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Cholera emergence is strongly linked to local environmental and ecological context. The 1991–2004 pandemic emerged in Perú and spread north into Ecuador’s El Oro province, making this a key site for potential re-emergence. Machala, El Oro, is a port city of 250,000 inhabitants, near the Peruvian border. Many livelihoods depend on the estuarine system, from fishing for subsistence and trade, to domestic water use. In 2014, we conducted biweekly sampling for 10 months in five estuarine locations, across a gradient of human use, and ranging from inland to ocean. We measured water-specific environmental variables implicated in cholera growth and persistence: pH, temperature, salinity, and algal concentration, and evaluated samples in five months for pathogenic and non-pathogenic Vibrio cholerae, by polymerase chain reaction (PCR). We found environmental persistence of pandemic strains O1 and O139, but no evidence for toxigenic strains. Vibrio cholerae presence was coupled to algal and salinity concentration, and sites exhibited considerable seasonal and spatial heterogeneity. This study indicates that environmental conditions in Machala are optimal for cholera re-emergence, with risk peaking during September, and higher risk near urban periphery low-income communities. This highlights a need for surveillance of this coupled cholera–estuarine system to anticipate potential future cholera outbreaks.

Gene-based vaccination using prime/boost regimens protects animals and humans against malaria, inducing cell-mediated responses that in animal models target liver stage malaria parasites. We tested a DNA prime/adenovirus boost malaria vaccine in a Phase 1 clinical trial with controlled human malaria infection. The vaccine regimen was three monthly doses of two DNA plasmids (DNA) followed four months later by a single boost with two non-replicating human serotype 5 adenovirus vectors (Ad). The constructs encoded genes expressing P. falciparum circumsporozoite protein (CSP) and apical membrane antigen-1 (AMA1). The regimen was safe and well-tolerated, with mostly mild adverse events that occurred at the site of injection. Only one AE (diarrhea), possibly related to immunization, was severe (Grade 3), preventing daily activities. Four weeks after the Ad boost, 15 study subjects were challenged with P. falciparum sporozoites by mosquito bite, and four (27%) were sterilely protected. Antibody responses by ELISA rose after Ad boost but were low (CSP geometric mean titer 210, range 44 817; AMA1 geometric mean micrograms/milliliter 11.9, range 1.5 102) and were not associated with protection. Ex vivo IFN-c ELISpot responses after Ad boost were modest (CSP geometric mean spot forming cells/million peripheral blood mononuclear cells 86, range 13 408; AMA1 348, range 88 1270) and were highest in three protected subjects. ELISpot responses to AMA1 were significantly associated with protection (p = 0.019). Flow cytometry identified predominant IFN-c mono-secreting CD8+ T cell responses in three protected subjects. No subjects with high pre-existing anti-Ad5 neutralizing antibodies were protected but the association was not statistically significant. The DNA/Ad regimen provided the highest sterile immunity achieved against malaria following immunization with a gene-based subunit vaccine (27%)., Published in PLoS ONE, v8 n2 article ID e55571, 14 Feb 2013. Prepared in collaboration with the US Military Malaria Vaccine Program, Naval Medical Research Center, Silver Spring, MD. Prepared in cooperation with the Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands, GenVec, Inc., Gaithersburg, MD, and USAID, Washington, D. C. The original document contains color images.

The parasite Plasmodium vivax is the most frequent cause of malaria outside of sub-Saharan Africa, but efforts to develop viable vaccines against P. vivax so far have been inadequate. We recently developed pathogen-mimicking polymeric vaccine nanoparticles composed of the FDA-approved biodegradable polymer poly(lactide-co-glycolide) acid (PLGA) “enveloped” by a lipid membrane. In this study, we sought to determine whether this vaccine delivery platform could be applied to enhance the immune response against P. vivax sporozoites. A candidate malaria antigen, VMP001, was conjugated to the lipid membrane of the particles, and an immunostimulatory molecule, monophosphoryl lipid A (MPLA), was incorporated into the lipid membranes, creating pathogen-mimicking nanoparticle vaccines (VMP001-NPs). Vaccination with VMP001-NPs promoted germinal center formation and elicited durable antigen-specific antibodies with significantly higher titers and more balanced Th1/Th2 responses in vivo, compared with vaccines composed of soluble protein mixed with MPLA. Antibodies raised by NP vaccinations also exhibited enhanced avidity and affinity toward the domains within the circumsporozoite protein implicated in protection and were able to agglutinate live P. vivax sporozoites. These results demonstrate that these VMP001-NPs are promising vaccines candidates that may elicit protective immunity against P. vivax sporozoites., United States. Dept. of Defense (contract W911NF-07-D-0004), Ragon Institute of MGH, MIT and Harvard

The parasite Plasmodium vivax is the most frequent cause of malaria outside of sub-Saharan Africa, but efforts to develop viable vaccines against P. vivax so far have been inadequate. We recently developed pathogen-mimicking polymeric vaccine nanoparticles composed of the FDA-approved biodegradable polymer poly(lactide-co-glycolide) acid (PLGA) “enveloped” by a lipid membrane. In this study, we sought to determine whether this vaccine delivery platform could be applied to enhance the immune response against P. vivax sporozoites. A candidate malaria antigen, VMP001, was conjugated to the lipid membrane of the particles, and an immunostimulatory molecule, monophosphoryl lipid A (MPLA), was incorporated into the lipid membranes, creating pathogen-mimicking nanoparticle vaccines (VMP001-NPs). Vaccination with VMP001-NPs promoted germinal center formation and elicited durable antigen-specific antibodies with significantly higher titers and more balanced Th1/Th2 responses in vivo, compared with vaccines composed of soluble protein mixed with MPLA. Antibodies raised by NP vaccinations also exhibited enhanced avidity and affinity toward the domains within the circumsporozoite protein implicated in protection and were able to agglutinate live P. vivax sporozoites. These results demonstrate that these VMP001-NPs are promising vaccines candidates that may elicit protective immunity against P. vivax sporozoites., United States. Dept. of Defense (contract W911NF-07-D-0004), Ragon Institute of MGH, MIT and Harvard

The Plasmodium protein Cell-traversal protein for ookinetes and sporozoites (CelTOS) plays an important role in cell traversal of host cells in both, mosquito and vertebrates, and is required for successful malaria infections. CelTOS is highly conserved among the Plasmodium species, suggesting an important functional role across all species. Therefore targeting the immune response to this highly conserved protein and thus potentially interfering with its biological function may result in protection against infection even by heterologous species of Plasmodium. Methodology/Principal Findings: To test this hypothesis, we developed a recombinant codon-harmonized P. falciparum CelTOS protein that can be produced to high yields in the E. coli expression system. Inbred Balb/c and outbred CD-1 mice were immunized with various doses of the recombinant protein adjuvanted with Montanide ISA 720 and characterized using in vitro and in vivo analyses. Conclusions/Significance: Immunization with PfCelTOS resulted in potent humoral and cellular immune responses and most importantly induced sterile protection against a heterologous challenge with P. berghei sporozoites in a proportion of both inbred and outbred mice. The biological activity of CelTOS-specific antibodies against the malaria parasite is likely linked to the impairment of sporozoite motility and hepatocyte infectivity. The results underscore the potential of this antigen as a pre-erythrocytic vaccine candidate and demonstrate for the first time a malaria vaccine that is cross-protective between species., Pub. in PLoS One, v5 n8, p1-9, Aug 2010.

Cutaneous Leishmania major has affected many travelers including military personnel in Iraq and Afghanistan. Optimal treatment for this localized infection has not been defined, but interestingly the parasite is thermosensitive. Participants with parasitologically confirmed L. major infection were randomized to receive intravenous sodium stibogluconate (SSG) 20mg/kg/day for ten doses or localized ThermoMed (TM) device heat treatment (applied at 50uC for 30 seconds) in one session. Those with facial lesions, infection with other species of Leishmania, or more than 20 lesions were excluded. Primary outcome was complete re-epithelialization or visual healing at two months without relapse over 12 months. Fifty-four/56 enrolled participants received intervention, 27 SSG and 27 TM. In an intent to treat analysis the per subject efficacy at two months with 12 months follow-up was 54% SSG and 48% TM (p = 0.78), and the per lesion efficacy was 59% SSG and 73% TM (p = 0.053). Reversible abdominal pain/pancreatitis, arthralgias, myalgias, headache, fatigue, mild cytopenias, and elevated transaminases were more commonly present in the SSG treated participants, whereas blistering, oozing, and erythema were more common in the TM arm. Skin lesions due to L. major treated with heat delivered by the ThermoMed device healed at a similar rate and with less associated systemic toxicity than lesions treated with intravenous SSG., Published in PLOS Neglected Tropical Diseases v4 i3 March 2010.

Background:This study advances the clinical development of the RTS,S/AS01B candidate malaria vaccine to malaria endemic populations. As a primary objective it compares the safety and reactogenicity of RTS,S/AS01B to the more extensively evaluated RTS,S/AS02A vaccine.Methodology:A Phase IIb, single centre, double-blind, controlled trial of 6 months duration with a subsequent 6 month single-blind follow-up conducted in Kisumu West District, Kenya between August 2005 and August 2006. 255 healthy adults aged 18 to 35 years were randomized (1:1:1) to receive 3 doses of RTS,S/AS02A, RTS,S/AS01B or rabies vaccine (Rabipur®; Chiron Behring GmbH) at months 0, 1, 2. The primary objective was the occurrence of severe (grade 3) solicited or unsolicited general (i.e. systemic) adverse events (AEs) during 7 days follow up after each vaccination.Principal Findings:Both candidate vaccines had a good safety profile and were well tolerated. One grade 3 systemic AE occurred within 7 days of vaccination (RTS,S/AS01B group). No unsolicited AEs or SAEs were related to vaccine. A marked increase in anti-CS antibody GMTs was observed post Dose 2 of both RTS,S/AS01B (31.6 EU/mL [95% CI: 23.9 to 41.6]) and RTS,S/AS02A (16.7 EU/mL [95% CI: 12.9 to 21.7]). A further increase was observed post Dose 3 in both the RTS,S/AS01B (41.4 EU/mL [95% CI: 31.7 to 54.2]) and RTS,S/AS02A (21.4 EU/mL [95% CI: 16.0 to 28.7]) groups. Anti-CS antibody GMTs were significantly greater with RTS,S/AS01B compared to RTS,S/AS02A at all time points post Dose 2 and Dose 3. Both candidate vaccines produced strong anti-HBs responses. Vaccine efficacy in the RTS,S/AS01B group was 29.5% (95% CI: −15.4 to 56.9, p = 0.164) and in the RTS,S/AS02A group 31.7% (95% CI: −11.6 to 58.2, p = 0.128).Conclusions:Both candidate malaria vaccines were well tolerated over a 12 month surveillance period. A more favorable immunogenicity profile was observed with RTS,S/AS01B than with RTS,S/AS02A.

Objective:RTS,S, a candidate vaccine for malaria, is a recombinant protein expressed in yeast containing part of the circumsporozoite protein (CSP) sequence of 3D7 strain of Plasmodium falciparum linked to the hepatitis B surface antigen in a hybrid protein. The RTS,S antigen is formulated with GSK Biologicals' proprietary Adjuvant Systems AS02A or AS01B. A recent trial of the RTS,S/AS02A and RTS,S/AS01B vaccines evaluated safety, immunogenicity and impact on the development of parasitemia of the two formulations. Parasite isolates from this study were used to determine the molecular impact of RTS,S/AS02A and RTS,S/AS01B on the multiplicity of infection (MOI) and the csp allelic characteristics of subsequent parasitemias.Design:The distribution of csp sequences and the MOI of the infecting strains were examined at baseline and in break-through infections from vaccinated individuals and from those receiving a non-malarial vaccine.Setting:The study was conducted in Kombewa District, western Kenya.Participants:Semi-immune adults from the three study arms provided isolates at baseline and during break-through infections.Outcome:Parasite isolates used for determining MOI and divergence of csp T cell–epitopes were 191 at baseline and 87 from break-through infections.Results:Grouping recipients of RTS,S/AS01A and RTS,S/AS02B together, vaccine recipients identified as parasite-positive by microscopy contained significantly fewer parasite genotypes than recipients of the rabies vaccine comparator (median in pooled RTS,S groups: 3 versus 4 in controls, P = 0.0313). When analyzed separately, parasitaemic individuals in the RTS,S/AS01B group, but not the RTS,S/AS02A group, were found to have significantly fewer genotypes than the comparator group. Two individual amino acids found in the vaccine construct (Q339 in Th2R and D371 in Th3R) were observed to differ in incidence between vaccine and comparator groups but in different directions; parasites harboring Q339 were less

Objective:The antigen, falciparum malaria protein 1 (FMP1), represents the 42-kDa C-terminal fragment of merozoite surface protein-1 (MSP-1) of the 3D7 clone of P. falciparum. Formulated with AS02 (a proprietary Adjuvant System), it constitutes the FMP1/AS02 candidate malaria vaccine. We evaluated this vaccine's safety, immunogenicity, and efficacy in African children.Methods:A randomised, double-blind, Phase IIb, comparator-controlled trial.The trial was conducted in 13 field stations of one mile radii within Kombewa Division, Nyanza Province, Western Kenya, an area of holoendemic transmission of P. falciparum. We enrolled 400 children aged 12–47 months in general good health.Children were randomised in a 1:1 fashion to receive either FMP1/AS02 (50 µg) or Rabipur® rabies vaccine. Vaccinations were administered on a 0, 1, and 2 month schedule. The primary study endpoint was time to first clinical episode of P. falciparum malaria (temperature ≥37.5°C with asexual parasitaemia of ≥50,000 parasites/µL of blood) occurring between 14 days and six months after a third dose. Case detection was both active and passive. Safety and immunogenicity were evaluated for eight months after first immunisations; vaccine efficacy (VE) was measured over a six-month period following third vaccinations.Results:374 of 400 children received all three doses and completed six months of follow-up. FMP1/AS02 had a good safety profile and was well-tolerated but more reactogenic than the comparator. Geometric mean anti-MSP-142 antibody concentrations increased from1.3 µg/mL to 27.3 µg/mL in the FMP1/AS02 recipients, but were unchanged in controls. 97 children in the FMP1/AS02 group and 98 controls had a primary endpoint episode. Overall VE was 5.1% (95% CI: −26% to +28%; p-value = 0.7).Conclusions:FMP1/AS02 is not a promising candidate for further development as a monovalent malaria vaccine. Future MSP-142 vaccine development should focus on othe

Persistence of vaccinia at vaccination sites may help determine the risk associated with secondary transmission. Culture, PCR, and antigen detection were performed on serial vaccination site swab specimens. On day 21 after vaccination, 37% of volunteers were culture positive, most of whom had received vaccine for the first time. Vaccinia is detectable at least through day 21 after vaccination., Published in Clinical Infectious Diseases, v46 n1 p101-102, 1 Jan 2008.