The taxonomy of yeasts is continually evolving, and currently yeasts that are of medical importance belong to two classes: the Saccharomycetes (previously Hemiascomycetes or Endomycetes), which contains Candida species, and the Tremellomycetes (previously Heterobasidiomycetes), which contains the basidiomycetous fungi Trichosporon and Cryptococcus. A number of species have been merged into Candida albicans, including Candida claussenii and Candida langeronii. There is evidence that C. albicans var. africana may be a new cause of vaginitis that is often misidentified as C. albicans on the Vitek 2 ID-YST system. Candida species are ubiquitous yeasts, being found on many plants and as the normal biota of the alimentary tract of mammals and mucocutaneous membranes of humans. In nearly 45% of AIDS patients, cryptococcosis was reported as the first AIDS-defining illness. Because none of the presenting signs or symptoms of cryptococcal meningitis (such as headache, fever, and malaise) are sufficiently characteristic to distinguish it from other infections that occur in patients with AIDS, determining cryptococcal antigen titers and culturing blood and cerebrospinal fluid (CSF) are useful in making a diagnosis. Not surprisingly, little effort has been undertaken to develop serologic tests for detection of invasive infections caused by yeasts other than Candida and Cryptococcus species. In order to develop a universal genetic identification system for yeasts, the most reproducible, specific, and sensitive sequence for comparison must be identified, and the majority of research has focused on the rDNA regions.

Candida, Cryptococcus, and other yeasts of medical importance.

Candida glabrata has emerged as a common cause of fungal infection. This yeast has intrinsically low susceptibility to azole antifungals such as fluconazole, and mutation to frank azole resistance during treatment has been documented. Potential resistance mechanisms include changes in expression or sequence of ERG11 encoding the azole target. Alternatively, resistance could result from upregulated expression of multidrug transporter genes; in C. glabrata these include CDR1 and PDH1. By RNA hybridization, 10 of 12 azole-resistant clinical isolates showed 6- to 15-fold upregulation of CDR1 compared to susceptible strains. In 4 of these 10 isolates PDH1 was similarly upregulated, and in the remainder it was upregulated three- to fivefold, while ERG11 expression was minimally changed. Laboratory mutants were selected on fluconazole-containing medium with glycerol as carbon source (to eliminate mitochondrial mutants). Similar to the clinical isolates, six of seven laboratory mutants showed unchanged ERG11 expression but coordinate CDR1-PDH1 upregulation ranging from 2- to 20-fold. Effects of antifungal treatment on gene expression in susceptible C. glabrata strains were also studied: azole exposure induced CDR1-PDH1 expression 4- to 12-fold. These findings suggest that these transporter genes are regulated by a common mechanism. In support of this, a mutation associated with laboratory resistance was identified in the C. glabrata homolog of PDR1 which encodes a regulator of multidrug transporter genes in Saccharomyces cerevisiae. The mutation falls within a putative activation domain and was associated with PDR1 autoupregulation. Additional regulatory factors remain to be identified, as indicated by the lack of PDR1 mutation in a clinical isolate with coordinately upregulated CDR1-PDH1.

Azole resistance in Candida glabrata : coordinate upregulation of multidrug transporters and evidence for a Pdr1-like transcription factor

The focus of this review is the evolution of biochemical phenotypic yeast identification methods with emphasis on conventional approaches, rapid screening tests, chromogenic agars, comprehensive commercial methods, and the eventual migration to genotypic methods. As systemic yeast infections can be devastating and resistance is common in certain species, accurate identification to the species level is paramount for successful therapy and appropriate patient care.

Yeast identification--past, present, and future methods.

The objective of the present study was one step extracellular biosynthesis of silver nanoparticles (AgNPs) using supernatant of Candida glabrata isolated from oropharyngeal mucosa of human immunodeficiency virus (HIV) patients and evaluation of their antibacterial and antifungal potential against human pathogenic bacteria and fungi. The mycosynthesized AgNPs were characterized by color visualization, ultraviolet-visible (UV) spectroscopy, fourier transform infrared spectroscopy (FTIR), and transmission electron microscopy (TEM). The FTIR spectra revealed the binding and stabilization of nanoparticles with protein. The TEM analysis showed that nanoparticles were well dispersed and predominantly spherical in shape within the size range of 2⁻15 nm. The antibacterial and antifungal potential of AgNPs were characterized by determining minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC)/ minimum fungicidal concentration (MFC), and well diffusion methods. The MBC and MFC were found in the range of 62.5⁻250 μg/mL and 125⁻500 μg/mL, which revealed that bacterial strains were more susceptible to AgNPs than fungal strains. These differences in bactericidal and fungicidal concentrations of the AgNPs were due to the differences in the cell structure and organization of bacteria and yeast cells. The interaction of AgNPs with C. albicans analyzed by TEM showed the penetration of nanoparticles inside the Candida cells, which led the formation of "pits" and "pores" that result from the rupturing of the cell wall and membrane. Further, TEM analysis showed that Candida cells treated with AgNPs were highly deformed and the cells had shrunken to a greater extent because of their interaction with the fungal cell wall and membrane, which disrupted the structure of the cell membrane and inhibited the normal budding process due to the destruction and loss of membrane integrity and formation of pores that may led to the cell death.

Biosynthesis of Silver Nanoparticles from Oropharyngeal Candida glabrata Isolates and Their Antimicrobial Activity against Clinical Strains of Bacteria and Fungi

Rapid (30-s) trehalase tests done with material from colonies of 482 yeasts suspended in a drop of trehalose solution on a commercially supplied glucose test strip were positive for 225 (99.1%) of 227 Candida glabrata isolates grown on either of two differential media, Candida ID medium or CandiSelect medium. The test was positive for only 3 (1.2%) and 12 (4.7%) of 255 isolates of other medically important yeast species grown on the same two media, respectively. A rapid maltase test done with a subset of 255 yeast isolates was negative for all but 1 of 64 trehalase-positive C. glabrata isolates, raising the specificity of the rapid testing for C. glabrata to 98.4 to 100%, depending on the isolation medium used. Rapid trehalase and maltase tests done independently in two laboratories with 217 yeast isolates showed sensitivities of 96.0 to 98.0% and specificities of 98.2 to 99.4% for identification of C. glabrata from colonies grown on Candida ID medium. The specificity was much lower because of frequent false-positive trehalose test results when the source of colonies was Sabouraud agar formulated with 4% glucose. We conclude that direct recognition of C. albicans as blue colonies on Candida ID isolation medium coupled with the performance of the 30-s trehalase and maltase tests for C. glabrata among the white colonies on this medium will allow the rapid presumptive identification of the two yeast species most commonly encountered in clinical samples.

Identification of Candida glabrata by a 30-Second Trehalase Test

Aims: To evaluate the rapid identification of Candida glabrata using a one minute trehalase and maltase test in four clinical laboratories.

Method: The test was evaluated with 944 freshly isolated yeasts comprising 572 C glabrata and 372 non-C glabrata strains. These strains were isolated on one of three differential media—Candida ID, CHROMagar Candida, or Albicans ID2 medium—and all strains were fully identified using standard methods.

Results: The trehalase and maltase test allowed the overall identification of 550 of 572 C glabrata strains (sensitivity, 96.2%) and only 11 of 372 isolates of other yeast species yielded a false positive result (specificity, 96.8 %). Sensitivity and specificity were consistent from one laboratory to another. Using Candida ID medium, the rapid trehalase and maltase test showed a sensitivity of 95% and specificity of 96.2%. Using CHROMagar Candida, sensitivity and specificity were 95.6% and 98.1%, respectively. Using Albicans ID2 medium (tested by two laboratories), the sensitivity was 100% and 98.5% and specificity was 98.1% and 98.2%. In 60% of cases, the test could be performed directly from the primary isolation medium, thus reducing the time for identification.

Conclusion: The rapid trehalase and maltase test was highly reliable for the presumptive identification of C glabrata on primary isolation using three different chromogenic media. Direct recognition of C albicans by means of their characteristic colour on chromogenic media coupled with one minute trehalase maltase testing performed only on suspect colonies of C glabrata allowed for rapid presumptive identification of the two yeast species most commonly encountered in clinical samples.

https://jcp.bmj.com/content/jclinpath/56/9/687.full.pdf

Routine use of a one minute trehalase and maltase test for the identification of Candida glabrata in four laboratories.

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Identification of Candida glabrata by a 30-Second Trehalase Test

Routine use of a one minute trehalase and maltase test for the identification of Candida glabrata in four laboratories.