Culture requirements of Ascaris suum larvae using a stationary multi-well system: increased survival, development and growth with cholesterol.

Ascaris suum third-stage larvae (L3) were recovered from infected rabbits and cultured in a stationary multi-well plate system. Several different culture media including Medium 199, NCTC-135, Minimum Essential Medium-Eagle, Dulbecco's Modified Eagle's Medium, RPMI 1640, McCoy's 5A and Neuman-Tytell medium were tested to determine which was best for overall larval survival, development and growth. Larvae developed from L3 to fourth-stage (L4) in all media tested, but larval survival and the yield and growth of L4 were superior in RPMI 1640. Pyruvate is an important medium component because its addition to RPMI 1640 enhanced the yield and growth of L4 while its removal from DMEM reduced larval survival and the yield and growth of L4. Cholesterol markedly enhanced larval survival and the yield and growth of L4 when added to RPMI 1640 as either a soluble supplement in Tween 80 or to a lesser degree, as a component of liposomes. The multi-well culture plate system is a convenient method for determining the effects of different media and changes in media composition on larvae in vitro.