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Applied and Environmental Microbiology, September 1999, p. 4181-4188, Vol. 65, No. 9
School of Biological Sciences, University of
Nebraska, Lincoln, Nebraska
Received 15 September 1998/Accepted 9 June 1999
Municipal effluent is the largest reservoir of human enteric
bacteria. Its public health significance, however, depends upon the
physiological status of the wastewater bacterial community. A novel
immunofluorescence assay was developed and used to examine the
bacterial growth state during wastewater disinfection. Quantitative levels of three highly conserved cytosolic proteins (DnaK, Dps, and
Fis) were determined by using enterobacterium-specific antibody fluorochrome-coupled probes. Enterobacterial Fis homologs were abundant
in growing cells and nearly undetectable in stationary-phase cells. In
contrast, enterobacterial Dps homologs were abundant in
stationary-phase cells but virtually undetectable in growing cells. The
range of variation in the abundance of both proteins was at least
100-fold as determined by Western blotting and immunofluorescence analysis. Enterobacterial DnaK homologs were nearly invariant with
growth state, enabling their use as permeabilization controls. The
cellular growth states of individual enterobacteria in wastewater samples were determined by measurement of Fis, Dps, and DnaK abundance (protein profiling). Intermediate levels of Fis and Dps were evident and occurred in response to physiological transitions. The results indicate that chlorination failed to kill coliforms but rather elicited
nutrient starvation and a reversible nonculturable state. These studies
suggest that the current standard procedures for wastewater analysis
which rely on detection of culturable cells likely underestimate fecal
coliform content.
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Bacterial Growth State Distinguished by Single-Cell
Protein Profiling: Does Chlorination Kill Coliforms in Municipal
Effluent?
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Corresponding author. Mailing address: E234 Beadle
Center for Genetics, University of Nebraska, Lincoln, NE 68588-0666. Phone: (402) 472-2769. Fax: (402) 472-8722. E-mail:
pblum{at}biocomp.unl.edu.
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