Spatial distribution of persistent organic pollutants (POPs) in sediments contaminated by fiber material from pulp and paper industries

Sweden has had a long history of forestry activities together with pulp and paper operations. Historical discharges of organic-rich wastewater from pulp and paper factories have resulted in formation of large fiberbanks on the seafloor, which lately have been found to be contaminated with persistent organic pollutants (POPs). The aim of this study was to determine the levels of POPs in fiberbank sediments from two known contaminated sites (Väja and Sandviken) located in the Ångermanälven estuary in the county of Västernorrland, Sweden. The objective was to investigate the spatial distribution of POPs in these fiberbanks on both a horizontal and vertical axis. The sediment samples (n= 43) were collected on SGU’s survey vessel Ocean Surveyor in autumn 2017. The sediment samples were extracted using a Soxhlet followed by a multilayered silica gel clean-up column, prior to instrumental analysis with GC-MS/MS. The sediment’s concentration of hexachlorobenzene (HCB), 20 polychlorinated biphenyls (PCBs) and dichlorodiphenyltrichloroethane (DDT) and its transformation products (DDE and DDD) together referred to as DDX, were measured. At both locations, surface sediment samples (0-4 cm depth) generally presented low concentrations of POPs and organic carbon (TOC), likely due to recent sedimentation of fine clay material. At increased depth, contents of TOC, as well as, the levels of PCBs and DDX (sum of 6 DDT congeners) tended to increase. However, spatial distribution was found to be irregular within both fiberbank and fiber-rich sediment areas. The PCBs compositions, for each location, were found to be related to known technical grade products by using hierarchical cluster analysis. The toxicity of the target pollutants, found in the studied areas, was also assessed using the Norwegian quality standards for sediments. It was found that the HCB levels, for all the samples analyzed, were classified as having no effects on biota (Class II). In both areas, the state of the surface sediment, regarding the Ʃ p,p’-DDX and o,p’-DDT, was classified as good (Class II). However, as the sampling depth increased, many sites contained levels causing chronic effects of long-term exposure (Class III). Most of the levels of Ʃ7PCB in surface sediment samples were classified as having no toxic effects (Class II) with few samples classified as causing chronic effects of long-term exposure (Class III). For both Sandviken and Väja, the levels of Ʃ7PCB increased with sampling depth, and some sites contained levels which are classified as having acute toxic effects of short-term exposure (Class IV). To conclude, this study has contributed in generating important information about the distribution of hazardous contaminants in fiberbanks, and the risk on the immediate environments. The outcome from this study demonstrated how heterogeneous the contaminant levels can be in the vicinity of a pulp and paper factory. This should be taken into consideration when defining remediation strategies for fiber-impacted areas, like defining the site of a remediation project

Polluted lignocellulose-bearing sediments as a resource for marketable goods—a review of potential technologies for biochemical and thermochemical processing and remediation

Lignocellulose-bearing sediments are legacies of the previously unregulated wastewater discharge from the pulp and paper industry, causing large quantities of toxic organic waste on the Baltic Sea floor and on the bottom of rivers and lakes. Several km2 are covered with deposits of lignocellulosic residues, typically heavily contaminated with complex mixtures of organic and inorganic pollutants, posing a serious threat to human and ecological health. The high toxicity and the large volume of the polluted material are challenges for remediation endeavours. The lignocellulosic material is also a considerable bioresource with a high energy density, and due to its quantity, it could appeal to commercialization as feedstock for various marketable goods. This study sets out to explore the potential of using this polluted material as a resource for industrial production at the same time as it is detoxified. Information about modern production methods for lignocellulosic material that can be adapted to a polluted feedstock is reviewed. Biochemical methods such as composting, anaerobic digestion, as well as, thermochemical methods, for instance, HTC, HTL, pyrolysis, gasification and torrefaction have been assessed. Potential products from lignocellulose-bearing sediment material include biochar, liquid and gaseous biofuels, growing substrate. The use of a contaminated feedstock may make the process more expensive, but the suggested methods should be seen as an alternative to remediation methods that only involve costs. Several experiments were highlighted that support the conception that combined remediation and generation of marketable goods may be an appropriate way to address polluted lignocellulose-bearing sediments. Graphic abstract: [Figure not available: see fulltext.]

Genome-wide mouse mutagenesis reveals CD45-mediated T cell function as critical in protective immunity to HSV-1.

International audienceHerpes simplex encephalitis (HSE) is a lethal neurological disease resulting from infection with Herpes Simplex Virus 1 (HSV-1). Loss-of-function mutations in the UNC93B1, TLR3, TRIF, TRAF3, and TBK1 genes have been associated with a human genetic predisposition to HSE, demonstrating the UNC93B-TLR3-type I IFN pathway as critical in protective immunity to HSV-1. However, the TLR3, UNC93B1, and TRIF mutations exhibit incomplete penetrance and represent only a minority of HSE cases, perhaps reflecting the effects of additional host genetic factors. In order to identify new host genes, proteins and signaling pathways involved in HSV-1 and HSE susceptibility, we have implemented the first genome-wide mutagenesis screen in an in vivo HSV-1 infectious model. One pedigree (named P43) segregated a susceptible trait with a fully penetrant phenotype. Genetic mapping and whole exome sequencing led to the identification of the causative nonsense mutation L3X in the Receptor-type tyrosine-protein phosphatase C gene (Ptprc(L3X)), which encodes for the tyrosine phosphatase CD45. Expression of MCP1, IL-6, MMP3, MMP8, and the ICP4 viral gene were significantly increased in the brain stems of infected Ptprc(L3X) mice accounting for hyper-inflammation and pathological damages caused by viral replication. Ptprc(L3X) mutation drastically affects the early stages of thymocytes development but also the final stage of B cell maturation. Transfer of total splenocytes from heterozygous littermates into Ptprc(L3X) mice resulted in a complete HSV-1 protective effect. Furthermore, T cells were the only cell population to fully restore resistance to HSV-1 in the mutants, an effect that required both the CD4⁺ and CD8⁺ T cells and could be attributed to function of CD4⁺ T helper 1 (Th1) cells in CD8⁺ T cell recruitment to the site of infection. Altogether, these results revealed the CD45-mediated T cell function as potentially critical for infection and viral spread to the brain, and also for subsequent HSE development

The HSV-1 susceptibility associated with the <i>Ptprc^L3X</i> genotype is independent of the infection route and HSV-1 strain.

<p>(<b>A</b>) <i>Ptprc^L3X</i> and heterozygous littermates were infected i.n. with 5×10³ pfu of HSV-1 and monitored for survival. (<b>B and C</b>) <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice were infected i.p. with 1×10⁴ pfu of either McIntyre (<b>B</b>) or F strain (<b>C</b>) and monitored for survival. n≥6 for each group and data represent at least two independent experiments. (<b>A, B and C</b>).</p

ENU-screening strategy.

<p>(<b>A</b>) Schematic representation of the breeding strategy used to produce G3 pedigrees. Details of the breeding procedure are described in the <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003637#s4" target="_blank">Materials and Methods</a> (<i>ENU mutagenesis and breeding</i> section). (<b>B</b>) G3 pedigrees were screened for their susceptibility to HSV-1 infection. Mice were infected i.p. with 1×10⁴ pfu of HSV-1 strain 17 and survival was monitored for two weeks post-infection; susceptible A/J and BALB/c control mice are represented by a blue line, resistant C57BL/6J and C57BL/10J control mice are represented by a green line, all G3 mice by a red line and <i>P43</i> G3s derived from two G2 daughters (G2a and Gb) and one G1 male by brown and black lines. “n” indicates the number of infected mice for each group.</p

CD4⁺, without CD8⁺ T cells, are incapable to protect mice against lethal HSV-1 infection.

<p>(<b>A</b>) Schematic representation of the adoptive transfer strategy. Details of this procedure are described in the <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003637#s4" target="_blank">Materials and Methods</a> (<i>Cell transfer experiments and NK depletion</i> section). (<b>B</b>) <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice received 2×10⁷ total splenocytes from either <i>Ptprc^L3X/+</i> or <i>Ptprc^L3X</i> mice. After two hours, these mice were infected i.p. with 1×10⁴ pfu of HSV-1 and their survival was monitored for two weeks (n = 6). (<b>C</b>) <i>Ptprc^L3X/+</i> mice were treated with either anti-asialo GM1 antibody or PBS. After 24 hours, these mice were infected i.p. with 1×10⁴ pfu of HSV-1 and their survival was monitored for two weeks (n = 6). The injection of either anti-asialo GM1 antibody or PBS was performed every three days until the experimental endpoint. (<b>D</b>) <i>Ptprc^L3X</i> mice received either 5×10⁶ T cells or 1.2×10⁷ B cells from <i>Ptprc^L3X/+</i> mice. <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice that received only PBS respectively correspond to the positive and negative controls. After two hours, all mice were infected i.p. with 1×10⁴ pfu of HSV-1 and survival was monitored for two weeks (n≥6). (<b>E</b>) <i>Ptprc^L3X</i> mice received either 2.5×10⁶ CD8⁺ or 2.5×10⁶ CD4⁺ T cells from either <i>Ptprc^L3X/+</i> or <i>B6.H2-D^bK^b</i> knock-out (only for the CD4⁺ T cells transfer) mice, whose <i>H2-D^bK^b</i> make them depleted in CD8⁺ T cells (CD8^−/−). <i>Ptprc^L3X/+</i> mice that received only PBS correspond to the positive control. After two hours, all mice were infected i.p. with 1×10⁴ pfu of HSV-1 and survival was monitored for two weeks (n≥6). Data represent two independent experiments. (<b>B–E</b>).</p

Expression of viral and inflammatory host genes in brain tissue collected from <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> infected mice.

<p>(<b>A</b>) <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice were infected i.p. with 1×10⁴ pfu of HSV-1. At the indicated day (D3, D6 and D10), total brains were harvested and mechanically homogenized for a plaque assay (n≥3). Viral titers are presented as pfu/gram of brain. The dotted line indicates the threshold of detection. (<b>B and C</b>) <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice were infected i.p. with 1×10⁴ pfu of HSV-1 (n = 9). Following infection, these mice were weighed two times daily. The brain stems of <i>Ptprc^L3X</i> mice that had lost at least 15% of their pre-infection weight were harvested. <i>Ptprc^L3X/+</i> mice were sacrificed and their brain stems were collected at days 7, 9, and 11 p.i. (n = 3 for each time point). The expression of the <i>ICP4</i> viral gene (<b>B</b>) and the indicated cellular genes (<b>C, upper panels</b>) was normalized to that of <i>hprt</i>. Data are presented as a fold increase relative to infected B6 samples. *, p-value (p)<0.05 and **, p<0.005. Correlations of expression levels were determined by comparing <i>ICP4</i> and the indicated cellular genes (<b>C, lower panels</b>).</p

Important effect of IFN-γ production by the CD4⁺ T cells.

<p>(<b>A, B</b>) <i>Ptprc^L3X</i> mice received 2.5×10⁶ CD4⁺ T cells from <i>Ptprc^L3X/+</i> mice. <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice that received only PBS correspond to the positive and negative controls, respectively. After two hours and when indicated, mice were infected i.p. with 1×10⁴ pfu of HSV-1 and sacrificed at days 2 and 7 p.i (n = 3 for each group). At the indicated day, peritoneal cells were collected, stained for CD45.2, CD3, CD4 and CD8, quantified by FACS and gated on CD45.2⁺CD3⁺ (A, left panels, only shown for <i>Ptprc^L3X</i> mice at day 7 p.i.); they were then analyzed based on their CD4 and CD8 expression profiles (<b>A, right panel,</b> only shown for <i>Ptprc^L3X</i> infected mice that received CD4+ T cells from <i>Ptprc^L3X/+</i> mice, day 7 p.i.). Proportion of CD45.2⁺CD3⁺ T cells was also represented as a percentage of total peritoneal cells (<b>B, left panel</b>), while expression of CD4 and CD8 were represented as a percentage of total CD45.2⁺CD3⁺ T cells (<b>B, right panel, left axis</b>). Triangles and squares correspond to <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice, respectively. The expression of the <i>ICP4</i> viral gene (<b>B, right panel, right axis</b>) was determined in total peritoneal cells collected from <i>Ptprc^L3X/+</i> mice at days 2 and 7 p.i. <i>ICP4</i> expression was normalized to that of <i>hprt</i>. Data are presented as a fold increase relative to infected <i>Ptprc^L3X/+</i> samples at day 7 p.i. (<b>C</b>) <i>Ptprc^L3X/+</i> and <i>IFN-γ^−/−</i> mice were infected i.p. with 1×10⁴ pfu of HSV-1 and sacrificed at day 7 p.i. Then, peritoneal cells were collected and were gated on CD45.2⁺CD3⁺. CD4 and CD8 expression are presented as a percentage of total CD45.2⁺CD3⁺ cells and are averaged over three animals. (<b>D</b>) <i>Ptprc^L3X</i> mice received 2.5×10⁶ CD4⁺ T cells from <i>IFN-γ^−/−</i> mice. <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice that received only PBS respectively correspond to the positive and negative controls. After two hours, all mice were infected i.p. with 1×10⁴ pfu of HSV-1 and survival was monitored for two weeks (n≥5).</p

Immunological phenotyping.

<p>(<b>A, B, C and D</b>) The spleen of <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice was collected and specific immune cell populations were analyzed by FACS (n = 3). Isolated splenocytes were stained for CD3, DX5 and CD19 as well as IgD and IgM; their expressions were quantified by FACS and represented as a percentage of total cells. (<b>E</b>) Thymocytes isolated from <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> mice were stained for CD4 and CD8 and their expression were represented as a percentage of total cells. (<b>F</b>) Thymocytes were gated on CD4⁻CD8⁻ (double negative, DN); they were then analyzed based on their CD25 and CD44 expression profiles (DN1: CD25⁻CD44⁺; DN2: CD25⁺CD44⁺; DN3: CD25⁺CD44⁻; DN4: CD25⁻CD44⁻). CD25 and CD44 expression are presented as a percentage of total CD4⁻CD8⁻ double negative cells and are averaged over three animals. *, p<0.05; **, p<0.005 and ***, p<0.0005. <i>Ptprc^L3X/+</i> and <i>Ptprc^L3X</i> are shown in white and grey, respectively (<b>A, B, C, D, E and F</b>).</p