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BALB/c mice are about 2,000 times less susceptible to sporozoites of Plasmodium berghei than to Plasmodium yoelii. Associated with this is the innate cellular response mounted after injection with P. berghei. Host inflammatory cells do not normally attack P. yoelii during their development as exoerythrocytic forms (EEFs) in the liver. We used P. berghei sporozoites to induce host inflammation that might act against developing P. yoelii EEFs. Mice injected with P. berghei sporozoites followed 1 hr later with P. yoelii had a 58% reduction in P. yoelii EEFs. To establish whether this was due to events that occurred before vs. after invasion of hepatocytes by P. yoelii sporozoites, mice received P. yoelii sporozoites that were allowed to invade for 1 hr before subsequent injection with P. berghei; these mice showed minimal reduction in P. yoelii EEFs. Thus, most of the deleterious effects of P. berghei sporozoites appear to have been directed against P. yoelii sporozoites prior to their invasion of hepatocytes. Plasmodium yoelii that had already invaded were relatively unaffected. Further timing experiments showed that this effect was induced only by viable P. berghei sporozoites, which may thus induce rapid changes in sinusoid physiology leading to host resistance against P. yoelii sporozoites.

After injection of Plasmodium berghei sporozoites into Norway-Brown rats, we were able to localize these sporozoites and the early hepatic trophozoites developing from them in histological sections of the liver stained with a sensitive immunogold-silver procedure. Sporozoites invading hepatocytes released substantial quantities of circumsporozoite protein into the hepatocyte cytoplasm. This intrahepatic cytoplasmic distribution reached a maximal level at about 4 h post-sporozoite injection. As the hepatic parasites continued to differentiate, circumsporozoite protein became undetectable within the cytoplasm of the hepatocytes and became localized around the periphery of each parasite. There was generalized cellular inflammation within the liver of the host, which first became evident at around 4 h post-sporozoite injection and progressed to the formation of well-defined granulomas by 24 h. Such histopathological changes were not seen in rats injected with killed sporozoites, indicating that the cellular inflammation was induced by viable, infective sporozoites. We did not observe cellular infiltration specifically associated with any of the developing hepatic stages that we observed, even up to 28 h post-sporozoite inoculation. These results indicate that viable sporozoites induced rapid and generalized hepatic inflammation in host rats. However, sporozoites that successfully invaded hepatocytes and then proceeded to develop further did not appear to be the target of inflammatory cells until a period beginning at around 40 h post-sporozoite inoculation.

Inflammatory responses to Plasmodium hepatic schizonts within the livers of non-immunized animals have long been assumed to be initiated only after the parasites have matured and begun to burst. However, recent reports of inflammatory responses around hepatic schizonts suggested a re-examination of this issue. We injected Norway-Brown rats of various ages intravenously with Plasmodium berghei sporozoites and studied subsequent liver histopathology. We found that the ability of these rats to mount an inflammatory response is age-dependent. Young (4 weeks) rats had weak inflammatory responses against hepatic schizonts, whereas older (8-10 weeks) rats mounted a strong response. Older rats had many granulomatous reactions within the liver; eosinophils represented a pioneer component of the cellular infiltrate. There was a reduction in the numbers of surviving hepatic schizonts in the older rats, suggesting that these granulomatous and eosinophilic reactions had effectively destroyed some of the hepatic schizonts. We found clear evidence of inflammatory cells (eosinophils) infiltrating hepatic schizonts as early as 40 hours post-injection with sporozoites, a time well before any hepatic schizonts could have burst within the liver. This presents histological evidence that inflammatory cells can recognize and infiltrate intact hepatocytes containing schizonts in immunologically naive animals.

We found BALB/c mice to be on the order of 2,000 times more susceptible to Plasmodium yoelii than Plasmodium berghei sporozoites, as measured by the ability of these sporozoites to differentiate into microscopically detectable hepatic schizonts in the livers of immunologically naive mice. One of the factors that determine the relative insusceptibility of mice to P. berghei sporozoites is the innate cellular inflammatory response that the mice mount after injection with sporozoites. The cellular inflammatory response against P. berghei is initiated soon after sporozoite injection; by 24 h, substantial histopathological changes have developed within the liver. There is considerably less of a cellular inflammatory response against P. yoelii; significant histopathological changes within the liver are not observed until well after hepatic schizonts have begun to rupture at around 44 h postinjection of sporozoites. These differences in the cellular inflammatory response against two different, closely related species of sporozoites are of considerable interest. The data strongly suggest that the BALB/c-P. berghei sporozoite system is a relatively poor biological model for sporozoite immunization studies.