The pathobiology and immunology of cryptosporidiosis

Abstract

There were three aims of this study: firstly, to investigate cryptosporidiosis in a symptomatic laboratory animal model where diarrhoea was the most important clinical sign; secondly, to compare the infection in this model with that in lambs; thirdly, to determine the neutralizing ability of specific immunoglobulins

Strains of rats and mice were each infected with three isolates of Cryptosporidium parvum. Using histopathological scoring methods it was shown that the distribution of infection depended more on the host species than on the isolates used. The inbred Lister rat strain was selected as a suitable laboratory animal host because infection was symptomatic with a distribution and age susceptibility which paralleled that in lambs

The infection in mice was asymptomatic and in the small intestine occurred principally in the ileum. Excystation conditions were shown to be more favourable in the mouse ileum by comparing the number of sporozoites collected from segments of intestine after oral inoculation of oocysts. In vitro experiments showed that exposure of oocysts to a o! shift from approximately 3.6 to 8.4 was an important excystation stimulus. The parasite's asexual reproductive capacity was examined by counting merozoites in mouse ileal surface mucus. It was shown that the amount of surface mucus increased during infection in association with increasing numbers of merozoites. A method was described which enabled merozoites to be collected for use as antigen.

The kinetics and specificity of antibodies produced in response to Cryptosporidium infection were examined in rats and lambs using IFA and immunoblotting techniques respectively. For rats and lambs specific serum antibodies were detected within one week of oocyst inoculation and titres did not decrease over an observation period of approximately 40 days. Higher serum titres were found in rats infected at 15 days compared to those infected at 4 days of age. This may have reflected the postnatal development of the gut associated lymphoid tissue in rats whereby the younger animals were immunologically immature at the time of infection.

Numerous oocyst antigens, with molecular weights greater that 45Kd.,v/ere recognized by antibody in convalescent rat and lamb sera. In addition, two antigens with estimated molecular weights of 23Kd. and between 12.3 and 17.3Kd. were features on serum immunoblots. It was shown in lambs that declining oocyst output was associated with increasing titres of IgA and IgM in faecal extracts and that both antibodies had similar oocyst antigen specificity. Most of the merozoite antigens recognized by IgA appeared in a molecular weight range from 66 to 180Kd., as did most of the oocyst antigen: recognized by this antibody. However, a 23Kd. oocyst antigen and others between 45 and 66Kd. recognized by IgA were not detected by this antibody on merozoite blots. These antigenic differences may be helpful in determining which stage of the parasite is most important to the development of protective immunity.

Mucus extracts from infected and uninfected lambs were fractionated on Bio-Gel A-1.5m. In vitro sporozoite agglutination was associated not only with fractions containing specific IgA but also glycoprotein fractions devoid of detectable specific antibody. Pretreatment of sporozoites with IgG, separated from hyperimmune lamb serum by affinity chromatography, was shown to neutralize their capacity to infect 5-day old rats. This may have important implications for passive protection of young ruminants.