Halogenated organic compounds are naturally occurring in subsurface environments; however, accumulation of the degradative intermediate cis-1,2-dichloroethene (cDCE) at soil and groundwater sites contaminated with xenobiotic chlorinated ethenes is a global environmental and public health issue. Identifying microorganisms capable of cDCE degradation in these environments is of interest because of their potential application to bioremediation techniques. In this study, we sequenced, assembled, and analyzed the complete genome of Acinetobacter pittii CEP14, a strain isolated from chloroethene-contaminated groundwater, that has demonstrated the ability for aerobic cometabolic degradation of cDCE in the presence of n-hexane, phenol, and toluene. The A. pittii CEP14 genome consists of a 3.93 Mbp-long chromosome (GenBank accession no. CP084921) with a GC content of 38.9% and three plasmids (GenBank accession no. CP084922, CP084923, and CP084924). Gene function was assigned to 83.4% of the 3,930 coding DNA sequences. Functional annotation of the genome revealed that the CEP14 strain possessed all genetic elements to mediate the degradation of a range of aliphatic and aromatic compounds, including n-hexane and phenol. In addition, it harbors gene clusters involved in cytosol detoxification and oxidative stress resistance, which could play a role in the mitigation of toxic chemical intermediates that can arise during the degradation of cDCE. Gene clusters for heavy metal and antibiotic resistance were also identified in the genome of CEP14. These results suggest that CEP14 may be a versatile degrader of xenobiotic compounds and well-adapted to polluted environments, where a combination of heavy metal and organic compound pollution is often found.

• Keywords
• Acinetobacter pittii, Chlorinated ethenes (CEs), Cometabolism, Oxygenase, Phenol, Whole-genome shotgun sequencing, cis-1,2-dichloroethene (cDCE) biodegradation,
• MeSH
• Acinetobacter MeSH
• Biodegradation, Environmental MeSH
• Dichloroethylenes MeSH
• Phenols * MeSH
• Genomics MeSH
• Xenobiotics * MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• 1,2-dichloroethylene MeSH Browser
• Dichloroethylenes MeSH
• Phenols * MeSH
• Xenobiotics * MeSH

In this study, the diversity of bphA genes was assessed in a 13C-enriched metagenome upon stable isotope probing (SIP) of microbial populations in legacy PCB-contaminated soil with 13C-biphenyl (BP). In total, 13 bphA sequence variants (SVs) were identified in the final amplicon dataset. Of these, one SV comprised 59% of all sequences, and when it was translated into a protein sequence, it exhibited 87, 77.4, and 76.7% identity to its homologs from Pseudomonas furukawaii KF707, Cupriavidus sp. WS, and Pseudomonas alcaliphila B-367, respectively. This same BphA sequence also contained unusual amino acid residues, Alanine, Valine, and Serine in region III, which had been reported to be crucial for the substrate specificity of the corresponding biphenyl dioxygenase (BPDO), and was accordingly designated BphA_AVS. The DNA locus of 18 kbp containing the BphA_AVS-coding sequence retrieved from the metagenome was comprised of 16 ORFs and was most likely borne by Paraburkholderia sp. The BPDO corresponding to bphAE_AVS was cloned and heterologously expressed in E. coli, and its substrate specificity toward PCBs and a spectrum of flavonoids was assessed. Although depleting a rather narrow spectrum of PCB congeners, the efficient transformation of flavone and flavanone was demonstrated through dihydroxylation of the B-ring of the molecules. The homology-based functional assignment of the putative proteins encoded by the rest of ORFs in the AVS region suggests their potential involvement in the transformation of aromatic compounds, such as flavonoids. In conclusion, this study contributes to the body of information on the involvement of soil-borne BPDOs in the metabolism of flavonoid compounds, and our paper provides a more advanced context for understanding the interactions between plants, microbes and anthropogenic compounds in the soil.

• Keywords
• aromatic ring-hydroxylating dioxygenases, biphenyl dioxygenase, flavonoids, functional metagenomics, polychlorinated biphenyls, secondary plant metabolites,
• Publication type
• Journal Article MeSH

A bacterial species is best characterized after its isolation in a pure culture. This is an arduous endeavor for many soil microorganisms, but it can be simplified by several techniques for improving culturability: for example, by using growth-promoting factors. We investigated the potential of a Micrococcus luteus culture supernatant containing resuscitation-promoting factor (SRpf) to increase the number and diversity of cultured bacterial taxa from a nutrient-rich compost soil. Phosphate-buffered saline and inactivated SRpf were included as controls. After agitation with SRpf at 28°C for 1 day, the soil suspension was diluted and plated on two different solid, oligotrophic media: tenfold diluted Reasoner's 2A agar (R2A) and soil extract-based agar (SA). Colonies were collected from the plates to assess the differences in diversity between different treatments and cultivation media. The diversity on both R2A and SA was higher in the SRpf-amended extracts than the controls, but the differences on R2A were higher. Importantly, 51 potentially novel bacterial species were isolated on R2A and SA after SRpf treatment. Diversity in the soil extracts was also determined by high-throughput 16S rRNA amplicon sequencing, which showed an increase in the abundance of specific taxa before their successful cultivation. Conclusively, SRpf can effectively enhance the growth of soil bacterial species, including those hitherto uncultured.

• Keywords
• Micrococcus luteus, extracellular organic matter, increased culturability, non-cultured bacteria, oligotrophic medium, resuscitation-promoting factor, viable but non-culturable state,
• Publication type
• Journal Article MeSH

The involvement of bacterial aromatic ring-hydroxylating dioxygenases (ARHDs) in the degradation of aromatic pollutants, such as polychlorinated biphenyls (PCBs), has been well studied. However, there is considerable speculation as to the origin of this ability. One hypothesis is centered on a connection between the ability to degrade aromatic pollutants and the necessity of soil bacteria to cope with and/or utilize secondary plant metabolites (SPMs). To investigate this connection, we researched the involvement of biphenyl 2,3-dioxygenase (BPDO), an ARHD essential for the degradation of PCBs, in the metabolism of SPMs in the soil bacterium Pseudomonas alcaliphila JAB1, a versatile degrader of PCBs. We demonstrated the ability of the strain JAB1 to transform a variety of SPMs, namely the flavonoids apigenin, flavone, flavanone, naringenin, fisetin, quercetin, morin, and catechin, caffeic acid, trans-cinnamic acid, and the monoterpenes (S)-limonene and (R)-carvone. Of those, the transformation of flavone, flavanone, and (S)-limonene was conditioned by the activity of JAB1-borne BPDO and thus was researched in more detail, and we found evidence for the limonene monooxygenase activity of the BPDO. Furthermore, the bphA gene in the strain JAB1 was demonstrated to be induced by a wide range of SPMs, with monoterpenes being the strongest inducers of the SPMs tested. Thus, our findings contribute to the growing body of evidence that ARHDs not only play a role in the catabolism of aromatic pollutants, but also of natural plant-derived aromatics, and this study supports the hypothesis that ARHDs participate in ecological processes mediated by SPMs.

• Keywords
• aromatic ring-hydroxylating dioxygenases, biphenyl dioxygenase, monoterpenes, phenolics, secondary plant metabolites,
• Publication type
• Journal Article MeSH

Certain industrial chemicals accumulate in the environment due to their recalcitrant properties. Bioremediation uses the capability of some environmental bacteria to break down these chemicals and attenuate the pollution. One such bacterial strain, designated Pvy, was isolated from sediment samples from a lagoon in Romania located near an oil refinery due to its capacity to degrade dibenzofuran (DF). The genome sequence of the Pvy strain was obtained using an Oxford Nanopore MiniION platform. According to the consensus 16S rRNA gene sequence that was compiled from six 16S rRNA gene copies contained in the genome and orthologous average nucleotide identity (OrthoANI) calculation, the Pvy strain was identified as Pseudomonas veronii, which confirmed the identification obtained with the aid of MALDI-TOF mass spectrometry and MALDI BioTyper. The genome was analyzed with respect to enzymes responsible for the overall biodegradative versatility of the strain. The Pvy strain was able to derive carbon from naphthalene (NP) and several aromatic compounds of natural origin, including salicylic, protocatechuic, p-hydroxybenzoic, trans-cinnamic, vanillic, and indoleacetic acids or vanillin, and was shown to degrade but not utilize DF. In total seven loci were found in the Pvy genome, which enables the strain to participate in the degradation of these aromatic compounds. Our experimental data also indicate that the transcription of the NP-dioxygenase α-subunit gene (ndoB), carried by the plasmid of the Pvy strain, is inducible by DF. These features make the Pvy strain a potential candidate for various bioremediation applications.

• Keywords
• Pseudomonas veronii strain Pvy, biodegradation, denitrification, dibenzofuran, dioxygenase, heavy-metal tolerance, nanopore technology, organic phosphate mineralization, whole-genome sequencing,
• MeSH
• Biodegradation, Environmental MeSH
• Dibenzofurans * MeSH
• Genomics * MeSH
• Pseudomonas MeSH
• RNA, Ribosomal, 16S MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Dibenzofurans * MeSH
• RNA, Ribosomal, 16S MeSH

The complete genome sequence of Rhodococcus sp. WAY2 (WAY2) consists of a circular chromosome, three linear replicons and a small circular plasmid. The linear replicons contain typical actinobacterial invertron-type telomeres with the central CGTXCGC motif. Comparative phylogenetic analysis of the 16S rRNA gene along with phylogenomic analysis based on the genome-to-genome blast distance phylogeny (GBDP) algorithm and digital DNA-DNA hybridization (dDDH) with other Rhodococcus type strains resulted in a clear differentiation of WAY2, which is likely a new species. The genome of WAY2 contains five distinct clusters of bph, etb and nah genes, putatively involved in the degradation of several aromatic compounds. These clusters are distributed throughout the linear plasmids. The high sequence homology of the ring-hydroxylating subunits of these systems with other known enzymes has allowed us to model the range of aromatic substrates they could degrade. Further functional characterization revealed that WAY2 was able to grow with biphenyl, naphthalene and xylene as sole carbon and energy sources, and could oxidize multiple aromatic compounds, including ethylbenzene, phenanthrene, dibenzofuran and toluene. In addition, WAY2 was able to co-metabolize 23 polychlorinated biphenyl congeners, consistent with the five different ring-hydroxylating systems encoded by its genome. WAY2 could also use n-alkanes of various chain-lengths as a sole carbon source, probably due to the presence of alkB and ladA gene copies, which are only found in its chromosome. These results show that WAY2 has a potential to be used for the biodegradation of multiple organic compounds.

• Keywords
• PAH, PCB, Rhodococcus, biodegradation, complete genome, hydrocarbons,
• MeSH
• AlkB Enzymes genetics   metabolism   MeSH
• Biodegradation, Environmental MeSH
• Phylogeny MeSH
• Naphthalenes metabolism   MeSH
• Polychlorinated Biphenyls chemistry   MeSH
• Rhodococcus classification   genetics   growth & development   MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Whole Genome Sequencing methods   MeSH
• Cluster Analysis MeSH
• High-Throughput Nucleotide Sequencing MeSH
• Xylenes metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• AlkB Enzymes MeSH
• Naphthalenes MeSH
• naphthalene MeSH Browser
• Polychlorinated Biphenyls MeSH
• RNA, Ribosomal, 16S MeSH
• Xylenes MeSH

Extended soil contamination by polychlorinated biphenyls (PCBs) represents a global environmental issue that can hardly be addressed with the conventional remediation treatments. Rhizoremediation is a sustainable alternative, exploiting plants to stimulate in situ the degradative bacterial communities naturally occurring in historically polluted areas. This approach can be enhanced by the use of bacterial strains that combine PCB degradation potential with the ability to promote plant and root development. With this aim, we established a collection of aerobic bacteria isolated from the soil of the highly PCB-polluted site "SIN Brescia-Caffaro" (Italy) biostimulated by the plant Phalaris arundinacea. The strains, selected on biphenyl and plant secondary metabolites provided as unique carbon source, were largely dominated by Actinobacteria and a significant number showed traits of interest for remediation, harbouring genes homologous to bphA, involved in the PCB oxidation pathway, and displaying 2,3-catechol dioxygenase activity and emulsification properties. Several strains also showed the potential to alleviate plant stress through 1-aminocyclopropane-1-carboxylate deaminase activity. In particular, we identified three Rhodococcus strains able to degrade in vitro several PCB congeners and to promote lateral root emergence in the model plant Arabidopsis thaliana in vivo. In addition, these strains showed the capacity to colonize the root system and to increase the plant biomass in PCB contaminated soil, making them ideal candidates to sustain microbial-assisted PCB rhizoremediation through a bioaugmentation approach.

• MeSH
• Arabidopsis growth & development   microbiology   MeSH
• Bacterial Proteins genetics   metabolism   MeSH
• Biodegradation, Environmental MeSH
• Gene Expression MeSH
• Catechol 2,3-Dioxygenase genetics   metabolism   MeSH
• Plant Roots growth & development   microbiology   MeSH
• Soil Pollutants metabolism   MeSH
• Carbon-Carbon Lyases genetics   metabolism   MeSH
• Oxidation-Reduction MeSH
• Phalaris growth & development   microbiology   MeSH
• Polychlorinated Biphenyls metabolism   MeSH
• Soil chemistry   MeSH
• Soil Microbiology MeSH
• Rhodococcus enzymology   genetics   MeSH
• Secondary Metabolism genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 1-aminocyclopropane-1-carboxylate deaminase MeSH Browser
• Bacterial Proteins MeSH
• Catechol 2,3-Dioxygenase MeSH
• Soil Pollutants MeSH
• Carbon-Carbon Lyases MeSH
• Polychlorinated Biphenyls MeSH
• Soil MeSH