A total of 516 slaughter-age ostrich sera were collected in Japan during 2006-2009. Sixty-one of five hundred and sixteen were positive by virus neutralization (VN) test and the titer of most positive samples was low level. Within the 61 positive sera, 35 sera were collected from unvaccinated ostriches. This result implies that these ostriches might have been infected naturally with low-virulent Newcastle disease virus (NDV). Within the 455 negative samples, 125 samples were from vaccinated ostriches. Since ostrich farmers use live attenuated vaccines, it is reasonable that the titer decreased to below detection level by 1 or 1.5 year-old. The above data indicate that NDV has infiltrated into ostrich farms in Japan, and that the efficacy of ostrich ND vaccination is often time-limited.