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

A simple bioluminescence procedure for early warning detection of coliform bacteria in drinking water

  • 339 Accesses

  • 6 Citations


Traditional cultivation-dependent tests for coliform bacteria in food and drinking water take 18–24 h to complete. Bioluminescence-based enzyme assays can potentially reduce analysis time for indicator bacteria such as coliforms. In the present study, we developed a simple presence/absence (P/A) bioluminescence procedure for rapid detection of coliform bacteria in groundwater-based drinking water. The bioluminescence procedure targeting β-d-galactosidase activity in coliform bacteria was based on hydrolysis of 6-O-β-galactopyranosyl-luciferin. Bacteria immobilized on membrane filters were enriched for 6–8 h in selective media containing isopropyl-β-d-thiogalactopyranoside (IPTG) to induce β-d-galactosidase activity in coliform bacteria. The equivalent of approximately 300 E. coli cells was required for bioluminescence detection of β-d-galactosidase activity. In comparison, PCR based detection of E. coli in drinking water required approximately 30 target cells. Analysis of contaminated drinking water samples showed comparable results for coliform bacteria using traditional multiple-tube fermentation, Colilert-18, and the bioluminescence procedure. Aeromonas hydrophila or indigenous groundwater bacteria did not interfere with the procedure. The bioluminescence procedure can be combined with commercial substrates such as Fluorocult or Colilert-18, and will allow the detection of one coliform in 100 ml drinking water within one working day. The results suggest the bioluminescence assays targeting β-d-galactosidase activity may be used for or for early warning screening of drinking water and/or rapid identification of contaminated drinking water wells.

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

Fig. 1
Fig. 2
Fig. 3


  1. Apte SC, Cheryl M, Davies MP, Sharon MP (1995) Rapid detection of faecal coliforms in sewage using a colorimetric assay of β-d-galactosidase. Water Res 29:1803–1806

  2. American Public Health Association (2005) Standard methods for the examination of water and wastewater. American Public Health Association, Washington, DC

  3. Bagley ST, Seidler RJ, Talbot Jr HW, Morrow JE (1978) Isolation of Klebsielleae from within living wood. Appl Environ Microbiol 36:178–185

  4. Bronstein I, Edwards B, Voyta JC (1989) 1,2-dioxetanes: novel chemiluminescent enzyme substrates. Applications to immunoassays. J Biolumin Chemilumin 4:99–111

  5. Geissler K, Manafi M, Amoros I, Alonso JL (2000) Quantitative determination of total coliforms and Escherichia coli in marine waters with chromogenic and fluorogenic media. J Appl Microbiol 88:280–285

  6. George I, Crop P, Servais P (2001) Use of beta-d-galactosidase and beta-d-glucuronidase activities for quantitative detection of total and fecal coliforms in wastewater. Can J Microbiol 47:670–675

  7. Heron SJE, Wilkinson JF, Duffus CM (1993) Enterobacteria associated with grass and silages. J Appl Bacteriol 75:13–17

  8. Landre JP, Gavriel AA, Lamb AJ (1998) False-positive coliform reaction mediated by Aeromonas in the Colilert defined substrate technology system. Lett Appl Microbiol 26:352–354

  9. Leclerc H, Mossel DAA, Edberg SC, Struijk CB (2001) Advances in the bacteriology of the coliform group: their suitability as markers of microbial water safety. Ann Rev Microbiol 55:201–234

  10. Masuda-Nishimura I, Fukuda S, Sano A, Kasai K, Tatsumi H (2000) Development of a rapid positive/absent test for coliforms using sensitive bioluminescence assay. Lett Appl Microbiol 30:130–135

  11. Rompré A, Servais P, Baudart J, de-Roubin MR, Laurent P (2002) Detection and enumeration of coliforms in drinking water: current methods and emerging approaches. J Microbiol Methods 49:31–54

  12. Sabat G, Rose P, Hickey WJ, Harkin JM (2000) Selective and sensitive method for PCR amplification of Escherichia coli 16S rRNA genes in soil. Appl Environ Microbiol 66:844–849

  13. Serra B, Dolores Moralez M, Zhang J, Julio Reviejo A, Hall EH, Pingarron JM (2005) In-a-day electrochemical detection of coliforms in drinking water using a tyrosinase composite biosensor. Anal Chem 77:8115–8121

  14. Tryland I, Samset ID, Hermansen L, Berg JD, Rydberg H (2001) Early warning of faecal contamination of water: a dual mode, automated system for high- (<1 h) and low-levels (6–11 h). Water Sci Technol 43:217–220

  15. Van Poucke SO, Nelis HJ (1995) Development of a sensitive chemiluminometric assay for the detection of beta-galactosidase in permeabilized coliform bacteria and comparison with fluorometry and colorimetry. Appl Environ Microbiol 61:4505–4509

  16. Van Poucke SO, Nelis HJ (1997) Limitations of highly sensitive enzymatic presence-absence tests for detection of waterborne coliforms and Escherichia coli. Appl Environ Microbiol 63:771–774

  17. Van Poucke SO, Nelis HJ (2000) Rapid detection of fluorescent and chemiluminescent total coliforms and Escherichia coli on membrane filters. J Microbiol Methods 42:233–244

  18. World Health Organization (2004) Guidelines for drinking-water quality. Sun Fung, Hong Kong

Download references


We appreciate the technical assistance of Kirsten Maagaard. We thank IDEXX Europe for providing equipment for the Colilert assays. We also thank Viggo Larsen (Miljøcenter Vendsyssel) for valuable help in collecting water samples. This work was supported financially by The Danish Ministry of Science, Technology and Innovation (projects BAKMAT and BIOTEKS).

Author information

Correspondence to Peter Roslev.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Bastholm, S., Wahlstrøm, L., Bjergbæk, L.A. et al. A simple bioluminescence procedure for early warning detection of coliform bacteria in drinking water. World J Microbiol Biotechnol 24, 2323–2330 (2008). https://doi.org/10.1007/s11274-008-9747-x

Download citation


  • Bioluminescence
  • β-d-Galactosidase
  • Coliform bacteria
  • Drinking water
  • Groundwater
  • Presence/absence