P477 In vitro synergy of combination therapy against antifungal-resistant Candida spp. isolated from captive bottlenose dolphins (Tursiops truncatus)
Authors: Matteucci Giorgia , Biancani Barbara , Binanti Diana 
Published: September 2022

Abstract

Poster session 1, September 21, 2022, 12:30 PM - 1:30 PM
The study aimed to evaluate antifungal resistance in Candida spp. isolated from captive bottlenose dolphins (Tursiops truncatus). Due to the need to find a therapy for symptomatic animals and to the presence of azole-resistant isolates, in vitro synergy of 3 antifungal combinations has been assayed.

A total of seven captive bottlenose dolphins were examined. Two dolphins showed mild aspecific symptoms, one was receiving nystatin due to gastric candidiasis, and the other four animals were asymptomatic. The presence of Candida spp. was investigated in fecal, gastric fluid, and blow samples from each dolphin twice.

Samples were cultured on Sabouraud Dextrose Agar and ChromAgar Candida at 30°C for 5 days. CHROMagar colonies consistent with Candida spp. were identified.

Susceptibility tests were performed according to CLSI standard with amphotericin B (AB), 5-flucytosine (FC), itraconazole (IZ), fluconazole (FZ), miconazole (MCZ), posaconazole (PZ), voriconazole (VOR), caspofungin (CAS), anidulafungin (AND), and micafungin (MF). In addition, minimum inhibitory concentration (MIC) for nystatin (NYS) and terbinafine (TER) was determined.

Based on the most common therapies in dolphins, assessment of the interaction between FZ and TER, VOR and TER, and VOR and NYS was carried out determining the fractional inhibitory concentration (FIC) index by checkerboard assay.

Out of 42 samples analyzed, 30 were positive for Candida spp. identified as C. albicans (n=21), C. tropicalis (n=6), and C. glabrata (n=3).

All the isolates showed resistance to azoles. A total of 50% of the isolates showed resistance to FC and 23.3% to AMB. No isolate showed resistance to echinocandins.

MIC values ranged from 16 to 128 µg/ml for TER (arithmetic mean 107.8 µg/ml) and from 8 to 128 µg/ml (arithmetic mean 51.8 µg/ml) for NYS.

The FIC index value for FLU and TER showed a synergistic effect on 71.4% and an additive effect on 28.6% of the tested isolates.

For VOR and TER an additive effect on 71.4% and a synergistic effect on 28.6% of the isolates were detected.

For VOR and NYS an antagonistic effect on 71.4% and an additive effect on 28.6% of the isolates were detected.

According to the results, combination therapy with FLU and TER was started. Symptoms resolved completely.

Candida spp. has been isolated from mucous membranes of free-living asymptomatic dolphins. However, a high load of Candida spp. may cause clinical signs.

As azoles are the most used antifungals in dolphins, azole-resistance is increasing. Since the tested dolphins had no recent history of azole treatment, it could be speculated that resistance might come from the environment. Thus, further studies and screening of multidrug-resistant Candida spp. in animals and their environment are needed to better understand resistance transmission in a One Health approach.

Moreover, susceptibility testing is important to select the appropriate therapy. In case of azole-resistant Candida spp., antifungals other than azoles alone or in combination should be considered.

Our study demonstrates that fluconazole exhibits in vitro synergistic antifungal activity with terbinafine against azole-resistant strains of C. albicans, C. tropicalis and C. glabrata isolated from captive bottlenose dolphins.