Phagotrophic Protists Modulate Copper Resistance of the Bacterial Community in Soil.

Protist predation is a crucial biotic driver modulating bacterial populations and functional traits. Previous studies using pure cultures have demonstrated that bacteria with copper (Cu) resistance exhibited fitness advantages over Cu-sensitive bacteria under the pressure of protist predation. However, the impact of diverse natural communities of protist grazers on bacterial Cu resistance in natural environments remains unknown. Here, we characterized the communities of phagotrophic protists in long-term Cu-contaminated soils and deciphered their potential ecological impacts on bacterial Cu resistance. Long-term field Cu pollution increased the relative abundances of most of the phagotrophic lineages in Cercozoa and Amoebozoa but reduced the relative abundance of Ciliophora. After accounting for soil properties and Cu pollution, phagotrophs were consistently identified as the most important predictor of the Cu-resistant (CuR) bacterial community. Phagotrophs positively contributed to the abundance of a Cu resistance gene (copA) through influencing the cumulative relative abundance of Cu-resistant and -sensitive ecological clusters. Microcosm experiments further confirmed the promotion effect of protist predation on bacterial Cu resistance. Our results indicate that the selection by protist predation can have a strong impact on the CuR bacterial community, which broadens our understanding of the ecological function of soil phagotrophic protists.

Biotechnological potential of Bacillus sp. S26 for alleviation of abiotic and biotic stresses in vine.

Grapevine (Vitis spp.) is one of the most cultivated fruit plants in the world. Vineyard growers apply copper-based products in these crops to prevent fungal diseases, generating worries about Cu contamination in soils and food. In this context, this study identifies prokaryotic communities associated with grapevine plants grown under different levels of Cu-contaminated soils. Moreover, the study isolates new bacteria to improve Cu resistance in plants. Soil Cu content correlated inversely with operational taxonomic units (OTUs) belonging to the groups Acidobacteria (SubGroup 2), Latescibacteria, Pedosphaeraceae, and Candidatus Udaeobacter. A total of 14 new bacterial isolates were obtained from copper-contaminated soils. These isolates produced Indolic Compounds (IC) in a range of 25 to 96 µg mL- 1, highlighting bacterial strains S20 and S26 as the highest producers. These new bacteria also produced siderophores, highlighting strains S19 and S26, which removed 58 and 59% of Fe ions from the CAS complex, respectively. From the in vitro antagonistic activity against Colletotrichum spp. strains, the authors identified some bacterial strains that inhibited phytopathogen growth. Bacterial strain Bacillus sp. S26 was chosen for inoculation experiments in grapevine plants. This bacterial isolate improved the growth of grapevine plants in Cu-contaminated soils. However, growth promotion did not occur in unstressed plants. More studies are necessary for developing a new bioinoculant containing S26 cells aiming to reduce biotic and abiotic stresses in grapevine.

Effects of micro-/nano-hydroxyapatite and phytoremediation on fungal community structure in copper contaminated soil.

Micro-/nano-hydroxyapatite (MHA/NHA) has been used to reduce the concentration of available heavy metals and increase soil pH in the remediation of heavy metal-contaminated soils. However, little is known about the effects of MHA and NHA on soil fungal communities and function. In this study, fungal community composition was characterized from copper-contaminated soils amended with MHA, NHA and three other classic amendments combined with Elsholtzia splendens during a 3-year immobilization experiment. High-throughput sequencing results showed that applications of MHA increased the richness and diversity of the fungal community, which was opposite the results of NHA. SIMPER analysis indicated that both the relative abundance of fungi associated with biosorption and plant growth promotion increased, whereas the relative abundance of fungi related to bioleaching and potential pathogens decreased after applying MHA. Redundancy (RDA) analysis revealed that the soil pH was a crucial environmental factor in the succession of fungal communities. In addition, the results of functional prediction via FUNGuild suggested that the application of MHA had the potential to reduce the risk of pathogens infecting animals and plants in the soil but that NHA had some environmental risks. Overall, fungal community showed a synergistic effect of immobilization with the test amendments, and MHA was better for the remediation of heavy metal-contaminated soils than the other test amendments.

Contribution of Piriformospora indica on improving the nutritional quality of greenhouse tomato and its resistance against cu toxicity after humic acid addition to soil.

Protected cultivation has a significant contribution in vegetable production. We investigated whether humic acid addition to soil and Piriformospora indica can improve the nutritional quality of greenhouse tomato. We conducted a pot experiment, in which the effects of P. indica inoculation, humic acid addition, and Cu spiking to soil (0, 120, 240, and 500 ppm Cu) were tested. Humic acid addition to soil spiked with 500 ppm Cu decreased the Cu concentration in the fruits of plants inoculated with P. indica from 0.65 to 0.40 mg 100 g Fw-1, which is still above the maximum allowed limits of Cu in tomato by World Health Organization (WHO). The lycopene and ascorbic acid content of tomato fruits were consistently improved by humic acid addition and P. indica inoculation. The antioxidant enzymes' activity changed in response to humic acid addition, Cu spiking to soil, and P. indica inoculation. With increasing Cu level up to 240 ppm, the activity of superoxide dismutase (SOD) and peroxidase (POD) increased significantly. However, with spiking more Cu to soil, the activity of antioxidant enzymes reduced and the MDA content increased significantly. Addition of humic acid to soil and/or presence of P. indica increased the activity of antioxidant enzymes when the soil spiked with 500 ppm Cu. This study indicated that addition of P. indica and humic acid to the soil can enhance the nutritional quality of greenhouse tomato by reduction of Cu toxicity as a common pollutant in the greenhouse media and increasing the antioxidant content of fruits.

Co-Amendment of S and Si Alleviates Cu Toxicity in Rice (Oryza Sativa L.) Grown on Cu-Contaminated Paddy Soil.

With irrigation using waste water, application of sewage sludge, and development of mine exploration, copper (Cu) contamination in some paddy fields has become increasingly serious. A greenhouse pot experiment was conducted using a factorial design with three sulfur (S) application rates (i.e., 0, 0.013, and 0.026 g S kg-1 soil) and three silicon (Si) application rates (i.e., 0, 0.05, and 0.1 g Si kg-1 soil) to test the effect of co-amendment of S and Si on alleviating Cu contamination in paddy soil. There were significant interaction effects between S and Si on soil Cu speciation and Cu uptake by rice plants (except brown rice). Sulfur addition decreased the content of soil-exchangeable Cu, whereas Si addition decreased the content of soil-reducible Cu, suggesting that co-amendment of S and Si generally reduced Cu availability. Copper was biominimized in the soil-rice plant system and rice root had the greatest Cu concentration (163⁻285 mg kg-1). Co-amendment of S and Si decreased the translocation of Cu from soil to rice root, possibly due to decreased soil Cu mobility and enhancement of the formation of iron plaque on rice root. Co-amendment of S-Si at a rate of 0.013 (S)⁻0.1 (Si) g kg-1 soil, respectively, was the optimal among all treatments.

Fingerprinting two metal contaminants in streams with Cu isotopes near the Dexing Mine, China.

Transition metal isotope signatures are becoming useful for fingerprinting sources in surface waters. This study explored the use of Cu isotope values to trace dissolved metal contaminants in stream water throughout a watershed affected by mining by-products of the Dexing Mine, the largest porphyry Cu operation in Asia. Cu isotope values of stream water were compared to potential mineral sources of Cu in the mining operation, and to proximity to the known Cu sources. The first mineral source, chalcopyrite, CuFeS2 has a 'tight' cluster of Cu isotope values (-0.15‰ to +1.65‰; +0.37 ± 0.6‰, 1σ, n=10), and the second mineral source, pyrite (FeS2), has a much larger range of Cu isotope values (-4‰ to +11.9‰; 2.7 ± 4.3‰, 1σ, n=16). Dissolved Cu isotope values of stream water indicated metal derived from either chalcopyrite or pyrite. Above known Cu mineralization, stream waters are approximately +1.5‰ greater than the average chalcopyrite and are interpreted as derived from weathering of chalcopyrite. In contrast, dissolved Cu isotope values in stream water emanating from tailings piles had Cu isotope values similar to or greater than pyrite (>+6‰, a common mineral in the tailings). These values are interpreted as sourced from the tailings, even in solutions that possess significantly lower concentrations of Cu (<0.05 ppm). Elevated Cu isotope values were also found in two soil and two tailings samples (δ(65)Cu ranging between +2 to +5‰). These data point to the mineral pyrite in tailings as the mineral source for the elevated Cu isotope values. Therefore, Cu isotope values of waters emanating from a clearly contaminated drainage possess different Cu isotope values, permitting the discrimination of Cu derived from chalcopyrite and pyrite in solution. Data demonstrate the utility of Cu isotopic values in waters, minerals, and soils to fingerprint metallic contamination for environmental problems.

Impact of copper toxicity on stone-head cabbage (Brassica oleracea var. capitata) in hydroponics.

Arable soils are frequently subjected to contamination with copper as the consequence of imbalanced fertilization with manure and organic fertilizers and/or extensive use of copper-containing fungicides. In the present study, the exposure of stone-head cabbage (Brassica oleracea var. capitata) to elevated Cu(2+) levels resulted in leaf chlorosis and lesser biomass yield at ≥2 µ M. Root nitrate content was not statistically affected by Cu(2+) levels, although it was substantially decreased at ≥5 µ M Cu(2+) in the shoot. The decrease in nitrate contents can be related to lower nitrate uptake rates because of growth inhibition by Cu-toxicity. Shoot sulfate content increased strongly at ≥2 µ M Cu(2+) indicating an increase in demand for sulfur under Cu stress. Furthermore, at ≥2 µM concentration, concentration of water-soluble non-protein thiol increased markedly in the roots and to a smaller level in the shoot. When exposed to elevated concentrations of Cu(2+) the improved sulfate and water-soluble non-protein thiols need further studies for the evaluation of their direct relation with the synthesis of metal-chelating compounds (i.e., phytochelatins).

Bile and liver metallothionein behavior in copper-exposed fish.

The present study analyzed metallothionein (MT) excretion from liver to bile in Nile Tilapia (Oreochromis niloticus) exposed to sub-lethal copper concentrations (2mgL(-1)) in a laboratory setting. MTs in liver and bile were quantified by spectrophotometry after thermal incubation and MT metal-binding profiles were characterized by size exclusion high performance liquid chromatography coupled to ICP-MS (SEC-HPLC-ICP-MS). Results show that liver MT is present in approximately 250-fold higher concentrations than bile MT in non-exposed fish. Differences between the MT profiles from the control and exposed group were observed for both matrices, indicating differential metal-binding behavior when comparing liver and bile MT. This is novel data regarding intra-organ MT comparisons, since differences between organs are usually present only with regard to quantification, not metal-binding behavior. Bile MT showed statistically significant differences between the control and exposed group, while the same did not occur with liver MT. This indicates that MTs synthesized in the liver accumulate more slowly than MTs excreted from liver to bile, since the same fish presented significantly higher MT levels in liver when compared to bile. We postulate that bile, although excreted in the intestine and partially reabsorbed by the same returning to the liver, may also release MT-bound metals more rapidly and efficiently, which may indicate an efficient detoxification route. Thus, we propose that the analysis of bile MTs to observe recent metal exposure may be more adequate than the analysis of liver MTs, since organism responses to metals are more quickly observed in bile, although further studies are necessary.