Organ-specific distribution and size-dependent toxicity of polystyrene nanoplastics in Australian bass (Macquaria novemaculeata).

Micro- and nano-plastics (MNPs) are emerging contaminants found in air, water, and food. Ageing and weathering processes convert aquatic plastics into MNPs which, due to their small size, can be assimilated by organisms. The accumulation of MNPs in aquatic life (e.g., fish, oysters, and crabs) will, in turn, pose risks to the health of ecosystems and human. This study focuses on the uptake, biodistribution, and size-dependent toxicity of polystyrene nano-plastics (PS-NPs) in a commercially important food web, the Australian Bass (Macquaria novemaculeata). Fish were fed artemia containing PS-NPs of various sizes (ranging from 50 nm to 1 µm) for durations of 5 and 7 days. The findings revealed that smaller NPs (50 nm) accumulated in the brain and muscle tissues at higher concentrations, whereas larger NPs (1 µm) were primarily found in the gills and intestines. In addition, an inverse correlation was observed between the size of NPs and the rate of trophic transfer, with smaller PS-NPs resulting in a higher transfer rate from artemia to fish. Polystyrene NPs caused both activation of the enzyme superoxide dismutase and damage to the DNA of fish tissues. These effects were size dependent. Metabolomic analysis revealed that indirect exposure to different-sized PS-NPs resulted in altered metabolic profiles within fish intestines, potentially impacting lipid and energy metabolism. These results offer novel perspectives on the size-specific toxic impacts of NPs on fish and the transfer of plastics through the food chain.

Occurrence and distribution of microplastics in long-term biosolid-applied rehabilitation land: An overlooked pathway for microplastic entry into terrestrial ecosystems in Australia.

Wastewater treatment plants (WWTPs) efficiently eliminate over 98% of microplastics (MPs) from wastewater discharge, subsequently accumulating them in sludge. This sludge is frequently employed as fertilizer in agricultural practices or land rehabilitation. While there is significant research on biosolid application in agriculture, the discussion regarding its application in rehabilitating industrial zones and MPs contamination is limited. The current study investigates the abundance, distribution, and composition of MPs in rehabilitation land with long-term biosolid-application in Australia. Three minesite fields (designated 1-3), each with distinct biosolid application histories since 2011, 2012, and 2017, and a control field without any biosolid application history, were chosen for this study. The abundances of MPs in biosolid-applied fields 1-3 (6.04 ± 1.92 x 102 MP kg-1; 4.94 ± 0.73 x 102 MP kg-1; 2.48 ± 0.70 x 102 MP kg-1) were considerably higher compared to non-biosolid-applied field (0.70 ± 0.63 x 102 MP kg -1). This indicates that the application of biosolids significantly contributes to the presence of MPs in the soil. Moreover, the results suggest that with each successive application, the abundance of MPs increases. The abundance and size of MPs in both biosolid and non-biosolid soils decreased as the soil depth increased. Microbeads were dominant in soils where biosolids were applied (up to 61.9%), while fibres were dominant in non-biosolid soils (accounting for 85.7%). The distribution of plastic polymer types varied among fields and soil depths. Most MPs were microbeads of polyamide (PA), fragments of polyethylene (PE), foam of polystyrene (PS), and fibres of rayon. This research presents evidence that the extended utilization of biosolids results in elevated MP pollution in minesite rehabilitation land, highlighting a frequently overlooked origin of MP contamination in terrestrial settings. Additional evaluations needed to understand ecological risks of MPs in soil ecosystems affected by biosolid application.

Borophene Embedded Cellulose Paper for Enhanced Photothermal Water Evaporation and Prompt Bacterial Killing.

Solar-driven photothermal water evaporation is considered an elegant and sustainable technology for freshwater production. The existing systems, however, often suffer from poor stability and biofouling issues, which severely hamper their prospects in practical applications. Conventionally, photothermal materials are deposited on the membrane supports via vacuum-assisted filtration or dip-coating methods. Nevertheless, the weak inherent material-membrane interactions frequently lead to poor durability, and the photothermal material layer can be easily peeled off from the hosting substrates or partially dissolved when immersed in water. In the present article, the discovery of the incorporation of borophene into cellulose nanofibers (CNF), enabling excellent environmental stability with a high light-to-heat conversion efficiency of 91.5% and water evaporation rate of 1.45 kg m-2 h-1 under simulated sunlight is reported. It is also demonstrated that borophene papers can be employed as an excellent active photothermal material for eliminating almost 100% of both gram-positive and gram-negative bacteria within 20 min under three sun irradiations. The result opens a new direction for the design of borophene-based papers with unique photothermal properties which can be used for the effective treatment of a wide range of wastewaters.

Metal(loid) uptake and partitioning within the saltmarsh halophyte, Juncus kraussii.

An investigation was conducted over three estuaries in SE Australia with a gradient in metal(loid) contamination to assess metal(loid) (Cu, Zn, As, Se, Cd and Pb) accumulation and transport within the halophytic saltmarsh rush, Juncus kraussii. Sydney Olympic Park exhibited the most elevated metal(loid) contamination, followed by Hunter Wetlands and Lake Macquarie. J. kraussii exhibited a strong ability to restrict metal(loid) movement into the root system, with the exception of cadmium (BCFs < 1.0) and unrestricted flow from root to culm excepting Se, Cd (TFs < 1). Pb and Zn exhibited elevated translocation between roots and culms (TF 4.4 and 7.3, respectively). Despite barriers for uptake into the below-ground tissues, most metal(loid)s were accumulated to the roots with environmental dose (except for Cu and Cd) and linear relationships were present between the root and culm (for As and Se) and the sediment and culm (for As, Se, Cd, and Pb).

Secondary treatment phase of tertiary wastewater treatment works significantly reduces estrogenic load.

Estrogenic compounds enter waterways via effluents from wastewater treatment works (WWTW), thereby indicating a potential risk to organisms inhabiting adjacent receiving waters. However, little is known about the loads or concentrations of estrogenic compounds that enter Australian WWTWs, the efficiency of removing estrogenic compounds throughout the various stages of tertiary WWTW processes (which are common in Australia), nor the concentrations released into estuarine or marine receiving waters, and the associated risk for aquatic taxa residing in these environments. Therefore, seven estrogenic compounds, comprising the natural estrogens estrone (E1), 17ß-estradiol (E2) and estriol (E3), the synthetic estrogen (EE2), and the industrial chemicals bisphenol A (BPA), 4-t-octyl phenol (4-t-OP) and 4-nonyl phenol (4-NP), in wastewater samples were quantified via liquid chromatographic-mass spectrometry (LC-MS) after solid-phase extraction at different stages of wastewater treatment and associated receiving waters. The concentrations of the target compounds in wastewater ranged from < LOQ (limit of quantification) to 158 ng/L for Tanilba Bay WWTW and < LOQ to 162 ng/L for Belmont WWTW. Most target compounds significantly declined after the secondary treatment phase. Appreciable removal efficiency throughout the treatment process was observed with removal from 39.21 to 99.98% of influent values at both WWTWs. The reduction of the natural estrogens (E1, E2 and E3) and 4-t-OP were significantly greater than EE2, BPA, and 4-NP in both WWTWs. Risk quotients (RQs) were calculated to assess potential ecological risks from individual estrogenic compounds. In predicted diluted effluents, no targeted compounds showed any ecological risk (RQ ≤1.65 × 10-2) at both WWTWs. Similarly, all RQs for shore samples at both WWTWs were below 1. Finally, the hazard index (HI), which represents combined estrogenic contaminants' ecological risk, indicated no mentionable risk for predicted diluted effluents (HI = 0.0097 to 0.0218) as well as shoreline samples (HI = 0.393 to 0.522) in the receiving estuarine or marine waters.

Biofilms Enhance the Adsorption of Toxic Contaminants on Plastic Microfibers under Environmentally Relevant Conditions.

Microplastics (MPs) exposed to the natural environment provide an ideal surface for biofilm formation, which potentially acts as a reactive phase facilitating the sorption of hazardous contaminants. Until now, changes in the contaminant sorption capacity of MPs due to biofilm formation have not been quantified. This is the first study that compared the capacity of naturally aged, biofilm-covered microplastic fibers (BMFs) to adsorb perfluorooctane sulfonate (PFOS) and lead (Pb) at environmentally relevant concentrations. Changes in the surface properties and morphology of aged microplastic fibers (MF) were studied by surface area analysis, infrared spectroscopy, and scanning electron microscopy. Results revealed that aged MFs exhibited higher surface areas because of biomass accumulation compared to virgin samples and followed the order polypropylene>polyethylene>nylon>polyester. The concentrations of adsorbed Pb and PFOS were 4-25% and 20-85% higher in aged MFs and varied among the polymer types. The increased contaminant adsorption was linked with the altered surface area and the hydrophobic/hydrophilic characteristics of the samples. Overall, the present study demonstrates that biofilms play a decisive role in contaminant-plastic interactions and significantly enhance the vector potential of MFs for toxic environmental contaminants. We anticipate that knowledge generated from this study will help refine the planetary risk assessment of MPs.

Accumulation and distribution of metal(loid)s in the halophytic saltmarsh shrub, Austral seablite, Suaeda australis in New South Wales, Australia.

We examined the patterns of uptake and partitioning of metal(loid)s in Suaeda australis from three highly urbanised estuaries (Sydney Olympic Park, Hunter Wetlands and Lake Macquarie) in NSW, Australia. Of these, Sydney Olympic Park was found to be the most contaminated estuary in terms of combined sediment metal(loid) load, followed by Hunter Wetlands and lowest in Lake Macquarie (via PERMANOVA). Uptake in roots was greater for the essential metals Cu and Zn along with the non-essential metal Cd and the metalloid Se (root BCFs >1) and lower for Pb and As (root BCFs <1). Substantial barriers for translocation from roots to stems were identified for all metal(loid)s (stem TFs; 0.07-0.68). Conversely, unrestricted flow from stems to leaves was observed for all metal(loid)s at unity or higher (leaf TFs ≥ 1). Strong linear relationships between sediment and root for Zn and Pb were observed, indicating roots as a useful bioindicator.

Exposure to estrogenic mixtures results in tissue-specific alterations to the metabolome of oysters.

The current study investigated the effect of environmentally relevant mixtures of estrogens at levels representative of receiving waters on the metabolome of the Sydney rock oyster, Saccostrea glomerata. Oysters were exposed to a "low" and a "high" mixture of (xeno) estrogens (representative of Australian and global receiving waters respectively) for 7 days and digestive gland, gill, and gonad tissue were sampled for quantification of polar metabolites by 1H NMR spectroscopy. Exposure to both mixtures lowered body mass and altered the metabolite profile in the digestive glands. Comparatively, gills, and ovaries demonstrated lesser sensitivity to the mixtures, with significant metabolomic alterations observed only for the high mixture. The male gonad did not respond to either estrogenic exposure. In the responsive tissues, major metabolites including amino acids, carbohydrates, intermediates of the tricarboxylic acid cycle and ATP were all down-regulated and exhibited tissue-specific patterns of down-regulation with the greatest proportion of metabolites down-regulated due to estrogenic exposure in the digestive gland. Exposure to (xeno) estrogen mixtures representative of concentrations reported in receiving waters in Australia and globally can impact the metabolome and associated energy metabolism, especially in the digestive gland, translating to lower pools of available ATP energy for potential cellular homeostasis, somatic maintenance and growth, reproduction and fitness.

Quantitative biomonitoring of polycyclic aromatic compounds (PACs) using the Sydney rock oyster (Saccostrea glomerata).

Increasing our understanding of the bioavailable fractions of polycyclic aromatic compounds (PACs) in an aquatic environment is important for the assessment of the environmental and human health risks posed by PACs. More importantly, the behaviour of polar polycyclic aromatic hydrocarbons (polar PAHs), which are metabolites of legacy PAHs, are yet to be understood. We, therefore, carried out a study involving Sydney rock oysters (Saccostrea glomerata) sourced from two locations, that had been exposed to PAH contamination, within an Australian south-east estuary. Biomonitoring of these oysters, following relocation from the estuary to a relatively isolated waterway, was done at 24 and 72 h after deployment and subsequently at 7, 14, 28, 52 and 86 days. Control samples from Camden Haven River were sampled for PAC analyses just before deployment, after 28 days and at the end of the study (day 86). Lipid-normalised concentrations in oyster tissues across the 86-day sampling duration, elimination rate constants (k2), biological half-lives (t1/2) and time required to reach 95% of steady-state (t95) were reported for parent PAHs and the less-monitored polar PAHs including nitrated/oxygenated/heterocyclic PAHs (NPAHs, oxyPAHs and HPAHs) for the three differently sourced oyster types. Most of the depurating PAHs and NPAHs, as well as 9-FLO (oxyPAH), had k2 values significantly different from zero (p < 0.05). All other oxyPAHs and HPAHs showed no clear depuration, with their concentrations remaining similar. The non-depuration of polar PAHs from oyster tissues could imply greater human health risk compared to their parent analogues.

Bioavailability of polycyclic aromatic compounds (PACs) to the Sydney rock oyster (Saccostrea glomerata) from sediment matrices of an economically important Australian estuary.

Improving risk assessment and remediation rests on better understanding of contaminant bioavailability. Despite their strong toxicological attributes, little is known about the partitioning behaviour and bioavailability of polar polycyclic aromatic hydrocarbons (PAHs) in aquatic environments. The present study provides an insight into the bioavailable fractions of polar PAHs and their parent analogues in the tissues of the Sydney rock oyster, Saccostrea glomerata, a model aquatic bio-indicator organism. The concentration and distribution patterns of parent and polar PAHs including oxygenated PAHs (oxyPAHs), nitrated PAHs (NPAHs) and heterocyclic PAHs (HPAHs) were determined in water, sediment and oysters from an ecologically and economically important estuary of New South Wales, Australia. Total concentrations of PAHs, oxyPAHs, NPAHs and HPAHs were higher in sediments compared to oyster tissue and water. For most polar PAHs, total concentrations for water, sediment and oyster samples were <1 µg/g (µg/l for water) while parent PAH concentrations were several orders of magnitude higher. Computed biota-sediment accumulation factors (BSAFs) on lipid-normalized oyster concentrations revealed that while ∑oxyPAHs and ∑HPAHs exhibited low accumulation from sediment to oyster tissues (BSAF <1), ∑PAHs and ∑NPAH were found to be accumulated at high levels (BSAF >1). BSAF individual computation showed that bioaccumulation of nine investigated HPAHs in oyster tissues were relatively low and only 2-EAQ (oxyPAH) and 1N-NAP (NPAH) showed high levels of accumulation in oyster tissues, similar to parent PAHs. To the best of our knowledge, this is the first known study on the bioavailability of polar and non-polar PAHs in an Australian aquatic environment. The outcome of this study might be a useful indicator of the potential risks of polar PAHs to humans and other living organisms.

Accumulation and partitioning of metals and metalloids in the halophytic saltmarsh grass, saltwater couch, Sporobolus virginicus.

Remnant endangered saltmarsh communities in Australia often occur in urbanised estuaries where industrial processes have contaminated sediments with metal(loid)s. Despite this issue, virtually nothing is known on local plant species exposure to metal contaminants, nor their ability to uptake and translocate metal(loid)s from contaminated estuarine sediment. In the current study, we assessed the accumulation and partitioning of the metal(loid)s Zn, Cu, Pb, Cd and Se in the dominant saltmarsh primary producer, Sporobolus virginicus, across three urbanised estuaries in NSW Australia. Lake Macquarie was the most contaminated estuary, while Sydney Olympic Park, Port Jackson exhibited intermediate metal(loid) loadings and Hunter Wetlands exhibited the lowest loadings among estuaries. Essential metals (Zn and Cu) were more mobile, with sediment:root bioconcentration factors (BCFs) greater than unity and translocation among plant organs greater than, or equal to, unity. Other metal(loid)s were less mobile, with BCFs equal to unity and translocation factors among organs much reduced. Despite these barriers to translocation, all metal(loid)s were accumulated to roots with dose, and further accumulative relationships between metal(loid)s in roots and culms, and culms and leaves, were evidenced (with the exception of Cu). Along with sediment metal(loid)s, increases in sediment pH predicted Cu uptake in roots and increases in soil organic matter predicted Se uptake in roots. Although significant positive linear relationships were observed between sediment metal(loid)s and plant organ metal(loid)s(withholding Cu), the variance explained was low to intermediate for most metal(loid)s suggesting employing S. virginicus as an accumulative bioindicator would be impractical.

Characterisation of the metallothionein gene in the Sydney rock oyster and its expression upon metal exposure in oysters with different prior metal exposure histories.

The metal-binding protein metallothionein (MT) is widely used as a biomarker of metal contamination. In this study, we cloned a MT gene (sgMT) from the Sydney rock oyster Saccostrea glomerata. The gene encodes a MT-I protein with a classical αß domain structure and is expressed as two transcripts resulting from alternative polyadenylation. The gene promoter contains two putative metal-responsive elements (MREs) which are known to be required for metal-inducible transcription. A specific and efficient qPCR assay was developed to quantify sgMT mRNA expression. Further, we assessed whether prior metal exposure history influences sgMT mRNA expression upon subsequent metal exposure. Oysters with varying prior metal exposure histories (contaminated and reference) were exposed to Cu, Cd and Zn. Expression of sgMT generally increased with metal dose, and oysters with an elevated past metal exposure history exhibited higher sgMT expression under Cd and Zn stress, representing a potential acclimatory response to prior metal exposure.

The utility of vitellogenin as a biomarker of estrogenic endocrine disrupting chemicals in molluscs.

Estrogenic endocrine disrupting chemicals (EDCs) are natural hormones, synthetic compounds or industrial chemicals that mimic estrogens due to their structural similarity with estrogen's functional moieties. They typically enter aquatic environments through wastewater treatment plant effluents or runoff from intensive livestock operations. Globally, most natural and synthetic estrogens in receiving aquatic environments are in the low ng/L range, while industrial chemicals (such as bisphenol A, nonylphenol and octylphenol) are present in the µg to low mg/L range. These environmental concentrations often exceed laboratory-based predicted no effect concentrations (PNECs) and have been evidenced to cause negative reproductive impacts on resident aquatic biota. In vertebrates, such as fish, a well-established indicator of estrogen-mediated endocrine disruption is overexpression of the egg yolk protein precursor vitellogenin (Vtg) in males. Although the vertebrate Vtg has high sensitivity and specificity to estrogens, and the molecular basis of its estrogen inducibility has been well studied, there is growing ethical concern over the use of vertebrate animals for contaminant monitoring. The potential utility of the invertebrate Vtg as a biomonitor for environmental estrogens has therefore gained increasing attention. Here we review evidence providing support that the molluscan Vtg holds promise as an invertebrate biomarker for exposure to estrogens. Unlike vertebrates, estrogen signalling in invertebrates remains largely unclarified and the classical genomic pathway only partially explains estrogen-mediated activation of Vtg. In light of this, in the latter part of this review, we summarise recent progress towards understanding the molecular mechanisms underlying the activation of the molluscan Vtg gene by estrogens and present a hypothetical model of the interplay between genomic and non-genomic pathways in the transcriptional regulation of the gene.

The constitutively active estrogen receptor (ER) binds and activates the promoter of the vitellogenin (Vtg) gene in the Sydney rock oyster, Saccostrea glomerata.

Vitellogenin (Vtg) is a well-established biomarker of estrogenic exposure in aquatic animals. In vertebrates, Vtg gene transcription is controlled by the estrogen receptors (ERs). Although an ER ortholog is present in molluscs, its role as a transcriptional regulator remains elusive. Here, we tested the hypothesis that in the Sydney rock oyster, Saccostrea glomerata, the ER ortholog activates Vtg gene transcription through specific interaction with its promoter. Luciferase reporter assays indicated that sgER activated both a minimal promoter containing the consensus estrogen-responsive elements (EREs) and the sgVtg promoter in an estrogen-independent manner. The sgVtg promoter-luciferase activation was significantly reduced when any of three putative ERE half sites (½EREs) in the promoter were mutated. Electrophoretic mobility shift assay (EMSA) confirmed that sgER binds specifically to a 68-bp promoter sequence where these ½EREs reside. Overall, the results suggest that sgER is a constitutively active transcription factor that binds and activates the sgVtg promoter.

Potential mechanisms underlying estrogen-induced expression of the molluscan estrogen receptor (ER) gene.

In vertebrates, estrogens and estrogen mimicking chemicals modulate gene expression mainly through a genomic pathway mediated by the estrogen receptors (ERs). Although the existence of an ER orthologue in the mollusc genome has been known for some time, its role in estrogen signalling has yet to be deciphered. This is largely due to its constitutive (ligand-independent) activation and a limited mechanistic understanding of its regulation. To fill this knowledge gap, we cloned and characterised an ER cDNA (sgER) and the 5'-flanking region of the gene from the Sydney rock oyster Saccostrea glomerata. The sgER cDNA is predicted to encode a 477-amino acid protein that contains a DNA-binding domain (DBD) and a ligand-binding domain (LBD) typically conserved among both vertebrate and invertebrate ERs. A comparison of the sgER LBD sequence with those of other ligand-dependent ERs revealed that the sgER LBD is variable at several conserved residues known to be critical for ligand binding and receptor activation. Ligand binding assays using fluorescent-labelled E2 and purified sgER protein confirmed that sgER is devoid of estrogen binding. In silico analysis of the sgER 5'-flanking sequence indicated the presence of three putative estrogen responsive element (ERE) half-sites and several putative sites for ER-interacting transcription factors, suggesting that the sgER promoter may be autoregulated by its own gene product. sgER mRNA is ubiquitously expressed in adult oyster tissues, with the highest expression found in the ovary. Ovarian expression of sgER mRNA was significantly upregulated following in vitro and in vivo exposure to 17ß-estradiol (E2). Notably, the activation of sgER expression by E2 in vitro was abolished by the specific ER antagonist ICI 182, 780. To determine whether sgER expression is epigenetically regulated, the in vivo DNA methylation status of the putative proximal promoter in ovarian tissues was assessed using bisulfite genomic sequencing. The results showed that the promoter is predominantly hypomethylated (with 0-3.3% methylcytosines) regardless of sgER mRNA levels. Overall, our investigations suggest that the estrogen responsiveness of sgER is regulated by a novel ligand-dependent receptor, presumably via a non-genomic pathway(s) of estrogen signalling.

Mechanistic insights into induction of vitellogenin gene expression by estrogens in Sydney rock oysters, Saccostrea glomerata.

Marine molluscs, such as oysters, respond to estrogenic compounds with the induction of the egg yolk protein precursor, vitellogenin (Vtg), availing a biomarker for estrogenic pollution. Despite this application, the precise molecular mechanism through which estrogens exert their action to induce molluscan vitellogenesis is unknown. As a first step to address this question, we cloned a gene encoding Vtg from the Sydney rock oyster Saccostrea glomerata (sgVtg). Using primers designed from a partial sgVtg cDNA sequence available in Genbank, a full-length sgVtg cDNA of 8498bp was obtained by 5'- and 3'-RACE. The open reading frame (ORF) of sgVtg was determined to be 7980bp, which is substantially longer than the orthologs of other oyster species. Its deduced protein sequence shares the highest homology at the N- and C-terminal regions with other molluscan Vtgs. The full-length genomic DNA sequence of sgVtg was obtained by genomic PCR and genome walking targeting the gene body and flanking regions, respectively. The genomic sequence spans 20kb and consists of 30 exons and 29 introns. Computer analysis identified three closely spaced half-estrogen responsive elements (EREs) in the promoter region and a 210-bp CpG island 62bp downstream of the transcription start site. Upregulation of sgVtg mRNA expression was observed in the ovaries following in vitro (explants) and in vivo (tank) exposure to 17ß-estradiol (E2). Notably, treatment with an estrogen receptor (ER) antagonist in vitro abolished the upregulation, suggesting a requirement for an estrogen-dependent receptor for transcriptional activation. DNA methylation of the 5' CpG island was analysed using bisulfite genomic sequencing of the in vivo exposed ovaries. The CpG island was found to be hypomethylated (with 0-3% methylcytosines) in both control and E2-exposed oysters. However, no significant differential methylation or any correlation between methylation and sgVtg expression levels was observed. Overall, the results support the possible involvement of an ERE-containing promoter and an estrogen-activated receptor in estrogen signalling in marine molluscs.