The in vitro activity of the novel fungal Cyp51 inhibitor VT-1129 was evaluated against a large panel of C. neoformans and C. gattii isolates. VT-1129 demonstrated potent activity against both Cryptococcus species as demonstrated by low MIC50 and MIC90 values. Against C. gattii, the in vitro potency was maintained against all genotypes. In addition, significantly lower geometric mean MICs were observed for VT-1129 compared to fluconazole against C. neoformans, including isolates with reduced fluconazole susceptibility.

MIC values were generated for caspofungin, micafungin, and anidulafungin against 106 isolates of C. lusitaniae, and these values were compared to established epidemiologic cutoff values. The majority of isolates were wild type both by MIC value as well as by FKS1 hotspot sequencing. Although C. lusitaniae isolates have MIC values to the echinocandins that are elevated compared to other common species, with regard to known mechanisms of resistance to the echinocandins, isolates with MIC values at or below the epidemiological cutoff values of 0.5 and 1mug/mL for micafungin and anidulafungin, respectively, should be considered wild type.

Candida glabrata is the second leading cause of candidemia in US hospitals. Current guidelines suggest that an echinocandin be used as primary therapy for C. glabrata due to the high rate of resistance to fluconazole. Recent case reports indicate that C. glabrata resistance to echinocandins may be increasing. We performed susceptibility testing on 1380 isolates of C. glabrata collected between 2008 and 2013 from four US cities, Atlanta, Baltimore, Knoxville and Portland. Our analysis showed that 3.1%, 3.3%, and 3.6% of the isolates were resistant to anidulafungin, caspofungin and micafungin, respectively. We screened 1032 of these isolates, including all 77 that had either a resistant or intermediate MIC value to one or more echinocandin, for mutations in the hotspot regions of FKS1 and FKS2, the major mechanism of echinocandin resistance. Fifty-one isolates were identified with hotspot mutations, 16 in FKS1 and 35 in FKS2. All but one of the isolates with an FKS mutation were resistant to one or more echinocandin by susceptibility testing. Of the isolates resistant to one or more echinocandin, 36% were also resistant to fluconazole. Echinocandin resistance among US C. glabrata isolates is a concern especially in light of the fact that a third of those isolates may be multidrug resistant. Further monitoring of US C. glabrata isolates for echinocandin resistance is warranted.

Echinocandins are the recommended treatment for invasive candidiasis due to Candida glabrata. Resistance to echinocandins is known to be caused by non-synonymous mutations in the hotspot-1 (HS1) regions of the FKS1 and FKS2 genes which encode a subunit of the beta-1,3-glucan synthase, the target of the echinocandins. Here, we describe the development of a microsphere-based assay using Luminex MagPix technology to identify mutations in the FKS1 and FKS2 HS1 domains which confer in vitro echinocandin resistance in C. glabrata isolates. The assay is rapid and can be performed with high throughput. The assay was validated using 102 isolates that had FKS1-HS1 and FKS2-HS1 domains previously characterized by DNA sequencing. The assay was 100% concordant with DNA sequencing results. The assay was then used for high throughput screening of 1032 C. glabrata surveillance isolates. Sixteen new isolates with mutations were identified, as well as a mutation new to our collection, del659F. This assay provides a rapid and cost-effective way to screen C. glabrata isolates for echinocandin resistance.

Between 2008 and 2011, population-based candidemia surveillance was conducted in Atlanta, GA and Baltimore, MD. Surveillance had been previously performed in Atlanta in 1992-1993 and in Baltimore in 1998-2000, making this the first population-based candidemia surveillance conducted over multiple time points in the US. From 2,675 identified cases of candidemia in the current surveillance, 2,329 Candida isolates were collected. Candida albicans no longer comprised the majority of isolates but remained the most frequently isolated species (38%), followed by C. glabrata (29%), C. parapsilosis (17%) and C. tropicalis (10%). The species distribution has changed over time; in both Atlanta and Baltimore the proportion of C. albicans decreased and the proportion of C. glabrata increased, while the proportion of C. parapsilosis increased in Baltimore only. There were 98 multi-species episodes, with C. albicans and C. glabrata the most frequently encountered combination. The new species-specific CLSI Candida MIC breakpoints were applied to these data. With the exception of C. glabrata (11.9% resistant), resistance to fluconazole was very low (2.3% for C. albicans, 6.2% for C.tropicalis and 4.1% for C. parapsilosis). There was no change in the proportion of fluconazole resistance between surveillance periods. Overall echinocandin resistance was low (1%) but was higher for C. glabrata isolates, ranging from 2.1% resistant to caspofungin in Baltimore to 3.1% resistant to anidulafungin in Atlanta. Given the increase at both sites and the higher echinocandin resistance, C. glabrata should be closely monitored in future surveillance.

Cryptococcus gattii causes infection in tropical and subtropical regions worldwide but has garnered increased attention since its 1999 emergence in North America. C. gattii can be divided into 4 molecular types that may represent cryptic species. Recent evidence has shown that azole antifungal MIC values differ among these molecular types. We tested a large collection of C. gattii isolates for susceptibility to 4 azole drugs. We found that isolates of molecular type VGII have the highest geometric mean (GM) fluconazole MIC values (8.6 mcg/mL), while isolates of molecular type VGI have the lowest (1.7 mcg/mL). For fluconazole, itraconazole, and voriconazole GM MIC values, VGI < VGIII < VGIV < VGII. The GM MIC values for posaconazole were similarly represented across molecular types, with the exception that VGII < VGIII and VGIV. We used the MIC values to establish preliminary epidemiologic cutoff values for each azole and molecular type of C. gattii.

Flucytosine and itraconazole are the only antifungal agents that continue to have a Clinical Laboratory and Standards Institute recommendation for MIC breakpoint readings at 48h only. Here we show good essential and categorical agreement between MIC readings for flucytosine made at 48h and those made at 24h for Candida species.