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Sudhaharan S, Sundarapu N A R, Pamidimukkala U. Comparison of Vitek 2C antifungal susceptibility testing with broth microdilution testing for Candida species. mljgoums 2024; 18 (5) :4-7
URL: http://mlj.goums.ac.ir/article-1-1656-en.html
URL: http://mlj.goums.ac.ir/article-1-1656-en.html
1- Nizam’s Institute of Medical Sciences, Hyderabad, Telangana, India , sukanyavimala@gmail.com
2- Nizam’s Institute of Medical Sciences, Hyderabad, Telangana, India
2- Nizam’s Institute of Medical Sciences, Hyderabad, Telangana, India
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Conclusion
The majority of Vitek 2C antifungal susceptibility testing results were consistent with those obtained using BMD. There were MiE in the tested Candida spp. except for C. auris, which showed ME for fluconazole and amphotericin B. Thus, the Vitek 2 system provides reliable results in a shorter time, aiding clinicians in initiating timely and appropriate antifungal therapy.
Acknowledgement
This study was done as a part of NIMS-ICMR-AMR surveillance network.
Funding sources
Nil.
Ethical statement
Since this study included isolates from samples routinely sent to the Department of Microbiology and did not involve any patient details or clinical trial, ethical approval was not applicable.
Conflicts of interest
The authors declare no conflict of interest.
Author contributions
SS: designed the concept, conducted the literature review, wrote the manuscript, analyzed the results, and edited the manuscript. NARS: handled the technical aspects and reviewed the manuscript. UP: contributed to the concept design and reviewed the manuscript.
Full-Text: (228 Views)
Introduction
Candida infections are one of the most common causes of invasive fungal infections in hospitals, leading to high morbidity and mortality, particularly in patients in intensive care units (1,2). In recent years, azole-resistant non-Candida albicans and multidrug-resistant Candida auris have emerged as important pathogens, resulting in treatment failures (2,3). Hence antifungal susceptibility testing (AFST) is important to monitor the development of resistance in these species. We need a faster and more reliable method for AFST. According to the Clinical and Laboratory Standard Institute (CLSI) guidelines, the reference method for AFST is Broth microdilution (BMD). However, the fully automated system, Vitek 2C system may reduce the workload and observer bias associated with manual BMD. It has also been updated for AFST of Candida species and interpreted as per the latest CLSI guidelines. The aim of this study was to compare the antifungal susceptibility results of Candida spp. with Vitek 2 AST-YS08 card and standard BMD method.
Methods
A total of 50 isolates of Candida spp. from blood cultures were included in the study from January 2022 to December 2022. Identification was done by the Vitek 2C system using the YST card (bioMérieux, Marcy l’Etoile, France). The antifungal susceptibility testing was done both by Vitek 2C using the YS08 (bioMérieux, Marcy l’Etoile, France) card, and BMD was done according to the CLSI guidelines M27M44S-Ed3 (4). The CLSI quality control isolate Candida parapsilosis ATCC 22019 was used as a control for each run.
The following drugs were tested, and the range of the concentration was 0.125-64 µg/L for fluconazole, and 0.03-16 µg/L for voriconazole, caspofungin, micafungin, and amphotericin B. The antifungal powders were procured from Sigma-Aldrich, United States. The test was done in duplicate. Discrepant results were repeated by both Vitek 2C and BMD. The BMD plates were incubated at 35 ○C for 24 hrs. The results were interpreted as per CLSI guidelines M27M44S-Ed3 (5). Vitek 2C does not display MIC for C. auris. The results of MIC were extrapolated in Vitek 2C by choosing any non-Candida albicans. The breakpoints recommended by the CDC were used for C. auris (6).
Discrepancies among MIC endpoints of more than two dilutions (Two wells) were used to calculate the essential agreement (EA) between the MICs determined with the Vitek 2 system and by the reference BMD (2). Categorical agreement (CA) was defined as the percentage of isolates classified in the same category by the reference procedures and the test method (7). Very major errors (VME) were defined when the isolate was considered resistant by the reference procedure but susceptible by the VITEK 2 system. Major errors (ME) occurred when the isolate was susceptible by the reference method but resistant by the VITEK 2 system, and minor error (MiE) were identified when the results by one of the methods included susceptible or resistant and susceptible-dose dependent/intermediate by the other method (8).
Results
The isolates of Candida included seven Candida albicans, nine Candida tropicalis, fourteen Candida parapsilosis, eleven Candida glabrata, and nine Candida auris. For C. albicans, the categorical agreement was 85.8%, 71.5%, 85.8%, and 100% for fluconazole, voriconazole, caspofungin, and micafungin, respectively. The MiE of 14.2% was detected for fluconazole and caspofungin, as well as 28.5% for voriconazole, in C. albicans. In C. glabrata, the CA was 100% for micafungin and voriconazole, while 63.7% for caspofungin. An MiE of 36.3% for caspofungin were detected. C. parapsilosis showed 100% CA for fluconazole, caspofungin, micafungin and 85.8% for voriconazole. There were 14.2% of MiEs for voriconazole. In C. tropicalis, a 100% CA was observed for fluconazole, micafungin, and caspofungin, and 88.9% for voriconazole. Also, an MiE of 11.1% (1/9) was observed for voriconazole. In C. auris, there was a 100% CA observed for caspofungin and micafungin, 77.8% for fluconazole, and 66.7% for amphotericin B. There was an ME of 22.2% for fluconazole and 33.3% for amphotericin B.
The susceptibility pattern of the Candida spp. by BMD and Vitek 2C is shown in Table 1. The agreement between the results of Vitek 2C antifungal susceptibility testing with broth microdilution was high for all tested Candida spp. (Table 2).
Discussion
Antifungal susceptibility testing will help clinicians start appropriate treatment for invasive Candidal infections. The standard method for AFST is BMD, which is time-consuming and difficult to perform (9). The Vitek 2 YS08 card has been updated for AFST according to the CLSI guidelines. It is easy to perform and the time consumed is shorter compared to BMD (1). In the present study, AFST was done for 50 isolates of Candida spp. against fluconazole, voriconazole, caspofungin, and micafungin. Many studies have compared Vitek 2 results with CLSI BMD at 24 hrs and 48 hrs of incubation (9,10,11). In the present study, the BMD plates were read at 24hrs as per CLSI guidelines (6). In the present study, the EA for fluconazole, caspofungin and micafungin was 100% across all tested Candida spp., while it was lower for voriconazole in C. albicans and C. parapsilosis. The comparison of the EA, CA, and various errors from different studies is shown in Table 3.
There was an MiE of 14.2% in case of C. albicans and ME of 22.2% in case of C. auris for fluconazole. Vitek 2 does not show the MIC for fluconazole in C. glabrata as the modified fluconazole formulation has not been validated for the species. Hence, this drug was not compared in the present study for C. glabrata (1). In the present study, we reported MiE for voriconazole for the Candida spp. Other studies have reported both minor and major errors for voriconazole (Table 3). For voriconazole, there are no breakpoints for C. glabrata in CLSI (5) and for C. auris in CDC (4). In the case of C. glabrata, there is insufficient data to correlate between in vitro susceptibility testing and clinical outcomes for voriconazole (5).
Minor errors (MiEs) were observed with caspofungin in C. albicans and C. glabrata in the present study. In other studies, there were both minor and major errors for caspofungin (Table 3). In the case of C. glabrata, micafungin results are more reliable in YS08 than caspofungin as there is a high chance of false resistance (12). Eight isolates of C. glabrata were resistant and one isolate had intermediate susceptibility to caspofungin in our study. According to the CLSI guidelines, caspofungin resistance should be confirmed with additional testing such as micafungin/anidulafungin and DNA sequence analysis of FKS genes (5). In the current study, no minor or major errors were observed with micafungin for other Candida spp., while there was an ME with micafungin in a study from Korea (1). The breakpoints for C. parapsilosis are applicable only in regions with a low prevalence of cryptic species (5).
In the case of C. auris, the EA was low for amphotericin B and there was an ME of 33.3%. Hence, automated systems should be used with caution and results should be confirmed with BMD while doing AFST for C. auris (13). There are only two studies that assessed the clinical performance of YS08 cards compared with BMD using clinical isolates from the world (1,14,15). Two studies had compared YS08 with sensititer (12,16). To the best of our knowledge, this is probably the second study from India.
Candida infections are one of the most common causes of invasive fungal infections in hospitals, leading to high morbidity and mortality, particularly in patients in intensive care units (1,2). In recent years, azole-resistant non-Candida albicans and multidrug-resistant Candida auris have emerged as important pathogens, resulting in treatment failures (2,3). Hence antifungal susceptibility testing (AFST) is important to monitor the development of resistance in these species. We need a faster and more reliable method for AFST. According to the Clinical and Laboratory Standard Institute (CLSI) guidelines, the reference method for AFST is Broth microdilution (BMD). However, the fully automated system, Vitek 2C system may reduce the workload and observer bias associated with manual BMD. It has also been updated for AFST of Candida species and interpreted as per the latest CLSI guidelines. The aim of this study was to compare the antifungal susceptibility results of Candida spp. with Vitek 2 AST-YS08 card and standard BMD method.
Methods
A total of 50 isolates of Candida spp. from blood cultures were included in the study from January 2022 to December 2022. Identification was done by the Vitek 2C system using the YST card (bioMérieux, Marcy l’Etoile, France). The antifungal susceptibility testing was done both by Vitek 2C using the YS08 (bioMérieux, Marcy l’Etoile, France) card, and BMD was done according to the CLSI guidelines M27M44S-Ed3 (4). The CLSI quality control isolate Candida parapsilosis ATCC 22019 was used as a control for each run.
The following drugs were tested, and the range of the concentration was 0.125-64 µg/L for fluconazole, and 0.03-16 µg/L for voriconazole, caspofungin, micafungin, and amphotericin B. The antifungal powders were procured from Sigma-Aldrich, United States. The test was done in duplicate. Discrepant results were repeated by both Vitek 2C and BMD. The BMD plates were incubated at 35 ○C for 24 hrs. The results were interpreted as per CLSI guidelines M27M44S-Ed3 (5). Vitek 2C does not display MIC for C. auris. The results of MIC were extrapolated in Vitek 2C by choosing any non-Candida albicans. The breakpoints recommended by the CDC were used for C. auris (6).
Discrepancies among MIC endpoints of more than two dilutions (Two wells) were used to calculate the essential agreement (EA) between the MICs determined with the Vitek 2 system and by the reference BMD (2). Categorical agreement (CA) was defined as the percentage of isolates classified in the same category by the reference procedures and the test method (7). Very major errors (VME) were defined when the isolate was considered resistant by the reference procedure but susceptible by the VITEK 2 system. Major errors (ME) occurred when the isolate was susceptible by the reference method but resistant by the VITEK 2 system, and minor error (MiE) were identified when the results by one of the methods included susceptible or resistant and susceptible-dose dependent/intermediate by the other method (8).
Results
The isolates of Candida included seven Candida albicans, nine Candida tropicalis, fourteen Candida parapsilosis, eleven Candida glabrata, and nine Candida auris. For C. albicans, the categorical agreement was 85.8%, 71.5%, 85.8%, and 100% for fluconazole, voriconazole, caspofungin, and micafungin, respectively. The MiE of 14.2% was detected for fluconazole and caspofungin, as well as 28.5% for voriconazole, in C. albicans. In C. glabrata, the CA was 100% for micafungin and voriconazole, while 63.7% for caspofungin. An MiE of 36.3% for caspofungin were detected. C. parapsilosis showed 100% CA for fluconazole, caspofungin, micafungin and 85.8% for voriconazole. There were 14.2% of MiEs for voriconazole. In C. tropicalis, a 100% CA was observed for fluconazole, micafungin, and caspofungin, and 88.9% for voriconazole. Also, an MiE of 11.1% (1/9) was observed for voriconazole. In C. auris, there was a 100% CA observed for caspofungin and micafungin, 77.8% for fluconazole, and 66.7% for amphotericin B. There was an ME of 22.2% for fluconazole and 33.3% for amphotericin B.
The susceptibility pattern of the Candida spp. by BMD and Vitek 2C is shown in Table 1. The agreement between the results of Vitek 2C antifungal susceptibility testing with broth microdilution was high for all tested Candida spp. (Table 2).
Discussion
Antifungal susceptibility testing will help clinicians start appropriate treatment for invasive Candidal infections. The standard method for AFST is BMD, which is time-consuming and difficult to perform (9). The Vitek 2 YS08 card has been updated for AFST according to the CLSI guidelines. It is easy to perform and the time consumed is shorter compared to BMD (1). In the present study, AFST was done for 50 isolates of Candida spp. against fluconazole, voriconazole, caspofungin, and micafungin. Many studies have compared Vitek 2 results with CLSI BMD at 24 hrs and 48 hrs of incubation (9,10,11). In the present study, the BMD plates were read at 24hrs as per CLSI guidelines (6). In the present study, the EA for fluconazole, caspofungin and micafungin was 100% across all tested Candida spp., while it was lower for voriconazole in C. albicans and C. parapsilosis. The comparison of the EA, CA, and various errors from different studies is shown in Table 3.
There was an MiE of 14.2% in case of C. albicans and ME of 22.2% in case of C. auris for fluconazole. Vitek 2 does not show the MIC for fluconazole in C. glabrata as the modified fluconazole formulation has not been validated for the species. Hence, this drug was not compared in the present study for C. glabrata (1). In the present study, we reported MiE for voriconazole for the Candida spp. Other studies have reported both minor and major errors for voriconazole (Table 3). For voriconazole, there are no breakpoints for C. glabrata in CLSI (5) and for C. auris in CDC (4). In the case of C. glabrata, there is insufficient data to correlate between in vitro susceptibility testing and clinical outcomes for voriconazole (5).
Minor errors (MiEs) were observed with caspofungin in C. albicans and C. glabrata in the present study. In other studies, there were both minor and major errors for caspofungin (Table 3). In the case of C. glabrata, micafungin results are more reliable in YS08 than caspofungin as there is a high chance of false resistance (12). Eight isolates of C. glabrata were resistant and one isolate had intermediate susceptibility to caspofungin in our study. According to the CLSI guidelines, caspofungin resistance should be confirmed with additional testing such as micafungin/anidulafungin and DNA sequence analysis of FKS genes (5). In the current study, no minor or major errors were observed with micafungin for other Candida spp., while there was an ME with micafungin in a study from Korea (1). The breakpoints for C. parapsilosis are applicable only in regions with a low prevalence of cryptic species (5).
In the case of C. auris, the EA was low for amphotericin B and there was an ME of 33.3%. Hence, automated systems should be used with caution and results should be confirmed with BMD while doing AFST for C. auris (13). There are only two studies that assessed the clinical performance of YS08 cards compared with BMD using clinical isolates from the world (1,14,15). Two studies had compared YS08 with sensititer (12,16). To the best of our knowledge, this is probably the second study from India.
Table 1. Susceptibility pattern of Candida spp. by BMD and Vitek 2C.PNG) Table 2. Comparison of results of Vitek 2C antifungal susceptibility testing with BMD .PNG) |
Table 3. Comparison with other similar studies.PNG) |
Conclusion
The majority of Vitek 2C antifungal susceptibility testing results were consistent with those obtained using BMD. There were MiE in the tested Candida spp. except for C. auris, which showed ME for fluconazole and amphotericin B. Thus, the Vitek 2 system provides reliable results in a shorter time, aiding clinicians in initiating timely and appropriate antifungal therapy.
Acknowledgement
This study was done as a part of NIMS-ICMR-AMR surveillance network.
Funding sources
Nil.
Ethical statement
Since this study included isolates from samples routinely sent to the Department of Microbiology and did not involve any patient details or clinical trial, ethical approval was not applicable.
Conflicts of interest
The authors declare no conflict of interest.
Author contributions
SS: designed the concept, conducted the literature review, wrote the manuscript, analyzed the results, and edited the manuscript. NARS: handled the technical aspects and reviewed the manuscript. UP: contributed to the concept design and reviewed the manuscript.
Research Article: Research Article |
Subject:
Mycology
Received: 2023/04/20 | Accepted: 2024/09/23 | Published: 2025/04/16 | ePublished: 2025/04/16
Received: 2023/04/20 | Accepted: 2024/09/23 | Published: 2025/04/16 | ePublished: 2025/04/16
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