In the same way, vp1s from 14 CA16 strains isolated in this study, 14 sequences obtained from GenBank and EV71 strain BrCr used as an outgroup for phylogenetic tree analysis showed that lineage B2 of CA16 circulated in Beijing during 2007 to 2009 (Figure 1C). The phylogenetic analysis of complete CA16 vp4s including
1 sequences isolated in this study, 14 sequences obtained from GenBank and EV71 strain BrCr used as an outgroup showed that Alisertib manufacturer the CA16 viruses isolated in Beijing belonged to lineage C (Figure 1D), which was consistent with results from vp1s. Figure 1 Phylogenetic analysis based on EV71 vp1s (A), EV71 vp4s (B), CA16 vp1s (C) and CA16 vp4s (D). The unrooted phylogenetic trees were generated by the neighbor-joining SB273005 method on the basis of a multiple alignment of the nucleotide sequences of EV71 vp1s, EV71 vp4s, CA16 vp1s and CA16 vp4s. The sequences in the dendrograms marked by red circle (○), green triangle
(Δ) and blue square (□) were isolated in this research (additional file 2) while other sequences were obtained from GenBank (additional file 1). CA16 strain G-10 was used as an outgroup in Figure 1A and Figure 1B while EV71 strain BrCr was used as an outgroup in Figure 1C and Figure 1D. Detection of IgM and IgG against EV71 and CA16 in serum samples by Western blot using expressed VP1 and VP4 as antigens The VP4s of EV71 (amplified from specimen s67) and CA16 (amplified from specimen s401) as well as VP1s
of EV71 (amplified from specimen s108) and CA16 (amplified from specimen s390) were expressed in E. coli BL21 and used as antigen by Western Blot to detect specific IgM antibodies in serum samples collected from children with acute enterovirus (EV) infections (Figure 2). Out of 14 serum samples from children with acute EV71 infection, 12 were positive for VP1 of s108 (EV71) and 1 for VP1 of s390 (CA16). Out of 12 serum samples from children with acute CA16 infections, the number of positive serum samples for s108 VP1 and s390 VP1 were 3 and 7, respectively. This result suggested that VP1s from EV71 and CA16 could Urease be used for the detection of IgM specific antibodies in serum samples from patients with acute infections (Table 2). When expressed VP4s of s67 (EV71) and s401 (CA16) were used as antigen to detect specific IgM, all of these 26 serum samples were negative, which raised the question about the antigenicity of the expressed VP4s from EV71 and CA16. Figure 2 Part of the results of the detection of IgM against s108 (EV71) VP1 (A), s67 (EV71) VP4 (B), s390 (CA16) VP1 (C) and s401 (CA16) VP4 (D) by Western Blot. Western blot assay using goat anti-human IgM as secondary LEE011 antibody.