First isolation and identification of Zika virus in China

Abstract

Objective To establish a method for the isolation of Zika virus and to gather experiences for viral isolation. Methods Suckling mice at age 1-3 days were inoculated with serum samples positive for Zika virus through intracranial injection. All mice were sacrificed 6 days after the injection. Viral nucleic acids were extracted from brain, heart, liver, spleen, lung, kidney, muscle, skin and intestine tissue samples and analyzed by real-time RT-PCR. The supernatants of brain tissues positive for Zika virus were used for subculturing. Nested PCR was performed to amplify the NS5 gene of the isolated virus. The sequences of NS5 gene were analyzed by using MEGA6.0 software. Results All of the tissue samples were positive for Zika virus. Higher viral loads were detected in heart and brain tissue samples with cycle threshold (Ct) values of 24.4 and 25.3, respectively. The second generation of Zika virus was identified in suckling mice brain tissues 2 days after infection by using real-time RT-PCR. The amplified product of nested PCR was 972 bp in length. Sequencing analysis showed that the isolated Zika virus (GDZ16002 strain) belonged to the Asian lineage. Conclusion A strain of Zika virus was successfully isolated in China by using intracranial injection via a suckling mouse model. The isolated Zika virus belonged to the Asian lineage. Key words: Zika virus; Viral isolation; Arbovirus; Suckling mice