In this study, 6 lepidopteran cell lines derived from Trichoplusia ni (Tn5), Lymantria dispar (LdElta), Bombyx mori (Bm-5), Spodoptera frugiperda (Sf21), Spodoptera exigua (Se-1), and Choristoneura fumiferana (Cf-124T) were investigated for susceptibility to Malacosoma neustria nucleopolyhedrovirus (ManeNPV) isolated in Turkey. Infection of Malacosoma disstria (Md203) cell line by ManeNPV was used as a control. In ManeNPV-Md203 infection, polyhedral inclusion bodies (PIBs) were detected at 36 h p.i. and observed in all infected cells by 120 h p.i., and the infectious virus titer increased 10(3)-fold. At 24 h p.i., the efficient replication of viral DNA was detected. in this Study, we determined that Tn5 and LdElta cells were productive to ManeNPV, and the replication of viral DNA occurred at 24 h p.i. PIB productions were determined in both cell lines at 36 h p.i., and the infectious virus titer in both cell lines increased 10-fold. By contrast, infections of Sf21 cells were not found productive. However, infected cells had apoptosis-like structures. Finally, no cytopathic effects, inclusion bodies, and viral DNA replication were detected in infected Bm-5, Se-1, and Cf-124T cell lines. Therefore, these cell lines were accepted as non-productive. This is the first Study on the host specifity of ManeNPV in vitro. Our results suggest that Tn5 and LdElta cell lines can provide an important model for studying ManeNPV in vitro system as in the Md203 cell line system.