Using IHC, the parasite was detected in only one experimentally
infected animal. In approximate calculation from the qPCR
results, the number of parasites per 1 g of tissue sample of
amygdala was about 110 parasites on average. Assuming a specific
gravity of brain 1, calculated simply, 1 g of tissue sample could
provide 2,500 pieces of 1-cm2 tissue sections when sliced 4 m
thick. This means that at least 23 sections are needed to find one
parasite even when the parasites are distributed evenly in tissue. In
general, the parasites would be sparsely distributed in the tissue as
colonies of tachyzoites or cysts. Therefore, detection of the parasite
by histopathological and immunohistochemical techniques is
difficult with low sensitivity. In contrast, qPCR is highly sensitive
and specific, especially in tissues with low levels of parasites.
In the present study, the importance of sampling and analyzing
brain tissue was confirmed in cattle experimentally infected with
N. caninum. Our results show that the amygdala, hippocampus,
and hypothalamus samples are most appropriate for qPCR assays.
Additionally, sections from the corpus striatum and diencephalon
were most useful in histopathological analysis. Our findings show
that these areas are most useful for evaluating the extent of N.
caninum infection, investigating the pathogenesis of neosporosis,
and evaluating antiparasitic drugs and vaccines against such organisms.
Although the Nc-1 isolate has a lower ability for cystformation, this isolate has been used in many studies of the pathogenesis
of neosporosis and can induce fetal death in experimentally
infected cattle (20–22). On the other hand, cystogenic isolates,
such as Nc-Liv, may show some differences in brain parasite
burdens and pathological changes. Thus, in future work, comparative
study of different isolates of N. caninum will be important to
understand the pathogenesis.