Journal of Industrial Microbiology

, Volume 16, Issue 6, pp 331–341

Tracing the interaction of bacteriophage with bacterial biofilms using fluorescent and chromogenic probes

  • M M Doolittle
  • J J Cooney
  • D E Caldwell
Article

DOI: 10.1007/BF01570111

Cite this article as:
Doolittle, M.M., Cooney, J.J. & Caldwell, D.E. Journal of Industrial Microbiology (1996) 16: 331. doi:10.1007/BF01570111

Abstract

Phages T4 and E79 were fluorescently-labeled with rhodamine isothiocyanate (RITC), fluoroscein isothiccyanate (FITC), and by the addition of 4′6-diamidino-2-phenylindole (DAPI) to phage-infected host cells ofEscherichia coli andPseudomonas aeruginosa. Comparisons of electron micrographs with scanning confocal laser microscope (SCLM) images indicated that single RITC-labeled phage particles could be visualized. Biofilms of each bacterium were infected by labeled phage. SCLM and epifluorescence microscopy were used to observe adsorption of phage to single-layer surface-attached bacteria and thicker biofilms. The spread of the recombinant T4 phage, YZA1 (containing an rll-LacZ fusion), within alac E. coli biofilm could be detected in the presence of chromogenic and fluorogenic homologs of galactose. Infected cells exhibited blue pigmentation and fluorescence from the cleavage products produced by the phage-encoded β-galactosidase activity. Fluorescent antibodies were used to detect nonlabeled progeny phage. Phage T4 infected both surface-attached and surface-associatedE. coli while phage E79 adsorbed toP. aeruginosa cells on the surface of the biofilm, but access to cells deep in biofilms was somewhat restricted. Temperature and nutrient concentration did not affect susceptibility to phage infection, but lower temperature and low nutrients extended the time-to-lysis and slowed the spread of infection within the biofilm.

Keywords

biofilms bacteriophages T4 and E79 Escherichia coli Pseudomonas aeruginosa SCLM fluorescence 

Copyright information

© Society for Industrial Microbiology 1996

Authors and Affiliations

  • M M Doolittle
    • 1
  • J J Cooney
    • 1
  • D E Caldwell
    • 2
  1. 1.Environmental Sciences ProgramUniversity of MassachusettsBostonUSA
  2. 2.Department of Applied Microbiology and Food ScienceUniversity of SaskatchewanSaskatoonCanada

Personalised recommendations