Characterisation, minimum inhibitory concentration, thermotolerance and virulence of Candida isolated from environmental plastic pollution, Scotland, 2023

At each site, plastic was collected using sterile forceps and placed into sterile ziplock bags. Candida species were recovered from replicate composite samples of each plastic type from each site using selective media. Twenty-seven colonies were isolated, and glycerol stocks prepared and frozen at -20°C.

Colony PCR was carried out with primers targeting the ITS region to identify the five common pathogenic species of Candida, i.e., C. albicans, C. glabrata, C. krusei, C. parapsilosis, and C. tropicalis. Different amplicon sizes were used to differentiate between distinct Candida species and three reference clinical pathogenic strains of C. albicans (strain SC 5314), C. glabrata (strain ATCC 2001) and C. tropicalis (strain CAY676), were included as positive controls.

To further confirm the identity of each isolate, the ITS1 region was amplified and sequenced. All purified PCR products underwent Sanger sequencing and species identification was confirmed using NCBI’s Basic Local Alignment Search Tool.

Each Candida isolate were subjected to minimum inhibitory concentration (MIC) analysis to determine antifungal resistance following the European Committee on Antimicrobial susceptibility testing (EUCAST) antifungal MIC method for yeasts. Resistance to four antifungals at ten concentrations was examined: amphotericin B (0.008 – 4 mg/L), caspofungin (0.008 – 4 mg/L), fluconazole (0.125 – 64 mg/L), voriconazole (0.008 – 4 mg/L).
To determine the thermotolerance profile, each isolate underwent an initial incubation at 18°C (simulating an environmental temperature) or 38°C (simulating human body temperature) for 24 h, before being moved to one of three different temperatures (18, 28 or 38°C) where their growth was measured. Absorbance at 570 nm was measured before and after incubation in a spectrophotometer to determine the growth of the isolates.

Galleria melonella was used as an infection model to investigate the pathogenicity of the Candida isolates. Groups of 10 larvae were injected with 10 µL of Candida cells (105 CFU/larvae) into the hemocoel via the last right pro-limb. All experiments were conducted in biological triplicate. An inoculation of 10 µL PBS was used as a control to account for mortality caused by physical injury or infection by contamination. Following injection, larvae were incubated at 37°C, and survival evaluated every 24 h for a total of 120 h. Larvae were considered dead when they did not respond to a touch stimulus.

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This dataset is included in the following collections

The survival, persistence, and ecology of microbial pathogens on the surfaces of environmental plastic pollution