Development of a simple and sensitive fluorimetric method for isolation of coumaphos-hydrolysing bacteria

R. L. Harcourt, Irene Horne, T. D. Sutherland, B. D. Hammock, R. J. Russell, J. G. Oakeshott

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26 Scopus citations

Abstract

Aims: To develop a simple, rapid and sensitive fluorimetric assay to detect, isolate and characterize a soil bacterium capable of degrading the organophosphorus pesticide, coumaphos. Methods and Results: A high throughput microtitre plate-based method was used to quantify coumaphos hydrolysis by the bacterium. The fluorescent hydrolysis product of coumaphos, chlorferon, was detected at levels as low as 10 nmol l-1. Incorporation of coumaphos into agar plates allowed the rapid detection of coumaphos-hydrolysing bacteria when exposed to an excitation wavelength of approximately 340 nm. The coumaphos-hydrolysing enzyme could be visualized when bacterial cell extracts were separated on SDS-PAGE, incubated with coumaphos and exposed to an excitation source as above. Conclusions: This method is 100-fold more sensitive than the currently used spectrophotometric method for coumaphos. Significance and Impact of the Study: This is a unique and versatile tool to screen for bacteria possessing phosphotriesterase activity.

Original languageEnglish (US)
Pages (from-to)263-268
Number of pages6
JournalLetters in Applied Microbiology
Volume34
Issue number4
DOIs
StatePublished - 2002

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ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Water Science and Technology
  • Fluid Flow and Transfer Processes
  • Microbiology
  • Ecology

Cite this

Harcourt, R. L., Horne, I., Sutherland, T. D., Hammock, B. D., Russell, R. J., & Oakeshott, J. G. (2002). Development of a simple and sensitive fluorimetric method for isolation of coumaphos-hydrolysing bacteria. Letters in Applied Microbiology, 34(4), 263-268. https://doi.org/10.1046/j.1472-765x.2002.01078.x