This study aimed to investigate the transcriptomic changes in the midgut and salivary glands of Aedes aegypti mosquitoes infected with Zika virus (ZIKV), in order to explore the molecular mechanisms underlying the interaction between the virus and the mosquito vector. Aedes aegypti from Jiegao (JG) and Mengding (MD) in China were experimentally infected with ZIKV, and the midgut and salivary gland tissues were collected at 2-, 4- and 6 days post-infection (dpi). High-throughput sequencing was performed to analyze the transcriptomic changes between ZIKV-infected and non-infected Ae. aegypti midgut and salivary gland tissues. Bioinformatics tools were employed for further analysis of the transcriptomic data. The expression levels of 8 significantly differentially expressed genes (DEGs) were validated using RT-qPCR. A conjoint analysis of small RNA-seq and mRNA-seq was performed to screen interactional miRNA-mRNA pairs during ZIKV infection. Using the Search Tool for the Retrieval of Interacting Genes, we constructed a protein-protein interaction network of genes and subsequently identified hub genes. The most significant transcriptional changes in Ae. aegypti occurred at 2 dpi. On 2, 4 and 6 dpi, 11 genes showed significant changes in both the midgut and salivary glands of the same mosquito strain, while 25 genes exhibited significant changes in the same tissue between the JG and MD strains. The expression tendencies of 8 DEGs obtained by RNA-Seq were similar to those detected by RT-qPCR. Furthermore, we individually identified 10 hub genes in the midgut and salivary glands. Based on previous miRNA research, we discovered the involvement of 9 miRNAs in the regulation of these hub genes. Our findings demonstrate that Ae. aegypti exhibit distinct transcriptomic changes in response to ZIKV infection. The identification of the hub genes and their regulatory miRNAs provides valuable insights into the molecular mechanisms underlying ZIKV infection in mosquitoes. This study contributes to a better understanding of the pathogen-vector interactions and may aid in the development of targeted strategies for ZIKV control.

Zika virus (ZIKV) has emerged as a globally important arbovirus. The virus is primarily transmitted to humans through the bite of an infective Aedes albopictus in temperate area. Vertical transmission of ZIKV by Ae. albopictus is determined and has been suggested to be a means by which the virus could persist in nature. Ae. albopictus undergoes a well-characterized photoperiodic diapause. Viruses are harbored by overwintering mosquitoes in diapause that contributes to the resurgence of vertebrate diseases in the following spring, yet little is known about the impact of diapause on the regulation of viral replication and survival. The purpose of this study is to determine that Ae. albopictus in Beijing are highly susceptible to ZIKV (92.3%), and viable virus is passed to their organs of progeny via vertical transmission. Moreover, diapause eggs (diapause incidence 97.8%) had significantly lower minimum infection rates and filial infection rates of the first gonotrophic cycle than those of the second gonotrophic cycle in the short-day photoperiod group. Regarding the development of diapause eggs, the minimum infection rates and ZIKV RNA copy number increased significantly, suggesting that virus RNA replication occurred in the diapause eggs. Meanwhile, eggs from the ZIKV-infected mosquitoes had a significantly lower hatching rate compared with uninfected mosquitoes, implying an intriguing interaction between diapause eggs and virus. The findings here suggest that vertical transmission of ZIKV from diapause eggs to progeny may have a critical epidemiological role in the dissemination and maintenance of ZIKV circulating in the vector.

白纹伊蚊Aedes albopictus是登革病毒、寨卡病毒等多种病原体的重要传播媒介,广泛分布于我国温带和亚热带地区,以滞育卵越冬.一旦虫媒病毒扩散到我国北亚热带地区,非流行季节时期病毒的保存方式,将成为虫媒病毒研究中的重要问题.本研究开展了白纹伊蚊滞育卵的生态学研究,发现在短光照下不同地理株白纹伊蚊滞育卵的滞育率从产卵后7 d逐渐上升到21 d,至40 d趋于平缓并达到最高,滞育率在滞育前期和滞育后期统计学上差异显著.在滞育中后期,北京株白纹伊蚊滞育率相比杭州株白纹伊蚊和广州株白纹伊蚊滞育率明显较高.卵孵化后,不同地理株白纹伊蚊的卵的孵化率在滞育前期和滞育后期有显著差异.在短光照下滞育后期北京株白纹伊蚊的卵孵化率(21.45％)明显比广州株白纹伊蚊的卵孵化率(8.15％)高,提示位于温带地区的北京株白纹伊蚊较位于亚热带地区的杭州株和广州株白纹伊蚊滞育率和孵化率更高,说明位于温带地区的白纹伊蚊品系的滞育卵更能在冬季不良环境如低温等下存活,有利于增加越冬存活率.通过白纹伊蚊以滞育卵实现种群扩散、定殖的重要生物学基础,为探索新型蚊媒的控制途径提供科学依据.

埃及伊蚊和白纹伊蚊是登革热的主要传播媒介,蚊虫孳生情况的持续调查和分析,有助于对种群变化进行实时监控,从而为登革热的防治提供理论依据.2017-2020年,每年9-10月在云南省西双版纳州(勐腊县、景洪市、勐海县),临沧市(镇康县南伞镇、耿马县孟定镇),德宏州(瑞丽市)共6个市(县)调查埃及伊蚊和白纹伊蚊孳生地类型并捕捞幼虫,对4龄幼虫分类鉴定以观察两种伊蚊特征.结果 显示,此次在所调查的6个市(县)均发现两种伊蚊孳生地,共560个.孳生地类型以家用水桶类最多,占比44.82％.边境各地区居民区调查发现埃及伊蚊孳生地占比33.64％,商业区埃及伊蚊占比48.89％,两种调查地埃及伊蚊数量差异存在统计学意义,在商业区埃及伊蚊占比高于居民区.2017年西双版纳州和瑞丽市白纹伊蚊孳生地占比分别为75.51％和67.44％,2020年为36.67％和38.10％,说明两地白纹伊蚊构成比下降.两地区埃及伊蚊孳生地占比由2017年的18.37％和16.28％增长为2020年的43.33％和42.85％,说明埃及伊蚊构成比上升.结果 表明,由于埃及伊蚊比白纹伊蚊传播登革热的能力更强,2020年的西双版纳州和瑞丽市较2017年,商业区较居民区暴发登革热的潜在风险更高.

Abstract Background Zika virus (ZIKV) is transmitted to humans primarily by Aedes aegypti. Previous studies on Ae. aegypti from Jiegao (JG) and Mengding (MD) in Yunnan province, China have shown that these mosquitoes are able to transmit ZIKV to their offspring through vertical transmission, indicating that these two Ae. aegypti strains pose a potential risk for ZIKV transmission. However, the vector competence of these two Ae. aegypti strains to ZIKV has not been evaluated and the molecular mechanisms influencing vector competence are still unclear. Methods Aedes aegypti mosquitoes from JG and MD were orally infected with ZIKV, and the infection rate (IR), dissemination rate (DR), transmission rate (TR) and transmission efficiency (TE) of these two mosquito strains were explored to evaluate their vector competence to ZIKV. On 2, 4 and 6 days post-infection (dpi), the small RNA profiles between ZIKV-infected and non-infected Ae. aegypti midgut and salivary gland tissues were compared to gain insights into the molecular interactions between ZIKV and Ae. aegypti. Results There were no significant differences in the IR, DR, TR and TE between the two Ae. aegypti strains (P > 0.05). However, ZIKV RNA appeared 2 days earlier in saliva of the JG strain, which indicated a higher competence of the JG strain to transmit ZIKV. Significant differences in the microRNA (miRNA) expression profiles between ZIKV-infected and non-infected Ae. aegypti were found in the 2-dpi libraries of both the midgut and salivary gland tissues from the two strains. In addition, 27 and 74 miRNAs (|log2 fold change| > 2) were selected from the miRNA expression profiles of ZIKV-infected and non-infected midgut and salivary gland tissues from the JG and MD strains, respectively. Conclusions Our results provide novel insights into the ZIKV–mosquito interactions and build a foundation for future research on how miRNAs regulate the vector competence of mosquitoes to this arbovirus. Graphical abstract

Zika virus (ZIKV) is a mosquito-borne pathogen classified in the genusFlavivirusof the family Flaviviridae. Vertical transmission is considered to be the primary way to maintain some arboviruses under adverse natural conditions, which play a critical epidemiological role in arbovirus spread and maintenance.Aedes aegyptiis the primary vector for ZIKV. In this study, we demonstrated vertical transmission in twoAe. aegyptistrains from Jiegao (JG) and Mengding (MD) in the border area of Yunnan province. The minimum infection rate of F1 adult progeny from JGAe. aegyptistrain was significantly higher than that of MDAe. aegyptistrain in the second gonotrophic cycle (1:14.29 and 1:200, respectively,p < 0.05). The cytopathic effect was observed in C6/36 cells after infection of ZIKV isolated from the progeny. The results suggest thatAe. aegyptimosquitoes from JG and MD play potential roles in ZIKV spread and maintenance. Therefore, more adult and eggs control methods should be implemented to control mosquitoes if a Zika epidemic occurs.