Although the large epidemic of CHIK across the islands of the Indian Ocean is now in decline, outbreaks of the Indian Ocean strains were reported in many other countries, and opportunities for the introduction of CHIKV to China were not limited.
In this study, for the first five cases of CHIK detected in China in 2008, the possible transmission of the virus carried by the travelers was monitored. Although, of the five imported cases, secondary cases were detected among the travelers from groups I and II at eight and five days after the onset of the first case in the group, respectively, we believe that the infections originated in Sri Lanka and Malaysia, respectively, rather than in China, as the disease has an incubation period of about three to twelve days  and no other local close contacts were found to be infected. For the two close contacts of the cases from group III (from Malaysia) who presented with a mild fever and were suspected to have a CHIKV infection, laboratory analysis did not support the suspicion and additional samples were not collected because the individuals left China soon after the onset of the first case in the travel group.
The amino acid differences detected among the imported CHIKVs might be related to their biological or pathogenic characteristics. To examine the genetic variation among the imported CHIKVs, a whole-genome analysis was performed based on 22 published CHIKV sequences. To prevent viral genome mutations caused by cell passage, we obtained our whole-genome sequences directly from clinical serum samples (FD080008, FD080178, and FD080231) or viral isolates (SD08P) and passaged in Vero cells only once.
Nonstructural proteins (nsPs) are involved in viral replication [12, 13], and studies of other alphaviruses have suggested a strong effect for point mutations or deletions in nsP1 and nsP3 on the neurovirulence of Sindbis virus . Strain SD08Pan, which was passaged only once in Vero cells, showed four amino acid changes in its nsPs (nsP1-Q120R, nsP2-G577R, nsP2-N632S, and nsP3- D372N); in comparison, the other sequences identified directly from patient serum showed only one or two amino acid changes (nsP3-M394I in FD080008, nsP2-L539S and nsP4-P181S in FD080178, and nsP2-L539S in FD080231). These may indicate the evolutionary potential of the virus.
In alphaviruses, structural proteins E2 and E1 occur as a closely associated heterodimer on the surface of the virion, with E2 projecting outward and over E1, covering the E1 fusion loops [15, 16]. The fusion of flaviviruses and alphaviruses with host cell membranes requires activation by proteolytic cleavage and a reduced pH as a trigger for conformational change. When alphavirus particles are exposed to a low pH, protein packing in the icosahedral surface lattice changes substantially [17, 18]. The E1-E2 heterodimers dissociate and E1 trimerizes, causing the E2 subunits to move away, permitting clustering of E1 to initiate fusion. Mutations around a hydrophobic pocket could alter the threshold pH for flavivirus fusion clustering . Residue 243 in E2 is likely to be the major determinant of neurovirulence within the structural proteins ; near this position, both FD080178 and FD080231 showed a unique change (E2 K252Q) in which a strongly basic amino acid was changed to a neutral one. This might alter the fusion ability of FD080178 and FD080231. Other changes in structural proteins (e.g., E2-R178H and 6K-V31I in FD08008 and C T8A and E1-A306V in FD080178) were detected; however, the effect on viral maturation and pathogenesis is unclear.
In conclusion, the data reported here confirm that CHIKV was imported into China, although transmission from the travelers' carrying CHIKV did not occur. In addition, the nucleic acid and amino acid changes described here may indicate recent evolution in the ability of the virus to cause an infection. Thus, clinician attention and public health laboratory surveillance should be enhanced in China.