Response of Immunized and Nonimmunized Calves to Cecal Inoculation of First-Generation Merozoites of Eimeria bovis

Abstract

Four experiments were conducted, in each of which 0.4 to 1.9 billion first-generation merozoites of Eimeria bovis were inoculated by laparotomy into the ligated cecum of a nonimmunized calf and a calf immunized with 25,000 or 50,000 oocysts. These merozoites were obtained from calves killed 15 days after inoculation with 2.0 million oocysts. An initial biopsy was performed at the time merozoites were inoculated, and three or four additional biopsies were done in each calf at intervals of 3 to 24 hr during the following 2 or 3 days. The ligature was removed 1 to 2 days after inoculation, and fecal samples collected daily were examined for oocysts. Approximately 1 week after the last biopsy in each experiment, the calves were challenged with an oral inoculation of 1.0 million oocysts and were killed 19 days later. Moderate infection occurred after the inoculation of merozoites in the nonimmunized calves of Experiments 1 and 2, but only slight infections occurred in the nonimmunized calves of Experiments 3 and 4. No infection could be detected in any of the immunized calves, except for a few parasites found in one calf at 24 hr. These findings suggest that the penetration of host cells by first-generation merozoites is inhibited in immunized calves. All of the immunized calves and two of the four nonimmunized calves showed little or no infection after challenge, but the nonimmunized calves of Experiments 3 and 4 became moderately infected. These results indicate that infection caused by inoculation into the cecum of first-generation merozoites stimulates the development of immunity. Although immunity against coccidiosis is well known in both mammals and birds, little has been learned about the way in which the parasites are affected by the immune reaction. The occurrence of unusually large first-generation schizonts in E. bovis, with the resulting ready availability of merozoites, makes this species suitable for investigation of this problem. The present study was undertaken to determine whether first-generation merozoites of E. bovis inoculated into the cecum of immunized and nonimmunized calves would be able to invade the epithelium and initiate an infection, and to determine whether infections so produced would result in immunity. Received for publication 22 January 1964. * Supported in part by research grant E-2374 from the Institute of Allergy and Infectious Diseases and by a fellowship (GPM-18,586) from the Division of General Medical Sciences, Public Health Service. Published as Journal Paper No. 385, Utah Agricultural Experiment Station. t Present address: College of Veterinary Medicine, University of Illinois, Urbana, Illinois. MATERIALS AND METHODS Twelve Holstein-Friesian male calves were used in a series of four biopsy experiments. Four of these calves were used as sources of merozoites for inoculation. The methods of housing, feeding, inoculating, sampling for oocysts, collecting and concentrating merozoites, and performing the biopsies were as described previously (Hammond, Andersen, and Miner, 1963a, b). In each experiment, one calf was immunized by inoculating it when 1 to 2 weeks old with 25,000 or 50,000 E. bovis oocysts. Approximately 1 month later, this animal and a nonimmunized calf of similar age were each given a cecal inoculation with 0.4 to 1.9 billion first-generation merozoites of E. bovis. These merozoites were obtained by scraping the intestinal mucosa of a calf killed 15 days after inoculation with 2.0 million oocysts. More than 90% of these oocysts were E. bovis; the remainder consisted chiefly of E. ellipsoidalis and E. auburnensis. Gametocytes of these two species were seen, in addition to E. bovis schizonts, in the intestinal sections of the calves used as sources of merozoites. However, the cecal inoculations resulted in pure E. bovis infections, as shown by the finding of oocysts of this species only in the fecal samples. In each experiment equal numbers of merozoites were introduced by laparotomy into the ceca of the two calves. Three or four additional biopsies