Fig. 2. CYV-ZIKV propagation in A. aegypti.

a–f Five- to six-day-old females were blood-fed with CYV-ZIKV or CYV diluted to 1 × 10⁸ FFU/ml. Control (Con.) was fed with medium. Data are presented as means ± SD of triplicate measurements. Total RNA from the group of 20 midguts (a, n = 3), 20 fat bodies (b, n = 3), 20 legs (c, n = 3), 20 thoraxes (d, n = 3), or 20 heads (e, n = 3) were extracted at different times post-infection, and the viral RNA was detected by real-time PCR. f The viral titers of saliva in 80 mosquitoes were determined by TCID₅₀ assay (n = 3). g, h Total RNA from a single mosquito was extracted, and the viral RNA level was measured by real-time PCR. A dot represents a mosquito. Con. indicates control uninfected mosquitoes. Data are as means ± SD. The P-value was determined by a two-sided unpaired t-test. g Venereal transmission of the virus between females and males (n = 17). h The transovarial transmission between infected females and F1 generation (n = 27). The dashed line represents the detection limit. i, j, k Genetic stability of CYV-ZIKV in mosquitoes during passage 1 (P1) to passage 5 (P5). P0 (passage 0), P2 (passage 2), P3 (passage 3), P4 (passage 4). i The viral titers in the saliva of 80 mosquitoes from P1 to P5 were determined by the TCID₅₀ assay (n = 3). Data are presented as means ± SD of triplicate measurements. Con. indicates uninfected mosquito group. j Mutations of CYV-ZIKV emerged during passage in mosquitoes. (k) Susceptibility of CYV-ZIKV from passage 5 in mosquito (C6/36) and vertebrate cells (293T, BHK-21, and Vero cells) incubated at 28 °C or 37 °C (MOI = 0.1). The ZIKV E protein was immunolabeled with the 4G2 antibody (green staining), and the nucleus was stained with Hoechst 33342 (blue staining) at 3 dpi. The bar indicates 50 μm. Similar results were obtained in three independent experiments. Source data are provided as a Source Data file.