Diversity of bacterial phenol hydroxylase-encoding genes from gasoline-contaminated silt soil




Abstract. Vasandani SR, Tan WA. 2022. Diversity of bacterial phenol hydroxylase-encoding genes from gasoline-contaminated silt soil. Biodiversitas 23: 5502-5506. Phenol is an aromatic compound often used as a raw material or intermediate in various industries. Improper handling and disposal may lead to the accumulation of this hazardous compound. Bioremediation is the most viable method to remove phenol from contaminated environment. In this study, the diversity of genes that encode for phenol hydroxylase, a key enzyme in phenol degradation, was assessed in gasoline-contaminated soil from a commercial gas station in Central Jakarta, Indonesia. Partial phenol hydroxylase-encoding gene library was constructed using a pair of universal primer in the pGEM®-Teasy vector. A total of 30 recombinant clones were obtained and sequenced to analyze the genetic diversity of this gene. Obtained clones were 86-99% identical to phenol hydroxylase-related proteins. Phylogenetic tree analysis on amino acid sequences derived from our library revealed that 56.7% of the cloned fragments were closely related to Proteobacteriodota and 20.0% of them were clustered with Actinomycetota. The rest 23.3% of the clones formed a cluster separate from any of the reference sequences, possibly indicating the presence of novel phenol hydroxylase genes.


Anku WW, Mamo MM, Govender PP. 2017. Phenolic compounds in water: sources, reactivity, toxicity and treatment methods. In: Soto-Hernandez M, Palma-Tenango M, Garcia-Mateos R , editors. Phenolic Compounds - Natural Sources, Importance and Applications. London (GB): IntechOpen, Page 419-443. doi: 10.5772/66927.
Bai Y, Müller DB, Srinivas G, Garrido-Oter R, Potthoff E, Rott M, Dombrowski N, Münch PC, Spaepen S, Remus-Emsermann M, Hüttel B, McHardy AC, Vorholt JA, Schulze-Lefert P. 2015. Functional overlap of the Arabidopsis leaf and root microbiota. Nature. 528: 364-369. doi:10.1038/nature16192.
Barka EA, Vatsa P, Sanchez L, Gaveau-Vaillant N, Jacquard C, Klenk HP, Clément C, Ouhdouch Y, van Wezel GP. 2016. Taxonomy, physiology, and natural products of Actinobacteria. Microbiol Mol Biol Rev. 80:1– 43. doi:10.1128/M MBR.00019-15.
Barton N, Horbal L, Starck S, Kohlstedt M, Luzhetskyy A, Wittmann C. 2018. Enabling the valorization of guaiacol-based lignin: Integrated chemical and biochemical production of cis,cis-muconic acid using metabolically engineered Amycolatopsis sp ATCC 39116. MBE 45:200-210. doi: 10.1016/j.ymben.2017.12.001.
Bernhardt ES, Rosi EJ, Gessner MO. 2017. Synthetic chemicals as agents of global change. Front Ecol Environ. 15:84–90. doi:10.1002/fee.1450.
Cai S, Chen LW, Ai YC, Qiu JG, Wang CH, Shi C, He J, Caib TM. 2017. Degradation of diphenyl ether in Sphingobium phenoxybenzoativorans SC_3 is initiated by a novel ring cleavage dioxygenase. Appl Environ Microbiol. 83(10):104-117. doi:10.1128/AEM.00104-17.
Choi EJ, Jin HM, Lee SH, Math RK, Madsen EL, Jeon CO. 2012. Comparative genomic analysis and benzene, toluene, ethylbenzene, and o-, m-, and p-xylene (BTEX) degradation pathways of Pseudoxanthomonas spadix BD-a59. Appl Environ Microbiol. 79(2):663-671. doi:10.1128/AEM.02809-12.
Duan W, Meng F, Cui H, Lin Y, Wang G, Wu J. 2018. Ecotoxicity of phenol and cresols to aquatic organisms: A review. Ecotoxicol Environ Saf. 157(March):441–456. doi:10.1016/j.ecoenv.2018.03.089.
Emelyanova EV, Solyanikova IP. 2020. Evaluation of phenol-degradation activity of Rhodococcus opacus 1CP using immobilized and intact cells. Int J Environ Sci Technol 17(4):2279-94. doi: 10.1007/s13762-019-02609-8
Fierer N. 2017. Embracing the unknown: disentangling the complexities of the soil microbiome. Nat Rev Microbiol. 15(10): 579–590. doi:10.1038/nrmicro.2017.87.
Futamata H, Harayama S, Watanabe K. 2001. Group-specific monitoring of phenol hydroxylase genes for a functional assessment of phenol-stimulated trichloroethylene bioremediation. Appl Environ Microbiol. 67:4671–7. doi:10.1128/AEM.67.10.4671-4677.2001.
Hugerth LW, Wefer HA, Lundin S, Jakobsson HE, Lindberg M, Rodin S, Engstrand L, Andersson AF. 2014. DegePrime, a program for degenerate primer design for broad-taxonomic-range PCR in microbial ecology studies. Appl Environ Microbiol. 80:5116–5123. doi:10.1128/AEM.01403-14.
Khor N. 2014. Generation of a Zn2+ - free oxygenase of phenol hydroxylase from Pseudomonas sp. strain CF600 [Thesis]. Quebec: Department of Chemistry and Biochemistry, Concordia University.
Krastanov A, Alexieva Z, Yemendzhiev H. 2012. Microbial degradation of phenol and phenolic derivatives. Eng Life Sci 13: 76-87. Doi: 10.1002/elsc.201100227.
Kumar S, Stecher G, Tamura K. 2016. MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol and Evol. 33:1870–1874. doi:10.1093/molbev/msw054.
Linder JU. 2015. The YHS-domain of an adenylyl cyclase from Mycobacterium phlei is a probable copper-sensor module. PLoS ONE. 10:1–11. doi:10.1371/journal.pone.0141843.
Messing J. 1983. New M13 vectors for cloning. Method Enzymol. 101:20-78. Doi:10.1016/0076-6879(83)01005-8.
Nešvera J, Rucká L, Pátek M. 2015. Catabolism of phenol and its derivatives in bacteria: genes, their regulation, and use in the biodegradation of toxic pollutants. Adv Appl Microbiol. 93:107–160. doi:10.1016/bs.aambs.2015.06.002.
Nogina T, Fomina M, Dumanskaya T, Zelena L, Khomenko L, Mikhalovsky S, Podgorskyi V, Gadd GM. A new Rhodococcus aetherivorans strain isolated from lubricant-contaminated soil as a prospective phenol-biodegrading agent. Appl Microbiol Biotechnol 104: 3611–3625. doi: 10.1007/s00253-020-10385-6.
Otsuka NY, Muramatsu Y, Nakagawa Y, Matsuda M, Nakamura M, Murata H. 2011. Burkholderia oxyphilla sp. nov., a bacterium isolated from acidic forest soil that catabolises (+)-cathechin and its putative aromatic derivatives. Int J Syst Evol Microbiol. 61: 249-254. doi:10.1099/ijs.0.017368-0.
Petkevi?ius V, Vaitek?nas J, Vaitkus D, ??nas N, Meškys R. 2019. Tailoring a Soluble Diiron Monooxygenase for Synthesis of Aromatic N-oxides. Catalysts. 9(4):356. Doi:10.3390/catal9040356.
Reichert K, Lipski A, Pradela S, Stackebrandt E, Altendorf K. 1998. Pseudonocardia asaccharolytica sp. nov and Pseudonocardia sulfidoxydans sp. nov., two new dimethyl disulphide-degrading actinomycetes and emended description of the genus Pseudonocardia. Int J Syst Bacteriol. 48:441-449. doi:10.1099/00207713-48-2-441.
Sarwade VD, Gawai KR. 2014. Biodegradation of phenol by alkaliphiic Bacillus badius D1. IOSR-JESTFT. 8(5):28-35. doi:10.9790/2402-08522835.
Silva CC, Hayden H, Sawbridge T, Mele P, De Paula SO, Silva LCF, Vidigal PMP, Vincentini M, Sousa MP, Torres APR, et al. 2013. Identification of genes and pathways related to phenol degradation in metagenomic libraries from petroleum refinery wastewater. PLoS ONE 8(4): e61811. doi:10.1371/journal.pone.0061811.
Sun JQ, Xu L, Liu XY, Zhao GF, Cai H, Nie Y, Wu XL. 2018. Functional genetic diversity and culturability of petroleum-degrading bacteria isolated from oil-contaminated soils. Front Microbiol. 9:1332. doi:10.3389/fmicb.2018.01332
Surkatti R, El-Naas MH. 2017. Competitive interference during the biodegradation of cresols. Int J Environ Sci Technol. 15(2):301–308. doi:10.1007/s13762-017-1383-2.
Tan WA, Kusuma F. 2021. Genetic diversity of phenol hydroxylase-encoding genes among wastewater sludge bacteria. Biodiversitas J Biol Diversity. 22(10):4291-4297.doi:10.13057/biodiv/d221021
Tan WA, Parales RE. 2016. Application of aromatic hydrocarbon dioxygenases. In: Patel RN, Editor. Green Biocatalysis. Hoboken (US): John Wiley & Sons. Page: 457-472. doi:10.1002/9781118828083.ch17.
Thakurta GS, Aakula M, Chakrabarty J, Dutta S. 2018. Bioremediation of phenol from synthetic and real wastewater using Leptolyngbya sp.: a comparison and assessment of lipid production. 3 Biotech. 8(4):206. doi:10.1007/s13205-018-1229-8.
Zhao L, Wu Q, Aijin M. 2017. Biodegradation of phenolic contaminants: current status and perspectives. IOP Conf Ser: Earth Environ Sci. 111: 1-5. doi :10.1088/1755-1315/111/1/012024.