Extreme resistancewas confirmed
in 38 strains of Z. bailii to several weak-acid preservatives. Using the brewing yeast Saccharomyces cerevisiae as a
control, tests showed that Z. bailii was ~3-fold more resistant to a variety of weak-acids but was not more resistant
to alcohols, aldehydes, esters, ethers, ketones, or hydrophilic chelating acids. Theweak acids were chemically
very diverse in structure, making it improbable that the universal resistance was caused by degradation or
metabolism. Examination of Z. bailii cell populations showed that extreme resistance to sorbic acid, benzoic
acid and acetic acid was limited to a few cells within the population, numbers decreasing with concentration
ofweak acid to b1 in 1000. Re-inoculation of resistant sub-populations into weak-acid-containingmedia showed
that all cells now possessed extreme resistance. Resistant sub-populations grown in any weak-acid preservative
also showed ~100% cross-resistance to other weak-acid preservatives. Tests using 14C-acetic acid showed that
weak-acid accumulation was much lower in the resistant sub-populations. Acid accumulation is caused by
acid dissociation in the higher pH of the cytoplasm. Tests on intracellular pH (pHi) in the resistant
sub-population showed that the pHwasmuch lower, ~ pH 5.6, than in the sensitive bulk population. The hypothesis
is proposed that extreme resistance toweak-acid preservatives in Z. bailii is due to population heterogeneity,
with a small proportion of cells having a lower intracellular pH. This reduces the level of accumulation of any
weak acid in the cytoplasm, thus conferring resistance to all weak acids, but not to other inhibitors.