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The radiation-attenuated Plasmodium falciparum sporozoites (PfSPZ) Vaccine has demonstrated safety and immunogenicity in 5-month-old to 50-year-old Africans in multiple trials. Except for one, each trial has restricted enrollment to either infants and children or adults < 50 years old. This trial was conducted in Equatorial Guinea and assessed the safety, tolerability, and immunogenicity of three direct venous inoculations of 1.8 × 106 or 2.7 × 106 PfSPZ, of PfSPZ Vaccine, or normal saline administered at 8-week intervals in a randomized, double-blind, placebo-controlled trial stratified by age (6–11 months and 1–5, 6–10, 11–17, 18–35, and 36–61 years). All doses were successfully administered. In all, 192/207 injections (93%) in those aged 6–61 years were rated as causing no or mild pain. There were no significant differences in solicited adverse events (AEs) between vaccinees and controls in any age group (P ≥ 0.17). There were no significant differences between vaccinees and controls with respect to the rates or severity of unsolicited AEs or laboratory abnormalities. Development of antibodies to P. falciparum circumsporozoite protein occurred in 67/69 vaccinees (97%) and 0/15 controls. Median antibody levels were highest in infants and 1–5-year-olds and declined progressively with age. Antibody responses in children were greater than in adults protected against controlled human malaria infection. Robust immunogenicity, combined with a benign AE profile, indicates children are an ideal target for immunization with PfSPZ Vaccine.

Plasmodium falciparum sporozoite (PfSPZ) Vaccine (radiation-attenuated, aseptic, purified, cryopreserved PfSPZ) and PfSPZ-CVac (infectious, aseptic, purified, cryopreserved PfSPZ administered to subjects taking weekly chloroquine chemoprophylaxis) have shown vaccine efficacies (VEs) of 100% against homologous controlled human malaria infection (CHMI) in nonimmune adults. Plasmodium falciparum sporozoite-CVac has never been assessed against CHMI in African vaccinees. We assessed the safety, immunogenicity, and VE against homologous CHMI of three doses of 2.7 × 106 PfSPZ of PfSPZ Vaccine at 8-week intervals and three doses of 1.0 × 105 PfSPZ of PfSPZ-CVac at 4-week intervals with each arm randomized, double-blind, placebo-controlled, and conducted in parallel. There were no differences in solicited adverse events between vaccinees and normal saline controls, or between PfSPZ Vaccine and PfSPZ-CVac recipients during the 6 days after administration of investigational product. However, from days 7–13, PfSPZ-CVac recipients had significantly more AEs, probably because of Pf parasitemia. Antibody responses were 2.9 times higher in PfSPZ Vaccine recipients than PfSPZ-CVac recipients at time of CHMI. Vaccine efficacy at a median of 14 weeks after last PfSPZ-CVac dose was 55% (8 of 13, P = 0.051) and at a median of 15 weeks after last PfSPZ Vaccine dose was 27% (5 of 15, P = 0.32). The higher VE in PfSPZ-CVac recipients of 55% with a 27-fold lower dose was likely a result of later stage parasite maturation in the liver, leading to induction of cellular immunity against a greater quantity and broader array of antigens.

Plasmodium falciparum sporozoite (PfSPZ) Vaccine is composed of radiation-attenuated, aseptic, purified cryopreserved PfSPZ. Multiple clinical trials empirically assessing two to six doses have shown multi-dose priming (two to four doses the first week) to be optimal for protection in both 4- and 16-week regimens. In this randomized, double-blind, normal saline (NS) placebo-controlled trial, four groups (G) of 18- to 32-year-old Equatoguineans received multi-dose priming regimens with or without a delayed final dose at 4 or 16 weeks. The regimens were G1: days 1, 3, 5, 7, and 113; G2: days 1, 3, 5, and 7; G3: days 1, 3, 5, 7, and 29; and G4: days 1, 8, and 29. All doses were 9 × 105 PfSPZ. Tolerability, safety, immunogenicity, and vaccine efficacy (VE) against homologous controlled human malaria infection (CHMI) 6–7 weeks after vaccination were assessed to down-select the best regimen. All four regimens were safe and well tolerated, with no significant differences in adverse events (AEs) between vaccinees (N = 84) and NS controls (N = 20) or between regimens. Out of 19 controls, 13 developed Pf parasitemia by quantitative polymerase chain reaction (qPCR) after CHMI. Only the vaccine regimen administered on study days 1, 8, and 29 gave significant protection (7/21 vaccinees versus 13/19 controls infected, VE 51.3%, P = 0.03, Barnard’s test, two-tailed). There were no significant differences in antibodies against Pf circumsporozoite protein (PfCSP), a major SPZ antigen, between protected and nonprotected vaccinees or controls pre-CHMI. The six controls not developing Pf parasitemia had significantly higher antibodies to blood stage antigens Pf exported protein 1 (PfEXP1) and Pf merozoite surface protein 1 (PfMSP1) than the controls who developed parasitemia, suggesting naturally acquired immunity against Pf limited infections in controls. This study identified a safe, protective, 4-week, multi-dose prime vaccination regimen for assessment in future trials of PfSPZ Vaccine.

In 2016, there were more cases and deaths caused by malaria globally than in 2015. An effective vaccine would be an ideal additional tool for reducing malaria’s impact. Sanaria® PfSPZ Vaccine, composed of radiation-attenuated, aseptic, purified, cryopreserved Plasmodium falciparum (Pf) sporozoites (SPZ) has been well tolerated and safe in malaria-naïve and experienced adults in the United States and Mali and protective against controlled human malaria infection with Pf in the United States and field transmission of Pf in Mali, but had not been assessed in younger age groups. We, therefore, evaluated PfSPZ Vaccine in 93 Tanzanians aged 45 years to 6 months in a randomized, double-blind, normal saline placebo-controlled trial. There were no significant differences in adverse events between vaccinees and controls or between dosage regimens. Because all age groups received three doses of 9.0 × 105 PfSPZ of PfSPZ Vaccine, immune responses were compared at this dosage. Median antibody responses against Pf circumsporozoite protein and PfSPZ were highest in infants and lowest in adults. T-cell responses were highest in 6–10-year olds after one dose and 1–5-year olds after three doses; infants had no significant positive T-cell responses. The safety data were used to support initiation of trials in > 300 infants in Kenya and Equatorial Guinea. Because PfSPZ Vaccine–induced protection is thought to be mediated by T cells, the T-cell data suggest PfSPZ Vaccine may be more protective in children than in adults, whereas infants may not be immunologically mature enough to respond to the PfSPZ Vaccine immunization regimen assessed.

Fifteen years of investment in malaria control on Bioko Island, Equatorial Guinea (EG), dramatically reduced malaria-associated morbidity and mortality, but the impact has plateaued. To progress toward elimination, EG is investing in the development of a malaria vaccine. We assessed the unique public–private partnership that has had such a significant impact on malaria on Bioko Island and now added a major effort on malaria vaccine development. As part of a $79M commitment, the EG government (75%) and three American energy companies (25%) have invested since 2012 greater than $55M in the Equatoguinean Malaria Vaccine Initiative (EGMVI) to support clinical development of Sanaria® PfSPZ vaccines (Sanaria Inc., Rockville, MD). In turn, the vaccine development program is building human capital and physical capacity. The EGMVI established regulatory and ethical oversight to ensure compliance with the International Conference on Harmonization and Good Clinical Practices for the first importation of investigational product, ethical approval, and conduct of a clinical trial in Equatoguinean history. The EGMVI has completed three vaccine trials in EG, two vaccine trials in Tanzania, and a malaria incidence study, and initiated preparations for a 2,100-volunteer clinical trial. Personnel are training for advanced degrees abroad and have been trained in Good Clinical Practices and protocol-specific methods. A new facility has established the foundation for a national research institute. Biomedical research and development within this visionary, ambitious public–private partnership is fostering major improvements in EG. The EGMVI plans to use a PfSPZ Vaccine alongside standard malaria control interventions to eliminate Pf malaria from Bioko, becoming a potential model for elimination campaigns elsewhere.

In 2015 and in 2016, there were an estimated 429,000–730,500 deaths caused by malaria.1–3 Plasmodium falciparum (Pf) is the cause of > 98% of malaria deaths and > 80% of malaria cases in sub-Saharan Africa. Our goal is to field a vaccine that will prevent infection with Pf and thereby prevent all manifestations of Pf malaria and parasite transmission from humans to mosquitoes.4 Plasmodium falciparum sporozoites (SPZ) are the only immunogens that have ever prevented Pf infection in > 90% of subjects.5–7 Sanaria® PfSPZ Vaccine (Sanaria Inc., Rockville, MD) is composed of radiation-attenuated, aseptic, purified, cryopreserved PfSPZ.8,9 When administered by rapid intravenous injection, PfSPZ Vaccine protected 100% (6/6) of malaria-naive subjects in the United States against mosquito bite–controlled human malaria infection (CHMI) with Pf parasites similar to those in the vaccine (homologous) 3 weeks after the last immunization,10 and 65% at 24 weeks.11 Protection was durable against homologous mosquito bite CHMI for at least 59 weeks12 and heterologous (parasites different than in vaccine) mosquito bite CHMI for at least 33 weeks.13 PfSPZ Vaccine also prevented naturally transmitted heterogeneous Pf in adults in Mali for at least 24 weeks (vaccine efficacy [VE] 52% by time to event and 29% by proportional analysis).14 We used the same dosage regimen as in the United States and Mali to evaluate the tolerability, safety, immunogenicity, and VE of PfSPZ Vaccine in young adult male Tanzanians. Previously, we had conducted the first modern CHMI in Africa and showed that injection of aseptic, purified, cryopreserved PfSPZ, Sanaria® PfSPZ Challenge, consistently infected Tanzanian volunteers and subsequently repeated in multiple other countries.15–21 In this study, we took advantage of this capability to assess VE of PfSPZ Vaccine by CHMI with PfSPZ Challenge (NF54). The same PfSPZ Vaccine dosage regimen was less immunogenic and protective in Tanzanians than in Americans,11 and VE against homologous CHMI in Tanzania was lower (or similar) to VE against intense field exposure to heterogeneous Pf parasites in Mali.14

Equatorial Guinea (EG) has implemented a successful malaria control program on Bioko Island. A highly effective vaccine would be an ideal complement to this effort and could lead to halting transmission and eliminating malaria. Sanaria® PfSPZ Vaccine (Plasmodium falciparum sporozoite Vaccine) is being developed for this purpose. To begin the process of establishing the efficacy of and implementing a PfSPZ Vaccine mass vaccination program in EG, we decided to conduct a series of clinical trials of PfSPZ Vaccine on Bioko Island. Because no clinical trial had ever been conducted in EG, we first successfully established the ethical, regulatory, quality, and clinical foundation for conducting trials. We now report the safety, tolerability, and immunogenicity results of the first clinical trial in the history of the country. Thirty adult males were randomized in the ratio 2:1 to receive three doses of 2.7 × 105 PfSPZ of PfSPZ Vaccine (N = 20) or normal saline placebo (N = 10) by direct venous inoculation at 8-week intervals. The vaccine was safe and well tolerated. Seventy percent, 65%, and 45% of vaccinees developed antibodies to Plasmodium falciparum (Pf) circumsporozoite protein (PfCSP) by enzyme-linked immunosorbent assay, PfSPZ by automated immunofluorescence assay, and PfSPZ by inhibition of sporozoite invasion assay, respectively. Antibody responses were significantly lower than responses in U.S. adults who received the same dosage regimen, but not significantly different than responses in young adult Malians. Based on these results, a clinical trial enrolling 135 subjects aged 6 months to 65 years has been initiated in EG; it includes PfSPZ Vaccine and first assessment in Africa of PfSPZ-CVac. ClinicalTrials.gov identifier: NCT02418962.

Rapid diagnostic tests (RDTs) represent an alternative to microscopy for malaria diagnosis and have shown high sensitivity and specificity in a variety of study settings. Current World Health Organization (WHO) guidelines for quality control of RDTs provide detailed instructions on pre-field testing, but offer little guidance for quality assurance once RDTs are deployed in health facilities. From September 2006 to April 2007, we introduced a histidine-rich protein II (HRP2)-based RDT (Paracheck) for suspected malaria cases five years of age and older in nine health facilities in Rufiji District, Tanzania, to assess sensitivity and specificity of RDTs in routine use at rural health facilities. Thick blood smears were collected for all patients tested with RDTs and stained and read by laboratory personnel in each facility. Thick smears were subsequently reviewed by a reference microscopist to determine RDT sensitivity and specificity. In all nine health facilities, there were significant problems with the quality of staining and microscopy. Sensitivity and specificity of RDTs were difficult to assess given the poor quality of routine blood smear staining. Mean operational sensitivity of RDTs based on reference microscopy was 64.8%, but varied greatly by health facility, range 18.8-85.9%. Sensitivity of RDTs increased with increasing parasite density. Specificity remained high at 87.8% despite relatively poor slide quality. Institution of quality control of RDTs based on poor quality blood smear staining may impede reliable measurement of sensitivity and specificity and undermine confidence in the new diagnostic. There is an urgent need for the development of alternative quality control procedures for rapid diagnostic tests that can be performed at the facility level.

Malaria's relationship with socioeconomic status at the macroeconomic level has been established. This is the first study to explore this relationship at the microeconomic (household) level and estimate the direction of association. Malaria prevalence was measured by parasitemia, and household socioeconomic status was measured using an asset based index. Results from an instrumental variable probit model suggest that socioeconomic status is negatively associated with malaria parasitemia. Other variables that are significantly associated with parasitemia include age of the individual, use of a mosquito net on the night before interview, the number of people living in the household, whether the household was residing at their farm home at the time of interview, household wall construction, and the region of residence. Matching estimators indicate that malaria parasitemia is associated with reduced household socioeconomic status.

Prior to the 2001 malarial treatment policy change in Tanzania, we conducted trials to assess the efficacy of sulfadoxine-pyrimethamine (SP) and the usefulness of molecular markers in monitoring resistance. A total of 383 uncomplicated Plasmodium falciparum malaria patients (between 6 and 59 months old) were treated with SP and their responses were assessed. Mutations in the P. falciparum dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes in admission day blood samples were analyzed. Results indicated that 85.6% of the patients showed an adequate clinical response, 9.7% an early treatment failure, and 4.7% a late treatment failure. The quintuple mutant genotype (pfdhfr 51 Ile, 59 Arg, and 108 Asn and pfdhps 437 Gly and 540 Glu) showed an association with treatment outcome (odds ratio = 2.1; 95% confidence interval = 0.94-4.48, P = 0.045). The prevalence of the triple pfdhfr mutant genotype (51 Ile, 59 Arg, and 108 Asn) at a site of high SP resistance (23.6%) was four times higher compared with that observed at sites of moderate SP resistance (6.8-14.4%) (P = 0.000001). The genotype failure index calculated by using this marker was invariable (1.96-2.1) at sites with moderate SP resistance, but varied (3.4) at a site of high SP resistance. In conclusion, our clinical and molecular findings suggest that SP may have a short useful therapeutic life in Tanzania; thus, its adoption as an interim first-line antimalarial drug. The findings also point to the potential of the triple pfdhfr mutant genotype as an early warning tool for increasing SP resistance. These data form the baseline SP efficacy and molecular markers profile in Tanzania prior to the policy change.

Malaria's relationship with socioeconomic status at the macroeconomic level has been established. This is the first study to explore this relationship at the microeconomic (household) level and estimate the direction of association. Malaria prevalence was measured by parasitemia, and household socioeconomic status was measured using an asset based index. Results from an instrumental variable probit model suggest that socioeconomic status is negatively associated with malaria parasitemia. Other variables that are significantly associated with parasitemia include age of the individual, use of a mosquito net on the night before interview, the number of people living in the household, whether the household was residing at their farm home at the time of interview, household wall construction, and the region of residence. Matching estimators indicate that malaria parasitemia is associated with reduced household socioeconomic status.

Prior to the 2001 malarial treatment policy change in Tanzania, we conducted trials to assess the efficacy of sulfadoxine-pyrimethamine (SP) and the usefulness of molecular markers in monitoring resistance. A total of 383 uncomplicated Plasmodium falciparum malaria patients (between 6 and 59 months old) were treated with SP and their responses were assessed. Mutations in the P. falciparum dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes in admission day blood samples were analyzed. Results indicated that 85.6% of the patients showed an adequate clinical response, 9.7% an early treatment failure, and 4.7% a late treatment failure. The quintuple mutant genotype (pfdhfr 51 Ile, 59 Arg, and 108 Asn and pfdhps 437 Gly and 540 Glu) showed an association with treatment outcome (odds ratio = 2.1; 95% confidence interval = 0.94-4.48, P = 0.045). The prevalence of the triple pfdhfr mutant genotype (51 Ile, 59 Arg, and 108 Asn) at a site of high SP resistance (23.6%) was four times higher compared with that observed at sites of moderate SP resistance (6.8-14.4%) (P = 0.000001). The genotype failure index calculated by using this marker was invariable (1.96-2.1) at sites with moderate SP resistance, but varied (3.4) at a site of high SP resistance. In conclusion, our clinical and molecular findings suggest that SP may have a short useful therapeutic life in Tanzania; thus, its adoption as an interim first-line antimalarial drug. The findings also point to the potential of the triple pfdhfr mutant genotype as an early warning tool for increasing SP resistance. These data form the baseline SP efficacy and molecular markers profile in Tanzania prior to the policy change.