An electrochemical RNA hybridization assay for detection of the fecal indicator bacterium Escherichia coli

• Published in: Marine pollution bulletin 2005-11, Vol.50 (11), p.1251-1261
• Main Authors: LaGier, Michael J., Scholin, Christopher A., Fell, Jack W., Wang, Joseph, Goodwin, Kelly D.
• Format: Article
• Language: English
• Subjects: Analysis methods; Applied sciences; Biological and medical sciences; Biological Assay - methods; Biosensor; Biosensors; Biotechnology; Earth sciences; Earth, ocean, space; Electrochemistry - methods; Engineering and environment geology. Geothermics; Environmental Monitoring - methods; Escherichia coli - genetics; Escherichia coli - metabolism; Exact sciences and technology; Fecal indicators; Fecal pollution; Feces - microbiology; Fundamental and applied biological sciences. Psychology; Guanine Nucleotides - metabolism; Methods. Procedures. Technologies; Microspheres; Monitoring; Natural water pollution; Nucleic Acid Hybridization; Oligonucleotide Probes - genetics; Pollution; Pollution, environment geology; RNA; RNA, Ribosomal, 16S - isolation & purification; Seawaters, estuaries; Sensitivity and Specificity; Various methods and equipments; Water Microbiology; Water quality; Water treatment and pollution
• ISSN: 0025-326X; 1879-3363
• DOI: 10.1016/j.marpolbul.2005.04.034

Monitoring waters for indicator bacteria is required to protect the public from exposure to fecal pollution. Our proof-of-concept study describes a method for detecting fecal coliforms. The coliform Escherichia coli was used as a model fecal indicator. DNA probe-coated magnetic beads in combination with the electrochemical monitoring of the oxidation state of guanine nucleotides should allow for direct detection of bacterial RNA. To demonstrate this concept, we used voltammetry in connection with pencil electrodes to detect isolated E. coli 16S rRNA. Using this approach, 10 7 cells of E. coli were detected in a quantitative, reproducible fashion in 4 h. Detection was achieved without a nucleic acid amplification step. The specificity of the assay for coliforms was demonstrated by testing against a panel of bacterial RNA. We also show that E. coli RNA can be detected directly from cell extracts. The method could be used for on-site detection and shows promise for adaptation into automated biosensors for water-quality monitoring.