Widespread and active piezotolerant microorganisms mediate phenolic compound degradation under high hydrostatic pressure in hadal trenches

Abstract

Phenolic compounds, as well as other aromatic compounds, have been reported to be abundant in hadal trenches. Although high-throughput sequencing studies have hinted at the potential of hadal microbes to degrade these compounds, direct microbiological, genetic and biochemical evidence under in situ pressures remain absent. Here, a microbial consortium and a pure culture of Pseudomonas, newly isolated from Mariana Trench sediments, efficiently degraded phenol under pressures up to 70 and 60 MPa, respectively, with concomitant increase in biomass. By analyzing a high-pressure (70 MPa) culture metatranscriptome, not only was the entire range of metabolic processes under high pressure generated, but also genes encoding complete phenol degradation via ortho- and meta-cleavage pathways were revealed. The isolate of Pseudomonas also contained genes encoding the complete degradation pathway. Six transcribed genes (dmpKLMNOP _sed) were functionally identified to encode a multicomponent hydroxylase catalyzing the hydroxylation of phenol and its methylated derivatives by heterogeneous expression. In addition, key catabolic genes identified in the metatranscriptome of the high-pressure cultures and genomes of bacterial isolates were found to be all widely distributed in 22 published hadal microbial metagenomes. At microbiological, genetic, bioinformatics, and biochemical levels, this study found that microorganisms widely found in hadal trenches were able to effectively drive phenolic compound degradation under high hydrostatic pressures. This information will bridge a knowledge gap concerning the microbial aromatics degradation within hadal trenches.

Keywords

Hadal trench / High hydrostatic pressure / Phenolic compounds degradation / Piezotolerant microorganisms / Widespread distribution

References

[1] Anku WW, Mamo MA, Govender PP (2017) Phenolic compounds in water: sources, reactivity, toxicity and treatment methods. In: Phenolic compounds—natural sources, importance and applications. InTech, pp 419–443

[2] Bertani G. Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli. J Bacteriol, 1951, 62: 293-300,

[3] Bertani G. Lysogeny at mid-twentieth century: P1, P2, and other experimental systems. J Bacteriol, 2004, 186: 595-600,

[4] Bu D, Luo H, Huo P, Wang Z, Zhang S, He Z, Wu Y, Zhao L, Liu J, Guo J, Fang S, Cao W, Yi L, Zhao Y, Kong L. KOBAS-i: intelligent prioritization and exploratory visualization of biological functions for gene enrichment analysis. Nucleic Acids Res, 2021, 49: W317-W325,

[5] Buchfink B, Xie C, Huson DH. Fast and sensitive protein alignment using diamond. Nat Methods, 2015, 12: 59-60,

[6] Chen S, Zhou Y, Chen Y, Gu J. Fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics, 2018, 34: i884-i890,

[7] Chen P, Zhou H, Huang Y, Xie Z, Zhang M, Wei Y, Li J, Ma Y, Luo M, Ding W, Cao J, Jiang T, Nan P, Fang J, Li X. Revealing the full biosphere structure and versatile metabolic functions in the deepest ocean sediment of the Challenger Deep. Genome Biol, 2021, 22: 207,

[8] Chen X, Sheng Y, Wang G, Guo L, Zhang H, Zhang F, Yang T, Huang D, Han X, Zhou L. Microbial compositional and functional traits of BTEX and salinity co-contaminated shallow groundwater by produced water. Water Res, 2022, 215: 118277,

[9] Crooks GE, Hon G, Chandonia JM, Brenner SE. WebLogo: a sequence logo generator. Genome Res, 2004, 14: 1188-1190,

[10] Cui J, Yu Z, Mi M, He L, Sha Z, Yao P, Fang J, Sun W. Occurrence of halogenated organic pollutants in hadal trenches of the Western Pacific Ocean. Environ Sci Technol, 2020, 54: 15821-15828,

[11] Dasgupta S, Peng X, Chen S, Li J, Du M, Zhou YH, Zhong G, Xu H, Ta K. Toxic anthropogenic pollutants reach the deepest ocean on earth. Geochem Perspect Lett, 2018, 7: 22-26,

[12] Doukyu N, Toyoda K, Aono R. Indigo production by Escherichia coli carrying the phenol hydroxylase gene from Acinetobacter sp strain ST-550 in a water-organic solvent two-phase system. Appl Microbiol Biotechnol, 2003, 60: 720-725,

[13] Duan W, Meng F, Cui H, Lin Y, Wang G, Wu J. Ecotoxicity of phenol and cresols to aquatic organisms: a review. Ecotoxicol Environ Saf, 2018, 157: 441-456,

[14] Fuchs G, Boll M, Heider J. Microbial degradation of aromatic compounds—from one strategy to four. Nat Rev Microbiol, 2011, 9: 803-816,

[15] Gao ZM, Huang JM, Cui GJ, Li WL, Li J, Wei ZF, Chen J, Xin YZ, Cai DS, Zhang AQ, Wang Y. In situ meta-omic insights into the community compositions and ecological roles of hadal microbes in the Mariana Trench. Environ Microbiol, 2019, 21: 4092-4108,

[16] Glud RN, Wenzhofer F, Middelboe M, Oguri K, Turnewitsch R, Canfield DE, Kitazato H. High rates of microbial carbon turnover in sediments in the deepest oceanic trench on earth. Nat Geosci, 2013, 6: 284-288,

[17] González-Gaya B, Martínez-Varela A, Vila-Costa M, Casal P, Cerro-Gálvez E, Berrojalbiz N, Lundin D, Vidal M, Mompeán C, Bode A, Jiménez B, Dachs J. Biodegradation as an important sink of aromatic hydrocarbons in the oceans. Nat Geosci, 2019, 12: 119-125,

[18] Haas BJ, Papanicolaou A, Yassour M, Grabherr M, Blood PD, Bowden J, Couger MB, Eccles D, Li B, Lieber M, MacManes MD, Ott M, Orvis J, Pochet N, Strozzi F, Weeks N, Westerman R, William T, Dewey CN, Henschel R, et al.. De novo transcript sequence reconstruction from RNA-seq using the trinity platform for reference generation and analysis. Nat Protoc, 2013, 8: 1494-1512,

[19] Hinteregger C, Leitner R, Loidl M, Ferschl A, Streichsbier F. Degradation of phenol and phenolic compounds by Pseudomonas putida EKII. Appl Microbiol Biotechnol, 1992, 37: 252-259,

[20] Hyatt D, Chen GL, Locascio PF, Land ML, Larimer FW, Hauser LJ. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics, 2010, 11: 119,

[21] Ichino MC, Clark MR, Drazen JC, Jamieson A, Jones DOB, Martin AP, Rowden AA, Shank TM, Yancey PH, Ruhl HA. The distribution of benthic biomass in hadal trenches: A modelling approach to investigate the effect of vertical and lateral organic matter transport to the seafloor. Deep-Sea Res Part I, 2015, 100: 21-33,

[22] Ismail WM, Ye Y, Tang H. Gene finding in metatranscriptomic sequences. BMC Bioinformatics, 2014, 15: S8,

[23] Itou M, Matsumura I, Noriki S. A large flux of particulate matter in the deep Japan Trench observed just after the 1994 Sanriku-Oki earthquake. Deep-Sea Res PT I, 2000, 47: 1987-1998,

[24] Jamieson AJ, Fujii T, Mayor DJ, Solan M, Priede IG. Hadal trenches: the ecology of the deepest places on earth. Trends Ecol Evol, 2010, 25: 190-197,

[25] Jamieson AJ, Malkocs T, Piertney SB, Fujii T, Zhang Z. Bioaccumulation of persistent organic pollutants in the deepest ocean fauna. Nat Ecol Evol, 2017, 1: 51,

[26] Jebbar M, Franzetti B, Girard E, Oger P. Microbial diversity and adaptation to high hydrostatic pressure in deep-sea hydrothermal vents prokaryotes. Extremophiles, 2015, 19: 721-740,

[27] Kanehisa M, Sato Y, Morishima K. BlastKOALA and GhostKOALA: KEGG tools for functional characterization of genome and metagenome sequences. J Mol Biol, 2016, 428: 726-731,

[28] Kopylova E, Noe L, Touzet H. SortMeRNA: fast and accurate filtering of ribosomal RNAs in metatranscriptomic data. Bioinformatics, 2012, 28: 3211-3217,

[29] Kukor JJ, Olsen RH. Complete nucleotide sequence of tbuD, the gene encoding phenol/cresol hydroxylase from Pseudomonas pickettii PKO1, and functional analysis of the encoded enzyme. J Bacteriol, 1992, 174: 6518-6526,

[30] Laalami S, Zig L, Putzer H. Initiation of mRNA decay in bacteria. Cell Mol Life Sci, 2014, 71: 1799-1828,

[31] Letunic I, Bork P. Interactive Tree Of Life (iTOL) v5: an online tool for phylogenetic tree display and annotation. Nucleic Acids Res, 2021, 49: W293-W296,

[32] Ley Y, Cheng XY, Ying ZY, Zhou NY, Xu Y. Characterization of two marine lignin-degrading consortia and the potential microbial lignin degradation network in nearshore regions. Microbiol Spectr, 2023, 11: e0442422,

[33] Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R Genome Project Data Processing S. The sequence alignment/map format and SAMtools. Bioinformatics, 2009, 25: 2078-2079,

[34] Li P, Tao J, Lin J, He C, Shi Q, Li X, Zhang C. Stratification of dissolved organic matter in the upper 2000 m water column at the Mariana Trench. Sci Total Environ, 2019, 668: 1222-1231,

[35] Li T, Xu J, Brower AL, Xu ZJ, Xu Y, Spain JC, Zhou NY. Molecular basis and evolutionary origin of 1-nitronaphthalene catabolism in Sphingobium sp. strain JS3065. Appl Environ Microbiol, 2023, 89: e0172822,

[36] Liao Y, Smyth GK, Shi W. Feature counts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics, 2014, 30: 923-930,

[37] Liu J, Zheng Y, Lin H, Wang X, Li M, Liu Y, Yu M, Zhao M, Pedentchouk N, Lea-Smith DJ, Todd JD, Magill CR, Zhang WJ, Zhou S, Song D, Zhong H, Xin Y, Yu M, Tian J, Zhang X-H. Proliferation of hydrocarbon-degrading microbes at the bottom of the Mariana Trench. Microbiome, 2019, 7: 47,

[38] Liu R, Wang Z, Wang L, Li Z, Fang J, Wei X, Wei W, Cao J, Wei Y, Xie Z. Bulk and active sediment prokaryotic communities in the Mariana and Mussau Trenches. Front Microbiol, 2020, 11: 1521,

[39] Liu R, Wei X, Song W, Wang L, Cao J, Wu J, Thomas T, Jin T, Wang Z, Wei W, Wei Y, Zhai H, Yao C, Shen Z, Du J, Fang J. Novel Chloroflexi genomes from the deepest ocean reveal metabolic strategies for the adaptation to deep-sea habitats. Microbiome, 2022, 10: 75,

[40] Lochab B, Shukla S, Varma IK. Naturally occurring phenolic sources: monomers and polymers. RSC Adv, 2014, 4: 21712-21752,

[41] Martin M. Cutadapt removes adapter sequences from high-throughput sequencing reads. Embnet J, 2011, 17: 10-12,

[42] Nesvera J, Rucka L, Patek M. Catabolism of phenol and its derivatives in bacteria: genes, their regulation, and use in the biodegradation of toxic pollutants. Adv Appl Microbiol, 2015, 93: 107-160,

[43] Nunoura T, Takaki Y, Hirai M, Shimamura S, Makabe A, Koide O, Kikuchi T, Miyazaki J, Koba K, Yoshida N, Sunamura M, Takai K. Hadal biosphere: insight into the microbial ecosystem in the deepest ocean on earth. Proc Natl Acad Sci USA, 2015, 112: E1230-1236,

[44] Nunoura T, Nishizawa M, Hirai M, Shimamura S, Harnvoravongchai P, Koide O, Morono Y, Fukui T, Inagaki F, Miyazaki J, Takaki Y, Takai K. Microbial diversity in sediments from the bottom of the Challenger Deep, the Mariana Trench. Microbes Environ, 2018, 33: 186-194,

[45] Oger PM, Jebbar M. The many ways of coping with pressure. Res Microbiol, 2010, 161: 799-809,

[46] Pathom-Aree W, Stach JE, Ward AC, Horikoshi K, Bull AT, Goodfellow M. Diversity of actinomycetes isolated from Challenger Deep sediment (10,898 m) from the Mariana Trench. Extremophiles, 2006, 10: 181-189,

[47] Pathom-Aree W, Nogi Y, Sutcliffe IC, Ward AC, Horikoshi K, Bull AT, Goodfellow M. Dermacoccus abyssi sp. nov., a piezotolerant actinomycete isolated from the Mariana Trench. Int J Syst Evol Microbiol, 2006, 56: 1233-1237,

[48] Powlowski J, Shingler V. Genetics and biochemistry of phenol degradation by Pseudomonas sp. CF600. Biodegradation, 1994, 5: 219-236,

[49] Saito Y, Sato T, Nomoto K, Tsuji H. Identification of phenol- and p-cresol-producing intestinal bacteria by using media supplemented with tyrosine and its metabolites. FEMS Microbiol Ecol, 2018, 94: fiy125,

[50] Sazinsky MH, Dunten PW, McCormick MS, DiDonato A, Lippard SJ. X-ray structure of a hydroxylase-regulatory protein complex from a hydrocarbon-oxidizing multicomponent monooxygenase, Pseudomonas sp. OX1 phenol hydroxylase. Biochemistry, 2006, 45: 15392-15404,

[51] Schneider TD, Stephens RM. Sequence logos: a new way to display consensus sequences. Nucleic Acids Res, 1990, 18: 6097-6100,

[52] Shigemitsu M, Yokokawa T, Uchida H, Kawagucci S, Murata A. Sedimentary supply of humic-like fluorescent dissolved organic matter and its implication for chemoautotrophic microbial activity in the Izu-Ogasawara Trench. Sci Rep, 2021, 11: 19006,

[53] Shingler V, Powlowski J, Marklund U. Nucleotide sequence and functional analysis of the complete phenol/3,4-dimethylphenol catabolic pathway of Pseudomonas sp. strain CF600. J Bacteriol, 1992, 174: 711-724,

[54] Simoneit BRT (2018) Hydrothermal petroleum. In: Hydrocarbons, oils and lipids: diversity, origin, chemistry and fate, pp 1–35

[55] Steiner PA, De Corte D, Geijo J, Mena C, Yokokawa T, Rattei T, Herndl GJ, Sintes E. Highly variable mRNA half-life time within marine bacterial taxa and functional genes. Environ Microbiol, 2019, 21: 3873-3884,

[56] Tamegai H, Li L, Masui N, Kato C. A denitrifying bacterium from the deep sea at 11,000-m depth. Extremophiles, 1997, 1: 207-211,

[57] Tarn J, Peoples LM, Hardy K, Cameron J, Bartlett DH. Identification of free-living and particle-associated microbial communities present in hadal regions of the Mariana Trench. Front Microbiol, 2016, 7: 665,

[58] Tian JW, Fan L, Liu HD, Liu JW, Li Y, Qin QL, Gong Z, Chen HT, Sun ZB, Zou L, Wang XC, Xu HZ, Bartlett D, Wang M, Zhang YZ, Zhang XH, Zhang CLL. A nearly uniform distributional pattern of heterotrophic bacteria in the Mariana Trench interior. Deep Sea Res 1 Oceanogr Res Pap, 2018, 142: 116-126,

[59] Wang Y, Yang J, Lee OO, Dash S, Lau SC, Al-Suwailem A, Wong TY, Danchin A, Qian PY. Hydrothermally generated aromatic compounds are consumed by bacteria colonizing in Atlantis II Deep of the Red Sea. ISME J, 2011, 5: 1652-1659,

[60] Wei Z-F, Li W-L, Huang J-M, Wang Y. Metagenomic studies of SAR202 bacteria at the full-ocean depth in the Mariana Trench. Deep Sea Res 1 Oceanogr Res Pap, 2020, 165: 103396,

[61] Wenzhöfer F, Oguri K, Middelboe M, Turnewitsch R, Toyofuku T, Kitazato H, Glud RN. Benthic carbon mineralization in hadal trenches: assessment by in situ O₂ microprofile measurements. Deep-Sea Res Part I, 2016, 116: 276-286,

[62] Widdel F, Bak F (1992) Gram-negative mesophilic sulfate-reducing bacteria

[63] Wood DE, Lu J, Langmead B. Improved metagenomic analysis with kraken 2. Genome Biol, 2019, 20: 257,

[64] Xu YP, Ge HM, Fang JS. Biogeochemistry of hadal trenches: Recent developments and future perspectives. Deep Sea Res Part II Top Stud Oceanogr, 2018, 155: 19-26,

[65] Xue CX, Liu J, Lea-Smith DJ, Rowley G, Lin H, Zheng Y, Zhu XY, Liang J, Ahmad W, Todd JD, Zhang XH. Insights into the vertical stratification of microbial ecological roles across the deepest seawater column on Earth. Microorganisms, 2020, 8: 1309,

[66] Yan F, Fang J, Cao J, Wei Y, Liu R, Wang L, Xie Z. Halomonas piezotolerans sp. nov., a multiple-stress-tolerant bacterium isolated from a deep-sea sediment sample of the New Britain Trench. Int J Syst Evol Microbiol, 2020, 70: 2560-2568,

[67] Yang S, Li X, Xiao X, Zhuang G, Zhang Y. Sphingomonas profundi sp. nov., isolated from deep-sea sediment of the Mariana Trench. Int J Syst Evol Microbiol, 2020, 70: 3809-3815,

[68] Zhang X, Xu W, Liu Y, Cai M, Luo Z, Li M. Metagenomics reveals microbial diversity and metabolic potentials of seawater and surface sediment from a hadal biosphere at the Yap Trench. Front Microbiol, 2018, 9: 2402,

[69] Zhang X, Xu Y, Xiao W, Zhao M, Wang Z, Wang X, Xu L, Luo M, Li X, Fang J, Fang Y, Wang Y, Oguri K, Wenzhöfer F, Rowden AA, Mitra S, Glud RN. The hadal zone is an important and heterogeneous sink of black carbon in the ocean. Commun Earth Environ, 2022, 3: 25,

[70] Zhou Z, Liu Y, Pan J, Cron BR, Toner BM, Anantharaman K, Breier JA, Dick GJ, Li M. Gammaproteobacteria mediating utilization of methyl-, sulfur- and petroleum organic compounds in deep ocean hydrothermal plumes. ISME J, 2020, 14: 3136-3148,

[71] Zhou YL, Mara P, Cui GJ, Edgcomb VP, Wang Y. Microbiomes in the challenger deep slope and bottom-axis sediments. Nat Commun, 2022, 13: 1515,

[72] Zídková L, Szőköl J, Rucká L, Pátek M, Nešvera J. Biodegradation of phenol using recombinant plasmid-carrying Rhodococcus erythropolis strains. Int Biodeterior Biodegradation, 2013, 84: 179-184,