Cryptosporidium parvum oocyst DNA samples (n ؍ 184) from humans with cryptosporidiosis contracted during foreign travel or during outbreaks in the United Kingdom were characterized genetically and categorized by single-strand conformation polymorphism (SSCP)-based analysis of the small-subunit gene (pSSU) (ϳ300 bp) and second internal transcribed spacer (pITS-2) (ϳ230 bp) of nuclear ribosomal DNA. The two recognized genotypes (types 1 and 2) of C. parvum could be readily differentiated by a distinct electrophoretic shift in the pSSU SSCP profile, associated with a nucleotide difference of ϳ1.3 to 1.7%. Of the 102 samples from cases contracted during foreign travel, 88 (86.3%) were identified as C. parvum type 1 and 14 (13.7%) were identified as type 2. For outbreak samples, unequivocal differentiation between type 1 (n ؍ 20; one child nursery outbreak) and type 2 (n ؍ 62; two waterborne outbreaks) was also achieved. Nucleotide variation in pITS-2 (both within and among samples representing each genotype) was substantially greater (10 to 13 different profiles for each genotype, relating to sequence differences of ϳ1 to 42%) than that in pSSU. SSCP analysis of pITS-2 for all samples revealed that some profiles had a broad geographical distribution whereas others were restricted to particular locations, suggesting a link between some subgenotypes and the geographical origin or source. Comparative denaturing polyacrylamide gel electrophoretic analysis revealed the same genotypic identification and a similar subgenotypic classification of samples as SSCP analysis. The findings of this study, particularly the detection of intragenotypic variation by SSCP, should have significant diagnostic implications for investigating transmission patterns and the monitoring of outbreaks.Species of Cryptosporidium are oocyst-forming apicomplexan parasites which infect a wide range of vertebrate hosts. Intestinal disease caused by these parasites is called cryptosporidiosis and is transmitted via the fecal-oral route (e.g., in child day care centers) and can be associated with waterborne outbreaks (27). The clinical signs of cryptosporidiosis in humans are mainly diarrhea, dehydration, malabsorption, weight loss, and/or wasting (27). In many cases, infection is self-limiting and resolves spontaneously, although chronic infections may develop in high-risk patient groups, particularly neonates and immunocompromised individuals (those with congenital or acquired immunodeficiencies or those undergoing immunosuppressive chemotherapy). The accurate identification of Cryptosporidium to the species or "strain" (i.e., genotypic) level is important for studying transmission patterns and genetic structures and is central to the control of cryptosporidiosis (9, 26, 34). Currently, 11 species of Cryptosporidium are recognized (9, 10), of which C. parvum is of major significance in humans. However, these species cannot be distinguished from one another simply on the basis of host occurrence and/or parasite morphology (9, 27). Consequently, a...