Tomato spotted wilt virus(TSWV) is transmitted byFrankliniella occidentalisin a persistent propagative manner. Despite the extensive replication of TSWV in midgut and salivary glands, there is little to no pathogenic effect onF. occidentalis. We hypothesize that the first-instar larva (L1) ofF. occidentalismounts a response to TSWV that protects it from pathogenic effects caused by virus infection and replication in various insect tissues. A partial thrips transcriptome was generated using 454-Titanium sequencing of cDNA generated fromF. occidentalisexposed to TSWV. Using these sequences, the L1 thrips proteome that resolved on a two-dimensional gel was characterized. Forty-seven percent of the resolved protein spots were identified using the thrips transcriptome. Real-time quantitative reverse transcriptase PCR (RT-PCR) analysis of virus titer in L1 thrips revealed a significant increase in the normalized abundance of TSWV nucleocapsid RNA from 2 to 21 h after a 3-h acquisition access period on virus-infected plant tissue, indicative of infection and accumulation of virus. We compared the proteomes of infected and noninfected L1s to identify proteins that display differential abundances in response to virus. Using four biological replicates, 26 spots containing 37 proteins were significantly altered in response to TSWV. Gene ontology assignments for 32 of these proteins revealed biological roles associated with the infection cycle of other plant- and animal-infecting viruses and antiviral defense responses. Our findings support the hypothesis that L1 thrips display a complex reaction to TSWV infection and provide new insights toward unraveling the molecular basis of this interaction.

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The plant-pathogenic virus tomato spotted wilt virus (TSWV) encodes a structural glycoprotein (GN) that, like with other bunyavirus/vector interactions, serves a role in viral attachment and possibly in entry into arthropod vector host cells. It is well documented that Frankliniella occidentalis is one of nine competent thrips vectors of TSWV transmission to plant hosts. However, the insect molecules that interact with viral proteins, such as GN, during infection and dissemination in thrips vector tissues are unknown. The goals of this project were to identify TSWV-interacting proteins (TIPs) that interact directly with TSWV GN and to localize the expression of these proteins in relation to virus in thrips tissues of principal importance along the route of dissemination. We report here the identification of six TIPs from first-instar larvae (L1), the most acquisition-efficient developmental stage of the thrips vector. Sequence analyses of these TIPs revealed homology to proteins associated with the infection cycle of other vector-borne viruses. Immunolocalization of the TIPs in L1 revealed robust expression in the midgut and salivary glands of F. occidentalis, the tissues most important during virus infection, replication, and plant inoculation. The TIPs and GN interactions were validated using protein-protein interaction assays. Two of the thrips proteins, endocuticle structural glycoprotein and cyclophilin, were found to be consistent interactors with GN. These newly discovered thrips protein-GN interactions are important for a better understanding of the transmission mechanism of persistent propagative plant viruses by their vectors, as well as for developing new strategies of insect pest management and virus resistance in plants. IMPORTANCE Thrips-transmitted viruses cause devastating losses to numerous food crops worldwide. For negative-sense RNA viruses that infect plants, the arthropod serves as a host as well by supporting virus replication in specific tissues and organs of the vector. The goal of this work was to identify thrips proteins that bind directly to the viral attachment protein and thus may play a role in the infection cycle in the insect. Using the model plant bunyavirus tomato spotted wilt virus (TSWV), and the most efficient thrips vector, we identified and validated six TSWV-interacting proteins from Frankliniella occidentalis first-instar larvae. Two proteins, an endocuticle structural glycoprotein and cyclophilin, were able to interact directly with the TSWV attachment protein, GN, in insect cells. The TSWV GN-interacting proteins provide new targets for disrupting the viral disease cycle in the arthropod vector and could be putative determinants of vector competence.

Mosquito-borne viruses are emerging or re-emerging globally, afflicting millions of people around the world. Aedes aegypti, the yellow fever mosquito, is the principal vector of dengue, Zika, and chikungunya viruses, and has well-established populations across tropical and subtropical urban areas of the Americas, including the southern United States. While intense arboviral epidemics have occurred in Mexico and further south in the Americas, local transmission in the United States has been minimal. Here, we study Ae. aegypti and Culex quinquefasciatus host feeding patterns and vertebrate host communities in residential environments of South Texas to identify host-utilization relative to availability. Only 31% of Ae. aegypti blood meals were derived from humans, while 50% were from dogs and 19% from other wild and domestic animals. In Cx. quinquefasciatus, 67% of blood meals were derived from chicken, 22% came from dogs, 9% from various wild avian species, and 2% from other mammals including one human, one cat, and one pig. We developed a model for the reproductive number, R0, for Zika virus (ZIKV) in South Texas relative to northern Mexico using human disease data from Tamaulipas, Mexico. We show that ZIKV R0 in South Texas communities could be greater than one if the risk of human exposure to Ae. aegypti bites in these communities is at least 60% that of Northern Mexico communities. The high utilization of non-human vertebrates and low risk of human exposure in South Texas diminishes the outbreak potential for human-amplified urban arboviruses transmitted by Ae. aegypti.

Aedes aegypti is the main vector of arboviral diseases such as dengue, chikungunya and Zika. A key feature for disease transmission modeling and vector control planning is adult mosquito dispersal. We studied Ae aegypti adult dispersal by conducting a mark-capture study of naturally occurring Ae. aegypti from discarded containers found along a canal that divided two residential communities in Donna, texas, USA. Stable isotopes were used to enrich containers with either 13 c or 15 n. Adult mosquitoes were collected outdoors in the yards of households throughout the communities with BG Sentinel 2 traps during a 12-week period. Marked mosquito pools with stable isotopes were used to estimate the mean distance travelled using three different approaches (Net, Strip or Circular) and the probability of detecting an isotopically marked adult at different distances from the larval habitat of origin. We consistently observed, using the three approaches that male (Net: 220 m, Strip: 255 m, Circular: 250 m) Ae. aegypti dispersed further in comparison to gravid (Net: 135 m, Strip: 176 m, Circular: 189 m) and unfed females (Net: 192 m, Strip: 213 m, Circular: 198 m). We also observed that marked male capture probability slightly increased with distance, while, for both unfed and gravid females, such probability decreased with distance. Using a unique study design documenting adult dispersal from natural larval habitat, our results suggest that Ae. aegypti adults disperse longer distances than previously reported. These results may help guide local vector control authorities in their fight against Ae. aegypti and the diseases it transmits, suggesting coverage of 200 m for the use of insecticides and innovative vector control tools.

A major bottleneck in identifying therapies to control citrus greening and other devastating plant diseases caused by fastidious pathogens is our inability to culture the pathogens in defined media or axenic cultures. As such, conventional approaches for antimicrobial evaluation (genetic or chemical) rely on time-consuming, low-throughput and inherently variable whole-plant assays. Here, we report that plant hairy roots support the growth of fastidious pathogens like Candidatus Liberibacter spp., the presumptive causal agents of citrus greening, potato zebra chip and tomato vein greening diseases. Importantly, we leverage the microbial hairy roots for rapid, reproducible efficacy screening of multiple therapies. We identify six antimicrobial peptides, two plant immune regulators and eight chemicals which inhibit Candidatus Liberibacter spp. in plant tissues. The antimicrobials, either singly or in combination, can be used as near- and long-term therapies to control citrus greening, potato zebra chip and tomato vein greening diseases.

Abstract Effective mosquito surveillance and management depend on a thorough understanding of the biology and feeding patterns unique to species and sex. Given that a propensity to sugar feed is necessary for some mosquito surveillance and newer control strategies, we sought to document the amount of total sugar in wild Aedes aegypti (L.) and Culex quinquefasciatus (Say) captured from five different locations in the Lower Rio Grande Valley (LRGV) of South Texas over 2 yr. We used Biogents Sentinel 2 (BGS2) traps in year 1 and aspirators, BGS2, and CDC resting traps in years 2 and 3 to collect adult mosquitoes. The hot anthrone test was used to quantify total sugar content in each mosquito. Additionally, the cold and hot anthrone tests were used to distinguish fructose content from total sugars for mosquitoes captured in 2019. Overall, Ae. aegypti females had significantly lower total sugar content than Ae. aegypti males as well as both sexes of Cx. quinquefasciatus. However, the percentage of Ae. aegypti positive for fructose consumption was four to eightfold higher than Ae. aegypti previously reported in other regions. The difference between locations was significant for males of both species, but not for females. Seasonality and trapping method also revealed significant differences in sugar content of captured mosquitoes. Our results reinforce that sugar feeding in female Ae. aegypti is less than Cx. quinquefasciatus, although not absent. This study provides necessary data to evaluate the potential effectiveness of sugar baits in surveillance and control of both Ae. aegypti and Cx. quinquefasciatus mosquitoes.