PURPOSE. To develop panfungal and Candida albicans species-specific polymer
ase chain reaction (PCR) assays to screen donor eyes for fungal contaminati
on before corneal excision.
METHODS. PCR primers were designed for either the broad-spectrum detection
of fungal DNA or the specific detection of C albicans DNA. Their sequences
were based on rDNA regions highly conserved among and specific to fungi and
C albicans, respectively. PCR conditions with the two primer sets were opt
imized and tested for sensitivity using purified C. albicans genomic DNA an
d a plasmid containing the relevant region of C albicans DNA. The specifici
ty of the primer sets was established using higher eukaryotic, fungal, prok
aryotic, and viral DNAs as PCR templates. Donor eye swab specimens were col
lected before corneal excision. DNA was extracted from the specimens and te
sted by both PCR assays.
RESULTS. The lower limit of detection for both primer sets was consistently
10(3) genome equivalents, when using genomic DNA as a template and 10(2) c
opies of plasmid. The fungal PCR assay amplified DNA from all fungal specie
s tested but did not amplify any of the selected mammalian, bacterial, or v
iral DNA. The C. albicans PCR detected the C albicans DNA but was negative
for all other DNA substrates, including the other fungal templates. Thirty-
five percent of the donor eye samples tested were positive for fungus, and
19% were positive for C albicans DNA.
CONCLUSIONS. The PCR assays allowed the rapid screening of DNA extracted fr
om specimens collected from corneal donors for potential fungal contaminati
on. The assay was highly sensitive and specific for screening corneal surfa
ces. The results suggest that approximately one-third of donor eyes tested
harbor fungi on the ocular surface.