Hepatitis B prevalence and incidence in the fishing communities of Lake Victoria, Uganda: a retrospective cohort study.

INTRODUCTION: Hepatitis B is a serious potentially fatal hepatocellular disease caused by the hepatitis B virus. In the fishing communities of Lake Victoria Uganda, the hepatitis B virus infection burden is largely unknown. This study assessed the prevalence and incidence of hepatitis B in these communities. METHODS: This was a retrospective cohort study that tested serum samples collected from 13 to 49-year-old study participants that were residing in two Ugandan Lake Victoria fishing communities of Kasenyi (a mainland) and Jaana (an island). The samples were collected between 2013 and 2015 during the conduct of an HIV epidemiological cohort study in these communities. A total of 467 twelve-month follow-up and 50 baseline visit samples of participants lost to follow-up were tested for hepatitis B serological markers to determine prevalence. To determine hepatitis B virus incidence, samples that were hepatitis B positive at the follow-up visit had their baseline samples tested to identify hepatitis B negative samples whose corresponding follow-up samples were thus incident cases. RESULTS: The baseline mean age of the 517 study participants was 31.1 (SD ± 8.4) years, 278 (53.8%) of whom were females. A total of 36 (7%) study participants had hepatitis B virus infection, 22 (61.1%) of whom were male. Jaana had a higher hepatitis B virus prevalence compared to Kasenyi (10.2% vs 4.0%). In total, 210 (40.6%) study participants had evidence of prior hepatitis B virus infection while 48.6% had never been infected or vaccinated against this disease. A total of 20 (3.9%) participants had results suggestive of prior hepatitis B vaccination. Hepatitis B incidence was 10.5 cases/100PY (95% CI: 7.09-15.53). Being above 25 years of age and staying in Jaana were significant risk factors for hepatitis B virus acquisition (AOR 1.6, 95% CI: 1.1-2.2; p < 0.01 and 1.4, 95% CI: 1.1-1.8; p < 0.01 respectively). CONCLUSION: Hepatitis B virus incidence in Lake Victoria fishing communities of Uganda is very high, particularly in the islands. Interventions to lower hepatitis B virus transmission in these communities are urgently needed.

An assessment of fishing communities around Lake Victoria, Uganda, as potential populations for future HIV vaccine efficacy studies: an observational cohort study.

BACKGROUND: An effective HIV vaccine is still elusive. Of the 9 HIV preventive vaccine efficacy trials conducted to-date, only one reported positive results of modest efficacy. More efficacy trials need to be conducted before one or more vaccines are eventually licensed. We assessed the suitability of fishing communities in Uganda for future HIV vaccine efficacy trials. METHODS: A community-based cohort study was conducted among a random sample of 2191 participants aged 18-49 years. Data were collected on socio-demographic characteristics, HIV risky behaviors, and willingness to participate in future HIV vaccine trials (WTP). Venous blood was collected for HIV serological testing. Retention/follow rates and HIV incidence rates per 100 person years at-risk (pyar) were estimated. Adjusted prevalence proportion ratios (PPRs) of retention and odds ratios (ORs) of lack of WTP were estimated using log-binomial and logistic regression models respectively. RESULTS: Overall retention rate was 76.9% (1685/2191), highest (89%) among participants who had spent 5+ years in the community and lowest (54.1%) among those with <1 year stay. Significant predictors of retention included tribe/ethnicity, baseline HIV negative status, and longer than 1 year stay in the community. Overall WTP was 89.1% (1953/2191). Lack of WTP was significantly higher among women than men [adj.OR = 1.51 (95% CI, 1.14- 2.00)] and among participants who had stayed in fishing communities for 10 or more years relative to those with less than one year [adj.OR = 1.78 (95% CI, 1.11 - 2.88)]. Overall HIV incidence rate per 100 pyar was 3.39 (95% CI; 2.55 - 4.49). Participants aged 25-29 years had highest incidence rates (4.61 - 7.67/100 pyar) and high retention rates between 78.5 and 83.1%. In a combined analysis of retention and incidence rates participants aged 30+ years had retention rates ~80% but low incidence rates (2.45 - 3.57 per 100 pyar) while those aged 25-29 years had the highest incidence rates (4.61 - 7.67/100 pyar) and retention rates 78.5 - 83.1%. CONCLUSIONS: There is high HIV incidence, retention and WTP among fishing communities around L. Victoria, Uganda which make these communities appropriate for future HIV prevention efficacy studies including vaccine trials.

High proportion of Ugandans with pre-pandemic SARS-CoV-2 cross-reactive CD4+ and CD8+ T-cell responses.

The estimated mortality rate of the SARS-CoV-2 pandemic varied greatly around the world with multiple countries in East, Central, and West Africa having significantly lower rates of COVID-19 related fatalities than many resource-rich nations with significantly earlier wide-spread access to life-saving vaccines. One possible reason for this lower mortality could be the presence of pre-existing cross-reactive immunological responses in these areas of the world. To explore this hypothesis, stored peripheral blood mononuclear cells (PBMC) from Ugandans collected from 2015-2017 prior to the COVID-19 pandemic (n=29) and from hospitalized Ugandan COVID-19 patients (n=3) were examined using flow-cytometry for the presence of pre-existing SARS-CoV-2 cross-reactive CD4+ and CD8+ T-cell populations using four T-cell epitope mega pools. Of pre-pandemic participants, 89.7% (26/29) had either CD4+ or CD8+, or both, SARS-CoV-2 specific T-cell responses. Specifically, CD4+ T-cell reactivity (72.4%) and CD8+ T-cell reactivity (65.5%) were relatively similar, and 13 participants (44.8%) had both types of cross-reactive types of T-cells present. There were no significant differences in response by sex in the population. The rates of cross-reactive T-cell populations in these Ugandans is higher than previous estimates from resource-rich countries like the United States (20-50% reactivity). It is unclear what role, if any, this cross-reactivity played in decreasing COVID-19 related mortality in Uganda and other African countries, but does suggest that a better understanding of global pre-existing immunological cross-reactivity could be an informative data of epidemiological intelligence moving forward.

Proportion of Ugandans with pre-pandemic SARS-CoV-2 cross-reactive CD4+ and CD8+ T-cell responses: A pilot study.

The estimated mortality rate of the SARS-CoV-2 pandemic varied greatly around the world. In particular, multiple countries in East, Central, and West Africa had significantly lower rates of COVID-19 related fatalities than many resource-rich nations with significantly earlier wide-spread access to life-saving vaccines. One possible reason for this lower mortality could be the presence of pre-existing cross-reactive immunological responses in these areas of the world. To explore this hypothesis, an exploratory study of stored peripheral blood mononuclear cells (PBMC) from Ugandans collected from 2015-2017 prior to the COVID-19 pandemic (n = 29) and from hospitalized Ugandan COVID-19 patients (n = 3) were examined using flow-cytometry for the presence of pre-existing SARS-CoV-2 cross-reactive CD4+ and CD8+ T-cell populations using four T-cell epitope mega pools. Of pre-pandemic participants, 89.7% (26/29) had either CD4+ or CD8+, or both, SARS-CoV-2 specific T-cell responses. Specifically, CD4+ T-cell reactivity (72.4%) and CD8+ T-cell reactivity (65.5%) were relatively similar, and 13 participants (44.8%) had both types of cross-reactive types of T-cells present. There were no significant differences in response by sex in the population, however this may be in part due to the limited sample size examined. The rates of cross-reactive T-cell populations in this exploratory Ugandan population appears higher than previous estimates from resource-rich countries like the United States (20-50% reactivity). It is unclear what role, if any, this cross-reactivity played in decreasing COVID-19 related mortality in Uganda and other African countries, but does suggest that a better understanding of global pre-existing immunological cross-reactivity could be an informative data of epidemiological intelligence moving forward.

Schistosoma mansoni infection alters the host pre-vaccination environment resulting in blunted Hepatitis B vaccination immune responses.

The impact of endemic infections on protective immunity is critical to inform vaccination strategies. In this study, we assessed the influence of Schistosoma mansoni infection on host responses in a Ugandan fishing cohort given a Hepatitis B (HepB) vaccine. Concentrations of schistosome-specific circulating anodic antigen (CAA) pre-vaccination showed a significant bimodal distribution associated with HepB titers, which were lower in individuals with high CAA. We established that participants with high CAA had significantly lower frequencies of circulating T follicular helper (cTfh) subpopulations pre- and post-vaccination and higher regulatory T cells (Tregs) post-vaccination. Polarization towards higher frequencies of Tregs: cTfh cells can be mediated by changes in the cytokine environment favoring Treg differentiation. In fact, we observed higher levels of CCL17 and soluble IL-2R pre-vaccination (important for Treg recruitment and development), in individuals with high CAA that negatively associated with HepB titers. Additionally, alterations in pre-vaccination monocyte function correlated with HepB titers, and changes in innate-related cytokines/chemokine production were associated with increasing CAA concentration. We report, that by influencing the immune landscape, schistosomiasis has the potential to modulate immune responses to HepB vaccination. These findings highlight multiple Schistosoma -related immune associations that could explain abrogated vaccine responses in communities with endemic infections. Author Summary: Schistosomiasis drives host immune responses for optimal pathogen survival, potentially altering host responses to vaccine-related antigen. Chronic schistosomiasis and co-infection with hepatotropic viruses are common in countries where schistosomiasis is endemic. We explored the impact of Schistosoma mansoni ( S. mansoni ) infection on Hepatitis B (HepB) vaccination of individuals from a fishing community in Uganda. We demonstrate that high schistosome-specific antigen (circulating anodic antigen, CAA) concentration pre-vaccination, is associated with lower HepB antibody titers post-vaccination. We show higher pre-vaccination levels of cellular and soluble factors in instances of high CAA that are negatively associated with HepB antibody titers post-vaccination, which coincided with lower frequencies of circulating T follicular helper cell populations (cTfh), proliferating antibody secreting cells (ASCs), and higher frequencies of regulatory T cells (Tregs). We also show that monocyte function is important in HepB vaccine responses, and that high CAA is associated with alterations in the early innate cytokine/chemokine microenvironment. Our findings suggest that in individuals with high CAA and likely high worm burden, schistosomiasis creates and sustains an environment that is polarized against optimal host immune responses to the vaccine, which puts many endemic communities at risk for infection against HepB and other diseases that are preventable by vaccines.

Schistosoma mansoni infection alters the host pre-vaccination environment resulting in blunted Hepatitis B vaccination immune responses.

Schistosomiasis is a disease caused by parasitic flatworms of the Schistosoma spp., and is increasingly recognized to alter the immune system, and the potential to respond to vaccines. The impact of endemic infections on protective immunity is critical to inform vaccination strategies globally. We assessed the influence of Schistosoma mansoni worm burden on multiple host vaccine-related immune parameters in a Ugandan fishing cohort (n = 75) given three doses of a Hepatitis B (HepB) vaccine at baseline and multiple timepoints post-vaccination. We observed distinct differences in immune responses in instances of higher worm burden, compared to low worm burden or non-infected. Concentrations of pre-vaccination serum schistosome-specific circulating anodic antigen (CAA), linked to worm burden, showed a significant bimodal distribution associated with HepB titers, which was lower in individuals with higher CAA values at month 7 post-vaccination (M7). Comparative chemokine/cytokine responses revealed significant upregulation of CCL19, CXCL9 and CCL17 known to be involved in T cell activation and recruitment, in higher CAA individuals, and CCL17 correlated negatively with HepB titers at month 12 post-vaccination. We show that HepB-specific CD4+ T cell memory responses correlated positively with HepB titers at M7. We further established that those participants with high CAA had significantly lower frequencies of circulating T follicular helper (cTfh) subpopulations pre- and post-vaccination, but higher regulatory T cells (Tregs) post-vaccination, suggesting changes in the immune microenvironment in high CAA could favor Treg recruitment and activation. Additionally, we found that changes in the levels of innate-related cytokines/chemokines CXCL10, IL-1ß, and CCL26, involved in driving T helper responses, were associated with increasing CAA concentration. This study provides further insight on pre-vaccination host responses to Schistosoma worm burden which will support our understanding of vaccine responses altered by pathogenic host immune mechanisms and memory function and explain abrogated vaccine responses in communities with endemic infections.

Performance of International AIDS Vaccine Initiative African clinical research laboratories in standardised ELISpot and peripheral blood mononuclear cell processing in support of HIV vaccine clinical trials.

BACKGROUND: Standardisation of procedures for performing cellular functional assays across laboratories participating in multicentre clinical trials is key for generating comparable and reliable data. OBJECTIVE: This article describes the performance of accredited laboratories in Africa and Europe on testing done in support of clinical trials. METHODS: For enzyme-linked immunospot assay (ELISpot) proficiency, characterised peripheral blood mononuclear cells (PBMCs) obtained from 48 HIV-negative blood donors in Johannesburg, South Africa, were sent to participating laboratories between February 2010 and February 2014. The PBMCs were tested for responses against cytomegalovirus, Epstein Barr and influenza peptide pools in a total of 1751 assays. In a separate study, a total of 1297 PBMC samples isolated from healthy HIV-negative participants in clinical trials of two prophylactic HIV vaccine candidates in Kenya, Uganda, Rwanda and Zambia were analysed for cell viability, cell yield and cell recovery from frozen PBMCs. RESULTS: Most (99%) of the 1751 ELISpot proficiency assays had data within acceptable ranges with low responses to mock stimuli. No significant statistical difference were observed in ELISpot responses at the five laboratories actively conducting immunological analyses. Of the 1297 clinical trial PBMCs processed, 94% had cell viability above 90% and 96% had cell yield above 0.7 million per mL of blood in freshly isolated cells. All parameters were within the predefined acceptance criteria. CONCLUSION: We demonstrate that multiple laboratories can generate reliable, accurate and comparable data by using standardised procedures, having regular training, having regular equipment maintenance and using centrally sourced reagents.

Characterization of CD4+ T cell subsets and HIV susceptibility in the inner and outer foreskin of Ugandan men.

PROBLEM: Biological mechanisms of foreskin HIV acquisition are poorly defined. The inner foreskin is preferentially infected in explant models, so we hypothesized that this site would be enriched for HIV-susceptible CD4+ T cells and proinflammatory/chemoattractant cytokines. METHOD OF STUDY: A total of 42 HIV-uninfected Ugandan men without genital symptoms provided foreskin tissues and swabs at the time of elective penile circumcision. The immune phenotype of foreskin-derived CD4+ T cells and entry of a CCR5-tropic HIV pseudovirus was characterized, and specific cytokine levels assayed by multiplexed chemiluminescent ELISA. RESULTS: Unexpectedly, outer foreskin CD4+ T cells more frequently expressed CCR5 (median 29.2% vs 22.9%, P = 0.01) and CD69 (median 36.5% vs 15%, P < 0.01), and on a per-cell basis, HIV entry was higher. However, overall CD4+ T cell density was approximately twofold higher in the inner foreskin, and several highly susceptible T cell subsets were increased at this site, including Th17 cells (20.0% vs 14.1%, P = 0.0021). Specific pro-inflammatory cytokine levels were also higher on the inner foreskin surface (IL-17, IL-8, RANTES and IL-1ß; all P < 0.05). CONCLUSION: There was marked heterogeneity in CD4+ T cell populations and immune milieu between inner and outer foreskin tissues. Despite higher per-cell viral entry into CD4+ T cells from the outer foreskin, the higher target cell density and enriched pro-inflammatory cytokines of the inner foreskin suggest that this may be a preferential site for HIV acquisition.

Schistosoma mansoni Infection in Ugandan Men Is Associated with Increased Abundance and Function of HIV Target Cells in Blood, but Not the Foreskin: A Cross-sectional Study.

BACKGROUND: Schistosoma mansoni infection has been associated with an increased HIV prevalence in humans and SHIV incidence in primate models. We hypothesized that immune activation from this gastrointestinal mucosa infection would increase highly HIV-susceptible CD4 T cell subsets in the blood and the foreskin through common mucosal homing. METHODOLOGY/PRINCIPAL FINDINGS: Foreskin tissue and blood were obtained from 34 HIV- and malaria-uninfected Ugandan men who volunteered for elective circumcision, 12 of whom were definitively positive for S. mansoni eggs in stool and 12 definitively negative for both S. mansoni eggs and worm antigen. Tissue and blood T cell subsets were characterized by flow cytometry and immunohistochemistry (IHC). Th17 and Th1 cells from both the blood and foreskin expressed higher levels of CCR5 and were more activated than other CD4 T cell subsets. S. mansoni-infected men had a higher frequency of systemic Th1 cells (22.9 vs. 16.5% of blood CD4 T cells, p<0.05), Th17 cells (2.3 vs. 1.5%, p<0.05), and Th22 cells (0.5 vs. 0.3%, p<0.01) than uninfected men. Additionally, Th17 cells in the blood of S. mansoni-infected men demonstrated enhanced function (28.1 vs. 16.3% producing multiple cytokines, p = 0.046). However, these immune alterations were not observed in foreskin tissue. CONCLUSIONS/SIGNIFICANCE: S. mansoni infection was associated with an increased frequency of highly HIV-susceptible Th1, Th17 and Th22 cell subsets in the blood, but these T cell immune differences did not extend to the foreskin. S. mansoni induced changes in T cell immunology mediated through the common mucosal immune system are not likely to increase HIV susceptibility in the foreskin.

Immune activation alters cellular and humoral responses to yellow fever 17D vaccine.

BACKGROUND: Defining the parameters that modulate vaccine responses in African populations will be imperative to design effective vaccines for protection against HIV, malaria, tuberculosis, and dengue virus infections. This study aimed to evaluate the contribution of the patient-specific immune microenvironment to the response to the licensed yellow fever vaccine 17D (YF-17D) in an African cohort. METHODS: We compared responses to YF-17D in 50 volunteers in Entebbe, Uganda, and 50 volunteers in Lausanne, Switzerland. We measured the CD8+ T cell and B cell responses induced by YF-17D and correlated them with immune parameters analyzed by flow cytometry prior to vaccination. RESULTS: We showed that YF-17D-induced CD8+ T cell and B cell responses were substantially lower in immunized individuals from Entebbe compared with immunized individuals from Lausanne. The impaired vaccine response in the Entebbe cohort associated with reduced YF-17D replication. Prior to vaccination, we observed higher frequencies of exhausted and activated NK cells, differentiated T and B cell subsets and proinflammatory monocytes, suggesting an activated immune microenvironment in the Entebbe volunteers. Interestingly, activation of CD8+ T cells and B cells as well as proinflammatory monocytes at baseline negatively correlated with YF-17D-neutralizing antibody titers after vaccination. Additionally, memory T and B cell responses in preimmunized volunteers exhibited reduced persistence in the Entebbe cohort but were boosted by a second vaccination. CONCLUSION: Together, these results demonstrate that an activated immune microenvironment prior to vaccination impedes efficacy of the YF-17D vaccine in an African cohort and suggest that vaccine regimens may need to be boosted in African populations to achieve efficient immunity. TRIAL REGISTRATION: Registration is not required for observational studies. FUNDING: This study was funded by Canada's Global Health Research Initiative, Defense Threat Reduction Agency, National Institute of Allergy and Infectious Diseases, Bill & Melinda Gates Foundation, and United States Agency for International Development.