Species-specific Clinical Breakpoints Research Articles

Nationwide Surveillance of Antifungal Resistance of Candida Bloodstream Isolates in South Korean Hospitals: Two Year Report from Kor-GLASS.

We incorporated nationwide Candida antifungal surveillance into the Korea Global Antimicrobial Resistance Surveillance System (Kor-GLASS) for bacterial pathogens. We prospectively collected and analyzed complete non-duplicate blood isolates and information from nine sentinel hospitals during 2020–2021, based on GLASS early implementation protocol for the inclusion of Candida species. Candida species ranked fourth among 10,758 target blood pathogens and second among 4050 hospital-origin blood pathogens. Among 766 Candida blood isolates, 87.6% were of hospital origin, and 41.3% occurred in intensive care unit patients. Adults > 60 years of age accounted for 75.7% of cases. Based on species-specific clinical breakpoints, non-susceptibility to fluconazole, voriconazole, caspofungin, micafungin, and anidulafungin was found in 21.1% (154/729), 4.0% (24/596), 0.1% (1/741), 0.0% (0/741), and 0.1% (1/741) of the isolates, respectively. Fluconazole resistance was determined in 0% (0/348), 2.2% (3/135, 1 Erg11 mutant), 5.3% (7/133, 6 Pdr1 mutants), and 5.6% (6/108, 4 Erg11 and 1 Cdr1 mutants) of C. albicans, C. tropicalis, C. glabrata, and C. parapsilosis isolates, respectively. An echinocandin-resistant C. glabrata isolate harbored an F659Y mutation in Fks2p. The inclusion of Candida species in the Kor-GLASS system generated well-curated surveillance data and may encourage global Candida surveillance efforts using a harmonized GLASS system.

One-Year Candida Data of the Central Mycology Laboratory: Which Sample, Which Species, How Resistant?

The incidence of fungal infections particularly Candida species, is increasing gradually as a result of the increased life expectancy associated with the advances in the diagnosis and treatment of diseases, and increased number of patients in the risk group over the years. In addition, the incidence of fungal infection types that are resistant to antifungal drugs has been increasing, and rare fungal species have been reported to be isolated more frequently. For this reason, it is indicated that identification to the species level will contribute to the early initiation of an accurate and effective treatment. In this study, it was aimed to define the Candida species isolated from various clinical specimens and to document the performance of antifungal sensitivity tests. The Candida isolates sent to the central mycology laboratory in 2019 for identification and antifungal susceptibility tests were included in the study. The definition of the fungi to the species level was carried out using matrix-assisted laser desorption ionization-time of fl ight mass spectrometry (MALDI-TOF MS) and conventional methods. In vitro antifungal drug susceptibilities were analyzed using the The Clinical and Laboratory Standarts Institute (CLSI, M27-A3) reference broth microdilution method. The minimum inhibitory concentration (MIC) results were interpreted in accordance with the species-specific clinical breakpoints (CBPs) cited in the CLSI-M60 guidelines, and according to the epidemiological cut-off value (ECV) when no CBP was mentioned. The distribution of the species of the total 813 Candida isolates included in the study were as follows: Candida albicans (n= 312 ), Candida parapsilosis (n= 202), Candida tropicalis (n= 92), Candida glabrata (n= 71), Candida dubliniensis (n=28), Candida lusitaniae (n= 26), Candida kefyr (n= 22), Candida utilis (n= 17), Candida krusei (n= 14), Candida orthopsilosis (n= 7), Candida inconspicua (n= 7), Candida guilliermondii (n= 5), Candida metapsilosis (n= 4), Candida norvegensis (n= 4), Candida lambica (n= 1) and Candida lipolytica (n= 1). The evaluation of the results of the antifungal susceptibility tests according to the CBPs revealed that one C.albicans isolate and 60 C.parapsilosis (29.7%) isolates were resistant, and seven C.parapsilosis (3.5%) isolates were dose-dependent susceptible to fluconazole; 32 C.parapsilosis (15.8%) isolates were intermediately susceptible to voriconazole; one C.parapsilosis (0.5%) was resistant and one C.krusei (7.1%) was intermediately susceptible to anidulafungin; and one C.parapsilosis (0.5%) was resistant and one C.krusei (7.1%) isolate was intermediately susceptible to micafungin. In terms of ECVs, one C.lusitaniae isolate for fluconazole and one of each C.lusitaniae and C.kefyr isolates were evaluated as a non-wild type. In the present study, 61 of 813 isolates were found to be resistant to fluconazole and seven were dose dependently susceptible, 32 were intermediately susceptible to voriconazole, one was resistant to anidulafungin, one was intermediately susceptible, and one was resistant to micafungin and one was intermediately susceptible to micafungin. In conclusion, the increased number of non- albicans Candida species and increased levels of resistance to antifungal drugs further establish the importance of early diagnosis at a species level alongside antifungal susceptibility tests.

A Multicenter Study of Antifungal Use and Species Distribution and Antifungal usceptibilities of Candida Isolates in South Korea

Background: Candidiasis control should include monitoring the epidemiology and resistance to various antifungal agents. In this study, the researchers investigated the Candida species recovered from clinical specimens at particular geographic areas or hospitals. Objective: The present study is geared toward the evaluation of antifungal drug usage at Korean hospitals in 2016. It is also essential that species distribution and antifungal susceptibilities of Candida isolates should be looked into to provide important data that can help devise therapeutic strategies to control the disease. Methods: Systemic antifungal agent usage over a one-year period was investigated at 10 Korean hospitals. Identification and antifungal susceptibility tests were performed on clinical isolates of the Candida species, which were collected over a three-month period. Results: The total antifungal usage in each hospital ranged from 7.7 to 158.9 defined daily doses (DDDs) per 1,000 patient days. Fluconazole was most commonly used (37.1%), followed by amphotericin B (30.6%), itraconazole (9.7%), echinocandins (8.8%), voriconazole (7.5%), and posaconazole (6.3%), respectively. Among 274 Candida isolates, C. albicans was the most frequently recovered (51.1%), followed by C. glabrata (15.7%), C. tropicalis (15.0%), and C. parapsilosis (13.5%), respectively. Through the application of either species-specific clinical breakpoints or epidemiological cutoff values to Candida isolates, the non-susceptibility rates to fluconazole, voriconazole, amphotericin B, and micafungin were found in 20.7%, 5.6%, 0%, and 0% of isolates, respectively. Conclusion: This nationwide multicenter study showed that total antifungal use varied considerably according to each hospital. Non-susceptibility to fluconazole should be further monitored, considering the drug's frequent use in Korea.

Antimicrobial Resistance in Corynebacterium spp., Arcanobacterium spp., and Trueperella pyogenes.

There is currently only limited information on the antimicrobial susceptibility and resistance of Corynebacterium spp., Arcanobacterium spp., and Trueperella pyogenes from animals. The comparability of the data is hampered by the use of different antimicrobial susceptibility testing methods and interpretive criteria. To date, standard broth microdilution methods and clinical breakpoints that are approved by the Clinical and Laboratory Standards Institute and are applicable to Corynebacterium spp., Arcanobacterium spp., and T. pyogenes are available. The lack of species-specific clinical breakpoints for the different animal species reduces the explanatory power of the data. Among the isolates of the three genera, elevated MICs for different classes of antimicrobial agents (e.g., β-lactams, macrolides, lincosamides, tetracyclines, aminoglycosides, phenicols, sulfonamides/diaminopyrimidines, and fluoroquinolones) have been described. The most comprehensive data set is available for T. pyogenes, which also includes information about genes and mutations involved in antimicrobial resistance. In T. pyogenes isolates, the macrolide-lincosamide-streptogramin B resistance genes erm(B) and erm(X) were identified. Tetracycline resistance in T. pyogenes was based on the resistance genes tet(W), tet(Z), and tet(33), whereas the aminoglycoside resistance genes aacC, aadA1, aadA2, aadA5, aadA24, and aadB have been described in T. pyogenes. So far, only single genes conferring either phenicol resistance (cmlA6), trimethoprim resistance (dfrB2a), or β-lactam resistance (blaP1) are known to occur in T. pyogenes isolates. Various 23S rRNA mutations, including A2058T, A2058G, and G2137C, were identified in macrolide/lincosamide-resistant T. pyogenes.

Etest and Sensititre YeastOne Susceptibility Testing of Echinocandins against Candida Species from a Single Center in Austria.

Candida species were tested for susceptibility to caspofungin, anidulafungin, and micafungin in order to evaluate the roles of Etest and Sensititre YeastOne in antifungal susceptibility testing for daily routines and to survey resistance. A total of 104 Candida species isolates detected from blood cultures were investigated. With EUCAST broth microdilution as the reference method, essential agreement (EA), categorical agreement (CA), very major errors (VME), major errors (ME), and minor (MIN) errors were assessed by reading MICs at 18, 24, and 48 h. By use of EUCAST broth microdilution and species-specific clinical breakpoints (CBPs), echinocandin resistance was not detected during the study period. Using EUCAST CBPs, MIC readings at 24 h for the Etest and Sensititre YeastOne resulted in CA levels of 99% and 93% for anidulafungin and 99% and 97% for micafungin. Using revised CLSI CBPs for caspofungin, CA levels were 92% and 99% for Etest and Sensititre YeastOne. The Etest proved an excellent, easy-to-handle alternative method for testing susceptibility to anidulafungin and micafungin. Due to misclassifications, the Etest is less suitable for testing susceptibility to caspofungin (8% of isolates falsely tested resistant). The CA levels of Sensititre YeastOne were 93% and 97% for anidulafungin and micafungin (24 h) by use of EUCAST CBPs and increased to 100% for both antifungals if CLSI CBPs were applied and to 100% and 99% if Sensititre YeastOne epidemiological cutoff values (ECOFFs) were applied. No one echinocandin could be demonstrated to be superior to another in vitro Since resistance was lacking among our Candida isolates, we cannot derive any recommendation from accurate resistance detection by the Etest and Sensititre YeastOne.

Antifungal susceptibility profiles of Candida species to triazole: application of new CLSI species-specific clinical breakpoints and epidemiological cutoff values for characterization of antifungal resistance

The Clinical and Laboratory Standards Institute (CLSI) Subcommittee on Antifungal Susceptibility Testing has newly introduced species-specific clinical breakpoints (CBPs) for fluconazole and voriconazole. When CBPs can not be determined, wild-type minimal inhibitory concentration (MIC) distributions are detected and epidemiological cutoff values (ECVs) provide valuable means for the detection of emerging resistance. The aim of this study is to determine triazole resistance patterns in Candida species by the recently revised CLSI CBPs. A total of 140 Candida strains isolated from blood cultures of patients with invasive candidiasis hospitalized in various intensive care units in Turkey and sent to our reference laboratory between 2011-2012, were included in the study. The isolates were identified by conventional methods, and susceptibility testing was performed against fluconazole, itraconazole and voriconazole, by the 24-h CLSI broth microdilution (BMD) method. Azole resistance rates for all Candida species were determined using the new species-specific CLSI CBPs and ECVs criteria, when appropriate. The species distribution of the isolates were as follows; C.parapsilosis (n= 31 ), C.tropicalis (n= 26 ), C.glabrata (n= 21), C.albicans (n= 18), C.lusitaniae (n= 16), C.krusei (n= 16), C.kefyr (n= 9), C.guilliermondii (n= 2), and C.dubliniensis (n= 1). According to the newly determined CLSI CBPs for fluconazole and C.albicans, C.parapsilosis, C.tropicalis [susceptible (S), ≤ 2 µg/ml; dose-dependent susceptible (SDD), 4 µg/ml; resistant (R), ≥ 8 µg/ml], and C.glabrata (SDD, ≤ 32 µg/ml; R≥ 64 µg/ml) and for voriconazole and C.albicans, C.parapsilosis, C.tropicalis (S, ≤ 0.12 µg/ml; SDD, 0.25-0.5 µg/ml; R, ≥ 1 µg/ml), and C.krusei (S, ≤ 0.5 µg/ml; SDD, 1 µg/ml; R, ≥ 2 µg/ml), it was found that three of C.albicans, one of C.parapsilosis and one of C.glabrata isolates were resistant to fluconazole, while two of C.albicans and two of C.tropicalis were resistant to voriconazole. The ECVs of 0.5 µg/ml for voriconazole and C.glabrata were used to differentiate wild-type (MIC ≤ ECV) from non-wild-type (MIC > ECV) strains. Five of C.glabrata were non-WT for voriconazole. Due to the lack of CBPs for the less common species, the ECVs for fluconazole, itraconazole and voriconazole, respectively, were used for C.lusitaniae (2 µg/ml, 0.5 µg/ml, 0.03 µg/ml), C.guilliermondii (8 µg/ml, 1 µg/ml, 0.25 µg/ml), C.dubliniensis (0.5 µg/ml, 0.25 µg/ml, 0.03 µg/ml), and C.kefyr (1 µg/ml, 0.015 µg/ml) to categorize isolates of these species as wild- and non-wild-type. When the ECVs were used for fluconazole, one each of C.lusitaniae, C.dubliniensis and C.kefyr; for voriconazole, three of C.lusitaniae and one of C.kefyr were detected as non-wild-type. Overall, a total of five Candida species were resistant to fluconazole and four to voriconazole and among these species one each of C.parapsilosis, C.tropicalis, C.glabrata, C.lusitaniae, C.kefyr and three of C.albicans exhibited cross-resistance at least against two azoles. It was concluded that, the strains identified as resistant and non-wild-type in this in vitro study should be supported by molecular and in vivo studies for the determination of their clinical validity.

Epidemiology, clinical characteristics, and outcome of candidemia in a tertiary referral center in Italy from 2010 to 2014.

We evaluated the epidemiology, clinical characteristics and outcome of candidemia in a single institution from 2010 to 2014. A retrospective observational study of all cases of candidemia was carried out at a University Hospital in Central Italy including five intensive care units (ICUs), 11 medical and 11 surgical wards. Data regarding demographic characteristics and clinical risk factors were collected from the patient's medical records. Antifungal susceptibility testing was performed and MIC results were interpreted according to species-specific clinical breakpoints. A total of 270 episodes of candidemia were identified. Overall incidence rate was 1.5 episodes/1000 hospital admissions. Although Candida albicans represented the most commonly isolated species, its percentage significantly decreased from 68 to 48 % (p = 0.040). The overall 30-day mortality was 35 %. The variables independently associated with a significant higher risk of mortality were: older age; being hospitalized in ICU or in medical wards vs surgical wards; being infected with C. albicans vs other species; the occurrence of septic shock, pneumonia and acute renal failure; the presence of a solid organ tumor or a chronic pulmonary disease. Conversely, an appropriate treatment was confirmed to be significantly associated with a lower risk of mortality. The overall resistance was low and it was noted only among triazoles. Our study shows that candidemia is a significant source of morbidity and mortality. The identification of risk factors associated with mortality along with the knowledge of local susceptibility may lead to a better management in terms of preventive and therapeutic measures.

Isavuconazole, micafungin, and 8 comparator antifungal agents' susceptibility profiles for common and uncommon opportunistic fungi collected in 2013: temporal analysis of antifungal drug resistance using CLSI species-specific clinical breakpoints and proposed epidemiological cutoff values

The in vitro activities of isavuconazole, micafungin, and 8 comparator antifungal agents were determined for 1613 clinical isolates of fungi (1320 isolates of Candida spp., 155 of Aspergillus spp., 103 of non-Candida yeasts, and 35 non-Aspergillus molds) collected during a global survey conducted in 2013. The vast majority of the isolates of the 21 different species of Candida, with the exception of Candida glabrata (MIC90, 2μg/mL), Candida krusei (MIC90, 1μg/mL), and Candida guilliermondii (MIC90, 8μg/mL), were inhibited by ≤0.25μg/mL of isavuconazole. C. glabrata and C. krusei were largely inhibited by ≤1μg/mL of isavuconazole. Resistance to fluconazole was seen in 0.5% of Candida albicans isolates, 11.1% of C. glabrata isolates, 2.5% of Candida parapsilosis isolates, 4.5% of Candida tropicalis isolates, and 20.0% of C. guilliermondii isolates. Resistance to the echinocandins was restricted to C. glabrata (1.3–2.1%) and C. tropicalis (0.9–1.8%). All agents except for the echinocandins were active against 69 Cryptococcus neoformans isolates, and the triazoles, including isavuconazole, were active against the other yeasts. Both the mold active triazoles as well as the echinocandins were active against 155 Aspergillus spp. isolates belonging to 10 species/species complex. In general, there was low resistance levels to the available systemically active antifungal agents in a large, contemporary (2013), global collection of molecularly characterized yeasts and molds. Resistance to azoles and echinocandins was most prominent among isolates of C. glabrata, C. tropicalis, and C. guilliermondii.

In vitro assessment of chloramphenicol and florfenicol as second-line antimicrobial agents in dogs.

The aim of this study was to evaluate the potential of chloramphenicol and florfenicol as second-line antimicrobial agents for treatment of infections caused by methicillin-resistant Staphylococcus pseudintermedius (MRSP) and extended-spectrum β-lactamase (ESBL)-producing Escherichia coli in dogs, through a systematic in vitro assessment of the pharmacodynamic properties of the two drugs. Minimum inhibitory concentrations (MIC) and phenicol resistance genes were determined for 169 S. pseudintermedius and 167 E. coli isolates. Minimum bactericidal concentrations (MBC), time-killing kinetics, and postantibiotic effect (PAE) of both agents against wild-type isolates of each species were assessed. For S. pseudintermedius, the chloramphenicol MIC90 was 32 μg/mL. No florfenicol resistance was detected in this species (MIC90 = 4 μg/mL). The MIC90 of both agents against E. coli was 8 μg/mL. Resistance genes found were catpC221 in S. pseudintermedius and catA1 and/or floR in E. coli. The phenicols displayed a time-dependent, mainly, bacteriostatic effect on both species. Prolonged PAEs were observed for S. pseudintermedius, and no PAEs were detected for E. coli. More research into determination of PK/PD targets of efficacy is needed to further assess the clinical use of chloramphenicol and florfenicol as second-line agents in dogs, optimize dosage regimens, and set up species-specific clinical break points.

SUSCEPTIBILITY OF Candida spp. ISOLATED FROM BLOOD CULTURES AS EVALUATED USING THE M27-A3 AND NEW M27-S4 APPROVED BREAKPOINTS

The high mortality rates associated with candidemia episodes and the emergence of resistance to antifungal agents necessitate the monitoring of the susceptibility of fungal isolates to antifungal treatments. The new, recently approved, species-specific clinical breakpoints (SS-CBPs)(M27-S4) for evaluating susceptibility require careful interpretation and comparison with the former proposals made using the M27-A3 breakpoints, both from CLSI. This study evaluated the susceptibility of the different species of Candida that were isolated from candidemias based on these two clinical breakpoints. Four hundred and twenty-two isolates were identified and, among them, C. parapsilosis comprised 46.68%, followed by C. albicans (35.78%), C. tropicalis (9.71%), C. glabrata (3.55%), C. lusitaniae (1.65%), C. guilliermondii (1.65%) and C. krusei (0.94%). In accordance with the M27-A3 criteria, 33 (7.81%) non-susceptible isolates were identified, of which 16 (3.79%) were resistant to antifungal agents. According to SS-CBPs, 80 (18.95%) isolates were non-susceptible, and 10 (2.36%) of these were drug resistant. When the total number of non-susceptible isolates was considered, the new SS-CBPs detected 2.4 times the number of isolates that were detected using the M27-A3 interpretative criteria. In conclusion, the detection of an elevated number of non-susceptible species has highlighted the relevance of evaluating susceptibility tests using new, species-specific clinical breakpoints (SS-CBPs), which could impact the profile of non-susceptible Candida spp. to antifungal agents that require continuous susceptibility monitoring.

Nosocomial bloodstream Candida infections in a tertiary-care hospital in South Brazil: a 4-year survey.

The aims of this study were to evaluate the epidemiology of nosocomial candidemia in a tertiary hospital in South Brazil and the in vitro antifungal susceptibility of isolates. Blood strains from 108 patients were identified by PCR-based method. Some 30.5 % of candidemia were caused by Candida tropicalis, 28.7 % were due to Candida albicans, 24.1 % with Candida parapsilosis sensu stricto, 8.3 % with Candida glabrata sensu lato, 1.8 % involved Candida krusei and 6.6 % with other species. Candidemia was more common in intensive care unit settings (66 %). In vitro susceptibility to antifungal drugs was determined by a microdilution method; and new species-specific clinical breakpoints for fluconazole and voriconazole were applied. Overall susceptibility rates were 100 % for itraconazole, 91 % for fluconazole, 98 % for voriconazole and 99 % for amphotericin B. Fluconazole resistance was mostly among C. parapsilosis sensu stricto isolates (26.9 %). Most of the findings reported here agreed with epidemiological features common to other tertiary hospitals in Brazil; but also revealed some peculiarities, such as a high frequency of C. tropicalis associated with candidemia. Besides, high rate of fluconazole resistance among C. parapsilosis stricto sensu isolates was obtained when applying the new species-specific clinical breakpoints.

Minocycline pharmacokinetics and pharmacodynamics in dogs: dosage recommendations for treatment of meticillin-resistant Staphylococcus pseudintermedius infections.

Although minocycline is not licensed for use in dogs, this tetracycline has therapeutic potential against meticillin-resistant Staphylococcus pseudintermedius. The aim of this study was to establish rational dosage recommendations for minocycline use in dogs. Specific objectives were to generate and analyse minocycline pharmacokinetic (PK) data on plasma and interstitial fluid (ISF) concentrations, plasma protein binding and pharmacodynamic (PD) data on antimicrobial activity against S.pseudintermedius. Six healthy dogs from a research colony were used in this study. Dogs were administered 5mg/kg intravenously and 10mg/kg orally (p.o.) of minocycline hydrochloride in separate crossover experiments. In vivo drug concentrations in plasma and in ISF collected by ultrafiltration were measured by high-performance liquid chromatography. Pharmacokinetic analysis was performed on plasma and ISF concentrations. PK/PD analysis was completed using in vitro data on plasma protein binding and minocycline susceptibility in 168 S.pseudintermedius isolates. Minocycline distributed to the ISF to a higher degree than predicted by the protein-unbound fraction in plasma. A large volume of distribution after oral administration, with plasma and ISF elimination half-lives of 4.1 and 7.4h, respectively, demonstrated that the ISF serves as a drug reservoir for sustained tissue concentrations. Monte Carlo simulation, used to assess target attainment at different drug dosages, indicated that p.o. administration of 5mg/kg twice daily is sufficient to inhibit S.pseudintermedius strains with minimal inhibitory concentrations ≤0.25μg/mL. Besides dosage recommendations for therapy of meticillin-resistant Staphylococcus pseudintermedius infections in dogs, the study also provides PK/PD data necessary to consider species-specific clinical breakpoints for minocycline susceptibility testing.

Support for the EUCAST and revised CLSI fluconazole clinical breakpoints by Sensititre(R) YeastOne(R) for Candida albicans: a prospective observational cohort study

Species-specific clinical breakpoints (CBPs) for Candida spp. were established following consideration of clinical outcomes in patients with oesophageal candidiasis. We sought to further determine the validity of the current CBPs based on data from a prospective candidaemia study. All Candida albicans candidaemia episodes in patients enrolled in the Australian Candidaemia Studyand who were treated with fluconazole monotherapy were included. Fluconazole MICs were established using Sensititre(®) YeastOne(®). Two hundred and seventeen evaluable episodes were identified, 93.5% of which occurred in adult patients. Fluconazole was commenced within 72 h of blood culture positivity in 96.3% (209/217) of episodes. Fluconazole doses were appropriate in 89.9% (195/217) of episodes and the median duration of therapy was 14 days (IQR 8-21 days) for the whole cohort. The all-cause 30 day mortality was 19.8% (43/217), with 37.2% (16/43) of deaths attributed to candidaemia. Classification and regression tree (CART) analysis identified a fluconazole MIC target of ≥2 mg/L for infection-related mortality and ≥4 mg/L for overall 30 day mortality. Overall mortality was no different in episodes with isolates above or below the identified MIC target, although there was a trend towards significance (P = 0.051). On univariate analysis, infection-related mortality was significantly increased in C. albicans episodes with an MIC ≥2 mg/L compared with those below this MIC target (20.6% versus 4.9%; P = 0.001). This target remained an independent predictor of infection-related mortality (OR 8.2; 95% CI 2.3-29.7; P = 0.001). We observed a direct relationship between infection-related mortality and rising fluconazole MIC for C. albicans candidaemia; overall, the data support the EUCAST and revised CLSI fluconazole clinical breakpoints.

Fungemia in a Spanish hospital: the role of Candida parapsilosis over a 15-year period.

Candida parapsilosis is one of the main causes of fungemia in tertiary-care hospitals. Few studies have analysed the changes in its distribution over a long period. We compared the distribution of C. parapsilosis with that of other fungi over a 15-y period in a tertiary hospital. The susceptibility of C. parapsilosis was analysed using the new species-specific clinical breakpoints. The C. parapsilosis complex species were differentiated molecularly. From January 1997 to December 2011, 360 isolates causing 350 episodes of fungemia were isolated. C. parapsilosis was the second most frequently isolated species (20%); only 1 C. orthopsilosis was identified and there were no C. metapsilosis. The remaining episodes were caused by C. albicans (43.1%), C. tropicalis (14.4%), C. glabrata (11.7%), and other fungal species (10.8%). The incidence of candidemia increased more than two-fold between 2009 and 2011 (from 3.3 to 7.4 cases/100,000 population), and C. parapsilosis and C. glabrata fungemia increased throughout the period. C. parapsilosis was the most frequent species in children under 15 y (57.1%). All C. parapsilosis isolates were susceptible to anidulafungin, micafungin, flucytosine, amphotericin B, and posaconazole, while 98.5% were susceptible to caspofungin, 97.1% to voriconazole, 95.6% to fluconazole, and 76.5% to itraconazole. This long-term study showed a slight increase in the incidence of candidemia during the years of the study and a trend towards an increase in C. parapsilosis. Because of its high frequency and intrinsic low susceptibility to echinocandins, the prevalence and susceptibility of C. parapsilosis should be monitored, especially in children.

Multicenter Evaluation of the New Vitek 2 Yeast Susceptibility Test Using New CLSI Clinical Breakpoints for Fluconazole

A fully automated antifungal susceptibility test system recently updated to reflect the new species-specific clinical breakpoints (CBPs) of fluconazole for Candida (Vitek 2 AF03 yeast susceptibility test; bioMérieux, Inc., Durham, NC) was compared in three different laboratories with the Clinical and Laboratory Standards Institute (CLSI) reference broth microdilution (BMD) method by testing 2 quality control strains, 10 reproducibility strains (4 Candida species and 6 Cryptococcus neoformans strains), and 746 isolates of Candida species (702 isolates, 13 species) and 44 isolates of C. neoformans against fluconazole. Excellent essential agreement (EA) (within 2 dilutions) between the reference and Vitek 2 MICs was observed for fluconazole and Candida species (94.0%). The EA was lower for fluconazole and C. neoformans at 86.4%. The mean times to a result with the Vitek 2 test were 9.1 h for Candida species and 12.1 h for C. neoformans. Categorical agreement (CA) between the two methods was assessed by using the new species-specific CBPs. For less common species without fluconazole CBPs, the epidemiological cutoff values (ECVs) were used to differentiate wild-type (WT; MIC, ≤ ECV) from non-WT (MIC, >ECV) strains. The CAs between the two methods were 92.0% for Candida species (0.3% very major errors [VME] and 2.6% major errors [ME]) and 84.1% for C. neoformans (4.5% VME and 11.4% ME). The updated Vitek 2 AF03 IUO yeast susceptibility system is comparable to the CLSI BMD reference method for testing the susceptibility of clinically important yeasts to fluconazole when using the new (lower) CBPs and ECVs.

Nosocomial bloodstream infections due to Candida spp. in the USA: species distribution, clinical features and antifungal susceptibilities

Candida spp. are among the most frequent nosocomial pathogens, contributing significantly to morbidity and mortality. Longitudinal data on the epidemiology of Candida bloodstream infections (BSIs) are still limited. Isolates and clinical data from 1218 episodes of Candida BSI were prospectively collected from patients in 52 hospitals in the USA between 1998 and 2006. Susceptibilities to amphotericin B, flucytosine, fluconazole, posaconazole, voriconazole, anidulafungin, caspofungin and micafungin were determined for 1077 Candida isolates by the CLSI reference broth microdilution method using the recently published species-specific clinical breakpoints. Candida albicans was the most prevalent species (50.7%), followed by Candida parapsilosis (17.4%), Candida glabrata (16.7%) and Candida tropicalis (10.2%). The prevalence of non-albicans Candida spp. increased over time. Patients had a mean age of 51 years and a mean length of hospital stay prior to BSI of 22 days. The main underlying conditions were gastrointestinal (20.1%) and pulmonary (13.0%) diseases. Intravenous catheters (19.1%) and the urinary tract (8.0%) were the most frequently determined likely sources, whilst in the majority of patients (61.1%) no source could be identified. Overall mortality was 38.1%. Of the isolates studied, 0.8% of C. albicans, 100.0% of C. glabrata, 2.9% of C. parapsilosis and 4.9% of C. tropicalis were non-susceptible to fluconazole, and 0.6% of C. albicans, 5.0% of Candida krusei, 7.6% of C. parapsilosis and 9.8% of C. tropicalis were non-susceptible to voriconazole. All echinocandins showed good activity against most Candida spp., including the majority of C. parapsilosis isolates, but only 38.1% of C. glabrata tested susceptible to caspofungin.

Species distribution and antifungal susceptibility of bloodstream fungal isolates in paediatric patients in Mexico: a nationwide surveillance study

To establish the species distribution and in vitro susceptibilities of 358 bloodstream fungal isolates from paediatric patients in Mexico. Isolates were collected during a 2 year surveillance programme in 14 medical centres in 10 Mexican states. A molecular approach was used to determine the Candida parapsilosis species complex. In vitro susceptibility to amphotericin B, fluconazole, voriconazole, itraconazole, posaconazole, caspofungin, anidulafungin and micafungin was determined according to CLSI procedures. Species-specific clinical breakpoints for fluconazole, voriconazole and echinocandins were applied. Candida spp. accounted for 98.33% of fungaemias, including 127 Candida albicans isolates, 127 C. parapsilosis complex isolates (121 C. parapsilosis sensu stricto, 4 Candida orthopsilosis and 2 Candida metapsilosis strains) and 72 Candida tropicalis isolates. C. albicans and C. parapsilosis complex were the species predominant in neonates (48 cases each; 41.02%). C. parapsilosis complex was also the predominant species in patients 1 month to <2 years of age (P = 0.007). In contrast, C. albicans was the most frequent species in patients aged 2 to <12 years (P = 0.003). Antifungal resistance was rare among the subset of isolates. Candida glabrata showed the highest resistance rate to amphotericin B (1/9 isolates), fluconazole (1/9 isolates) and itraconazole (2/9 isolates). The species distribution differed with the age of the patients, with C. albicans and C. parapsilosis complex being the most commonly isolated species. C. glabrata showed the highest resistance rate to amphotericin B, fluconazole and itraconazole. This is the first study of fungaemia episodes in Mexican children.

Comparison of the Vitek 2 yeast susceptibility system with CLSI microdilution for antifungal susceptibility testing of fluconazole and voriconazole against Candida spp., using new clinical breakpoints and epidemiological cutoff values

A commercially available, fully automated yeast susceptibility test system (Vitek 2; bioMérieux, Marcy d’Etoile, France) was compared in 3 different laboratories with the Clinical and Laboratory Standards Institute (CLSI) reference microdilution (BMD) method by testing 2 quality control strains, 10 reproducibility strains, and 425 isolates of Candida spp. against fluconazole and voriconazole. Reference CLSI BMD MIC endpoints and Vitek 2 MIC endpoints were read after 24 hours and 9.1–27.1 hours incubation, respectively. Excellent essential agreement (within 2 dilutions) between the reference and Vitek 2 MICs was observed for fluconazole (97.9%) and voriconazole (96.7%). Categorical agreement (CA) between the 2 methods was assessed using the new species-specific clinical breakpoints (CBPs): susceptible (S) ≤2 μg/mL, susceptible dose-dependent (SDD) 4 μg/mL, and resistant (R) ≥8 μg/mL for fluconazole and Candida albicans, Candida tropicalis, and Candida parapsilosis and ≤32 μg/mL (SDD), ≥64 μg/mL (R) for Candida glabrata; S ≤0.12 μg/mL, SDD 0.25–0.5 μg/mL, R ≥1 μg/mL for voriconazole and C. albicans, C. tropicalis, and C. parapsilosis, and ≤0.5 μg/mL (S), 1 μg/mL (SDD), ≥2 μg/mL (R) for Candida krusei. The epidemiological cutoff value (ECV) of 0.5 μg/mL for voriconazole and C. glabrata was used to differentiate wild-type (WT; MIC ≤ ECV) from non-WT (MIC > ECV) strains of this species. Due to the lack of CBPs for the less common species, the ECVs for fluconazole and voriconazole, respectively, were used for Candida lusitaniae (2 μg/mL and 0.03 μg/mL), Candida dubliniensis (0.5 μg/mL and 0.03 μg/mL), Candida guilliermondii (8 μg/mL and 0.25 μg/mL), and Candida pelliculosa (4 μg/mL and 0.25 μg/mL) to categorize isolates of these species as WT and non-WT. CA between the 2 methods was 96.8% for fluconazole and 96.5% for voriconazole with less than 1% very major errors and 1.3–3.0% major errors. The Vitek 2 yeast susceptibility system remains comparable to the CLSI BMD reference method for testing the susceptibility of Candida spp. when using the new (lower) CBPs and ECVs.

Species distribution and drug susceptibilities of Candida isolates in TSARY 2010

Susceptibilities to antifungal drugs of 1083 Candida isolates collected in Taiwan Surveillance of Antimicrobial Resistance of Yeasts in 2010 were determined. There were 422 (39%) C. albicans, 270 (24.9%) C. tropicalis, 258 (23.8%) C. glabrata, 87 (8%) C. parapsilosis, 18 (1.7%) C. krusei, and 28 (2.6%) of 13 other species. In the present study, we have applied species-specific clinical breakpoints for common species and epidemiological cutoff values for rare species. We found that majority of isolates were susceptible to tested drugs. A total of 15, 3, 2, and 0 isolates were not susceptible to fluconazole, voriconazole, amphotericin B, and anidulafungin, respectively. We found that three of the four fluconazole non-susceptible C. albicans isolates were resistant to voriconazole. Hence, there is an issue of cross-resistance among azole-type drugs.

Comparison of the Sensititre YeastOne colorimetric antifungal panel with CLSI microdilution for antifungal susceptibility testing of the echinocandins against Candida spp., using new clinical breakpoints and epidemiological cutoff values

A commercially prepared dried colorimetric microdilution panel (Sensititre Yeast One, TREK Diagnostic Systems, Cleveland, OH, USA) was compared in 3 different laboratories with the Clinical and Laboratory Standards Institute (CLSI) reference microdilution method by testing 2 quality control strains, 25 reproducibility strains, and 404 isolates of Candida spp. against anidulafungin, caspofungin, and micafungin. Reference CLSI BMD MIC end points and YeastOne colorimetric end points were read after 24 h of incubation. Excellent (100%) essential agreement (within 2 dilutions) between the reference and colorimetric MICs was observed. Categorical agreement (CA) between the 2 methods was assessed using the new species-specific clinical breakpoints (CBPs): susceptible (S), ≤0.25 μg/mL; intermediate (I), 0.5 μg/mL; and resistant (R), ≥1 μg/mL, for C. albicans, C. tropicalis, and C. krusei, and ≤2 μg/mL (S), 4 μg/mL (I), and ≥8 μg/mL (R) for C. parapsilosis and all 3 echinocandins. The new CBPs for anidulafungin and caspofungin and C. glabrata are ≤0.12 μg/mL (S), 0.25 μg/mL (I), and ≥0.5 μg/mL (R), whereas those for micafungin are ≤0.06 μg/mL (S), 0.12 μg/mL (I), and ≥0.25 μg/mL (R). Due to the lack of CBPs for any of the echinocandins and C. lusitaniae, the epidemiological cutoff values (ECVs) were used for this species to categorize the isolates as wild-type (WT; MIC ≤ECV) and non-WT (MIC >ECV), respectively, for anidulafungin (≤2 μg/mL/>2 μg/mL), caspofungin (≤1 μg/mL/>1 μg/mL), and micafungin (≤0.5 μg/mL/>0.5 μg/mL). CA ranged from 93.6% (caspofungin) to 99.6% (micafungin) with less than 1% very major or major errors. The YeastOne colorimetric method remains comparable to the CLSI BMD reference method for testing the susceptibility of Candida spp. to the echinocandins when using the new (lower) CBPs and ECVs. Further study using defined fks mutant strains of Candida is warranted.