## 5 Conclusions  
The highest tested concentrations of polystyrene NPIs dispersion (1200 and 1500 mg NPIs/kg), 21 d exposure via soil, had an impact on the _E. cryptic_s survival and reproduction (decreased). On the other hand, the exposure via water showed that polystyrene NPIs dispersion induced 100% of mortality at concentrations \(\geq\) 150 mg NPIs/L. However, these effects seem mainly associated with tween 20 and NaN\({}_{3}\) present in the polystyrene NPIs dispersion and not with NPIs themselves. Nevertheless, there was a tendency of the organisms to avoid the polystyrene NPIs dispersion spiked soils, being this response relevant at the lowest tested concentration (0.015 mg NPIs/kg). Indeed, organisms decreased the avoidance responses with the polystyrene NPIs concentration increase. A NPIs dispersion toxicity dependent on the medium of exposure (soil versus water) and NPIs concentration was found in our study. The obtained results show that there is an urgent requirement to focus on research about this topic, and to provide more knowledge about environmental behaviour, as well as ecotoxicological data about NPIs (exposed alone and with other compounds) in terrestrial ecosystems.  
The following are available online at [[http://www.mdpi.com/2076-3417/10/21/7674/s1](http://www.mdpi.com/2076-3417/10/21/7674/s1)]([http://www.mdpi.com/2076-3417/10/21/7674/s1](http://www.mdpi.com/2076-3417/10/21/7674/s1)), Figure S1: Representative images of _Enchytraus crypticus_ adults when exposed during 5 days in ISO water to polystyrene nanobulstics (NPIs). Black arrows indicate NPIs agglomeration/aggregation or adsorption in the ditellum/cocon area, Table S1: Number of _Enchytraus crypticus_ juveniles counted after 21 d exposure to polystyrene nanoplastics (NPIs) in LUFA 2.2 soil. Results are expressed as average value (AV) \(\pm\) standard error (SE). * Significant differences to control (p \(<\) 0.05), Table S2: Number of _Enchytraus crypticus_ juveniles counted after 21 d exposure to the dispersant--tween 20 and sodium azide (NaN3)--in LUFA 2.2 soil. Results are expressed as average value (AV) \(\pm\) standard error (SE). * Significant differences to control (p \(<\) 0.05), Table S3: Effect Concentrations (ECx), applying the 2-parameters Logistic model, for survival and reproduction of _Enclytraus crypticus_ after 21 d exposure to polystyrene nanobulstics (NPIs) in LUFA 2.2 soil. EC 20, 50. 80: Concentration that causes 20, 50 and 80% of effect, respectively. Results are presented as estimated value \(\pm\) standard error.  
Conceptualization: A.B., J.S. and V.L.M.; Methodology: A.B., J.S. and V.L.M.; Formal Analysis: A.B., J.S. and V.L.M.; Investigation: A.B., J.S. and V.L.M.; Resources: M.J.B.A. and V.L.M.; Writing--Original Draft Preparation: A.B.; Writing--Review and Editing: A.B., J.S., M.J.B.A. and V.L.M.; Project Administration: V.L.M.; Funding Acquisition: V.L.M. All authors have read and agreed to the published version of the manuscript.  
This research was funded by CESAM (UIDB/50017/2020+UIDP/50017/2020), FCT/MEC through national funds, and the co-funding by the FEDER (POCI-01-0145-FEDER-00763), within the PT2020 Partnership Agreement and Compete 2020. Work done under the project UNRAFEL (POCI-01-0145-FEDER-029035) financed by FEDER, through COMPETE2020 - POCI, and by national funds (QE), through FCT/MCTES national funds (PIDDAC), the co-funding by the FEDER, within the PT2020 Partnership Agreement and Compete 2020. [PERSON] and [PERSON] have a contract researcher and a fellowship from the project (POCI-01-0145-FEDER-029035), respectively. [PERSON] is funded by national funds (OE), through FCT, in the scope of the framework contract foreseen in the numbers 4, 5 and 6 of the article 23, of the Decree-Law 57/2016, of August 29, changed by Law 57/2017, of July 19.  
The authors would like to thank to Professor [PERSON] (Chemistry Department, CICECO, UA) for the opportunity given to the physicochemical characterization of nanoplastics.  
The authors declare no conflict of interest.