We previously demonstrated that ducks immunized with recombinant VP2 protein (rVP2) of parvovirus in conjunction with CpG oligodeoxynucleotides (ODN) containing three copies of GACGTT motifs had significantly ([Formula: see text]) increased antibody titer, splenocyte proliferation index, amount of CD4[Formula: see text] and CD8[Formula: see text] cells, and mRNA expression of cytokines (IFN-[Formula: see text], IFN-[Formula: see text], IL-6, and IL-8) in comparison with those immunized with rVP2 plus aluminum hydroxide. In the present study, ducklings hatched from eggs produced by ducks immunized with rVP2 and various adjuvants were used to analyze the passive immunity of maternal antibodies followed by an experimental challenge with live parvovirus. Results indicated that the titer of VP2-specific antibodies and the body weight of ducklings produced from ducks immunized with rVP2[Formula: see text]CpG ODN were significantly ([Formula: see text]) higher than those of ducklings from other groups at day 7 after the challenge. The autopsy report revealed that ducklings in groups other than rVP2[Formula: see text]CpG ODN and rVP2[Formula: see text]mock had hemorrhage in the liver and intestinal mucous at 11 days after the challenge, and the presence of parvovirus was confirmed by PCR using parvovirus-specific primers. Therefore, immunizing ducks with rVP2[Formula: see text]CpG ODN containing three copies of GACGTT motifs is able to enhance the passive immunity, which protects their ducklings from parvovirus infection.

Adjuvants are important components of vaccine formulations. Effective adjuvants line innate and adaptive immunity by signaling through pathogen recognition receptors. Synthetic cytosine-phosphate-guanine (CpG) oligodeoxynucleotides (ODNs) have been shown to have potentials as adjuvants for vaccines. However, the immunostimulatory effect of CpG is species-specific and depends on the sequence of CpG motifs. A CpG ODN (2135), containing 3 identical copies of GTCGTT motif, was previously reported to have the strongest effects on bovine peripheral blood mononuclear cells (PBMC). Based on the sequence of 2135, we replaced the GTCGTT motif with 11 other sequences containing CG and investigated their effects on bovine lymphocyte proliferation. Results showed that the CpG ODNs containing 3 copies of GACGTT motif had the highest lymphocyte stimulation index (7.91±1.18), which was significantly (P<0.05) higher than that of 2135 (4.25±0.56). The CpG ODNs containing 3 copies of GACGTT motif also significantly increased the mRNA expression of interferon (IFN)-α, interleukin (IL)-12, and IL-21 in bovine PBMC. When dairy cows were immunized with the keyhole limpet hemocyanin (KLH) antigen formulated with CpG ODNs containing 3 copies of GACGTT, production of KLH-specific antibodies in serum and in milk whey was significantly (P<0.05) enhanced. IFN-γ in whole blood stimulated by KLH was also significantly (P<0.05) increased in cows immunized with KLH plus CpG ODNs. Our results indicate that CpG ODNs containing 3 copies of the GACGTT motifs is a potential adjuvant for bovine vaccines.

Recombinant parvovirus VP2 (rVP2) was formulated with different types of adjuvant, including aluminum adjuvant and CpG oligodeoxynucleotides (ODNs), and the immunological responses after vaccination in ducks were examined. In comparison with the control group, production of rVP2-specific antibodies, expression of cytokines in peripheral blood mononuclear cells (PBMC) stimulated by rVP2, and percentage of CD4(+)/CD8(+) cells in PBMC were significantly increased in ducks immunized with rVP2 formulated with CpG ODNs containing 3 copies of GACGTT motif. CpG ODNs with GACGTT motifs might be used to improve the efficacy of vaccines for ducks.