A female mosquito salivary protein-driven influx of myeloid cells facilitates flavivirus transmission

Mosquitoes transmit many flaviviruses of global public health significance. Efficient viral transmission to mammalian hosts requires mosquito salivary factors that modulate local host responses, such as recruitment of virus-permissive myeloid cells to the bite sites. However, the specific salivary components facilitating viral transmission and their mechanisms of action remain largely unknown. Here, we showed that a female mosquito salivary gland-specific protein, named Aedes aegypti Neutrophil Recruitment Protein (AaNRP), acts as a key salivary component to facilitate the transmission of Zika (ZIKV) and dengue (DENV) viruses. AaNRP promotes a rapid influx of neutrophils followed by virus-susceptible myeloid cells toward mosquito bite sites, which facilitate establishment of local infection and systemic dissemination. Mechanistically, AaNRP engages TLR1 and TLR4 of skin resident macrophages and activates MyD88-dependent NF-κB signaling to induce the expression of neutrophil chemoattractants. Inhibition of MyD88-NF-κB with dietary resveratrol, a phytochemical, neutralizes the AaNRP effects, thus reducing flavivirus transmission by mosquitoes. This study offers mechanistic insight into saliva-aided viral transmission and provides a potential prophylactic target. We investigated the underlying mechanism of the mosquito bite-mediated promotion of neutrophil infiltration by a single-cell RNA sequencing (scRNA-Seq). The skin of mouse footpads at 4 hpb or without mosquito bites was isolated for enzymatic digestion. The total skin CD45+ immunocytes were flow-sorted and subjected to 10x genomics scRNA-Seq.

蚊虫可传播多种具有全球公共卫生重要性的黄病毒（flaviviruses）。病毒若要高效传播至哺乳动物宿主，需依赖蚊虫唾液因子调节宿主局部免疫应答，例如招募允许病毒增殖的髓系细胞至叮咬位点。然而，介导病毒传播的特定唾液成分及其作用机制仍未被充分阐明。
本研究发现，一种雌性埃及伊蚊唾液腺特异性蛋白——埃及伊蚊嗜中性粒细胞招募蛋白（Aedes aegypti Neutrophil Recruitment Protein，AaNRP），可作为关键唾液成分促进寨卡病毒（Zika virus，ZIKV）与登革病毒（Dengue virus，DENV）的传播。AaNRP可促使嗜中性粒细胞及病毒易感髓系细胞快速向蚊虫叮咬位点聚集，从而助力局部感染的建立与全身播散。
从机制上而言，AaNRP可结合皮肤驻留巨噬细胞的Toll样受体1（TLR1）与Toll样受体4（TLR4），激活依赖髓系分化因子88（Myeloid differentiation primary response 88，MyD88）的核因子κB（NF-κB）信号通路，进而诱导嗜中性粒细胞趋化因子的表达。通过膳食摄入的植物化学物质白藜芦醇（resveratrol）抑制MyD88-NF-κB通路，可中和AaNRP的上述作用，从而降低蚊虫传播黄病毒的能力。
本研究为唾液辅助的病毒传播机制提供了机制性见解，并提出了潜在的预防靶点。我们通过单细胞RNA测序（single-cell RNA sequencing，scRNA-Seq）探究了蚊虫叮咬介导嗜中性粒细胞浸润的潜在机制：分离叮咬后4小时（4 hpb）或未被蚊虫叮咬的小鼠足垫皮肤，进行酶消化处理；对总皮肤CD45+免疫细胞进行流式分选后，采用10x Genomics平台开展单细胞RNA测序。