## 4 Discussion  
### Liquid Medium Growth Assay  
The growth and activity of _P. ostreatus_ and _P. erygii_ in the presence of TCE in liquid broth were evaluated as preliminary step in the assessment of the removal of TCE from soil. The results showed that both species of _Pleurotus_ tolerated and grew in the presence of the highest concentration of TCE (140 mg L\({}^{-1}\)) and developed functional pellets visually similar in shape and size to those in the control (Figure 1). Other researchers have reported that some species of ligninolytic fungi were not affected by the presence of TCE, such as the \begin{table}
\begin{tabular}{c c c c c} \hline  & **Week 1** & **Week 2** & **Week 3** & **Week 4** \\ \hline _P. ostreatus_ & 12.8 \(\pm\) 6.8 & 2.6 \(\pm\) 1.5 & 4.7 \(\pm\) 2.9 & 3.1 \(\pm\) 1.9 \\ _P. erygii_ & 10.5 \(\pm\) 4.8 & n.d. & 4.2 \(\pm\) 1.5 & 2.4 \(\pm\) 0.5 \\ \hline n.d.: not detected. & & & & \\ \end{tabular}
\end{table} 
Table 2: MnP (U kg\({}^{-1}\)) activity in TCE-polluted soil with mycoremediation using _P. ostreatus_ and _P. erygii_. Data present the mean \(\pm\) standard deviation of three replicates. Figure 4: Percentage of TCE removal in polluted soil with TCE at 140 mg kg\({}^{-1}\) for the control and mycoremediation treatments (_P. ostreatus_ and _P. erygii_). Error bars represent the standard deviation (\(n=3\)). Different letters indicate significant differences among treatments at the same sampling time ([PERSON] post hoc test, \(p\) < 0.05). n.s. : not significant. case of _T. versicolor_ (2-20 mg L\({}^{-1}\)) [26], _P. chryssopporum_ (10-75 mg L\({}^{-1}\)) [27], _G. lucidum_ and _I. lacticus_ (5 and 10 mg L\({}^{-1}\)) [28]. However, the concentration evaluated in this work was much higher, reaching 140 mg L\({}^{-1}\). Therefore, the results of this first test in liquid culture were promising because the two fungi were able to grow at a very high concentration of TCE. However, the observations carried out at a microscopical level using SEM showed that the hyphae of _P. ostreatus_ appeared structurally affected by the presence of TCE at 140 mg L\({}^{-1}\), exhibiting hyphae with morphologically altered tips and a loss of their tubular structure. However, despite this they did not appear to have lost the ability to function properly, as was evaluated by the production of extracellular enzymes (Figure 3A). [PERSON] et al. [27] reported a slight inhibition of the growth of _P. chryssopporum_ at TCE 100 mg L\({}^{-1}\). It would therefore appear that a high concentration of TCE produces toxic effects toward some white rot fungi. By contrast, the presence of this contaminant did not produce these effects on the structure of _P. erygii_ at any of the concentrations used (Figure 2). _P. erygii_ secreted higher levels of extracellular laccase and MnP than _P. ostreatus_. Laccase was not detected in the control by _P. ostreatus_. The highest levels of both laccase and MnP were produced in the presence of TCE 140 mg L\({}^{-1}\) at day 14. The production of laccase by the two _Pleurotus_ species seemed to be induced by the presence of TCE. This finding is in accordance with those of [PERSON] et al. [26], who stated that TCE could induce the production of laccase in _T. versicolor_ because the laccase activity appeared to increase with increased initial TCE concentrations. Furthermore, the fungal growth did not seem to be greatly affected by the concentration of the pollutant, as the mycelial dry weight was only 15% lower in the highest TCE concentration treatment (20 mg L\({}^{-1}\)). However, the fact that laccase was produced in the presence of TCE did not mean the two _Pleurotus_ species would be able to degrade it. For example, _P. chryssopporum_ produced ligninolytic enzymes in the presence of TCE, but these were not involved in the degradation of this contaminant. The highest rates of degradation were obtained when _P. chryssopporum_ was cultured in a high N medium, which is well known for suppressing ligninolytic enzyme production [27]. Similarly, [PERSON] et al.