http://www.ncbi.nlm.nih.gov/pubmed/16923951?ordinalpos=1&itool=EntrezSystem2.PEn
trez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_Discovery_RA

   Candida albicans biofilms produce antifungal-tolerant persister
cells.
   LaFleur MD, Kumamoto CA, Lewis K.

   Department of Biology, Northeastern University, 360 Huntington
Ave., 134 Mugar Hall, Boston, MA 02115, USA.

   Fungal pathogens form biofilms that are highly recalcitrant to
antimicrobial therapy. The expression of multidrug resistance pumps in
young biofilms has been linked to increased resistance to azoles, but
this mechanism does not seem to underlie the resistance of mature
biofilms that is a model of in vivo infection. The mechanism of drug
resistance of mature biofilms remains largely unknown. We report that
biofilms formed by the major human pathogen Candida albicans exhibited
a strikingly biphasic killing pattern in response to two microbicidal
agents, amphotericin B, a polyene antifungal, and chlorhexidine, an
antiseptic, indicating that a subpopulation of highly tolerant cells,
termed persisters, existed. The extent of killing with a combination
of amphotericin B and chlorhexidine was similar to that observed with
individually added antimicrobials. Thus, surviving persisters form a
multidrug-tolerant subpopulation. Interestingly, surviving C. albicans
persisters were detected only in biofilms and not in exponentially
growing or stationary-phase planktonic populations. Reinoculation of
cells that survived killing of the biofilm by amphotericin B produced
a new biofilm with a new subpopulation of persisters. This suggests
that C. albicans persisters are not mutants but phenotypic variants of
the wild type. Using a stain for dead cells, rare dark cells were
visible in a biofilm after amphotericin B treatment, and a bright and
a dim population were physically sorted from this biofilm. Only the
dim cells produced colonies, showing that this method allows the
isolation of yeast persisters. Given that persisters formed only in
biofilms, mutants defective in biofilm formation were examined for
tolerance of amphotericin B. All of the known mutants affected in
biofilm formation were able to produce normal levels of persisters.
This finding indicates that attachment rather than formation of a
complex biofilm architecture initiates persister formation. Bacteria
produce multidrug-tolerant persister cells in both planktonic and
biofilm populations, and it appears that yeasts and bacteria have
evolved analogous strategies that assign the function of survival to a
small part of the population. In bacteria, persisters are dormant
cells. It remains to be seen whether attachment initiates dormancy
that leads to the formation of fungal persisters. This study suggests
that persisters may be largely responsible for the multidrug tolerance
of fungal biofilms.