The native microflora associated with mosquitoes have important roles in mosquito development and vector competence. Sequencing of bacterial V3 region from 16S rRNA genes across the developmental stages of Culex mosquitoes (early and late larval instars, pupae and adults) was used to test the hypothesis that bacteria found in the larval stage of Culex are transstadially transmitted to the adult stage, and to compare the microbiomes of field-collected versus laboratory-reared mosquitoes. Beta diversity analysis revealed that bacterial community structure differed among three life stages (larvae, pupae and adults) of Culex tarsalis. Although only ~2 % of the total number of bacterial OTUs were found in all stages, sequences from these OTUs accounted for nearly 82 % of the total bacterial sequences recovered from all stages. Thorsellia (Gammaproteobacteria) was the most abundant bacterial taxon found across all developmental stages of field-collected Culex mosquitoes, but was rare in mosquitoes from laboratory-reared colonies. The proportion of Thorsellia sequences in the microbiomes of mosquito life stages varied ontogenetically with the greatest proportions recovered from the pupae of C. tarsalis and the lowest from newly emerged adults. The microbiome of field-collected late instar larvae was not influenced significantly by differences in the microbiota of the habitat due to habitat age or biopesticide treatments. The microbiome diversity was the greatest in the early instar larvae and the lowest in laboratory-reared mosquitoes. Bacterial communities in early instar C. tarsalis larvae were significantly more diverse when compared to late instar larvae, pupae and newly emerged adults. Some of the bacterial OTUs found in the early instar larvae were also found across developmental stages. Thorsellia dominated the bacterial communities in field-collected immature stages but occurred at much lower relative abundance in adults. Differences in microbiota observed in larval habitats did not influence bacterial community profiles of late instar larvae or adults. However, bacterial communities in laboratory-reared C. tarsalis larvae differed significantly from the field. Determining the role of Thorsellia in mosquitoes and its distribution across different species of mosquitoes warrants further investigation.

Species-specific differences in the responses of egg-laying mosquitoes (Diptera: Culicidae) to the presence of fish exudates in oviposition sites in laboratory bioassays have been related to the likelihood of encountering mosquito-eating fish in natural oviposition sites. We examined the responses of egg-laying mosquitoes to the presence of larvivorous fish in oviposition sites to test this hypothesis in the field. The number of Culex tarsalis Coquillett egg rafts laid on mesocosms (15.5 m²; 8 m³) containing caged mosquitofish (0.066 Gambusia affinis per liter) was reduced by 84% relative to mesocosms lacking fish. Egg-laying Culex quinquefasciatus Say did not differentiate significantly between comparatively small (0.11 m²) oviposition sites containing water conditioned with mosquitofish (0.3 G. affinis per liter) versus aged reservoir water. Cx. quinquefasciatus egg rafts were not collected from the larger mesocosms, and Cx. tarsalis egg rafts were rarely collected from the smaller oviposition sites. Oviposition preferences for characteristics of aquatic habitats that lack fish (e.g., small size, semipermanence, and high levels of organic enrichment in which hypoxic conditions are prevalent) may limit the coexistence of immature stages of some mosquito species such as Cx. quinquefasciatus and insectivorous fish. Mosquito species such as Cx. tarsalis that also use comparatively large water bodies as developmental sites may have evolved the ability to detect the presence of predatory fish.

Commercially available formulations of two entomopathogenic fungi, Beauveria bassiana (Bals.-Criv.) Vuill. (Hypocreales: Clavicipitaceae) and Metarhizium anisopliae (Metchnikoff) Sorokin (Hypocreales: Clavicipitaceae), were assessed for control of Culex quinquefasciatus Say (Diptera: Culicidae) in underground storm drain systems (USDS) in the Coachella Valley of southern California. Each of three treatments, the two fungi or a water control, was applied to 1 m² of vertical wall at eight USDS sites in spring and autumn of 2015. Fungal infectivity and lethality were assessed at 1 d and 1, 2, and 4 wk post-application. Overnight bioassays using adult lab-reared female mosquitoes were carried out on the treated USDS wall areas and then mosquitoes were held in the laboratory for up to 21 d to allow fungal infections to be expressed. Postmortem fungal sporulation was assessed up to 2 wk at 100% humidity. Mosquito–fungal interactions also were assessed in bioassays of the three treatments on filter paper exposed to USDS conditions during autumn. Metarhizium anisopliae killed mosquitoes faster than B. bassiana; nevertheless, both freshly applied formulations caused greater than 80% mortality. Fungal persistence declined significantly after 1 wk under USDS conditions, but some infectivity persisted for more than 4 wk. Beauveria bassiana was more effective against Cx. qinquefasciatus in the spring, while M. anisopliae was more effective in the cooler conditions during autumn. USDS environmental conditions (e.g., temperature, relative humidity, standing water) influenced fungal-related mortality and infection of Cx. quinquefasciatus. The utility of these fungal formulations for mosquito abatement in the Coachella Valley and implications for fungal control agents in USDS environments are discussed.

Considerable previous research has focused on predator-associated semiochemicals and how they affect mosquito oviposition behavior. However, most of this work has been done without taking into consideration either the natural density of the predators or how other semiochemicals in aquatic environments might affect the responses of gravid mosquitoes. The influence of mosquitofish density, source water (tap vs pond), presence of freshly laid egg rafts, and removal of a putative source of semiochemicals (bacteria) on oviposition by Culex tarsalis Coquillett (Diptera: Culicidae) was studied in laboratory bioassays. Culex tarsalis females were deterred from laying egg rafts on water that contained semiochemicals associated with Gambusia affinis (Baird & Girard), but this deterrence was not strongly associated with the density of fish used to condition aged tap water. The number of egg rafts laid onto Gambusia-exudate water made with either tap water (density < 1 fish per liter) or pond water was typically ≥ 50% of that onto water that did not house mosquitofish. Gravid mosquitoes tested individually did not reduce oviposition onto Gambusia-exudate water as compared to controls. Likewise, oviposition by females with ablated wings did not differ significantly between Gambusia-exudate water and controls. Oviposition onto filter-sterilized Gambusia-exudate water was reduced relative to unfiltered water, suggesting that semiochemicals deterring egg-laying were still present after bacteria were removed. Taken together, these findings suggest that the responses of gravid Cx. tarsalis to chemicals from habitats containing mosquitofish are complex and the origin of the semiochemicals present in the Gambusia-exudate water needs to be elucidated.

Gravid mosquito collections were compared among several large-volume (infusion volume ≥35 liters) gravid trap designs and the small-volume (infusion volume = 6 liters) Centers for Disease Control and Prevention (CDC) gravid trap used routinely by vector control districts for vector and pathogen surveillance. The numbers of gravid Culex quinquefasciatus, Cx. tarsalis, and Cx. stigmatosoma collected by large gravid traps were greater than by the CDC gravid trap during nearly all overnight trials. Large-volume gravid traps collected on average 6.6-fold more adult female Culex mosquitoes compared to small-volume CDC gravid traps across 3 seasons during the 3 years of the studies. The differences in gravid mosquito collections between large-versus small-volume gravid traps were greatest during spring, when 8- to 56-fold more Culex individuals were collected using large-volume gravid traps. The proportion of gravid females in collections did not differ appreciably among the more effective trap desig...