Evaluation of Agar Candida ID2 (bioMerieux) a chromogenic medium for yeasts differentiation

Invasive diagnostic and therapeutic methods, widespread antibiotic therapy and rising percent of the immunocompromised patients cause incrementation of frequency of occurrence of the yeast infection. C. albicans is the most commonly isolated species of Candida from clinical samples. However, recently growth of frequency of isolation Candida non - albicans from clinical specimens have been observed. Yeast-like fungi different from C. albicans have become serious clinical problem. Conventional methods of identification of the yeast-like fungi carry away a lot time enough. Employment of chromogenic agar shortens latency on result. We decided to examine the usefulness ofAgar Candida ID2 (CAN2) (bioMérieux) in the identification of Candida species. The subjects within the study were 146 of Candida spp strains which were isolated from the clinical specimens of patients hospitalized at the University Hospital in Bydgoszcz. Germ tube test. Api 20C AUX test (bioMérieux) and Agar Candida ID2 (bioMérieux) were used. We have ascertained correspondence of identifying species amounted to 82.2% of analyzed Candida species between API 20C AUX test and kind of growth on CAN2 medium. Divergence of results received between CAN2 medium and API 20C AUX test suggests necessity of conducting of verification data with other methods. In conclusion, our study shows that Agar Candida ID2 is an effective medium for the isolation yeast-like fungi and in preliminary identification of Candida species direct from clinical materials.     

Evaluation of the Pro-Lab ID ring system for the identification of medically important yeasts

We evaluated 151 coded isolates of medically important yeast species belonging to the genera Candida, Cryptococcus, Geotrichum, Rhodoturula, Saccharomyces and Torulopsis using the newly developed rapid Pro-Lab Identification Ring, PL 960 system (PLID-Ring). All isolates were concurrently identified by the API 20C and conventional procedures comprising macro- and micromorphology, assimilation and fermentation of various carbon and nitrogen compounds. The PLID-Ring system identified isolates of Candida albicans, C. kefyr, C. krusei, C. lusitaniae, C. parapsilosis, Rhodotorula rubra, and Torulopsis glabrata with 100% accuracy in 24 h. This system identified C guilliermondii and S. cerevisiae isolates with an accuracy of 90% and 86%, respectively, while those belonging to Cr. neoformans, T. candida (= C. famata), C. rugosa and C. tropicalis were identified with 38.4%, 50%, 12.5% and 50% accuracy, respectively. Three isolates of Cr. laurentii were not identified by the PLID-Ring system. The overall accuracy of the PLID-Ring system was 81.45% (123 of 151 isolates). However, the system does not include species such as Cr. laurentii in its data base. When these three Cr. laurentii isolates were excluded from the evaluation, the accuracy of the PLID-Ring system increased from 81.45% to 83.1%.     

Identification of Candida albicans using the chromogenic medium Albicans ID

The correct identification of the microrganism is the base for epidemiological studies and treatment of infections. The aim of our study was to evaluate the efficacy of the chromogenic media Albicans ID (bioMerieux, France) in the identification of Candida albicans. A total of 190 yeasts strains were evaluated in the study. A rate of 100% of all C. albicans (80) and Candida dubliniensis (five) strains exhibited blue color. Nevertheless, the blue color was also observed with cultures of Candida rugosa (3/5) and Candida tropicalis (3/17). Albicans ID cromogenic media presented specificity rate of 90% and positive and negative predictive values of 88% and 100%, respectively, in the identification of C. albicans.     

Evaluation of Chromalbicans Agar for presumptive identification of Candida albicans

The utility of Chromalbicans Agar (Biolife Italiana, Milano, Italy) was evaluated with 723 clinical isolates and type culture collection strains from different genera including Candida, Cryptococcus, Pichia, Rhodotorula, Saccharomyces, Trichosporon y Zygosaccharomyces. Presumptive identification was confirmed by germ tube test and carbohydrate assimilation on API-ATB ID 32C (bioMerieux, France). Growth on Chromalbicans Agar was very useful for the presumptive identification of C. albicans isolates, and sensitivity and specificity values were significantly high (>97%), since a very low number of isolates were found to be false negative or false positive.     

Evaluation of a new chromogenic agar medium for isolation and identification of Group B streptococci

AIMS: To evaluate a new chromogenic agar as a screening medium for the isolation of Group B streptococci from high vaginal swabs from pregnant women. METHODS AND RESULTS: The medium was evaluated with 195 high vaginal swabs referred for antenatal screening and compared with blood agar and Granada medium. The new chromogenic medium showed 100% sensitivity for the detection of Group B streptococci, and also showed a positive predictive value of 100%. Granada medium also showed excellent sensitivity and specificity and both media were superior to blood agar. CONCLUSIONS: The new chromogenic medium showed excellent performance for the detection of Group B streptococci from clinical samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first chromogenic medium described for the detection of Group B streptococci. The medium offers an effective and convenient alternative to conventional media, currently used in clinical laboratories.     

Evaluation of SOC for the presumptive identification of Candida albicans and Cryptococcus neoformans

SOC, a fungal growth medium composed of Solryth^R, oxgall, and caffeic acid, was evaluated as a medium to provide rapid, differential identification of Candida albicans and Cryptococcus neoformans. Using a variety of common isolation media to produce the yeast inocula, the germ tube methods tested ranked in the following order of decreasing sensitivity: SOC (97%±1), serum (92%±5), rabbit coagulase plasma with EDTA in combination with tryptic soy broth (89%±5), TOC (89%±6), and rabbit coagulase plasma with EDTA (83%±4). In chlamydospore production, SOC also proved to be the most sensitive after 24 h incubation: SOC (96%±2), TOC (80%±2), and cornmeal-Tween 80 agar (14%±3). Other medically important yeasts showed normal patterns of growth within 24 h on SOC, thus assisting in their identification.Eighty strains of Cryptococcus neoformans showed characteristic brown pigmentation on SOC and TOC within 18 h, while all other species of the genus Cryptococcus and 229 Candida isolates did not show a change in pigmentation.     

A simple medium for isolation and identification of Candida albicans directly from clinical specimens

A simple and specific medium consisting of chitosan, trypticase, Tween-80 and agar is devised to isolate the organisms directly from the clinical specimens and to produce germ tubes and chlamydospores for rapid differentiation and identification of Candida albicans from other closely related Candida species. By manipulating the incubating conditions, the specific phase of the organism can be produced in liquid or on solid medium at different time intervals to study the physiology of the organism.Many methods and media have been proposed in the past for identification of Candida albicans and to differentiate this from the closely related species of Candida (5–8, 15). Taschdjian, Burchall&Kozinn (15) showed that C. albicans produces germ tube within an hour or two when it is grown in human or animal serum or serum substitutes. The specificity of this germ tube test was later confirmed by various workers by using different media (3–5). The distinctive feature that differentiates C. albicans from other species is the production of chlamydospores (14). However, in all these studies three types of media were required to isolate the organisms from clinical specimens and to produce germ tubes and chlamydospores for identification. Recently studies have shown that a single medium can be employed to produce both structural components of the organism from the primary isolation medium but the preparation of the medium is more exhaustive (1) and time consuming (13) than the medium to be described here. The present investigation was therefore undertaken to develop a simple and specific medium to isolate the organism directly from the clinical specimens and to produce various morphological phases of Candida albicans to differentiate from other closely related Candida species for clinical diagnosis and to provide a medium to study the physiology and metabolism of the organism under in vitro conditions.Supported in part by Grant CA 20917, National Cancer Institute, National Institutes of Health and ALSAC.     

New chromogenic agar medium for the identification of Candida spp

A new chromogenic agar medium (Candida diagnostic agar [CDA]) for differentiation of Candida spp. is described. This medium is based on Sabouraud dextrose agar (Oxoid CM41) and contains (per liter) 40.0 g of glucose, 10.0 g of mycological peptone, and 15.0 g of agar along with a novel chromogenic glucosaminidase substrate, ammonium 4-(2-[4-(2-acetamido-2-deoxy-beta-D-glucopyranosyloxy)-3-methoxyphenyl]-vinyl)-1-(propan-3-yl-oate)-quinolium bromide (0.32 g liter(-1)). The glucosaminidase substrate in CDA was hydrolyzed by Candida albicans and Candida dubliniensis, yielding white colonies with deep-red spots on a yellow transparent background after 24 to 48 h of incubation at 37 degrees C. Colonies of Candida tropicalis and Candida kefyr were uniformly pink, and colonies of other Candida spp., including Candida glabrata and Candida parapsilosis, were white. CDA was evaluated by using 115 test strains of Candida spp. and other clinically important yeasts and was compared with two commercially available chromogenic agars (Candida ID agar [bioMerieux] and CHROMagar Candida [CHROMagar Company Ltd.]). On all three agars, colonies of C. albicans were not distinguished from colonies of C. dubliniensis. However, for the group containing C. albicans plus C. dubliniensis, both the sensitivity and the specificity of detection when CDA was used were 100%, compared with values of 97.6 and 100%, respectively, with CHROMagar Candida and 100 and 96.8%, respectively, with Candida ID agar. In addition, for the group containing C. tropicalis plus C. kefyr, the sensitivity and specificity of detection when CDA was used were also 100%, compared with 72.7 and 98.1%, respectively, with CHROMagar Candida. Candida ID agar did not differentiate C. tropicalis and C. kefyr strains but did differentiate members of a broader group (C. tropicalis, C. kefyr, Candida lusitaniae plus Candida guilliermondii); the sensitivity and specificity of detection for members of this group were 94.7 and 93.8%, respectively. In addition to the increased sensitivity and/or specificity of Candida detection when CDA was used, differentiation of colony types on CDA (red spotted, pink, or no color) was unambiguous and did not require precise assessment of colony color.     

Evaluation of a new chromogenic agar medium for the identification of urinary tract pathogens

This study evaluated the performance of CPS ID2 (bioMérieux) compared to that of conventionally used selective agar for the identification of bacteria responsible for urinary tract infections. This medium detects bacterial enzymes using chromogenic substrates. Two hundred and nineteen samples of urine were tested in order to evaluate new CPS ID2 in comparison to blood agar and MacConkey agar.
According to our results, the CPS ID2 agar is an easy, rapid and sensitive method for the screening of colonies suspected of being Escherichia coli , reducing the number of biochemical tests needed. Also, enterococci and Proteae can be easily detected. Other micro-organisms require further identification. The greatest value of this medium is the accurate identification of polymicrobial cultures.     

In-vitro activity of 5-fluorocytosine against 1,021 Spanish clinical isolates of Candida and other medically important yeasts

The aim of this study was to determine the prevalence of primary resistance to 5-fluorocytosine (5FC) among clinical isolates of yeasts in Spain where this drug is not currently available for therapy. We have tested the in vitro activity of 5FC against 1,021 recent yeast clinical isolates, including 522 Candida albicans, 140 Candida parapsilosis, 68 Candida glabrata, 41 Candida dubliniensis, 50 Candida guilliermondii, 34 Candida tropicalis, 28 Candida krusei, 20 Candida famata, 11 Cryptococcus neoformans, 5 Cryptococcus albidus, 43 Rhodotorula spp., 24 Trichosporon spp., 5 Saccharomyces cerevisiae, 9 Pichia spp., and 21 isolates from other 11 yeast species. The MICs were determined by the ATB Fungus agar microdilution test (bioMerieux, France) and the following interpretive breakpoints were used: susceptible, > 4 microg/ml; intermediate, 8 to 16 microg/ml; resistant, > 32 microg/ml. 5FC was very active against Candida spp. and other medically important yeasts as 852 (83.4%) of the studied isolates were susceptible (MIC < 4 microg/ml). The species most susceptible to 5FC were C. dubliniensis (100%of isolates; MIC90, 0.25 microg/ml), C. famata (100% of isolates; MIC90, 0.25 microg/ml), C. guilliermondii (98%of isolates; MIC90, 0.25 microg/ml), C. glabrata (95.5% of isolates; MIC90, 0.25 microg/ml), and C. neoformans (90.9% of isolates; MIC90, 2 microg/ml). Primary resistance to 5FC was very uncommon, and a MIC > 32 microg/ml, indicator of in vitro resistance, was observed in 106 isolates (10.4%): 77 C. albicans (16.5% of isolates; MIC90, > 128 microg/ml), 9 C. parapsilosis (6.4% of isolates; MIC90, 8 microg/ml), 4 C. albidus (80% of isolates, MIC50, > 128 microg/ml), 3 C. glabrata (4.4% of isolates; MIC90, 0.25 microg/ml), 3 C. tropicalis (8.8% of isolates; MIC90, 4 microg/ml), 2 C. krusei (7.1% of isolates; MIC90, 8 microg/ml), 2 Rhodotorula spp. (4.6% of isolates, MIC90, 1 microg/ml), 8 Trichosporon spp. (33.3% of isolates; MIC90, 64 microg/ml), and 1 C. lipolytica (50% of isolates). Interestingly, most C. albicans (67 out of 77 isolates) resistant to 5FC were serotype B isolates.     

Utility of albicans ID plate for rapid identification of Candida albicans in clinical samples

Albicans ID (bioMérieux, Marcy l'Etoile, France) is a ready-to-use medium that contains a chromogenic substrate that allows rapid detection and specific identification of Candida albicans. We have evaluated its clinical performance by culturing 846 clinical specimens from pregnant women and neonates. A 99.2% sensitivity and a 100% specificity were observed in the identification of C. albicans isolates from primary culture.     

Comparative evaluation of four commercial tests for presumptive identification of Candida albicans

Four commercially available tests (Albicans ID2, Chromalbicans Agar, CHROMagar Candida, and BactiCard Candida) and the germ tube (GT) test for presumptive identification of Candida albicans were evaluated using clinical isolates of C. albicans (n=89) and of non-albicans yeasts (n=107). Sensitivities and specificities of all tests regarding the identification of C. albicans were greater than 92%, except for Chromalbicans Agar plates (88.7% after 48 h) and their specificity was 86%. Overall, the four commercial systems were easy to use and are good systems for the routine identification of C. albicans.     

A new chromogenic medium for the isolation of Listeria spp

A new medium is described for the isolation of Listeria spp. from foods and environmental samples. It is based on a modified Oxford medium in which 3,4-cyclohexenoesculetin-beta-D-glucoside replaces aesculin. Positive colonies are intensely black with the advantage that the pigment does not diffuse into the medium. The medium, when tested alongside the US Department of Agriculture (spiked samples) and Food and Drug Administration (naturally contaminated samples) isolation procedures, performed significantly better than the current formulations (34% more confirmed positives from naturally contaminated samples) with a reduction of 1 d in the assay time for most samples.     

SST broth,a new serum free germ tube induction medium for identification of Candida albicans

Three serum free media viz, sucrose solution, starch solution and SST broth have been formulated. The objective of the present study was to evaluate these three different serum free media for induction of germ tubes by Candida albicans and to compare their efficacy with the pooled human serum. Out of 50 C. albicans isolates 47 (94 %) and 49 (98 %) produced germ tubes in pooled human serum and SST broth, respectively. Germ tube production was positive in 40 (80 %) and 36 (72 %) isolates, respectively in sucrose solution and starch solution. This study reports SST broth as a new stable and less expensive germ tube induction medium, which requires less time for preparation and can be used without any safety concerns. SST broth is found to be more effective than pooled human serum for induction of germ tubes by C. albicans isolates.     

Evaluation of a new presumptive medium for group D streptococci.

A new medium designated as D streptococcus-enterococcus broth was formulated and evaluated for the enrichment and isolation of strains of serological group D streptococci. This medium was made by modifying Todd-Hewitt broth. Most-probable-number multiple-tube and membrane filter techniques were employed to estimate the numbers of enterococci in known cultures, wastewater, and other samples. Preliminary most-probable-number counts with this medium were as much as 3 logs higher than those counts obtained from four other media with which it was compared. The methodology for using this medium to estimate the numbers of group D streptococci in water is discussed.     

Evaluation of a new presumptive medium for group D streptococci.

A new medium designated as D streptococcus-enterococcus broth was formulated and evaluated for the enrichment and isolation of strains of serological group D streptococci. This medium was made by modifying Todd-Hewitt broth. Most-probable-number multiple-tube and membrane filter techniques were employed to estimate the numbers of enterococci in known cultures, wastewater, and other samples. Preliminary most-probable-number counts with this medium were as much as 3 logs higher than those counts obtained from four other media with which it was compared. The methodology for using this medium to estimate the numbers of group D streptococci in water is discussed.     

DNA probes for the rapid identification of medically important Candida species using a multianalyte profiling system

Recently, a new flow cytometric technology to detect multiple DNA target sequences in a single microtiter well plate was developed [multianalyte profiling (MAP) System, Luminex Corp., Austin, TX]. DNA probes, directed to the internal transcribed spacer 2 region of ribosomal DNA, were therefore designed to detect and differentiate PCR amplicons from six medically important Candida species using this system. Each probe was covalently linked to one of 100 available microsphere (bead) sets. Biotinylated PCR amplicons were then hybridized to the complementary probe on each bead set. Bound amplicons were detected fluorometrically using a streptavidin-linked reporter dye, R-phycoerythrin. Specific hybridization was noted for all six Candida species probes (mean sample-to-background ratio+/-standard error: Candida albicans, 58.7+/-1.2; Candida tropicalis, 53.2+/-3.8; Candida glabrata, 46.9+/-2.1; Candida parapsilosis, 59.9+/-1.6; Candida krusei, 54.7+/-3.7 vs. 0.9+/-0.03 for all heterologous Candida species DNA targets and vs. 1.0+/-0.1 for samples containing water instead of DNA; P < 0.001). The limit of test sensitivity was 0.5 pg of DNA. A sample could be processed and analyzed within 1 h post-PCR amplification. Therefore, the multianalyte profiling system was rapid, sensitive and specific for the detection and differentiation of the most medically important species of Candida.     

Evaluation of chromogenic medium (ORSAB) for routine identification of methicillin resistant Staphylococcus aureus (MRSA)

Performance of chromogenic medium (ORSAB) for routine detection of methicillin resistant S. aureus (MRSA) was evaluated on 510 specimens collected from patients suspected of MRSA infection or colonization. Addition of ORSAB plates to the routine protocol allowed MRSA identification in 24 hours from samples plating. In 18 samples MRSA colonies were identified only on ORSAB plates, those cases would have been missed by routine protocol alone.     

Ammonium assimilation by Candida albicans and other yeasts: evidence for activity of glutamate synthase

Activities and properties of the ammonium assimilation enzymes NADP+-dependent glutamate dehydrogenase (GDH), glutamate synthase (GOGAT) and glutamine synthetase (GS) were determined in batch and continuous cultures of Candida albicans. NADP+-dependent GDH activity showed allosteric kinetics, with an S0.5 for 2-oxoglutarate of 7.5 mM and an apparent Km for ammonium of 5.0 mM. GOGAT activity was affected by the buffer used for extraction and assay, but in phosphate buffer, kinetics were hyperbolic, yielding Km values for glutamine of 750 microM and for 2-oxoglutarate of 65 microM. The enzymes GOGAT and NADP+-dependent GDH were also assayed in batch cultures of Saccharomyces cerevisiae and three other pathogenic Candida spp.: Candida tropicalis, Candida pseudotropicalis and Candida parapsilosis. Evidence is presented that GS/GOGAT is a major pathway for ammonium assimilation in Candida albicans and that this pathway is also significant in other Candida species.     

Ammonium assimilation by Candida albicans and other yeasts: a 13N isotope study.

Nuclear magnetic resonance spectra of cultures of Candida albicans incubated in the presence of 15N-labelled ammonium demonstrated that glutamine and glutamate were the only initial products of ammonium assimilation. The nature of the route of assimilation in the yeasts Candida albicans, Saccharomyces cerevisiae, and Candida tropicalis was further examined by the use of the short-lived isotope 13N. [13N]ammonium was generated in the reaction 16O(p,alpha)13N, induced by proton bombardment of water in tandem accelerator. High-pressure liquid chromatography was used to separate and identify the products of assimilation, and radioactivity was detected and corrected for decay, using a computer-linked NaI scintillation detector. In the three yeasts studied, the labelled ammonium was assimilated into the acid-extractable fraction of cell suspensions within 1 min, and over 75% was converted to glutamine and glutamate. Subsequent to exhaustion of the labelled ammonium, the stoichiometry of the distribution of radiolabel was consistent with a net transfer of radiolabel from glutamine to glutamate, confirming the operation of glutamate synthase (EC 1.4.1.14) in these yeasts. Initial assimilation of label was mostly into glutamine (at a maximal rate within 10 s in C. albicans), whereas accumulation in glutamate did not occur at maximal rate until more than 70% of the labelled ammonium had been assimilated (between 30 and 60 s in C. albicans). We conclude that the glutamine synthetase-glutamate synthase pathway is the major route of ammonium assimilation in C. albicans and also in nitrogen-starved cultures of S. cerevisiae and C. tropicalis.