In vitro culture of equine strongylidae to the fourth larval stage in a cell-free medium.

An efficient and reliable method is described for the culture of equine strongyles from the third (L3) to the fourth (L4) larval stage. Medium consists of 50% fetal calf serum and 50% NCTC with additions of L-glutamine, NaHCO3, yeast extract, bactopeptone, and dextrose. The gas phase used is of prime importance; it is a mixture of 10% CO2, 5% O2, and 85% N2. Strongylus vulgaris, Strongylus edentatus, Strongylus equinus, Triodontophorus brevicauda, Triodontophorus serratus, Triodontophorus tenuicollis, Oesophagodontus robustus, Cylicocyclus insigne, and mixed species of cyathostomes were cultured to the L4 stage. Oesophagodontus robustus was cultured to the fifth larval stage. Depending on species, 44-95% of Strongylinae L3 inoculated into this system molted to L4. Although some development of the Cyathostominae L3 occurred, only a small portion (1%) completed ecdysis to L4. Viability 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. robustus L4 enhanced development and prolonged viability of these larvae. Hemin had no effect on cultures of S. edentatus or S. equinus, and it was not tested in cultures of other species.