Culture -independent Pathogenic Bacterial Communities in Bottled Mineral Water

Hamdy A. Hassan


Bottled mineral water (BMW) is an alternative to mains water and consider it to be better and safer. Access to safe BMW from the bacteria involving potential health hazard is essential to health. Cultivation-independent technique PCR-based single-strand conformation polymorphism (SSCP) for genetic profiling of PCR-amplified 16S rRNA genes was performed using Com primer set targeting the 16S rRNA genes for detection of pathogenic bacteria in bottled mineral water from the final product of six factories for bottled mineral drinking water in Wadi El-natron region- Egypt. These factories use often ozone technology to treat large quantities of water because of its effectiveness in purifying and conditioning water. A total of 27 single products were isolated from the profiles by PCR re-amplification and cloning. Sequence analysis of 27 SSCP bands revealed that the 16S rRNA sequences were clustered into seven operational taxonomic units (OTUs) and the compositions of the communities of the six samples were all common. The results showed that most communities from phyla Alphaproteobacteria and certainly in the Sphingomonas sp. Culture-independent approaches produced complementary information, thus generating a more accurate view for the bacterial community in the BMW, particularly in the disinfection step, as it constitutes the final barrier before BMW distribution to the consumer


Water Resources, Bottled mineral water, Pathogenic bacteria, Culture- independent, PCR-SSCP, 16S rRNA genes

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Amann, R.I., W. Ludwig, and K.H. Schleifer. 1995. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev. 59: 143-169.

Anon. (2000) Bacterial nomenclature up-to-date.

Bassam, B. J., G. Caetano-Anolles, and P. M. Gresshoff. 1991. Fast and sensitive silver staining of DNA in polyacrylamide gels. Anal. Biochem 196: 80–83.

Blumenroth, P., and I. Wagner-Dobler. 1998. Survival of inoculants in polluted sediments: effect of strain origin and carbon source competition. Microb. Ecol. 35:279–288

Cavicchioli, R., F. Fegatella, M. Ostrowski, M. Eguchi and J. Gottschal. 1999. Sphingomonas from marine environments. Journal of Industrial Microbiology and Biotechnology 23: 268-272.

Ferreira, A.C., P.V. Morais and M. Da Costa. 1994. Alterations in total bacteria, iodonitrophenyltetrazolium (1NT)-positive bacteria, and heterotrophic plate counts of bottled mineral water. Canadian Journal of Microbiology 40: 72-77.

Ferreira, A.C., P.V. Morais, C. Gomes, and M.S. da Costa. 1996. Computer-aided comparison of protein electrophoretic patterns for grouping and identi®cation of heterotrophic bacteria from mineral water. Journal of Applied Bacteriology 80: 479-486.

Gusman V., Z. Jelesic, M. Mihajlovic-Ukropina, D. Medic, G. Pavlovic, and B. Radosavljevic. 2010. Isolation of Sphingomonas paucimobilis from drinking water using novel automated system. HealthMED. 4:1068-1071.

Gauthier, V., S. Redercher, and J. C. Block. 1999. Chlorine inactivation of Sphingomonas cells attached to goethite particles in drinking water. . Appl Environ Microbiol 65: 355-357.

Hannun, Y. and R.M. Bell. 1989. Function of sphingolipids and sphingolipid breakdown products in cellular regulation. Science 243: 500-507.

Hayashi, K. 1991. PCR-SSCP: a simple and sensitive method for detection of mutations in the genomic DNA. PCR Methods Appl. 1:34-38.

Jean-Luc J. and B. Georges. 2008. Culture-Independent Methods for

Identifying Microbial Communities in Cheese. Food Microbiology 25: 839-848

Keskes S, F. Hmaied, H. Gannoun, H . Bouallagui, J. J. Godon and M. Hamdi. 2012. Performance of a submerged membrane bioreactor for the aerobic treatment of abattoir wastewater. Bioresour Technol. 103:28-34.

Kalmbach, S., W. Manz, and U. Szewzyk. 1997. Isolation of new bacterial species from drinking water biofilms and proof of their in situ dominance with highly specific 16S rRNA probes. . Appl Environ Microbiol 63: 4164-4170.

Kawahara, K., H. Kuraishi, and U. ZaÈhringer. 1999 Chemical structure and function of glycosphingolipids of Sphingomonas spp. and their distribution among members of the alfa-4 subclass of Proteobacteria. Journal of Industrial Microbiology and Biotechnology 23:408-413.

Krziwon, C., U. ZaÈhringer, K. Kawahara, B. Weidemann, S. Kusumoto, E. Rietschel, H. D. Flad, and A.J. Ulmer. 1995. Glycosphingolipids from Sphingomonas paucimobilis induce monokine production in human mononuclear cells. Infection and Immunity 63: 2899-2905.

Laskin, A.I. and D.C. White. 1999. Special issue on the genus Sphingomonas. Journal of Industrial Microbiology and Biotechnology 23: 231±408

Lee, D.H., Y.G. Zu, and S.J. Kim. 1996. Nonradioactive method to study genetic profiles of natural bacterial communities by PCR-single strand conformation polymorphism. Appl. Environ. Microbiol. 62:3112-3120.

Männistö, M., Tiirola, M., Salkinoja-Salonen, M., Kulomaa, M.and Puhakka, J. (1999) Diversity of chlorophenol-degrading bacteria isolated from contaminated boreal groundwater. Archives of Microbiology 171, 189±197.

Meng-zhi WANG, Hong-rong WANG, Heng-chun CAO, Guo-xiang LI and Jie ZHANG. (2008). Effects of Limiting Amino Acids on Rumen Fermentation and Microbial Community. In vitro. Agricultural Sciences in China 7: 1524-1531.

Miyazaki, Y., S. Oka, S. Yamaguchi, S. Mizuno, and I. Yano. 1995. Stimulation of phagosytosis and phagosome-lysosome fusion by sphingolipids from Sphingomonas paucimobilis. Journal of Biochemistry 118: 271-277.

Neef, A., R. Witzenberg, and P. KaÈmpfer. 1999. Detection of sphingomonads and in situ identification in activated sludge using 16S rRNA-targeted oligonucleotide probes. Journal of Industrial Microbiology and Biotechnology 23: 261-267.

Oie, S., M. Oomaki, K. Yorioka, T. Tatsumi, M. Amasaki, T. Fukuda, H. Hakuno, K. Nagano, M. Matsuda, N. Hirata, N. Miyano, and A. Kamiya. 1998. Microbial contamination of `sterile water' used in Japanese hospitals. Journal of Hospital Infection 38: 61-65.

Orita, M., H. Iwahana, H. Kanazawa, K. Hayashi, and T. Sekyia. 1989. Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms. Proc. Nat. Acad. Sci. USA 86:2766-2770.

Papapetropoulou M., A. Tsintzou and A. Vantarakis. 1997. Environmental mycolacteria in botled table waters in Greece. Can. J. Microbiology 43: 499 - 502.

Peters, S., S. Koschinsky, F. Schwieger, and C.C. Tebbe. 2000. Succession of microbial communities during hot composting asdetected by PCR-single-strand-conformation polymorphism-based genetic profiles of small-subunit rRNA genes. Appl Environ Microbiol. 66:930-936.

Sambrook, J., E. F. Fritsch, and T. Maniatis. 1989. Molecular cloning: a laboratory manual. Second edition Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, USA.

Schmalenberger, A., F. Schwieger, and C.C. Tebbe. 2000. Effect of primers hybridizing to different evolutionarily conserved regions of the small-subunit rRNA gene in PCR-based microbial community analyses and genetic profiling. Appl Environ Microbiol. 67:3557-3563.

Schmalenberger, A. and C.C. Tebbe. 2003. Bacterial diversity in maize rhizospheres: conclusions on the use of genetic profiles based on PCR-amplified partial small subunit rRNA genes in ecological studies. Mol Ecol. 12:251-262.

Schwieger, F., and C. C. Tebbe. 1998. A new approach to utilize PCR-single-strand-conformation polymorphism for 16S rRNA gene-based microbial community analysis. Appl. Environ. Microbiol. 64:4870-4876.

Schwieger, F. and C.C. Tebbe. 2000. Effect of field of inoculation with Sinorhizobium meliloti L33 on the composition of bacterial communities in rhizospheres of a target plant (Medicago sativa) and a non-target plant (Chenopodium album)-linking of 16S rRNA gene-based single-strand conformation polymorphism community profiles to the diversity of cultivated bacteria. Appl Environ Microbiol. 66:3556-3565.

Stach, J . E. M . and R.G. Burns. 2002. Enrichment versus biofilm culture: a functional and phylogenetic comparison of polycyclic aromatic hydrocarbon- degrading microbial communities. Environ Microbiol. 4:169-182.

The Council of the European Union. 1998. Council directive on the quality of water intended for human consumption. Council Directive 98/83/EC 3 November, 1998. The Official Journal of the European Communities L 330, 32-34.

Tomoyuki H., H. Shin, U. Yoshiyuki, I. Masaharu and I. Yasuo. 2006. Direct comparison of single-strand conformation polymorphism (SSCP) and denaturing gradient gel electrophoresis (DGGE) to characterize a microbial community on the basis of 16S rRNA gene fragments. Journal of Microbiological Methods 66: 165-169.

Warburton, D. W. 1993. A review of the microbiological quality of bottled water sold in Canada. Part 2. The need for more stringent standards and regulations. Can. J. Microbiology 39: 158–168.

Wiese, A., Reiners, J., Brandenburg, K., Kawahara, K., ZaÈhringer, U. and Seydel, U. (1996) Planar asymmetric lipid bilayers of glycosphingolipid or lipopolysaccharide on one side and phospholipids on the other: Membrane potential, porin function, and complement activation. Biophysical Journal 70: 321-329



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Renewable Energy and Sustainable Development

E-ISSN: 2356-8569

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Arab Academy for Science, Technology and Maritime Transport (AASTMT)

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