Infectivity and Virulence of Cryptosporidium Genotype H Oocysts in Healthy Adult VolunteersEPA Grant Number: R828035
Title: Infectivity and Virulence of Cryptosporidium Genotype H Oocysts in Healthy Adult Volunteers
Investigators: Chappell, Cynthia L. , Okhuysen, Pablo C. , Tzipori, Saul , Widmer, Giovanni
Institution: The University of Texas Health Science Center at San Antonio
EPA Project Officer: Nolt-Helms, Cynthia
Project Period: March 1, 2000 through March 1, 2003 (Extended to February 28, 2005)
Project Amount: $503,884
RFA: Drinking Water (1999) RFA Text | Recipients Lists
Research Category: Water , Drinking Water
Cryptosporidium parvum is now recognized to be made up of a genetically heterogeneous population of organisms, which can be divided into those infecting humans (genotype 1/H) and those which are transmitted between human and animal hosts (genotype 2/C). Recent studies have shown that stable genotype 1/H isolates can be maintained in gnotobiotic (GNB) pigs. This significant advance will make studies of these isolates possible in human hosts. Experimental cryptosporidiosis in healthy adults has previously revealed variability in infectivity, outcome, and the immune response to geographically-diverse genotype 1/C isolates. Further, a comparison of serologically negative versus serologically positive individuals showed a significant increase in the ID50 in those with pre-existing serum IgG. These antibody positive individuals excreted many fold fewer oocysts than the antibody negatives, suggesting that secondary infections would be less likely to occur with this population. Although, the pre-sensitized volunteers were relatively resistant to low level exposures, high oocyst concentrations resulted in infection and diarrheal illness in a number of the volunteers. Indeed, some illness measures indicated that the diarrhea in antibody positives was more severe than in the antibody negatives.
The proposed experiments will extend these studies to two genotype 1/H isolates. Specifically, the study can be divided into three parts: 1) establish the infectious dose (ID50), clinical outcomes and intensity of infection for two Cryptosporidium genotype 1/H isolates in seronegative individuals; 2) investigate the antibody and cellular responses in volunteers to genotype 1/H isolates; and 3) identify parasite genotype-specific differences and the stability of DNA markers prior to and after passage in pigs and humans.
Two stable genotype H (genotype 1) isolates from HIV-negative donors will be identified and amplified in a gnotobiotic pig and/or a IFN gamma knock out mouse model. Pre- and post- amplified oocysts (in pigs and volunteers) will be analyzed to document genotype stability. The dose response studies will be carried out in healthy adult volunteers. Each volunteer will be given a single oocyst dose and followed for 6 weeks. Clinical signs and symptoms will be documented, and fecal oocysts will be quantitated. Blood and saliva will be collected at specific time points for immunological studies. Endoscopic biopsies will be collected before and after challenge in those who wish to participate in this part of the study. The antibody and cellular responses in peripheral blood will be examined at various time points using crude oocyst extracts and recombinant proteins. New genetic probes will seek to identify parasite factors indicative of differences in volunteer infectivity (if different ID50's are found).
Documenting the outcomes of infection with the 1/H genotype isolates will significantly advance the understanding of community acquired cryptosporidiosis and allow the development of more accurate risk assessment models. Comparison of the two genotype 1/H isolates will provide evidence for potential variations in virulence characteristics, a phenomenon seen in genotype 2/C isolates. Antibody and cellular studies will examine the immune response to exposure and/or infection and provide samples for study of the degree of cross-reactivity between antigens from the two genotypes. These studies will be important in understanding Cryptosporidium infection in the community and will have significant implications for water quality assessments.