title: Failure to propagate Cryptosporidium spp. in cell-free culture.
abstract: The successful propagation of Cryptosporidium parvum in cell-free culture medium was recently reported. To investigate whether this phenomenon could be broadened to include other C. parvum isolates, as well as Cryptosporidium hominis, we attempted to propagate 3 isolates in cell-free medium under reported culture conditions. Cryptosporidium oocysts from C. parvum strains Moredun (MD) or IOWA or C. hominis strain TU502 were added to media containing coagulated newborn calf serum. The cultures were sampled at various times throughout a 45 (IOWA) or 78 (MD, TU502)-day period and were microscopically examined for various life stages of Cryptosporidium. Cell-free cultures harvested on days 45 and 68 postinoculation were tested for in vitro infectivity on Madrin-Darby bovine kidney cells. In vivo infectivity testing was performed using either infant or 2-wk-old immunosuppressed C57BL mice with cell-free cultures harvested on days 52 and 78. Fecal and gut samples collected from mice were examined by modified acid-fast staining. Data from wet mounts, electron microscopy, and in vitro and in vivo infectivity testing showed that the original oocysts did not complete their life cycle and produce new, viable, infectious oocysts in cell-free culture. Thus, we conclude that this is not a universal phenomenon or readily accomplished.
authors.affiliation: Division of Infectious Diseases, Tufts University School of Veterinary Medicine, 200 Westboro Road, Grafton, Massachusetts 01536, USA.
authors.forename: Donald
authors.initials: D
authors.lastname: Girouard
authors.forename: Jennifer
authors.initials: J
authors.lastname: Gallant
authors.forename: Donna E
authors.initials: DE
authors.lastname: Akiyoshi
authors.forename: John
authors.initials: J
authors.lastname: Nunnari
authors.forename: Saul
authors.initials: S
authors.lastname: Tzipori
mesh_terms: D000818:Animals; D002417:Cattle; D003457:Cryptosporidiosis / Q000469:parasitology*; D003458:Cryptosporidium / Q000235:genetics / Q000254:growth & development* / Q000472:pathogenicity / Q000648:ultrastructure; D003470:Culture Media; D016054:DNA, Protozoan / Q000032:analysis; D019084:Fluorescent Antibody Technique, Indirect; D007165:Immunosuppression Therapy; D008018:Life Cycle Stages / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008854:Microscopy, Electron; D033661:Oocysts / Q000648:ultrastructure; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length
chemical_list: D003470:Culture Media; D016054:DNA, Protozoan