Molecular Epidemiological, Serological, and Pathogenic Analysis of EV-B75 Associated With Acute Flaccid Paralysis Cases in Tibet, China

Abstract

Enterovirus B75 (EV-B75) is a newly identified serotype of the enterovirus B species. To date, only 112 cases related to EV-B75 have been reported worldwide, and research on EV-B75 is still limited with only two full-length genome sequences available in GenBank. The present study reported seven EV-B75 sequences from a child with acute flaccid paralysis and six asymptomatic close contacts in Shigatse, Tibet. Phylogenetic analysis revealed that the Tibetan strain was possibly imported from neighboring India. Seroepidemiological analyses indicated that EV-B75 has not yet caused a large-scale epidemic in Tibet. Similarity plots and boot scanning analyses revealed frequent intertypic recombination in the non-structural region of all seven Tibet EV-B75 strains. All seven Tibetan strains were temperature-sensitive, suggesting their poor transmissibility in the environment. Overall, though the seven Tibetan strains did not cause large-scale infection, prevention and control of the novel enterovirus cannot be underestimated.

Keywords: Tibet; enterovirus B75; phylogenetic analyses; seroprevalence; temperature sensitivity.

FIGURE 1

Time, region, and clinical distribution of EV-B75 worldwide.

FIGURE 2

Maximum likelihood phylogenetic tree based on the entire VP1 coding region sequences of EV-B75 available from the GenBank database. The seven Tibetan EV-B75 strains isolated in this study are indicated by red circles, and the prototype of EV-B75 is indicated by a green circle. The scale bars indicate the substitution per site per year. The numbers at the nodes indicate the bootstrap support for the node (percentage of 1,000 bootstrap replicates).

FIGURE 3

Maximum likelihood phylogenetic tree based on the VP1, P1, P2, and P3 coding regions of the prototype sequence of all EV-B in the GenBank database and the seven Tibet EV-B75 strains in this study. The seven Tibet EV-B75 strains in this study are indicated by red circles, and the prototype of EV-B75 is indicated by a green circle. The scale bars indicate the substitution per site per year. The numbers at the nodes indicate the bootstrap support for the node (percentage of 1,000 bootstrap replicates). Coding sequences of (A) VP1, (B) P1, (C) P2, and (D) P3 are shown.

FIGURE 4

Similarity and boot scanning analysis of the seven Tibetan EV-B75 strains with the prototype of EV-B75, a field EV-107 strain (AB426609-TN-94-0349), and a field CVB1 strain (JN797615-1167438). All seven Tibetan EV-B75 strains were set in a series as a query sequence.

FIGURE 5

Temperature sensitivity test curves of the seven Tibetan EV-B75 strains. Blue and orange lines represent the growth trends of the viruses on RD cells at 36 and 39.5°C, respectively. The Xinjiang EV-B85 strain (HTYT-ARL-AFP02F/XJ/CHN/2011, showing no temperature sensitivity) and the EV-B106 strain (KS-MGTH90F/XJ/CHN/2011, showing temperature sensitivity) were used as experimental controls. (A) Strain Y16/XZ/CHN/2007; (B) strain Y17/XZ/CHN/2007; (C) strain Y18/XZ/CHN/2007; (D) strain Y20/XZ/CHN/2007; (E) strain Y24/XZ/CHN/2007; (F) strain Y25/XZ/CHN/2007; (G) strain Y26/XZ/CHN/2007; (H) strain HTYT-ARL-AFP02F/XJ/CHN/2011 (EV-B85); (I) strain KS-MGTH90F/XJ/CHN/2011 (EV-B106).