Fig 1.
Illustration of WEEV constructs and growth kinetics in vitro.
A panel of recombinant WEEV constructs (A) were used to study the salivary gland escape barrier in Culex tarsalis. The viability of each recombinant virus was assessed using growth kinetics in C6/36 (B) cells. Arrows indicate the native and duplicated sub-genomic promoters. HPI: Hours post-infection.
Fig 2.
Epifluorescent imaging of Cx. tarsalis salivary glands and rates of salivary gland infection.
Salivary glands were compared to negative control epifluorescent (A) and light (D) images of non-infected Cx. tarsalis salivary glands at 20x magnification. Cx. tarsalis salivary glands 7 days post intrathoracic injection with 500 TCID50 McM-mCherry (B) and IMP181-mCherry (C). IFA of Cx. tarsalis salivary glands. Cx. tarsalis salivary glands stained immunohistochemically for WEEV 7 days post intrathoracic injection with 500 TCID50 McMillan (F) and IMP181 (H) along with light microscopy images of the same salivary glands (E and G). Salivary gland infection rates were calculated using the number of fluorescent mosquitoes compared to total injected mosquitoes at 7 (I) and 14 (J) DPI. DL, distal-lateral lobe; M, medial lobe; PL, proximal-lateral lobe. DPI: Days post-infection.
Fig 3.
Salivary gland infection rates and infectious virus concentration in saliva.
Salivary gland infection rates at 7 (A) and 14 (B) days were compared between IMP181-mCherry and McM-mCherry infected mosquitoes injected with 125, 250, or 500 TCID50. Infectious viral titers were also compared between IMP181 and McMillan derived viruses in saliva collected at 7 days post-injection with 500 TCID50 with IMP181 6K/E1 genes associated with an increase in expectorated virus (C). Transmission rates were also assessed for IMP181 and McMillan derived viruses (D).