Development Of An Indirect VP2-ELISA To Detect Antibodies Of Infected FMDV And A 8BF-ELISA Kit Using Tandem Epitopes Of 3B To Differentiate Infected From Vaccinated Cattle

Foot-and-mouth disease (FMD) caused by foot-and-mout disease virus (FMDV) is a highly contagious and acutely infected disease of cattle, pigs, sheep, goats and wild ruminant species. It has an economically devastating impact on affected countries, where it creates great losses to productivity and considerable economic losses to the husbandry industry. Whenever the disease has happened; it is must reported to the OIE. Since the Asia 1 FMDV first reported in 2005 in china, there are several outbreaks with the Asia 1, O and A serotypes. There is a complex policy to control the FMD mainly by the preventive vaccination in china, where the most animals are vaccinated 2-4 times every year. So the accurate and in-time diagnosis and routine surveillance become very important for the disease control. Thus, diagnostic methods play an important role for FMD control.The complete gene encoding the structural protein VP2 of FMDV was subcloned into expression vector pPROEXTM HTb and expressed in DH5αcells. Purified VP2 protein reacted positively with serotype-specific cattle sera of 5 serotypes of FMDV (O, A, C, SAT 2 and Asia 1) by western blot. An indirect ELISA (VP2-ELISA) was developed using purified protein to detect FMDV antibodies in cattles. The protein had no cross-reaction with the positive sera of other bovine diseases. The specificity was 100% to detect the negative cattle serum from Canada where was free of FMD without vaccination. The sensitivity was 97.3% to detect infected sera. Comparison with four commercial kit showed a coincidence rate of 69.0%,95.0%,90.4% and 86.8% against vaccinated cattle serum, respectively. Thus the VP2-ELISA can be used to detect the infected and vaccinated cattle serum.To differentiate antibodies induced by FMDV infection from those induced by vaccination, an indirect ELISA was established then the kit was developed, using purified tandem epitopes of 3B protein as antigen. A large number of sera from naive, vaccinated and infected cattle were examined for determination of the cut-off value of the method. The cut-off value was established as follows: positive, S/P≥0.3; suspicious, 0.2≤value<0.3; negative, S/P <0.2. Seroreactivity to 8BF in some infected cattle was maintained through 10 months. The rate of agreement with Ceditest? FMDV-NS ELISA and the 3ABC-I-ELISA was up of 95% by detection of 101 field cattle serum. The performance of this assay was further validated by 3ABC-I-ELISA kits. The coincident rate was 94.5% (499/528), respectively. The number of 2026 cattle sera derived from with different situations (na?ve, unknown, immune and infectious status) were detected. The prevalence of 8BF antibodies reached 72.2% in some diseased cattle herds. The described 8BF-ELISA is safe, cheap and also easy to perform in large scale of serological surveys. The high specificity and sensitivity makes this test a good tool for better distinguishing between infected and vaccinated cattle.