Affiliation:
1. Laboratoire des Sciences Végétales, UFR des Sciences Pharmaceutiques et Biologiques, Université Paris 5, 4 avenue de l'Observatoire, 75006 Paris, France
Abstract
ABSTRACT
In a previous work, we described the possible relationship between a defect of purine-cytosine permease and the acquisition of a cross-resistance to the antifungal combination flucytosine (5FC) and fluconazole (FLC) in
Candida lusitaniae
(T. Noël, F. François, P. Paumard, C. Chastin, D. Brethes, and J. Villard, Antimicrob. Agents Chemother. 47:1275-1284, 2003). Using degenerate PCR and chromosome walking, we cloned two
FCY2
-like genes in
C. lusitaniae
. Northern blot analysis revealed that only one gene was expressed; it was named
FCY2
. The other one behaved as a pseudogene and was named
FCY21
. In order to better characterize the possible role of
FCY2
in cross-resistance to 5FC-FLC, disruption experiments with auxotrophic strain 6936
ura3
(
D95V
)
FCY2
with an integrative vector carrying the
URA3
gene and a partial sequence of the
C. lusitaniae FCY2
gene were undertaken. Southern blot analysis revealed that homologous recombination events occurred in all transformants analyzed at rates of 50% at resident locus
FCY2
and 50% at resident locus
URA3
, resulting in the genotypes
ura3 fcy2
::
URA3
and
ura3
::
URA3 FCY2
, respectively. It was then demonstrated that only transformants harboring a disrupted
fcy2
gene were resistant to 5FC, susceptible to FLC, and resistant to the 5FC-FLC combination. Finally, complementation experiments with a functional
FCY2
gene restored 5FC and FLC susceptibilities to the wild-type levels. The results of this study provide molecular evidence that inactivation of the sole
FCY2
gene promotes cross-resistance to the antifungal association 5FC-FLC in
C. lusitaniae
.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Pharmacology (medical),Pharmacology
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