Development of ELISA for detection of antibody to African swine fever virus
The P72 and P30 structural proteins of African swine fever virus (ASFV) were expressed in the prokaryotic system. The expressed recombinant proteins were purified by Ni-NTA column, and each protein was expressed at MW of 72 and 30 kDa, respectively, and they were all soluble proteins. The Western blot test showed that the two recombinant proteins can be recognized by positive pig serum from African swine fever virus infection. Another tested by the ELISA method, the P72 recombinant protein showed better antigen-antibody reactivity to ASFV-positive serum than P30. The P72 recombinant protein was purified and coated to the microplate to develop the indirect-ELISA method for African swine fever antibody detection. 1,163 African swine fever negative sera collected from Taiwan pig farms were used to test the efficiency of the homemade indirect-ELISA kit, and the imported ASFV-positive serum was selected as a control serum. The results showed that taking the mean of negative serum plus 3 folds of standard deviations (3SD) as the cut-off value (Cut-off), the specificity of homemade indirect antibody-ELISA was 97.5%. A commercial African swine fever blocking ELISA kit was used to test 255 African swine fever negative swine sera from domestic pig farms. The results showed that the specificity of the detection was 100%.
Surveillance on Ephemeral Fever Virus Disease
Phylogenetic analysis of the surface glycoprotein (G) gene of 21 Taiwanese bovine ephemeral fever (BEF) virus isolates revealed that the identities of the nucleotide sequences ranged from 96.2% to 99.5%. Moreover, among the isolates identified in Hualien County, the isolates identified in 2019 were genetically more similar to the isolates from 2012 than those from 2014. Furthermore, the results suggest that the 2019 isolate strain came from the 1996-2012 endogenous groups. A cross neutralization test between the 2019 strain and the vaccine strain resulted in an R value of 0.88 b, indicating that the commercially available BEF vaccine still confers good protection against the 2019 strain. To evaluate the effectiveness of different adjuvants, available data from the BEF serological surveillance database were compared for four farms. Two weeks after administering vaccines adjuvanted with either phosphate aluminum gel or long-term W/O/W, the mean antibody titers were 142.03±35.39 and 430.54±257.25, respectively. The W/O/W-adjuvanted vaccine was more effective, not only eliciting 3.36X more antibody titers than the aluminum-gel, but also providing titers ≥ 4096 in 33.9% of vaccinated cattle. On the other hand, common side effects of this W/O/W adjuvanted vaccine included fever, anorexia for 2-3 days, and a drop in milk production in 5-9%of vaccinated cattle, with these side effects usually disappearing after 10 days. To try to minimize these side effects, one farm only administered a half dose booster shot to heifers. The heifers were found to be healthy and their mean antibody titer was 304.44±152.03 at day 81, which was roughly the same as the previous field tests where full doses were administered. We thus suggest that a reduction in either the volume of vaccine adjuvant, or the vaccine concentration administered in a single dose should make the new BEV vaccine safer as well as allow it to provide better lasting long-term immunity.