SciELO - Scientific Electronic Library Online

vol.34 issue1 author indexsubject indexarticles search
Home Pagealphabetic serial listing  

Services on Demand



Related links

  • On index processCited by Google
  • On index processSimilars in Google


Water SA

On-line version ISSN 1816-7950
Print version ISSN 0378-4738

Water SA vol.34 n.1 Pretoria Jan. 2008


Construction and evaluation of a gfp-tagged Salmonella Typhimurium strain for environmental applications*



LM BurkeI; VS BrözelII; SN VenterIII

IDivision of Natural Resources and the Environment, CSIR, Pretoria 0001, South Africa
IIDepartment of Biology and Microbiology, South Dakota State University, Brookings, SD 57007, USA
IIIDepartment of Microbiology and Plant Pathology, University of Pretoria, Pretoria 0002, South Africa





Salmonella enterica ser. Typhimurium was isolated from freshwater sediments and chromosomally labelled with a stable variant of the green fluorescent protein (GFP). The pUT mini-Tn5 Km transposon was used to introduce the gfp gene onto the chromosome of the
S. Typhimurium strain by tri-parental mating. Southern Blot hybridisation confirmed that the gene had integrated into the chromosome. The gfp gene was stably maintained and the labelled strain was not growth-rate impaired. The incorporation of the gfp gene did not convey any significant loss of phenotype which would affect the survival and behaviour of the tagged strains. The tagged S. Typhimurium strain was used to spike an established drinking water biofilm and was able to colonise and persist within the biofilm. The tagged strain was also successfully used to study the survival of S. Typhimurium in natural sediments under different temperatures. These tagged strains can therefore be used to study the fate and survival of different Salmonella strains in water environments.

Keywords: biofilm, green fluorescent protein, Salmonella Typhimurium, survival


"Full text available in PDF"



AMANN RIW, LUDWIG W and SCHLEIFER K-H (1995) Phylo-genetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev. 59 143-169.         [ Links ]

ANDERSON KL, WHITLOCK JE and HARWOOD VJ (2005) Persistence and differential survival of fecal indicator bacteria in subtropical waters and sediments. Appl. Environ. Microbiol. 71 3041-3048.         [ Links ]

ANDERSON JB, STERNBERG C, POULSEN LS, BJ0RN SP, GIVSKOV M and MOLIN S (1998) New unstable variants of GFP for studies of transient gene expression in bacteria. Appl. Environ. Microbiol. 64 2240-2246.         [ Links ]

BARCINA I, LEBARON P and VIVES-REGO J (1997) Survival of allochtonous bacteria in aquatic systems: a biological approach. FEMS Microbiol. Ecol. 23 1-9.         [ Links ]

BAUDART J, LEMARCHAND K, BRISABOIS A and LEBARON P (2000) Diversity of Salmonella strains isolated from the aquatic environment as determined by serotyping and amplification of the ribosomal DNA spacer regions. Appl. Environ. Microbiol. 66 1544-1552.         [ Links ]

BELIAEFF B and MARY J-Y (1993) The "most probable number" estimate and its confidence limits. Water Res. 27 799-805.         [ Links ]

CAMPER A, BURR M, ELLIS B, BUTTERFIELD P and ABERNATHY C (1999) Development and structure of drinking water biofilms and techniques for their study. J. Appl. Microbiol. Symp. Supplement 85 1S-12S.         [ Links ]

CAMPER A, LECHAVELLIER MW, BROADAWAY SC and McFETERS GA (1985) Evaluation of procedures to desorb bacteria from granular activated carbon. J. Microbiol. Methods 3 187-198.         [ Links ]

CHALFIE M, TU Y, EUSKIRCHEN G, WARD WW and PRASHER DC (1994) Green fluorescent protein as a marker for gene expression. Sci. 263 802-805.         [ Links ]

CLOETE TE and DE BRUYN EE (2001) The effect of culture media on antigenic expression in sulphate-reducing bacteria. Curr. Microbiol. 42 305-309.         [ Links ]

DE LORENZO V, HERRERO M, SANCHEZ JM and TIMMIS KN (1998) Mini-transposons in microbial ecology and environmental biotechnology. FEMS Microbiol. Ecol. 27 211-224.         [ Links ]

DE LORENZO V, HERRERO M and TIMMIS KN (1990) Mini-Tn5 transposon derivatives for insertion mutagenesis, promoter probing and chromosomal insertion of cloned DNA in Gram-negative bacteria. J. Bacteriol. 172 6568-6572.         [ Links ]

EBERL L, SCHUKZE R, AMMENDOLA A, GEISENBERGER O, ERHART R, STERNBERG C, MOLIN S and AMANN R (1997) Use of green fluorescent protein as a marker for ecological studies of activated sludge communities. FEMS Microbiol. Lett. 149 77-83.         [ Links ]

FIGURSKI DH and HELINSKI DR (1979) Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc. Natl. Acad. Sci. USA 76 1648-1652.         [ Links ]

FRANA TS and CARLSON SA (2001) Development and use of a plasmid encoding green fluorescent protein in multiple antibiotic resistant Salmonella. BioTechniques 30 28-32.         [ Links ]

GANNON VP, GRAHAM TA, READ S, ZIEBELL K, MUCKLE A, MORI J, THOMAS J, SELINGER B, TOWNSHEND I and BYRNE J (2004) Bacterial pathogens in rural water supplies in Southern Alberta. Canada. J. Toxicol. Environ. Health A 67 1643-1653.         [ Links ]

HO BSW and TAM T-Y (2000) Rapid enumeration of Salmonella in environmental waters and wastewater. Water Res. 43 2397-2399.         [ Links ]

HUNTER PR (1997) Water-Borne Diseases: Epidemiology and Ecology. John Wiley and Sons, Chichester.         [ Links ]

ISLAM M, MORGEN J, DOYLE MP, PHATAK SC, MILLNER P, JIANG X (2004) Persistence of Salmonella enterica serovar Typhimurium on lettuce and parsley and in soils on which they were grown in fields treated with contaminated manure composts or irrigation water. Foodborne Pathog. Dis 1 27-35.         [ Links ]

LIM CH and FLINT KP (1989) The effects of nutrients in the survival of Escherichia coli in lake water. J. Appl. Bacteriol. 66 559-569.         [ Links ]

MEIKLE A, GLOVER LA, KILLHAM K and PROSSER JI (1994) Potential luminescence as an indicator of activation of genetically modified Pseudomonas fluoresceins in liquid culture and in soil. Soil Biol. Biochem. 26 747-755.         [ Links ]

MOGANEDI KLM, GOYVAERTS EMA, VENTER SN and SIBARA MM (2007) Optimisation of PCR-invA primers for the detection of Salmonella in drinking and surface waters following a pre-cultivation step. Water SA 33 (2) 195-202.         [ Links ]

MÖLLER S, STERNBERG C, ANDERSEN JB, CHRISTENSEN BB, RAMOS JL, GIVSKOV M and MOLIN S (1998) In situ gene expression in mixed culture biofilms evidence of metabolic interaction between community members. Appl. Environ. Microbiol. 64 721-732.         [ Links ]

MOORE BC, MARTINEZ E, GAY JM and RICE DH (2003) Survival of Salmonella enterica in freshwater and sediments and transmission by the aquatic midge Chironomus tentans (Chironomidae: Diptera). Appl. Environ. Microbiol. 69 (8) 4556-4560.         [ Links ]

NATVIG EE, INGHAM SC, INGHAM BH, COOPERBAND LR, ROPER TR (2002) Salmonella enterica serovar Typhimurium and Escherichia coli contamination of root and leaf vegetables grown in soils with incorporated bovine manure Appl. Environ. Microbiol. 68 2737-2744        [ Links ]

O'SHEA ML and FIELD R (1992) Detection and disinfection of pathogens in storm-generated flows. Can. J. Microbiol. 38 267-276.         [ Links ]

SOBECKY PA, SCHELL MA, MORAN MA and HODSON RE (1992) Adaptation of model genetically engineered microorganisms to lake water: Growth rate enhancements and plasmid loss. Appl. Environ. Microbiol. 58 3630-3637.         [ Links ]

STERNBERG C, CHRISTENSEN BB, JOHANSEN T, NIELSEN AT, ANDERSEN JB, GIVSKOV M and MOLIN S (1999) Distribution of bacterial growth activity in flow chamber biofilms. Appl. Environ. Microbiol. 65 (9) 4108-4117.         [ Links ]

SZEWZYK U, SZEWZYK R, MANZ W and SCHLEIFER K-H (2000) Microbiological safety of drinking water. Annu. Rev. Microbiol. 54 81-127.         [ Links ]

TOMBOLINI R, UNGE A, ELLEN D, DE BRUIJN F and JANSSON JK (1997) Flow cytometric and microscopic analysis of GFP-tagged Pseudomonas fluorescens bacteria. FEMS Microbiol. Ecol. 22 17-28.         [ Links ]

VALDIVIA RH, HROMOCKYI AE, MONACK D, RAMAKRISHNAN L and FALKOW S (1996) Applications for the green fluorescent protein (GFP) in the study of host-pathogen interaction. Gene. 173 47-52.         [ Links ]

VENTER SN (2005) Origin, Fate and Clinical Relevance of Water-borne Pathogens Present in Surface Waters. WRC Report No. 1398/1/05. Water Research Commission, Pretoria, South Africa.         [ Links ]

WINFIELD MD and GROISMAN EA (2003) Role of nonhost environments in the lifestyles of Salmonella and Escherichia coli. Appl. Environ. Microbiol. 69 (7) 3687-3694.         [ Links ]



+27 12 420-4100;
Fax: +27 12 420-3266

Received 10 September 2007;
Accepted in revised form 11 December2007.



* Journal series publication 3595 from the South Dakota Agricultural Experiment Station.

Creative Commons License All the contents of this journal, except where otherwise noted, is licensed under a Creative Commons Attribution License