Jonathan R. Davis

Clinical Manifestations of Plasmodium falciparum Malaria Experimentally Induced by Mosquito Challenge

To determine the characteristics of clinical illness accompanying Plasmodium falciparum infection induced by controlled exposure to infected mosquitoes, records of 118 volunteers participating in studies conducted between 1985 and 1992 were reviewed. One hundred fourteen volunteers (97%) reported at least one symptom attributable to malaria, with fatigue, myalgias or arthralgias, headache, and chills most commonly reported. The median duration of symptoms was 3 days. Fever was recorded in 61% of volunteers; 4 volunteers had temperatures >40 degrees C. Neutropenia and thrombocytopenia were present in 9% and 12% of volunteers, respectively. Despite counts as low as 658/microL (neutrophils) or 73,000/microL (platelets), no secondary infectious or hemorrhagic complications occurred. In all cases, volunteers recovered completely and laboratory values returned to baseline after specific antimalarial therapy. Recrudescence did not occur in any volunteer. In this model, mosquito inoculation of P. falciparum is a reliable, safe, and well-tolerated method of experimental challenge.

COMPARISON OF SAMPLING ANOPHELINE MOSQUITOES BY LIGHT TRAP AND HUMAN BAIT COLLECTIONS INDOORS AT BAGAMOYO, TANZANIA

Abstract. The mosquito sampling efficiency of CDC miniature light‐traps, relative to night‐biting collections, was evaluated indoors at two sites in coastal Tanzania. We found that the total number of anophelines captured overnight by light‐traps (hung beside a bednet in use) was 1.23 times the number of anophelines captured by human‐bait collections. This relationship was not affected significantly by changes in the mosquito density, order of trapping method, date of sampling, or number of household occupants. Malaria sporozoite rates were twice as high among mosquitoes captured by light‐trap as compared to those captured by night‐biting collection. This was attributed to the tendency of light‐traps to capture a larger proportion of gravid mosquitoes, which also had high sporozoite rates. The differences in sporozoites rates according to abdominal stage indicates that unfed mosquitoes captured by light‐traps may define more precisely the human‐biting activity and sporozoite rates as seen by night‐biting collections. Our study shows that light‐traps, when used in combination with night‐biting collections, can be an effective and sensitive means for measuring human‐biting activity and the sporozoite rate.

Serum cytokine profiles in experimental human malaria. Relationship to protection and disease course after challenge.

Serum cytokine profiles were evaluated in immunized and nonimmunized human volunteers after challenge with infectious Plasmodium falciparum sporozoites. Three volunteers had been immunized with x-irradiated sporozoites and were fully protected from infection. Four nonimmune volunteers all developed symptomatic infection at which time they were treated. Sera from all volunteers were collected at approximately 20 time points during the 28-d challenge period; levels of IL-1 alpha, IL-1 beta, IL-2, IFN-gamma, tumor necrosis factor-alpha, IL-4, IL-6, granulocyte macrophage-colony-stimulating factor, and soluble CD4, CD8, and IL-2 receptor (sCD4, sCD8, and sIL-2R, respectively) were determined by ELISA. C-reactive protein (CRP) was assayed by radial immunodiffusion. Parasitemic subjects developed increases in CRP and IFN-gamma, with less marked increases in sIL-2R and sCD8; the other cytokines tested did not change. CRP increases were abrupt and occurred at the onset of fever (day 14 after challenge). IFN-gamma increases were also abrupt, preceding those of fever and CRP by one day. Increases in sIL-2R and sCD8 were more gradual. Increases in fever, CRP, IFN-gamma, and sCD8 were concordant in each volunteer. Early IL-6 increases were noted in the protected vaccinees. Thus, after challenge with virulent P. falciparum, unique systemic cytokine profiles were detectable both in immunized, nonparasitemic volunteers and in unvaccinated, parasitemic subjects. The contrasting cytokine profiles in the two groups may relate to mechanisms of protection and immunopathology in experimental human malaria.

Malaria infection potential of anopheline mosquitoes sampled by light trapping indoors in coastal Tanzanian villages

Abstract. Anopheline mosquito populations were studied during 1992 in seven villages south of Bagamoyo, coastal Tanzania, prior to malaria control intervention using insecticide treated bednets. To collect mosquitoes, CDC light traps were used in ten houses per village fortnightly for 12 months. Anopheles females were identified and checked by ELISA for the presence of malaria sporozoite antigen and source of bloodmeal. An. funestus peaked in June‐July after the long rains. Three members of the An. gambiae complex had different seasonality: An. arabiensis, An. gambiae and small numbers of An. merus were collected.

Plasmodium falciparum: in vitro characterization and human infectivity of a cloned line.

The culture-adapted NF54 isolate of Plasmodium falciparum was subjected in vitro to three sequential limiting dilution titrations and the resulting clone was given the designation CVD1. DNA sequence analysis of the gene encoding the circumsporozoite (CS) protein revealed differences between CVD1 and the published NF54 CS gene. CVD1 had 1191 bp, 397 amino acids, and 42 repeat units while NF54 had 1218 bp, 405 amino acids, and 44 repeat units. The CVD1 clone was more sensitive to chloroquine than was the parental line, in vitro. Anopheles stephensi mosquitoes were infected equally by the cloned and uncloned parasites. Volunteers were readily infected by NF54 and CVD1 following infectious mosquito bites. The availability of a well-characterized, chloroquine-sensitive clone which safety infects humans should facilitate performance of experimental challenge studies to assess vaccine efficacy.

Estimate of anti-plasmodium falciparum sporozoite activity in humans vaccinated with synthetic circumsporozoite protein (NANP)3

A mathematical model was defined to estimate the degree of in vivo activity against Plasmodium falciparum sporozoites expressed by volunteers vaccinated with a synthetic peptide comprising the immunodominant epitope of the circumsporozoite protein. Relative to the course of infection in non-immunized controls, infections in vaccinated volunteers corresponded to the neutralization or delay of development of greater than 99% of challenge sporozoites.

Urinary neopterin in volunteers experimentally infected with Plasmodium falciparum

To investigate the kinetics of monocyte/macrophage activation in falciparum malaria we determined urinary neopterin values serially in experimentally infected volunteers. Three subjects who had been immunized with irradiated sporozoites via mosquito bites served as controls. These individuals remained aparasitaemic, afebrile and without a rise in neopterin after challenge by infective mosquitoes. Four non-immune subjects developed Plasmodium falciparum parasitaemia, fever (3 of 4) and sharp rises in neopterin. Parasite densities reached 10-100 parasitized erythrocytes per microliter before elevations in temperature or neopterin levels were detected. Onset of fever preceded the rise in neopterin excretion by one day. Prompt chemotherapy was associated with the clearance of parasites from the blood and the return of temperature and neopterin levels to normal.

Laboratory methods for the conduct of experimental malaria challenge of volunteers.

Experimental malaria challenge trials of volunteers are conducted to determine whether candidate sporozoite vaccines generate protective immunity against the bites of infected mosquitoes. The absence of patent erythrocytic-stage infection and clinical sequelae suggests sterile immunity among vaccinated volunteers. Sterile immunity is validated through laboratory studies that (1) demonstrate the capacity of erythrocytes and serum obtained from each volunteer before challenge to support parasite growth in vitro; (2) confirm parasitaemia by blood culture, and (3) evaluate each volunteers blood for surreptitious use of antimalarial agents. The bites of experimentally infected laboratory-reared anophelines must elicit a 100% attack rate among non-immunized volunteers. This report presents guidelines for the laboratory methods used to validate the clinical findings during experimental Plasmodium falciparum anti-sporozoite vaccine efficacy trials.

Continuation of chloroquine-susceptible Plasmodium falciparum parasitemia in volunteers receiving chloroquine therapy.

Volunteers infected with a chloroquine-susceptible line of Plasmodium falciparum were administered standard oral chloroquine therapy at the first detection of parasites in the blood. Parasitemias progressed in the face of therapy for up to 5 days and to levels up to 100-fold greater than those at the initiation of treatment. Thereafter, infections cleared without a requirement for additional chemotherapy. This course of infection and response to treatment has not been previously reported and may have been detected because volunteers were exposed to an unusually large number of sporozoites. The observations are consistent with the hypothesis that prolonged parasitemia resulted from the continued release of merozoites from liver.