The effects of multiple metal contamination on ectomycorrhizal Scots pine (Pinus sylvestris) seedlings

Experiments were conducted to investigate the effects of single and multiple metal contamination (Cd, Pb, Zn, Sb, Cu) on Scots pine seedlings colonised by ectomycorrhizal (ECM) fungi from natural soil inoculum. Seedlings were grown in either contaminated field soil from the site of a chemical accident, soils amended with five metals contaminating the site, or in soil from an uncontaminated control site. Although contaminated and metal-amended soil significantly inhibited root and shoot growth of the Scots pine seedlings, total root tip density was not affected. Of the five metals tested in amended soils, Cd was the most toxic to ECM Scots pine. Field-contaminated soil had a toxic effect on ECM fungi associated with Scots pine seedlings and caused shifts in ECM species composition on ECM seedlings. When compared to soils amended with only one metal, soils amended with a combination of all five metals tested had lower relative toxicity and less accumulation of Pb, Zn and Sb into seedlings. This would indicate that the toxicity of multiple metal contamination cannot be predicted from the individual toxicity of the metals investigated.

In vitro mesenchymal stem cell responses on laser-welded NiTi alloy

The biocompatibility of NiTi after laser welding was studied by examining the in vitro (mesenchymal stem cell) MSC responses at different sets of time varying from early (4 to 12 h) to intermediate phases (1 and 4 days) of cell culture. The effects of physical (surface roughness and topography) and chemical (surface Ti/Ni ratio) changes as a consequence of laser welding in different regions (WZ, HAZ, and BM) on the cell morphology and cell coverage were studied. The results in this research indicated that the morphology of MSCs was affected primarily by the topographical factors in the WZ: the well-defined and directional dendritic pattern and the presence of deeper grooves. The morphology of MSCs was not significantly modulated by surface roughness. Despite the possible initial Ni release in the medium during the cell culture, no toxic effect seemed to cause to MSCs as evidenced by the success of adhesion and spreading of the cells onto different regions in the laser weldment. The good biocompatibility of the NiTi laser weldment has been firstly reported in this study.

Intravitreal Injection of Normal Saline Induces Retinal Degeneration in the C57BL/6J Mouse

Purpose: To investigate the adverse effect of intravitreal injection of normal saline (NS) and phosphate buffered saline (PBS) in mouse eyes.

Methods: NS or PBS was injected intravitreally into C57BL/6J mouse eyes. Retinal lesions were monitored by fundus imaging, spectral-domain optical coherence tomography (SD-OCT), and histological investigations. Retinal immune gene expression was determined by real-time polymerase chain reaction (PCR). The toxic effect of NS and PBS or retinal protein from NS- or PBS-injected eyes on retinal pigment epithelium (RPE) was tested in B6-RPE-07 mouse RPE cell cultures.

Results: Intravitreal injection of NS dose-dependently induced localized retinal lesion in mice. Histological investigations revealed multiple vacuoles in photoreceptor outer segments and RPE cells. The lesions recovered over time and by 3 weeks post injection the majority of lesions vanished in eyes receiving 1 μl NS. Inflammatory genes, including TNF-α, IL-1β, IL-6, iNOS, and VEGF were upregulated in NS injected eyes. Intravitreal injection of PBS did not cause any pathology. The treatment of B6-RPE07 cells with 30% PBS or 30% NS did not affect RPE viability. However, incubation of 1-μg/ml retinal protein from NS-injected eyes, but not PBS-injected eyes induced RPE cell death.

Conclusion: NS is toxic to the C57BL/6J mouse retina and should not be used as a vehicle for intraocular injection. PBS is not toxic to the retina and is a preferred vehicle.

Translational Relevance: NS is not a physiological solution for intraocular injection in the C57BL/6J mice and questions its suitability for intraocular injection in other species, including human.

Cytotoxic assessment of the regulated, co-existing mycotoxins aflatoxin B1, fumonisin B1 and ochratoxin, in single, binary and tertiary mixtures

Aflatoxin B1 (AFB1), ochratoxin A (OTA) and fumonisin B1 (FB1) are contaminants which have been shown to regularly co-occur in a range of foods. However, only a small number of studies have evaluated the interactive effect of binary and tertiary mycotoxins. The present study evaluated the effects of low levels of each mycotoxin in combination at their EU regulatory limits. Toxic effect with respect to cell viability was measured by MTT and neutral red assays, assessing mitochondria and lysosome integrities respectively. Individual toxicity showed that OTA (10 μg/ml) was the most cytotoxic mycotoxin in all three cell lines studied (caco-2, MDBK and raw 264.7). Binary combinations were cytotoxic to the MDBK cell line in the order [OTA/FB1] > [AFB1/FB1] > [AFB1/OTA], whilst all effects observed were classified as being additive. Tertiary combinations of AFB1, FB1 and OTA at the EU regulatory limits were tested and not found to exhibit measurable cytotoxicity in MDBK, caco-2 or raw 264.7 cells. However by increasing these concentrations above the legal limits to OTA (3 μg/ml), FB1 (8 μg/ml) and AFB1 (1.28 μg/ml), cytotoxicity was observed with up to 26% reduction in cell viability and synergistic effects were evident with regard to mitochondrial integrity. © 2014 Elsevier Ltd. All rights reserved.

Biosensor based toxicity dissection of tannery and associated environmental samples

Kenyan tannery and associated environmental samples were selected for ecotoxicological assessment. A tool-kit of techniques was developed, including whole-cell biosensor and chemical assays. A luminescence based bacterial biosensor (Escherichia coli HB101 pUCD607) (via a multi-copy plasmid) was used for toxicity assessment. Samples were manipulated prior to biosensor interrogation to identify the nature of the toxic contaminants. Untreated samples (before any manipulations) showed a strong toxic effect at the discharge point in comparison to other sampling points. Sparging was used to identify toxicity associated with volatile organics. The toxicity of contaminants, removed by treatment with activated charcoal was identified for all the sampling points except for those upstream of effluent discharges. Filtration identified toxicity associated with suspended solids. Changes in availability of toxic contaminants due to pH adjustment of most samples from the tannery effluent treatment pits were also associated with the extreme pH values (4.0 and 8.0). The approach used has highlighted the complexicity of toxic pollutants in effluent from the tanning industry and the dissection of toxicity points to possible remediation strategies for effluents from the tanning industry.

Bioavailability of 2,4-dichlorophenol associated with soil water-soluble humic material

Interaction of organic xenobiotics with soil water-soluble humic material (WSHM) may influence their environmental fate and bioavailability. We utilized bacterial assays (lux-based toxicity and mineralization by Burkholderia sp. RASC) to assess temporal changes in the bioavailability of [¹⁴C]-2,4-dichlorophenol (2,4-DCP) in soil water extracts (29.5 μg mL^-1 2,4-DCP; 840.2 μg mL^-1 organic carbon). HPLC determined and bioavailable concentrations were compared. Gel permeation chromatography (GPC) was used to confirm the association of a fraction (>50%) of [¹⁴C]-2,4-DCP with WSHM. Subtle differences in parameters describing 2,4-DCP mineralization curves were recorded for different soil-2,4-DCP contact times. Problems regarding the interpretation of mineralization data when assessing the bioavailability of toxic compounds are discussed. The lux-bioassay revealed a time-dependent reduction in 2,4-DCP bioavailability: after 7 d, less than 20% was bioavailable. However, GPC showed no quantitative difference in the amount of WSHM-associated 2,4-DCP over this time. These data suggest qualitative changes in the nature of the 2,4-DCP-WSHM association and that associated 2,4-DCP may exert a toxic effect. Although GPC distinguished between free- and WSHM-associated 2,4-DCP, it did not resolve the temporal shift in bioavailability revealed by the lux biosensor. These results stress that assessment of risk posed by chemicals must be considered using appropriate biological assays.

Retention of toxic chromium from aqueous phase by H3PO4-activated lignin: Effect of salts and desorption studies

In this study, the feasibility of using H₃PO₄-activated lignin for hexavalent chromium adsorption has been investigated. The composite of activated lignin was characterized using FTIR, XRD and SEM with EDAX analysis. It was observed that the pH had a strong effect on the adsorption capacity; adsorption of Cr(VI) was more favorable at acidic pH with maximum uptake at pH 2. The adsorption equilibrium data were best represented by Koble-Corrigan isotherm. The monolayer sorption capacity obtained from the Langmuir model was found to be 77.85 mg/g. Adsorption showed pseudo-second order rate kinetics and the process involving the rate-controlling step is complex as it involves both film and intraparticle diffusion processes. The NaOH desorbing agent was able to release approximately 84% of metal ions. Thermodynamic parameters showed that the sorption process is exothermic and non-spontaneous. The overall Cr(VI) retention on the activated lignin surface perhaps includes both the physical adsorption of Cr(VI) and the consequent reduction of Cr(VI) to Cr(III). (C) 2011 Elsevier B.V. All rights reserved.

Biosorption of toxic chromium from aqueous phase by lignin: mechanism, effect of other metal ions and salts

Surface reaction methodology was implicated in the optimization of hexavalent chromium removal onto lignin with respect to the process parameters. The influence of altering the conditions for removal of chromium(VI), for instance; solution pH, ionic strength, initial concentration, the dose of biosorbent, presence of other metals (Zn and Cu), presence of salts and biosorption-desorption studies, were investigated. It was found that the biosorption capacity of lignin depends on solution pH, with a maximum biosorption capacity for chromium at pH 2. Experimental equilibrium data were fitted to five different isotherm models by non-linear regression method, however, the biosorption equilibrium data were well interpreted by the Freundlich isotherm. The maximum biosorption capacities (q(max)) obtained using Dubinin-Radushkevich and Khan isotherms for Cr(VI) biosorption are 31.6 and 29.1 mg/g. respectively. Biosorption showed pseudo second order rate kinetics at different initial concentrations of Cr(VI). The intraparticle diffusion study indicated that film diffusion may be involved in the current study. The percentage removal of chromium on lignin decreased significantly in the presence of NaHCO3 and K2P2O7 salts. Desorption data revealed that nearly 70% of the Cr(VI) adsorbed on lignin could be desorbed using 0.1 M NaOH. It was evident that the biosorption mechanism involves the attraction of both hexavalent chromium (anionic) and trivalent chromium (cationic) onto the surface of lignin. (C) 2011 Elsevier B.V. All rights reserved.

The effect of pesticides on Byssus formation in the common mussel, Mytilus edulis

Seed mussels (Mytilus edulis) were exposed to a range of pesticides and PCBs, several of which caused a reduction in byssal attachment at higher concentrations. In queen scallops (Chlamys opercularis) byssus formation was similarly affected although this species was more sensitive than M. edulis. The sensitivity of mussels was greater at higher temperatures and decreased with increase in size. Of the compounds tested Endosulfan (organochlorine) was the most toxic, causing a 50% reduction in byssal attachment after 24 h at 0•45 mg/l. Trichlorphon (organophosphate) was the least toxic and did not affect byssal attachment at concentrations up to 30 mg/l. The probable cause of decreased byssal attachment is a reduction in pedal activity, although it is possible that direct interference with the synthesis or combination of byssus components may be involved. It is suggested that byssogenesis tests offer a rapid and convenient technique for the routine screening of potential marine pollutants.

Using SILAC proteomics to investigate the effect of the mycotoxin, alternariol, in the human H295R steroidogenesis model

The mycotoxin alternariol (AOH) is an important contaminant of fruits and cereal products. The current study sought to address the effect of a non-toxic AOH concentration on the proteome of the steroidogenic H295R cell model. Quantitative proteomics based on stable isotope labeling by amino acids in cell culture (SILAC) coupled to 1D-SDS-PAGE-LC-MS/MS was applied to subcellular-enriched protein samples. Gene ontology (GO) and ingenuity pathway analysis (IPA) were further carried out for functional annotation and identification of protein interaction networks. Furthermore, the effect of AOH on apoptosis and cell cycle distribution was also determined by the use of flow cytometry analysis. This work identified 22 proteins that were regulated significantly. The regulated proteins are those involved in early stages of steroid biosynthesis (SOAT1, NPC1, and ACBD5) and C21-steroid hormone metabolism (CYP21A2 and HSD3B1). In addition, several proteins known to play a role in cellular assembly, organization, protein synthesis, and cell cycle were regulated. These findings provide a new framework for studying the mechanisms by which AOH modulates steroidogenesis in H295R cell model. 

Toxic effects of cadmium and zinc on ectomycorrhizal colonization of scots pine (Pinus sylvestris L.) from soil inoculum

Scots pine seedlings colonized by ectomycorrhizal (ECM) fungi from natural soil inoculum were exposed to a range of Cd or Zn concentrations to investigate the effects of metals on ECM fungi-Scots pine associations in a realistic soil environment. Experiments focused on the relationship between the sensitivity of ECM fungi and their host plants, the influence of metals on ECM community dynamics on Scots pine roots, and the effects of metal exposure on ECM colonization from soil-borne propagules. Ectomycorrhizal colonization was inhibited by Cd and Zn, with a decrease in the proportion of ECM-colonized root tips. Shoot and root biomass, total root length, and total root-tip density, however, were unaffected by Cd or Zn. A decrease in the diversity of ECM morphotypes also occurred, which could have a negative effect on tree vigor. Overall, colonization by ECM fungi was more sensitive than seedling growth to Cd and Zn, and this could have serious implications for successful tree establishment on metal-contaminated soils.

Toxic interactions of metal ions (Cd2+, Pb2+, Zn2+ and Sb3- on in vitro biomass production of ectomycorrhizal fungi

A number of ectomycorrhizal (ECM) fungi, from sites uncontaminated by toxic metals, were investigated to determine their sensitivity to Cd^2-, Pb²⁺, Zn²⁺ and Sb^3-, measured as an inhibition of fungal biomass production. Isolates were grown in liquid media amended with the metals, individually (over a range of concentrations) and in combination (at single concentrations) to determine any significant interactions between the metals. Significant interspecific variation in sensitivity to Cd²⁺ and Zn²⁺ was recorded, while Pb²⁺ and Sb^3- individually had little effect. The presence of Pb²⁺ and Sb^3- in the media did however, ameliorate Cd²⁺ and Zn²⁺ toxicity in some circumstances. Interactions between Cd²⁺ and Zn²⁺ were investigated further over a range of concentrations. Zn²⁺ was found to significantly ameliorate the toxicity of Cd²⁺ to three of the four isolates tested. The influence of Zn²⁺ varied between ECM species and with the concentrations of metals tested.