scholarly journals Comparative Evaluation of FUNGITEST and Broth Microdilution Methods for Antifungal Drug Susceptibility Testing of Candida Species and Cryptococcus neoformans

1998 ◽  
Vol 36 (4) ◽  
pp. 926-930 ◽  
Author(s):  
Kate G. Davey ◽  
Ann D. Holmes ◽  
Elizabeth M. Johnson ◽  
Adrien Szekely ◽  
David W. Warnock
Keyword(s):  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Susceptibility Testing ◽  
Clinical Laboratory ◽  
Drug Susceptibility ◽  
Drug Susceptibility Testing ◽  
National Committee ◽  
Broth Microdilution ◽  
Fluconazole Resistant

The FUNGITEST method (Sanofi Diagnostics Pasteur, Paris, France) is a microplate-based procedure for the breakpoint testing of six antifungal agents (amphotericin B, flucytosine, fluconazole, itraconazole, ketoconazole, and miconazole). We compared the FUNGITEST method with a broth microdilution test, performed according to National Committee for Clinical Laboratory Standards document M27-A guidelines, for determining the in vitro susceptibilities of 180 isolates ofCandida spp. (50 C. albicans, 50C. glabrata, 10 C. kefyr, 20C. krusei, 10 C. lusitaniae, 20C. parapsilosis, and 20 C. tropicalisisolates) and 20 isolates of Cryptococcus neoformans. Overall, there was 100% agreement between the methods for amphotericin B, 95% agreement for flucytosine, 84% agreement for miconazole, 83% agreement for itraconazole, 77% agreement for ketoconazole, and 76% agreement for fluconazole. The overall agreement between the methods exceeded 80% for all species tested with the exception ofC. glabrata (71% agreement). The poorest agreement between the results for individual agents was seen with C. glabrata (38% for fluconazole, 44% for ketoconazole, and 56% for itraconazole) and C. tropicalis (50% for miconazole). The FUNGITEST method misclassified as susceptible 2 of 12 (16.6%) fluconazole-resistant isolates, 2 of 10 (20%) itraconazole-resistant isolates, and 4 of 8 (50%) ketoconazole-resistant isolates of several Candida spp. Further development of the FUNGITEST procedure will be required before it can be recommended as an alternative method for the susceptibility testing of Candida spp. or C. neoformans.

scholarly journals Comparison of E-Test and Broth Microdilution Methods for Antifungal Drug Susceptibility Testing of Molds

1999 ◽  
Vol 37 (5) ◽  
pp. 1480-1483 ◽  
Author(s):  
Adrien Szekely ◽  
Elizabeth M. Johnson ◽  
David W. Warnock
Keyword(s):  
Amphotericin B ◽  
Clinical Laboratory ◽  
Drug Susceptibility ◽  
Drug Susceptibility Testing ◽  
National Committee ◽  
Broth Microdilution ◽  
Suitable Alternative ◽  
Absidia Corymbifera ◽  
Broth Microdilution Method

We compared the E test with a broth microdilution method, performed according to National Committee for Clinical Laboratory Standards document M27-A guidelines, for determining the in vitro susceptibilities of 90 isolates of pathogenic molds (10 Absidia corymbifera, 10 Aspergillus flavus, 10Aspergillus fumigatus, 10 Aspergillus niger, 10Aspergillus terreus, 10 Exophiala dermatitidis, 10 Fusarium solani, 10 Scedosporium apiospermum, 5 Scedosporium prolificans, and 5Scopulariopsis brevicaulis). Overall, there was 71% agreement between the results of the two methods for amphotericin B (E-test MICs within ±2 log2 dilutions of broth microdilution MICs) and 88% agreement with the results for itraconazole. The overall levels of agreement (within ±2 log2 dilutions) were ≥80% for 5 of the 10 species tested against amphotericin B and 8 of the 10 species tested against itraconazole. The best agreement between the results was seen withA. fumigatus and A. terreus (100% of results for both agents within ±2 log2 dilutions). The poorest agreement was seen with S. apiospermum, S. prolificans, and S. brevicaulis tested against amphotericin B (20% of results within ±2 log2 dilutions). In every instance, this low level of agreement was due to isolates for which the broth microdilution MICs were low but for which the E-test MICs were much higher. The E test appears to be a suitable alternative procedure for testing the susceptibility of Aspergillusspp. and some other molds to amphotericin B or itraconazole.

scholarly journals Evaluation of Amphotericin B Interpretive Breakpoints for Candida Bloodstream Isolates by Correlation with Therapeutic Outcome

2006 ◽  
Vol 50 (4) ◽  
pp. 1287-1292 ◽  
Author(s):  
Benjamin J. Park ◽  
Beth A. Arthington-Skaggs ◽  
Rana A. Hajjeh ◽  
Naureen Iqbal ◽  
Meral A. Ciblak ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Susceptibility Testing ◽  
Clinical Laboratory ◽  
National Committee ◽  
Restricted Range ◽  
Broth Microdilution ◽  
Therapeutic Success ◽  
In Vitro Susceptibility ◽  
In Vitro Susceptibility Testing

ABSTRACT One hundred seven Candida bloodstream isolates (51 C. albicans, 24 C. glabrata, 13 C. parapsilosis, 13 C. tropicalis, 2 C. dubliniensis, 2 C. krusei, and 2 C. lusitaniae strains) from patients treated with amphotericin B alone underwent in vitro susceptibility testing against amphotericin B using five different methods. Fifty-four isolates were from patients who failed treatment, defined as death 7 to 14 days after the incident candidemia episode, having persistent fever of ≥5 days' duration after the date of the incident candidemia, or the recurrence of fever after two consecutive afebrile days while on antifungal treatment. MICs were determined by using the Clinical Laboratory Standards Institute (formally National Committee for Clinical Laboratory Standards) broth microdilution procedure with two media and by using Etest. Minimum fungicidal concentrations (MFCs) were also measured in two media. Broth microdilution tests with RPMI 1640 medium generated a restricted range of MICs (0.125 to 1 μg/ml); the corresponding MFC values ranged from 0.5 to 4 μg/ml. Broth microdilution tests with antibiotic medium 3 produced a broader distribution of MIC and MFC results (0.015 to 0.25 μg/ml and 0.06 to 2 μg/ml, respectively). Etest produced the widest distribution of MICs (0.094 to 2 μg/ml). However, none of the test formats studied generated results that significantly correlated with therapeutic success or failure.

scholarly journals Trends in Antifungal Drug Susceptibility of Cryptococcus neoformans Isolates in the United States: 1992 to 1994 and 1996 to 1998

2001 ◽  
Vol 45 (11) ◽  
pp. 3065-3069 ◽  
Author(s):  
Mary E. Brandt ◽  
Michael A. Pfaller ◽  
Rana A. Hajjeh ◽  
Richard J. Hamill ◽  
Peter G. Pappas ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Antifungal Agents ◽  
Clinical Laboratory ◽  
The United States ◽  
Drug Susceptibility ◽  
Antifungal Drug ◽  
National Committee ◽  
Broth Microdilution Method

ABSTRACT The antifungal drug susceptibilities of two collections ofCryptococcus neoformans isolates obtained through active laboratory-based surveillance from 1992 to 1994 (368 isolates) and 1996 to 1998 (364 isolates) were determined. The MICs of fluconazole, itraconazole, and flucytosine were determined by the National Committee for Clinical Laboratory Standards broth microdilution method; amphotericin B MICs were determined by the E-test. Our results showed that the MIC ranges, the MICs at which 50% of isolates are inhibited (MIC50s), and the MIC90s of these four antifungal agents did not change from 1992 to 1998. In addition, very small numbers of isolates showed elevated MICs suggestive of in vitro resistance. The MICs of amphotericin B were elevated (≥2 μg/ml) for 2 isolates, and the MICs of flucytosine were elevated (≥32 μg/ml) for 14 isolates. Among the azoles, the fluconazole MIC was elevated (≥64 μg/ml) for 8 isolates and the itraconazole MIC (≥1 μg/ml) was elevated for 45 isolates. Analysis of 172 serial isolates from 71 patients showed little change in the fluconazole MIC over time. For isolates from 58 patients (82% of serial cases) there was either no change or a twofold change in the fluconazole MIC. In contrast, for isolates from seven patients (12% of serial cases) the increase in the MIC was at least fourfold. For isolates from another patient there was a 32-fold decrease in the fluconazole MIC over a 1-month period. We conclude that in vitro resistance to antifungal agents remains uncommon in C. neoformans and has not significantly changed with time during the past decade.

scholarly journals The Trailing End Point Phenotype in Antifungal Susceptibility Testing Is pH Dependent

1999 ◽  
Vol 43 (6) ◽  
pp. 1383-1386 ◽  
Author(s):  
Kieren A. Marr ◽  
Tige R. Rustad ◽  
John H. Rex ◽  
Theodore C. White
Keyword(s):  
Minimal Medium ◽  
Susceptibility Testing ◽  
Clinical Laboratory ◽  
Ph Effect ◽  
Antifungal Susceptibility ◽  
Drug Susceptibility ◽  
Drug Susceptibility Testing ◽  
National Committee ◽  
End Points

ABSTRACT The interpretation of end points in azole antifungal drug susceptibility testing is problematic, in part due to incomplete growth inhibition of Candida species. Such trailing growth can cause the MICs of fluconazole for some isolates to be low (<1 μg/ml) after 24 h of growth but much higher (>64 μg/ml) after 48 h. Isolates having this type of growth have been described as having a low-high phenotype. Although these isolates would be considered resistant by current National Committee of Clinical Laboratory Standards definitions, growing evidence suggests that they are susceptible in vivo. To further characterize these isolates in vitro, microdilution susceptibility testing comparing the complex defined medium RPMI 1640 to a defined minimal medium (yeast nitrogen broth) was performed. Isolates having trailing growth in MOPS (morpholinepropanesulfonic acid)-buffered RPMI 1640 (pH 7.0) were found to have clear end points in the minimal medium at its native pH of 4.5. The pH of the medium influenced the low-high phenotype, as these same isolates trailed in minimal medium adjusted to a pH of ≥6.0 but did not trail in RPMI 1640 adjusted to a pH of ≤5.0. This pH effect was independent of the medium buffering capacity, as trailing was decreased in both minimal medium and RPMI 1640 (pH 4.5) buffered in citrate. Adjustment in the pH of MOPS-buffered RPMI 1640 reduced trailing in multiple strains of Candida albicans without affecting the MICs for isolates having known susceptible (low-low) and resistant (high-high) phenotypes. Adjustment of the medium pH could be considered to eliminate trailing in azole drug susceptibility testing.

Use of spectrophotometric reading for in vitro antifungal susceptibility testing ofAspergillusspp.

1999 ◽  
Vol 45 (10) ◽  
pp. 871-874 ◽  
Author(s):  
Eric Dannaoui ◽  
Florence Persat ◽  
Marie-France Monier ◽  
Elisabeth Borel ◽  
Marie-Antoinette Piens ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Susceptibility Testing ◽  
Clinical Laboratory ◽  
Antifungal Susceptibility ◽  
Reference Method ◽  
Antifungal Susceptibility Testing ◽  
Aspergillus Species ◽  
National Committee ◽  
Broth Microdilution Method

A comparative study of visual and spectrophotometric MIC endpoint determinations for antifungal susceptibility testing of Aspergillus species was performed. A broth microdilution method adapted from the National Committee for Clinical Laboratory Standards (NCCLS) was used for susceptibility testing of 180 clinical isolates of Aspergillus species against amphotericin B and itraconazole. MICs were determined visually and spectrophotometrically at 490 nm after 24, 48, and 72h of incubation, and MIC pairs were compared. The agreement between the two methods was 99% for amphotericin B and ranged from 95 to 98% for itraconazole. It is concluded that spectrophotometric MIC endpoint determination is a valuable alternative to the visual reference method for susceptibility testing of Aspergillus species.Key words: antifungal, susceptibility testing, Aspergillus, spectrophotometric reading.

scholarly journals Interactions between Triazoles and Amphotericin B against Cryptococcus neoformans

2000 ◽  
Vol 44 (9) ◽  
pp. 2435-2441 ◽  
Author(s):  
Francesco Barchiesi ◽  
Anna M. Schimizzi ◽  
Francesca Caselli ◽  
Andrea Novelli ◽  
Stefania Fallani ◽  
...  
Keyword(s):  
Combination Therapy ◽  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Clinical Laboratory ◽  
Fungal Infections ◽  
National Committee ◽  
Positive Interaction ◽  
Pronounced Increase

ABSTRACT The interaction of amphotericin B (AmB) and azole antifungal agents in the treatment of fungal infections is still a controversial issue. A checkerboard titration broth microdilution-based method that adhered to the recommendations of the National Committee for Clinical Laboratory Standards was applied to study the in vitro interactions of AmB with fluconazole (FLC), itraconazole (ITC), and the new investigational triazole SCH 56592 (SCH) against 15 clinical isolates ofCryptococcus neoformans. Synergy, defined as a fractional inhibitory concentration (FIC) index of ≤0.50, was observed for 7% of the isolates in studies of the interactions of both FLC-AmB and ITC-AmB and for 33% of the isolates in studies of the SCH-AmB interactions; additivism (FICs, >0.50 to 1.0) was observed for 67, 73, and 53% of the isolates in studies of the FLC-AmB, ITC-AmB, and SCH-AmB interactions, respectively; indifference (FICs, >1.0 to ≤2.0) was observed for 26, 20, and 14% of the isolates in studies of the FLC-AmB, ITC-AmB, and SCH-AmB interactions, respectively. Antagonism (FIC >2.0) was not observed. When synergy was not achieved, there was still a decrease, although not as dramatic, in the MIC of one or both drugs when they were used in combination. To investigate the effects of FLC-AmB combination therapy in vivo, we established an experimental model of systemic cryptococcosis in BALB/c mice by intravenous injection of cells of C. neoformans 2337, a clinical isolate belonging to serotype D against which the combination of FLC and AmB yielded an additive interaction in vitro. Both survival and tissue burden studies showed that combination therapy was more effective than FLC alone and that combination therapy was at least as effective as AmB given as a single drug. On the other hand, when cells of C. neoformans 2337 were grown in FLC-containing medium, a pronounced increase in resistance to subsequent exposures to AmB was observed. In particular, killing experiments conducted with nonreplicating cells showed that preexposure to FLC abolished the fungicidal activity of the polyene. However, this apparent antagonism was not observed in vivo. Rather, when the two drugs were used sequentially for the treatment of systemic murine cryptococcosis, a reciprocal potentiation was often observed. Our study shows that (i) the combination of triazoles and AmB is significantly more active than either drug alone against C. neoformans in vitro and (ii) the concomitant or sequential use of FLC and AmB for the treatment of systemic murine cryptococcosis results in a positive interaction.

scholarly journals In Vitro Susceptibilities of Candida dubliniensisIsolates Tested against the New Triazole and Echinocandin Antifungal Agents

1999 ◽  
Vol 37 (3) ◽  
pp. 870-872 ◽  
Author(s):  
M. A. Pfaller ◽  
S. A. Messer ◽  
S. Gee ◽  
S. Joly ◽  
C. Pujol ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Antifungal Agents ◽  
Clinical Laboratory ◽  
National Committee ◽  
Mucosal Disease ◽  
Fluconazole Resistant ◽  
Resistant Strains ◽  
Investigational Agents ◽  
Multidrug Resistance Transporters

Candida dubliniensis is a newly recognized fungal pathogen causing mucosal disease in AIDS patients. Although preliminary studies indicate that most strains of C. dubliniensis are susceptible to established antifungal agents, fluconazole-resistant strains have been detected. Furthermore, fluconazole-resistant strains are easily derived in vitro, and these strains exhibit increased expression of multidrug resistance transporters, especially MDR1. Because of the potential for the development of resistant strains of C. dubliniensis, it is prudent to explore the in vitro activities of several of the newer triazole and echinocandin antifungals against isolates of C. dubliniensis. In this study we tested 71 isolates of C. dubliniensis against the triazoles BMS-207147, Sch 56592, and voriconazole and a representative of the echinocandin class of antifungal agents, MK-0991. We compared the activities of these agents with those of the established antifungal agents fluconazole, itraconazole, amphotericin B, and 5-fluorocytosine (5FC) by using National Committee for Clinical Laboratory Standards microdilution reference methods. Our findings indicate that the vast majority of clinical isolates of C. dubliniensis are highly susceptible to both new and established antifungal agents. Strains with decreased susceptibilities to fluconazole remained susceptible to the investigational agents as well as to amphotericin B and 5FC. The increased potencies of the new triazole and echinocandin antifungal agents may provide effective therapeutic options for the treatment of infections due to C. dubliniensis.

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