Evaluation of Spectrophotometric Broth Microdilution Method to Determine the Fluconazole MIC of the Candida Species / 대한진단검사의학회지

BACKGROUND: Although the broth microdilution method has been recently established for antifungal susceptibility testing of the Candida species, there is still an argue in the interpretation of the trailing endpoint. We evaluated the spectrophotometric broth microdilution method (SBM) to determine the fluconazole MICs from five different Candida species. METHODS: A total of 252 clinical isolates of five Candida species (144 C. albicans, 42 C. tropicalis, 32 C. glabrata, 28 C. parapsilosis, and 6 C. krusei) were tested for fluconazole susceptibility with the broth microdilution method. The MICs were spectrophotometrically determined at 80% (Spec-80%) and 50% (Spec-50%) decrease in absorbance as compared with growth control, respectively. The results were compared with the fluconazole MICs tested by the National Committee for the Clinical Laboratory Standards (NCCLS) macrodilution method. RESULTS: When MICs were obtained by Spec-80%, the agreements of SBM and the NCCLS macro dilution method within two doubling dilutions were 92.4% (220/238) at 24 h and 78.6% (198/252) at 48 h for all Candida species. Using the Spec-50%, those were increased to 97.9% (233/238) at 24 h and 98.8% (249/252) at 48 h (P<0.01). Especially, for C. albicans and C. tropicalis, the agreement of the Spec-50% was significantly higher than those of the Spec-80% at 48 h; 97.9% vs. 75.0%, for C. albicans (P<0.01), and 100% vs. 57.1%, for C. tropicalis (P<0.01). CONCLUSIONS: These data suggest that the SBM using Spec-50% can provide a more precise and objective mean for fluconazole susceptibility testing, especially for C. albicans and C. tropicalis.

Evaluation of a Spectrophotometric Broth Microdilution Method for Determining Fluconazole Susceptibility of Candida albicans: Influence of RPMI and RPMI-2% Glucose on the Selection of Endpoint Criteria / 대한진단검사의학회지

BACKGROUND: Minimum inhibitory concentration (MIC) endpoint determination is the major varia-tion source for the fluconazole susceptibility test, especially for Candida albicans. In this study, we evaluated spectrophotometric broth microdilution methods using RPMI 1640 and RPMI supple-mented with 18 g of glucose per liter (RPMI-2% glucose) for determining fluconazole susceptibility of C. albicans. METHODS: A total of 129 clinical isolates of C. albicans were tested by the broth microdilution method using RPMI and RPMI-2% glucose. The MIC endpoint was calculated objectively with the spectrophotometer set at 405 nm. These results were compared to those by the National Commit-tee for Clinical Laboratory Standards (NCCLS) macrodilution method and the agar dilution method. RESULTS: The mean absorbances in the drug-free wells in RPMI and RPMI-2% glucose were 0.208 +/- 0.014 and 0.316 +/- 0.061, respectively, at 24 h and 0.339 +/- 0.094 and 0.530 +/- 0.104, respectively, at 48 h (P < 0.01). The agreement of the microdilution method with the RPMI within two doubling dilutions of the macrodilution reference were 91.5% (118/129) at 24 h and 76.7% (99/129) at 48 h. The percentage of agreement in the microdilution method with the RPMI-2% glucose were significantly higher: 100% (129/129) at 24 h and 99.2% (128/129) at 48 h (P < 0.01). In addition, the MIC endpoints were easier to detect in RPMI-2% glucose, because of the greater difference in absorbance in between grown wells and fluconazole-inhibited wells (P < 0.01). CONCLUSIONS: The spectrophotometric microdilution method with RPMI-2% glucose may have an excellent agreement with the NCCLS broth macrodilution method and may provide more easily determined MIC endpoints for fluconazole susceptibility testing for C. albicans.

Clinical Significance of Pastorex Candida Antigen Assay in Patients with Candidemia / 대한임상병리학회지

BACKGROUND: The incidence of candidemia has increased, and an early differentiation of transient or central venous catheter (CVC)-related candidemia from deep-seated invasive candidiasis is often difficult. The Pastorex Candida antigen assay (Sanofi Diagnostics Pasteur, Marnes-la-Coquette, France) is known to be an useful and specific tool for the diagnosis of invasive candidiasis. We assessed the clinical significance of Pastorex Candida antigen assay in patients with candidemia. METHODS: Eighty-five sera from 27 patients with candidemia and 42 control sera (32 patients with superficial Candida colonization and 10 healthy subjects) were tested. The Pastorex Candida latex agglutination test was performed to evaluate the presence of Candida mannan antigen. Candidemia was divided into 3 categories; (i) transient, (ii) CVC-related, and (iii) non-CVC-related persistent types. RESULTS: Thirty-two patients with superficial Candida colonization and 10 healthy subjects were negative for the Pastorex Candida antigen. Of the 85 sera from 27 patients with candidemia, 14 (16.4%) were positive for the Pastorex Candida antigen. The Pastorex Candida antigen was detected neither in 6 patients with transient candidemia nor 15 patients with CVC-related candidemia. Conversely, it was detected in at least one serum sample of 5 of the 6 (83.3%) patients with non-CVC-related persistent candidemia. Of the 24 sera from 6 patients with non-CVC-related persistent candidemia, 14 (58.3%) were positive for the Pastorex Candida antigen. Overall, the sensitivity and specificity of the Pastorex Candida antigen assay for the diagnosis of non-CVC-related persistent candidemia were 83.3% and 100%, respectively. CONCLUSIONS: Our data suggest that the Pastorex Candida antigen assay has a potential for the differential diagnosis of non-CVC-related persistent candidemia from transient or CVC-related candidemia.

Detection of Mycobacterium tuberculosis using BACTEC Mycobacteria Growth Indicator Tube(MGIT) 960 system: Comparison with BACTEC 460 TB system and Ogawa Media / 대한임상병리학회지

BACKGROUND: BACTEC MGIT 960 system(Becton Dickinson, USA; MGIT 960) is a fully automated, noninvasive culture system for mycobacteria, which has been regarded as a sensitive and least labor-intensive method. This study was purposed to evaluate the performance of MGIT 960 compared to BACTEC 460 TB radiometric system(Becton Dickinson, USA; BACTEC 460) and Ogawa media. METHODS: A total of 1,067 clinical specimens submitted from April to June in 1999 was cultured for acid fast bacilli(AFB). All specimens were digested, decontaminated by the 6% sodium hydroxide(final concentration of 1.5%) and 0.5% N-acetyl-L-cysteine method. All specimens were inoculated into three kinds of media: a MGIT, a BACTEC 12B, and an Ogawa medium. The AFB recovered from cultures were identified to M. tuberculosis complex and MOTT by NAP test. RESULTS: Of 106 isolates of M. tuberculosis recovered from all culture systems, 101(95.3 %) were detected in the MGIT 960, 95(89.6%) in the BACTEC 460 and 76(71.7%) on Ogawa media. MGIT 960 plus Ogawa media detected 104(98.1%) isolates and BACTEC 460 plus Ogawa media recovered 96(90.6%) isolates. The mean time required for detection of M. tuberculosis was 12.7+/-5.8 days with MGIT 960, 16.2+/-7.7 days with BACTEC 460, and 22.8+/-9.5 days with Ogawa media. The contamination rate were 5.1% for MGIT 960, 2.7% for BACTEC 460, and 6.7% for Ogawa media. CONCLUSIONS: MGIT 960 is a sensitive and rapid method to isolate M. tuberculosis.