Article pubs.acs.org/est
The Role of Metal Nanoparticles in Influencing Arbuscular Mycorrhizal Fungi Effects on Plant Growth Youzhi Feng,† Xiangchao Cui,† Shiying He,‡ Ge Dong,§ Min Chen,† Junhua Wang,† and Xiangui Lin†,* †
State Key Laboratory of Soil and Sustainable Agriculture, Institute of Soil Science, Chinese Academy of Sciences, Nanjing, 210008, Jiangsu Province, P.R. China ‡ Insititute of Agricultural Resources and Environment, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, Jiangsu Province, P.R. China § School of Biological Science and Medical Engineering, Southeast University, Nanjing, 210096, Jiangsu Province, P.R. China S Supporting Information *
ABSTRACT: A knowledge gap still remains concerning the in situ influences of nanoparticles on plant systems, partly due to the absence of soil microorganisms. Arbuscular mycorrhizal fungi (AMF) can form a mutualistic symbiosis with the roots of over 90% of land plants. This investigation sought to reveal the responses of mycorrhizal clover (Trifolium repens) to silver nanoparticles (AgNPs) and iron oxide nanoparticles (FeONPs) along a concentration gradient of each. FeONPs at 3.2 mg/kg significantly reduced mycorrhizal clover biomass by 34% by significantly reducing the glomalin content and root nutrient acquisition of AMF. In contrast, no negative effects of AgNPs at concentrations over 0.1 mg/kg were observed; however, AgNPs at 0.01 mg/kg inhibited mycorrhizal clover growth. In response to the elevated AgNPs content, the ability of AMF to alleviate AgNPs stress (via increased growth and ecological behaviors) was enhanced, which decreased Ag content and the activities of antioxidant enzymes in plants. These results were further supported by X-ray microcomputed tomography. Our findings suggest that in soil ecosystem, the influence of nanometals on plant systems would be more complicated than expected, and more attention should be focused on plant responses in combination with those of soil microorganisms.
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INTRODUCTION Nanoparticles, which measure from 1 to 100 nm in diameter, have a greater surface area than bulk materials, yielding a higher proportion of atoms on the surface relative to the interior and resulting in a higher surface reactivity.1 Due to their unique physicochemical properties, nanoparticles are increasingly used in a wide range of technical applications and consumer products. For example, silver nanoparticles (AgNPs) are currently being used in numerous consumer products including textiles, personal care products, food storage containers, laundry additives, home appliances, paints, and even food supplements.2 Iron oxide nanoparticles (FeONPs) are also one of the most widely used nanoparticles in biomedical and environmental applications such as magnetic resonance imaging, targeted delivery of drugs, targeted destruction of tumor tissue, magnetic bioseparations, and pollution remediation.3−6 As nanoparticles enter large-scale applications, their release to the environment is inevitable. Because the high reactivity of nanoparticles may interfere with many natural processes in the ecosystem, biosafety studies are encouraged.7 Plants are an essential component of the terrestrial ecosystem and play critical roles in the fate and transport of nanoparticles throughout the environment via uptake and bioaccumulation.8 Therefore, plant responses to nanoparticles are of great interest © 2013 American Chemical Society
and aid our understanding of the consequences of introducing nanoparticles into ecosystems.9−13 Stampoulis et al.14 reported that AgNPs (20 μm spatial resolution, we can use red and green colors to represent the mycorrhizal root system and the sand matrix (Figure 6). Consistent with the results observed for the belowground biomass, a less developed root system was observed in plants treated with a low concentration of AgNPs compared with those treated with AMF. The density of the red color obviously increased with an increase in AgNPs content, indicating that the mycorrhizal root system was more developed. Concomitantly, the porosity rate was decreased (Table 2), implying that AMF optimizes the structure of the medium for the growth of mycorrhizal plants. These results support our speculation that elevated levels of AgNPs can enhance the effectiveness of AMF stress alleviation. Relatively speaking, FeONPs amendments had limited influence on the root system; a pairwise comparison revealed that mycorrhizal root systems treated with FeONPs were relatively better developed than those treated with AgNPs (Figure 6). The density of the red color appeared to decrease, and the root system (Figure 6) was less developed only under high concentrations of FeONPs. These images imply that FeONPs do not influence AMF growth. However, the porosity rate of the growth medium was enhanced with an increase in FeONPs concentration (Table 2). This inconsistency might result from the decrease in AMF-excreted glomalin (Figure 4a), which is not beneficial for the optimization of the growth medium or for plant growth. These results also support our speculation that FeONPs inhibit plant growth by adversely affecting AMF-excreted glomalin. In conclusion, we observed the differential responses of mycorrhizal plants to two types of metal nanoparticles. Specifically, two nanoparticles exerted negative effects on mycorrhizal plant growth but in an opposite way: FeONPs were observed at high concentration (3.2 mg/kg) and AgNPs were at low concentration (0.01 mg/kg). Both responses are dramatically different from the current knowledge regarding the influence of nanoparticles on plant growth. It is possible that metal nanoparticles exert effects on AMF and therefore substantially alter plant growth. In this investigation, a sand medium was used instead of a soil medium, and only one type of soil microorganism was present. The actual situation in a
elevated levels of certain heavy metal species can enhance the effectiveness of AMF for stress alleviation.54 For example, Vogel-Mikus et al.41 found positive correlations between AMF colonization and total soil Zn, Cd and Pb concentrations, which indicates that the ability of mycorrhizal plants to alleviate heavy metal stress becomes more efficient. Therefore, in response to higher concentrations of AgNPs, increases were noted in the AMF infection rate (Figure 3b), the extractable GRSP content and root P acquisition (Figure 4b), compared to a low concentration of AgNPs. Due to the increased growth and enhanced ecological functions of AMF, the physiological toxicity of AgNPs for mycorrhizal plants was minimized. Specifically, the plant Ag content significantly decreased (p < 0.05) from 89.5 ± 7.6 ng/g to 10.4 ± 6.8 ng/g (Table 1), and the activities of three antioxidant enzymes decreased (Figure 5). Therefore, high levels of AgNPs did not exert negative effects on mycorrhizal clover biomass (Figure 2b). FeONPs Inhibit Mycorrhizal Plant Growth by Adversely Affecting AMF-Excreted Glomalin. Contrary to what was observed for AgNPs, the growth-promoting effect of AMF on mycorrhizal plants was weakened by FeONPs. Fe has long been recognized as a physiological requirement for life, which is not the case for many other heavy metals. AMF are a Fe modulator during the plant growth. In Fe-deficient environments, AMF change the form of Fe species to increase its bioavailability,55 and ultimately facilitate the Fe uptake by the plant.56 In Fe-excess environments, AMF are able to hold Fe in roots, and alleviate the unfavorable effect of Fe on the plant growth.54 In our experiments, Fe was already sufficient due to the addition of Hoagland’s nutrient solution. Therefore, additional Fe 2 O 3 bulk amendments did not influence mycorrhizal plant growth (Figure 2a) or Fe absorption (Table 1). When on FeONPs treatment, however, an adverse effect has been observed for the plants, especially at a high concentration (Figure 2a). Initially, we assumed that this could be a result of the characteristics of FeONPs - the small size allows them to permeate the cell wall and eventually reach the cytoplasm of microorganisms.57 Consequently, the growthpromoting effect of AMF on plants is greatly offset. However, ICP-MS revealed that FeONPs amendments did not increase Fe content in mycorrhizal plants (Table 1), which indicates that FeONPs are not directly phytotoxic to AMF and plants. Additionally, AMF infection rates did not decrease under FeONPs amendments (Figure 3a), implying that FeONPs do not inhibit AMF growth. Therefore, another underlying mechanism could be responsible for the negative influence of FeONPs on mycorrhizal plants. It is possible that FeONPs adversely affect AMF-excreted glomalin. Glomalin, a glycoprotein produced by AMF, is a component of the hyphal wall58 that accumulates in soils.59 Glomalin greatly contributes to soil aggregation, which is central to soil and ecosystem functioning because it controls fluxes of water, gases and nutrients. FeONPs have the most active surface sites (mainly the Fe−OH site) with high affinity for organic compounds.60 Due to this property, FeONPs might bind glomalin and restrain its ecological behavior. This speculation is supported by the data, which reveal a decrease 9502
dx.doi.org/10.1021/es402109n | Environ. Sci. Technol. 2013, 47, 9496−9504
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natural soil system is definitely more complicated. Therefore, our results imply that in a soil ecosystem, the influence of nanometals on plant systems is more complicated than expected, and more attention should be focused on the responses of soil microorganisms when evaluating the biological effect of nanomaterials on plants, the environment and ecology. Currently, there is still a large information gap that prevents our comprehensive understanding of the destiny of nanomaterials in ecosystems.
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ASSOCIATED CONTENT
S Supporting Information *
Additional material as noted in the text. This material is available free of charge via the Internet at http://pubs.acs.org.
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AUTHOR INFORMATION
Corresponding Author
*Phone:+086-025-86881589; fax:+086-025-86881000; e-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS We thank Professor Joselito M. Arocena for his pertinent suggestions and English improvement. This work was supported by the National Natural Science Foundation of China (Project No. 41071168, 41271256, 41001142, 61127002, and 61179035).
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