gammadelta T cells are a component of early immunity against preerythrocytic malaria parasites

Abstract

We tested the hypothesis that gammadelta T cells are a component of an early immune response directed against preerythrocytic malaria parasites that are required for the induction of an effector alphabeta T-cell immune response generated by irradiated-sporozoite (irr-spz) immunization. gammadelta T-cell-deficient (TCRdelta(-/-)) mice on a C57BL/6 background were challenged with Plasmodium yoelii (17XNL strain) sporozoites, and then liver parasite burden was measured at 42 h postchallenge. Liver parasite burden was measured by quantification of parasite-specific 18S rRNA in total liver RNA by quantitative-competitive reverse transcription-PCR and by an automated 5' exonuclease PCR. Sporozoite-challenged TCRdelta(-/-) mice showed a significant (P < 0.01) increase in liver parasite burden compared to similarly challenged immunocompetent mice. In support of this result, TCRdelta(-/-) mice were also found to be more susceptible than immunocompetent mice to a sporozoite challenge when blood-stage parasitemia was used as a readout. A greater percentage of TCRdelta(-/-) mice than of immunocompetent mice progressed to a blood-stage infection when challenged with five or fewer sporozoites (odds ratio = 2.35, P = 0.06). TCRdelta(-/-) mice receiving a single irr-spz immunization showed percent inhibition of liver parasites comparable to that of immunized immunocompetent mice following a sporozoite challenge. These data support the hypothesis that gammadelta T cells are a component of early immunity directed against malaria preerythrocytic parasites and suggest that gammadelta T cells are not required for the induction of an effector alphabeta T-cell immune response generated by irr-spz immunization.

FIG. 1

Quantification of P. yoelii 18S rRNA in total liver RNA by two unique assays. Groups of four to five mice were challenged with increasing numbers of sporozoites (5.0 × 10⁴ to 4.0 × 10⁵), and parasite rDNA was measured in total liver cDNA by quantitative-competitive RT-PCR (A) and by 5′ exonuclease PCR (Taq Man) (B). Error bars indicate 1 standard deviation of the mean.

FIG. 2

Variability in sporozoite viability and/or infectivity between sporozoites isolated from different batches of infected mosquitoes. Groups of mice were challenged with 5.0 × 10⁴ sporozoites, and liver parasite burden was measured by Taq Man quantification of parasite-specific 18S rDNA at 42 h postinfection (A). Each experiment represents a group of mice challenged with sporozoites isolated from a different batch of infected mosquitoes. Equal cDNA synthesis between groups of mice was ensured by quantification of the housekeeping gene GAPDH (B). Experiment 1 represents data from immunocompetent C57BL/6 mice challenged with 5.0 × 10⁴ sporozoites also displayed in Fig. 1. Experiment 3 represents data from immunocompetent C57BL/6 mice challenged with 5.0 × 10⁴ sporozoites also displayed in experiment 1 of Table 1. Experiment 4 represents data from immunocompetent C57BL/6 mice challenged with 5.0 × 10⁴ sporozoites also displayed in Fig. 4 and experiment 2 of Table 1.

FIG. 3

Liver parasite burden in γδ T-cell-deficient mice receiving a single irr-spz immunization. TCRδ^−/− and C57BL/6 mice received a single irr-spz immunization with 7.5 × 10⁴ sporozoites and were challenged with 10⁵ sporozoites 7 days later. As a mock immunization control, some groups of mice were given an equivalent volume of medium alone. Liver parasite burden was measured by quantitative-competitive RT-PCR amplification of parasite-specific 18S rDNA in total liver cDNA at 42 h postinfection. Error bars indicate 1 standard deviation of the mean.

FIG. 4

Increased liver parasite burden in αβ T-cell-deficient mice. Groups of five C57BL/6, TCRδ^−/−, TCRβ^−/−, class I-deficient (B2M^−/−), and class II-deficient (A_β^b−/−) mice were challenged with 5.0 × 10⁴ sporozoites, and liver parasite burden was measured by quantitative-competitive RT-PCR of parasite-specific 18S rDNA. Error bars indicate 1 standard deviation of the mean. ∗, significant increase (P < 0.05) in liver parasite burden compared to the liver parasite burden of C57BL/6 mice.