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This study investigates the potential of an indigenous estuarine microbial consortium to degrade two polycyclic aromatic hydrocarbons (PAHs), naphthalene and fluoranthene, under nitrate-reducing conditions. Two physicochemically diverse sediment samples from the Lima Estuary (Portugal) were spiked individually with 25 mg L -1 of each PAH in laboratory designed microcosms. Sediments without PAHs and autoclaved sediments spiked with PAHs were run in parallel. Destructive sampling at the beginning and after 3, 6, 12, 30 and 63 weeks incubation was performed. Naphthalene and fluoranthene levels decreased over time with distinct degradation dynamics varying with sediment type. Next-generation sequencing (NGS) of 16 S rRNA gene amplicons revealed that the sediment type and incubation time were the main drivers influencing the microbial community structure rather than the impact of PAH amendments. Predicted microbial functional analyses revealed clear shifts and interrelationships between genes involved in anaerobic and aerobic degradation of PAHs and in the dissimilatory nitrate-reducing pathways (denitrification and dissimilatory nitrate reduction to ammonium - DNRA). These findings reinforced by clear biogeochemical denitrification signals (NO 3 - consumption, and NH 4 + increased during the incubation period), suggest that naphthalene and fluoranthene degradation may be coupled with denitrification and DNRA metabolism. The results of this study contribute to the understanding of the dissimilatory nitrate-reducing pathways and help uncover their involvement in degradation of PAHs, which will be crucial for directing remediation strategies of PAH-contaminated anoxic sediments., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Denitrification and anammox are key processes for reducing the external nitrogen loads delivered to coastal ecosystems, and these processes can be affected by pollutants. In this study, we investigated the effect of crude oil on denitrification and anammox. Controlled laboratory experiments were performed using sediment slurries from the Lima Estuary (NW Portugal). Anammox and denitrification rates were measured using (15)N-labeled NO3(-), and the production of (29)N2 and (30)N2 quantified by membrane inlet mass spectrometry. Results revealed that while denitrification rates were stimulated between 10 and 25 000 times after crude oil amendment, anammox activity was partially (between 2 and 5 times) or completely inhibited by the addition of crude oil when comparing to rates in unamended controls. Similar results were observed across four estuarine sediment types, despite their different physical-chemical characteristics. Moreover, N2O production was reduced by 2-36 times following crude oil addition. Further work is required to fully understand the mechanism(s) of the observed reduction in N2O production. This study represents one of the first contributions to the understanding of the impact of crude oil pollution on denitrification and anammox, with profound implications for the management of aquatic ecosystems regarding eutrophication (N-removal)., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

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 © 2014 Elsevier Ltd. All rights reserved.)

Microbial communities play an important role in the biodegradation of organic pollutants in sediments, including hydrocarbons. The aim of this study was to evaluate the response of temperate salt marsh microbial communities to petroleum contamination, in terms of community structure, abundance and capacity to degrade hydrocarbons. Sediments un-colonized and colonized (rhizosediments) by Juncus maritimus, Phragmites australis and Triglochin striata were collected in a temperate estuary (Lima, NW Portugal), spiked with petroleum under variable nutritional conditions, and incubated for 15 days. Results showed that plant speciation emerged as the major factor for shaping the rhizosphere community structure, overriding the petroleum influence. Moreover, when exposed to petroleum contamination, the distinct salt marsh microbial communities responded similarly with (i) increased abundance, (ii) changes in structure, and (iii) decreased diversity. Communities, particularly those associated to J. maritimus and P. australis roots displayed a potential to degrade petroleum hydrocarbons, with degradation percentages between 15% and 41%, depending on sediment type and nutritional conditions. In conclusion, distinct salt marsh microbial communities responded similarly to petroleum contamination, but presented different pace, nutritional requirements, and potential for its biodegradation, which should be taken into account when developing bioremediation strategies., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The influence of Juncus maritimus, Phragmites australis, and Triglochin striata on hydrocarbon degrading microorganisms (HD) in Lima River estuary (NW Portugal) was investigated through a year-long plant life cycle. Sediments un-colonized and colonized (rhizosediments) by those salt marsh plants were sampled for HD, total cell counts (TCC), and total petroleum hydrocarbons (TPHs) assessment. Generally, TCC seemed to be markedly thriving by the presence of roots, but without significant (p > 0.05) differences among rhizosediments. Nevertheless, plants seemed to have a distinct influence on HD abundance, particularly during the flowering season, with higher HD abundance in the rhizosediments of the fibrous roots plants (J. maritimus < P. australis < T. striata). Our data suggest that different plants have distinct influence on the dynamics of HD populations within its own rhizosphere, particularly during the flowering season, suggesting a period of higher rhizoremediation activity. Additionally, during the vegetative period, plants with fibrous and dense root system tend to retain hydrocarbons around their belowground tissues more efficiently than plants with adventitious root system. Overall results indicate that fibrous root plants have a higher potential to promote hydrocarbons degradation, and that seasonality should be taken into account when designing long-term rhizoremediation strategies in estuarine areas.

Estuaries are often considered sinks for contaminants and the cleanup of salt marshes, sensitive ecosystems with a major ecological role, should be carried out by means of least intrusive approaches, such as bioremediation. This study was designed to evaluate the influence of plant-microorganisms associations on petroleum hydrocarbons fate in salt marshes of a temperate estuary (Lima River, NW Portugal). Sediments un-colonized and colonized (rhizosediments) by different plants (Juncus maritimus, Phragmites australis, Triglochin striata and Spartina patens) were sampled in four sites of the lower and middle estuary for hydrocarbon degrading microorganisms (HD), total cell counts (TCC) and total petroleum hydrocarbons (TPHs) assessment. In general, TPHs, HD and TCC were significantly higher (P < 0.05) in rhizosediments than in un-colonized sediments. Also recorded were differences on the abundance of hydrocarbon degraders among the rhizosediment of the different plants collected at the same site (J. maritimus < P. australis < T. striata), with statistically significant differences (P < 0.05) between J. maritimus and T. striata. Moreover, strong positive correlations-0.81 and 0.84 (P < 0.05), between biotic (HD) and abiotic (organic matter content) parameters and TPHs concentrations were also found. Our data clearly suggest that salt marsh plants can influence the microbial community, by fostering the development of hydrocarbon-degrading microbial populations in its rhizosphere, an effect observed for all plants. This effect, combined with the plant capability to retain hydrocarbons around the roots, points out that salt marsh plant-microorganisms associations may actively contribute to hydrocarbon removal and degradation in estuarine environments.