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The presence of emergent antibiotics, in livestock wastewater may affect constructed wetlands (CWs) performance in the removal of other pollutants. The main objective of this study was to evaluate the influence of two antibiotics commonly used in livestock industry, enrofloxacin and ceftiofur, on metal removal by CWs. Microcosms (0.4 m×0.3 m×0.3 m), simulating CWs, were constructed with Phragmites australis to treat livestock wastewater spiked or not with 100 µg/L of enrofloxacin or ceftiofur (individually or in mixture). Wastewater was treated during 20 one-week cycles. After one-week cycle wastewater was removed and replaced by new wastewater (with or without spiking). At weeks 1, 2, 4, 8, 14, 18 and 20, treated wastewater was analysed to determine the removal rates of metals (Zn, Cu, Fe and Mn) and of each antibiotic. At weeks 1, 8 and 20 portions of the plant root substrate were collected and metals determined. At the end of the experiment metal levels were also determined in plant tissues. Removal rate of Fe from wastewater was 99%. Removal rates of Cu and Zn were higher than 85% and 89%, respectively, whereas for Mn removal rates up to 75% were obtained. In general, no significant differences were observed through time in the removals of the different metals, indicating that the systems maintained their functionality during the experimental period. Antibiotics did not interfere with the system depuration capacity, in terms of metals removals from wastewater, and ceftiofur even promoted metal uptake by P. australis . Therefore, CWs seem to be a valuable alternative to remove pollutants, including antibiotics and metals, from livestock wastewaters, reducing the risk the release of these wastewaters might pose into the environment, although more research should be conducted with other antibiotics in CWs. [ABSTRACT FROM AUTHOR]

Metal exposure is known to induce the production and secretion of substances, such as aliphatic low molecular weight organic acids (ALMWOAs), into the rhizosphere by plant roots. Knowledge on this matter is extensive for soil plants but still considerably scarce regarding marsh plants roots adapted to high salinity media. Phragmites australis and Halimione portulacoides , two marsh plants commonly distributed in European estuarine salt marshes, were used to assess the response of roots of both species, in terms of ALMWOAs exudation, to Cu, Ni and Cd exposure (isolated and in mixture since in natural environment, they are exposed to mixture of metals). As previous studies were carried out in unrealistic and synthetic media, here a more natural medium was selected. Therefore, in vitro experiments were carried out, with specimens of both marsh plants, and in freshwater contaminated with two different Cu, Ni and Cd concentrations (individual metal and in mixture). Both marsh plants were capable of liberating ALMWOAs into the surrounding medium. Oxalic, citric and maleic acids were found in P. australis root exudate solutions and oxalic and maleic acids in H. portulacoides root exudate solutions. ALMWOA liberation by both plants was plant species and metal-dependent. For instance, Cu affected the exudation of oxalic acid by H. portulacoides and of oxalic and citric acids by P. australis roots. In contrast, Ni and Cd did not stimulate any specific response. Regarding the combination of all metals, H. portulacoides showed a similar response to that observed for Cu individually. However, in the P. australis case, at high metal concentration mixture, a synergetic effect led to the increase of oxalic acid levels in root exudate solution and to a decrease of citric acid liberation. A correlation between ALMWOAs exudation and metal accumulation could not be established. P. australis and H. portulacoides are considered suitable metal phytoremediators of estuarine impacted areas. Understanding the mechanisms developed by these plants which allow them to tolerate and remediate metal-contaminated sediments is important to potentiate their use in phytoremediation purpose. This work provides new knowledge regarding the H. portulacoides and P. australis ability to exude ALMWOAs in response to metal contamination. [ABSTRACT FROM AUTHOR]

The increased use of metallic nanoparticles (NPs) raises the probability of finding NPs in the environment. A lot of information exists already regarding interactions between plants and metals, but information regarding interactions between metallic NPs and plants, including salt marsh plants, is still lacking. This work aimed to study interactions between CuO NPs and the salt marsh plants Halimione portulacoides and Phragmites australis . In addition, the potential of these plants for phytoremediation of Cu NPs was evaluated. Plants were exposed for 8 days to sediment elutriate solution doped either with CuO or with ionic Cu. Afterwards, total metal concentrations were determined in plant tissues. Both plants accumulated Cu in their roots, but this accumulation was 4 to 10 times lower when the metal was added in NP form. For P. australis , metal translocation occurred when the metal was added either in ionic or in NP form, but for H. portulacoides no metal translocation was observed when NPs were added to the medium. Therefore, interactions between plants and NPs differ with the plant species. These facts should be taken in consideration when applying these plants for phytoremediation of contaminated sediments in estuaries, as the environmental management of these very important ecological areas can be affected. [ABSTRACT FROM AUTHOR]

Metals are known to induce oxidative stress in plant cells. Antioxidant thiolic compounds are known to play an important role in plants׳ defence mechanisms against metal toxicity but, regarding salt marsh plants, their role is still very poorly understood. In this work, the involvement of non-protein thiols (NPT), such as cysteine (Cys), reduced glutathione (GSH), oxidised glutathione (GSSG) and total acid-soluble SH compounds (total thiols), in the tolerance mechanisms of the marsh plant Phragmites australis against Cu and Cd toxicity was assessed. Specimens of this plant, freshly harvested in an estuarine salt marsh, were exposed, for 7 days, to rhizosediment soaked with the respective elutriate contaminated with Cu (0, 10 and 100 mg/L) or Cd (0, 1, 10 mg/L). In terms of NPT production, Cu and Cd contamination induced different responses in P. australis . The content of Cys increased in plant tissue after plant exposure to Cu, whereas Cd contamination led to a decrease in GSSG levels. In general, metal contamination did not cause a significant variation on GSH levels. Both metals influenced, to some extent, the production of other thiolic compounds. Despite the accumulation of considerable amounts of Cu and Cd in belowground tissues, no visible toxicity signs were observed. So, antioxidant thiolic compounds were probably involved in the mechanisms used by P. australis to alleviate metal toxicity. As P. australis is considered suitable for phytostabilising metal-contaminated sediments, understanding its tolerance mechanisms to toxic metals is important to optimise the conditions for applying this plant in phytoremediation procedures. [ABSTRACT FROM AUTHOR]

Abstract: This work aimed to evaluate, in vitro, the capability of roots of two salt marsh plants to release low molecular weight organic acids (LMWOAs) and to ascertain whether Cu contamination would stimulate or not organic acids exudation. The sea rush Juncus maritimus and the sea-club rush Scirpus maritimus, both from the lower Douro river estuary (NW Portugal), were used. Plants were collected seasonally, four times a year in 2004, during low tide. After sampling, plant roots were washed for removal of adherent particles and immersed for 2 h in a solution that matched salinity (3) and pH (7.5) of the pore water from the same location to obtain plant exudates. In one of the seasons, similar experiments were carried out but spiking the solution with different amounts of Cu in order to embrace the range between 0 and 1600 nM. In the final solutions as well as in sediment pore water LMWOAs were determined by high performance liquid chromatography. Plants were able to release, in a short period of time, relatively high amounts of LMWOAs (oxalate, citrate, malate, malonate, and succinate). In the sediment pore water oxalate, succinate and acetate were also detected. Therefore, plant roots probably contributed to the presence of some of these organic compounds in pore water. Exudation differed between the plant species and also showed some seasonally variation, particularly for S. maritimus. The release of oxalate by J. maritimus increased with Cu increase in the media. However, exudation of the other LMWOAs did not seem to be stimulated by Cu contamination in the media. This fact is compatible with the existence of alternative internal mechanisms for Cu detoxification, as denoted by the fact that in media contaminated with Cu both plant species accumulated relatively high amounts (29–83%) of the initially dissolved Cu. This study expands our knowledge on the contribution of globally dominant salt marsh plants to the release of LMWOAs into the environment. [Copyright &y& Elsevier]

Abstract: The influence of each of four organic pollutants selected from among those commonly found in coastal areas, 1,1-dichloro-2,2-bis(p-chlorophenyl) ethylene (DDE), monobutyltin (MBT), Triton X-100 and polycyclic aromatic hydrocarbons (PAHs), on Cu accumulation by Halimione portulacoides was investigated. Experiments were carried out in a laboratory setting, either in hydroponics (sediment elutriate) or in a salt marsh sediment (Cávado River, NW Portugal) soaked in elutriate. Groups of H. portulacoides were exposed to media for 6 days spiked with 10mg/L Cu(II) and with one of the selected pollutants, at an environmentally realistic concentration. DDE and MBT did not cause any major change on Cu accumulation by H. portulacoides, whereas PAHs slightly increased accumulation only in hydroponics i.e. in the absence of sediment. On the other hand, the non-ionic surfactant Triton X-100 markedly favoured Cu accumulation on plant roots both in the presence and absence of sediment. The addition of DDE, MBT and Triton X-100 also favoured Cu solubility from sediments. Therefore, the simultaneous presence of pollutants from different nature (inorganic and organic) in the estuarine environment may result in a composition of water column, pore water, sediment or biota different of that expected considering the effect of each individual pollutant. [Copyright &y& Elsevier]

Abstract: This work aimed to evaluate, in vitro, the capability of roots of salt marsh plants to release strong Cu-complexing ligands and to ascertain whether Cu contamination would stimulate ligands'' exudation or not. The sea rush Juncus maritimus and the sea-club rush Scirpus maritimus, both from the lower Douro river estuary (NW Portugal), were used. Plants were collected seasonally, four times a year in 2004, during low tide. After sampling, plant roots were washed for removal of adherent particles and immersed for 2h in a solution that matched salinity (3) and pH (7.5) of the pore water from the same location and spiked with Cu2+ in the range 0–1600nM to obtain plant exudates. In the final solutions as well as in sediment pore water total dissolved Zn and Cu, Cu-complexing ligand concentrations and the respective conditional stability constants (K CuL ′) values were determined by voltammetry. This study demonstrated that plants are able to release, in a short period of time, relatively high amounts of strong Cu-complexing ligands (56–265nmolgroot −1), which differed among plants and sampling site but were independent of the season. Cu contamination did not stimulate exudation of Cu-complexing ligands. On the other hand, in media contaminated with Cu both plants accumulated relatively high amounts (29–83%) of the initially dissolved Cu, indicating that they have alternative internal mechanisms for Cu detoxification. Cu exchange between roots and medium (either accumulation in contaminated medium or release in the absence of Cu) was more intense for S. maritimus than for J. maritimus. It was observed that exudate solutions obtained in the absence of added Cu and sediment pore water (the densities of roots observed inside the salt marsh where comparable to those used in the in vitro experiments), displayed similarities in terms of total dissolved metals, Cu-complexing ligands concentrations, values of K CuL ′ (12

Abstract: The aim of this work was to identify a variety of low molecular weight organic acids exuded by the sea rush Juncus maritimus collected at two locations with different sediment characteristics (sandy and muddy) and to examine whether specific differences in physico-chemical sediment characteristics influenced plant exudation. Just after collection, plant roots were rinsed and put in contact with deionised water for 2h. In the obtained solution the organic acids, exuded by the plants, were determined by high performance liquid chromatography. Juncus maritimus was shown to be capable of releasing malonate and oxalate. Sediments and rhizosediments (sediment in contact with the plant roots and rhizomes, corresponding to the area of higher belowground biomass) from the areas where the plants had been collected were characterised in terms of physical and chemical composition, including acid volatile sulphide and total-recoverable metals (Pb, Cr, Cu, Zn, Ni and Cd). It was found that the extent of exudation varied markedly between sites. The identified organic acids were used as extractants of metals from sediments and rhizosediments and the results were compared with those provided by a very commonly used sequential extraction approach, which was carried out in parallel. This work demonstrates that J. maritimus can release organic compounds that can act as complexing agents of trace metal and therefore organic exudates should be accounted for when dealing with estuarine environment quality. [Copyright &y& Elsevier]

Abstract: Degradation of petroleum hydrocarbons in colonized and un-colonized sediments by salt marsh plants Juncus maritimus and Phragmites australis collected in a temperate estuary was investigated during a 5-month greenhouse experiment. The efficiency of two bioremediation treatments namely biostimulation (BS) by the addition of nutrients, and bioaugmentation (BA) by addition of indigenous microorganisms was tested in comparison with hydrocarbon natural attenuation in un-colonized and with rhizoremediation in colonized sediments. Hydrocarbon degrading microorganisms and root biomass were assessed as well as hydrocarbon degradation levels. During the study, hydrocarbon degradation in un-colonized sediments was negligible regardless of treatments. Rhizoremediation proved to be an effective strategy for hydrocarbon removal, yielding high rates in most experiments. However, BS treatments showed a negative effect on the J. maritimus potential for hydrocarbon degradation by decreasing the root system development that lead to lower degradation rates. Although both plants and their associated microorganisms presented a potential for rhizoremediation of petroleum hydrocarbons in contaminated salt marsh sediments, results highlighted that nutrient requirements may be distinct among plant species, which should be accounted for when designing cleanup strategies. [Copyright &y& Elsevier]

Abstract This study aimed to investigate the potential of microbial communities from the rhizosediment of two plants - Phragmites australis and Juncus maritimus - occurring in an estuarine area subjected to a high anthropogenic impact, to biodegrade ENR, a commonly used veterinary antibiotic. An enrichment process with 1 mgL−1 of ENR was conducted during ca. 9 months, using acetate as a co-substrate. After this, the enriched microbial consortia were challenged with higher ENR concentrations of 2 and 3 mgL−1. Microbial cultures enriched with 1 mgL−1 of ENR were capable of biodegrading this antibiotic, though not completely. By the end of the enrichment phase, microbial cultures were defluorinating an average of 50% of the ENR supplemented. Higher ENR concentrations led to lower biodegradation performances, suggesting a possible toxic/inhibitory effect in the microbial cultures. Phylogenetic identification of the microorganisms isolated from microbial cultures enriched with ENR revealed a high taxonomical diversity, with microorganisms belonging mainly to Proteobacteria and Bacteroidetes phyla. Assemblage of the obtained isolated strains (according to the enriched cultures from which they were isolated) revealed that the resulting consortia were also capable of degrading ENR, indicating that the main microbial players in the biodegradation of this antibiotic were isolated. These consortia also showed to be more robust to degrade higher concentrations of ENR than the corresponding enriched cultures. This study shows that microorganisms derived from rhizosediments of the selected plants, exhibit capacity to biodegrade ENR, though not completely for the concentrations tested, and may be further explored for the development of bioremediation strategies for the treatment of this antibiotic. Highlights • Capacity of microbes from rhizosediment of estuary plants to degrade ENR was assessed. • Enriched microbial cultures were able to degrade ENR though not completely. • Exposure to higher ENR concentrations led to lower biodegradation performances. • Degrading strains predominantly belonged to Proteobacteria and Bacteroidetes phyla. • Assemblage of isolates from enriched cultures also resulted in ENR biodegradation. [ABSTRACT FROM AUTHOR]

Reusing reclaimed wastewater is needed to fight water scarcity, reduce freshwater consumption and conserve water resources, but one must ensure that hazardous substances are fully removed/eliminate before that reuse. The potential of lab-scale constructed wetlands (CWs) for the removal of chemical and biological contaminants from livestock wastewater, while maintaining nutrient levels for fertilization, was assessed, evaluating changes in microbial communities, with particular focus on potential pathogens. CW microcosms with two different substrates (lava rock or light expanded clay aggregate), both planted with Phragmites australis , were tested. After 15 days of treatment, removal rates were higher than 80% for Cd, Cr, Cu, Fe, Pb and Zn, in general with no significant differences between the two different substrates. Organic matter and nutrients were also removed but their levels still allowed the used of the treated wastewater as a fertilizer Removal of bacterial contamination was estimated through enumeration of cultivable bacteria. High removal rates of fecal indicator bacteria were observed, reaching >95% for enterococci and >98% for enterobacteria after 15 days of treatment, decreasing hazardous biological contaminants initially present in the wastewater. In addition, the microbial communities in the initial and treated wastewater, and in the plant roots bed substrate, were characterized by using 16SrRNA gene amplicon sequencing. Microbial communities in the CW systems showed a clear shift comparatively with the initial wastewater showing system adaptation and removal potentialities. This also revealed an important removal of the most represented potential pathogenic genus, Clostridium , which relative abundance decreased from 33% to 1% through the treatment. Overall, CWs showed potential to be efficient in removing chemical and biological contaminants, while maintaining moderated levels of nutrients, allowing the reuse of reclaimed water in agriculture, namely as fertilizer. Current results will contribute for the optimization and use of CWs for a sustainable treatment of liquid wastes, promoting the circular economy. • Removal rates >80% for Cd, Cr, Cu, Fe, Pb and Zn in livestock wastewater treatment CWs. • Fecal indicator bacteria removal >95% for enterococci and >98% for enterobacteria after CW treatment. • CW systems microbial communities clearly shifted comparatively with the initial wastewater. • CW systems microbial communities showed system adaptation and removal potentialities. • CW treatment maintain moderated levels of nutrients, allowing reclaimed water reuse in agriculture. [ABSTRACT FROM AUTHOR]

Aquaculture is the fastest growing animal food-producing sector. Water is the central resource for aquaculture, and it is essential that its quality be preserved. Micropollutants (MPs) can reach aquaculture through anthropogenic addition or inlet water, and may cause harmful effects such as endocrine disruption and antibiotic resistance, adversely affecting the fish species being farmed. Furthermore, the discharge of aquaculture effluents into the environment may contribute to the deterioration of water courses. In this sense, the implementation of environmentally responsible measures in aquaculture farms is imperative for the protection of ecosystems and human health. The European Commission (EC) has recently launched a guiding document promoting ecological aquaculture practices; however, options for water treatment are still lacking. Conventional processes are not designed to deal with MPs; this review article consolidates relevant information on the application of advanced oxidation technologies (AOTs) and constructed wetlands (CWs) as potential strategies in this regard. Although 161 studies on the application of AOTs or CWs in aquaculture have already been published, only 34 focused on MPs (28 on AOTs and 6 on CWs), whereas the others reported the removal of contaminants such as bacteria, organic matter, solids and inorganic ions. No study coupling both treatments has been reported to date for the removal of MPs from aquaculture waters. AOTs and CWs are prospective alternatives for the treatment of aquacultural aqueous matrices. However, the type of aquaculture activity and the specifications of these available technologies should be considered while selecting the most suitable treatment option. [Display omitted] • Micropollutants in aquaculture water is a concern of the European Commission. • Chemical and biological treatments to remove these micropollutants are reviewed. • Studies with real matrices are scarce for advanced oxidation technologies (AOTs). • More studies conducted at full scale are needed for constructed wetlands (CWs). • Both AOTs and CWs can remove conventional pollutants from aquaculture waters. [ABSTRACT FROM AUTHOR]

Abstract: Several salt marsh plant species have shown to be able to uptake and concentrate metals in their tissues, showing potential for metal phytoremediation. However, studies in controlled conditions, mimicking as much as possible the plants natural environment, are needed to confirm this potential. For the present study, Juncus maritimus and Phragmites australis were collected in an estuary together with the sediment surrounding their roots, put in vessels and maintained in greenhouses under estuarine tidal simulation. After 3 weeks of acclimation, vessels were spiked with two different cadmium concentrations. After 2 months, cadmium was assessed in plant tissues and sediments. Results indicate that both plant species were able to uptake and translocate cadmium into their tissues, contributing also to retain it in rhizosediments and thus reducing the available amount of metal in the environment. Metal was preferentially accumulated in belowground structures, in concentrations not directly proportional to the amount of cadmium present in the sediment. Although no visual toxicity signs were observed, some defence mechanisms were triggered as observed by the changes in carotenoids, lignin, total soluble phenolic compounds and thiolic compounds levels, this response differing between plant species. This work shows that these two salt marsh plants can contribute for the retention of cadmium in salt marshes being useful for the phytostabilization of this metal in estuarine environments. [Copyright &y& Elsevier]

The recovery of estuarine environments is in need. Phytoremediation could be a valid option to reduce pollution while preserving natural biodiversity. In this work, estuarine sediments colonized by Juncus maritimus or Phragmites australis were spiked with cadmium in the absence and in the presence of an autochthonous microbial consortium resistant to the metal. The aim of this study was to increase the potential for cadmium phytoremediation that these two halophyte plants have shown. Experiments were carried out in greenhouses with an automatic irrigation system that simulated estuarine tidal cycles. After 2 months, Cd concentration in P. australis stems increased up to 7 times when the rhizosphere was inoculated with the microbial consortium. So, P. australis phytoextraction potential was increased through autochthonous bioaugmentation. As for J. maritimus, up to 48% more Cd (total amount) was observed in its belowground tissues after being subjected to autochthonous bioaugmentation. Therefore, the phytostabilization potential of this plant was promoted. For both plants this increase in cadmium uptake did not cause significant signs of toxicity. Therefore, the addition of autochthonous microorganisms resistant to cadmium seems to be a valuable strategy to potentiate phytoremediation of this metal in saltmarshes, being useful for the recovery of moderately impacted estuaries. This will contribute for an effective management of these areas. Research on this topic regarding estuarine ecosystems, especially saltmarshes, is, to our knowledge, inexistent. [Copyright &y& Elsevier]

Abstract: Salt marshes are important sinks for contaminants, namely metals that tend to accumulate around plant roots and could eventually be taken up in a process known as phytoremediation. On the other hand, microbial communities display important roles in the salt marsh ecosystems, such as recycling of nutrients and/or degradation of organic contaminants. Thus, plants can benefit from the microbial activity in the phytoremediation process. Nevertheless, above certain levels, metals are known to be toxic to microorganisms, fact that can eventually compromise their ecological functions. In this vein, the aim of present study was to investigate, in the laboratory, the effect of selected metals (Cd, Cu and Pb) on the microbial communities associated to the roots of two salt marsh plants. Sediments colonized by Juncus maritimus and Phragmites australis were collected in the River Lima estuary (NW Portugal), and spiked with each of the metals at three different Effects Range-Median (ERM) concentrations (1, 10×, 50×), being ERM the sediment quality guideline that indicates the concentration above which adverse biological effects may frequently occur. Spiked sediments were incubated with a nutritive saline solution, being left in the dark under constant agitation for 7 days. The results showed that, despite the initial sediments colonized by J. maritimus and P. australis displayed significant (p < 0.05) differences in terms of microbial community structure (evaluated by ARISA), they presented similar microbial abundances (estimated by DAPI). Also, in terms of microbial abundance, both sediments showed a similar response to metal addition, with a decrease in number of cells only observed for the higher addition of Cu. Nevertheless, both Cu and Pb, at intermediate metals levels promote a shift in the microbial community structure, with possibly effect on the ecological function of these microbial communities in salt marshes. These changes may affect plants phytoremediation potential and further work on this subject is in need. [Copyright &y& Elsevier]