Candida dubliniensis is a newly described species that is closely related phylogenetically to Candida albicans and that is commonly associated with oral candidiasis in human immunodeficiency virus-positive patients. Several recent studies have attempted to elucidate phenotypic and genotypic characteristics of use in separating the two species. However, results obtained with simple phenotypic tests were too variable and tests that provided more definitive data were too complex for routine use in the clinical laboratory setting. The objective of this study was to determine if reproducible identification of C. dubliniensis could be obtained with commercial identification kits. The substrate reactivity profiles of 80 C. dubliniensis isolates were obtained by using the API 20C AUX, ID 32 C, RapID Yeast Plus, VITEK YBC, and VITEK 2 ID-YST systems. The percentages of C. dubliniensisisolates capable of assimilating or hydrolyzing each substrate were compared with the percentages from the C. albicans profiles in each kit's database, and the results were expressed as percentC. dubliniensis and percent C. albicans. Any substrate that showed >50% difference in reactivity was considered useful in differentiating the species. In addition, assimilation of methyl-α-d-glucoside (MDG), d-trehalose (TRE), and d-xylose (XYL) by the same isolates was investigated by the traditional procedure of Wickerham and Burton (L. J. Wickerham and K. A. Burton, J. Bacteriol. 56:363–371, 1948). At 48 h (the time recommended by the manufacturer for its new database), we found that the assimilation of four carbohydrates in the API 20C AUX system could be used to distinguish the species, i.e., glycerol (GLY; 88 and 14%), XYL (0 and 88%), MDG (0 and 85%), and TRE (15 and 97%). Similarly, results with the ID 32 C system at 48 h showed that XYL (0 and 98%), MDG (0 and 98%), lactate (LAT; 0 and 96%), and TRE (30 and 96%) could be used to separate the two species. Phosphatase (PHS; 9 and 76%) and α-d-glucosidase (23 and 94%) proved to be the most useful for separation of the species in the RapID Yeast Plus system. While at 24 h the profiles obtained with the VITEK YBC system showed that MDG (10 and 95%), XYL (0 and 95%), and GLY (26 and 80%) could be used to separate the two species, at 48 h only XYL (6 and 95%) could be used to separate the two species. The most useful substrates in the VITEK 2 ID-YST system were TRE (1 and 89%), MDG (1 and 99%), LAT (4 and 98%), and PHS (83 and 1%). While the latter kit was not yet commercially available at the time of the study, it would appear to be the most valuable for the identification of C. dubliniensis. Although assimilation of MDG, TRE, and XYL proved to be the most useful for species differentiation by the majority of commercial systems, the results with these carbohydrates by the Wickerham and Burton procedure were essentially the same for both species, albeit following protracted incubation. Thus, it is the rapidity of the assimilation achieved with the commercial systems that allows the differentiation of C. dubliniensis from C. albicans.

Hyphal-wall preparations of Aspergi//us fumigatus have been analysed by sequential treatment with KOH, nitrous acid and again with KOH. By acidification of the alkali-soluble extract, a polyglucose was precipitated which * Department of Plant Biology, University of Groningen, The Netherlands showed an X-ray diffraction pattern similar to that of (1 -+ 3)-a-glucan. The remainder of the alkali-soluble fraction was precipitated with ethanol; it contained all the mannose, galactose and protein of the wall and, in addition, 6 2 YO of the amino sugars. This wall-associated glycoprotein, following SDS-PAGE and immunoblotting, reacted with antisera raised against several mycelial extracts of A. fumigatus. Sera from patients with aspergilloma have antibodies which recognize components of this glycoprotein. The glycoprotein nature of these antigens was shown by their ability to bind Lens culinaris lectin. In addition, the antigedantibody binding could be disrupted by exposure of antigen to periodate oxidation, hydrolysis with dilute acid or pretreatment with a large excess of an exo-/?-D-galactofuranosidase. The alkaliinsoluble fraction consisted of a covalently linked glucan-chitin complex. Nitrous acid treatment, which specifically disrupts glycosidic linkages involving glucosamine, did not solubilize much material but changed the X-ray diffraction pattern from diffuse to a pattern showing the characteristic lines of crystalline (1 + 3)-g=glucan and chitin. Most of the glucan became alkali-soluble after this treatment, and the insoluble residue appeared to contain crystalline chitin.

A previously undescribed, immunoreactive, membranous spherule outer wall (SOW) fraction produced by Coccidioides immitis (strains 634 and 735) grown in culture was isolated. Both this fraction and intact spherules were reactive with sera from coccidioidomycosis patients, as demonstrated by immunofluorescence microscopy. The serological activity of SOW was also demonstrated by its reactivity with human anti-C. immitis tube precipitin in a standardized immunodiffusion assay. Extraction of SOW with the nonionic detergent N-octyl-P-D-glucopyranoside (OG) permitted the isolation of an OG-soluble fraction which was reactive in the immunodiffusion assay. Rabbit antisera raised against the OG-soluble fraction were used in immunofluorescence and immunoelectron-microscopic studies of the parasitic cycle to confirm that the immunoreactive components of the solubilized fraction of SOW were associated with the inner and outer layers of the spherule wall as well as with distinct cytoplasmic organelles observed in thin sections of spherules. The immunoreactivity of SOW with sera from patients suggested that infected individuals are exposed to this surface wall material isolated from in vitro-grown spherules.decanted onto sterile Whatman no. 1 filter paper and subjected to aspirator-vacuum filtration. The filtrate was centrifuged (10,000 x g, 15 min, 4°C), and the supernatant was 2686 on August 5, 2020 by guest http://iai.asm.org/ Downloaded from 479-485.

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