of interest are typically higher in pH and Ca signals. Here, using H. elegans, we present a protocol to characterize the presence of calcium carbonate structures in a biological specimen on the scanning electron microscope (SEM), using energy-dispersive X-ray spectroscopy (EDS) to visualize elemental composition, using electron backscatter diffraction (EBSD) to determine the presence of crystalline structures, and using transmission electron microscopy (TEM) to analyze the composition and structure of the material. In this protocol, a focused ion beam (FIB) is used to isolate samples with dimension suitable for TEM analysis. As FIB is a site specific technique, we demonstrate how information from the previous techniques can be used to identify the region of interest, where Ca signals are highest.
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http://dx.doi.org/10.3791/55164DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5408784PMC
February 2017

Barnacle biology before, during and after settlement and metamorphosis: a study of the interface.

J Exp Biol 2017 01 3;220(Pt 2):194-207. Epub 2016 Nov 3.

Duke University Marine Laboratory, Marine Science and Conservation, 135 Duke Marine Lab Road, Beaufort, NC 28516, USA.

Mobile barnacle cypris larvae settle and metamorphose, transitioning to sessile juveniles with morphology and growth similar to that of adults. Because biofilms exist on immersed surfaces on which they attach, barnacles must interact with bacteria during initial attachment and subsequent growth. The objective of this study was to characterize the developing interface of the barnacle and substratum during this key developmental transition to inform potential mechanisms that promote attachment. The interface was characterized using confocal microscopy and fluorescent dyes to identify morphological and chemical changes to the interface and the status of bacteria present as a function of barnacle developmental stage. Staining revealed patchy material containing proteins and nucleic acids, reactive oxygen species amidst developing cuticle, and changes in bacteria viability at the developing interface. We found that as barnacles metamorphose from the cyprid to juvenile stage, proteinaceous materials with the appearance of coagulated liquid were released into and remained at the interface. It stained positive for proteins, including phosphoprotein, as well as nucleic acids. Regions of the developing cuticle and the patchy material itself stained for reactive oxygen species. Bacteria were absent until the cyprid was firmly attached, but populations died as barnacle development progressed. The oxidative environment may contribute to the cytotoxicity observed for bacteria and has the potential for oxidative crosslinking of cuticle and proteinaceous materials at the interface.
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http://dx.doi.org/10.1242/jeb.145094DOI Listing
January 2017

Synergistic roles for lipids and proteins in the permanent adhesive of barnacle larvae.

Nat Commun 2014 Jul 11;5:4414. Epub 2014 Jul 11.

Biological Sciences, Clemson University, 132 Long Hall, Clemson, South Carolina 29634, USA.

Thoracian barnacles rely heavily upon their ability to adhere to surfaces and are environmentally and economically important as biofouling pests. Their adhesives have unique attributes that define them as targets for bio-inspired adhesive development. With the aid of multi-photon and broadband coherent anti-Stokes Raman scattering microscopies, we report that the larval adhesive of barnacle cyprids is a bi-phasic system containing lipids and phosphoproteins, working synergistically to maximize adhesion to diverse surfaces under hostile conditions. Lipids, secreted first, possibly displace water from the surface interface creating a conducive environment for introduction of phosphoproteins while simultaneously modulating the spreading of the protein phase and protecting the nascent adhesive plaque from bacterial biodegradation. The two distinct phases are contained within two different granules in the cyprid cement glands, implying far greater complexity than previously recognized. Knowledge of the lipidic contribution will hopefully inspire development of novel synthetic bioadhesives and environmentally benign antifouling coatings.
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http://dx.doi.org/10.1038/ncomms5414DOI Listing
July 2014

Noradrenaline-functionalized hyperbranched fluoropolymer-poly(ethylene glycol) cross-linked networks as dual-mode, anti-biofouling coatings.

ACS Nano 2012 Feb 25;6(2):1503-12. Epub 2012 Jan 25.

Department of Chemistry, Texas A&M University, College Station, Texas 77842, United States.

The strategy of decorating antibiofouling hyperbranched fluoropolymer-poly(ethylene glycol) (HBFP-PEG) networks with a settlement sensory deterrent, noradrenaline (NA), and the results of biofouling assays are presented. This example of a dual-mode surface, which combines both passive and active modes of antibiofouling, works in synergy to improve the overall antibiofouling efficiency against barnacle cyprids. The HBFP-PEG polymer surface, prior to modification with NA, was analyzed by atomic force microscopy, and a significant distribution of topographical features was observed, with a nanoscopic roughness measurement of 110 ± 8 nm. NA attachment to the surface was probed by secondary ion mass spectrometry to quantify the extent of polymer chain-end substitution with NA, where a 3- to 4-fold increase in intensity for a fragment ion associated with NA was observed and 39% of the available sites for attachment were substituted. Cytoskeletal assays confirmed the activity of tethered NA on adhering oyster hemocytes. Settlement assays showed deterrence toward barnacle cyprid settlement, while not compromising the passive biofouling resistance of the surface. This robust strategy demonstrates a methodology for the incorporation of actively antibiofouling moieties onto a passively antibiofouling network.
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http://dx.doi.org/10.1021/nn204431mDOI Listing
February 2012

Toxicity of aqueous C70-gallic acid suspension in Daphnia magna.

Environ Toxicol Chem 2012 Jan 23;31(1):215-20. Epub 2011 Nov 23.

Institute of Environmental Toxicology, Clemson University, Pendleton, South Carolina, USA.

The present study assessed the toxic effects of stable aqueous colloidal suspensions of gallic-acid-stabilized C(70) fullerene on Daphnia magna. The suspensions were stabilized through noncovalent surface modification with gallic acid. In addition to whole-organism responses, changes in antioxidative processes in D. magna were quantified. Acute toxicity was observed with 96LC50 for C(70) -gallic acid of 0.4 ± 0.1 mg/L C(70) . Daphnia magna fecundity was significantly reduced in 21-d bioassays at C(70) -gallic aqcid concentrations below quantifiable limits. Antioxidant enzyme activities of glutathione peroxidase and superoxide dismutase as well as lipid peroxidation suggested that exposed organisms experienced oxidative stress. Microscopic techniques used to determine cellular toxicity via apoptosis proved unsuccessful.
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http://dx.doi.org/10.1002/etc.727DOI Listing
January 2012

Electrochemical characterization of a bioceramic material: The shell of the Eastern oyster Crassostrea virginica.

Bioelectrochemistry 2011 Jun 16;81(2):91-8. Epub 2011 Apr 16.

Nonstructural Materials Division, University of Dayton Research Institute, University of Dayton, Dayton, OH 45469, USA.

The shell of the Eastern oyster (Crassostrea virginica) is composed of multiple incongruent mineralized layers. This bioceramic composite material was investigated to determine the effects of shell thickness, orientation and layer composition on its electrochemical behavior using electrochemical impedance spectroscopy, potentiodynamic polarization and scanning electron microscopy-energy dispersive spectroscopy. SEM-EDS analysis of the oyster shell revealed that the multilayered biocomposite material is composed of calcium carbonate (CaCO(3)). EIS measurements in 3.5wt.% NaCl indicated that the impedance of the whole oyster shell in the low frequency region exhibited high impedance values which exhibited a decreasing trend with increasing immersion time. In terms of overall shell thickness, limiting currents measured by potentiodynamic techniques through the shell were observed to increase when the outer layers of the shell were sequentially removed by grinding, thus decreasing the shell thickness. These limiting current values remained relatively constant when the inner layers of the shell were removed. The impedance values of the oyster shell material as measured by EIS were shown to decrease with decreasing shell thickness. These findings suggest that the prismatic (outermost) shell layer in combination with the soluble organic matrix between all shell layers may influence the ionic conductivity through the oyster shell.
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http://dx.doi.org/10.1016/j.bioelechem.2011.04.002DOI Listing
June 2011

Visualization of shell matrix proteins in hemocytes and tissues of the Eastern oyster, Crassostrea virginica.

J Exp Zool B Mol Dev Evol 2008 May;310(3):227-39

Department of Biological Sciences, Clemson University, Clemson, South Carolina, USA.

The tissues of the oyster were examined for the presence of shell matrix proteins (SMPs) using a combination of Western, proteomic, and epi-fluorescent microscopy techniques. SMP, including 48 and 55 kDa phosphoproteins, was detected in the epithelial cells of mantle, gill, heart, and adductor muscle and linings of arteries and veins. The 48 kDa SMP circulates continuously within the hemolymph, and is present in the immune system hemocytes. It appears to be secreted from hemocytes on induction of shell repair. We suggest that the 48 and 55 kDa proteins are multifunctional and bridge the process of soft tissue repair and shell formation by mediating cellular activities during immune response as well as interacting with the mineral phase during deposition.
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http://dx.doi.org/10.1002/jez.b.21206DOI Listing
May 2008

In vivo biomodification of lipid-coated carbon nanotubes by Daphnia magna.

Environ Sci Technol 2007 Apr;41(8):3025-9

Department of Biological Sciences, University of North Texas, Denton, Texas 76203, USA.

This study examined the interactions between Daphnia magna and a water-soluble, lysophophatidylcholine coated single-walled carbon nanotube. D. magna were able to ingest the nanotubes through normal feeding behavior and utilize the lysophophatidylcholine coating as a food source. D. magna were able to modify the solubility of the nanotube, likely through digestion of the lipid coating. This study provides evidence of biomodification of a carbon-based nanomaterial by an aquatic organism. The modification significantly altered the physical properties of the nanomaterial in freshwater. Acute toxicity was observed only in the highest test concentrations. These are important findings related to determining the behavior and potential toxicity of coated nanomaterials released into the environment.
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http://dx.doi.org/10.1021/es062572aDOI Listing
April 2007

Translocation of C60 and its derivatives across a lipid bilayer.

Nano Lett 2007 Mar 23;7(3):614-9. Epub 2007 Feb 23.

Department of Mechanical Engineering, Clemson University, Clemson, South Carolina 29634, USA.

Obtaining an understanding, at the atomic level, of the interaction of nanomaterials with biological systems has recently become an issue of great research interest. Here we report on the molecular dynamics study of the translocation of fullerene C60 and its derivative C60(OH)20 across a model cell membrane (dipalmitoylphosphatidylcholine or DPPC bilayer). The simulation results indicate that, although a pristine C60 molecule can readily "jump" into the bilayer and translocate the membrane within a few milliseconds, the C60(OH)20 molecule can barely penetrate the bilayer. Indeed, the mean translocation time via diffusion for the C60(OH)20 molecule is several orders of magnitude longer than for the former. It was also determined that the two different forms of fullerenes, when adsorbed into/onto the bilayer, affected the membrane structure differently. This study offers a mechanistic explanation of that difference and for the reduced acute toxicity of functionalized fullerenes.
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http://dx.doi.org/10.1021/nl062515fDOI Listing
March 2007

Coating single-walled carbon nanotubes with phospholipids.

J Phys Chem B 2006 Feb;110(6):2475-8

Single-walled carbon nanotubes (SWNTs), being hydrophobic by nature, aggregate in water to form large bundles. However, isolated SWNTs possess unique physical and chemical properties that are desirable for sensing and biological applications. Conventionally isolated SWNTs can be obtained by wrapping the tubes with biopolymers or surfactants. The binding modes proposed for these solubilization schemes, however, are less than comprehensive. Here we characterize the efficacies of solubilizing SWNTs through various types of phospholipids and other amphiphilic surfactants. Specifically, we demonstrate that lysophospholipids, or single-chained phospholipids offer unprecedented solubility for SWNTs, while double-chained phospholipids are ineffective in rendering SWNTs soluble. Using transmission electron microscopy (TEM) we show that lysophospholipids wrap SWNTs as striations whose size and regularity are affected by the polarity of the lysophospholipids. We further show that wrapping is only observed when SWNTs are in the lipid phase and not the vacuum phase, suggesting that the environment has a pertinent role in the binding process. Our findings shed light on the debate over the binding mechanism of amphiphilic polymers and cylindrical nanostructures and have implications on the design of novel supramolecular complexes and nanodevices.
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http://dx.doi.org/10.1021/jp057252cDOI Listing
February 2006

Hemocyte-mediated shell mineralization in the eastern oyster.

Science 2004 Apr;304(5668):297-300

Department of Biological Sciences, Clemson University, Clemson, SC 29634, USA.

The growth of molluscan shell crystals is usually thought to be initiated from solution by extracellular organic matrix. We report a class of granulocytic hemocytes that may be directly involved in shell crystal production for oysters. On the basis of scanning electron microscopy (SEM) and x-ray microanalysis, these granulocytes contain calcium carbonate crystals, and they increase in abundance relative to other hemocytes following experimentally induced shell regeneration. Hemocytes are observed at the mineralization front using vital fluorescent staining and SEM. Some cells are observed releasing crystals that are subsequently remodeled, thereby at least augmenting matrix-mediated crystal-forming processes in this system.
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http://dx.doi.org/10.1126/science.1090506DOI Listing
April 2004