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Subtractive hybridization and random arbitrarily primed PCR analyses of a benzoate-assimilating bacterium, Desulfotignum balticum

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Abstract

Subtractive hybridization (SH) and random arbitrarily primed PCR (RAP-PCR) were used to detect genes involved in anaerobic benzoate degradation by Desulfotignum balticum. Through SH, we obtained 121 DNA sequences specific for D. balticum but not for D. phosphitoxidans (a non-benzoate-assimilating species). Furthermore, RAP-PCR analysis showed that a 651-bp DNA fragment, having 55% homology with the solute-binding protein of the ABC transporter system in Methanosarcina barkeri, was expressed when D. balticum was grown on benzoate, but not on pyruvate. By shotgun sequencing of the fosmid clone (38,071 bp) containing the DNA fragment, 33 open reading frames (ORFs) and two incomplete ORFs were annotated, and several genes within this region corresponded to the DNA fragments obtained by SH. An 11.3-kb gene cluster (ORF10–17) revealed through reverse transcription-PCR showed homology with the ABC transporter system and TonB-dependent receptors, both of which are presumably involved in the uptake of siderophore/heme/vitamin B12, and was expressed in response to growth on benzoate.

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References

  1. Akopyants NS, Fradkov A, Diatchenko L, Hill JE, Siebert PD, Lukyanov SA, Sverdlov ED, Berg DE (1998) PCR-based subtractive hybridization and differences in gene content among strains of Helicobacter pylori. Proc Natl Acad Sci U S A 95:13108–13113

  2. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3420

  3. Bak F, Widdel F (1986) Anaerobic degradation of phenol and phenol derivatives by Desulfobacterium phenolicum sp. nov. Arch Microbiol 146:177–180

  4. Boll M, Fuchs G, Heider H (2002) Anaerobic oxidation of aromatic compounds and hydrocarbons. Curr Opin Chem Biol 6:604–611

  5. Cord-Ruwisch R, Carcia JL (1985) Isolation and characterization of an anaerobic benzoate-degrading spore-forming sulfate-reducing bacterium, Desulfotomaculum spomandens sp. nov. FEMS Microbiol Lett 29:325–330

  6. Drzyzga O, Küver J, Blotevogel K-H (1993) Complete oxidation of benzoate and 4-hydroxybenzoate by a new sulfate-reducing bacterium resembling Desulfoarculus. Arch Microbiol 159:109–113

  7. Galushko A, Minz D, Schink B, Widdel F (1999) Anaerobic degradation of naphthalene by a pure culture of a novel type of marine sulfate-reducing bacterium. Environ Microbiol 1:415–420

  8. Hammann B, Kutsver HJ (1998) Key enzymes for the degradation of benzoate, m- and p-hydroxybenzoate by some members of the order Actinomycetales. J Basic Microbiol 38:207–220

  9. Harms G, Zengler K, Rabus R, Aeckersberg F, Minz D, Rosselló-Mora R, Widdel F (1999) Anaerobic oxidation of o-xylene, m-xylene, and homologous alkylbenzenes by new types of sulfate-reducing bacteria. Appl Environ Microbiol 65:999–1004

  10. Harwood CS, Burchardt G, Herrmann H, Fuchs G (1999) Anaerobic metabolism of aromatic compounds via the benzoyl-CoA pathway. FEMS Microbiol Rev 22:439–458

  11. Hosoda A, Kasai Y, Hamamura N, Takahata Y, Watanabe K (2005) Development of a PCR method for the detection and quantification of benzoyl-CoA reductase genes and its application to monitored natural attenuation. Biodegradation 16:591–601

  12. Joo LM, Macfarlane-Smith LR, Okeke IN (2007) Error-prone DNA repair system in enteroaggregative Escherichia coli identified by subtractive hybridization. J Bacteriol 189:3793–3803

  13. Kerpola RE, Ames GFL (1992) Topology of the hydrophobic membrane-bound components of the histidine periplasmic permeases. J Biol Chem 267:2329–2336

  14. Kniemeyer O, Fischer T, Wilkes H, Glöckner FO, Widdel F (2003) Anaerobic degradation of ethylbenzene by a new type of marine sulfate-reducing bacterium. Appl Environ Microbiol 69:760–768

  15. Koebnik R (2005) TonB-dependent trans-envelope signaling: the exception or the rule. Trends Microbiol 13:343–347

  16. Köster W (2001) ABC transporter-mediated uptake of iron, siderophores, heme and vitamin B12. Res Microbiol 152:291–301

  17. Kuever J, Könneke M, Galushko A, Drzyzga O (2001) Reclassification of Desulfobacterium phenolicum as Desulfobacula phenolica comb. nov. and description of strain SaxT as Desulfotignum balticum gen. nov., sp. nov. Int J Syst Evol Microbiol 51:171–177

  18. Lee C-T, Amaro C, Sanjuán E, Hor L-I (2005) Identification of DNA sequences specific for Vibrio vulnificus biotype 2 strains by suppression subtractive hybridization. Appl Environ Microbiol 71:5593–5597

  19. Morasch B, Schink B, Tebbe CC, Meckenstock RU (2004) Degradation of o-xylene and m-xylene by a novel sulfate-reducer belonging to the genus Desulfotomaculum. Arch Microbiol 181:407–417

  20. Müller JA, Galushko AS, Kappler A, Schink B (1999) Anaerobic degradation of m-cresol by Desulfobacterium cetonicum is initiated by formation of 3-hydroxybenzylsuccinate. Arch Microbiol 172:287–294

  21. Müller JA, Galushko AS, Kappler A, Schink B (2001) Initiation of anaerobic degradation of p-cresol by formation of 4-hydroxybenzylsuccinate in Desulfobacterium cetonicum. J Bacteriol 183:752–757

  22. Nojiri H, Sekiguchi H, Maeda K, Urata M, Nakai S, Yoshida T, Habe H, Omori T (2001) Genetic characterization and evolutionary implications of a car gene cluster in the carbazole degrader Pseudomonas sp. strain CA10. J Bacteriol 183:3663–3679

  23. Peters F, Rother M, Boll M (2004) Selenocystein-containing proteins in anaerobic benzoate metabolism of Desulfococcus multivorans. J Bacteriol 186:2156–2163

  24. Peters F, Shinoda Y, McInerney MJ, Boll M (2007) Cyclohexa-1,5-diene-1- carbonyl-coenzyme A (CoA) hydratases of Geobacter metallireducens and Syntrophus aciditrophicus: evidence for a common benzoyl-CoA degradation pathway in facultative and strict anaerobes. J Bacteriol 189:1055–1060

  25. Proff C, Kölling R (2001) Functional asymmetry of the two nucleotide binding domains in the ABC transporter Ste6. Mol Gen Genet 264:883–893

  26. Rabus R, Nordhaus R, Ludwig W, Widdel F (1993) Complete oxidation of toluene under strictly anoxic conditions by a new sulfate-reducing bacterium. Appl Environ Microbiol 59:1444–1451

  27. Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edrd edn. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY

  28. Schink B, Thiemann V, Laue H, Friedrich MW (2002) Desulfotignum phosphitoxidans sp. nov., a new marine sulfate reducer that oxidizes phosphite to phosphate. Arch Microbiol 177:381–391

  29. Shepard BD, Gilmore MS (1999) Identification of aerobically and anaerobically induced genes in Enterococcus faecalis by random arbitrarily primed PCR. Appl Environ Microbiol 65:1470–1476

  30. Smoot LM, Franke DD, McGillivary G, Actis LA (2002) Genomic analysis of the F3031 Brazilian purpuric fever clone of Haemophilus influenzae biogroup aegyptus by PCR-based subtractive hybridization. Infect Immun 70:2694–2699

  31. Walker JE, Saraste M, Runswick MJ, Gay NJ (1982) Distantly related sequences in the a- and b-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. EMBO J 1:945–951

  32. Widdel F, Pfennig N (1984) Dissimilatory sulfate- or sulfur-reducing bacteria. In: Krieg NR, Holt JG (eds) Bergey’s manual of systematic bacteriology, IX edn., vol. I. Williams & Wilkins, Baltimore, MD, pp 663–679

  33. Widdel F, Kohring GW, Mayer F (1983) Entides on dissimilatory sulfate-reducing bacteria that decompose fatty acids. II. Characterization of the filamentous gliding Desulfonema limicola gen. nov., sp. nov., and Desulfonema magnum sp. nov. Arch Microbiol 134:286–294

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Acknowledgments

This work was carried out as a part of “The Project for Development of Technologies for Analyzing and Controlling the Mechanism of Biodegrading and Processing,” which was supported by the New Energy and Industrial Technology Development Organization (NEDO).

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Correspondence to Hiroshi Habe.

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Habe, H., Kobuna, A., Hosoda, A. et al. Subtractive hybridization and random arbitrarily primed PCR analyses of a benzoate-assimilating bacterium, Desulfotignum balticum . Appl Microbiol Biotechnol 79, 87–95 (2008). https://doi.org/10.1007/s00253-008-1414-5

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Keywords

  • Sulfate-reducing bacteria
  • ABC transporter
  • Desulfotignum balticum
  • Benzoate assimilation