Rapid identification of Candida dubliniensis by indirect immunofluorescence based on differential localization of antigens on C. dubliniensis blastospores and Candida albicans germ tubes

Abstract

There is a clear need for the development of a rapid and reliable test for the identification of Candida dubliniensis and for the discrimination of this species from Candida albicans. In the present study we have investigated the potential use of C. dubliniensis-specific antigens as a basis for its identification. We produced an anti-C. dubliniensis serum which, after adsorption with C. albicans blastospores, was found to differentially label C. dubliniensis isolates in an indirect immunofluorescence test. In this test, the antiserum reacted with blastospores and germ tubes of C. dubliniensis and with blastospores of Candida krusei and Rhodotorula rubra but did not react with blastospores of several other Candida species including C. albicans. The antiserum also reacted with C. albicans germ tubes. The anti-C. dubliniensis adsorbed serum reacted with specific components of 25, 28, 37, 40, 52, and 62 kDa in the C. dubliniensis extract and with a variety of antigens from other yeast species. The antigens from non-C. dubliniensis yeasts showing reactivity with the anti-C. dubliniensis adsorbed serum are mostly expressed within the cell walls of these yeast species, and this reactivity does not interfere with the use of the anti-C. dubliniensis adsorbed serum in an indirect immunofluorescence test for the rapid identification of C. dubliniensis.

FIG. 1

Immunofluorescence photographs of C. dubliniensis NCPF 3949 blastospores (a) and germ tubes (d) stained with the anti-C. dubliniensis adsorbed serum. Immunofluorescence (b and c) and phase-contrast (e and f) photographs, respectively, of the same microscopic fields, show C. krusei NCPF 3100 blastospores (b and e) and C. albicans NCPF 3153 germ tubes (c and f) stained with the anti-C. dubliniensis adsorbed serum. Magnification, ×1,000.

FIG. 2

Western blots of 10% slab gels loaded with extracts from C. dubliniensis NCPF 3949 blastospores grown at 37°C (lanes A1 and B2) or 24°C (lane B1) and C. albicans NCPF 3153 germ tubes (lanes A2 and B4) or blastospores (lane B3) stained with the anti-C. dubliniensis serum (A) or with the anti-C. dubliniensis adsorbed serum (B). The molecular masses of standard proteins (in kilodaltons) are listed to the left of the gels. Relevant antigenic bands are indicated by arrowheads.

FIG. 3

Western blots of 10% slab gels loaded with extracts from C. krusei NCPF 3100 (lane 1), C. tropicalis NCPF 3111 (lane 2), C. parapsilosis NCPF 3104 (lane 3), C. guilliermondii NCPF 3099 (lane 4), C. glabrata NCPF 3203 (lane 5), and R. rubra UPV94921 (lane 6) stained with the anti-C. dubliniensis adsorbed serum. The molecular masses of standard proteins (in kilodaltons) are listed to the left of the gel.

FIG. 4

Western blots of 10% (wt/vol) acrylamide slab gels loaded with extracts from C. dubliniensis NCPF 3949 grown at 24°C (lane 1) and 37°C (lane 2), C. albicans NCPF 3153 grown at 24°C (lane 3) and 37°C (lane 4), C. krusei NCPF 3100 (lane 5), and R. rubra UPV94921 (lane 6) stained with the anti-C. dubliniensis cell wall surface-specific serum. The molecular masses of standard proteins (in kilodaltons) are listed to the left of the gel.