Bioaugmentation with zeolite-immobilized bacterial consortium OPK results in a bacterial community shift and enhances the bioremediation of crude oil-polluted marine sandy soil microcosms.

A pyrene-degrading consortium OPK containing Mycolicibacterium strains PO1 and PO2, Novosphingobium pentaromativorans PY1 and Bacillus subtilis FW1 effectively biodegraded medium- and long-chain alkanes as well as mixed hydrocarbons in crude oil. The detection of alkB and CYP153 genes in the genome of OPK members supports its phenotypic ability to effectively degrade a broad range of saturated hydrocarbons in crude oil. Zeolite-immobilized OPK was developed as a ready-to-use bioproduct and it exhibited 74% removal of 1000 mg L-1 crude oil within 96 h in sterilized seawater without nutrient supplementation and maintained high crude oil-removal activity under a broad range of pH values (5.0-9.0), temperatures (30-40 °C) and salinities (20-60‰). In addition, the immobilized OPK retained a high crude oil removal efficacy in semicontinuous experiments and showed reusability for at least 5 cycles. Remarkably, bioaugmentation with zeolite-immobilized OPK in sandy soil microcosms significantly increased crude oil (10,000 mg kg-1 soil) removal from 45% to 80.67% within 21 days compared to biostimulation and natural attenuation. Moreover, bioaugmentation with exogenous immobilized OPK stimulated an increase in the relative abundances of Alcanivorax genus, indigenous hydrocarbon-degrading bacteria, which in turn enhanced removal efficiency of crude oil contamination from sandy soil microcosms. The results indicate positive interactions between the bioaugmented immobilized consortium, harboring Mycolicibacterium as a key player, and indigenous Alcanivorax, which exhibited crucial functions for improving crude oil removal efficacy. The knowledge obtained forms an important basis for further synthesis and handling of a promising bio-based product for enhancing the in situ bioremediation of crude oil-polluted marine environments.

Effects of environmental factors and coexisting substrates on PAH degradation and transcriptomic responses of the defined bacterial consortium OPK.

The present study showed that syntrophic associations in a defined bacterial consortium, named OPK, containing Mycolicibacterium strains PO1 and PO2, Novosphingobium pentaromativorans PY1 and Bacillus subtilis FW1, led to effective pyrene degradation over a wide range of pH values, temperatures and salinities, as well as in the presence of a second polycyclic aromatic hydrocarbon (PAH). Anthracene, phenanthrene or fluorene facilitated complete pyrene degradation within 9 days, while fluoranthene delayed pyrene degradation. Interestingly, fluoranthene degradation was enhanced in the presence of pyrene. Transcriptome analysis confirmed that Mycolicibacterium strains were the key PAH-degraders during the cometabolism of pyrene and fluoranthene. Notably, the transcription of genes encoding pyrene-degrading enzymes were shown to be important for enhanced fluoranthene degradation. NidAB was the major initial oxygenase involved in the degradation of pyrene and fluoranthene mixture. Other functional genes encoding ribosomal proteins, an iron transporter, ABC transporters and stress response proteins were induced in strains PO1 and PO2. Furthermore, an intermediate pyrene-degrading Novosphingobium strain contributed to protocatechuate degradation. The results demonstrated that synergistic interactions among the bacterial members (PO1, PO2 and PY1) of the consortium OPK promoted the simultaneous degradation of two high molecular weight (HMW) PAHs.

Synergistic degradation of pyrene by five culturable bacteria in a mangrove sediment-derived bacterial consortium.

A pyrene-degrading microbial consortium was obtained after enrichment with mangrove sediment collected from Thailand. Five cultivable bacteria (Mycobacterium spp. PO1 and PO2, Novosphingobium pentaromativorans PY1, Ochrobactrum sp. PW1, and Bacillus sp. FW1) were successfully isolated from the consortium. Draft genomes of them showed that two different morphotypes of Mycobacterium (PO1 and PO2), possessed a complete gene set for pyrene degradation. PY1 contained genes for phthalate assimilation via protocatechuate, a central intermediate, by meta-cleavage pathway, and PW1 possessed genes for protocatechuate degradation via ortho-cleavage pathway. The occurrence of biosurfactant-producing genes in FW1 suggests the involvement in enhancing the pyrene bioavailability. Biotransformation experiments revealed that Mycobacterium completely degraded 100mgL-1 pyrene within six days, whereas no significant degradation was observed with the others. Notably, PY1 and PW1 exhibited higher activity for protocatechuate degradation than the others. The artificially reconstructed consortia containing Mycobacterium with the other three strains (PY1, PW1 and FW1) showed three-fold higher degradation rate for pyrene than the individual Mycobacterium. The enhanced pyrene biodegradation achieved in the consortium was due to the cooperative interaction of bacterial mixture. Our findings showing that synergistic degradation of pyrene in the consortium will facilitate the application of the defined bacterial consortium in bioremediation.