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The composition and structure of microbial communities that inhabit the mosquito midguts are poorly understood despite their well-documented potential to impede pathogen transmission. We used MiSeq sequencing of the 16S rRNA gene to characterize the bacterial communities of field-collected populations of 12 mosquito species. After quality filtering and rarefaction, the remaining sequences were assigned to 181 operational taxonomic units (OTUs). Approximately 58% of these OTUs occurred in at least two mosquito species but only three OTUs: Gluconobacter (OTU 1), Propionibacterium (OTU 9), and Staphylococcus (OTU 31) occurred in all 12 mosquito species. Individuals of different mosquito species shared similar gut microbiota and it was common for individuals of the same species from the same study site and collection date to harbor different gut microbiota. On average, the microbiota of Aedes albopictus was the least diverse and significantly less even compared to Anopheles crucians, An. quadrimaculatus, Ae. triseriatus, Ae. vexans, Ae. japonicus, Culex restuans, and Culiseta inornata. The microbial community of Cx. pipiens and Ae. albopictus differed significantly from all other mosquitoes species and was primarily driven by the dominance of Wolbachia. These findings expand the range of mosquito species whose gut microbiota has been characterized and sets the foundation for further studies to determine the influence of these microbiota on vector susceptibility to pathogens.

Aedes aegypti, the mosquito vector of dengue viruses, utilizes its innate immune system to ward off a variety of pathogens, some of which can cause disease in humans. To date, the features of insects' innate immune defenses against viruses have mainly been studied in the fruit fly Drosophila melanogaster, which appears to utilize different immune pathways against different types of viruses, in addition to an RNA interference-based defense system. We have used the recently released whole-genome sequence of the Ae. aegypti mosquito, in combination with high-throughput gene expression and RNA interference (RNAi)-based reverse genetic analyses, to characterize its response to dengue virus infection in different body compartments. We have further addressed the impact of the mosquito's endogenous microbial flora on virus infection. Our findings indicate a significant role for the Toll pathway in regulating resistance to dengue virus, as indicated by an infection-responsive regulation and functional assessment of several Toll pathway-associated genes. We have also shown that the mosquito's natural microbiota play a role in modulating the dengue virus infection, possibly through basal-level stimulation of the Toll immune pathway.

Abstract Gut microbial communities of mosquitoes can influence vector susceptibility to pathogens, yet the factors that govern their composition remain poorly understood. We investigated the impact of host blood-meal source on gut microbiota of Aedes aegypti L. Adult mosquitoes were fed on human, rabbit or chicken blood and their gut microbiota compared to those of sugar-fed and newly emerged adults. Microbial diversity was significantly reduced in blood-fed and sugar-fed mosquitoes but was restored to the levels of newly emerged adults at 7-days post-blood meal. Microbial composition was strongly influenced by host blood-meal source. Leucobacter spp., Chryseobacterium spp., Elizabethkingia spp. and Serratia spp. were characteristic of newly emerged adults and adults fed on chicken, rabbit and human blood, respectively. Sugar-fed mosquitoes had higher abundance of Pseudomonas spp. and unclassified Acetobacteraceae. Shifts in gut microbial communities in response to host blood-meal source may fundamentally impact pathogen transmission given the well-documented link between specific bacterial taxa and vector susceptibility to a variety of mosquito-borne pathogens and may be a key determinant of individual and population variation in vector competence. Gut bacterial communities of Aedes aegypti shift in response to host blood-meal source.

Development of alternative vector control strategies are becoming more pressing given the rapid evolution of insecticide resistance and the rise of vector borne pathogens affecting public health such as dengue, chikungunya and Zika. Fungal-based biopesticides are promising alternatives to synthetic insecticides because they are ecofriendly and are highly effective at infecting insects through contact. This study evaluated the susceptibility of the yellow fever mosquito Ae . aegypti to a range of entomopathogenic fungal strains from the genus Isaria . We observed a diverse variation in the virulence of the Isaria strains tested, with two strains showing high pathogenicity towards adult mosquitoes. Mosquito susceptibility to fungal infection was further corroborated through the molecular quantification of fungal loads and the transcript evaluation of a fungal-specific pathogen recognition molecule in the mosquito body. Moreover, quantitative analysis of transcript abundance coupled with enzymatic assays revealed strain-specific subversion of the melanization cascade, an important immune response component. Our study contributes critical insights for a better understanding of fungal-mosquito interactions.

Synthetic pesticides are the cornerstone of vector-borne disease control, but alternatives are urgently needed to tackle the growing problem of insecticide resistance and concerns over environmental safety. Leptospermum scoparium J.R. Forst and G. Forst (manuka) essential oil and its four fractions were analyzed for chemical composition and toxicity against Aedes aegypti larvae. The use of bio-based amylose-N-1-hexadecylammonium chloride inclusion complexes (Hex-Am) as an emulsifier for L. scoparium essential oil was also investigated. Fraction 1 was inactive, fractions 2 (LC50 = 12.24 ppm) and 3 (LC50 = 20.58 ppm) were more toxic than the whole essential oil (LC50 = 47.97 ppm), and fraction 4 (LC50 = 35.87 ppm) had similar toxicity as the whole essential oil. Twenty-one chemical constituents were detected in L. scoparium essential oil compared to 16, 5, 19 and 25 chemical constituents in fractions, 1, 2, 3 and 4 respectively. The two most dominant chemical constituents were calamenene (17.78%) and leptospermone (11.86%) for L. scoparium essential oil, calamenene (37.73%) and ledene (10.37%) for fraction 1, leptospermone (56.6%) and isoleptospermone (19.73) for fraction 2, cubenol (24.30%) and caryophyllene oxide (12.38%) for fraction 3, and γ-gurjunene (21.62%) and isoleptospermone (7.88%) for fraction 4. Alpha-pinene, ledene, and aromandendrene were 2-7 times less toxic than the whole essential suggesting that the toxicity of L. scoparium essential oil was either due to other chemical constituents that were not tested or due synergist interactions among chemical constituents. Leptospermum scoparium essential oil-Hex-Am emulsion (LC50 = 29.62) was more toxic than the whole essential oil. These findings suggest that L. scoparium essential oil is a promising source of mosquito larvicide and that Hex-Am is an excellent emulsifier for L. scoparium essential oil for use as a larvicide.

Animal agriculture is under pressure to increase efficiency, sustainability, and innovation to meet the demands of a rising global population while decreasing adverse environmental effects. Feed cost and availability are 2 of the biggest hurdles to sustainable production. Current diets depend on sources of grain and animal byproduct protein for essential amino acids which have limited sustainability. Insects have arisen as an attractive, sustainable alternative protein source for animal diets due to their favorable nutrient composition, low space and water requirements, and natural role in animal diets. Additionally, insects are capable of bioremediating waste streams including agricultural and food waste, manure, and plastics helping to increase their sustainability. The insect rearing industry has grown rapidly in recent years and shows great economic potential. However, state-of-the-art research is urgently needed to overcome barriers to adoption in commercial animal diets such as regulatory restrictions, production scale issues, and food safety concerns. To address this need, the USDA Agricultural Research Service “MINIstoc: Model for INsect Inclusion” project was created to bring together diverse scientists from across the world to synergistically advance insect meal production and inclusion in animal diets. Here, we provide a short review of insects as feed while describing the MINIstock project which serves as the inspiration for the Journal of Economic Entomology Special Collection “Insects as feed: sustainable solutions for food waste and animal production practices.”

Microbial control of mosquitoes via the use of symbiotic or pathogenic microbes, such as Wolbachia and entomopathogenic fungi, are promising alternatives to synthetic insecticides to tackle the rapid increase in insecticide resistance and vector-borne disease outbreaks. This study evaluated the susceptibility and host responses of two important mosquito vectors, Ae . albopictus and Cx . pipiens , that naturally carry Wolbachia , to infections by entomopathogenic fungi. Our study indicated that while Wolbachia presence did not provide a protective advantage against entomopathogenic fungal infection, it nevertheless influenced the bacterial / fungal load and the expression of select anti-microbial effectors and phenoloxidase cascade genes in mosquitoes. Furthermore, although host responses from Ae . albopictus and Cx . pipiens were mostly similar, we observed contrasting phenotypes with regards to susceptibility and immune responses to fungal entomopathogenic infection in these two mosquitoes. This study provides new insights into the intricate multipartite interaction between the mosquito host, its native symbiont and pathogenic microbes that might be employed to control mosquito populations.

Dengue fever is the most frequent arthropod-borne viral disease of humans, with almost half of the world's population at risk of infection. The high prevalence, lack of an effective vaccine, and absence of specific treatment conspire to make dengue fever a global public health threat. Given their compact genomes, dengue viruses (DENV-1-4) and other flaviviruses probably require an extensive number of host factors; however, only a limited number of human, and an even smaller number of insect host factors, have been identified. Here we identify insect host factors required for DENV-2 propagation, by carrying out a genome-wide RNA interference screen in Drosophila melanogaster cells using a well-established 22,632 double-stranded RNA library. This screen identified 116 candidate dengue virus host factors (DVHFs). Although some were previously associated with flaviviruses (for example, V-ATPases and α-glucosidases), most of the DVHFs were newly implicated in dengue virus propagation. The dipteran DVHFs had 82 readily recognizable human homologues and, using a targeted short-interfering-RNA screen, we showed that 42 of these are human DVHFs. This indicates notable conservation of required factors between dipteran and human hosts. This work suggests new approaches to control infection in the insect vector and the mammalian host.

Malaria is endemic in Darién and an assessment of the different factors affecting its epidemiology is crucial for the development of adequate strategies of surveillance, prevention, and disease control. The objective of this study was to determine the main characteristics of the epidemiological behavior of malaria in the Darien region. This research was comprised of a retrospective analysis to determine the incidence and malaria distribution in the Darien region from 2015 to 2017. We evaluated malaria indicators, disease distribution, incidence (by age group and sex), diagnostic methods, treatment, and control measures. In addition, we examined the cross-border migration activity and its possible contribution to the maintenance and distribution of malaria. During the period of 2015-2017, we examined 41,141 thick blood smear samples, out of which 501 tested positive for malaria. Plasmodium vivax was responsible for 92.2% of those infections. Males comprised 62.7% of the total diagnosed cases. Meanwhile, a similar percentage, 62.7%, of the total cases were registered in economically active ages. The more frequent symptoms included fever (99.4%) and chills (97.4%), with 53.1% of cases registering between 2,000 and 6,000 parasites/μl of blood. The annual parasitic incidence (API) average was 3.0/1,000 inhabitants, while the slide positivity rate (SPR) was 1.2% and the annual blood examination rate (ABER) 22.5%. In Darién there is a constant internal and cross-border migration movement between Panama and Colombia. Malaria control measures consisted of the active and passive search of suspected cases and of the application of vector control measures. This study provides an additional perspective on malaria epidemiology in Darién. Additional efforts are required to intensify malaria surveillance and to achieve an effective control, eventually moving closer to the objective of malaria elimination. At the same time, there is a need for more eco-epidemiological, entomological and migratory studies to determine how these factors contribute to the patterns of maintenance and dissemination of malaria.

Background Microbial communities that inhabit the mosquito body play an import role in host biology and may have potential for mosquito control. However, the forces that shape these microbial communities are poorly understood. Methods To gain a better understanding of how host location influences the composition and diversity of mosquito microbiota, we performed a survey of microbial communities in mosquito samples collected from six USA states using HiSeq sequencing of the 16S rRNA gene. Results A total of 284 bacterial operational taxonomic units (OTUs) belonging to 14 phyla were detected in nine mosquito species, with Proteobacteria, Firmicutes and Actinobacteria accounting for 95% of total sequences. OTU richness varied markedly within and between mosquito species. The microbial composition and diversity was heavily influenced by the site of mosquito collection, suggesting that host location plays an important role in shaping the mosquito microbiota. Conclusions Variation in microbial composition and diversity between mosquitoes from different locations may have important implications on vector competence and transmission dynamics of mosquito-borne pathogens. Future studies should investigate the environmental factors responsible for these variations and the role of key bacteria characterized in this study on mosquito biology and their potential application in symbiotic control of mosquito-borne diseases.