## 2 Materials and Methods
### 4-n-NP Biodegradation in Solution  
By inoculating the NP-degrading bacterial strains, 4-n-NP biodegradation studies were carried out in a solution. All the microcosm components were sterilized (autoclavedfor one cycle at 120 \({}^{\circ}\)C and a pressure of 101 kPa for 20 min, steam sterilizer model S100, Matachana, Casteldefells, Spain). 4-n-NP biodegradation in the solution was carried out in triplicate. Briefly, 4-n-NP was added to Corex glass centrifuge tubes containing 3 mL of the MSM + micronutrient solution to obtain a final concentration of 4-n-NP 10 mg L\({}^{-1}\) (about 7 mg L\({}^{-1}\) should be dissolved and the rest in solid state, according to the aqueous solubility of 4-n-NP). In the previous study where these selected strains were isolated [36], it was observed that the degradation of 4-n-NP by _B. safensis_ CN12 increased in the presence of glucose, and the same was observed in the present study for the rest of the isolated strains. The results of these studies are included in Figure S1 (Supplementary Material). For this reason, MSM was supplemented with glucose (1 g L\({}^{-1}\)) as an external carbon source. The containers were inoculated with 10\({}^{8}\) CFUs mL\({}^{-1}\) of each bacterial strain, and non-inoculated sterile controls were also prepared. Microcosms were incubated on a rotary shaker (150 rpm, 30 \(\pm\) 1 \({}^{\circ}\)C) and were periodically opened to ensure sufficient oxygen supply. To determine the remaining 4-n-NP in the system, at periodic intervals, some containers were removed from the incubator, frozen, and thawed three times (to also determine 4-n-NP accumulated in the microbial biomass [39]), and a liquid-liquid extraction assay was carried out by adding dichloromethane (1:1, \(v/v\)). The concentration of 4-n-NP in the dichloromethane extract was determined by HPLC with a fluorescence detector (RF-10 AXL Shimadzu, Kyoto, Japan), a C-18 column (4 mm diameter, 150 mm length), and the acetonitrile mobile phase: water (90:10), as well as 222 and 305 nm as the excitation and emission wavelengths. 4-n-NP transformation products produced (Figure S2, Supplementary Material) were also analyzed by ultra-high-performance liquid chromatography (UHPLC, Agilent 1290 II Infinity LC system equipment (Agilent, CA, USA)) using an Infinity Lab Poroshell 120 EC-C18 (100 \(\times\) 4.6 mm i.d.; 4 \(\upmu\)m) analytical column (Agilent, CA, USA) thermostated at 35 \({}^{\circ}\)C. The mobile phase consisted of a mixture of a 10 mM ammonium acetate aqueous solution (solvent A) and methanol (solvent B) at a flow rate of 0.6 mL min\({}^{-1}\). The elution program was for 0-15 min, with a linear increment from 10% to 100% of solvent B, followed by 15-18 min in isocratic 100% solvent B. Mass spectrometry detection was performed on an Agilent 6495c triple-quadrupole-mass spectrometer equipped with an electrospray ionization source (more details in [PERSON] et al.) [36]. Precursor ions of \(m/z\) 219 corresponding to 4-n-NP, and values of 233, 235, and 237 corresponding to the ions [M\({}^{-1}\)] of nonylphenol, 4-(6-oxo-1,1-dimethylheptyl)phenol (TP1), 4-nonyl-4-hydroxycyclohexa-2,5-dienone (TP2), and 4-Nonyl-4-hydroxycyclohex-2-enone (TP3), respectively, were selected. The MS/MS spectra from m/z 50 to 500 were obtained using a scan time of 300 ms. The presence of the compounds was confirmed using Multiple Reaction Monitoring (MRM) mode. At least two transitions were selected for all compounds, except for 4-Nonyl-4-hydroxycyclohex-2-enone (only one transition was monitored). The experimental conditions used in MRM mode are shown in Table S2.