Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Use of plate-wash samples to monitor the fates of culturable bacteria in mercury- and trichloroethylene-contaminated soils

  • 177 Accesses

  • 9 Citations


With the ultimate aim of developing bioremediation technology that use the optimum bacterial community for each pollutant, we performed polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) and phylogenetic analysis and identified communities of culturable bacteria in HgCl2- and trichloroethylene (TCE)-contaminated soil microcosms. PCR-DGGE band patterns were similar at 0 and 1 ppm HgCl2, but changes in specific bands occurred at 10 ppm HgCl2. Band patterns appearing at 10 and 100 ppm TCE were very different from those at 0 ppm. Phylogenetic analysis showed four bacterial groups in the HgCl2-contaminatied cultures: Firmicutes, Actinobacteria, Proteobacteria, and Bacteroidetes. Most high-density bands, decreased-density bands, and common bands were classified into the phyla Proteobacteria, Actinobacteria, and Firmicutes, respectively; the effects of HgCl2 on culturable bacteria appeared to differ among phyla. Duganella violaceinigra [98.4% similarity to DNA Data Bank of Japan (DDBJ) strain], Lysobacter koreensis (98.2%), and Bacillus panaciterrae (98.6%) were identified as bacteria specific to HgCl2-contaminated soils. Bacteria specific to TCE-contaminated soils were distributed into three phyla (Firmicutes, Proteobacteria, and Actinobacteria), but there was no clear relationship between phylum and TCE effects on culturable bacteria. Paenibacillus kobensis (97.3%), Paenibacillus curdlanolyticus (96.3%), Paenibacillus wynnii (99.8%), and Sphingomonas herbicidovorans (99.4%) were identified as bacteria specific to TCE-contaminated soils. These bacteria may be involved in pollutant degradation.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3


  1. Cabirol N, Villemur R, Perrier J, Jacob F, Fouillet B, Chambon P (1998) Isolation of a methanogenic bacterium, Methanosarcina sp. strain FR, for its ability to degrade high concentrations of perchloroethylene. Can J Microbiol 44:1142–1147

  2. Chang BV, Yang CM, Cheng CH, Yuan SY (2004) Biodegradation of phthalate esters by two bacteria strains. Chemosphere 55:533–538

  3. De J, Remaiah N, Mesquita A, Verlekar XN (2003) Tolerance to various toxicants by marine bacteria highly resistant to mercury. Mar Biotechnol 5:185–193

  4. Demnerova K, Mackova M, Spevakova V, Beranova K, Kochankova L, Lovecka P, Ryslava E, Macek T (2005) Two approaches to biological decontamination of groundwater and soil polluted by aromatics-characterization of microbial populations. Int Microbiol 8:205–211

  5. Dixit V, Bini E, Drozda M, Blum P (2004) Mercury inactivates transcription and the generalized transcription factor TFB in the archaeon Sulfolobus solfataricus. Antimicrob Agents Chemother 48:1993–1999

  6. Ellis RJ, Morgan P, Weightman AJ, Fry JC (2003) Cultivation-dependent and -independent approaches for determining bacterial diversity in heavy-metal-contaminated soil. Appl Environ Microbiol 69:3223–3230

  7. Gannon F, Bridgeland ES, Jones KM (1977) l-serine dehydratase from Arthrobacter globiformis. Biochem J 161:345–355

  8. Habe H, Ide K, Yotsumoto M, Tsuji H, Hirano H, Widada J, Yoshida T, Nojiri H, Omori T (2001) Preliminary examinations for applying a carbazole-degrader, Pseudomonas sp. strain CA10, to dioxin-contaminated soil remediation. Appl Microbiol Biotechnol 56:788–795

  9. Haruta S, Cui Z, Huang Z, Li M, Ishii M, Igarashi Y (2002) Construction of a stable microbial community with high cellulose-degradation ability. Appl Microbiol Biotechnol 59:529–534

  10. Hashimoto A, Iwasaki K, Nakajima M, Yagi O (2001) Quantitative detection of trichloroethylene-degrading Mycobacterium sp. TA27 with a real-time PCR product detection system. Microb Environ 16:109–116

  11. Hashimoto A, Iwasaki K, Nakasugi N, Nakajima M, Yagi O (2002) Degradation pathways of trichloroethylene and 1,1,1-trichloroethane by Mycobacterium sp. TA27. Biosci Biotechnol Biochem 66:385–390

  12. Huang CC, Narita M, Yamagata T, Itoh Y, Endo G (1999) Structure analysis of a class II transposon encoding the mercury resistance of the gram-positive bacterium Bacillus megaterium MB1, a strain isolated from Minamata Bay, Japan. Gene 234:361–369

  13. Iwasaki K, Uchiyama H, Yagi O (1993) Survival and impact of genetically engineered Pseudomonas putida harboring mercury resistance gene in aquatic microcosms. Biosci Biotechnol Biochem 57:1264–1269

  14. Izaki K (1981) Enzymatic reduction of mercurous and mercuric ions in Bacillus cereus. Can J Microbiol 27:192–197

  15. Jonas RB, Gilmour CC, Stoner DL, Weir MM, Tuttle JH (1984) Comparison of methods to measure acute metal and organometal toxicity to natural aquatic microbial communities. Appl Environ Microbiol 47:1005–1011

  16. Kasai Y, Takahata Y, Hoaki T, Watanabe K (2005) Physiological and molecular characterization of a microbial community established in unsaturated, petroleum-contaminated soil. Environ Microbiol 7:806–818

  17. Kato S, Haruta S, Cui ZJ, Ishii M, Igarashi Y (2004) Effective cellulose degradation by a mixed-culture system composed of a cellulolytic Clostridium and aerobic non-cellulolytic bacteria. FEMS Microbiol Ecol 51:133–142

  18. Kimura M (1980) A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120

  19. Mera N, Iwasaki K (2006) Use of plate-wash samples to evaluate bacterial population dynamics in mercury- and trichloroethylene-contaminated soils. J Environ Biotechnol 6:115–122

  20. Mukerjee-Dhar G, Hatta T, Shimura M, Kimbara K (1998) Analysis of changes in congener selectivity during PCB degradation by Burkholderia sp. strain TSN101 with increasing concentrations of PCB and characterization of the bphBCD genes and gene products. Arch Microbiol 169:61–70

  21. Muyzer G, de Waal EC, Uitterlinden AG (1993) Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol 59:695–700

  22. Nakajima-Kambe T, Shigeno-Akutsu Y, Nomura N, Onuma F, Nakahara T (1999) Microbial degradation of polyurethane, polyester, polyurethanes and polyether polyurethanes. Appl Microbiol Biotechnol 51:134–140

  23. Neufeld JD, Mohn WW, de Lorenzo V (2006) Composition of microbial communities in hexachlorocyclohexane (HCH) contaminated soils from Spain revealed with a habitat-specific microarray. Environ Microbiol 8:126–140

  24. Oldenhuis R, Oedzes JY, van der Waarde JJ, Janssen DB (1991) Kinetics of chlorinated hydrocarbon degradation by Methylosinus trichosporium OB3 and toxicity of trichloroethylene. Appl Environ Microbiol 57:7–14

  25. Raj H, Liston J (1961) Detection and enumeration of fecal indicator organisms in frozen sea foods. I. Escherichia coli. Appl Microbiol 9:171–174

  26. Ramakrishnan V, Ogram AV, Lindner AS (2005) Impacts of co-solvent flushing on microbial populations capable of degrading trichloroethylene. Environ Health Perspect 113:55–61

  27. Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425

  28. Sakai M, Ezaki S, Suzuki N, Kurane R (2005) Isolation and characterization of a novel polychlorinated biphenyl-degrading bacterium, Paenibacillus sp. KBC101. Appl Microbiol Biotechnol 68:111–116

  29. Sasaki M, Maki J, Oshiman K, Matsumura Y, Tsuchido T (2005) Biodegradation of bisphenol A by cells and cell lysate from Sphingomonas sp. strain AO1. Biodegradation 16:449–459

  30. Shitashiro M, Kato J, Fukumura T, Kuroda A, Ikeda T, Takiguchi N, Ohtake H (2003) Evaluation of bacterial aerotaxis for its potential use in detecting the toxicity of chemicals to microorganisms. J Biotechnol 101:11–18

  31. Smiejan A, Wilkinson KJ, Rossier C (2003) Cd bioaccumulation by a freshwater bacterium, Rhodospirillum rubrum. Environ Sci Technol 37:701–706

  32. Sulistyaningdyah WT, Ogawa J, Li QS, Shinkyo R, Sakaki T, Inouye K, Schmid RD, Shimizu S (2004) Metabolism of polychlorinated dibenzo-p-dioxins by cytochrome P450 BM-3 and its mutant. Biotechnol Lett 26:1857–1860

  33. Suyama A, Iwakiri R, Kai K, Tokunaga T, Sera N, Furukawa K (2001) Isolation and characterization of Desulfitobacterium sp. strain Y51 capable of efficient dehalogenation of tetrachloroethene and polychloroethanes. Biosci Biotechnol Biochem 65:1474–1481

  34. Taira K, Hirose J, Hayashida S, Furukawa K (1992) Analysis of bph operon from the polychlorinated biphenyl-degrading strain of Pseudomonas pseudoalcaligenes KF707. J Biol Chem 267:4844–4853

  35. Tan W, Shelef LA (1999) Automated detection of Salmonella spp. in foods. J Microbiol Methods 37:87–91

  36. Uchiyama H, Nakajima T, Yagi O, Tabuchi T (1989) Aerobic degradation of trichloroethylene by a new type II methane-utilizing bacterium, strain M. Agric Biol Chem 53:2903–2907

  37. Watanabe N, Sato E, Ose Y (1985) Adsorption and desorption of polydimethylsiloxane, PCBs, cadmium nitrate, copper sulfate, nickel sulfate and zinc nitrate by river surface sediments. Sci Total Environ 41:153–161

  38. Wittich RM, Wilkes H, Sinnwell V, Francke W, Fortnagel P (1992) Metabolism of dibenzo-p-dioxin by Sphingomonas sp. strain RW1. Appl Environ Microbiol 58:1005–1010

  39. Yu SH, Ke L, Wong YS, Tam NFY (2005) Degradation of polycyclic aromatic hydrocarbons (PAHS) by a bacterial consortium enriched from mangrove sediments. Environ Int 31:149–154

  40. Zaitsev GM, Tsoi TV, Grishenkov VG, Plotnikova EG, Boronin AM (1991) Genetic control of degradation of chlorinated benzoic acids in Arthrobacter globiformis, Corynebacterium sepedonicum and Pseudomonas cepacia strains. FEMS Microbiol Lett 65:171–176

Download references


This work was supported in part by the Global Environment Research Fund.

Author information

Correspondence to Kazuhiro Iwasaki.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Mera, N., Iwasaki, K. Use of plate-wash samples to monitor the fates of culturable bacteria in mercury- and trichloroethylene-contaminated soils. Appl Microbiol Biotechnol 77, 437–445 (2007). https://doi.org/10.1007/s00253-007-1152-0

Download citation


  • Culturable bacteria
  • Plate wash
  • Mercury
  • Trichloroethylene
  • Phylogenetic analysis