Metabolic interactions in a bacterial co-culture accelerate phenanthrene degradation

doi:10.1016/j.jhazmat.2020.123825

Journal of Hazardous Materials

Volume 403, 5 February 2021, 123825

https://doi.org/10.1016/j.jhazmat.2020.123825 Get rights and content

Highlights

•
Rhodococcus sp. WB9 and Mycobacterium sp. WY10 showed synergistic PHE degradation.
•
Metabolic cross-feeding in co-culture promoted PHE degradation and mineralization.
•
1H2N and PHTA were accumulated extracellularly in monoculture by efflux transporters.
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The absence of transporters for metabolites uptake restricts metabolites degradation.

Abstract

A highly eﬀ ;ective phenanthrene (PHE)-degrading co-culture containing Rhodococcus sp. WB9 and Mycobacterium sp. WY10 was constructed and completely degraded 100 mg L⁻¹ PHE within 36 h, showing improved degradation rate compared to their monocultures. In the co-culture, strain WY10 played a predominant role in PHE degradation. 1-hydroxy-2-naphthoic acid was an end-product of PHE degradation by strain WB9 and accumulated in the culture medium to serve as a substrate for strain WY10 growth, thereby accelerating PHE degradation. In turn, strain WY10 degraded PHE and 1-hydroxy-2-naphthoic acid intracellularly to form phthalate and protocatechuate that were exported to the culture medium through efflux transporters. However, strain WY10 cannot take up extracellular phthalate due to the absence of phthalate transporters, restricting phthalate degradation and PHE mineralization. In the co-culture, phthalate and protocatechuate accumulated in the culture medium were taken up and degraded towards TCA cycle by strain WB9. Therefore, the metabolic cross-feeding of strains WB9 and WY10 accelerated PHE degradation and mineralization. These findings exhibiting the synergistic degradation of PHE in the bacterial co-culture will facilitate its bioremediation application.

Graphical abstract

Introduction

Polycyclic aromatic hydrocarbons (PAHs) are a class of chemical pollutants in various environmental niches, which are of concern due to their wide distribution, recalcitrance and toxicity (Shen et al., 2013; Hong et al., 2016; Kraiselburd et al., 2019; Subashchandrabose et al., 2019). Bacteria are a group of microorganisms actively involved removal of pollutants through their metabolism, and the potential of bacterial consortia to degrade PAHs is thus of interest (Lu et al., 2019; Rabodonirina et al., 2019). However, most of previous studies on PAHs biodegradation have focused on the single pure bacterial culture and the PAHs-degradation ability of a pure culture was found to be limited (Lafortune et al., 2009; Luo et al., 2014; Hennessee and Li, 2016; Wang et al., 2018b).

Many studies have explored that PAHs-biodegradation efficiency was enhanced by bacterial consortia due to their efficient synergism (Luan et al., 2006; Wang et al., 2006; Zhong et al., 2011; Wang et al., 2018a; Wanapaisan et al., 2018). The synergistic mechanisms in bacterial consortia during PAHs degradation generally involve cooperation of metabolic activities (Luan et al., 2006; Wang et al., 2006; Zhong et al., 2011; Wanapaisan et al., 2018). For instance, the cooperation within a bacterial consortium would possibly result in diverse PAHs metabolic pathways, thus accelerating PAHs degradation (Wang et al., 2006). In other PAHs-degrading consortia, some bacteria were able to catabolize PAHs metabolites accumulated by other members, which relieve the metabolite repression on the degradation of parent PAHs (Luan et al., 2006; Zhong et al., 2011).

Nevertheless, due to the diversity of bacterial consortia, more studies are needed to clarify the diverse mechanisms that govern PAHs metabolism by the consortia. Moreover, the composition of communities affects the rate of PAHs degradation as well as the identity of the final products. Thus, further study is needful to obtain the composition of bacterial consortia throughout PAHs degradation process and evaluate on members’ roles in PAHs degradation.

Rhodococcus sp. WB9 and Mycobacterium sp. WY10 isolated from PAHs-contaminated soils were both eﬀ ;ective phenanthrene (PHE)-degrading bacteria (Sun et al., 2019, 2020). The PHE metabolic pathways of both strains have been studied. The results showed that a large amount of metabolite 1-hydroxy-2-naphthoic acid (1H2N) was accumulated during PHE degradation by strain WB9, which restricted PHE mineralization by strain WB9 (Sun et al., 2020), however, strain WY10 had high ability to degrade 1H2N as the sole carbon source (Sun et al., 2019). The genomic analysis of strains WB9 and WY10 have revealed a lot of catabolic genes encoding enzymatic steps required for PHE metabolites catabolism (Sun et al., 2019, 2020). It is likely that the transportation of metabolites is playing an important role in PHE degradation and mineralization. However, the functional genes involved in the transport of PHE metabolites have not been studied.

Thus, in the present study, a bacterial co-culture containing Rhodococcus sp. WB9 and Mycobacterium sp. WY10 was constructed to enhance PHE degradation and mineralization. More significantly, the aim of this study is to clarify the synergistic mechanism of strains WB9 and WY10 as well as their respective roles in PHE degradation through chemical analysis and genomic analysis with respect to trans-membrane transport.

Section snippets

Chemicals and media

PHE (98 %) was procured from Sigma-Aldrich (Shanghai, China). Diphenic acid (DIPA, 98 %), 1H2N (98 %), phthalate (PHTA, 99.5 %) protocatechuate (POCA, 98 %) and salicylate (SALA, 99.5 %) were purchased from Aladdin Reagent (Shanghai, China). Acetone, methanol, and acetonitrile used in this work were all high performance liquid chromatography (HPLC)-grade. Lysogeny broth (LB) was obtained from BD Difco (Shanghai, China). The nutrient medium (NM) for strain WY10 was composed of 10 g L⁻¹ glucose,

PHE biodegradation and bacterial growth

The PHE-degradation ability of individual strain (WB9 or WY10) and the defined co-culture (WB9 + WY10) were examined over 144 h (Fig. 1). Results revealed that strain WY10 presented a higher degradation rate than did the reconstructed co-culture during the first 24 h (Fig. 1). However, the co-culture showed a sharp enhancement in PHE degradation thereafter and degraded 100 mg L⁻¹ PHE completely within 36 h. In comparison, over 60 h were required to completely degrade the equal amount of PHE by

Conclusion

A network of metabolic cross-feeding was revealed in the co-culture of Rhodococcus sp. WB9 and Mycobacterium sp. WY10. The end-product of PHE degradation by strain WB9, 1H2N, served as a growth substrate for strain WY10. The promoted strain WY10 growth accelerated PHE degradation. Meanwhile, strain WY10 degraded PHE and 1H2N to form PHTA and POCA that was accumulated in the culture medium and taken up by strain WB9, followed by the degradation towards TCA cycle. The metabolic cross-feeding of

CRediT authorship contribution statement

Shanshan Sun: Investigation, Methodology, Data curation, Writing - original draft. Haizhen Wang: Conceptualization, Writing - review & editing, Supervision, Funding acquisition. Kang Yan: Investigation, Validation. Jun Lou: Methodology. Jiahui Ding: Investigation, Validation. Shane A. Snyder: Writing - review & editing. Laosheng Wu: Writing - review & editing. Jianming Xu: Writing - review & editing, Resources.

Declaration of Competing Interest

This article has no conflict of interest.

Acknowledgements

This work was supported by the National Key Research and Development Program of China (2019YFC1803704); and the National Natural Science Foundation of China (41771344).

References (34)

S. Festa et al.
Bacterial diversity and functional interactions between bacterial strains from a phenanthrene-degrading consortium obtained from a chronically contaminated-soil
Int. Biodeterior. Biodegradation
(2013)
A. Janbandhu et al.
Biodegradation of phenanthrene using adapted microbial consortium isolated from petrochemical contaminated environment
J. Hazard. Mater.
(2011)
T.G. Luan et al.
Study of metabolites from the degradation of polycyclic aromatic hydrocarbons (PAHs) by bacterial consortium enriched from mangrove sediments
Chemosphere
(2006)
S. Rabodonirina et al.
Degradation of ﬂuorene and phenanthrene in PAHs-contaminated soil using Pseudomonas and Bacillus strains isolated from oil spill sites
J. Environ. Manage.
(2019)
R.L. Stingley et al.
Molecular characterization of a phenanthrene degradation pathway in Mycobacterium vanbaalenii PYR-1
Biochem. Biophys. Res. Commun.
(2004)
S.R. Subashchandrabose et al.
Bioremediation of soil long-term contaminated with PAHs by algal-bacterial synergy of Chlorella sp. MM3 and Rhodococcus wratislaviensis strain 9 in slurry phase
Sci. Total Environ.
(2019)
S.S. Sun et al.
Salicylate and phthalate pathways contributed diﬀ ;erently on phenanthrene and pyrene degradations in Mycobacterium sp. WY10
J. Hazard. Mater.
(2019)
S.S. Sun et al.
Non-bioavailability of extracellular 1-hydroxy-2-naphthoic acid restricts the mineralization of phenanthrene by Rhodococcus sp
WB9. Sci. Total Environ.
(2020)
P. Wanapaisan et al.
Synergistic degradation of pyrene by five culturable bacteria in a mangrove sedimentderived bacterial consortium
J. Hazard. Mater.
(2018)
B.B. Wang et al.
Effect of mixed soil microbiomes on pyrene removal and the response of the soil microorganisms
Sci. Total Environ.
(2018)

Y. Xu et al.

Heterologous expression and localization of gentisate transporter Ncg12922 from Corynebacterium glutamicum ATCC 13032

Biochem. Biophys. Res. Commun.

(2006)

Y. Zhong et al.

Production of metabolites in the biodegradation of phenanthrene, fluoranthene and pyrene by the mixed culture of Mycobacterium sp. and Sphingomonas sp

Bioresour. Technol.

(2011)

M.E. Acuña et al.

Microbiological and kinetic aspects of a biofilter for the removal of toluene from waste gases

Biotechnol. Bioeng.

(1999)

N.R. Draper et al.

Applied Regression Analysis

(1981)

H.P. Gu et al.

Biodegradation, biosorption of phenanthrene and its trans-membrane transport by Massilia sp. WF1 and Phanerochaete Chrysosporium

Front. Microbiol.

(2016)

C.T. Hennessee et al.

Effects of PAH mixtures on degradation, gene expression, and metabolite production in four Mycobacterium species

Appl. Environ. Microbiol.

(2016)

W. Hong et al.

Distribution, fate, inhalation exposure and lung cancer risk of atmospheric polycyclic aromatic hydrocarbons in some Asian countries

Environ. Sci. Technol.

(2016)

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Metabolic interactions in a bacterial co-culture accelerate phenanthrene degradation

Highlights

Abstract

Graphical abstract

Introduction

Section snippets

Chemicals and media

PHE biodegradation and bacterial growth

Conclusion

CRediT authorship contribution statement

Declaration of Competing Interest

Acknowledgements

Int. Biodeterior. Biodegradation

J. Hazard. Mater.

Chemosphere

J. Environ. Manage.

Biochem. Biophys. Res. Commun.

Sci. Total Environ.

J. Hazard. Mater.

WB9. Sci. Total Environ.

J. Hazard. Mater.

Sci. Total Environ.

Biochem. Biophys. Res. Commun.

Bioresour. Technol.

Microbiological and kinetic aspects of a biofilter for the removal of toluene from waste gases

Biotechnol. Bioeng.

Applied Regression Analysis

Biodegradation, biosorption of phenanthrene and its trans-membrane transport by Massilia sp. WF1 and Phanerochaete Chrysosporium

Front. Microbiol.

Effects of PAH mixtures on degradation, gene expression, and metabolite production in four Mycobacterium species

Appl. Environ. Microbiol.

Distribution, fate, inhalation exposure and lung cancer risk of atmospheric polycyclic aromatic hydrocarbons in some Asian countries

Environ. Sci. Technol.