Intracellular bioaccumulation of the rare earth element Gadolinium in ciliate cells resulting in biogenic particle formation and excretion.

Ciliates are abundant unicellular organisms capable of resisting high concentrations of metal ions in the environment caused by various anthropogenic activities. Understanding the cellular pathways involved in resistance to and detoxification of elements is required to predict the impact of ciliates on environmental element cycles. Here, we investigated the so far unknown process of tolerance, cellular uptake and bioaccumulation of the emerging rare earth element gadolinium (Gd) in the common ciliate Tetrahymena pyriformis. Gd treatment results in the intracellular formation and excretion of biogenic Gd-containing particles. This cellular process effectively removes dissolved Gd from the organic growth medium by 53.37% within 72 h. Based on light and electron microscopic observations, we postulate a detoxification pathway: Cells take up toxic Gd3+ ions from the medium by endocytosis, process them into stable Gd-containing particles within food vacuoles, and exocytose them. Stable biogenic particles can be isolated, which are relatively homogeneous and have a diameter of about 3 µm. They consist of the elements Gd, C, O, P, Na, Mg, K, and Ca. These findings broaden the view of metal ion accumulation by protists and are of relevance to understand environmental elemental cycles and may inspire approaches for metal recovery or bioremediation.

Mineralized scale patterns on the cell periphery of the chrysophyte Mallomonas determined by comparative 3D Cryo-FIB SEM data processing.

Unicellular protists can biomineralize spatially complex and functional shells. A typical cell of the photosynthetic synurophyte Mallomonas is covered by about 60-100 silica scales. Their geometric arrangement, the so-called scale case, mainly depends on the species and on the cell cycle. In this study, the scale case of the synurophyte Mallomonas was preserved in aqueous suspension using high-pressure freezing (HPF). From this specimen, a three-dimensional (3D) data set spanning a volume of about 25.6 µm × 19.2 µm × 4.2 µm with a voxel size of 12.5 nm × 12.5 nm × 25.0 nm was collected by Cryo-FIB SEM in 3 h and 24 min. SEM imaging using In-lens SE detection allowed to clearly differentiate between mineralized, curved scales of less than 0.2 µm thickness and organic cellular ultrastructure or vitrified ice. The three-dimensional spatial orientations and shapes of a minimum set of scales (N = 13) were identified by visual inspection, and manually segmented. Manual and automated segmentation approaches were comparatively applied to one arbitrarily selected reference scale using the differences in grey level between scales and other constituents. Computational automated routines and principal component analysis of the experimentally extracted data created a realistic mathematical model based on the Fibonacci pattern theory. A complete in silico scale case of Mallomonas was reconstructed showing an optimized scale coverage on the cell surface, similarly as it was observed experimentally. The minimum time requirements from harvesting the living cells to the final scale case determination by Cryo-FIB SEM and computational image processing are discussed.

Heavy metal mobility and valuable contents of processed municipal solid waste incineration residues from Southwestern Germany.

As conventional end-of-life disposal, municipal solid waste (MSW) incineration residues can be problematic due to potential release of toxic compounds into the environment. Using municipal solid waste incineration residues as urban-mine of valuable metals (e.g. precious metals) could provide a trash-to-treasure possibility. The objectives of the study are to (i) determine the contents of different contaminant metallic elements (Zn, Cu, Ba, Pb, Cr and Ni) in four size fractions of MSW incineration residues and discuss their mobility potential by using the modified BCR sequential extraction method; (ii) investigate the level of valuable critical contents (precious metals, rare earth elements, etc.) in these wastes. We also characterized mineralogy and elemental composition of four different grain size fractions (0-0.5, 0.5-2.0, 2.0-4.0 and 4.0-16.0 mm) of processed municipal solid waste incineration residue (PIR) from the Southwestern region of Germany, using X-ray fluorescence, X-ray powder diffraction and different spectroscopic techniques. Among all studied size fractions, grains smaller than 2 mm contained higher amounts of total extractable heavy metals in most cases. The most important finding of the study is that the total contents of Cu, Au and Pt in the incineration residues reached economically profitable levels (5.1 g/kg, 21.69 mg/kg and 17.45 mg/kg, respectively).

Aqueous ball milling of nacre constituents facilitates directional self-assembly of aragonite nanoparticles of the gastropod Haliotis glabra.

A ball-milling approach was developed to investigate the constituents of isolated nacre tablets of the gastropod Haliotis glabra in aqueous suspension without additional chemical additives. The obtained particle mixtures were characterized using X-ray crystallography as well as scanning and transmission electron microscopy. Aragonite nanoparticles retained their crystal structure even after 14 h of ball milling. The long-term stability of the particle mixtures varied as a function of the ball-milling duration. An increased milling time led to rod-like stable assemblies of aragonite nanoparticles. Selected area electron diffraction investigations revealed that the longitudinal axes in about one-third of these nanoparticle rods were oriented along the crystallographic c-axis of aragonite, indicating oriented attachment of the aragonite nanoparticles. These in vitro observations support the idea that a two-stage process, separated into crystallization of nanoparticles and oriented assembly of nanocrystals, could also occur in vivo.

Biogenic and Synthetic Peptides with Oppositely Charged Amino Acids as Binding Sites for Mineralization.

Proteins regulate diverse biological processes by the specific interaction with, e.g., nucleic acids, proteins and inorganic molecules. The generation of inorganic hybrid materials, such as shell formation in mollusks, is a protein-controlled mineralization process. Moreover, inorganic-binding peptides are attractive for the bioinspired mineralization of non-natural inorganic functional materials for technical applications. However, it is still challenging to identify mineral-binding peptide motifs from biological systems as well as for technical systems. Here, three complementary approaches were combined to analyze protein motifs consisting of alternating positively and negatively charged amino acids: (i) the screening of natural biomineralization proteins; (ii) the selection of inorganic-binding peptides derived from phage display; and (iii) the mineralization of tobacco mosaic virus (TMV)-based templates. A respective peptide motif displayed on the TMV surface had a major impact on the SiO2 mineralization. In addition, similar motifs were found in zinc oxide- and zirconia-binding peptides indicating a general binding feature. The comparative analysis presented here raises new questions regarding whether or not there is a common design principle based on acidic and basic amino acids for peptides interacting with minerals.

Silver nanoparticle toxicity in sea urchin Paracentrotus lividus.

Silver nanoparticles (AgNPS) are an important model system for studying potential environmental risks posed by the use of nanomaterials. So far there is no consensus as to whether toxicity is due to AgNPs themselves or Ag(+) ions leaching from their surfaces. In sea urchin Paracentrotus lividus, AgNPs cause dose dependent developmental defects such as delayed development, bodily asymmetry and shortened or irregular arms, as well as behavioural changes, particularly in swimming patterns, at concentration ∼0.3 mg/L AgNPs. It has been observed that AgNPs are more toxic than their equivalent Ag(+) ion dose.

Isolation of alveolar plates from Coleps hirtus.

In the ciliate Coleps hirtus, the alveoli contain rigid alveolar plates that are almost unstudied so far. Neither the exact composition nor the genesis and function are known. A necessary step to study the alveolar plates is to isolate these structures in an adequate amount. Therefore, culture conditions of C. hirtus were optimized to obtain an axenic and dense long-time culture. The protocol we developed to isolate C. hirtus alveolar plates is presented and clean alveolar plates were documented via scanning electron microscopy. The described procedure delivers alveolar plates of very good structure and integrity with preserved filigree details in sufficient amount. They can be analysed via a range of different material and biological characterisations. Since there are indications of a mineral phase within the alveolar plates, the presented results will allow to study C. hirtus alveolar plates also in the context of biomineralisation.

Genesis of amorphous calcium carbonate containing alveolar plates in the ciliate Coleps hirtus (Ciliophora, Prostomatea).

In the protist world, the ciliate Coleps hirtus (phylum Ciliophora, class Prostomatea) synthesizes a peculiar biomineralized test made of alveolar plates, structures located within alveolar vesicles at the cell cortex. Alveolar plates are arranged by overlapping like an armor and they are thought to protect and/or stiffen the cell. Although their morphology is species-specific and of complex architecture, so far almost nothing is known about their genesis, their structure and their elemental and mineral composition. We investigated the genesis of new alveolar plates after cell division and examined cells and isolated alveolar plates by electron microscopy, energy-dispersive X-ray spectroscopy, FTIR and X-ray diffraction. Our investigations revealed an organic mesh-like structure that guides the formation of new alveolar plates like a template and the role of vesicles transporting inorganic material. We further demonstrated that the inorganic part of the alveolar plates is composed out of amorphous calcium carbonate. For stabilization of the amorphous phase, the alveolar vesicles, the organic fraction and the element phosphorus may play a role.

Diversity and abundance of photosynthetic sponges in temperate Western Australia.

BACKGROUND: Photosynthetic sponges are important components of reef ecosystems around the world, but are poorly understood. It is often assumed that temperate regions have low diversity and abundance of photosynthetic sponges, but to date no studies have investigated this question. The aim of this study was to compare the percentages of photosynthetic sponges in temperate Western Australia (WA) with previously published data on tropical regions, and to determine the abundance and diversity of these associations in a range of temperate environments. RESULTS: We sampled sponges on 5 m belt transects to determine the percentage of photosynthetic sponges and identified at least one representative of each group of symbionts using 16S rDNA sequencing together with microscopy techniques. Our results demonstrate that photosynthetic sponges are abundant in temperate WA, with an average of 63% of sponge individuals hosting high levels of photosynthetic symbionts and 11% with low to medium levels. These percentages of photosynthetic sponges are comparable to those found on tropical reefs and may have important implications for ecosystem function on temperate reefs in other areas of the world. A diverse range of symbionts sometimes occurred within a small geographic area, including the three "big" cyanobacterial clades, Oscillatoria spongeliae, "Candidatus Synechococcus spongiarum" and Synechocystis species, and it appears that these clades all occur in a wide range of sponges. Additionally, spongin-permeating red algae occurred in at least 7 sponge species. This study provides the first investigation of the molecular phylogeny of rhodophyte symbionts in sponges. CONCLUSION: Photosynthetic sponges are abundant and diverse in temperate WA, with comparable percentages of photosynthetic to non-photosynthetic sponges to tropical zones. It appears that there are three common generalist clades of cyanobacterial symbionts of sponges which occur in a wide range of sponges in a wide range of environmental conditions.