A genotyping array for the globally invasive vector mosquito, Aedes albopictus.

BACKGROUND: Although whole-genome sequencing (WGS) is the preferred genotyping method for most genomic analyses, limitations are often experienced when studying genomes characterized by a high percentage of repetitive elements, high linkage, and recombination deserts. The Asian tiger mosquito (Aedes albopictus), for example, has a genome comprising up to 72% repetitive elements, and therefore we set out to develop a single-nucleotide polymorphism (SNP) chip to be more cost-effective. Aedes albopictus is an invasive species originating from Southeast Asia that has recently spread around the world and is a vector for many human diseases. Developing an accessible genotyping platform is essential in advancing biological control methods and understanding the population dynamics of this pest species, with significant implications for public health. METHODS: We designed a SNP chip for Ae. albopictus (Aealbo chip) based on approximately 2.7 million SNPs identified using WGS data from 819 worldwide samples. We validated the chip using laboratory single-pair crosses, comparing technical replicates, and comparing genotypes of samples genotyped by WGS and the SNP chip. We then used the chip for a population genomic analysis of 237 samples from 28 sites in the native range to evaluate its usefulness in describing patterns of genomic variation and tracing the origins of invasions. RESULTS: Probes on the Aealbo chip targeted 175,396 SNPs in coding and non-coding regions across all three chromosomes, with a density of 102 SNPs per 1 Mb window, and at least one SNP in each of the 17,461 protein-coding genes. Overall, 70% of the probes captured the genetic variation. Segregation analysis found that 98% of the SNPs followed expectations of single-copy Mendelian genes. Comparisons with WGS indicated that sites with genotype disagreements were mostly heterozygotes at loci with WGS read depth < 20, while there was near complete agreement with WGS read depths > 20, indicating that the chip more accurately detects heterozygotes than low-coverage WGS. Sample sizes did not affect the accuracy of the SNP chip genotype calls. Ancestry analyses identified four to five genetic clusters in the native range with various levels of admixture. CONCLUSIONS: The Aealbo chip is highly accurate, is concordant with genotypes from WGS with high sequence coverage, and may be more accurate than low-coverage WGS.

Phylogenomics reveals the history of host use in mosquitoes.

Mosquitoes have profoundly affected human history and continue to threaten human health through the transmission of a diverse array of pathogens. The phylogeny of mosquitoes has remained poorly characterized due to difficulty in taxonomic sampling and limited availability of genomic data beyond the most important vector species. Here, we used phylogenomic analysis of 709 single copy ortholog groups from 256 mosquito species to produce a strongly supported phylogeny that resolves the position of the major disease vector species and the major mosquito lineages. Our analyses support an origin of mosquitoes in the early Triassic (217 MYA [highest posterior density region: 188-250 MYA]), considerably older than previous estimates. Moreover, we utilize an extensive database of host associations for mosquitoes to show that mosquitoes have shifted to feeding upon the blood of mammals numerous times, and that mosquito diversification and host-use patterns within major lineages appear to coincide in earth history both with major continental drift events and with the diversification of vertebrate classes.

Vertical habitat stratification in Aedes triseriatus and Aedes hendersoni (Diptera: Culicidae): complications associated with sloped and flood-prone landscapes.

For the sibling mosquito species Aedes triseriatus Say and Ae. hendersoni Cockerell, success of offspring is highly dependent on oviposition site selection by gravid females, and the dynamics of that selection process have been the subject of much investigation. Patterns of vertical oviposition stratification were examined in this study by placing basal and canopy level oviposition traps in relatively steep and flat terrains, as well as in flood-prone riverbank vs upland settings. Strong stratification existed in both steep and flat terrains, with Ae. triseriatus favoring basal ovitraps and Ae. hendersoni favoring canopy ovitraps. Contrary to expectations, Ae. hendersoni displayed even more marked preference for canopy ovitraps in steeper terrain. Both species avoided basal ovitraps in flood-prone locations, where Ae. triseriatus oviposited only in canopy ovitraps and greatly outnumbered Ae. hendersoni in canopy ovitraps. In contrast, the expected stratification was found in nearby upland habitats.

Field Evidence of Mosquito Population Regulation by a Gregarine Parasite.

Although parasites are by definition costly to their host, demonstrating that a parasite is regulating its host abundance in the field can be difficult. Here we present an example of a gregarine parasite, Ascogregarina taiwanensis Lien and Levine (Apicomplexa: Lecudinidae), regulating its mosquito host, Aedes albopictus Skuse (Diptera: Culicidae), in Bermuda. We sampled larvae from container habitats over 2 yr, assessed parasite prevalence, and estimated host abundance from egg counts obtained in neighboring ovitraps. We regressed change in average egg count from 1 yr to the next on parasite prevalence and found a significant negative effect of parasite prevalence. We found no evidence of host density affecting parasite prevalence. Our results demonstrate that even for a parasite with moderate virulence, host regulation can occur in the field.

Vertical Habitat Stratification in Sympatric and Allopatric Populations of Aedes hendersoni and Aedes triseriatus (Diptera: Culicidae).

Vertical habitat stratification in populations of Aedes hendersoni Cockerell (Diptera: Culicidae) and Aedes triseriatus (Say) (Diptera: Culicidae) has been observed to varying degrees throughout the species' sympatric range, and potential causes of the phenomenon, including species competition and interaction, have been debated extensively. Stratification patterns in oviposition in allopatric, sympatric, and marginally sympatric populations of both species were investigated and compared in this research to detect any pattern differences related to species composition. Expected patterns were observed in sympatric populations, with Ae. hendersoni preferentially ovipositing in canopy habitats, whereas Ae. triseriatus preferred basal habitats. Allopatric populations presented a strong shift toward basal preference in the former and a slighter but significant shift toward canopy in the latter. Marginal populations of Ae. hendersoni showed intermediate height preferences, whereas preferences of marginal and sympatric Ae. triseriatus did not differ. The convergence of habitat selection in allopatric populations and corresponding divergence in sympatric populations support interspecific competition-based hypotheses regarding the origin of the stratification phenomenon, although plausible alternative or contributing explanations are numerous and warrant further study.

From ground pools to treeholes: convergent evolution of habitat and phenotype in Aedes mosquitoes.

BACKGROUND: Invasive mosquito species are responsible for millions of vector-borne disease cases annually. The global invasive success of Aedes mosquitoes such as Aedes aegypti and Aedes albopictus has relied on the human transport of immature stages in container habitats. However, despite the importance of these mosquitoes and this ecological specialization to their widespread dispersal, evolution of habitat specialization in this group has remained largely unstudied. We use comparative methods to evaluate the evolution of habitat specialization and its potential influence on larval morphology, and evaluate whether container dwelling and invasiveness are monophyletic in Aedes. RESULTS: We show that habitat specialization has evolved repeatedly from ancestral ground pool usage to specialization in container habitats. Furthermore, we find that larval morphological scores are significantly associated with larval habitat when accounting for evolutionary relationships. We find that Ornstein-Uhleinbeck models with unique optima for each larval habitat type are preferred over several other models based predominantly on neutral processes, and that OU models can reliably simulate real morphological data. CONCLUSIONS: Our results demonstrate that multiple lineages of Aedes have convergently evolved a key trait associated with invasive success: the use of container habitats for immature stages. Moreover, our results demonstrate convergence in morphological characteristics as well, and suggest a role of adaptation to habitat specialization in driving phenotypic diversity in this mosquito lineage. Finally, our results highlight that the genus Aedes is not monophyletic.

Differential response to mosquito host sex and parasite dosage suggest mixed dispersal strategies in the parasite Ascogregarina taiwanensis.

Mixed dispersal strategies are a form of bet hedging in which a species or population utilizes different dispersal strategies dependent upon biotic or abiotic conditions. Here we provide an example of a mixed dispersal strategy in the Aedes albopictus / Ascogregarina taiwanensis host/parasite system, wherein upon host emergence, the gregarine parasite is either carried with an adult mosquito leaving the larval habitat, or released back into the larval habitat. We show that the parasite invests a larger proportion of its dispersing (oocyst) life stage into adult female mosquitoes as opposed to adult male mosquitoes at low parasite exposure levels. However, as the exposure level of parasite increases, so does the parasite investment in adult males, whereas there is no change in the proportion of oocysts in the adult female, regardless of dose. Thus, A. taiwanensis is utilizing several dispersal strategies, depending upon host sex and intraspecific density. Furthermore, we demonstrate that this parasite reduces body size, increases time to emergence in females, and leads to a reduction in estimates of per capita growth rate of the host.

A parasite's modification of host behavior reduces predation on its host.

Parasite modification of host behavior is common, and the literature is dominated by demonstrations of enhanced predation on parasitized prey resulting in transmission of parasites to their next host. We present a case in which predation on parasitized prey is reduced. Despite theoretical modeling suggesting that this phenomenon should be common, it has been reported in only a few host-parasite-predator systems. Using a system of gregarine endosymbionts in host mosquitoes, we designed experiments to compare the vulnerability of parasitized and unparasitized mosquito larvae to predation by obligate predatory mosquito larvae and then compared behavioral features known to change in the presence of predatory cues. We exposed Aedes triseriatus larvae to the parasite Ascogregarina barretti and the predator Toxohrynchites rutilus and assessed larval mortality rate under each treatment condition. Further, we assessed behavioral differences in larvae due to infection and predation stimuli by recording larvae and scoring behaviors and positions within microcosms. Infection with gregarines reduced cohort mortality in the presence of the predator, but the parasite did not affect mortality alone. Further, infection by parasites altered behavior such that infected hosts thrashed less frequently than uninfected hosts and were found more frequently on or in a refuge within the microcosm. By reducing predation on their host, gregarines may be acting as mutualists in the presence of predation on their hosts. These results illustrate a higher-order interaction, in which a relationship between a species pair (host-endosymbiont or predator-prey) is altered by the presence of a third species.

Sexual harassment and feeding inhibition between two invasive dengue vectors.

Two invasive mosquitoes, Aedes aegypti and Aedes albopictus, have been interacting during the course of a rapid range expansion by A. albopictus. We investigated the potential for interspecific feeding interference by male mosquitoes interacting with females within and between these species. A. aegypti feeding on both sugar and blood was suppressed when females of this species were exposed to A. albopictus males, but no change was observed when exposed to conspecifics. A. albopictus feeding was not affected by males of either species. The potential consequences of these behaviors are discussed within the context of other known interspecific effects, all of which appear to favor the displacement of A. aegypti by A. albopictus.

Hatch plasticity in response to varied inundation frequency in Aedes albopictus.

Eggs of container-breeding mosquitoes are able to withstand drought conditions as an egg and hatch when submerged. Frequent rainfall can be simulated by frequent submersion, and drought conditions can be simulated by infrequent submersion. We examined the hatch response of Aedes albopictus (Skuse) eggs to simulated drought conditions. Ae. albopictus eggs from a strain originating outside Kobe, Japan, were subjected to one of three treatments; high-frequency hatch stimulation consisting of submerging the eggs in a nutrient broth mixture every 3 d, low-frequency hatch stimulation consisting of submerging the eggs every 7 d, and delayed high-frequency hatch stimulation. Eggs that were subjected to lower-frequency stimulation showed a significant decrease in hatch delay, which was the opposite of the predicted response. This decrease in hatch delay may be an example of hatch plasticity in response to drought conditions. This response could not be explained as a result of the difference in the ages of the eggs on any given stimulus. A decreased hatch delay response to potential drought conditions combined with rapid larval development may enable Ae. albopictus, whose eggs are not as desiccation resistant as some other container-breeding mosquitoes, to survive extended drought.

Climate and geographic trends in hatch delay of the treehole mosquito, Aedes triseriatus Say (Diptera: Culicidae).

Eggs of Aedes triseriatus mosquitoes are stimulated to hatch when inundated with water, but only a small fraction of eggs from the same batch will hatch for any given stimulus. Similar hatching or germination patterns are observed in desert plants, copepods, rotifers, insects, and many other species. Bet hedging theory suggests that parents stagger offspring emergence into vulnerable life history stages in order to avoid catastrophic reproductive failures. For Ae. triseriatus, a treehole breeding mosquito, immediate hatching of an entire clutch leaves all of the parent's progeny vulnerable to extinction in the event of a severe drought. Natural selection has likely favored parents that pursued a bet hedging strategy where the risk of reproductive failure is distributed over time. Considering treehole mosquitoes, bet hedging theory could be used to predict that hatch delay would be positively correlated with the likelihood of drought. To test this prediction, we collected Ae. triseriatus from habitats that varied widely in mean annual precipitation and exposed them to several hatch stimuli in the laboratory. Here we report that, as predicted, Ae. triseriatus eggs from high precipitation regions showed less hatch delay than areas of low precipitation. This strategy probably allows Ae. triseriatus to cope with the wide variety of climatic conditions that it faces in its extensive geographical range.

Experience-induced changes in sugar taste discrimination.

An apparent plasticity in glucose sensitivity was first noted while studying human taste variants, but the experimental design did not rule out regression to the mean. Since then, a human taste induction hypothesis that sensitivity for a taste stimulus increases with repeated exposure to it has been supported first by experience-induced changes in taste identification of monosodium glutamate and, subsequently, in sensory detection of glutaraldehyde, as well as in psychophysical and functional magnetic resonance imaging responses to novel taste stimuli. Yet, whether such plasticity occurs for the highly familiar taste of sugar remained unconfirmed. Therefore, we tested the taste induction hypothesis for sugar using a counterbalanced design, consisting of 3 pretreatment and 2 treatment conditions. The effects over time also were followed with an additional group of participants. The results showed that 1) experience with fructose significantly increases sensitivity for the taste of a sugar, glucose; 2) there are no significant differences in the sugar sensitivity between groups of randomly assigned participants before treatments; 3) a single session of 5 brief tastings of glucose has an effect on glucose sensitivity when tested 11 or 12 days later; and 4) without continued treatment, the increased sensitivity reverses within 33 or 34 days.

Field and laboratory comparison of hatch rates in Aedes albopictus (Skuse).

Laboratory experiments attempting to elicit a response based on a natural condition rely on the assumption that the laboratory treatment accurately mimics field conditions. With Aedes albopictus (Skuse), laboratory experiments analyzing hatch rates assume that the laboratory stimuli resemble those received by the eggs in field conditions. With the use of a colonized strain of Ae. albopictus, an analysis of the hatch rates comparing both field and laboratory settings was conducted. Additionally, hatch rates were compared for mosquitoes exposed to regular, periodic hatch stimulation (as usually seen in laboratory experiments) and random hatch stimulation (as seen in the field). In both experiments, laboratory treatments were not found to differ significantly from the field treatments, indicating that experimental results achieved in the lab are relevant to field situations.

Polymorphic Foraging Behavior Among Caenorhabditis elegans: Frequency- and Density-Dependent Selection.

Strains of Caenorhabditis elegans obtained from their natural soil environment exhibit one of two forms of foraging behavior. Some strains forage solitarily and disperse evenly on a bacterial lawn. Other strains move rapidly until they encounter groups of conspecifics, and then slow their movement and join the group. Strains expressing these behaviors are globally widespread and have been isolated from the same location, suggesting a foraging polymorphism. We hypothesized that density-dependent selection maintains both foraging alleles in populations. Alternatively, both foraging alleles could be retained in populations through frequency-dependent selection. We tested both of these hypotheses by manipulating strain density and frequency, and observing changes in population density over time. Our results indicated that neither density- nor frequency-dependent selection appears to be responsible for the observed polymorphism. The clumping strain consistently out-competed the solitary strain over all treatment levels. We suggest other potential factors that may maintain both alleles in populations.