Increasing attention has been paid to the toxicity and hazards of antibiotics on non-target organisms in soil ecosystems as excess antibiotics in the excretion of treated animals are being brought into the soil through manure and sewage irrigation.

The activated sludge process can effectively remove polycyclic aromatic hydrocarbons (PAHs) from wastewater via biodegradation. However, the degradable microorganisms and functional enzymes involved in this process remain unclear. In this study, we successfully employed a laboratory-scale sequential batch reactor to investigate variations in microbial community and protein expression in response to the addition of different PAHs and process time. The analysis of bacterial community structure by 454 pyrosequencing of the 16S rRNA gene indicated that bacteria from Burkholderiales order were dominant in PAHs treated sludge. Mass spectrometry performed with 2D protein profiles of all sludge samples demonstrated that most proteins exhibiting differential expression profiles during the process were derived from Burkholderiales populations; these proteins are involved in DNA replication, fatty acid and glucose metabolism, stress response, protein synthesis, and aromatic hydrocarbon metabolism. Nevertheless, the protein expression profiles indicated that naphthalene, but not anthracene, can induce the expression of PAH-degrading proteins and accelerate its elimination from sludge. Though only naphthalene and anthracene were added into our experimental groups, the differentially expressed enzymes involved in other PAHs (especially biphenyl) metabolism were also detected. This study provides apparent evidence linking the metabolic activities of Burkholderiales populations with the degradation of PAHs in activated sludge processes. Overall, our findings highlighted the successful application of metaproteomics integrated with microbial diversity analysis by high-throughput sequencing technique on the analysis of environmental samples, which could provide a convenience to monitor the changes in proteins expression profiles and their correlation with microbial diversity. View Article Onlinea NAP eff : average effluent naphthalene concentration; ANT eff : average effluent anthracene concentration; NAP sludge : naphthalene in sludge; ANT sludge : anthracene in sludge; E NAP : naphthalene removal efficiency; E ANT : anthracene removal efficiency; E-BIO NAP : biodegradation efficiency of naphthalene; E-BIO ANT : biodegradation efficiency of anthracene. b n ¼ 3, mean AE SD.22844 | RSC Adv., 2019,9,[22841][22842][22843][22844][22845][22846][22847][22848][22849][22850][22851][22852] This journal is View Article Online 356 in naphthalene 1,2-dioxygenase (N1,2-DO, EC 1.14.12.12) of Pseudomonas putida G7. Data showed that there was an efficient electron transfer between the BDO reductase and the N1,2-DO ferredoxin, the BDO ferredoxin could resubstitute N1,2-DO ferredoxin in part (22%), which indicated that BDO might participate in naphthalene degradation. 41 Furthermore, Yu et al. Fig. 3 2D PAGE profiles of proteins extracted from the experimental groups. Potential protein molecular mass ranges are indicated on the left and isoelectric point ranges are provided on the top of the gel images.22848 | RSC Adv., 2019,9,[22841][22842][22843][...

Many bacteria metabolize aromatic compounds via catechol as a catabolic intermediate, and possess multiple genes or clusters encoding catechol-cleavage enzymes. The presence of multiple isozyme-encoding genes is a widespread phenomenon that seems to give the carrying strains a selective advantage in the natural environment over those with only a single copy. In the naphthalene-degrading strain Pseudomonas putida ND6, catechol can be converted into intermediates of the tricarboxylic acid cycle via either the orthoor meta-cleavage pathways. In this study, we demonstrated that the catechol ortho-cleavage pathway genes (catB I C I A I and catB II C II A II) on the chromosome play an important role. The cat I and cat II operons are co-transcribed, whereas catA I and catA II are under independent transcriptional regulation. We examined the binding of regulatory proteins to promoters. In the presence of cis-cis-muconate, a well-studied inducer of the cat gene cluster, CatR I and CatR II occupy an additional downstream site, designated as the activation binding site. Notably, CatR I binds to both the cat I and cat II promoters with high affinity, while CatR II binds weakly. This is likely caused by a T to G mutation in the G/T-N 11-A motif. Specifically, we found that CatR I and CatR II regulate catB I C I A I and catB II C II A II in a cooperative manner, which provides new insights into naphthalene degradation.