IN-VITRO CULTURE OF EQUINE STRONGYLIDAE TO THE 4TH LARVAL STAGE IN A CELL-FREE MEDIUM

An efficient and reliable method is described for the culture of equin e strongyles from the third (L(3)) to the fourth (L(4)) larval stage. Medium consists of 50% fetal calf serum and 50% NCTC with additions of L-glutamine, NaHCO3, yeast extract, bactopeptone, and dextrose. The g as phase used is of prime importance; it is a mixture of 10% CO2, 5% O -2, and 85% N-2. Strongylus vulgaris, Strongylus edentatus, Strongylus equinus, Triodontophorus brevicauda, Triodontophorus serratus, Triodo ntophorus tenuicollis, Oesophagodontus robustus, Cylicocyclus insigne, and mixed species of cyathostomes were cultured to the L(4) stage. Oe sophagodontus robustus was cultured to the fifth larval stage. Dependi ng on species, 44-95% of Strongylinae L(3) inoculated into this system molted to L(4). Although some development of the Cyathostominae L(3) occurred, only a small portion (1%) completed ecdysis to L(4). Viabili ty in cultures of all species remained high (>60-70% larvae surviving) for at least 4 wk (cyathostomes) and as long as 6 mo (S. edentatus). The addition of equine hemin to cultures of S. vulgaris and O. robustu s L(4) enhanced development and prolonged viability of these larvae. H emin had no effect on cultures of S. edentatus or S. equinus, and it w as not tested in cultures of other species.