Skip to main content

16S rDNA Phylogenetic and RAPD–PCR Analyses of Petroleum Polycyclic Aromatic Hydrocarbons-Degrading Bacteria Enriched from Oil-Polluted Soils

Arabian Journal for Science and Engineering volume 41, pages 2095–2106 (2016)Cite this article

Abstract

Petroleum polycyclic aromatic hydrocarbons (PAHs) are health risks to human, as they can be toxic, mutagenic and carcinogenic. As the Kingdom of Saudi Arabia is one of the major petroleum-producing countries, they inevitably suffer from worsening this environmental problem. Therefore, removal of these compounds from the environment is a necessity for ensuring human health. In this study, 54 bacterial isolates were obtained by enrichment techniques from oil-contaminated soil samples. Out of them, seven gram-negative bacterial strains, KKU-J1, KKU-J2, KKU-J4, KKU-J7, KKU-J9, KKU-J14 and KKU-J17, exhibiting ability for petroleum PAHs degradation were selected. The isolates showing the highest growth during screening as demonstrated by the increase in their optical densities (OD600) and a concentration-dependent growth in all examined PAH compounds that grew in it, with strains KKU-J2, KKU-J7 and KKU-J17 were the best. The highest optimum growth rate of \({0.333 \pm 0.0, 0.364 \pm 0.016}\) and \({0.357 \pm 0.004 ({\rm OD}_{600})}\) was recorded for the strains KKU-J2, KKU-J7 and KKU-J17, respectively, when the level of phenanthrene was 100 mg/l. On the other hand, strain KKU-J2 was found to be the best one (\({0.413 \pm 0.0}\)) when the level of naphthalene was 100 mg/l. Molecular identification of the selected isolates was detected based on 16S rRNA gene amplification and partial sequence determination. Alignment results and the comparison of 16S rRNA gene sequences of the isolates to the 16S rRNA gene sequences available in GenBank database, as well as the phylogenetic analysis, confirmed the accurate position of the isolates as Sphingomonas paucimobilis KKU-J1, Pseudomonas alcaligenes KKU-J2, Micrococcus antarcticus KKU-J4, Arthrobacter oxydans KKU-J7, Stenotrophomonas rhizophila KKU-J9, Kocuria rhizophila KKU-J14 and Shinella zoogloeoides KKU-J17. RAPD–PCR fingerprinting was carried out for the seven isolates, and the DNA patterns revealed that there is no correlation between the RAPD profile and geographic origin sites where these isolates were collected from. This study indicates that the contaminated soil samples contain a diverse population of PAH-degrading bacteria, and the use of soil-associated microorganisms could be recommended for PAHs bioremediation in the environment.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

USD 39.95

Price includes VAT (USA)
Tax calculation will be finalised during checkout.

References

  1. Aitken M.D., Stringfellow W.T., Nagel R.D., Kazunga C., Chen S.-H.: Characteristics of phenanthrene-degrading bacteria isolated from soils contaminated with polycyclic aromatic hydrocarbons. Can. J. Microbiol. 44, 743–752 (1998)

    Article  Google Scholar 

  2. Molina-Barahona L., Rodrıguez-Vázquez R., Hernández-Velasco M., Vega-Jarquın C., Zapata-Pérez O., Mendoza-Cantú A., Albores A.: Diesel removal from contaminated soils by biostimulation and supplementation with crop residues. Appl. Soil Ecol. 27, 165–175 (2004)

    Article  Google Scholar 

  3. Mastrangelo G., Fadda E., Marzia V.: Polycyclic aromatic hydrocarbons and cancer in man. Environ. Health Perspect. 104, 1166–1170 (1996)

    Article  Google Scholar 

  4. Goldman R., Enewold L., Pellizzari E., Beach J.B., Bowman E.D., Krishnan S.S., Shields P.G.: Smoking increases carcinogenic polycyclic aromatic hydrocarbons in human lung tissue. Cancer Res. 61, 6367–6371 (2001)

    Google Scholar 

  5. Khan S., Cao Q., Lin A.-J., Zhu Y.-G.: Concentrations and bioaccessibility of polycyclic aromatic hydrocarbons in wastewater-irrigated soil using in vitro gastrointestinal test. Environ. Sci. Pollut. Res. 15, 344–353 (2008)

    Article  Google Scholar 

  6. Preuss R., Angerer J., Drexler H.: Naphthalene—an environmental and occupational toxicant. Int. Arch. Occup. Environ. Health 76, 556–576 (2003)

    Article  Google Scholar 

  7. Surridge, A.K.J.; Wehner, F.C.; Cloete, T.E.: Bioremediation of polluted soil. In: Singh, A.; Kuhad, R.C.; Ward, O.P. (eds.) Advances in Applied Bioremediation, pp. 103-121, Springer, Berlin, (2009) ISBN 978-3-540-89620-3

  8. Couling N.R., Towell M.G., Semple K.T.: Biodegradation of PAHs in soil: influence of chemical structure, concentration and multiple amendment. Environ. Pollut. 158, 3411–3420 (2010)

    Article  Google Scholar 

  9. Hesham, A.; Mawad, A.M.; Mostafa, Y.M.; Shoreit, A.: Biodegradation ability and catabolic genes of petroleum-degrading Sphingomonas koreensis strain ASU-06 isolated from Egyptian oily soil. BioMed Res. Int. 2014, 127674 (2014). doi:10.1155/2014/127674

  10. Hesham A., Mawad A.M., Mostafa Y.M., Shoreit A.: Study of enhancement and inhibition phenomena and genes relating to degradation of petroleum polycyclic aromatic hydrocarbons in isolated bacteria. Microbiology 83, 599–607 (2014)

    Article  Google Scholar 

  11. Hesham A., Wang Z., Zhang Y., Zhang J., Lv W., Yang M.: Isolation and identification of a yeast strain capable of degrading four and five ring aromatic hydrocarbons. Ann. Microbiol. 56, 109–112 (2006)

    Article  Google Scholar 

  12. Hesham A.E.L.: New safety and rapid method for extraction of genomic DNA from bacteria and yeast strains suitable for PCR amplifications. J. Pure Appl. Microbiol. 8, 383–388 (2014)

    Google Scholar 

  13. Lane D.J.: 16S/23S rRNA sequencing. In: Stackebrandt, E., Goodfellow, M. (eds.) Nucleic acid Techniques in Bacterial systematics, pp. 115–175. Wiley, New York (1991)

    Google Scholar 

  14. Martorell, R.; Behrman, J.R.; Flores, R.; Stein, A.D.: Rationale for a follow-up study focusing on economic productivity. Food Nut. Bull. 26(2), S5–S14 (2005)

  15. Yeh, F.C.; Boyle, T.J.B.: Population genetic analysis of codominant and dominant markers and quantitative traits. Belg. J. Bot. 129, 157–163 (1997)

  16. Hunter P.R., Gaston M.A.: Numerical index of the discriminatory ability of typing systems: an application of Simpson’s index of diversity. J. Clin. Microbiol. 26, 2465–2466 (1988)

    Google Scholar 

  17. Aitken M.D., Stringfellow W.T., Nagel R.D., Kazunga C., Chen S.-H.: Characteristics of phenanthrene-degrading bacteria isolated from soils contaminated with polycyclic aromatic hydrocarbons. Can. J. Microbiol. 44, 743–752 (1998)

    Article  Google Scholar 

  18. Gordon L., Dobson A.D.W.: Fluoranthene degradation in Pseudomonas alcaligenes PA-10. Biodegradation 12, 393–400 (2001)

    Article  Google Scholar 

  19. Arulazhagan P., Vasudevan N., Yeom I.: Biodegradation of polycyclic aromatic hydrocarbon by a halotolerant bacterial consortium isolated from marine environment. Environ. Sci. Technol. 7, 639–652 (2010)

    Google Scholar 

  20. Abd-El-Haleem D., Moawad H., Zaki E.A., Zaki S.: Molecular characterization of phenol-degrading bacteria isolated from different Egyptian ecosystems. Microbiol 43, 217–224 (2002)

    Google Scholar 

  21. Franzetti L., Scarpeluni M.: Characterisation of Pseudomonas spp. isolated from foods. Ann. Microbiol. 57, 39–47 (2007)

    Article  Google Scholar 

  22. Zhang M., Liu W., Nie X., Li C., Gu J., Zhang C.: Molecular analysis of bacterial communities in biofilms of a drinking water clearwell. Microbes Environ. 27, 443–448 (2012)

    Article  Google Scholar 

  23. Panda S., Bandyopadhyay P.K., Chatterjee S.N.: Characterization of Pseudomonas aeruginosa PB112 (JN996498) isolated from infected Labeo bata (Hamilton) by 16S rRNA gene sequence analysis and fatty acid methyl ester (FAME) analysis. Afr. J. Biotechnol. 12, 400–405 (2013)

    Google Scholar 

  24. Narasimhulu K., Setty Y.P.: Isolation and characterization of naphthalene degrading bacteria from soils in Warangal. Int. J. Appl. Biol. Pharm. Technol. 2, 94–103 (2011)

    Google Scholar 

  25. John R., Essien J., Akpan S., Okpokwasili G.: Polycyclic aromatic hydrocarbon-degrading bacteria from aviation fuel spill site at Ibeno, Nigeria. Bull. Environ. Contam. Toxicol. 88, 1014–1019 (2012)

    Article  Google Scholar 

  26. Rahman K.S.M., Banat I.M., Thahira J., Thayumanavan T., Lakshmanaperumalsamy P.: Bioremediation of gasoline contaminated soil by a bacterial consortium amended with poultry litter, coir pith and rhamnolipid biosurfactant. Bioresour. Technol. 81, 25–32 (2002)

    Article  Google Scholar 

  27. Leahy J.G., Tracy K.D., Eley M.H.: Degradation of mixtures of aromatic and chloroaliphatic hydrocarbons by aromatic hydrocarbon-degrading bacteria. FEMS Microbiol. Ecol. 43, 271–276 (2003)

    Article  Google Scholar 

  28. Stapleton R.D., Bright N.G., Sayler G.S.: Catabolic and genetic diversity of degradative bacteria from fuel-hydrocarbon contaminated aquifers. Microbial. Ecol. 39, 211–221 (2000)

    Google Scholar 

  29. Ntougias S., Melidis P., Navrozidou E., Tzegkas F.: Diversity and efficiency of anthracene-degrading bacteria isolated from a denitrifying activated sludge system treating municipal wastewater. Int. Biodeterior. Biodegrad. 97, 151–158 (2015)

    Article  Google Scholar 

  30. Juhasz A.L., Stanley G., Britz M.: Metabolite repression inhibits degradation of benzo [a] pyrene and dibenz [a, h] anthracene by Stenotrophomonas maltophilia VUN 10,003. J. Ind. Microbiol. Biotechnol. 28, 88–96 (2002)

    Article  Google Scholar 

  31. Ahmed R.Z., Ahmed N., Gadd G.M.: Isolation of two Kocuria species capable of growing on various polycyclic aromatic hydrocarbons. Afr. J. Biotechnol 9, 3611–3617 (2010)

    Google Scholar 

  32. Peng H., Yin H., Deng J., Ye J.-S., Chen S.-N., He B.-Y., Zhang N.: Biodegradation of Benzo[a]pyrene by Arthrobacter oxydans B4. Pedosphere 22, 554–561 (2012)

    Article  Google Scholar 

  33. Al-Thani R.F., Abd-El-Haleem D.A., Al-Shammri M.: Isolation and characterization of polyaromatic hydrocarbons-degrading bacteria from different Qatari soils. Afr. J. Microbiol. Res 3, 761–766 (2009)

    Google Scholar 

  34. Desai A.M., Autenrieth R.L., Dimitriou-Christidis P., McDonald T.J.: Biodegradation kinetics of select polycyclic aromatic hydrocarbon (PAH) mixtures by Sphingomonas paucimobilis EPA505. Biodegradation 19, 223–233 (2008)

    Article  Google Scholar 

  35. Guo C., Dang Z., Wong Y., Tam N.F.: Biodegradation ability and dioxgenase genes of PAH-degrading Sphingomonas and Mycobacterium strains isolated from mangrove sediments. Int. Biodeterior. Biodegrad. 64, 419–426 (2010)

    Article  Google Scholar 

  36. Molnar E., Baude A., Richmond S., Patel P., Somogyi P., McIlhinney R.: Biochemical and immunocytochemical characterization of antipeptide antibodies to a cloned GluR1 glutamate receptor subunit: cellular and subcellular distribution in the rat forebrain. Neuroscience 53, 307–326 (1993)

    Article  Google Scholar 

  37. Thanos M., Schonian G., Meyer W., Schweynoch C., Graser Y., Mitchell T., Presber W., Tietz H.: Rapid identification of Candida species by DNA fingerprinting with PCR. J. Clin. Microbiol. 34, 615–621 (1996)

    Google Scholar 

  38. Cancilla M.R., Powell I.B., Hillier A.J., Davidson B.E.: Rapid genomic fingerprinting of Lactococcus lactis strains by arbitrarily primed polymerase chain reaction with 32P and fluorescent labels. Appl. Environ. Microbiol. 58, 1772–1775 (1992)

    Google Scholar 

  39. Cocconcelli P.S., Porro D., Galandini S., Senini L.: Development of RAPD protocol for typing of strains of lactic acid bacteria and enterococci. Lett. Appl. Microbiol 21, 376–379 (1995)

    Article  Google Scholar 

  40. Drake L.R., Lin S., Rayson G.D., Jackson P.J.: Chemical modification and metal binding studies of Datura innoxia. Environ. Sci. Technol 30, 110–114 (1996)

    Article  Google Scholar 

  41. Migheli Q., Briatore E., Garibaldi A.: Use of random amplified polymorphic DNA (RAPD) to identify races 1, 2, 4 and 8 of Fusarium oxysporum f. sp. dianthi in Italy. Eur. J. Plant Pathol. 104, 49–57 (1998)

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Biology Department, College of Science, King Khalid University, P. O. Box 3100, Abha, 61471, Saudi Arabia

    Abd El-Latif Hesham, Sulaiman A. Alrumman & Jawaher A. Al-Amari

  2. Genetics Department, Faculty of Agriculture, Assiut University, Assiut, Egypt

    Abd El-Latif Hesham

Authors
  1. Abd El-Latif Hesham
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sulaiman A. Alrumman
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jawaher A. Al-Amari
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Abd El-Latif Hesham.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Hesham, A.EL., Alrumman, S.A. & Al-Amari, J.A. 16S rDNA Phylogenetic and RAPD–PCR Analyses of Petroleum Polycyclic Aromatic Hydrocarbons-Degrading Bacteria Enriched from Oil-Polluted Soils. Arab J Sci Eng 41, 2095–2106 (2016). https://doi.org/10.1007/s13369-015-1843-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13369-015-1843-2

Keywords

  • Enrichment culture
  • Isolation
  • 16S rRNA gene sequencing
  • Phylogenetic analysis
  • Petroleum PAHs-degrading bacteria
  • Oil-polluted soils