Cd, Hg, Pb, and As in European species of wild growing forest landscape fungi : a review

Kadmij (Cd), živo srebro (Hg), svinec (Pb) in arzen (As) so kovine, ki se naravno ali kot posledica človekove dejavnosti pojavljajo v okolju, tudi v gozdni krajini, kjer so rastišča številnih evropskih vrst gliv. Namen članka je bil pripraviti pregled vrst in količin izbranih kovin v trosnjakih gliv terprimerjati lastne raziskave, opravljene v različno onesnaženih območjih v Sloveniji (Zgornja Mežiška, Šaleška in Poljanska dolina), s podatki evropskih raziskav. Vsebnosti kovin v trosnjakih gliv iz neonesnaženih območij pravilomanajdemo v naslednjih intervalih: <0,5 mg/kg suhe teže (Cd), < 0,5 mg/kg do 10 mg/kg suhe teže (Hg), < 0,5 mg/kg do 5 mg/kg suhe teže (Pb) in < 0,5 mg/kg do 1 (2) mg/kg suhe teže (As). Na podlagi pregleda vsebnosti izbranih kovin v trosnjakih gliv ugotavljamo, da sta problematični kovini predvsem Cd in Hg. Omenjeni kovini lahko dosegata velike vsebnosti celo v glivah, ki rastejo v neonesnaženih območjih. Za vse analizirane kovin je značilno, da v trosnjakih gliv iz močno onesnaženih območji dosegajo velike, celo ekstemne vsebnosti, ki nekajkrat prekoračujejo vsebnosti iz neonesnaženih območij. Upoštevaje primerjavo z evropskimi raziskavami ugotavljamo, da je Zgornja Mežiška dolina obremenjena s Pb in Cd, Šaleška dolina pa s Cd in As.Metals, which originate from anthropogenic and natural activities, frequently occur in forest landscape with habitats of many European species of wild growing fungi. The presented review focuses on cadmium (Cd), mercury (Hg), lead (Pb), and arsenic (As) levels in fruiting bodies of wild growing European species of fungi of forest landscape. Furthermore, a comparison with studies of this kind performed in Slovenia was made with the aim to assess themetals levels in fungi from differently polluted areas in Slovenia (the Upper Meža Valley, the Šalek Valley, the Poljana Valley). The usual reported levels for most species grown in unpolluted areas are in the following ranges:Cd: < 0,5 mg/kg - 5 mg/kg dry weight (dw), Hg: < 0,5 mg/kg - 10 mg/kg dw, Pb: < 0,5 mg/kg - 5 mg/kg dw, As: < 0,5 mg/kg -1 (2) mg/kg dw (As), respectively. The presented data reveal that cadmium (Cd) and mercury (Hg) have probably been the most detrimental trace elements in fruiting bodies, which can reach increased levels even in unpolluted areas. It is evident for all analyzed trace elements that values can considerably increase in fungi picked in severely polluted areas. According to data regarding Slovene studies and comparison with other European studies, it is obvious that the Šalek Valley is enriched with Cd and As, while the Upper Meža Valley is considerably polluted with Pb and Cd

Expressions of PD-L1 on dog tumor cell lines.

<p>The expression of PD-L1 was evaluated by flow cytometric analysis using anti-PD-L1 mAb 4G12-C1. -; <3% positive, +; 3–30% positive, ++; 30–60% positive, +++; >60% positive.</p><p>*Cells were incubated with recombinant canine IFN-γ (100 ng/ml) for 24 h before the analysis.</p

Expression of PD-L1 on dog tumor cells.

<p>(A) Representative data for the analysis of PD-L1 expression on dog tumor cell lines (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098415#pone-0098415-t003" target="_blank">Table 3</a>). Cells maintained in the medium or those stimulated by IFN-γ (100 ng/mL) for 24 h were stained with anti-PD-L1 monoclonal antibody 4G12-C1 or isotype control antibody (rat IgG2a). Black line, medium/4G12-C1; red line, IFN-γ/4G12-C1; shaded area, medium/isotype control; vertical-striped area, IFN-γ/isotype control. (B) Tumor tissues excised surgically from clinical cases of dog tumors were treated with collagenase, and a tumor single cell suspension was obtained. To reduce the effect of collagenase on the degradation of PD-L1 and to restore the cell surface PD-L1, the tumor cells were cultured in the medium for 24 h before FACS analysis. Lymphocytes obtained from healthy dogs were used as a negative control to confirm that the collagenase and culture treatment would not upregulate the PD-L1 expression. The histogram shows the expression of PD-L1 on each tumor cell. Solid line, 4G12-C1; shaded area, isotype control. AS, angiosarcoma; HCC, hepatocellular carcinoma; SC, squamous carcinoma; and BA, breast adenocarcinoma. Details of each tumor samples are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098415#pone-0098415-t004" target="_blank">Table 4</a>.</p

The prevalence of <i>B. miyamotoi</i> and LD borreliae in <i>Ixodes</i> ticks in Hokkaido.

<p>The asterisk shows significant difference: * P<0.01, by Fisher's exact test.</p

Sequence analysis of canine PD-1.

<p>(A) Nucleic acid and deduced amino acid sequences of canine PD-1 cDNA. Canine PD-1 cDNA encodes for a 288 amino acid polypeptide. Predicted N-glycosylation sites in the amino acid sequence of canine PD-1 are doubly underlined. (B) Multiple sequence alignment of vertebrate PD-1 amino acid sequences. Predicted domains and motifs of canine PD-1 are shown in the figure. Signal peptide, 1–24; IgV domain, 39–145; transmembrane domain, 170–192; ITIM, 223–228; ITSM, 246–253. (C) Phylogenetic tree of the canine PD-1 sequence in relation to those of other vertebrate species. The bootstrap consensus tree was inferred from 1000 replicates (the numbers next to the branches indicate the bootstrap percentage). The scale indicates the divergence time. (D) Schematic image of predicted functional motifs in canine PD-1. Canine PD-1 consists of an extracellular region, a transmembrane region, and an intracellular region.</p

Prevalence of borreliae in ticks.

<p>*The numbers include mixed infections.</p

<i>Borrelia miyamotoi</i> strains used in this study.

<p><i>Borrelia miyamotoi</i> strains used in this study.</p

Monoclonal antibodies which recognize canine PD-L1.

<p>(A) Cross-reactivities of antibovine PD-L1 monoclonal antibodies. Binding abilities of recently established anti-boPD-L1 monoclonal antibodies to canine PD-L1 were examined by flow cytometry. Three anti-boPD-L1 monoclonal antibody clones, 4G12-C1 (rat IgG2a), 5A2-A1 (rat IgG1), and 6G7-E1 (rat IgM), were tested and all the three clones were found to recognize canine PD-L1. Rat IgG2a, rat IgG1, and rat IgM were used as isotype-matched negative controls. (a) cPD-L1–EGFP–expressing Cos7 cells and (b) dog PBMCs stimulated with PMA/ionomysin for 3 days were stained with anti-boPD-L1 monoclonal antibodies (10 µg/mL) or isotype-matched control antibodies. Red line, 4G12-C1; blue line, 5A2-A1; green line, 6G7-E1; shaded area, rat IgG2a; vertical-striped area, rat IgG1; horizontal-striped area, rat IgM. (B) Blockade of cPD-1/cPD-L1 binding by anti-PD-L1 monoclonal antibody 6G7-E1. cPD-L1–EGFP–expressing cells were preincubated with anti-PD-L1 antibody and then cPD-1–Ig bindings were evaluated by flow cytometry. (a) Blocking effect of anti-PD-L1 monoclonal antibody 6G7-E1 on cPD-1/cPD-L1 binding. Five microgram per milliliter of isotype-matched control antibody (rat IgM) could not affect the cPD-1/cPD-L1 binding (left panel), whereas the same concentration of 6G7-E1 significantly blocked the Ig binding (right panel). (b) Representative histogram of the flow cytometric analysis. Shaded area, isotype control (5 µg/mL); solid line, anti-PD-L1 monocolonal antibody 6G7-E1 (5 µg/mL). (c) Dose-dependent blocking effect of 6G7-E1 on cPD-1/cPD-L1 binding. Cells were preincubated with 6G7-E1 or isotype control antibody at various concentrations (0.5, 1.0, 2.5, 5.0 µg/mL) and Ig binding was analyzed by flow cytometry. Each point indicates the average value of relative MFI obtained from three independent experiments (compared to no antibody control, error bar; SEM). Statistical significance was evaluated by Tukey’s test (*<i>p</i><0.05, between the 0 µg/mL and the 1 µg/mL of anti-PD-L1 antibody treatment group and between the 1 µg/mL and the 5 µg/mL of anti-PD-L1 group. †<i>p</i><0.05, between the each concentration of anti-PD-L1 group and the same concentration of isotype control group).</p

Phylogenetic analysis of RF borreliae based on <i>16 S rDNA</i> of <i>Borrelia</i> spp.

<p>The phylogenetic tree of <i>16 S rDNA</i> was constructed. The phylogenetic branches were supported in >70% by the bootstrap analysis. The bar indicates the percentage of sequence divergence. Sequences in this study were shown in bold type. If possible, clone or strain name, isolation source, and country were described in the case of <i>B. miyamotoi</i>. Numbers in parentheses indicate Accession Numbers in GenBank.</p

PD-L1 expression on dog tumor tissues.

<p>Immunohistochemical analysis was performed using anti-PD-L1 monoclonal antibody 5A2-A1 or isotype control antibody (rat IgG1). (A–B) Representative immunohistochemical staining of melanoma. (C–D) Representative immunohistochemical staining of mastocytoma. (E–F) Representative immunohistochemical staining of renal cell carcinoma.</p