Mosquitoes host diverse microbial communities that influence many aspects of their biology including reproduction, digestion, and ability to transmit pathogens. Unraveling the composition, structure, and function of these microbiota can provide new opportunities for exploiting microbial function for mosquito-borne disease control. MiSeq® sequencing of 16S rRNA gene amplicons was used to characterize the microbiota of adult females of Culex pipiens L. and Cx. restuans Theobald collected from nine study sites in central Illinois. Out of 195 bacterial OTUs that were identified, 86 were shared between the two mosquito species while 16 and 93 OTUs were unique to Cx. pipiens and Cx. restuans, respectively. The composition and structure of microbial communities differed significantly between the two mosquito species with Cx. restuans hosting a more diverse bacterial community compared to Cx. pipiens. Wolbachia (OTU836919) was the dominant bacterial species in Cx. pipiens accounting for 91 % of total microbiota while Sphingomonas (OTU817982) was the dominant bacterial species in Cx. restuans accounting for 31 % of total microbiota. Only 3 and 6 OTUs occurred in over 60 % of individuals in Cx. pipiens and Cx. restuans, respectively. There was little effect of study site on bacterial community structure of either mosquito species. These results suggest that the two mosquito species support distinct microbial communities that are sparsely distributed between individuals. These findings will allow investigations of the role of identified microbiota on the spatial and temporal heterogeneity in WNV transmission and their potential application in disease control.
Alternative methods of mosquito control are needed to tackle the rising burden of mosquito-borne diseases while minimizing the use of synthetic insecticides, which are threatened by the rapid increase in insecticide resistance in mosquito populations. Fungal biopesticides show great promise as potential alternatives because of their ecofriendly nature and ability to infect mosquitoes on contact. Here we describe the temporospatial interactions between the mosquito Aedes aegypti and several entomopathogenic fungi. Fungal infection assays followed by the molecular assessment of infection-responsive genes revealed an intricate interaction between the mosquito immune system and entomopathogenic fungi. We observed contrasting tissue and time-specific differences in the activation of immune signaling pathways and antimicrobial peptide expression. In addition, these antifungal responses appear to vary according to the fungal entomopathogen used in the infection. Enzyme activity-based assays coupled with gene expression analysis of prophenoloxidase genes revealed a reduction in phenoloxidase (PO) activity in mosquitoes infected with the most virulent fungal strains at 3 and 6d post-fungal infection. Moreover, fungal infection led to an increase in midgut microbiota that appear to be attributed in part to reduced midgut reactive oxygen species (ROS) activity. This indicates that the fungal infection has far reaching effects on other microbes naturally associated with mosquitoes. This study also revealed that despite fungal recognition and immune elicitation by the mosquito, it is unable to successfully eliminate the entomopathogenic fungal infection. Our study provides new insights into this intricate multipartite interaction and contributes to a better understanding of mosquito antifungal immunity.
The succession of mosquito species and abiotic factors affecting their distribution and abundance in rice (Oryza spp.) fields was investigated over a 16-wk rice growing cycle covering the period between January and May 2006. Fifteen experimental rice plots were sampled for mosquito larvae and characterized based on rice height, number of tillers, floating vegetation cover, water depth, water temperature, turbidity, salinity, pH, dissolved oxygen, total dissolved solids, and conductivity. Microscopic identification of 3,025 larvae yielded nine mosquito species predominated by Anopheles arabiensis Patton (45.0%), Culex quinquefasciatus Say (35.8%), Anopheles pharoensis Theobald (9.0%) and Ficalbia splendens Theobald (7.1%). Other species, including Anopheles rufipes Gough, Anopheles coustani Laveran, Anonopheles maculipalpis Giles, Culex annulioris Theobald, and Culex poicilipes Theobald made up 3.1% of the total collection. Anopheles gambiae s.l., Cx. quinquefasciatus, and An. pharoensis occurred throughout the cycle, but they were more abundant up to 4 wk posttransplanting with peaks after fertilizer application. As rice plants became established, three groups of mosquitoes were recognized: the first groups included An. rufipes, Fl. splendens, and Cx. annulioris, which occurred throughout much of the second half of the rice cycle, whereas the second group included Cx. poicilipes, which was found in the middle of the rice cycle. An. coustani and An. maculipalpis formed the third group occurring toward the end of the cycle. Dissolved oxygen, number of tillers, and rice height were negatively associated with the abundance of An. arabiensis and Cx. quinquefasciatus larvae. In addition, Cx. quinquefasciatus also was associated with water depth (−ve) and turbidity (+ve). Abundance of An. pharoensis larvae was significantly associated with water temperature (+ve), the number of tillers (−ve), and rice height (−ve), whereas Fl. splendens was significantly associated with the number of tillers (+ve). The results demonstrate a complex nature of the interactions between some of the factors in the ecosystem and mosquito species abundance and calls for time-dependent and species-specific mosquito control operations.
Although many mosquito species develop within agricultural landscapes where they are potentially exposed to agricultural chemicals (fertilizers and pesticides), the effects of these chemicals on mosquito biology remain poorly understood. This study investigated the effects of sublethal concentrations of four agricultural chemicals on the life history traits of Anopheles arabiensis and Culex quinquefasciatus mosquitoes. Field and laboratory experiments were conducted to examine how sublethal concentrations of four agricultural chemicals: an insecticide (cypermethrin), a herbicide (glyphosate), and two nitrogenous fertilizers (ammonium sulfate and diammonium phosphate) alter oviposition site selection, emergence rates, development time, adult body size, and longevity of An. arabiensis and Cx. quinquefasciatus. Both mosquito species had preference to oviposit in fertilizer treatments relative to pesticide treatments. Emergence rates for An. arabiensis were significantly higher in the control and ammonium sulfate treatments compared to cypermethrin treatment, while emergence rates for Cx. quinquefasciatus were significantly higher in the diammonium phosphate treatment compared to glyphosate and cypermethrin treatments. For both mosquito species, individuals from the ammonium sulfate and diammonium phosphate treatments took significantly longer time to develop compared to those from cypermethrin and glyphosate treatments. Although not always significant, males and females of both mosquito species tended to be smaller in the ammonium sulfate and diammonium phosphate treatments compared to cypermethrin and glyphosate treatments. There was no significant effect of the agrochemical treatments on the longevity of either mosquito species. These results demonstrate that the widespread use of agricultural chemicals to enhance crop production can have unexpected effects on the spatial distribution and abundance of mosquito vectors of malaria and lymphatic filariasis.
We examined algorithms for malaria mapping using the impact of reflectance calibration uncertainties on the accuracies of three vegetation indices (VI)'s derived from QuickBird data in three rice agro-village complexes Mwea, Kenya. We also generated inferential statistics from field sampled vegetation covariates for identifying riceland Anopheles arabiensis during the crop season. All aquatic habitats in the study sites were stratified based on levels of rice stages; flooded, land preparation, post-transplanting, tillering, flowering/maturation and post-harvest/fallow. A set of uncertainty propagation equations were designed to model the propagation of calibration uncertainties using the red channel (band 3: 0.63 to 0.69 μm) and the near infra-red (NIR) channel (band 4: 0.76 to 0.90 μm) to generate the Normalized Difference Vegetation Index (NDVI) and the Soil Adjusted Vegetation Index (SAVI). The Atmospheric Resistant Vegetation Index (ARVI) was also evaluated incorporating the QuickBird blue band (Band 1: 0.45 to 0.52 μm) to normalize atmospheric effects. In order to determine local clustering of riceland habitats Gi*(d) statistics were generated from the ground-based and remotely-sensed ecological databases. Additionally, all riceland habitats were visually examined using the spectral reflectance of vegetation land cover for identification of highly productive riceland Anopheles oviposition sites. The resultant VI uncertainties did not vary from surface reflectance or atmospheric conditions. Logistic regression analyses of all field sampled covariates revealed emergent vegetation was negatively associated with mosquito larvae at the three study sites. In addition, floating vegetation (-ve) was significantly associated with immature mosquitoes in Rurumi and Kiuria (-ve); while, turbidity was also important in Kiuria. All spatial models exhibit positive autocorrelation; similar numbers of log-counts tend to cluster in geographic space. The spectral reflectance from riceland habitats, examined using the remote and field stratification, revealed post-transplanting and tillering rice stages were most frequently associated with high larval abundance and distribution. NDVI, SAVI and ARVI generated from QuickBird data and field sampled vegetation covariates modeled cannot identify highly productive riceland An. arabiensis aquatic habitats. However, combining spectral reflectance of riceland habitats from QuickBird and field sampled data can develop and implement an Integrated Vector Management (IVM) program based on larval productivity.
We determined changes in species composition and densities of immature stages of Anopheles arabiensis mosquitoes in relation to rice growth cycle in order to generate data for developing larval control strategies in rice ecosystems. Experimental rice paddies (6.3m × 3.15m) exposed to natural colonization of mosquitoes were sampled weekly for two rice growing cycles between February 2004 and March 2005. Overall, 21,325 Anopheles larvae were collected, of which 91.9% were 1st and 2nd instars and 8.1% were 3rd and 4th instars. An. arabiensis was the predominant species (84.1%) with other species, An. pharoensis (13.5%), An. funestus (2.1%), An. coustani (0.3%), and An. maculipalpis (0.1%) accounting for only a small proportion of the anophelines collected. Culex quinquefasciatus (65.7%) was the predominant species among the non-anopheline species. Others species collected included: C. annulioris (9.9%), C. poicilipes (7.3%), C. tigripes (7.2%), C. duttoni (0.6%), Aedes aegypti (5.3%), Ae. cumminsii (3.5%), and Ae. vittatus (0.7%). The densities of the major anopheline species were closely related to rice stage and condition of the rice field. An. arabiensis, the predominant species, was most abundant over a three-week period after transplanting. Low densities of larvae were collected during the late vegetative, reproductive, and ripening phases of rice. An increase in larval density ten days post-transplanting was found to correlate with the application of fertilizer (sulphate of ammonia). Culicine and aedine species densities were significantly higher during the post-harvesting period. Our results suggest that the transplanting stage is favorable for the growth of immature stages of An. arabiensis and provides a narrow window for targeted larval intervention in rice.
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