Browsing by Author "Abrahams, Bianca"

Now showing 1 - 2 of 2
  • No Thumbnail Available
    Item
    Open Access
    The effect of bacterial vaginosis on HIV infection
    (2024) Abrahams, Bianca; Kullin, Brian
    Human Immunodeficiency Virus-1 (HIV) and Bacterial Vaginosis (BV) are both among the most common diseases affecting young women in Sub-Saharan Africa. BV is characterised by dysbiosis in the female reproductive tract (FRT) when optimal Lactobacillus spp. such as L. crispatus, are displaced by anaerobes such as Gardnerella spp., consistently isolated from the FRT of BV-positive women. Gardnerella spp. produce a number of important virulence factors such as vaginolysin (VLY) and sialidase and are known to initiate the formation of biofilms and hence, contribute to the pathology associated with BV. As many studies have suggested that BV increases susceptibility to HIV infection, it stands to reason that Gardnerella spp. might either indirectly, (initiate the onset of BV and subsequent immune responses) or directly (via its virulence factors) play a role in enhancing HIV acquisition. This study investigated whether VLY, sialidase and biofilm formation played a role in HIV infection. Recently, it was discovered that Gardnerella spp. comprises G. leopoldii, G. vaginalis, G. piotii and G. swidsinskii as well as nine other genome species which vary in virulence potential. When we compared twenty strains isolated from BV-positive women belonging to G. vaginalis (n = 16), G. piotii (n = 2) and G. swidsinskii (n = 2), we found differences in the presence, expression, and activity of VLY and sialidase as well as biofilm-forming capacity between the strains, suggesting a wide range in virulence. However, there was no overt association between HIV infection and Gardnerella virulence factors. Gardnerella sialidase consists of three isoforms: NanH1, NanH2 and NanH3 and as the latter is responsible for the sialidase activity in the FRT, nanH3 was cloned, expressed in E.coli and purified by His-Tag affinity chromatography. We found that purified, recombinant NanH3 increased HIV infection in vitro, most likely by removing the sialic acid moieties on the surface of host cells, reducing the negative repulsive force between the viral Envelope and cell membrane. This may then facilitate virus accumulation at the cell surface, favouring attachment to CD4 and/or its co-receptors and thereby enhance HIV infection. Interestingly, only the two G. piotii strains expressed NanH3, suggesting that perhaps only some species of Gardnerella may play a role in enhancing HIV infection. This has important implications for diagnosis and treatment as BV-positive women shown to carry G. piotii strains, might benefit from preexposure prophylaxis with antiretroviral drugs.
  • Thumbnail Image
    Item
    Open Access
    To compare the expression, processing, incorporation and function of pseudoviruses and infectious molecular clones using different cell types and HIV backbones
    (2019) Abrahams, Bianca; Woodman, Zenda
    Understanding HIV transmission mechanisms is essential for the design and development of an efficacious, broadly acting vaccine that targets features common to transmitted viruses. However, there is a lack of consensus amongst current HIV studies characterising transmitted founders (TFs). When investigating the methods employed across studies, it becomes clear that methodologies are highly variable and thus, could be impacting research outcomes. This study therefore aimed to determine whether Envelope (Env) expression and processing affects function and whether cell type and/or expression system were responsible for these differences. Our data suggest that even though we did not observe differential expression of recombinant Env clones across cell types, when pseudovirus and infectious molecular clone (IMC) backbones were introduced, expression of Env decreased. We also found differences in processing in the form of cleavage, N-glycosylation and incorporation of Env across cell types. We conclude from this that methods used to study Env characteristics are highly sensitive to cell type and HIV backbone which suggests that a more standardised system is required to make meaningful comparisons between studies. The results of our functional Env analysis revealed high variation depending on the methodology used. We found that entry of TZM-bl cells by pseudovirus (PSV) is dependent on the cell line used to produce the viral particles. Unfortunately, due to low IMC titre, we had to expand the virus in PBMCs, negating the effect that cell type might have had on IMC expression. We could thus not directly compare PSVs to IMCs. However, PSV and IMC entry as well as IMC replication in PBMCs suggested that CHO cells were not suitable for robust viral production and better suited for recombinant Env expression. Overall, the findings in this project support previous findings that PSVs and IMCs are not directly comparable due to multiple factors that influence Env expression and virus production. We suggest that researchers who focus on HIV functional analysis, particularly Env, with the end-point of informing vaccine design, need regulated methods across laboratories, similar to the way that neutralisation assays were standardised.