Evidence for Mating of the "Asexual" Yeast Candida albicans in a Mammalian Host

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The construction of disruption plasmids for MTL, MTLα1, and ADE2 is described on ScienceOnline (25). Ura⁻Ade⁺ strains were created through counter selection on 5-fluoroorotic acid to eliminate the URA3 marker gene from the penultimate disruption strain. Ade⁻Ura⁺ strains were created by deleting both copies of ADE2 in Ura⁻ strains and allowing the cells to retain URA3 after the second round of transformation. Because C. albicans does not contain silent cassettes of MTL (15), the various MTL deletions would be expected to be sufficient to create strains analogous to a and α.

At least two female Balb-C mice for each test mating combination were injected with 2.5 × 10⁷ yeast cells into the lateral tail vain. Cells for injection were grown to saturation at 30°C and then back-diluted and grown for 4 hours to mid logarithmic phase. The cells were pelleted, resuspended in 0.9% saline, and counted by hemacytometer. Mice were euthanized after 24 hours, and the kidneys were removed, homogenized, and plated in fractions onto standard synthetic medium plus dextrose (SD)-Ade-Ura plates and incubated at 30°C. A fraction of each sample was also plated to medium composed of yeast extract, peptone, plus dextrose (YPD) for total colony counts of recovered cells.

C. M. Hull, R. M. Raisner, A. D. Johnson, unpublished data.

Data from additional test matings can be found on Science Online (25).

Supplemental information on the in vitro conditions tested is available on Science Online (25).

Supplemental data and protocol information on the FACS and DAPI analyses are available on Science Online (25).

Additional MTL and ADE2 Southern blot information is available on Science Online (25).

Supplemental information on the isolation of genomic DNA and Southern blot protocol and probes is available on ScienceOnline (25).

Chromosome information was obtained from alces.med.umn.edu/Candida.html. Specific assignments were made by the Stanford Candida albicans sequencing project. Sequence data for C. albicans were obtained from the Stanford DNA Sequencing and Technology Center Web site at www-sequence.stanford.edu/group/candida.

Supplemental data are available at www.sciencemag.org/feature/data/1049869.shl.

The authors acknowledge V. Nguyen and additional members of the J. Li laboratory (University of California, San Francisco) for assistance with the FACS analysis, A. Uhl, I. Herskowitz, D. Ganem, and R. Brazas for helpful comments, and D. Inglis and additional members of the Johnson Laboratory for continuing support and assistance. Sequencing of C. albicans by the Stanford DNA Sequencing and Technology Center was accomplished with the support of the NIDR and the Burroughs Wellcome Fund. This work was supported by NIH grant GM37049 and a Burroughs Wellcome Merit Award to A.D.J.