Medical Virology

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38 years after its discovery, Ebola virus spins out of control
(South African Centre for Epidemiological Modelling and Analysis (SACEMA), 2014-09) Preiser, Wolfgang
The evolutionary origin of Ebolaviruses is not very clear. The simple notion that these viruses have been circulating for many millennia in wildlife in tropical parts of Africa, occasionally spilling over into human populations, often brought on by human activities, may not be correct or at least incomplete. Over time a number of Ebola disease outbreaks reported and a pattern in the outbreak response seemed to have been established. A lot was also learnt about Ebola viruses, their epidemiology and ecology. However, the 2014 Ebola outbreak challenges our understanding in many respects.
Aberrant in vitro HLA-DR expression in patients with chronic fatigue
(Health and Medical Publishing Group (HMPG), 1990-08) Van Greune, C. H. J.; Bouic, P. J. D.
To the Editor: The chronic fatigue syndrome (CFS) is a clinical entity characterised by chronic fluctuating fatigue associated-with a multitude of related symptoms, which may vary between patients. It is of unknown causation, but usually follows a presumed acute viral infection.
Academic publishing in pandemic times
(ASSAf, 2020-09-21) Preiser, Wolfgang; Preiser, Rika
ENGLISH ABSTRACT: Even though it tends to feel like ages, it has not been that long since the final days of 2019, when cases of severe respiratory illness (now known as COVID-19), caused by a previously unknown coronavirus (since named SARSCoV- 2), were reported from China. The ongoing COVID-19 pandemic has brought unprecedented disruption to almost every area of our daily and professional lives. Science has not been spared, nor has scientific publishing. Most researchers have been unable to continue with their work, and many had to all but re-invent their teaching. Quite a few have re-invented themselves as coronavirus researchers.1 As biomedical researchers, we are astonished to see how much interest the public is taking in our findings. For no other disease do members of the public so fervently seek out reports in traditional and social media about the latest research findings. These reports often trigger controversial discussions, mostly on social media platforms, about rather complicated aspects of epidemiology, diagnostics, pathogenesis or therapy. Many of these issues are matters outside the realm of everyday life and normally left to experts to assess the evidence and translate it into practice.
Analyses of HIV-1 integrase sequences prior to South African national HIV-treatment program and availability of integrase inhibitors in Cape Town, South Africa
(Nature Publishing Group, 2018) Brado, Dominik; Obasa, Adetayo Emmanuel; Ikomey, George Mondinde; Cloete, Ruben; Singh, Kamalendra; Engelbrecht, Susan; Neogi, Ujjwal; Jacobs, Graeme Brendon
ENGLISH ABSTRACT: HIV-Integrase (IN) has proven to be a viable target for highly specific HIV-1 therapy. We aimed to characterize the HIV-1 IN gene in a South African context and identify resistance-associated mutations (RAMs) against available first and second generation Integrase strand-transfer inhibitors (InSTIs). We performed genetic analyses on 91 treatment-naïve HIV-1 infected patients, as well as 314 treatment-naive South African HIV-1 IN-sequences, downloaded from Los Alamos HIV Sequence Database. Genotypic analyses revealed the absence of major RAMs in the cohort collected before the broad availability of combination antiretroviral therapy (cART) and INSTI in South Africa, however, occurred at a rate of 2.85% (9/314) in database derived sequences. RAMs were present at IN-positions 66, 92, 143, 147 and 148, all of which may confer resistance to Raltegravir (RAL) and Elvitegravir (EVG), but are unlikely to affect second-generation Dolutegravir (DTG), except mutations in the Q148 pathway. Furthermore, protein modeling showed, naturally occurring polymorphisms impact the stability of the intasome-complex and therefore may contribute to an overall potency against InSTIs. Our data suggest the prevalence of InSTI RAMs, against InSTIs, is low in South Africa, but natural polymorphisms and subtype-specific differences may influence the effect of individual treatment regimens.
Analysis of HIV-1 diversity and inflammatory markers in HIV-associated neurocognitive disorders (HAND).
(Stellenbosch : Stellenbosch University, 2022-04) Ruhanya, Vurayai; Engelbrecht, Susan; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology.
ENGLISH SUMMARY: HIV-associated neurocognitive disorders (HAND), which involve impairment or disruption of neurocognitive functioning have become one of the most frequent complications in adult HIV-1 infections with global estimates ranging from 42% to 45%. The screening and diagnosis for HAND relies on multiple clinical and neuro-psychometric methods. However, these methods have a low reliability because they are not precise as most of possess inadequate psychometric properties and diagnostic accuracy. Therefore, this study aimed to describe and characterise viral and immunological determinants of HAND and evaluated their relationship with specific clinical, neuromedical and neuropsychological data to identify putative easy-to-measure biological markers for diagnosis of the condition. This study demonstrated that higher peripheral blood lymphocyte-derived HIV-1 proviral DNA is a predictor of global and domain-specific neurocognitive impairment among individuals infected with HIV-1 subtype C. The study also determined proviral load cut-off /threshold value for neurocognitive impairment and associated diagnostic accuracy. It also identified IP-10 and RANTES as a plasma chemokine bio-signature for HIV-associated neurocognitive impairment with diagnostic accuracy comparable to standard psychometric tests used to screen and describe severity of HAND. In addition, the study identified 3 viral genetic signatures for cognitive impairment, namely Lysine at codon 24, (24K) and Arginine at codon 29 (29R) on Tat protein and Tyrosine (Y) at position 45 (45Y) of Vpr. These three signature amino acids were related to classical markers for monitoring HIV infection. Finally, we identified 4 conserved fragment sequences, PEDQGPQREPYNEWTLE (5 to 21), LGQYIY (42 to 47), TYGDTW (49 to 54), PEDQGPQREPYNEW (5 to 18) on viral protein R, that were associated with higher plasma viral load and proviral load. The study has identified novel cytokine/chemokine and viral biomarker signatures for HIV associated neurocognitive impairment with low to moderate diagnostic accuracy. The findings demonstrated a need for interdisciplinary approach to elucidate possible molecular interactions between the peripheral blood immune markers, viral signatures and the CNS that are linked to observed clinical outcomes of neurodegradation in HIV infection. The identified biomarkers can be further investigated for use as screening tools and treatment endpoints for HAND.
Analysis of viral diversity in relation to the recency of HIV-1C infection in Botswana
(Public Library of Science, 2016) Moyo, Sikhulile; Vandormael, Alain; Wilkinson, Eduan; Engelbrecht, Susan; Gaseitsiwe, Simani; Kotokwe, Kenanao P.; Musonda, Rosemary; Tanser, Frank; Essex, Max; Novitsk, Vladimir; De Oliveira, Tulio
Background: Cross-sectional, biomarker methods to determine HIV infection recency present a promising and cost-effective alternative to the repeated testing of uninfected individuals. We evaluate a viral-based assay that uses a measure of pairwise distances (PwD) to identify HIV infection recency, and compare its performance with two serologic incidence assays, BED and LAg. In addition, we assess whether combination BED plus PwD or LAg plus PwD screening can improve predictive accuracy by reducing the likelihood of a false-recent result. Methods: The data comes from 854 time-points and 42 participants enrolled in a primary HIV-1C infection study in Botswana. Time points after treatment initiation or with evidence of multiplicity of infection were excluded from the final analysis. PwD was calculated from quasispecies generated using single genome amplification and sequencing. We evaluated the ability of PwD to correctly classify HIV infection recency within <130, <180 and <360 days post-seroconversion using Receiver Operator Characteristics (ROC) methods. Following a secondary PwD screening, we quantified the reduction in the relative false-recency rate (rFRR) of the BED and LAg assays while maintaining a sensitivity of either 75, 80, 85 or 90%. Results: The final analytic sample consisted of 758 time-points from 40 participants. The PwD assay was more accurate in classifying infection recency for the 130 and 180-day cut-offs when compared with the recommended LAg and BED thresholds. A higher AUC statistic confirmed the superior predictive performance of the PwD assay for the three cut-offs. When used for combination screening, the PwD assay reduced the rFRR of the LAg assay by 52% and the BED assay by 57.8% while maintaining a 90% sensitivity for the 130 and 180-day cut-offs respectively. Conclusion: PwD can accurately determine HIV infection recency. A secondary PwD screening reduces misclassification and increases the accuracy of serologic-based assays.
Anti-HIV-1 activity of salivary MUC5B and MUC7 mucins from HIV patients with different CD4 counts
(BioMed Central, 2010-10) Habte, Habtom H.; De Beer, Corena; Lotz, Zoe E.; Roux, Paul; Mall, Anwar S.
Background: We have previously shown that MUC5B and MUC7 mucins from saliva of HIV negative individuals inhibit HIV-1 activity by 100% in an in vitro assay. The purpose of this subsequent study was to investigate whether MUC5B and MUC7 from saliva of HIV patients or with full blown AIDS had a similar inhibitory activity against the virus. Methods Salivary MUC5B and MUC7 from HIV patients with different CD4 counts (< 200, 200-400 and > 400) were incubated with HIV-1 prior to infection of the human T lymphoblastoid cell line (CEM SS cells). Cells were then cultured and viral replication was measured by a qualitative p24 antigen assay. The size, charge and immunoreactivity of mucins from HIV negative and positive individuals was also analysed by SDS-PAGE, Western blot and ELISA respectively. Results: It was shown that irrespective of their CD4 counts both MUC5B and MUC7 from HIV patients, unlike the MUC5B and MUC7 from HIV negative individuals, did not inhibit HIV-1 activity. Size, charge and immunoreactivity differences between the mucins from HIV negative and positive individuals and among the mucins from HIV patients of different CD4 count was observed by SDS-PAGE, Western blot and ELISA. Conclusions: Purified salivary mucins from HIV positive patients do not inhibit the AIDS virus in an in vitro assay. Although the reason for the inability of mucins from infected individuals to inhibit the virus is not known, it is likely that there is an alteration of the glycosylation pattern, and therefore of charge of mucin, in HIV positive patients. The ability to inhibit the virus by aggregation by sugar chains is thus diminished.
An anti-SARS-CoV-2 quantitative study to determine the effect of time on the antibody titre, post-vaccination
(Stellenbosch : Stellenbosch University, 2024-01) Meyer, Burnet Adriaan; De Beer, Corena; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Medical Virology.
ENGLISH ABSTRACT: In 2019, the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) outbreak emerged in Wuhan, China, rapidly evolving to a pandemic. Vaccines such as messenger ribonucleic acid (mRNA)- based and Adenovirus-based, have been developed to counter severe Coronavirus Disease 2019 (COVID-19). However, many factors, for example, immunosuppression and prior infection can affect the antibody titre. This study evaluated the effectiveness in Anti-SARS-CoV-2 immunoglobulin (Ig) G production and decline over time, indicating the potential requirement for additional boosters. Furthermore, this study focused on how Adenovirus-based vaccine boosters and breakthrough infections affected the antibody titre. A total of 184 samples were collected from two cohorts which consisted of 67 participants. Samples were collected at baseline, week 3, and week 6 post-booster for Cohort A (laboratory workers at the Western Cape Blood Service). Cohort B (staff and students at the Faculty of Medicine and Health Sciences, Stellenbosch University) had samples collected at baseline, week 2, week 4 and week 8 post-booster. The samples were tested for the presence of the SARS-CoV-2 Anti-nucleocapsid (N) as well as Anti-spike (S) antibodies. The Anti-N antibodies indicated that a participant had a natural infection to SARS-CoV-2 while the Anti-S antibodies were utilised to examine the influence of age, gender and time on vaccine-induced antibodies. This findings of this study demonstrated that vaccine-induced antibodies were still detectable six months following the initial vaccination. A significant increase was observed for both cohorts, when comparing the baseline titre with the different timepoints, post-booster. Multiple samples were found to be Anti-N positive, suggesting that these participants were potential asymptomatic cases. Variations were found in the Anti-S antibody titre based on different age groups, although these variations were not statistically significant. Moreover, the results have indicated that gender does not affect the Anti-S antibody titre. This research enhances the understanding of antibody titre dynamics, contributing to a better grasp of immune longevity. Further studies are recommended to determine the impact of combining different boosters on the antibody titre, and assess the difference in antibody titre between monovalent boosters with bivalent boosters.
Antibody responses to vaccination among South African HIV-exposed and unexposed uninfected infants during the first 2 years of life
(American Society for Microbiology, 2013-01) Reikie, Brian A.; Naidoo, Shalena; Ruck, Candice E.; Slogrove, Amy L.; De Beer, Corena; La Grange, Heleen; Adams, Rozanne C. M.; Ho, Kevin; Smolen, Kinga; Speert, David P.; Cotton, Mark F.; Preiser, Wolfgang; Esser, Monika; Kollmann, Tobias R.
HIV-exposed but uninfected (HEU) infants born to HIV-infected mothers from areas in the world with a high burden of infectious disease suffer higher infectious morbidity and mortality than their HIV unexposed uninfected (HUU) peers. Vaccination provides protection from infection. The possibility exists that altered response to vaccination contributes to the higher rate of infection in HEU than in HUU infants. While short-term, cross-sectional studies support this notion, it is unclear whether or not HEU infants develop long-term protective immune responses following theWHOextended program on immunization (EPI). Vaccine-specific antibody responses were compared between HEU and HUU infants from 2 weeks until 2 years of age in a longitudinal South African cohort. Total IgG and antibodies specific for Bordetella pertussis, Haemophilus influenzae type b (Hib), tetanus toxoid, hepatitis B virus (HepB), and measles virus were measured at multiple time points throughout the first 2 years of life. Prevaccine antibodies (maternal antibodies passively acquired) specific for tetanus were lower in HEU than in HUU infants, while prevaccine antibodies to HepB were higher in HEU than in HUU infants. Both groups responded similarly to tetanus, Hib, and HepB vaccination. HEU demonstrated stronger pertussis vaccine responses, developing protective titers 1 year earlier than HUU patients, and maintained higher anti-tetanus titers at 24 months of age. Vaccine-induced antibodies to measles virus were similar in both groups at all time points. Our results suggest that the current EPI vaccination program as practiced in South Africa leads to the development of vaccine-specific antibody responses that are equivalent in HEU and HUU infants. However, our data also suggest that a large fraction of both HEU and HUU South African infants have antibody titers for several infectious threats that remain below the level of protection for much of their first 2 years of life.
The apoptotic potential of different HIV-1 subtype C Tat mutations in cell culture
(Stellenbosch : Stellenbosch University, 2013-03) Isaacs, Shahieda; Engelbrecht, Susan; Glashoff, Richard; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Medical Virology.
The efficiency in which HIV-1 can infect, spread and evade the attack of therapeutic agents can be attributed to a high mutation rate and frequent recombination events. These factors have collectively contributed to the diversity observed in HIV-1 and resulted in a multitude of subtypes, sub-subtypes, circulating recombinant forms (CRF’s) and unique recombinant forms (URF’s). The aim of this study was to investigate HIV-1 diversity in Cape Town using a small cohort of treatment naive patients being investigated for HIV Associated Neurocognitive Disorders (HAND). Four different genomic domains: gag, pol, accessory and gp41 genes were sequenced to subtype the virus. HIV-1 tat was further investigated because the dicysteine motif has been reported to play a role in HAND. Viral RNA and proviral DNA was extracted from 64 patients and used for the amplification and sequencing of the genes. Rega and jpHMM online tools were used to identify HIV-1 subtypes and recombinants while Neighbor-joining phylogenetic trees were constructed for phylogenetic analysis. The pol gene was further investigated using SCUEAL to detect possible intra-subtype recombination and was also screened for the presence of transmitted drug resistance. In addition tat sequence datasets retrieved from the Los Alamos sequence database were investigated and compared with the newly generated sequences for the detection of point mutations and amino acid signature patterns. Sequencing identified most of the samples as subtype C; however six inter-subtype recombinants (AE, A1G, A1CU and two BC) and 9 intra-subtype C recombinants were identified. In addition 13% of pol sequences were identified with resistance mutations. Signature pattern analysis identified a high level of variability in the tat sequences: 68% were identified with C30S31; 29% with the C30C31 mutation and a single sequence with a novel mutation C30A31. Functional analysis of these mutations indicated that all mutations investigated were capable of inducing apoptosis in cell culture. The C30C31 mutation generated the highest level of apoptosis, closely followed by the C30A31 mutation. However no statistical significance could be detected between tat mutations and the observed levels of apoptosis.
The application of wastewater-based epidemiology to Hepatitis E virus in South Africa
(Stellenbosch : Stellenbosch University, 2024-02) Roberts, Bronwyn; Maponga, Tongai; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Medical Virology.
ENGLISH ABSTRACT: Background Hepatitis E virus (HEV) has a global presence, but the highest burden of disease is found in Africa and Asia. Unfortunately, in Sub-Saharan Africa, there is a gap in knowledge on the molecular epidemiology of the disease, as often serological methods only are applied. In recent years a high seroprevalence of the virus has been detected in populations in the Western Cape province. Evidence of viraemia has been published on a few South African patients from the Western Cape. Viral ribonucleic acid (RNA) was also detected in swine samples from an abattoir and in porcine meat products in Cape Town. Based on the existing literature, HEV genotype 3 has been detected in patients and swine samples. In this study, the aim was to determine whether HEV was circulating in communities in the Western Cape by testing wastewater extracts for the presence of HEV RNA. The study also sought to determine the HEV genotype in wastewater sample extracts. Methods One hundred and forty-three wastewater extracts were tested for the presence of HEV RNA using real-time polymerase chain reaction (PCR). Samples were sourced from four locations: two wastewater treatment works and sewer manholes from two Stellenbosch University residences. Nested reverse-transcriptase PCR (RTPCR) targeting three regions of the HEV genome was applied to samples which tested positive for HEV RNA. The sequenced regions were a portion of open reading frame (ORF) 2 covering 347 base pairs (bp), a 286 bp region of ORF1 and a 126 bp region of the overlapping ORF2/3. The generated PCR products were then sequenced with Sanger sequencing. The generated consensus sequences were placed in a phylogenetic tree with reference sequences for HEV genotypes 1 to 8, to determine which genotype is likely circulating in the selected communities. Results Out of the 143 wastewater samples, 130 valid results were generated. Out of 130, 21 (16.2%%) were positive and 109 (83.9%) were negative. Of the 21 positive samples, 5 (23.8%) were collected from Athlone wastewater treatment works (WWTW), 9 (42.9%) from Zandvliet WWTW, 1 (4.8%) from Meerhoff residence in Tygerberg and 6 (28.6%) from Metanoia residence in Stellenbosch. The positive samples had a median cycle threshold value of 37.9 (interquartile range: 36.7-39.7). Out of the 21 positive samples, 4 (19.1%) sequences were obtained from the ORF2/3 region. The sequences clustered most closely with a sequence from a South African patient and HEV genotype 3 reference sequences. Conclusions Based on the real-time PCR results, it appears that HEV is circulating among communities in the Western Cape province. Based on the detection of HEV genotype 3 in the wastewater samples, it suggests that zoonotic transmission may be the mostly likely route of infections. Further investigation to identify porcine and other food products which may be contaminated with HEV are necessary to break chains of transmission.
B lymphocyte activation and exhaustion in chronic HIV : novel surrogate markers of generalised immune activation and selective modulation of aberrant B cell responses using vasoactive intestinal peptide (VIP)
(Stellenbosch : Stellenbosch University, 2015-04) Reid, Timothy Dawson; Glashoff, Richard H.; Ipp, Hayley; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Medical Virology.
ENGLISH ABSTRACT: Introduction: Chronic HIV-1 infection is characterized by immune activation and dysregulation of immune homeostasis, which impacts on multiple immune cell types. The B-cell compartment, which plays an important role in the producing neutralizing antibodies, is also dysregulated in HIV- 1 infection. In this study we investigated peripheral blood B-cell subset distribution, and changes in expression of cellular activation, inhibition, and apoptosis signaling markers in both untreated chronic HIV-1 infected individuals and healthy uninfected controls. The neuropeptide immune modulator, vasoactive intestinal peptide (VIP) is known to selectively down-regulate activation of CD4+ T-cells in various disease settings including HIV-1, however to our knowledge, no studies have investigated the effect of VIP inhibition on B-cell activation. Materials & Methods: A total of 21 HIV+ve (CD4 count >250 cells/µl), and 19 HIV-ve individuals were recruited from the Emavundleni voluntary testing and counseling clinic in Crossroads, Western Province, South Africa. Whole blood was stained to distinguish B-cell subsets (activated memory (AM: CD21-CD27+), resting memory (RM: CD21+CD27+), mature naïve (MN: CD21+CD27-), or tissue-like memory (TLM: CD21loCD27lo). In addition expression of markers of B-cell activation (CD126, CD86, CD38, CD284, CD287), inhibition (CD72, CD85j, CD300a, CD305, CD307d), and apoptosis signaling (CD95), was assessed ex vivo by flow cytometry (BD FACSCanto II). For determination of functional responsiveness isolated B-cells (RosetteSep, Stemcell Technologies) were cultured for 18h (37°C, 5%CO2) without stimulation or stimulated with TLR ligands (LPS or R848). Stimulation experiments were also performed in the presence or absence of VIP. Results: Chronic HIV-1 infection affected B-cell subset distribution. The percentage (%) of TLM was increased by 59.24%, and %RM was decreased by 22.73% (both p<0.01). Total expression of the VIP receptor VPAC2 was decreased by 47.35% (p=0.0296). Subsets had a mixed phenotype ex vivo; HIV infection upregulated CD38 (by 59.56%, p=0.0004), CD72 (by 60.70%, p=0.0396), CD307d (by 68.63%, p=0.0015) on AM, while RM B-cells had increased expression of TLR4 (by 107.04%, p=0.0057) and TLR7 (by 208.14%, p=0.0199). TLM B cells (i.e. exhausted phenotype) displayed upregulated TLR7 (by 550%, p=0.0128) and CD307d (by 72.40% p=0.045) expression. MN B-cells had increased CD72 expression (by 70.98%, p=0.0026). R848 upregulated CD86 expression by 42.20% on AM (p<0.01), and by 56.06% on RM B-cells (p<0.01), which was significantly downregulated with VIP inhibition (both p<0.05). Similarly, CD95 expression on RM, TLM, and MN B-cells increased by 31.10% (p<0.001), 21.46% (p<0.01), and 39.92% (p<0.01) with R848 stimulation respectively, which was also significantly downregulated with VIP inhibition. Conclusion: These data indicate that B-cells in untreated HIV infection display increased levels of activation, and also the potential for increased susceptibility to apoptosis as evidenced by increased FAS (CD95) expression. VIP significantly down-regulated markers of activation, inhibition, and apoptosis signaling. Dysregulation of B-cells is thus apparent in asymptomatic stable chronic HIV-1 infection, which may impact on both inefficient neutralizing antibody production and hypergammaglobulinemia. The ability of VIP to prevent stimulationassociated marker upregulation may indicate that VIP is a potential therapeutic agent. Its immuno-modulatory properties were demonstrated to limit B-cell hyperactivation, and selectively down-regulate apoptosis and mark it out for further investigation.
Barriers to HIV remission research in low- and middle-income countries
(Wiley Open Access, 2017) Rossouw, Theresa; Tucker, Joseph D.; Van Zyl, Gert U.; Sikwesi, Kenly; Godfrey, Catherine
Introduction: HIV eradication and remission research has largely taken place in high-income countries. In low- and middleincome countries (LMIC), there may be factors that have a substantial impact on the size of the latent HIV reservoir and the immunological response to infection. If a curative strategy is to be available to all HIV-infected individuals, these factors must be understood. Methods: We use a scoping review to examine the literature on biological factors that may have an impact on HIV persistence in LMIC. Three databases were searched without date restrictions. Results: Uncontrolled viral replication and higher coinfection prevalence may alter the immunological milieu of individuals in LMIC and increase the size of the HIV reservoir. Differences in HIV subtype could also influence the measurement and size of the HIV reservoir. Immune activation may differ due to late presentation to care, presence of chronic infections, increased gut translocation of bacterial products and poor nutrition. Conclusions: Research on HIV remission is urgently needed in LMIC. Research into chronic immune activation in resource poor environments, the immune response to infection, the mechanisms of HIV persistence and latency in different viral clades and the effect of the microbiological milieu must be performed. Geographic differences, which may be substantial and may delay access to curative strategies, should be identified.
Barriers to VCT despite 13 years of community-based awareness campaigns in a peri-urban township in northern Limpopo
(Health and Medical Publishing Group (HMPG), 2010) De Koker, Petra; Lefevre, Pierre; Matthys, Francine; Van der Stuyft, Patrick; Delva, Wim
Bimodal distribution and set point HBV DNA viral loads in chronic infection : retrospective analysis of cohorts from the UK and South Africa
(Wellcome Open Research, 2020-10-14) Downs, Louise O.; Vawda, Sabeehah; Bester, Phillip Armand; Lythgoe, Katrina A.; Wang, Tingyan; McNaughton, Anna L.; Smith, David A.; Maponga, Tongai; Freeman, Oliver; Várnai, Kinga A.; Davies, Jim; Woods, Kerrie; Fraser, Christophe; Barnes, Eleanor; Goedhals, Dominique; Matthews, Philippa C.
ENGLISH ABSTRACT: Hepatitis B virus (HBV) viral load (VL) is used as a biomarker to assess risk of disease progression, and to determine eligibility for treatment. While there is a well recognised association between VL and the expression of the viral e-antigen protein, the distributions of VL at a population level are not well described. We here present cross-sectional, observational HBV VL data from two large population cohorts in the UK and in South Africa, demonstrating a consistent bimodal distribution. The right skewed distribution and low median viral loads are different from the left-skew and higher viraemia in seen in HIV and hepatitis C virus (HCV) cohorts in the same settings. Using longitudinal data, we present evidence for a stable 'set-point' VL in peripheral blood during chronic HBV infection. These results are important to underpin improved understanding of HBV biology, to inform approaches to viral sequencing, and to plan public health interventions.
Biomedical research and capacity building : bilateral collaboration between research institutes in South Africa and Cameroon
(Health & Medical Publishing Group, 2016) Jacobs, Graeme Brendon; Ikomey, G. M.
No abstract available
Blood dendritic cells in chronically HIV-1 infected individuals in South Africa: Phenotype, function and immune modulation
(Stellenbosch : Stellenbosch University, 2016-12) De Swardt, Dalene; Glashoff, Richard H.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Medical Virology
ENGLISH ABSTRACT :HIV-1 infection detrimentally affects CD4 T lymphocytes as well as the blood plasmacytoid (pDC) and myeloid dendritic cell (mDC) compartment. DCs act as innate sensors and as initiators and directors of antigen-specific immune responses. Whereas, mDCs have the unique ability to prime naïve T-lymphocytes and activate adaptive immune responses, pDCs are primary producers of type 1 interferons (IFNs), playing a pivotal role in anti-viral immunity. In the current study both pDCs and mDCs from chronically HIV-1 infected South African individuals (on or naïve for ARV therapy) as well as with and without concurrent TB disease, were compared to matched uninfected controls. Similar to CD4 T lymphocytes, bothpDCs and mDCs, were significantly depleted during HIV-1 infection, (reduction of pDC, mDC and CD4 T lymphocyte was 63% (p 0.001), 80% (p 0.001) and 31% (p 0.01), respectively). In parallel, significantly higher levels of generalised immune activation and exhaustion (CD38+CD8+, PD-1+CD8+ and CD38+PD-1+CD8+ T lymphocytes) were observed. ARV treatment ( 1 year) did not result in DC number recovery despite a significant increase of CD4 T lymphocytes numbers (CD4 T lymphocyte number gain of 89% (p 0.01), it fell short of full recovery).TB co-infection did not exacerbate number loss. Phenotypic characterisation of DC populations in circulation during HIV-1 infection may indicate the underlying reasons for the loss from circulation. Phenotypic profiling by multiparameter flow cytometry included: markers of activation (CD86, CD80 and CD62L), maturation (CD83), apoptosis (TNF-R2, FAS, FASL and TRAIL R1-R4) and chemotaxis (CCR5, CCR7, CCR9 and CXCR6). HIV-infection was associated with a significantly higher percentage of CD86+mDCs which may be indicative of early maturation or transition to secondary lymphoid tissue. The frequency of the CD86+mDCs subset normalised upon ARV therapy. Also, HIV-1 infection influenced the distribution of TNF-R2+pDCs and TNF-R2+mDCs. Increased TNF-R2 expression in both subsets, may attest to enhanced survival function. Functionally, DCs of HIV-1 infected individuals were reactive to TLR-L stimulation and in some cases showed enhanced responses compared to uninfected individuals. A significantly higher frequency of TNF-R2+pDCs, IFN- +pDCs, and TNF +mDCs was observed in whole blood TLR cultures of HIV-1 infected individuals (TNF-R2+pDCs: LPS (p = 0.002) and R848 (p = 0.01), IFN- +pDCs: R848 (p = 0.04), TNF +mDCs: LPS (p = 0.003))s.In addition, whole blood TLR cultures of the ARV treated study group generally showed normalisation of the responses, however; in certain cases ARV therapy reduced responsiveness to levels significantly lower than the control study group (i.e.TNF-R2+pDCs and TNF-R2+mDCs in CpG ODN stimulation). In contrast, a significantly lower frequency of IL12p40+mDCs was observed during HIV-1 infection (p = 0.02). TLR-L cultures of the ARV treated study groups showed normalisation of IL12p40+mDCsfrequency. Notably, treatment with the immunomodulating peptide VIP induced a decline in IL12p40+mDCs to levels lower than the control study group.The frequency of TNF +pDCs in TLR-L whole blood cultures was similar between the healthy, untreated and treated HIV-1 infected study groups, however, significantly reduced frequencies were observed in these study groups upon VIP treatment. These data indicate unique phenotypic and functional changes in DC subsets in chronic HIV- 1 infection which may provide potential targets for immunotherapeutic intervention.
Candida species : species distribution and antifungal susceptibility patterns
(Health and Medical Publishing Group (HMPG), 2008) Arendse, Tracy; Orth, Heidi
[No abstract available]
The characterisation and expression of HIV-1 subtype C gag
(Stellenbosch : Stellenbosch University, 2002) Sampson, Candice Corene; Van Rensburg, E.J.; Engelbrecht, S.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Division of Medical Virology.
ENGLISH ABSTRACT: The gag gene of HIV-1 encodes for one of the major structural proteins, which contains several conserved cytotoxic T cell (CTL) epitopes. Gag specific CTL responses are important in controlling viral load during acute infection and asymptomatic stages of the infection. Currently, only one complete South African HIV-1 subtype C gag sequence has been published. The first aim of this study was to characterise the complete gag gene of 15 HIV-1 subtype C isolates, to be used as a set of reference sequences in the design of a South African HIV-1 subtype C vaccine. Fifteen HIV-1 subtype C isolates selected for this study, were isolated during 1998 and 1999 from the HIV-1 positive patients attending the Infectious Disease Clinic at Tygerberg Hospital. The gag gene of these isolates was amplified by PCR, cloned into mammalian expression vectors and sequenced. Restriction digest analyses as well as phylogenetic analyses were performed on the sequencing data. Previously published mutational analyses and CTL epitopes were compared to the predicted amino acid sequences of the gag clones. Sequences of 23 complete gag genes representing the 15 HIV-1 subtype C isolates as well as one complete sequence of an HIV-1 subtype B isolate were compiled. Subtyping by restriction fragment length polymorphism (RFLP) would have correctly identified 14 of the 15 subtype C isolates as subtype C and one as unidentifiable. The subtype B isolate would have also been correctly identified. Phylogenetic analyses showed that our subtype C isolates clustered with reference subtype C strains from various countries, including Botswana, India, Israel, Tanzania and Zambia. Strains from Ethiopia and Brazil formed a separate subtype C cluster. The diversity between our isolates was comparable to the diversity seen between all the HIV-1 subtype C strains. Comparisons of previously published mutational analyses and CTL epitopes to the predicted amino acid sequences of the gag clones, showed conservation in most of the clones throughout the sequence. A second aim was to establish transfection and Western Blot techniques in our laboratory for use in future studies. An in vitro transcription! translation assay was performed on the gag clones and the protein producing clones were used to transfect mammalian cells using electroporation. A Western blot was then used to screen for Gag protein expression in the transfected cell Iysates. The in vitro transcription! translation assay showed that seven of the 23 clones could produce a protein of -55 kDa in size. Four out of the seven of these clones gave a weak expression of a-55 kDa protein after transfection in a mammalian cell line. Since the completion of the experimental work of this study, other cloned HIV-1 genes have successfully been transfected into mammalian cells using the electroporation technique and the proteins produced were screened for by Western blot. To conclude with; the native form of the gag gene does not elicit strong expression of the protein, but studies have shown that expression can be improved by sequence-modification of the gag nucleotide sequence. Due to the conservation of gag, the sequence of any subtype C strain can be used for the development of a Southern African vaccine.
Characterisation of new full-length HIV-1 subtype D viruses from South Africa
(2004-12) Loxton, Andre Gareth; Janse van Rensburg, E.; Engelbrecht, S.
ENGLISH ABSTRACT: The first episode of HIV-1 in South Africa was documented in 1982. Homosexual transmission of the virus was the predominate mode of transmission in an epidemic of mainly HIV-1 subtype Band D infections. To date, no full-length sequences of Subtype D strains from South Africa has been reported. Here we describe the characterization and some of the unique features of the Tygerberg HIV-1 subtype D strains. A near full-length 9 kb fragment was obtained through a one step PCR using high molecular weight DNA. Cloning was done successfully with the pCR-XLTapa cloning kit. Large quantities of plasmid DNA was grown and sequenced on both strands of the DNA. ORF determination and subtyping was followed by standard phylogenetic methods to construct evolutionary phylogenetic trees. Subtyping and similarity plots revealed that the sequences from Tygerberg are pure subtype D. All the Tygerberg strains had intact genes with no premature stop codons. At the tip of the V3 loop, the Tygerberg strains have the GOGO motif. R214 has a more variable vpu gene than the rest of the Tygerberg strains, but is still subtype D in this region. No premature stop codons have been observed in the tat gene and the glycosilation of the strains are less than the subtype D consensus. We are the first to report full-length sequences of HIV-1 subtype D strains from South Africa. The sequences represent non-mosaic genomes of subtype D. Our results confirm that the subtype D sequences from the beginning of the HIV-1 epidemic differ from the Subtype D sequences from recent isolates.

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