Validation of an Adapted QuEChERS Method for the Simultaneous Analysis of Polycyclic Aromatic Hydrocarbons, Polychlorinated Biphenyls and Organochlorine Pesticides in Sediment by Gas Chromatography-Mass Spectrometry.

A Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) method has been adapted and validated for the simultaneous analysis of 16 PAHs, 12 PCBs and 9 OCPs in sediment. The sample preparation was adapted by modifying the nature of the extraction solvent, the extraction technique and the amount of sediment. The analytical performances were evaluated in terms of accuracy, linearity and quantification limits. The method was validated by the analysis of a reference marine sediment material (SRM 1941b). The obtained concentrations are in good agreement with the certified values with recoveries ranging 60 %-103 % for most of PAHs. Acceptable recoveries are obtained for PCBs, ranging 76 %-131 %, and for OCPs ranging 81 %-137 %. The method was applied to the analysis of sediments from the hydro-system Bizerte Lagoon/Ichkeul Lake (Tunisia). The Bizerte lagoon is mainly contaminated by PAHs whereas the Ichkeul lake is mainly by OCPs.

Impacts of bioremediation schemes for the mitigation of a low-dose anthracene contamination on free-living marine benthic nematodes.

A microcosm experiment was used to examine (1) the effects of different bioremediation schemes on degradation of anthracene and the structure of free-living marine nematodes in a lightly contaminated (4.5 µg g(-1)) sediment from Bizerte lagoon and (2) the responses of the nematode community upon an artificial spiking of a low dose anthracene (1 µg g(-1)). For that purpose sediment microcosms were incubated in laboratory for 40 days. Bioremediation techniques decreased the anthracene contamination, and interestingly, biodegradation were more efficient when anthracene was artificial supplied into the sediment suggesting that the addition of bioavailable anthracene stimulated the bacterial community to adjust towards a PAH-degrading community. Spiking with this low dose of anthracene provoked significant changes in the nematode community structure and abundance, with the elimination of specific species such as Mesacanthion diplechma, the decrease of the dominant species Oncholaimus campylocercoides and the increase in abundance of opportunistic species such as Spirinia parasitifera. This would suggest a low tolerance of the nematode community despite the presence of a weak anthracene contamination in the sediment that could have allow dominance of an anthracene tolerant nematode species. Anthracene toxicity was alleviated in biostimulation treatments, leading to a strong increase in nematode abundance, concomitantly with changes in the nematode community structure; Prochromadorella neapolitana became the most abundant species.

Effectiveness of Shewanella oneidensis bioaugmentation in the bioremediation of phenanthrene-contaminated sediments and possible consortia with omnivore-carnivore meiobenthic nematodes.

This study was conducted to assess the impact and efficiency of the bioaugmentation as a bioremediation technique in annoying effects of a polycyclic aromatic hydrocarbon (phenanthrene) on a community of free-living nematodes from Bizerte bay (Tunisia). For this purpose, closed microcosms were exposed to three doses of phananthrene (0.1 µg kg-1, 1 µg kg-1 and 10 µg kg-1), in combination or not with a strain of Shewanella oneidensis. After 40 days of the exposure, results were obtained at the numerical, taxonomic and feeding levels. The results of univariate analyses revealed significant decreases in most univariate indices for phenanthrene treated communities compared to controls, with a discernible increase in the proportion of epistrate feeders. After bioaugmentation, similar patterns were observed for univariate and multivariate analyses, with the exception of the highest treatment, which showed no difference from the controls. The results obtained showed that the bioaugmentation with Shewanellea oneidensis was highly effective in reducing the negative impact of the highest dose of phenanthrene (10 µg kg-1 Dry Weight) tested on meiobenthic nematodes. Furthermore, a combination of Shewanellea oneidensis and four omnivore-carnivore nematode taxa could be suggested as an effective method in the bioremediation of phenanthrene-contaminated sediment.

Microbial diversity alteration reveals biomarkers of contamination in soil-river-lake continuum.

Microbial communities inhabiting soil-water-sediment continuum in coastal areas provide important ecosystem services. Their adaptation in response to environmental stressors, particularly mitigating the impact of pollutants discharged from human activities, has been considered for the development of microbial biomonitoring tools, but their use is still in the infancy. Here, chemical and molecular (16S rRNA gene metabarcoding) approaches were combined in order to determine the impact of pollutants on microbial assemblages inhabiting the aquatic network of a soil-water-sediment continuum around the Ichkeul Lake (Tunisia), an area highly impacted by human activities. Samples were collected within the soil-river-lake continuum at three stations in dry (summer) and wet (winter) seasons. The contaminant pressure index (PI), which integrates Polycyclic aromatic hydrocarbons (PAHs), alkanes, Organochlorine pesticides (OCPs) and metal contents, and the microbial pressure index microgAMBI, based on bacterial community structure, showed significant correlation with contamination level and differences between seasons. The comparison of prokaryotic communities further revealed specific assemblages for soil, river and lake sediments. Correlation analyses identified potential "specialist" genera for the different compartments, whose abundances were correlated with the pollutant type found. Additionally, PICRUSt analysis revealed the metabolic potential for pollutant transformation or degradation of the identified "specialist" species, providing information to estimate the recovery capacity of the ecosystem. Such findings offer the possibility to define a relevant set of microbial indicators for assessing the effects of human activities on aquatic ecosystems. Microbial indicators, including the detection of "specialist" and sensitive taxa, and their functional capacity, might be useful, in combination with integrative microbial indices, to constitute accurate biomonitoring tools for the management and restoration of complex coastal aquatic systems.

Bacterial community structure of sediments of the bizerte lagoon (Tunisia), a southern Mediterranean coastal anthropized lagoon.

In order to estimate how pollution affects the bacterial community structure and composition of sediments, chemical and molecular approaches were combined to investigate eight stations around the Bizerte lagoon. Terminal restriction fragment length polymorphism (T-RFLP) analysis of PCR-amplified 16S rRNA genes revealed that each station was characterized by a specific bacterial community structure. The combination of this data with those of chemical analysis showed a correlation between the bacterial fingerprint and the pollutant content, principally with hydrocarbon pollution. The composition of the bacterial community of two contrasted stations related to the pollution revealed sequences affiliated to alpha, beta, gamma, delta, epsilon subclass of the Proteobacteria, Actinobacteria, and Acidobacteria in both stations although in different extent. Gamma and delta subclass of the Proteobacteria were dominant and represent 70% of clones in the heavy-metal-contaminated station and 47% in the polyaromatic hydrocarbon (PAH)-contaminated. Nevertheless, most of the sequences found were unaffiliated to cultured bacteria. The adaptation of the bacterial community mainly to PAH compounds demonstrated here and the fact that these bacterial communities are mainly unknown suggest that the Bizerte lagoon is an interesting environment to understand the capacity of bacteria to cope with some pollutants.

Bacterial community assemblages in sediments under high anthropogenic pressure at Ichkeul Lake/Bizerte Lagoon hydrological system, Tunisia.

Bacterial communities inhabiting sediments in coastal areas endure the effect of strong anthropogenic pressure characterized by the presence of multiple contaminants. Understanding the effect of pollutants on the organization of bacterial communities is of paramount importance in order to unravel bacterial assemblages colonizing specific ecological niches. Here, chemical and molecular approaches were combined to investigate the bacterial communities inhabiting the sediments of the Ichkeul Lake/Bizerte Lagoon, a hydrological system under anthropogenic pressure. Although the microbial community of the Ichkeul Lake sediment was different to that of the Bizerte Lagoon, common bacterial genera were identified suggesting a lake-lagoon continuum probably due to the hydrology of the system exchanging waters according to the season. These genera represent bacterial "generalists" maintaining probably general biogeochemical functions. Linear discriminant analysis effect size (LEfSe) showed significant differential abundance distribution of bacterial genera according to the habitat, the pollution type and level. Further, correlation analyses identified specific bacterial genera which abundance was linked with pesticides concentrations in the lake, while in the lagoon the abundance of specific bacterial genera was found linked with the concentrations of PAHs (Polycyclic aromatic hydrocarbons) and organic forms of Sn. As well, bacterial genera which abundance was not correlated with the concentrations of pollutants were identified in both lake and lagoon. These findings represent valuable information, pointing out specific bacterial genera associated with pollutants, which represent assets for developing bacterial tools for the implementation, the management, and monitoring of bioremediation processes to mitigate the effect of pollutants in aquatic ecosystems.

Distribution of organic contamination of sediments from Ichkeul Lake and Bizerte Lagoon, Tunisia.

Analyses of organochlorine pesticides (OCPs), polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), and butyl tins (BuSn) were conducted on sediments from Ichkeul Lake-Bizerte Lagoon watershed (Tunisia). A total of 59 compounds (16 PAHs, 12 PCBs, 22 OCPs and 9 BuSn) were measured in 40 surface sediment samples collected during two campaigns. High concentrations of total PAHs were identified in the lagoon ranging from 122 to 19600ng·g-1. Several OCPs, including endrin, dieldrin, and lindane (Hexachlorocyclohexane or HCH or BHC) were found in high concentrations in Ichkeul Lake, ranging from 28 to 2012ngg-1. PAHs and OCPs varied seasonally, in response to the complex hydrology of the watershed. The concentrations of total PCBs ranged between 0.04 and 10.653ngg-1 and suggests low total PCBs sediment contamination, when compared to most international criteria. Total BuSn concentrations range between 67 and 526ng·g-1, which are relatively low when compared to most international criteria and ecological risk assessments. This is the first study of organic contamination in Ichkeul Lake (RAMSAR and UNESCO World Heritage site).

Response of Core Microbial Consortia to Chronic Hydrocarbon Contaminations in Coastal Sediment Habitats.

Traditionally, microbial surveys investigating the effect of chronic anthropogenic pressure such as polyaromatic hydrocarbons (PAHs) contaminations consider just the alpha and beta diversity and ignore the interactions among the different taxa forming the microbial community. Here, we investigated the ecological relationships between the three domains of life (i.e., Bacteria, Archaea, and Eukarya) using 454 pyrosequencing on the 16S rRNA and 18S rRNA genes from chronically impacted and pristine sediments, along the coasts of the Mediterranean Sea (Gulf of Lion, Vermillion coast, Corsica, Bizerte lagoon and Lebanon) and the French Atlantic Ocean (Bay of Biscay and English Channel). Our approach provided a robust ecological framework for the partition of the taxa abundance distribution into 859 core Operational taxonomic units (OTUs) and 6629 satellite OTUs. OTUs forming the core microbial community showed the highest sensitivity to changes in environmental and contaminant variations, with salinity, latitude, temperature, particle size distribution, total organic carbon (TOC) and PAH concentrations as main drivers of community assembly. The core communities were dominated by Gammaproteobacteria and Deltaproteobacteria for Bacteria, by Thaumarchaeota, Bathyarchaeota and Thermoplasmata for Archaea and Metazoa and Dinoflagellata for Eukarya. In order to find associations among microorganisms, we generated a co-occurrence network in which PAHs were found to impact significantly the potential predator - prey relationship in one microbial consortium composed of ciliates and Actinobacteria. Comparison of network topological properties between contaminated and non-contaminated samples showed substantial differences in the network structure and indicated a higher vulnerability to environmental perturbations in the contaminated sediments.

Changes of benthic bacteria and meiofauna assemblages during bio-treatments of anthracene-contaminated sediments from Bizerta lagoon (Tunisia).

Sediments from Bizerta lagoon were used in an experimental microcosm setup involving three scenarios for the bioremediation of anthracene-polluted sediments, namely bioaugmentation, biostimulation, and a combination of both bioaugmentation and biostimulation. In order to investigate the effect of the biotreatments on the benthic biosphere, 16S rRNA gene-based T-RFLP bacterial community structure and the abundance and diversity of the meiofauna were determined throughout the experiment period. Addition of fresh anthracene drastically reduced the benthic bacterial and meiofaunal abundances. The treatment combining biostimulation and bioaugmentation was most efficient in eliminating anthracene, resulting in a less toxic sedimentary environment, which restored meiofaunal abundance and diversity. Furthermore, canonical correspondence analysis showed that the biostimulation treatment promoted a bacterial community favorable to the development of nematodes while the treatment combining biostimulation and bioaugmentation resulted in a bacterial community that advantaged the development of the other meiofauna taxa (copepods, oligochaetes, polychaetes, and other) restoring thus the meiofaunal structure. The results highlight the importance to take into account the bacteria/meiofauna interactions during the implementation of bioremediation treatment.

Microbial community responses to bioremediation treatments for the mitigation of low-dose anthracene in marine coastal sediments of Bizerte lagoon (Tunisia).

PURPOSE: The main goals of this study were to investigate (1) the behavior of microbial communities in response to low-dose bioavailable anthracene addition in lightly contaminated sediment from Bizerte Lagoon and (2) the effects of bioremediation treatments on microbial biomass, activity, and community structure. METHODS: Sediment microcosms amended with 1 ppm anthracene were incubated in triplicate during 30 days. Biostimulation (addition of nitrogen and phosphorus fertilizer) and bioaugmentation (inoculation of a hydrocarbonoclastic bacterium) were used as bioremediation treatments. Bacterial biomass was estimated using flow cytometry. Sediment oxygen consumption was measured with oxygen microelectrodes. Bacterial community structure was assessed by molecular fingerprints (terminal restriction fragment length polymorphism; T-RFLP) analysis. RESULTS: Anthracene contamination resulted in a significant reduction of bacterial abundance with an impact on cell integrity. Concomitantly, sediment oxygen consumption was strongly inhibited. Correspondence analysis on T-RFLP data indicated that bacterial community structures from anthracene-contaminated microcosms were different from that of the control. Interestingly, the changes observed in microbial biomass, structure, and activities as a result of anthracene contamination were not alleviated even with the use of biostimulation and combination of biostimulation and bioaugmentation strategy for anthracene bioremediation. Nevertheless, both treatment methods resulted in different community structures relative to the contaminated and control microcosms with the appearance of distinct populations. CONCLUSION: Anthracene spiking severely affected microbial communities, suggesting dominance of nontolerant populations in this lightly-contaminated sediment. Although biostimulation and/or bioaugmentation treatments did not alleviate the anthracene toxic effects, the changes observed in microbial population and structure suggest that the proposed treatments might be promising to promote bacterial growth. Further works are still required to propose a more efficient strategy to stimulate biodegradation that takes into account the complex interactions between species for resource access.