Studies on the Helminth Fauna of Alaska. XXXI. Observations on the Propagation of the Larval Echinococcus multilocularis Leuckart, 1863, In vitro

Abstract

Larval tissue of E. multilocularis, removed aseptically from the livers of experimentally infected rodents (Microtus pennsylvanicus, Clethrionomys rutilus, Lemmus sibiricus), was successfully propagated in culture tubes containing a medium of 40% human ascitic fluid in Hank''s basic salt solution. To this was added 100 units of penicillin and 100 ug of streptomycin per ml. Some tubes were inoculated with malignant human epithelial cells (HeLa strain) and, in some cases, extract of vole embryo (Microtus) or human plasma was added. The tubes were incubated at 35[degree] C. Masses of larval vesicles were removed and stained with Semichon''s acetic carmine (whole mounts) or embedded and stained with hematoxylineosin (sections). Small vesicles were injected intraperitoneally into susceptible hosts to determine whether in vitro development was normal. Larval E. multilocularis proliferates and produces scolices in vitro about as rapidly as in suitable intermediate hosts; and the cultured larval tissue, when injected into such hosts, proliferates and produces scolices. The scolices are presumably infective although no volume of material sufficient for infection of a canine final host has been obtained. In vitro conditions are apparently not detrimental, although proliferation ceases after a certain maximum size is attained. This is probably due to the adjacent medium becoming unfavorable. The HeLa cells formed a substrate for attachment of larval tissue, and it is possible that they stimulated proliferation. Proliferation of larvae in vitro takes place through exogenous budding, the vesicles assuming spherical shape. Metabolic and other studies are planned as well as the production of antigens (for serological diagnosis of hydatid disease) free of the undesirable components of host tissue.