Biology subjects

Farrer, R. A.

Publications and source records attributed to Farrer, R. A..

5 recordsLinked to original sources

bioRxiv2018

Transcriptional heterogeneity of Cryptococcus gattii VGII compared with non-VGII lineages underpins key pathogenicity pathways

Cryptococcus gattii is a pathogenic yeast of humans and other animals, which causes disease predominantly in immunocompetent hosts. Infection begins when aerosolized yeast or spores enter the body, triggering an immune response, including engulfment by macrophages. To understand the early transcriptional signals in both the yeast and its mammalian host, we performed a time-course dual RNA-seq experiment for four lineages of C. gattii (VGI-IV) interacting with mouse macrophages at 1hr, 3hr and 6hr post infection. Comparison of in vitro to ex vivo gene expression indicates lineage VGII is transcriptionally divergent to non-VGII lineages, including differential expression of genes involved in capsule synthesis, capsule attachment and ergosterol production. Various paralogs demonstrate sub-functionalisation between lineages including an upregulation of capsule biosynthesis-related gene CAP2, and downregulation of CAP1 in VGIII. Isolates also compensate for lineage-specific gene-losses by over-expression of genetically similar paralogs, including an over-expression of capsule gene CAS3 in VGIV having lost CAS31. Differential expression of one in five C. gattii genes was detected following co-incubation with mouse macrophages; all isolates showed high induction of oxidative-reduction functions and a downregulation of capsule attachment genes. We also show that VGII switches expression of two laccase paralogs (from LAC1 to LAC2) during co-incubation of macrophages. Finally, we found that mouse macrophages respond to all four lineages of C. gattii by upregulating FosB/Jun/Egr1 regulatory proteins at early time points. This study highlights the evolutionary breadth of expression profiles amongst the lineages of C. gattii and the diversity of transcriptional responses at this host-pathogen interface.\n\nImportanceThe transcriptional profiles of related pathogens and their response to host induced stresses underpin their pathogenicity. Expression differences between related pathogens during host interaction can indicate when and how these genes contribute to virulence, ultimately informing new and improved treatment strategies for those diseases. In this paper, we compare the transcriptional profiles of five isolates representing four lineages of C. gattii in rich media. Our analyses identified key processes including cell capsule, ergosterol production and melanin that are differentially expressed between lineages, and we find that VGII has the most distinct profile in terms of numbers of differentially expressed genes. All lineages have also undergone sub-functionalisation for various paralogs including capsule biosynthesis and attachment genes. Most genes appeared down-regulated during co-incubation with macrophages, with the largest decrease observed for capsule attachment genes, which appears coordinated with a stress response, as all lineages also upregulated oxidative stress response genes. Furthermore, VGII upregulated many genes that are linked to ergosterol biosynthesis and switched expression of the laccase LAC1 to LAC2 ex vivo. Finally, we saw a pronounced increase in the FosB/Jun/Egr1 regulatory proteins at early time points in bone marrow derived macrophages, marking a role in the host response to C. gattii. This work highlights the dynamic roles of key C. gattii virulence genes in response to macrophages.

genomics↗

bioRxiv2018

Genomic basis of multidrug-resistance, mating, and virulence in Candida auris and related emerging species

Candida auris is an emergent fungal pathogen of rising public health concern due to increasing reports of outbreaks in healthcare settings and resistance to multiple classes of antifungal drugs. While distantly related to the more common pathogens C. albicans and C. glabrata, C. auris is closely related to three rarely observed and often multidrug-resistant species, C. haemulonii, C. duobushaemulonii and C. pseudohaemulonii. Here, we generated and analyzed near complete genome assemblies and RNA-Seq-guided gene predictions for isolates from each of the four major C. auris clades and for C. haemulonii, C. duobushaemulonii and C. pseudohaemulonii. Our analyses mapped seven chromosomes and revealed chromosomal rearrangements between C. auris clades and related species. We found conservation of genes involved in mating and meiosis and identified both MTLa and MTL C. auris isolates, suggesting the potential for mating between clades. Gene conservation analysis highlighted that many genes linked to drug resistance and virulence in other pathogenic Candida species are conserved in C. auris and related species including expanded families of transporters and lipases, as well as mutations and copy number variants in ERG11 that confer drug resistance. In addition, we found genetic features of the emerging species that likely underlie differences in virulence and drug response between these and other Candida species, including genes involved in cell wall structure. To begin to characterize the species-specific genes important for antifungal response, we profiled the gene expression of C. auris in response to voriconazole and amphotericin B and found induction of several transporters and metabolic regulators that may play a role in drug resistance. This study provides a comprehensive view of the genomic basis of drug resistance, potential for mating, and virulence in this emerging fungal clade.

genomics↗

bioRxiv2018

A Global Analysis of Mutations Accompanying Microevolution in the Heterozygous Diploid Pathogen Candida albicans

Candida albicans is a heterozygous diploid yeast that is a commensal of the human gastrointestinal (GI) tract and a prevalent opportunistic pathogen. Here, whole-genome sequencing was performed on multiple C. albicans isolates passaged in different niches to characterize the complete spectrum of mutations arising during microevolution. We reveal that evolution during short time-scales (<600 generations) is driven by both de novo base substitutions and short-tract loss of heterozygosity (LOH) events. In contrast, large-scale chromosomal changes are relatively rare, although chromosome 7 trisomies repeatedly emerged during passaging in one GI colonization model. Both strain background and chromosomal features affected mutational patterns, with mutation rates being greatly elevated in regions adjacent to emergent LOH tracts. Mutation rates were also elevated during host infection where genomes showed strong evidence of purifying selection. These results establish the genetic events driving C. albicans evolution and that this heterozygous diploid is extensively shaped by purifying selection.

genomics↗

bioRxiv2018

Development and worldwide use of a non-lethal and minimal population-level impact protocols for the isolation of chytrids from amphibians

Parasitic chytrid fungi have emerged as a significant threat to amphibian species worldwide, necessitating the development of techniques to isolate these pathogens into sterile culture for research purposes. However, early methods of isolating chytrids from their hosts relied on killing amphibians. We modified a pre-existing protocol for isolating chytrids from infected animals to use toe clips and biopsies from toe webbing rather than euthanizing hosts, and distributed the protocol to interested researchers worldwide as part of the BiodivERsA project RACE - here called the RML protocol. In tandem, we developed a lethal procedure for isolating chytrids from tadpole mouthparts. Reviewing a database of use a decade after their inception, we find that these methods have been widely applied across at least 5 continents, 23 countries and in 62 amphibian species, and have been successfully used to isolate chytrids in remote field locations. Isolation of chytrids by the non-lethal RML protocol occured in 18% of attempts with 207 fungal isolates and three species of chytrid being recovered. Isolation of chytrids from tadpoles occured in 43% of attempts with 334 fungal isolates of one species (Batrachochytrium dendrobatidis) being recovered. Together, these methods have resulted in a significant reduction and refinement of our use of threatened amphibian species and have improved our ability to work with this important group of emerging fungal pathogens.

ecology↗

bioRxiv2017

Rapid genome sequencing for outbreak analysis of the emerging human fungal pathogen Candida auris

BackgroundCandida auris was first described in 2009, and has since caused nosocomial outbreaks, invasive infections and fungaemia across 11 countries in five continents. An outbreak of C. auris occurred in a specialised cardiothoracic London hospital between April 2015 and November 2016, which to date has been the largest outbreak reported worldwide, involving a total of 72 patients.\n\nMethodsTo understand the epidemiology of C. auris infection within this hospital, we sequenced the genomes of outbreak isolates using Oxford Nanopore Technologies and Illumina in order to type antifungal resistance alleles and to explore the outbreak within its local and global context.\n\nFindingsPhylogenomic analysis placed the UK outbreak in the India/Pakistan clade, demonstrating an Asian origin. The outbreak showed similar diversity to that of the entire clade and limited local spatiotemporal clustering was observed. One isolate displayed resistance to both echinocandins and 5-flucytosine; the former was associated with a serine to tyrosine amino acid substitution in the gene FKS1, and the latter was associated with a phenylalanine to isoleucine substitution in the gene FUR1. These mutations are novel for this pathogen.\n\nInterpretationMultiple differential episodic selection of antifungal resistant genotypes has occurred within a genetically heterogenous population across this outbreak, creating a resilient pathogen and making it difficult to define local-scale patterns of transmission as well as implementing outbreak control measures.\n\nFundingAntimicrobial Research Collaborative, Imperial College London

epidemiology↗