Jpn. J. Infect. Dis., 57, S19-S21, 2004
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Neutrophil Microbicidal Activity: Screening
Bacterial Mutants for Survival after Phagocytosis Using Quantitative
Henry Rosen*, Patrick J. Lewis and Cory M. L. Nitzel
Department of Medicine, University of Washington, Seattle,
*Corresponding author: email@example.com
SUMMARY: When a constant gene replacement sequence is introduced
into bacteria to produce mutants and the flanking chromosomal
sequences are known, it is possible to use a quantitative polymerase
chain reaction method (QPCR) to compare the concurrent survival
of the different bacterial mutants under identical conditions.
We describe Escherichia coli survival following neutrophil
phagocytosis among three mutants deleted respectively for araB,
dps or oxyR. Comparisons were made both by traditional
and QPCR methods with similar results and indicate that the survival
defect of an oxyR and oxyS mutant described previously
can be attributed to the loss of oxyR alone. Deletion of
dps, a prominent member of the regulon controlled by the
oxyR gene product does not engender a survival defect.
We suggest that QPCR analysis can readily compare the relative
survival of 10 or more mutants concurrently. QPCR analysis would
seem to be especially valuable when experimental conditions are
subject to a high degree of sample to sample variability or when
the stress producing system involves use of expensive or scarce
resources like rare patient cells, cells from children, or the
use of genetically modified animal hosts.