Sample collection
A
total of 33 samples were collected from small animal clinics between
2018 and 2020. When the dogs were tested positive using Rapid CPV/CCV Ag
Test Kit (Anigen, Korea) or showed some specific clinical symptoms,
their
fecal
swabs will be mixed with normal saline and preserved
in
centrifuge tubes, which will be packed and mailed to our department and
kept in a -80 °C refrigerator.
Virus
isolation
The
cell line used is the cat kidney F81 cell line. Added 1 mL PBS to each
centrifuge tube and
oscillated
using
an oscillator, then drew 500 μL liquid and filtered through a 0.22μm
filter (Millipore, USA). At the same time, the
cell
suspension was prepared, which was digested by 0.05% pancreatin and
maintained in Minimal Essential Medium (MEM, Gibco, USA) supplemented
with 10% calf
serum.
The cell suspension was added to the culture flask and
inoculated
with
CPV-2
with a proportion of 5%. The culture flask was kept in a
CO2 incubator (MEMMERT, German). We harvested the virus
when the cytopathic effect (CPE) was about 80%.
The
third passage of viruses was used for analysis.