Examining the inorganic elemental composition of lobster phyllosoma (Panulirus ornatus) with X-ray fluorescence microscopy.

The ornate spiny rock lobster, Panulirus ornatus, is an attractive candidate for aquaculture. The larval stages of spiny lobsters, known as phyllosoma, are complex with many developmental stages. Very little is known about the inorganic element composition of phyllosoma. In this study, a novel method using synchrotron X-ray fluorescence microscopy (XFM) was applied to investigate the distributions of metals potassium (K), calcium (Ca), copper (Cu), zinc (Zn), the metalloid arsenic (As), and nonmetal bromine (Br) within individual phyllosoma at stages 3, 4, and 8 of their development. For the first time, 1 µm resolution synchrotron XFM images of whole phyllosoma as well as closer examinations of their eyes, mouths, setae, and tails were obtained. Elements accumulated in certain locations within phyllosoma, providing insight into their likely biological role for these organisms. This information may be useful for the application of dietary supplementation in the future to closed larval cycle lobster aquaculture operations.

Natural heavy metal concentrations in seawater as a possible cause of low survival of larval mussels.

BACKGROUND: A period of seismic activity starting in 2010 coincided with a decline in commercial catches of wild seed mussels in a major aquaculture production region of New Zealand. Analyses of over 40 years of mussel seed catch data from in the Pelorus and Kenepuru Sounds, confirmed a marked decline since 2010 in catches of the preferred, green-lipped mussel (Perna canaliculus), the larvae of which is known to have low tolerance of heavy metals in seawater. METHODS: Heavy metal mean concentrations were measured throughout the Pelorus and Kenepuru Sounds. The concentrations ranged from < 0.60-3.24, < 16.94-74.35, < 1.47-4.00, 2.23-19.02, 1.86-3.29 and 0.12-0.52 µg L-1 for Cr, Fe, Cu, Zn, As, and Cd, respectively. Seawater from six locations in the Sounds, historically associated with high commercial catches of settling mussel larvae, was used for experimental rearing of green-lipped mussel larvae. RESULTS: No mussel embryos survived when incubated in these seawater samples. The mean concentrations of Cr, Fe, As, and Cd were significantly higher in the seawater from the Sounds than in the hatchery seawater. A higher concentration of one or a combination of these heavy metals could be the cause of the poor larval survival. These findings could be crucial for the sustainability of mussel farming in the area.

Balancing essential and non-essential metal bioavailability during hatchery rearing of Greenshell mussel (Perna canaliculus) larvae.

The use of ethylenediaminetetraacetic acid (EDTA) during bivalve hatchery production is thought to improve larval yields due to the reduced exposure to toxic metals (such as Cu); however, few studies have focused on the bioavailability of metals during the rearing process. Greenshell™ mussels (Perna canaliculus) were reared for 48 h with and without EDTA (12 µM) exposure and larvae were subsequently raised to 21 days post-fertilisation with and without EDTA exposure. Survival, shell length, algal ingestion rate, swimming activity, total metal concentration in water, bioavailable metal concentrations and larval metal accumulation were monitored for the 21 day period. Larval fitness (specifically D-yields) was improved on day 2 in the EDTA treatment, whereas an overall negative effect of EDTA treatment on fitness was observed on day 10 and 21. During the first 48 h, increased survival in the EDTA treatment is believed to be due to the reduction of bioavailable Zn concentrations in the rearing seawater. No other metal (essential or non-essential) displayed a consistent trend when comparing metal bioavailability to any of the fitness parameters measured throughout the experiment. Though the measured metal bioavailability was not clearly linked to fitness, the uptake of Al, P, Cr, Fe, Co, Ni, Zn, As, Cd, and Hg by P. canaliculus was reduced during the first 48 h, suggesting that the biological regulation of these elements is just as important as the bioavailability. Overall, treatment of the rearing seawater with 12 µM EDTA is effective for improving Greenshell™ mussel larval yields by decreasing metal bioavailability during the first two days of development but has minimal benefit between day 2 and 21.

Biodegradable chelating agent improves the survival of early larvae for shellfish aquaculture.

The toxicity of heavy metals commonly impacts the survival of crustacean and bivalve larvae in hatchery culture, and this has led to the widespread use of EDTA to decrease this toxicity. Since EDTA has a very poor biodegradability leading to potential persistent environmental effects, alternative methods to prevent heavy metal toxicity to shellfish larvae are needed. EDDS is a biodegradable potential alternative to EDTA for this application and was tested as a treatment of the seawater used for rearing aquaculture Greenshell™ mussel (Perna canaliculus) larval embryos in this study. Mussel embryos reared with EDTA or EDDS had significantly better survival than without. The concentrations and spatial distributions of heavy metals in D-veliger larvae as determined by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) and X-ray Fluorescence Microscopy (XFM) suggested that chelating agents increased the levels of calcium in larvae while they reduced the concentration of zinc. In addition, where decreased accumulation of the other heavy metals was not observed, chelating agents affected their distribution within the larvae, especially for copper and arsenic. This is the first study to test the use of EDDS for aquaculture hatchery application and shows that EDDS is an effective biodegradable alternative to EDTA that can mitigate the effects of heavy metals for shellfish larval rearing.

Structure of soft and hard protein corona around polystyrene nanoplastics-Particle size and protein types.

A major challenge in understanding nanoplastic toxicity (or nanoparticles in general) lies in establishing the causal relationships between its physical properties and biological impact. This difficulty can be attributed to surface alterations that follow the formation of a biological complex around the nanoplastic, as exemplified by protein coronae. The protein corona is known to be responsible for the biological response elicited, although its own structure and attributes remain unknown. We approach this knowledge gap by independently studying the structure of soft and hard coronae using neutron scattering techniques. We investigated the formation and the structure of corona proteins (human serum albumin and lysozyme) and the resulting protein corona complexes with polystyrene nanoplastics of different sizes (20 and 200 nm) and charges. Soft corona complexes (regardless of protein type) adopted a structure where the nanoplastics were surrounded by a loose protein layer (∼2-3 protein molecules thick). Hard corona complexes formed fractal-like aggregates, and the morphology of which is known to be harmful to cellular membranes. In most cases, hard-corona coated nanoplastics also formed fractal-like aggregates in solution. Nanoplastic size affected the structures of both the protein corona and the intrinsic protein: more significant conformational change was observed in the hard corona proteins around smaller nanoparticles compared to larger ones, as the self-association forces holding the nanoplastic/protein complex together were stronger. This also implies that protein-dependent biochemical processes are more likely to be disrupted by smaller polystyrene nanoplastics, rather than larger ones.

Examining the role of ethylenediaminetetraacetic acid (EDTA) in larval shellfish production in seawater contaminated with heavy metals.

Heavy metal pollution is a concern in many coastal locations where it is frequently deleterious to the survival of young shellfish. Consequently, a great number of commercial shellfish hatcheries around the world rely on the addition of ethylenediaminetetraacetic acid (EDTA) to seawater to ensure successful larval production. Despite the importance of this practice to global shellfish production the mode of action of EDTA in larval production remains undetermined. It is assumed EDTA chelates heavy metals in seawater preventing interference in larval development. Larval mussels (Perna canaliculus) raised in seawater with 3 µM EDTA had a 15 fold higher yield than those without EDTA. The concentration and spatial arrangement of heavy metals in larvae as determined by Inductively Coupled Plasma Mass Spectrometry (ICPMS) and X-ray Fluorescence Microscopy (XFM) was consistent with reduced bioavailability of several metals, especially copper and zinc. This is the first study to confirm the effectiveness of EDTA for managing metal pollution commonly encountered in coastal shellfish hatcheries.

Biomineralization of Calcium Phosphate and Calcium Carbonate within Iridescent Chitosan/Iota-Carrageenan Multilayered Films.

This work systematically explores the biomineralization of calcium phosphate (CaP) and carbonate (CaCO3) within chitosan/iota-carrageenan multilayer films. Multilayer films of chitosan and iota-carrageenan (up to 128-coupled layers) were prepared on glass substrates by a layer-by-layer dip-coating technique. Cryo-scanning electron microscopy revealed dense interfaces between the chitosan and iota-carrageenan layers with thicknesses in the range 250 and 350 nm in the hydrated state, accounting for the iridescent nature of multilayer films when wet. Immersion of the multilayered films in simulated body fluid or simulated seawater at 25 °C resulted in the mineralization of CaP and CaCO3, respectively, at the interfaces between the biopolymer layers and modified the iridescence of the films. Lamellar scattering features in small-angle neutron scattering measurements of the mineralized films provided evidence of the localized mineralization. Further evidence of this was found through the lack of change in the dynamic and static correlation lengths of the polymer networks within the bulk phase of the chitosan and iota-carrageenan layers. CaP mineralization occurred to a greater extent than CaCO3 mineralization within the films, evidenced by the higher lamellar density and greater rigidity of the CaP-mineralized films. Results provide valuable new insights into CaP and CaCO3 biomineralization in biopolymer networks.