Kinetics of mucosal ileal gamma-interferon response during cryptosporidiosis in immunocompetent neonatal mice.

The kinetics of serum and ileal interferon-gamma (IFN-gamma) content were determined during recovery from cryptosporidiosis in NMRI suckling mice. A total of 60 mice aged 4 days were inoculated by intragastric gavage with 10(4) cryptosporidia (n = 30) or phosphate-buffered saline (n = 30). Six animals per group were killed on days 0, 3, 6, 9 and 13 postinoculation. Blood samples and ileum were collected. Experimental infection was followed by a rise in parasite load in the ileum starting on day 3 postinfection, which peaked at day 6 postinoculation. Ileal IFN-gamma levels increased rapidly in parasitized mice from day 3 to day 6, then fell rapidly. These levels were significantly higher than the control values (day 3 P < 0.05, days 6 and 9 P < 0.001). IFN-gamma secretion began before parasite excretion, but the curves of these two parameters correlated positively. Recovery from cryptosporidiosis in immunocompetent neonatal mice is thus associated with an early and marked increase in ileal IFN-gamma content.

A high-yield outbred suckling mouse model of cryptosporidiosis.

Outbred suckling mice (NMRI strain) were used as hosts. They were initially inoculated with oocysts of human origin, and subsequently with parasites recovered from the mouse ileal mucosa. Cryptosporidia were counted in an aliquot of whole-ileum homogenate. Parasite load was expressed as cryptosporidia per centimeter of ileum. Serial passage of C. parvum in NMRI mouse litters led to a gradual amplification of parasite burden relative to animals initially inoculated with the human isolate.

Determination of anti-Cryptosporidium coproantibodies by time-resolved immunofluorometric assay.

The role of the mucosal immune response against Cryptosporidium has been suggested by studies on the therapeutic effects of hyperimmune colostrum. In order to study the intestinal response to this infection, we have developed a sandwich-type time-resolved immunofluorometric assay for the determination of anti-Cryptosporidium coproantibodies. This assay has the inherent sensitivity of an immunoassay without the problems due to background responses from other biological compounds, and is thus suitable for faecal samples. The intra-assay coefficients of variation (5.1%, 4.6%, and 5.8% for immunoglobulins (Ig) A, M and G respectively), inter-assay coefficients of variation (9.4%; 10.5% and 12.2% for IgA, IgM and IgG, respectively) and specificity (100% for all 3 isotypes) were all satisfactory. Using this assay to study 12 patients with the acquired immune deficiency syndrome (AIDS) who were infected with cryptosporidiosis, we found a marked elevation of anti-Cryptosporidium IgA and IgM coproantibody titres relative to 18 healthy control values, but no correlation with the gravity of the infection in terms of oocyst shedding. These results suggest that a non-protective mucosal immune response develops to Cryptosporidium in AIDS patients.

Sexual stage development of cryptosporidia in the Caco-2 cell line.

We used the spontaneously differentiated human intestinal epithelial cell line Caco-2 to develop an in vitro model of Cryptosporidium sp. infection. The mean cell infection rate was 3% +/- 2%. Asexual stages of cryptosporidia were observed on day 2 postinoculation. Transmission electron microscopy showed the presence of macrogametes at day 5. This cell line appears to be suited to the study of the mechanisms by which biological agents inhibit both sexual and asexual development of cryptosporidia.