Novel, anionic, antiviral septapeptides from mosquito cells also protect monkey cells against dengue virus

•Persistent dengue virus infections in mosquito cells produce antiviral septapeptides.•The peptides are strongly anionic.•The peptides protect both mosquito and Vero cells against dengue virus infection.•The peptide consensus sequence is D-D/E-X-X-X-Q-D.

We have shown previously that ultrafiltrates (5 kDa cutoff) of cell-free medium from mosquito cell cultures persistently infected with DENV serotype 2 (DENV-2) contained a novel antiviral agent (called viprolaxikine) that could protect pre-treated, naïve mosquito cells from DENV infection. Here, we show that viprolaxikine also reduced DENV-2 titers by almost 4 logs (>99.9%) when compared to Vero cells mock-treated with ultrafiltrates from cultures of uninfected mosquito cells. Protease treatment removed the anti-DENV-2 activity. Pre-incubation for 48-h was required to obtain the maximum, dose-dependent protection against DENV-2, indicating that the antiviral activity was based on the interaction between Vero cells and viprolaxikine rather than direct action of viprolaxikine on DENV-2. Activity was highest against DENV-2, but there was also significant activity against the 3 other DENV serotypes. LC-MS-MS analysis revealed that the active viprolaxikine fraction contained anionic, antiviral peptides, each comprised of 7 amino acids (DDHELQD, DETELQD and DEVMLQD or DEVLMQD) and with a common sequence motif of D-D/E-X-X-X-Q-D. These sequences do not occur in the dengue virus genome, suggesting that the peptides are produced by the host insect cells when persistently infected with DENV-2. These peptides represent a new class of anionic, insect-derived, antiviral peptides with activity against a flavivirus in both mammalian and insect cells.