## 2 Materials and Methods  
### Experimental Design  
The soil used in this study was collected (to a depth of 1 m, as previous information showed that the plume of contamination reached 1 m deep) from a chronically contaminated site near Bilbao, Spain. The soil was mechanically sieved (<20 mm mesh), air dried until constant weight, and finally sieved again to <6 mm. The analysis of soil contaminants was carried out by a certified laboratory (SYNLAB Analytics & Services B. V, Rotterdam, The Netherlands). PAHs were extracted in hexane/acetone by agitation and determined by Gas Chromatography-Mass Spectrometry (GC-MS) [27]. C10-C40 hydrocarbon fractions were extracted in hexane/acetone and determined by Gas Chromatography-Flame Ionization Detector [28]. C5-C10 hydrocarbon fractions were extracted in methanol and determined by GC-MS [29]. In order to facilitate the bioavailability and degradability of recalcitrant contaminant fractions, before placing the soil in the experimental pots (see below), rhamnids (10 mg kg\({}^{-1}\) DW soil, Sigma-Aldrich R90-10G) and sodium dodecyl benzenesulfonate (50 mg kg\({}^{-1}\) DW soil, Sigma-Aldrich 289957) were added to the soil and then hand-mixed thoroughly to ensure a homogeneous mixture. In this study, 12 treatments were tested (in triplicate; a total of 36 pots; Table 1). Two organic amendments were individually applied to the contaminated soil as biostimulating agents (bioremediation via _biostimulation_): composted horse manure (C: 34%, N: 2.0%; C/N ratio = 17) and dried cow slurry (C: 42%, N: 2.8%; C/N ratio = 15). Amendment doses were adjusted to reach a final organic matter (OM) concentration of 20%. A control soil with no organic amendment was also established. Experimental pots containing 2 kg of the biostimulated organically-contaminated industrial soil were placed in a growth chamber under the following controlled conditions: 25/22 \({}^{\circ}\)C day/night, 65% relative humidity, and 300 \(\upmu\)mol photon m\({}^{-2}\) s\({}^{-1}\), with a 14-h photoperiod. Half of the pots were planted with _Brassica napus_ for _rhizoremediation_ purposes. In a previous rhizoremediation experiment [30], we found _B. napus_ plants to have a key role in the recovery of the health of soils contaminated with metals and diesel. Approximately, 20 seeds per pot were superficially buried in the soil and, after seed germination, only 6 homogeneous plants per pot were left (the rest were manually removed). Fifty days after initiation of the treatments (to allow enough time for plant development), _bioaugmentation_ with a bacterial consortium was performed in half of the pots. The bacterial strains used for this consortium were: (i) _Burkholderia xenovorans_ LB400 DSM-17367, _Paenibacillus_ sp. and _Lysinibacillus_ sp., three strains previously characterized by their ability to degrade hydrocarbons; and (ii) four endophyte strains (one _Microbacterium_ sp. and three _Pseudomonas_ sp.) which, in previous studies [17], showed their plant growth-promoting traits. The strains were grown overnight in \begin{table}
\begin{tabular}{c c c} \hline \hline
**Rhizoremediation** & **Bioaugmentation** & **Biostimulation** \\ \hline _Brassica napus_ & Bioaugmented & Control \\  & & Dried cow slurry \\  & & Composed horse manure \\  & Non-bioaugmented & Control \\  & & Dried cow slurry \\  & & Composed horse manure \\ \hline Unplanted & Bioaugmented & Control \\  & & Dried cow slurry \\  & & Composed horse manure \\ \hline Non-bioaugmented & Control \\  & & Dried cow slurry \\  & & Composed horse manure \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Experimental treatments. Luria-Bertani broth (Sigma-Aldrich L3022) at 28 \({}^{\circ}\)C on a rotary shaker (125 rpm, Sartorius, Certomat(r) BS-1).