Selokela G. Selabe

Increased detection of HBV DNA in HBsAg-positive and HBsAg-negative South African HIV/AIDS patients enrolling for highly active antiretroviral therapy at a tertiary hospital.

This retrospective study investigated the prevalence of hepatitis B virus (HBV) in 192 stored sera from human immunodeficiency virus (HIV) positive South African patients initiating antiretroviral therapy (ART), and explored the implications of HBV–HIV co‐infection on laboratory diagnosis of HBV. HBV serology (HBsAg, anti‐HBs and anti‐HBc) and nested HBV PCR assays targeting the HBV polymerase gene were performed, with HBV DNA positive samples being quantified with Cobas Taqman HBV test 48 assay (Roche Diagnostics). The study found that 63% (121/192) of patients had past or present HBV infection, and 40.6% (78/192) had detectable HBV DNA. Also, 22.9% (44/192) of patients were HBsAg positive and HBV DNA positive, while 23% (34/148) of HBsAG negatives had occult HBV infections. Of the 78 HBV DNA positive samples, 62.8% had viral loads ranging from 102 to ≥108 IU/ml, and 37.2% had HBV viral loads <200 IU/ml. There was a statistically significant positive association between HBsAg‐positivity and high viral loads, with 27% (12/44) of HBsAg positives having HBV viral loads between 104 and ≥108 IU/ml, compared to only 5.9% (2/34) of HBsAg negatives (relative risk: 4.64; 95% confidence interval: 1.11, 19.35; chi‐square P‐valueu2009=u20090.015). The study shows that the majority of HIV/AIDS patients initiating ART have either acute or chronic HBV infections, and further confirms that HIV remains a risk factor for occult HBV infections in South African patients as previously shown. The findings strongly support HBV screening in all HIV‐positive patients initiating ART in South Africa, considering that current ART regimens include drugs with anti‐HBV activity (e.g., lamivudine). J. Med. Virol. 81:406–412, 2009.

Evidence for a change in the epidemiology of hepatitis B virus infection after nearly two decades of universal hepatitis B vaccination in South Africa

The hepatitis B vaccine has been part of the South African Expanded Program on Immunization since April 1995 but its long‐term impact remains unknown. This study tested 1,206 sera collected from patients aged 1–25 years from various health facilities across the country for HBV serological markers and HBV DNA. Based on the year the vaccine was introduced, samples were stratified by age into pre‐ and post‐vaccine introduction populations, which were then compared for evidence of immunity and chronic carriage using the Chi‐square test. Where HIV status was known, subset analyses were performed. Immunity to HBV infection increased from 13.0% in the pre‐ to 57.0% in the post‐vaccine introduction population (Pu2009<u20090.001). This decreased with increasing age within the post‐vaccine introduction population (76.1% for 1–5 years, 50.0% for 6–10 years, and 46.3% for 11–16 years). In addition, HBV chronic carriage was significantly (Pu2009=u20090.003) reduced in the post‐ (1.4%) compared to the pre‐vaccine introduction population (4.2%). The difference in prevalence of active HBV infection in the serologically exposed pre‐ and post‐vaccine introduction populations was not statistically significant. Subset analyses showed that evidence of immunity was significantly (Pu2009<u20090.001) higher in the HIV negative compared to the HIV positive subset in both populations. Universal hepatitis B vaccination has been a remarkable success, with a significant increase in immunity to HBV infection. The observation that HBV chronic carriage increases as immunity wanes over time calls into question whether the time has come to consider a pre‐adolescence vaccine booster dose policy. J. Med. Virol. 86:918–924, 2014.

Near full-length genome analysis of HCV genotype 5 strains from South Africa.

Hepatitis C virus (HCV) genotype 5 is the predominant genotype in South Africa. However, to date, only 2 full-length genotype 5 genomes have been sequenced and only one is from South Africa. This study characterized HCV genotype 5 sequences from South Africa, including six near full-length genomes, as well as the E1 region from an additional 12 genotype 5 samples. Phylogenetic analysis of these near full-length genome sequences revealed that all genotype 5 sequences formed a close cluster with high bootstrap support. Bayesian analysis of the E1 region was used to estimate the time of the most recent common ancestor (tMRCA). The tMRCA for HCV genotype 5a was estimated at 114-134 years before the last sampling date. In conclusion, this study provides six near full-length genotype 5 nucleotide sequences for use as references to design efficient vaccines and for the development of new antiviral agents, and provides further insight into the diversity of HCV genotypes circulating in South Africa.

Introduction of new subtypes and variants of hepatitis C virus genotype 4 in South Africa.

Hepatitis C virus (HCV) genotype is an important predictor of disease progression and treatment response. This descriptive study investigated the sequence diversity and genotypes of HCV in South Africa based on comparative analysis of the 5′ untranslated region (UTR), C/E1, and NS5B regions of 60 sequences from 52 patients. Genotype distribution in the studied population was as follows: 54% (28/52) were genotype 5, 19% (10/52) were genotype 1, 19% (10/52) were genotype 4, and 2% (1/52) were genotype 3. Three of 52 (6%) individuals were infected with multiple genotypes based on the 5′UTR. Phylogenetic analysis of the 5′UTR was accurate in determining genotypes, while the C/E1 and NS5B coding region was able to differentiate both genotypes and subtypes, including an outlier group. Furthermore, this study observed the existence of distinct variants of HCV which were divergent from confirmed genotype 4 subtypes. For the first time in South Africa, this analysis has shown the presence of HCV subtypes 4k, 4q, and 4r, as well as evidence of intragenotypic recombinant 4l/4q within NS5B. In conclusion, while genotype 5a remains the predominant genotype in South Africa, the current study indicates the introduction of new subtypes and existence of variants of genotype 4 in South Africa. J. Med. Virol. 84:601–607, 2012.

Hepatitis B virus infection in post-vaccination South Africa : occult HBV infection and circulating surface gene variants

BACKGROUND AND OBJECTIVEnThe aim of this study was to investigate the prevalence of occult hepatitis B virus (HBV) infection and the HBV surface (S) gene variants circulating in the South African population after nearly two decades of universal hepatitis B vaccination.nnnSTUDY DESIGNnFrom a previous serosurvey, 201 serum samples with serological evidence of exposure to HBV were identified and these were stratified into post- and pre-vaccine introduction populations. For all samples, HBV DNA was screened and quantified using a real-time PCR assay and results analysed together with HBV serological markers. Where HIV results were available, subset analysis was performed. The HBV S gene was PCR-amplified and sequences analysed for a total of 37 isolates.nnnRESULTSnThe prevalence of occult HBV infection reduced from 70.4% in the pre-vaccine introduction era to 66.0% post-vaccine introduction. There was an association between HIV infection and an increase in prevalence of occult HBV infection within the post-vaccine introduction population, although this was not statistically significant. Furthermore, sequence analysis revealed the following HBV subgenotypes; A1 (n=34), A2 (n=2) and a rare D4 isolate. HBV S gene variants, including diagnostic escape mutants were isolated.nnnCONCLUSIONnThere was a decline in the prevalence of occult HBV infection in post-vaccination South Africa, although the disease burden remains significant in the HIV co-infected population. After nearly two decades of a universal hepatitis B vaccination programme, no positive selection of vaccine escape mutants were observed.

Prediction of T-cell epitopes of hepatitis C virus genotype 5a

BackgroundHepatitis C virus (HCV) is a public health problem with almost 185 million people estimated to be infected worldwide and is one of the leading causes of hepatocellular carcinoma. Currently, there is no vaccine for HCV infection and the current treatment does not clear the infection in all patients. Because of the high diversity of HCV, protective vaccines will have to overcome significant viral antigenic diversities. The objective of this study was to predict T-cell epitopes from HCV genotype 5a sequences.MethodsHCV near full-length protein sequences were analyzed to predict T-cell epitopes that bind human leukocyte antigen (HLA) class I and HLA class II in HCV genotype 5a using Propred I and Propred, respectively. The Antigenicity score of all the predicted epitopes were analysed using VaxiJen v2.0. All antigenic predicted epitopes were analysed for conservation using the IEDB database in comparison with 406, 221, 98, 33, 45, 45 randomly selected sequences from each of the HCV genotypes 1a, 1b, 2, 3, 4 and 6 respectively, downloaded from the GenBank. For epitope prediction binding to common HLA alleles found in South Africa, the IEDB epitope analysis tool was used.ResultsA total of 24 and 77 antigenic epitopes that bind HLA class I and HLA class II respectively were predicted. The highest number of HLA class I binding epitopes were predicted within the NS3 (63%), followed by NS5B (21%). For the HLA class II, the highest number of epitopes were predicted in the NS3 (30%) followed by the NS4B (23%) proteins. For conservation analysis, 8 and 31 predicted epitopes were conserved in different genotypes for HLA class I and HLA class II alleles respectively. Several epitopes bind with high affinity for both HLA class I alleles and HLA class II common in South Africa.ConclusionThe predicted conserved T-cell epitopes analysed in this study will contribute towards the future design of HCV vaccine candidates which will avoid variation in genotypes, which in turn will be capable of inducing broad HCV specific immune responses.

Functional analysis of ‘a’ determinant mutations associated with occult HBV in HIV-positive South Africans

Occult hepatitis B is defined by the presence of hepatitis B virus (HBV) DNA in the absence of hepatitis B surface antigen (HBsAg). Occult HBV is associated with the development of hepatocellular carcinoma, reactivation during immune suppression, and virus transmission. Viral mutations contribute significantly to the occult HBV phenotype. Mutations in the a determinant of HBsAg are of particular interest, as these mutations are associated with immune escape, vaccine escape and diagnostic failure. We examined the effects of selected occult HBV-associated mutations identified in a population of HIV-positive South Africans on HBsAg production in vitro. Mutations were inserted into two different chronic HBV backbones and transfected into a hepatocyte-derived cell line. HBsAg levels were quantified by enzyme-linked immunosorbent assay (ELISA), while the detectability of mutant HBsAg was determined using an HA-tagged HBsAg expression system. Of the seven mutations analysed, four (S132P, C138Y, N146D and C147Y) resulted in decreased HBsAg expression in one viral background but not in the second viral background. One mutation (N146D) led to a decrease in HBsAg detected as compared to HA-tag, indicating that this mutation compromises the ability of the ELISA to detect HBsAg. The contribution of occult-associated mutations to the HBsAg-negative phenotype of occult HBV cannot be determined adequately by testing the effect of the mutation in a single viral background, and rigorous analysis of these mutations is required.

Active co‐infection with HBV and/or HCV in South African HIV positive patients due for cancer therapy

Human immunodeficiency virus (HIV), Hepatitis B virus (HBV) and Hepatitis C virus (HCV) share routes of transmission. There is limited data on the incidence of active co‐infection with HBV and/or HCV in cancer patients infected with HIV in Africa. This was a prospective study based on 34 patients with varied cancer diagnosis, infected with HIV and awaiting cancer therapy in South Africa. HIV viral load, CD4+ cell counts, Alanine‐aminotransferase and aspartate aminotransferase levels were tested. Exposure to HBV and HCV was assessed serologically using commercial kits. Active HBV and/or HCV co‐infection was detected using viral specific nested PCR assays. HCV 5′‐UTR PCR products were sequenced to confirm active HCV infection. Active viral infection was detected in 64.7% of patients for HBV, 38.2% for HCV, and 29.4% for both HBV and HCV. Occult HBV infection was observed in 63.6% of the patients, while seronegative HCV infection was found in 30.8% of patients. In addition, CD4+ cell countu2009<u2009350 cells/µl was not a risk factor for increased active HBV, HCV or both HBV and HCV co‐infections. A total of 72.7%, 18.2% and 9.1% of the HCV sequences were assigned genotype 5, 1 and 4 respectively.The study revealed for the first time a high active HBV and/or HCV co‐infection rate in cancer patients infected with HIV. The findings call for HBV and HCV testing in such patients, and where feasible, appropriate antiviral treatment be indicated, as chemotherapy or radiotherapy has been associated with reactivation of viral hepatitis and termination of cancer therapy. J. Med. Virol. 87:213–221, 2015.

Should routine serological screening for HCV be mandatory in HIV/AIDS patients enrolling for HAART in South Africa?

To the Editor: Globally, an estimated 170 million people (about 3% of the worlds population) have been infected with hepatitis C virus (HCV). HCV, a member of the Hepacivirus genus in the Flaviviridae family, possesses a single stranded, positive-sense RNA genome of approximately 9.6 kilobase (kb), and is the major cause of chronic hepatitis, cirrhosis and hepatocellular carcinoma. In Africa, HCV prevalence ranges from <3% in South Africa to <20% in Egypt. In South Africa, there is a low prevalence from 0.16% to 1.8%, but this is higher in high-risk individuals, e.g. 39.4% in haemophiliacs and 4.8% in chronic dialysis patients.

High prevalence of active and occult hepatitis B virus infections in healthcare workers from two provinces of South Africa

BACKGROUNDnHepatitis B (HB) is a vaccine-preventable liver disease caused by infection with the blood-borne hepatitis B virus (HBV). South African healthcare workers (HCWs) may be at high risk of occupational exposure to HBV infection, since previous studies have found suboptimal levels of protection against HBV in HCWs.nnnMETHODSnA descriptive prevalence study based on self-administered questionnaires with data on demographics and HB vaccination status, and stored serum samples collected from 2009 to 2012, from 333 HCWs working or studying in Gauteng and Mpumalanga province hospitals or nursing colleges, was conducted. Samples were tested for HB surface antigen (HBsAg), antibodies to HBsAg (anti-HBs), antibodies to HB core antigen (anti-HBc), and HBV deoxyribonucleic acid (DNA).nnnRESULTSnThe majority of HCWs from whom the serum samples were drawn were black (91.4% [298/326]), female (82.6% [275/333]) and had received at least one dose of HB vaccine (70.9% [236/333]). The average age was 38.8years (range: 19-62). Of the HCWs, 23.2% (73/314) were susceptible (negative for all markers); 9.6% (30/314) were infected (HBsAg and/or DNA positive); 29.0% (91/314) were exposed (positive for either HBsAg, anti-HBc, or DNA); 18.8% (59/314) were immune due to natural infection (anti-HBs and anti-HBc positive only); while 47.8% (150/314) were immune due to vaccination (anti-HBs positive only). Furthermore, HBV DNA was detected in 8.6% (27/314) and occult HBV infection (OBI) (HBV DNA positive but HBsAg negative) was found in 6.7% (21/314) of samples.nnnDISCUSSION AND CONCLUSIONnThis study, which is the first to report OBI in South African HCWs, found high rates of active HBV infection and sub-optimal protection against HBV in HCWs. There is a need to strengthen vaccination programmes through a policy that ensures protection for all HCWs and their patients.