Pathogenesis and clinical signs in animals.

In order to determine the pathogenicities of chimeric viruses of the seven serotypes in animals (pigs [n = 2], mice [n = 3], and suckling mice [n = 10]), two pigs each were inoculated with viruses of each serotype in the footpad, which is a region sensitive to FMDV, at 10^5.0 50% tissue culture infective doses (TCID₅₀)/0.1 ml and observed for 2 weeks. The clinical score was determined by the addition of points distributed as follows: (i) an elevated body temperature of 40°C (score of 1), >40.5°C (score of 2), or >41°C (score of 3); (ii) reduced appetite (1 point) or no food intake and food left over from the day before (2 points); (iii) lameness (1 point) or reluctance to stand (2 points); (iv) the presence of heat and pain after palpation of the coronary band (1 point) or not standing on the affected foot (2 points); (v) vesicles on the feet, dependent on the number of feet affected, with a maximum of 4 points; and (vi) visible mouth lesions on the tongue (1 point), gums or lips (1 point), or snout (1 point), with a maximum of 3 points (32). After challenge inoculation, levels of virus in nasal discharge and serum samples were monitored for 9 days by collecting the samples at 1-day intervals; viruses were detected by using the real-time RT-PCR method.

After intraperitoneal inoculation of 10^5.0 TCID₅₀/0.1 ml, fatality rates and weight changes in C57BL/6 mice (n = 3) were observed. After virus challenge, survival rates and weight changes in adult mice (C57BL/6) were observed until the 12th day after inoculation. Suckling mice (BALB/c) (n = 10) sensitive to FMDV were inoculated with 10^5.0 TCID₅₀/0.1 ml at 7 days of age, and only their survival rates were investigated for 8 days. The PrioCHECK FMDV NSP kit (Prionics AG, Schlieren-Zurich, Switzerland), an ELISA kit for the detection of FMDV NSP antibodies in serum samples of pigs and goats, was employed to detect NSP antibodies until the 12th day postinfection, because the antibody could generally be detected at 10 to ∼12 dpi.