Genetic diversity as a major driver of phenotypic variability and virulence in local outbreak strains of the emerging pathogen Candida auris

Candida auris has emerged as a nosocomial pathogen only a decade ago, and has developed into a global health threat with a mortality rate of ~50%. C. auris is often multidrug resistant, sticks to skin and medical devices, and is difficult to eradicate from high-dependency hospital units. Consequently, national health authorities, including the Centre for Disease Control in the USA (CDC) and Public Health England (PHA) have posted alerts on C. auris.
The Centre for Genome Biology and Medicine at Aberdeen undertook the whole genome sequencing of ten C. auris isolates covering the 4 main clades; this provides a major resource for answering these key questions. We also have been gifted unrestricted access to outbreak strains from PHE and the CDC.
(1) Understanding how genetic diversity evolves in C. auris and drives the outbreak of specific genotypes around the world
Our recent genome sequencing and analysis revealed that C. auris, contrary to previous report, has a complete mating type locus (MAT; the sex determination system in fungi). To elucidate whether C. auris is capable of sexual reproduction including meiosis, we will test whether the MATa-strain VPCI479/P/13 is sexually compatible with any of the characterized MATx-strains. We have already established that the C. auris genome is haploid. Using pulsed-field gel electrophoresis we will also determine whether - in addition to the genetic diversity revealed by the whole genomic sequencing - C. auris isolates display chromosome number and size differences. Differences in karyotype between individuals of the same species introduce a reproductive barrier, but can be a driver of genetic variability. This will guide our choice of mating combinations of isolates to be tested, and will contribute to fully assembled genomes of different C. auris isolates.
(2) Characterization of the cell wall's phenotypic diversity in clinical isolates of C. auris and determining how this underpins virulence
To elucidate why this species is hypervariable in its phenotype and why various isolates show significant differences in their virulence, this project will examine the genetic and physiological variation in cell wall composition and the response of the cell wall to antibiotic stress. This is relevant to the aggregating properties of the cells, as well as to immune recognition and altered sensitivity to echinocandin antifungal drugs which this species is often resistant to. Cell adhesion is as established virulence factor of Candida species and, intriguingly, C. auris infections are passed readily from person to person and are hard to eradicate. This suggests that adherence and virulence correlate well in C. auris. This objective has significant potential for elucidating aspects of the epidemiology of C. auris infections that are not yet understood.
(3) Characterization of the genetic diversity of mult-drug resistance determinants of C. auris clinical isolates and determining how these affect virulence
C. auris has a massively expanded family of genes encoding MDR drug efflux pumps, and it is crucially important to understand how these contribute to its overall multi-drug resistance phenotype. In this project bioinformatics analysis will reveal which drug efflux factors are present in which isolates. This information will be used to select representatives of each of the 4 clades whose drug efflux factors will be subjected to realtime-PCR analysis to determine which are upregulated in response to exposure to different classes of antifungals (azoles, echinocandins, amphothericin). Those genes that are upregulated in response to antifungal exposure will be selectively oblated by CRISPR-Cas9 technology, for which we now have extensive experience for C. albicans. This project has great potential to elucidate the origin of differences in drug resistance between clinical isolates of C. auris, and ultimately will provide guidance for treating patients infected with C. auris strains