Rapid Identification Of Candida Albicans Research Articles

Rapid identification of Candida albicans and its related species Candida stellatoidea and Candida dubliniensis by a single PCR amplification using primers specific for the repetitive sequence (RPS) of Candida albicans

Candidiasis is caused by several Candida species, of which Candida stellatoidea and C. dubliniensis are phenotypically close to C. albicans. Although current molecular biology-based techniques can distinguish between C. albicans and C. dubliniensis, a convenient tool that can distinguish C. stellatoidea from C. albicans has not yet been developed. To develop a system that can simply, rapidly and specifically distinguish C. albicans from the related Candida species C. stellatoidea and C. dubliniensis. Genomic DNAs were purified from various yeast species and amplified by primers specific for the repetitive sequence (RPS) of C. albicans. The PCR products were purified and sequenced in order to test the specificity of the PCR amplification. The PCR primers only amplified several products from C. albicans, C. stellatoidea and C. dubliniensis. Sequence analysis of the products revealed that C. stellatoidea and C. dubliniensis both had RPSs including alt repeats, similar to C. albicans. After the PCR amplification, each of the three Candida species showed a unique amplification profile. Furthermore, RFLP analysis of the PCR products using EcoRI and ClaI produced species-specific restriction profiles. This PCR-based technique targeting the alt repeats in the RPS is useful as a tool for the rapid identification and distinction of C. albicans, C. stellatoidea and C. dubliniensis.

Evaluation of latex reagents for rapid identification of Candida albicans and C. krusei colonies.

A total of 322 yeast strains and yeastlike organisms belonging to the genera Candida, Cryptococcus, Geotrichum, Saccharomyces, and Trichosporon were tested with the new monoclonal antibody-based Bichro-latex albicans and Krusei color latex tests. Comparison of results with those obtained by conventional identification methods showed 100% sensitivity for both latex tests and 100% and 95% specificity for the Bichro-latex albicans and Krusei color tests, respectively. Because the test is easy to read and quick to perform, the Bichro-latex albicans test may be useful for rapid identification of Candida albicans colonies in the clinical laboratory.

Rapid identification of Candida albicans, C. glabrata, C. parapsilosis and C. krusei by species-specific PCR of large subunit ribosomal DNA.

A rapid PCR-based method for the identification of four Candida species is described. Primers to conserved sequences in the V3 region of large subunit rDNA were used to amplify DNA from C. albicans, C. glabrata, C. parapsilosis and C. krusei. The sequences were aligned and areas of non-concordance were used to design four species-specific forward primers. In a blind study of 82 yeast strains, four PCRs based on these primers correctly identified nine C. albicans, 18 C. glabrata, 13 C. parapsilosis and 18 C. krusei strains after gel electrophoresis of amplified DNA. Furthermore, of 17 other Candida strains (10 species) and seven strains from other yeast genera (Saccharomyces, Cryptococcus and Trichosporon) only one false positive amplification product (with C. dubliniensis) was seen. These PCRs offer a rapid alternative to conventional techniques for the identification of C. albicans, C. glabrata, C. parapsilosis and C. krusei.

Rapid identification of Candida albicans by using Albicans ID and fluoroplate agar plates.

Two commercially available agar media, Albicans ID and Fluoroplate, that use a chromogenic or a fluorogenic substrate for the detection and identification of Candida albicans were evaluated. From 1,006 clinical samples containing 723 yeast strains, 352 C. albicans strains were detected with either of the two media. The sensitivity of each of the two media was 93.8% and the specificity was 98.6%, with five false-positive reactions for Candida tropicalis and no false-negative reactions.

Rapid identification of Candida albicans by fluoroplate candida agar

150 clinical isolates of Candida species, Trichosporon species and Pichia anomala as well as 32 yeasts isolated from dairy products were tested in order to evaluate the utilization of a specific fluorogenic substrate in direct detection of Candida albicans. Detection of N- acetyl-β- d-galactosaminidase (NAGase) was performed with fluoroplate candida agar (Merck, Darmstadt, FRG), which contained 4- methylumbelliferyl-N- aceryl-β — d-galactosaminide (4-MUAG). The API 20C yeast identification system was used as a reference identification method for all isolates. The results of our investigations showed that 99% of C. albicans and all strains of C. stellatoidea examined were NAGase-positive. There were two false positive reactions of four C. tropicalis isolates and one false positive reaction of three Trichosporon beigelii strains.

Rapid Identification ofCandida Albicansby Dot-Enzyme Immunoassay

A highly sensitive and specific dot-enzyme immunoassay for the rapid identification of Candida albicans was developed using a murine monoclonal antibody (Mab), which adsorbed to cell surface-exposed determinants. This Mab reacted with 28 of the 28 C. albicans strains tested including the serotypes A and B and 2 C. stellatoidea. It did not react with 32 other isolates representing eight other Candida species commonly encountered in human materials. All the test could be performed in four steps in less than an hour. The yeasts were directly spotted on a strip of immunodyne membrane. Then the strip was incubated for 5 min. with the Mab, for 15 min. with a peroxidase-conjugate and for 30 min. with the enzyme substrate and 4-chloro 1 naphthol. This test proved useful for rapid and easy identification of C. albicans.

Rapid identification of Candida albicans using 4-methylumbelliferyl N-acetyl-β-galactosaminide

Rapid identification of Candida albicans is performed mainly by the germ-tube test. However, recent reports have suggested that up to 5% of C. albicans species can give false negative results. We describe the use of 4-Methylumbelliferyl N-acetyl-β-D-galactosaminide (4-MAG) conjugate as an alternative to the germ-tube test. Our results indicate that, in comparison to the germ-tube test, the 4-MAG test has a sensitivity of 100% and a specificity of 92%. Candida tropicalis can give false-positive results, and that a further screening test is required to identify this species. Problems reading end-points were not encountered.

A simple laboratory method for the rapid identification of Candida albicans

A simple, relatively inexpensive method for distinguishing Candida albicans from other species of the genus Candida was developed. The organisms were first spread on a solid 2% agar medium which contained 1% bactopeptone, then incubated at 37d`C for three hours and finally examined microscopically. A coverslip placed over the organisms at the time of inoculation increased from 93.5 to 100% the proportion of isolates of C. albicans which formed parallel-sided germ tubes when tested in this way. Five other species of Candida (parapsilosis, krusei, tropicalis. pseudotropicalis and guilliermondii) were also tested but did not form germ tubes. Organisms of C. tropicalis sometimes formed elongated buds but these could be readily distinguished on direct microscopic examination from germ tubes by the constriction at their base.

Further studies of the germ-tube test for Candida albicans identification.

An evaluation of the optimal incubation time for the germ-tube test used in the rapid identification of Candida albicans revealed that a 2-hr. incubation will identify 78% of the strains, whereas a 3-hr. incubation will identify 94% of the strains tested. Extending the incubation to 4 hr. increases the strains identified by only 1%. A study of the comparative efficiency of human and sheep sera in producing germ tubes revealed that both were equally effective. However, the cost of commercially obtained sheep serum is about one third that of human serum; thus, it is more economical to use sheep serum.

Production of chlamydospores by Candida albicans cultivated on dilute oxgall agar.

A simple easily prepared clear medium consisting of 2% oxgall in 1.8% agar is described. This has proved to be excellent for the rapid identification ofCandida albicans in swabs and cultures, provided that the inoculated area is covered by a coverslip and the incubation temperature period is maintained at 28°C for 24 to 48 hours. No special technical skill is required to prepare this medium. The results presented show thatC. albicans will always produce chlamydospores on this medium if the recommended procedure is followed.

Simple rapid identification of Candida albicans with emphasis on the differentiation between Candida albicans and Candida stellatoidea.

Subcultures ofC. albicans, made from Sabouraud agar, grown at room temperature for 48 hours, were inoculated into a 10 times saline dilution of Sabouraud liquid medium and left in the incubator for 45–60 minutes at 37° C, transferred to corn meal agar plates and incubated at 37° C for 18–24 hours. Small portions of the surface agar containing the yeasts from these plates were pressed under cover glasses and examined under the oil immersion lens. Under these conditions,C. albicans cultures were observed to produce only yeast-like cells, whereasC. stellatoidea cultures contained predominantly abundant, long, thin mycelia.

Rapid identification of Candida albicans by filamentation on serum and serum substitutes.

It was noted by Hill and Gebhardt (1956), as well as by Young (1958), that Candida albicans will begin to form mycelia within 60 minutes after subcutaneous injection into mice. Hill and Gebhardt demonstrated that this phenomenon occurs only with C. albicans and C. stellatoidea, while other Candida species fail to form mycelia and are rapidly destroyed in the tissues. Johnson (1954) observed that C. albicans forms mycelia on human serum but does not mention the time interval of beginning filamentation. We noted that C. albicans spores produce short germ-tubes within 90 minutes on human serum at 37 to 42 C. This occurred on pooled and individual, hemolyzed and nonhemolyzed, and inactivated sera and on fresh as well as on deep-frozen stored material. Of other Candida species, only C. stellatoidea, whose species status is doubtful, produced these characteristic short germtubes. C. tropicalis showed long mycelia, carried over with the inoculum