Schistosoma mansoni infection impairs antimalaria treatment and immune responses of rhesus macaques infected with mosquito-borne Plasmodium coatneyi

Abstract

Malaria and schistosomiasis are the world's two most important parasitic infections in terms of distribution, morbidity, and mortality. In areas where Plasmodium and Schistosoma species are both endemic, coinfections are commonplace. Mouse models demonstrate that schistosomiasis worsens a malaria infection; however, just as mice and humans differ greatly, the murine-infecting Plasmodium species differ as much from those that infect humans. Research into human coinfections (Schistosoma haematobium-Plasmodium falciparum versus Schistosoma mansoni-P. falciparum) has produced conflicting results. The rhesus macaque model provides a helpful tool for understanding the role of S. mansoni on malaria parasitemia and antimalarial immune responses using Plasmodium coatneyi, a malaria species that closely resembles P. falciparum infection in humans. Eight rhesus macaques were exposed to S. mansoni cercariae. Eight weeks later, these animals plus 8 additional macaques were exposed to malaria either through bites of infected mosquitos or intravenous inoculation. When malaria infection was initiated from mosquito bites, coinfected animals displayed increased malaria parasitemia, decreased hematocrit levels, and suppressed malaria-specific antibody responses compared to those of malaria infection alone. However, macaques infected by intravenous inoculation with erythrocytic-stage parasites did not display these same differences in parasitemia, hematocrit, or antibody responses between the two groups. Use of the macaque model provides information that begins to unravel differences in pathological and immunological outcomes observed between humans with P. falciparum that are coinfected with S. mansoni or S. haematobium. Our results suggest that migration of malaria parasites through livers harboring schistosome eggs may alter host immune responses and infection outcomes.

Fig 1

Clinical responses in coinfected and malaria-only-infected macaques infected by sporozoite challenge. (A) Parasitemia was calculated as the number of P. coatneyi-infected red blood cells per the total number of uninfected red blood cells. Macaques were treated on days 17, 18, 19, 20, 21, and 22 with subcurative doses of quinine. Parasitemia is shown here from day 10 to day 25 after malaria exposure and is graphed on a linear scale. The difference between parasitemia of the coinfected and malaria-only-infected macaques was found to be significant (P < 0.0001; ■, malaria only or schistosomiasis negative; ▲, coinfected or schistosomiasis positive). (B) Parasites per microliter were defined as the total number of parasites present per microliter of blood, and these values from day 10 to day 25 after malaria exposure were graphed on a log scale. The difference between the curves was significant (P < 0.0001). (C) Hematocrit values were determined weekly after malaria exposure and graphed here on a linear scale. Hematocrit values between the coinfected and malaria-only-infected macaques were significantly different (P = 0.025). (D) Hemoglobin levels were also determined for the same time period as the hematocrit values, and these numbers are shown in grams per deciliter and graphed on a linear scale. The difference in hemoglobin levels was also determined to be significantly different (P = 0.027).

Fig 2

Antibody responses (IgG and IgM) to malaria antigen during a sporozoite infection. Plasma was collected at 0, 2, 3, 5, 7, 8, and 14 weeks after malaria exposure. Malaria was cured at week 8 after malaria exposure, and so week 14 corresponds to 6 weeks after malaria cure. The antibody values were determined from a standard curve using P. coatneyi immune plasma. The arrows indicate when drug treatment was started (day 17). (A) Malaria-specific IgG antibody responses were determined using P. coatneyi crude antigen. IgG responses for coinfected macaques are displayed by the black bars and those for malaria-only-infected macaques are displayed by the gray bars. The IgG responses for coinfected macaques are significantly lower than those of the malaria-only-infected macaques (P = 0.01). (B) Malaria-specific IgM responses were also determined using a P. coatneyi crude antigen, and IgM responses for coinfected macaques are displayed in black and those for malaria-only macaques are in gray. The IgM responses are significantly different, with malaria-only-infected macaques having significantly lower IgM responses (P = 0.01).

Fig 3

Parasitemia in malaria blood-stage inoculated macaques. (A) Parasitemia was defined as number of P. coatneyi-infected red blood cells per the total number of uninfected red blood cells. Macaques were treated on days 11, 12, 13, and 15 after exposure to malaria. Parasitemia from day 1 to day 15 is graphed on a linear scale for both coinfected and malaria-only-infected macaques (■, malaria-only or schistosomiasis negative; ▲, coinfected or schistosomiasis positive). Parasitemia is not significantly different between the two groups of macaques (P = 0.38). (B) Parasites per microliter were defined as the total number of parasites in a microliter of blood; this was graphed from day 4 to day 15 on a log scale. The values were not significantly different between the groups (P = 0.39). (C) Hematocrit was determined weekly after exposure to malaria until malaria cure; there is no significant difference between coinfected and malaria-only-infected macaques (P = 0.16). (D) Hemoglobin values (in grams per deciliter) were determined for the same time period as the hematocrit, and there is no significant difference between the groups (P = 0.17).

Fig 4

P. coatneyi antigen-specific immunoglobulin responses (IgG and IgM) in blood-stage-exposed macaques. Plasma was collected at weeks 0, 3, 6, 8, 11, and 15 after exposure to malaria. Malaria infection was cured at week 7 postexposure. Malaria-specific antibody responses were determined using P. coatneyi crude antigen. The unknown values were determined by assigning units based from a standard curve of P. coatneyi immune sera. The arrows indicate the time when drug treatment was started (day 11). Over the course of the infection, IgG responses (A) and IgM responses (B) to malaria antigen were not significantly different (P = 0.34 and P = 0.23) between the coinfected (black bars) and malaria-only-infected macaques (gray bars).

Fig 5

Schistosome parasite load for sporozoite and blood-stage malaria infection. Egg counts were determined microscopically from processed stool samples collected weekly until egg counts reach zero for a minimum of two consecutive weeks. The graph shows the average number of eggs per gram of stool determined for the four macaques that were exposed to malaria through mosquito bites (◆) and the average number of eggs per gram of stool determined for the four macaques exposed to malaria through blood-stage parasites (●). The difference between schistosome parasite load of the two groups over the course of the schistosomiasis infection is not statistically significant (P = 0.20).