Vaccine-Induced Seroreactivity Impacts the Accuracy of HIV Testing Algorithms in Sub-Saharan Africa: An Exploratory Study.

The detection of vaccine-induced HIV antibody responses by rapid diagnostic tests (RDTs) may confound the interpretation of HIV testing results. We assessed the impact of vaccine-induced seroreactivity (VISR) on the diagnosis of HIV in sub-Saharan Africa. Samples collected from healthy participants of HIVIS and TaMoVac HIV vaccine trials after the final vaccination were analyzed for VISR using HIV testing algorithms used in Mozambique and Tanzania that employ two sequential RDTs. The samples were also tested for VISR using Enzygnost HIV Integral 4 ELISA and HIV western blot assays. Antibody titers to subtype C gp140 were determined using an in-house enzyme-linked immunosorbent assay (ELISA). The frequency of VISR was 93.4% (128/137) by Enzygnost HIV Integral 4 ELISA, and 66.4% (91/137) by western blot assay (WHO interpretation). The proportion of vaccine recipients that would have been misdiagnosed as HIV-positive in Mozambique was half of that in Tanzania: 26.3% (36/137) and 54.0% (74/137), respectively, p < 0.0001. In conclusion, the HIV RDTs and algorithms assessed here will potentially misclassify a large proportion of the HIV vaccine recipients if no other test is used. Increased efforts are needed to develop differential serological or molecular tools for use at the point of care.

Effect of 10-Valent Pneumococcal Conjugate Vaccine on Streptococcus pneumoniae Nasopharyngeal Carriage Among Children Less Than 5 Years Old: 3 Years Post-10-Valent Pneumococcal Conjugate Vaccine Introduction in Mozambique.

BACKGROUND: Mozambique introduced 10-valent pneumococcal conjugate vaccine (PCV10) in 2013 with doses at ages 2, 3, and 4 months and no catch-up or booster dose. We evaluated PCV10 impact on the carriage of vaccine-type (VT), non-VT, and antimicrobial non-susceptible pneumococci 3 years after introduction. METHODS: We conducted cross-sectional carriage surveys among HIV-infected and HIV-uninfected children aged 6 weeks to 59 months: 1 pre-PCV10 (2012-2013 [Baseline]) and 2 post-PCV10 introductions (2014-2015 [Post1] and 2015-2016 [Post2]). Pneumococci isolated from nasopharyngeal swabs underwent Quellung serotyping and antimicrobial susceptibility testing. Non-susceptible isolates (intermediate or resistant) were defined using Clinical and Laboratory Standards Institute 2018 breakpoints. We used log-binomial regression to estimate changes in the pneumococcal carriage between survey periods. We compared proportions of non-susceptible pneumococci between Baseline and Post2. RESULTS: We enrolled 720 children at Baseline, 911 at Post1, and 1208 at Post2. Baseline VT carriage was similar for HIV-uninfected (36.0%, 110/306) and HIV-infected children (34.8%, 144/414). VT carriage was 36% (95% confidence interval [CI]: 19%-49%) and 27% (95% CI: 11%-41%) lower in Post1 vs baseline among HIV-uninfected and HIV-infected children, respectively. VT carriage prevalence declined in Post2 vs Post1 for HIV-uninfected but remained stable for HIV-infected children. VT carriage prevalence 3 years after PCV10 introduction was 14.5% in HIV-uninfected and 21.0% in HIV-infected children. Pneumococcal isolates non-susceptible to penicillin declined from 66.0% to 56.2% (P= .0281) among HIV-infected children. CONCLUSIONS: VT and antimicrobial non-susceptible pneumococci carriage dropped after PCV10 introduction, especially in HIV-uninfected children. However, VT carriage remained common, indicating ongoing VT pneumococci transmission.

Dried blood spots for Streptococcus pneumoniae and Haemophilus influenzae detection and serotyping among children < 5 years old in rural Mozambique.

BACKGROUND: Dried blood spots (DBS) have been proposed as potentially tool for detecting invasive bacterial diseases. METHODS: We evaluated the use of DBS for S. pneumoniae and H. influenzae detection among children in Mozambique. Blood for DBS and nasopharyngeal (NP) swabs were collected from children with pneumonia and healthy aged < 5 years. Bacterial detection and serotyping were performed by quantitative PCR (qPCR) (NP and DBS; lytA gene for pneumococcus and hpd for H. influenzae) and culture (NP). Combined detection rates were compared between children with pneumonia and healthy. RESULTS: Of 325 children enrolled, 205 had pneumonia and 120 were healthy. Pneumococci were detected in DBS from 20.5 and 64.2% of children with pneumonia and healthy, respectively; NP specimens were positive for pneumococcus in 80.0 and 80.8%, respectively. H. influenzae was detected in DBS from 22.9% of children with pneumonia and 59.2% of healthy; 81.4 and 81.5% of NP specimens were positive for H. influenzae, respectively. CONCLUSION: DBS detected pneumococcal and H. influenzae DNA in children with pneumonia and healthy. Healthy children were often DBS positive for both bacteria, suggesting that qPCR of DBS specimens does not differentiate disease from colonization and is therefore not a useful diagnostic tool for children.

Invasive disease potential of Streptococcus pneumoniae serotypes before and after 10-valent pneumococcal conjugate vaccine introduction in a rural area, southern Mozambique.

BACKGROUND: Invasive pneumococcal disease (IPD) is a significant cause of morbidity and mortality among children worldwide. In April 2013, Mozambique introduced 10-valent PCV (PCV10) into the National Expanded Program on immunization using a three-dose schedule at 2, 3, and 4 months of age. We aimed to evaluate the invasive disease potential of pneumococcal serotypes among children in our region before and after PCV10 introduction. METHODS: We used data from ongoing population-based surveillance for IPD and cross-sectional pneumococcal carriage surveys among children aged <5 years in Manhiҫa, Mozambique. To determine the invasive disease potential for each serotype pre- and post-PCV10 introduction, odds ratios (OR) and 95% confidence intervals (95% CI) were calculated comparing serotype-specific prevalence in IPD and in carriage. For each serotype, OR and 95% CI > 1 indicated high invasive disease potential and OR and 95% CI < 1 indicated low invasive disease potential. RESULTS: In the pre-PCV10 period, 524 pneumococcal isolates were obtained from 411 colonized children and IPD cases were detected in 40 children. In the post-PCV10 period, 540 pneumococcal isolates were obtained from 507 colonized children and IPD cases were detected in 30 children. The most prevalent serotypes causing IPD pre-PCV10 were 6A (17.5%), 6B (15.0%), 14 (12.5%), 23F (10.0%) and 19F (7.5%), and post-PCV10 were 6A (36.7%), 13 (10%), 1 (10.0%), 6B (6.7%) and 19A (6.7%). Serotypes associated with high invasive disease potential pre-PCV10 included 1 (OR:22.3 [95% CI 2.0; 251.2]), 6B (OR:3.1 [95% CI 1.2; 8.1]), 14 (OR: 3.4 [95% CI 1.2; 9.8]) and post-PCV10 included serotype 6A (OR:6.1[95% CI 2.7; 13.5]). CONCLUSION: The number of serotypes with high invasive disease potential decreased after PCV10 introduction. Serotype 6A, which is not included in PCV10, was the most common cause of IPD throughout the study and showed a high invasive potential in the post-PCV10 period.

Optimizing the immunogenicity of HIV prime-boost DNA-MVA-rgp140/GLA vaccines in a phase II randomized factorial trial design.

BACKGROUND: We evaluated the safety and immunogenicity of (i) an intradermal HIV-DNA regimen given with/without intradermal electroporation (EP) as prime and (ii) the impact of boosting with modified vaccinia virus Ankara (HIV-MVA) administered with or without subtype C CN54rgp140 envelope protein adjuvanted with Glucopyranosyl Lipid A (GLA-AF) in volunteers from Tanzania and Mozambique. METHODS: Healthy HIV-uninfected adults (N = 191) were randomized twice; first to one of three HIV-DNA intradermal priming regimens by needle-free ZetaJet device at weeks 0, 4 and 12 (Group I: 2x0.1mL [3mg/mL], Group II: 2x0.1mL [3mg/mL] plus EP, Group III: 1x0.1mL [6mg/mL] plus EP). Second the same volunteers received 108 pfu HIV-MVA twice, alone or combined with CN54rgp140/GLA-AF, intramuscularly by syringe, 16 weeks apart. Additionally, 20 volunteers received saline placebo. RESULTS: Vaccinations and electroporation did not raise safety concerns. After the last vaccination, the overall IFN-γ ELISpot response rate to either Gag or Env was 97%. Intradermal electroporation significantly increased ELISpot response rates to HIV-DNA-specific Gag (66% group I vs. 86% group II, p = 0.026), but not to the HIV-MVA vaccine-specific Gag or Env peptide pools nor the magnitude of responses. Co-administration of rgp140/GLA-AF with HIV-MVA did not impact the frequency of binding antibody responses against subtype B gp160, C gp140 or E gp120 antigens (95%, 99%, 79%, respectively), but significantly enhanced the magnitude against subtype B gp160 (2700 versus 300, p<0.001) and subtype C gp140 (24300 versus 2700, p<0.001) Env protein. At relatively low titers, neutralizing antibody responses using the TZM-bl assay were more frequent in vaccinees given adjuvanted protein boost. CONCLUSION: Intradermal electroporation increased DNA-induced Gag response rates but did not show an impact on Env-specific responses nor on the magnitude of responses. Co-administration of HIV-MVA with rgp140/GLA-AF significantly enhanced antibody responses.

Intradermal HIV-1 DNA Immunization Using Needle-Free Zetajet Injection Followed by HIV-Modified Vaccinia Virus Ankara Vaccination Is Safe and Immunogenic in Mozambican Young Adults: A Phase I Randomized Controlled Trial.

We assessed the safety and immunogenicity of HIV-DNA priming using Zetajet™, a needle-free device intradermally followed by intramuscular HIV-MVA boosts, in 24 healthy Mozambicans. Volunteers were randomized to receive three immunizations of 600 µg (n = 10; 2 × 0.1 ml) or 1,200 µg (n = 10; 2 × 0.2 ml) of HIV-DNA (3 mg/ml), followed by two boosts of 108 pfu HIV-MVA. Four subjects received placebo saline injections. Vaccines and injections were safe and well tolerated with no difference between the two priming groups. After three HIV-DNA immunizations, IFN-γ ELISpot responses to Gag were detected in 9/17 (53%) vaccinees, while none responded to Envelope (Env). After the first HIV-MVA, the overall response rate to Gag and/or Env increased to 14/15 (93%); 14/15 (93%) to Gag and 13/15 (87%) to Env. There were no significant differences between the immunization groups in frequency of response to Gag and Env or magnitude of Gag responses. Env responses were significantly higher in the higher dose group (median 420 vs. 157.5 SFC/million peripheral blood mononuclear cell, p = .014). HIV-specific antibodies to subtype C gp140 and subtype B gp160 were elicited in all vaccinees after the second HIV-MVA, without differences in titers between the groups. Neutralizing antibody responses were not detected. Two (13%) of 16 vaccinees, one in each of the priming groups, exhibited antibodies mediating antibody-dependent cellular cytotoxicity to CRF01_AE. In conclusion, HIV-DNA vaccine delivered intradermally in volumes of 0.1-0.2 ml using Zetajet was safe and well tolerated. Priming with the 1,200 µg dose of HIV-DNA generated higher magnitudes of ELISpot responses to Env.

First description of HTLV-1/2 seroprevalence in HIV-infected inmates in Mozambique.

No study has yet been conducted to estimate the burden of co-infection of HIV and HTLV-1/2 in inmates in sub-Saharan Africa. To investigate prevalence of co-infection in inmates in Mozambique, a total of 2140 inmates were screened for HIV, of which 515 were HIV seropositive. All HIV seropositive inmates were further screened for HTLV infection, and eight (1.55%) were co-infected. Co-infection was higher in females (3.45% [2/58; CI: 0.42-11.91]) as compared to males (1.35% [6/445; CI: 0.55-3.06]). Early screening of HTLV in prisons is urgently needed in Mozambique in order to improve the care provided to incarcerated individuals, including initiation of ART.

Incidence of HIV and the prevalence of HIV, hepatitis B and syphilis among youths in Maputo, Mozambique: a cohort study.

BACKGROUND: Prevalence of HIV in Mozambique among individuals aged 15-49 years is 11.5%. The HIV prevalence is higher in women than in men across the country, peaking at ages 25-29 years and 35-39 years, respectively. In this study, we aimed at determining the prevalence and incidence of HIV, prevalence of Hepatitis B (HBV), and prevalence of syphilis in youths. We also characterized a cohort of youths for future participation in phase I/II HIV vaccine trials. METHODS: The study was conducted at a youth clinic in Maputo Central Hospital from August 2009 to October 2011. Youths of both genders aged 18-24 years (n = 1380) were screened for HIV using a sequential algorithm of two immunochromatographic assays, HBV using an enzyme linked immunosorbant test, and syphilis using a treponemal immunochromatographic strip test. The HIV seronegative participants (n = 1309) were followed-up for 12 months with quarterly study visits. The clinical and behavioral data were collected using structured questionnaires. The HIV seroconversions were confirmed by a molecular assay. RESULTS: The study population was female dominant (76.8%). All participants had a formal education, with 44.6% studying for technical or higher education degrees. The mean age at sexual debut was 16.6 years (SD: ± 1.74), with 85.6% reporting more than one sexual partner in life. The screening showed the prevalence of HIV, HBV, and syphilis at 5.1% (95% CI: 3.97-6.31), 12.2% (95% CI 10.5%-14.0%), and 0.36% (95% CI 0.15%-0.84%), respectively. The HIV incidence rate was found to be 1.14/100 person years (95% CI: 0.67-1.92). Retention rates were stable throughout the study being 85.1% at the last visit. CONCLUSION: Incidence of HIV in this cohort of youths in Maputo was relatively low. Also, the prevalence of HIV and syphilis was lower than the national values in this age group. However, the HBV prevalence was higher than in previous reports in the country.

Reference values for clinical laboratory parameters in young adults in Maputo, Mozambique.

BACKGROUND: Clinical laboratory reference values from North American and European populations are currently used in most Africans countries due to the absence of locally derived reference ranges, despite previous studies reporting significant differences between populations. Our aim was to define reference ranges for both genders in 18 to 24 year-old Mozambicans in preparation for clinical vaccine trials. METHODS: A cross-sectional study including 257 volunteers (102 males and 155 females) between 18 and 24 years was performedat a youth clinic in Maputo, Mozambique. All volunteers were clinically healthy and human immunodeficiency virus, Hepatitis B virus and syphilis negative.Median and 95% reference ranges were calculated for immunological, hematological and chemistry parameters. Ranges were compared with those reported based on populations in other African countries and the US. The impact of applying US NIH Division of AIDS (DAIDS) toxicity tables was assessed. RESULTS: The immunology ranges were comparable to those reported for the US and western Kenya.There were significant gender differences in CD4+ T cell values 713 cells/µL in males versus 824 cells/µL in females (p<0.0001). Hematologic values differed from the US values but were similar to reports of populations in western Kenya and Uganda. The lower and upper limits of the ranges for hemoglobin, hematocrit, red blood cells, white blood cells and lymphocytes were somewhat lower than those from these African countries. The chemistry values were comparable to US values, with few exceptions. The upper limits for ALT, AST, bilirubin, cholesterol and triglycerides were higher than those from the US. DAIDStables for adverse events predicted 297 adverse events and 159 (62%) of the volunteers would have been excluded. CONCLUSION: This study is the first to determine normal laboratory parameters in Mozambique. Our results underscore the necessity of establishing region-specific clinical reference ranges for proper patient management and safe conduct of clinical trials.

Absolute and percent CD4+ T-cell enumeration by flow cytometry using capillary blood.

INTRODUCTION: CD4+ T-cell counting is usually performed on whole blood obtained from standard venipuncture. Venipuncture requires expertise, results in discomfort and generates biological waste. Capillary blood could be used to measure the levels of CD4+ T-cell in children, elderly and very ill patients. We studied the agreement between CD4+ T-cell counts and percent generated using venous blood with those obtained with capillary blood in HIV-infected adults and children in a resource-limited tropical setting. METHODS: This cross-sectional study consecutively enrolled a total of 152 adult and pediatric HIV-positive patients attending two outpatient clinics in Maputo City, Mozambique. We recruited individuals presenting for their routine clinical follow-up that included the determination of CD4+ T-cell counts in peripheral blood. For each subject, peripheral blood specimens were obtained by both venipuncture and finger prick. Specimens were tested using two flow cytometers, the FACSCount and the FACSCalibur. RESULTS: Absolute CD4+ T-cell counts obtained using capillary blood were in close agreement with those from venous blood both on the FACSCalibur (absolute bias=+12.3 cells/mm³, limits of agreement: -259.2 to +283.9, R²=0.96) and the FACSCount (absolute bias=+16.1 cells/mm³, limits of agreement: -209.2 to +241.5, R²=0.97). Percent CD4+ T-cell counts were measured only on the FACSCalibur also showed a good agreement with a bias of +0.6% and limits of agreement of -3.1 to +4.3. CONCLUSIONS: Absolute CD4+ T-cell counts and percent generated using capillary blood are in close agreement with those from venous blood. Point-Of-Care assays and standard flow cytometers can be deployed in a tiered laboratory network where both venous and capillary blood collection can be used for CD4+ T-cell enumeration.