1,721,074 research outputs found
Measuring the Success of HIV-1 Cure Strategies
Full text linkHIV-1 eradication strategies aim to achieve viral remission in the absence of antiretroviral therapy (ART). The development of an HIV-1 cure remains challenging due to the latent reservoir (LR): long-lived CD4 T cells that harbor transcriptionally silent HIV-1 provirus. The LR is stable despite years of suppressive ART and is the source of rebound viremia following therapy interruption. Cure strategies such as “shock and kill” aim to eliminate or reduce the LR by reversing latency, exposing the infected cells to clearance via the immune response or the viral cytopathic effect. Alternative strategies include therapeutic vaccination, which aims to prime the immune response to facilitate control of the virus in the absence of ART. Despite promising advances, these strategies have been unable to significantly reduce the LR or increase the time to viral rebound but have provided invaluable insight in the field of HIV-1 eradication. The development and assessment of an HIV-1 cure requires robust assays that can measure the LR with sufficient sensitivity to detect changes that may occur following treatment. The viral outgrowth assay (VOA) is considered the gold standard method for LR quantification due to its ability to distinguish intact and defective provirus. However, the VOA is time consuming and resource intensive, therefore several alternative assays have been developed to bridge the gap between practicality and accuracy. Whilst a cure for HIV-1 infection remains elusive, recent advances in our understanding of the LR and methods for its eradication have offered renewed hope regarding achieving ART free viral remission
Mycobacterium tuberculosis glycolipids and their effects on HIV-1 infection and replication
Full text linkHuman Immunodeficiency Virus type 1 (HIV-1) and Mycobacterium tuberculosis (Mtb) co-infection results in a rapid loss of numerous immunological functions and ultimately leads to death when not treated. The WHO estimated in 2015 that 13 million people were infected by both pathogens, with 390,000 deaths related to tuberculosis among HIV-infected individuals. Mtb and HIV-1 can induce profound changes in the host immune response resulting in Mtb infections causing active disease or the exacerbation of HIV-1 infection. For aiding in the development of new treatments a better understanding of the interactions between both pathogens and the host is crucial. This thesis follows this perspective by aiming to characterise the impact Mtb glycolipids have on modulating HIV-1 infection and to identify Mtb strains that differentially modulate HIV-1 infection and immune cell innate signalling
The Effects of HIV-1, HCV and Ebola Virus Envelope Proteins on Modulating Viral Phenotypes and Activity
Full text linkDevelopment of HIV-1 and SARS-CoV-2 Pseudo-Typed Viral Particle Systems for Studying Virus Infectivity
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Generation and Application of SARS-CoV-2 Pseudo-typed Viral Particle System to Assess the Neutralising Ability of Sera from Mild and Severe COVID-19 Study Cohorts in the First Year of the Pandemic
Full text linkSevere acute respiratory syndrome virus 2 (SARS-CoV-2) first emerged in Wuhan, China, at
the end of 2019 and quickly resulted in a global pandemic responsible for over seven million
deaths to date. A major advancement has been the development of efficient vaccines against
SARS-CoV-2, with more than 13.5 billion doses of vaccines administered worldwide.
However, without fundamental research to understand immune responses against SARS-CoV 2 infections, the rapid development and subsequent administration of vaccines would have been
halted. Specifically, quantification of the neutralising antibody (nAb) responses generated after
SARS-CoV-2 infection is crucial to fully understand long-term immunity and protection
against SARS-CoV-2. In this thesis, the nAb responses generated against SARS-CoV-2 were
studied. To assess nAb responses a lentiviral pseudo-typed viral particle (PVP) system
expressing SARS-CoV-2 spike protein was used. The first results chapter details the
optimisation of the PVP neutralisation assay and demonstrates the reliability of the assay. The
second results chapter describes the nAb responses within the sera of a cohort of hospitalised
patients recruited through the ISARIC4C consortium during the first wave of SARS-CoV-2
infections. It was observed that older and male patients experienced the most severe disease.
Strong nAb responses were produced very early on in infection, especially in patients who
experienced more severe disease but ultimately survived their infection. The third result chapter
studied the nAb responses within sera obtained from individuals who experienced mild SARS CoV-2 infection at the end of the first year of the pandemic. The nAb responses in this cohort
of individuals were much lower than the responses observed within the ISARIC4C cohort but
positively correlated with the number of symptoms experienced. Taken together, it is clear that
nAb responses generated following infection with SARS-CoV-2 range and the severity of
COVID-19 experienced plays a role in this. In the final results chapter, the use of filter paper
as a medium to study serological responses against the SARS-CoV-2 spike was investigated.
nAb responses in sera, venous blood and fingerstick blood stored on filter paper were measured
and good agreement was observed between the nAb responses measured in eluates and paired
sera. The stability of nAb responses was also observed in sera stored on filter paper at room
temperature for 28 days. The finding that filter paper can be effectively used to store serological
samples without loss of neutralising ability holds promise for a more convenient way to store
and ship samples. Overall, this thesis details the study of a novel emergent virus during its first
wave of infections within the UK population. The knowledge gained and techniques
established may be beneficial and informative for studying future emergent viruses
Measuring the HIV-1 Reservoir in the Context of Eradication and Pathogen Co-Infection
Full text linkInfection with human immunodeficiency virus type-1 (HIV-1) causes progressive destruction of the host immune system due to increased loss of the virus’s target cells, CD4 T lymphocytes. Natural HIV-1 infection progresses to a disease known as acquired immunodeficiency syndrome (AIDS), characterised by increased susceptibility to opportunistic infections and cancers. Whilst HIV-1/AIDS was previously a terminal diagnosis, the development of antiretroviral therapy (ART) has drastically reduced HIV-1 associated mortality by providing sustained suppression of virus replication. Nevertheless, ART is unable to eradicate HIV-1 infection due to long-lived cells that harbour transcriptionally silent but inducible provirus, known as the latent reservoir (LR). HIV-1 eradication strategies therefore target latently infected cells for clearance and such studies require robust and sensitive assays to detect small changes in the viral reservoir. Therefore, two prominent HIV-1 quantification assays and widely used calibration standards were compared for their ability to quantify intracellular HIV-1 DNA in patient samples. Based on this analysis, an assay was selected for improvement by redesigning primers to improve the specificity of the method. Further, this work demonstrated the potential for HIV-1 transcript quantification using nanopore based next generation sequencing. Due to overlapping transmission routes and shared regions of high prevalence, HIV-1 infection is frequently associated with co-infections of pathogens such as Mycobacterium tuberculosis (Mtb) and hepatitis C virus (HCV). In both cases, co-infection with HIV-1 leads to more severe prognosis and increased rate of disease progression through disruption of the immune response, however, the effect of these pathogens on HIV-1 disease outcome is not well characterised. To quantify the impact of Mtb on HIV-1 replication, virus production was measured in cells that were infected in the presence of liposomes that mimic the cell wall glycolipid of variant Mycobacteria strains. This analysis demonstrates heterogenous effects of Mycobacteria derived liposomes on HIV-1 infection that is dependent on the tropism of the virus, the strain of Mycobacteria and the method of infection. Based on previous research, which showed that HCV envelope (Env) glycoprotein expression causes downmodulation of HIV-1 LTR driven transcription through disruption of NF-κB, the aim was to measure the effect of exogenous HCV glycoprotein production on HIV-1 activation and replication. Additionally, the effect of HCV glycoprotein expression on the transcriptome of the host cell was investigated using nanopore based RNAseq. Through this analysis, clear upregulation of endoplasmic reticulum stress response pathways in the presence of the HCV Env was observed. These pathways, or individual genes in these pathways, such as the stress response gene, activating transcription factor 3 (ATF3), are potential mediators of HCV induced downmodulation of NF-κB signalling and suggest that through these pathways, HCV may inhibit host immune signalling
HCV and SARS-CoV-2 Envelope Glycoproteins Down-Modulate NF-κB Activity in Association with Induction of ER Stress
No full textComparative analysis and generation of a robust HIV-1 DNA quantification assay
Full text linkHIV-1 infection cannot be cured due to the presence of the latent reservoir (LR). Novel cure or treatment strategies, such as “shock and kill” or therapeutic vaccination, aim to reduce or eradicate the LR. Cure strategies utilise robust DNA quantification assays to measure the change in the LR in low copy scenarios. No standard assay exists, which impedes the reliable comparison of results from different therapy and vaccine trials and HIV-1 total DNA quantification methods have not been previously compared. The HIV-1 long terminal repeat (LTR) has been shown to be the best target for DNA quantification. We have analysed two HIV-1 quantification assays, both able to differentiate between the variant HIV-1 DNA forms via the use of pre-amplification and primers targeting LTR. We identify a strong correlation (r=0.9759, P<0.0001) between assays which is conserved in low copy samples (r=0.8220, P<0.0001) indicating that these assays may be used interchangeably. The RvS assay performed significantly (P=0.0021) better than the CV assay when quantifying HIV-1 total DNA in patient CD4+ T lymphocytes. Sequence analysis demonstrated that viral diversity can limit DNA quantification, however in silico analysis of the primers indicated that within the target region nucleotide miss-matches appear infrequently. Further in silico analysis using up to-date sequence information led to the improvement of primers and enabled us to establish a more broadly specific assay with significantly higher HIV-1 DNA quantification capacity in patient samples (p=0.0057, n=17)
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