Biology subjects

Harrison, T. S.

Publications and source records attributed to Harrison, T. S..

6 recordsLinked to original sources

bioRxiv2024

A composite subunit vaccine confers full protection against Buruli ulcer disease in the mouse footpad model of Mycobacterium ulcerans infection

Buruli ulcer (BU) disease, a neglected necrotizing tropical skin infection caused by Mycobacterium ulcerans, is the third most common mycobacterial disease after tuberculosis and leprosy. Infections mostly occur in remote, rural areas of Central and West Africa, but also in Australia, Japan and Papua New Guinea. There is currently no vaccine against Buruli ulcer disease and all previous attempts using closely related bacteria and subunit proteins have been partially successful only. Here, we tested in mice a composite subunit formulation incorporating the Mycobacterium ulcerans toxin mycolactone as the immunomodulator, and the antigens Ag85A and Polyketide Synthase Enzyme Ketoreductase A (KRA), formulated with Quil-A adjuvant ( Burulivac). Burulivac induced Ag85A and KRA antigen-specific antibodies, T cells and a mixed pro- and anti-inflammatory cytokine responses, which conferred absolute protection against Buruli ulcer disease in the mouse footpad model over a 14-week period of observation. This was superior to both live attenuated mycobacterial vaccines, that is, BCG and an avirulent M. ulcerans strain that lacks the mycolactone toxin (Mu{Delta}). Interleukin 10 was found to be strongly associated with protection. We suggest that Burulivac is a promising vaccine candidate against Buruli ulcer disease that warrants further exploration. Author summaryBuruli ulcer is a neglected tropical disease caused by skin infections by Mycobacterium ulcerans, an organism related to causative pathogens of tuberculosis and leprosy. The disease is endemic to parts of Central and West Africa but is also found in Australia, Japan and some other countries. While mortality is low, morbidity caused by extensive and unsightly ulceration of the skin is high. There currently is no vaccine against Buruli ulcer and all previous attempts to develop one have been unsuccessful. Here we developed a vaccine formulation termed Burulivac that incorporates three M. ulcerans antigens, two of which are proteins and the third the mycolactone toxin, much responsible for the pathology of the BU disease. Our rationale was that in a vaccine setting and using a controlled dose of the toxin, we could calibrate the immune responses to selected antigens, so that they mimic those induced during natural infection. Using an experimental mouse footpad model of infection, we demonstrate that immunisation with Burulivac completely prevented ulceration in the mouse footpad over an extended 14-weeks period of observation, and that this correlated with elevated levels of anti-inflammatory cytokine Interleukin-10. We propose that Burulivac is a promising vaccine candidate that merits further testing and development.

immunology↗

bioRxiv2024

In vitro host transcriptomics during Cryptococcus neoformans, Cryptococcus gattii, and Candida albicans infection of South African volunteers

Cryptococcus neoformans, Cryptococcus gattii and Candida albicans are opportunistic fungal pathogens associated with infections in immunocompromised hosts. Cryptococcal meningitis (CM) is the leading fungal cause of HIV-related deaths globally, with the majority occurring in Africa. The human immune response to C. albicans infection has been studied extensively in large genomics studies whereas cryptococcal infections, despite their severity, are comparatively understudied. Here we investigated the transcriptional response of immune cells after in vitro stimulation with in vitro C. neoformans, C. gattii and C. albicans infection of peripheral blood mononuclear cells (PBMCs) collected from healthy South African volunteers. We found a lower transcriptional response to cryptococcal stimuli compared to C. albicans and unique expression signatures from all three fungal stimuli. This work provides a starting point for further studies comparing the transcriptional signature of CM in immunocompromised patients, with the goal of identifying biomarkers of disease severity and possible novel treatment targets.

genomics↗

bioRxiv2018

Transcriptional Profiling of Patient Isolates Identifies a Novel TOR Regulatory Pathway in Cryptococcal Virulence

Human infection with Cryptococcus causes up to a quarter million AIDS-related deaths annually and is the most common cause of non-viral meningitis in the United States. As an opportunistic fungal pathogen, C. neoformans is distinguished by its ability to adapt to diverse host environments including plants, amoeba and mammals. In the present study, comparative transcriptomics of the fungus within human cerebrospinal fluid identified expression profiles representative of low-nutrient adaptive responses. Transcriptomics of fungal isolates from a cohort of HIV/AIDS patients identified a low nutrient-induced gene, an alternative carbon nutrient transporter STL1 associated with poor early fungicidal activity, an important clinical prognostic marker. Mouse modeling and pathway analysis demonstrated a role for STL1 in mammalian pathogenesis and revealed that STL1 expression is regulated by a novel target-of-rapamycin (TOR)-related multi-gene regulatory mechanism involving the CAC2 subunit of the chromatin assembly complex 1, CAF-1. In this pathway, the TOR-related RNA chaperone, VAD1 was found to transcriptionally regulate a cryptococcal homolog of a cytosolic protein Ecm15, in turn, required for nuclear transport of the Cac2 protein. Derepression of STL1 by the CAC2-containing CAF-1 complex was mediated by Cac2 and modulated binding and suppression of the STL1 enhancer element. Derepression of STL1 resulted in enhanced survival and growth of the fungus in the presence of low nutrient, alternative carbon sources, facilitating virulence in mice. The study underscores the utility of ex vivo expression profiling of fungal clinical isolates and provides fundamental genetic understanding of saprophyte adaption to the human host.\n\nAuthor summaryThe fungus Cryptococcus is a fungal pathogen that kills an estimated quarter of a million individuals yearly and is the most common cause of meningitis in the United States. The fungus is carried in about 10% of the adult population and, after re-activation, causes disease in a wide variety of individuals including HIV-infected as well as immunosuppression either from genetic defects or after immune suppressive treatments due to transplant conditioning, cancer therapy or treatment of autoimmune diseases. The fungus is widely carried in the soil and trees and can infect plants, single cell organisms and even dolphins. However, mechanisms for this widespread ability to infect a variety of hosts are poorly understood. The present study identified adaptation to low nutrients as a key property that allows the fungus to infect these diverse hosts and identified a nutrient transporter, STL1 to be associated with a marker of poor clinical outcome in a cohort of HIV/AIDS patients. Understanding molecular mechanisms involved in environmental adaptation may help to design better methods of control and treatment of widely dispersed fungal pathogens such as Cryptococcus.

microbiology↗

bioRxiv2017

The Cryptococcus neoformans Titan cell is an inducible and regulated morphotype underlying pathogenesis

Fungi undergo changes in cell shape in response to environmental stimuli that drive pathogenesis and niche adaptation, such as the yeast-to-hyphal transition of dimorphic fungi in response to changing temperature. The basidiomycete Cryptococcus neoformans undergoes an unusual morphogenetic transition in the host lung from haploid yeast to large, highly polyploid cells termed Titan cells. Titan cells influence fungal interaction with host cells, including through increased drug resistance, altered cell size, and altered Pathogen Associated Molecular Pattern exposure. Despite the important role these cells play in pathogenesis, understanding the environmental stimuli that drive the morphological transition, and the molecular mechanisms underlying their unique biology, has been hampered by the lack of a reproducible in vitro induction system. Here we demonstrate reproducible in vitro Titan cell induction in response to environmental stimuli consistent with the host lung. In vitro Titan cells exhibit all the properties of in vivo generated Titan cells, the current gold standard, including altered capsule, cell wall, size, high mother cell ploidy, and aneuploid progeny. We identify bacterial peptidoglycan as a serum compound associated with shift in cell size and ploidy, and demonstrate the capacity of bronchial lavage fluid and E. coli co-culture to induce Titanisation. Additionally, we demonstrate the capacity of our assay to identify established and previously undescribed regulators of Titanisation in vitro and investigate the Titanisation capacity of clinical isolates and their impact on disease outcome. Together, these findings provide new insight into the environmental stimuli and molecular mechanisms underlying the yeast-to-titan transition and establish an essential in vitro model for the future characterization of this important morphotype.\n\nAuthor SummaryChanges in cell shape underlie fungal pathogenesis by allowing immune evasion and dissemination. Aspergillus and Candida albicans hyphae drive tissue penetration. Histoplasma capsulatum and C. albicans yeast growth allows evasion and dissemination. As major virulence determinates, morphogenic transitions are extensively studied in animal models and in vitro. The pathogenic fungus Cryptococcus neoformans is a budding yeast that, in the host lung, switches to an unusual morphotype termed the Titan cell. Titans are large, polyploid, have altered cell wall and capsule, and produce haploid daughters. Their size prevents engulfment by phagocytes, yet they are linked to dissemination and altered immune response. Despite their important influence on disease, replicating the yeast-to-Titan switch in vitro has proved challenging. Here we show that Titans are induced by host-relevant stimuli, including serum and bronchio-alveolar lavage fluid. We identify bacterial peptidoglycan as a relevant inducing compound and predict an in vivo Titan defect for a clinical isolate. Genes regulating in vivo Titanisation also influence in vitro formation. Titanisation is a conserved morphogenic switch across the C. neoformans species complex. Together, we show that Titan cells are a regulated morphotype analogous to the yeast-to-hyphal transition and establish new ways to study Titans outside the host lung.

microbiology↗

bioRxiv2017

Population Genomics Of Cryptococcus neoformans var. grubii Reveals New Biogeographic Relationships And Finely Maps Hybridization

Cryptococcus neoformans var. grubii is the causative agent of cryptococcal meningitis, a significant source of mortality in immunocompromised individuals, typically HIV/AIDS patients from developing countries. Despite the worldwide emergence of this ubiquitous infection, little is known about the global molecular epidemiology of this fungal pathogen. Here we sequence the genomes of 188 diverse isolates and characterized the major subdivisions, their relative diversity and the level of genetic exchange between them. While most isolates of C. neoformans var. grubii belong to one of three major lineages (VNI, VNII, and VNB), some haploid isolates show hybrid ancestry including some that appear to have recently interbred, based on the detection of large blocks of each ancestry across each chromosome. Many isolates display evidence of aneuploidy, which was detected for all chromosomes. In diploid isolates of C. neoformans var. grubii (serotype A/A) and of hybrids with C. neoformans var. neoformans (serotype A/D) such aneuploidies have resulted in loss of heterozygosity, where a chromosomal region is represented by the genotype of only one parental isolate. Phylogenetic and population genomic analyses of isolates from Brazil revealed that the previously African VNB lineage occurs naturally in the South American environment. This suggests migration of the VNB lineage between Africa and South America prior to its diversification, supported by finding ancestral recombination events between isolates from different lineages and regions. The results provide evidence of substantial population structure, with all lineages showing multi-continental distributions demonstrating the highly dispersive nature of this pathogen.\n\nAuthor SummaryCryptococcus neoformans var. grubii is a human fungal pathogen of immunocompromised individuals that has global clinical impact, causing half a million deaths per year. Substantial genetic substructure exists for this pathogen, with two lineages found globally (VNI, VNII) whereas a third has appeared confined to sub-Saharan Africa (VNB). Here, we utilized genome sequencing of a large set of global isolates to examine the genetic diversity, hybridization, and biogeography of these lineages. We found that while the three major lineages are well separated, recombination between the lineages has occurred, notably resulting in hybrid isolates with segmented ancestry across the genome. In addition, we showed that isolates from South America are placed within the VNB lineage, formerly thought to be confined to Africa, and that there is phylogenetic separation between these geographies that substantially expands the diversity of these lineages. Our findings provide a new framework for further studies of the dynamics of natural populations of C. neoformans var. grubii.

genomics↗

bioRxiv2016

A population genomics approach to assessing the genetic basis of within-host microevolution underlying recurrent cryptococcal meningitis infection

Recurrence of meningitis due to Cryptococcus neoformans after treatment causes substantial mortality in HIV/AIDS patients across sub-Saharan Africa. In order to determine whether recurrence occurred due to relapse of the original infecting isolate or reinfection with a different isolate weeks or months after initial treatment, we used whole-genome sequencing to assess the genetic basis of infection in 17 HIV-infected individuals with recurrent cryptococcal meningitis. Comparisons revealed a clonal relationship for 15 pairs of isolates recovered before and after recurrence showing relapse of the original infection. The two remaining pairs showed high levels of genetic heterogeneity; in one pair we found this to be a result of infection by mixed genotypes, whilst the second was a result of nonsense mutations in the gene encoding the DNA mismatch repair proteins MSH2, MSH5 and RAD5. These nonsense mutations led to a hypermutator state, leading to dramatically elevated rates of synonymous and non-synonymous substitutions. Hypermutator phenotypes owing to nonsense mutations in these genes have not previously been reported in Cryptococcus neoformans and represent a novel pathway for rapid within-host adaptation and evolution of resistance to firstline antifungal drugs.

genomics↗