To evaluate neurotropism of APMVs, virus replication was evaluated in primary chicken neuronal cells in vitro. We first evaluated infection (MOI of 0.1 PFU) in neuronal cells by confocal microscopy analysis of cells that had been immunostained at 72 hpi with N protein against each respective APMV (Fig. 3A) . Expression of the N protein of APMV-1 strain BC was clearly detected in dendrites and axons. In contrast, APMV-1 strain LaSota failed to replicate in neuronal cells even after supplementation with allantoic fluid as a source of protease. Among serotypes 2-9, APMV-3 was able to replicate well without the addition of allantoic fluid, whereas the presence of APMV-7 and -8 antigens in infected cells was sporadically detected. Replication of the other APMVs was not detected up to 3 dpi. We also examined replication in the neuronal cell cultures by collecting supernatants from the cultures at 12 h intervals and assaying for infectious virus by titration. This confirmed the ability of APMV-3 to replicate in neuronal cells, although its titer was lower than that of neurovirulent APMV-1 strain BC (Fig. 3B) . Titers of BC and APMV-3 in neuronal cells increased gradually and reached 5.5 and 3 log TCID 50 /ml, respectively, at 72 hpi. In contrast, other APMVs, including APMV-1 strain LaSota, were not detected in the supernatants of neuronal cells, indicating their inability to replicate in these cultures of primary chicken neuronal cells.