Fig. 6
Glutamate-mediated GABA generation facilitates infection of mosquito vectors by mosquito-borne viruses. a The oral introduction of glutamic acid enhanced the prevalence of DENV-2 infection in A. aegypti. A mixture, which contained 1% sucrose (500 µl), supernatant from 6 × 105 p.f.u./ml of DENV-2-infected Vero cells (cultured in serum-free medium) (500 µl), and 10 µg/ml or 100 µg/ml glutamic acid, was used to feed A. aegypti via an in vitro blood feeding system. Mosquitoes fed this mixture without glutamic acid served as a negative control. b, c Knockdown of the AaGAD1 gene suppressed glutamate-mediated GABA generation b, resulting in reduced DENV infection c with sucrose feeding. AaGAD1 was silenced by inoculation with AaGAD1 dsRNA. Mosquitoes inoculated with GFP dsRNA served as negative controls. Three days post dsRNA treatment, a mixture that containing 1% sucrose (500 µl), supernatant from DENV-2-infected Vero cells (cultured in serum-free medium) (500 µl), and 100 µg/ml glutamic acid, was used to feed A. aegypti via an in vitro blood feeding system. d–f Sucrose feeding with glutamic acid enhanced the prevalence of SINV infection in A. aegypti d, JEV infection in C. pipiens pallens e and TAHV infection in C. pipiens pallens f. A mixture, which contained 1% sucrose (500 µl), supernatant from either 2 × 106 p.f.u. ml−1 of SINV-infected d, 5 × 105 p.f.u./ml of JEV-infected e and 5 × 106 p.f.u./ml of TAHV-infected f Vero cells (cultured in serum-free medium) (500 µl), and 100 µg/ml glutamic acid, was used to feed mosquitoes via an in vitro blood feeding system. Mosquitoes fed on this mixture without glutamic acid served as negative controls. b The numbers on the Y-axis represents the amount of GABA inoculated per mosquito. The results were reproduced twice. a, c–f Mosquito infectivity was determined by TaqMan qPCR or SYBR Green at 8 days post-blood meal. The number of infected mosquitoes relative to the total numbers of mosquitoes is shown at the top of each column. Each dot represents one mosquito. The data upper mosquito number are represented as the percentage of mosquito infection. Differences in the mosquito infective ratios were compared using Fisher’s exact test. a–f The data from at least two independent experiments were combined
