Clinical and epidemiological aspects of HIV and Hepatitis C virus co-infection in KwaZulu-Natal province of South Africa.

Thesis (M.Med.)-University of KwaZulu-Natal, Durban, 2008.HIV is known to affect the epidemiology, transmission, pathogenesis and natural history of HCV infection whilst studies on the effects of HCV on HIV have shown conflicting results and are confounded by the influence of intravenous drug use and anti-retroviral therapy. This study was conducted in KwaZulu-Natal Province in South Africa where HIV is predominantly a sexually transmitted infection. Intravenous drug use is rare in this region and the study population was naive to anti-retroviral therapy. For this study, specimens from selected sentinel sites submitted to a central laboratory for routine HIV testing were screened for anti-HCV IgG antibodies. HIV positive HCV-positive patients were compared to HIV-positive HCV-negative patients in a subgroup of patients within this cohort in order to determine if HCV sero-prevalence was associated with clinical outcomes in a linked anonymous retrospective chart survey. The prevalence of HCV was 6.4% and that of HIV, 40.2%. There was a significantly higher prevalence of HCV among HIV infected patients as compared to HIV negative patients (13.4% vs. 1.73% respectively). HCV-HIV co-infected patients had significantly increased mortality (8.3 vs. 21%). A significant association was found between HCV serostatus and abnormal urea and creatinine levels. Hepatitis B surface antigen seropo-sitivity was not found to be a confounding factor. This study has found that hepatitis C co-infection is more common in HIV positive individuals and is associated with an increased mortality and renal morbidity

Synthesis, characterization and biocompatibility of a multifunctional gold nanoparticle system for the delivery of single-stranded RNA to lymphocytes

The use of RNA macromolecules as therapeutic agents for HIV and other infectious diseases is promising but limited by suboptimal delivery to the target site. With HIV infection, this is particularly challenging since lymphocytes are particularly difficult to transfect. This paper describes an innovative strategy for the intracellular delivery of a novel single-stranded RNA (oligoribonucleotide) with putative anti-HIV activity. This strategy is based on a PEGylated gold nanoparticle scaffold covalently linked to the thiol-modified oligoribonucleotide via a cleavable N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) linker molecule. The nanoparticle was then coated with a cationic polymer (polyethyleneimine) to facilitate cell entry and endosomal escape. A synthetic anti-CD4 cyclic targeting peptide was attached to the polyethyleneimine-coated nanoparticle via an SPDP linker molecule, in an attempt to enhance uptake and selectivity. Synthesis, characterization, SPDP and RNA loading, cytotoxicity and antiviral activity of the nanoparticle are described. Approximately 45 000 strands of RNA were taken up per lymphocyte. Uptake was limited by relatively inefficient loading ofRNAonto the gold nanoparticle surface (1 strand per 4.8 nm2 of nanoparticle surface area) and significant aggregation of the nanoparticle in physiological solutions. No antiviral activity was demonstrated, possibly due to insufficient intracytoplasmic delivery of the RNA.Keywords: Gold nanoparticle, polyethyleneimine, transfection, RNA deliver

IN VITRO TESTING OF AFRICAN TRADITIONAL MEDICINES FOR CYTOTOXIC, IMMUNE MODULATORY AND ANTI-HIV ACTIVITIES

African Traditional Medicines (ATMs) serve as a major source of primary healthcare for African people. The reasons for their use range from easy access, affordability, beliefs in traditional systems and long term safety. ATMs have been used to treat individuals infected with HIV and therefore need scientific validation; a view supported by Traditional Health Practitioners (THPs). This study aimed to evaluate the in vitro cytotoxicity, immune modulatory and anti-HIV activities of traditional multiple herbal preparations from local THPs. Ugambu, Ihashi, Product Nene, Product Blue, SPNa and SDKc ATM were supplied by local THPs. Changes in adenosine triphosphate (ATP) & glutathione (GSH) over 24 hours were measured using luminometry. Changes in 12 cytokines were assayed using an ELISA-based absorbance assay. Protective effects against HIV killing of MT-4 cells were tested using the XTT assay and antiviral activity was measured using an HIV-1 viral load assay. Cyclosporine and AZT were used as positive controls. Ugambu, Ihashi, Product Nene and SDKc induced a dose dependent toxicity on treated PBMCs by reducing ATP and GSH at high doses (p< 0.001). These medicinal preparations, along with SPNa, showed immunomodulatory activity by significantly (p< 0.001) changing the secretion of pro-inflammatory cytokines. Product Blue stimulated the levels of ATP and GSH in treated PBMCs at all doses however this product did not show any immunomodulatory activity on cytokine secretion when compared to control cells. Ugambu, Ihashi, Product Nene showed promising anti-HIV activity relative to AZT (p< 0.01). This study has shown that some of these traditional medicinal preparations have at least one or all the properties of immunostimulation, immunomodulation or antiretroviral effects. The mechanism of action of the shown activities should further be investigated

Hepatitis B and HIV co-infection in pregnant women: Indication for routine antenatal hepatitis B virus screening in a high HIV prevalence setting

Background. Sub-Saharan Africa is endemic for hepatitis B virus (HBV) and human immunodeficiency virus (HIV) infections. HBV/HIV co-infection in women of reproductive age is of clinical and public health importance because these women constitute a significant reservoir for horizontal and perinatal HBV transmission. Childhood HBV vaccination from 6 weeks of age protects most children against chronic HBV infection. However, infants born to HBV/HIV co-infected women are more likely to be infected perinatally, with an increased risk of chronic hepatitis, than infants born to HBV mono-infected women.Objectives. The aim of our study was to establish the prevalence of HBV infection and HBV/HIV co-infection in pregnant women in KwaZulu-Natal, South Africa, to inform antenatal HBV screening and childhood immunisation policies in South Africa.Methods. Stored plasma specimens obtained from 570 pregnant women were tested for hepatitis B surface antigen (HBsAg) and HBV infectivity, as characterised by the presence of hepatitis B e antigen (HBeAg) and/or HBV DNA load.Results. The antenatal HIV prevalence and HBsAg prevalence in this study were 41.6% and 5.3% (95% confidence interval (CI) 3.4 - 7.1%), respectively. Overall, 3.1% (95% CI 1.7 - 4.6%) of pregnant women were HBV/HIV co-infected, with HBeAg positivity and the HBV DNA load being significantly higher in co-infected women.Conclusion. We report a 5.3% HBV prevalence and a 3.1% HBV/HIV co-infection prevalence in pregnant women from this HIV-endemic region. Routine antenatal HBV screening will allow early identification of neonates who require HBV active-passive immunoprophylaxis at birth. This strategy, together with antenatal antiretrovirals, will reduce the risk of perinatal HBV transmission, especially in high-risk HBV/HIV co-infected pregnant women

HIV-1 drug resistance by ultra-deep sequencing following short course zidovudine, single-dose nevirapine, and single-dose tenofovir with emtricitabine for prevention of mother-to-child transmission

Antiretroviral drug resistance following pMTCT strategies remains a significant problem. With rapid advancements in next generation sequencing technologies, there is more focus on HIV drug-resistant variants of low frequency, or the so-called minority variants. In South Africa, AZT monotherapy for pMTCT, similar to World Health Organization option A, has been used since 2008. In 2010, a single dose of co-formulated TDF/FTC was included in the strategy for prevention of resistance conferred by single-dose nevirapine (sd NVP). The study was conducted in KwaZulu-Natal, South Africa, among pMTCT participants who received AZT monotherapy from 14 weeks of gestation, intrapartum AZT and sd NVP, and postpartum sd TDF/FTC. Twenty-six specimens collected at 6 weeks post-delivery were successfully sequenced using 454 ultra-deep sequencing. Non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance was detected in 17 of 26 (65%) patients, 2 (7%) had Thymidine analogue mutations, and 3 (11%) had K65R. Of the 17 patients with NNRTI resistance, 11 (65%) had high-level NNRTI resistance, whereas 6 (35%) had intermediate NNRTI resistance. The levels of NNRTI resistance are much higher than would be expected, given the inclusion of antepartum AZT and postpartum TDF/FTC. This high level of NNRTI resistance could impact future NNRTI-containing treatment for a large proportion of pMTCT-exposed women. The detection of Thymidine analogue mutations highlights the need to understand the clinical impact of these on AZT-containing antiretroviral treatment in women exposed to AZT monotherapy

Nanotechnology and the Treatment of HIV Infection

Suboptimal adherence, toxicity, drug resistance and viral reservoirs make the lifelong treatment of HIV infection challenging. The emerging field of nanotechnology may play an important role in addressing these challenges by creating drugs that possess pharmacological advantages arising out of unique phenomena that occur at the “nano” scale. At these dimensions, particles have physicochemical properties that are distinct from those of bulk materials or single molecules or atoms. In this review, basic concepts and terms in nanotechnology are defined, and examples are provided of how nanopharmaceuticals such as nanocrystals, nanocapsules, nanoparticles, solid lipid nanoparticles, nanocarriers, micelles, liposomes and dendrimers have been investigated as potential anti-HIV therapies. Such drugs may, for example, be used to optimize the pharmacological characteristics of known antiretrovirals, deliver anti-HIV nucleic acids into infected cells or achieve targeted delivery of antivirals to the immune system, brain or latent reservoirs. Also, nanopharmaceuticals themselves may possess anti-HIV activity. However several hurdles remain, including toxicity, unwanted biological interactions and the difficulty and cost of large-scale synthesis of nanopharmaceuticals

Incidence, clinical spectrum, risk factors and impact of HIV-associated immune reconstitution inflammatory syndrome in South Africa.

Immune reconstitution inflammatory syndrome (IRIS) is a widely recognised complication of antiretroviral therapy (ART), but there are still limited data from resource-limited settings. Our objective was to characterize the incidence, clinical spectrum, risk factors and contribution to mortality of IRIS in two urban ART clinics in South Africa

The prevalence of hepatitis B virus infection in HIV-positive and HIV-negative infants: KwaZulu-Natal, South Africa

Background: The prevalence of hepatitis B virus (HBV) amongst South African infants and children has been reported in the pre-HIV era. Despite the reported high prevalence of HIV in the general population of South Africa, the rate of HIV/HBV co-infection amongst infants and children remains poorly reported. Objectives: We describe the prevalence of HBV infection amongst HIV-positive and HIV-negative infants by molecular methods of diagnosis using dried blood spot samples. Methods: This retrospective cross-sectional study was conducted between July 2011 and December 2011 in an academic referral laboratory offering viral diagnostic services to the entire KwaZulu-Natal province of South Africa. A total of 322 study samples were collected from discarded residual dried blood spot samples following routine infant diagnosis of HIV. Equal proportions of HIV-positive and HIV-negative infant specimens were studied. Statistical differences in the prevalence of HBV between the HIV-positive and HIV-negative samples were calculated using the Pearson chi-square test, and a p-value < 0.05 was considered statistically significant. Further testing for HBV DNA using a nested polymerase chain reaction method was performed. Results: The overall prevalence of HBV was 10%. In the HIV-positive group, 21 of 161 infants tested positive for HBV compared with 12 of 161 HIV-negative infants who tested positive for HBV. The proportion of infants infected with HBV was marginally higher amongst HIV positiveinfants (13.0%; 95% CI 6.8–19.9) compared with HIV-negative infants (7.5%; 95% C I2.5–13.7; P = 0.098), though not statistically significant. Conclusion: The finding of a 10% HBV prevalence in this infant cohort is clinically significant. The non-statistically significant difference in HBV prevalence between the HIV-positive and HIV-negative infants suggests that high prevalence of HBV infection in children may be a problem independent of HIV

Real-time polymerase chain reaction optimised for hepatitis C virus detection in dried blood spots from HIV-exposed infants, KwaZulu-Natal, South Africa

Background: There is a paucity of data on the prevalence of hepatitis C virus (HCV) in children, particularly in sub-Saharan Africa. A major obstacle in resource-limited settings for polymerase chain reaction (PCR) testing is the necessity for specimen transportation and storage at low temperatures. There are numerous recent studies of using real-time HCV PCR for diagnosis and screening of plasma and serum, but few have looked at using dried blood spot (DBS) specimens. Objectives: The aim of this study was to optimise a real-time HCV PCR method to detect HCV RNA from infant DBS specimens for use as a tool for HCV surveillance in KwaZulu-Natal, South Africa. Method: The LightCycler® 2.0 instrument was used for the HCV PCR using the LightCycler® RNA Master SYBR Green I kit. Template volume, primer concentration and primer annealing temperatures were optimised and the method was used on 179 DBS specimens from HIV-exposed infants in KwaZulu-Natal. Results: Primer concentrations adjusted to 0.25 µM and a template volume of 10 µL improved the PCR amplification. Primer annealing temperatures lowered from 65 °C to 58 °C resulted in higher quantities of amplified PCR product. The limit of detection of the optimised HCV PCR assay was between 1200 IU/mL and 3580 IU/mL of HCV RNA. HCV was not detected in any of the 179 DBS specimens. Conclusion: The optimised real-time HCV PCR on infant DBS specimens performed well, but HCV was not found in this surveillance study. HIV infection may have little impact on the vertical transmission of HCV in this region