Extensive Diversity and Prevalent Fluconazole Resistance among Environmental Yeasts from Tropical China

Abstract

Yeasts play important roles in both the environment and in human welfare. While some environmental yeasts positively contribute to nutrient cycling and food production, a significant number of yeast species are opportunistic human pathogens, including several that are tolerant/resistant to commonly used antifungal drugs. At present, most of our understanding of environmental yeasts has come from a few terrestrial environments in selected geographic regions. Relatively little is known about yeast diversity in tropical environments and their potential impacts on human health. Here, we characterize culturable yeasts in 968 environmental samples from eight regions in tropical China. Among the 516 soil, 273 freshwater, and 179 seawater samples, 71.5%, 85.7%, and 43.6% contained yeasts, respectively. A total of 984 yeast isolates were analyzed for their DNA barcode sequences and their susceptibilities to fluconazole. DNA sequence comparisons revealed that the 984 yeast isolates likely belonged to 144 species, including 106 known species and 38 putative novel species. About 38% of the 984 isolates belonged to known human pathogens and the most common species was Candida tropicalis, accounting for 21% (207/984) of all isolates. Further analyses based on multi-locus sequence typing revealed that some of these environmental C. tropicalis shared identical genotypes with clinical isolates previously reported from tropical China and elsewhere. Importantly, 374 of the 984 (38%) yeast isolates showed intermediate susceptibility or resistance to fluconazole. Our results suggest that these environmental yeasts could have significant negative impacts on human health.

Keywords: Candida; Candida tropicalis; Rhodotorula; environment yeast; fluconazole-resistance.

Figure 1

Geographic map of Hainan province in tropical China showing sites where environmental samples were collected for this study. The eight municipalities where samples were collected are marked by red stars.

Figure 2

Diversity and abundance of culturable environmental yeasts within each of the eight geographic samples. Labels on the X-axis represent the eight geographic regions. The yeast diversity of each geographic region is represented by a stacked bar plot. In the stacked bars, each color represents a unique species, and the height of the colored sections indicates the abundance of that species in the population. For detailed prevalence of each species at each region, see Table S1.

Figure 3

Maximum likelihood tree showing the relationships between ten putative novel Candida species and their closely related known species based on rDNA ITS sequences. The geographic origins of these putative novel species are indicated as two-letter abbreviations, where BT = Baoting, DZ = Danzhou, and WC = Wenchang. The tree was constructed using RaxML with 1000 bootstraps.

Figure 4

Maximum likelihood tree showing the relationships between nine putative novel Rhodotorula species and their closely related known species based on rDNA ITS sequences. The geographic origins of these putative novel species in our sample are indicated as two-letter abbreviations, where HK = Haikou, DZ = Danzhou, WC = Wenchang, and DF = Dongfang. The tree was constructed using RaxML with 1000 bootstraps.

Figure 5

Prevalence of opportunistic yeast pathogens in environmental samples from eight municipalities in Hainan province in tropical China. Highlighted in red are the number of strains of pathogenic yeast species, and those in various shades of gray are not known to be pathogenic to humans.

Figure 6

Relationships among strains of C. tropicalis from environmental and clinical sources in Hainan province in tropical China. Strains highlighted in red were from natural environments isolated in this study. The non-highlighted ones were from oral cavities of patients in Hainan reported in our previous study. For detailed strain information, see Ref. [22] and Table S4.

Figure 6

Relationships among strains of C. tropicalis from environmental and clinical sources in Hainan province in tropical China. Strains highlighted in red were from natural environments isolated in this study. The non-highlighted ones were from oral cavities of patients in Hainan reported in our previous study. For detailed strain information, see Ref. [22] and Table S4.

Figure 6

Relationships among strains of C. tropicalis from environmental and clinical sources in Hainan province in tropical China. Strains highlighted in red were from natural environments isolated in this study. The non-highlighted ones were from oral cavities of patients in Hainan reported in our previous study. For detailed strain information, see Ref. [22] and Table S4.

Figure 6

Relationships among strains of C. tropicalis from environmental and clinical sources in Hainan province in tropical China. Strains highlighted in red were from natural environments isolated in this study. The non-highlighted ones were from oral cavities of patients in Hainan reported in our previous study. For detailed strain information, see Ref. [22] and Table S4.