A New Pipeline for Removing Paralogs in Target Enrichment Data.

Target enrichment (such as Hyb-Seq) is a well-established high throughput sequencing method that has been increasingly used for phylogenomic studies. Unfortunately, current widely used pipelines for analysis of target enrichment data do not have a vigorous procedure to remove paralogs in target enrichment data. In this study, we develop a pipeline we call Putative Paralogs Detection (PPD) to better address putative paralogs from enrichment data. The new pipeline is an add-on to the existing HybPiper pipeline, and the entire pipeline applies criteria in both sequence similarity and heterozygous sites at each locus in the identification of paralogs. Users may adjust the thresholds of sequence identity and heterozygous sites to identify and remove paralogs according to the level of phylogenetic divergence of their group of interest. The new pipeline also removes highly polymorphic sites attributed to errors in sequence assembly and gappy regions in the alignment. We demonstrated the value of the new pipeline using empirical data generated from Hyb-Seq and the Angiosperms353 kit for two woody genera Castanea (Fagaceae, Fagales) and Hamamelis (Hamamelidaceae, Saxifragales). Comparisons of data sets showed that the PPD identified many more putative paralogs than the popular method HybPiper. Comparisons of tree topologies and divergence times showed evident differences between data from HybPiper and data from our new PPD pipeline. We further evaluated the accuracy and error rates of PPD by BLAST mapping of putative paralogous and orthologous sequences to a reference genome sequence of Castanea mollissima. Compared to HybPiper alone, PPD identified substantially more paralogous gene sequences that mapped to multiple regions of the reference genome (31 genes for PPD compared with 4 genes for HybPiper alone). In conjunction with HybPiper, paralogous genes identified by both pipelines can be removed resulting in the construction of more robust orthologous gene data sets for phylogenomic and divergence time analyses. Our study demonstrates the value of Hyb-Seq with data derived from the Angiosperms353 probe set for elucidating species relationships within a genus, and argues for the importance of additional steps to filter paralogous genes and poorly aligned regions (e.g., as occur through assembly errors), such as our new PPD pipeline described in this study. [Angiosperms353; Castanea; divergence time; Hamamelis; Hyb-Seq, paralogs, phylogenomics.].

Genetic evidence for the origin of Aedes aegypti, the yellow fever mosquito, in the southwestern Indian Ocean.

Aedes aegypti is among the best-studied mosquitoes due to its critical role as a vector of human pathogens and ease of laboratory rearing. Until now, this species was thought to have originated in continental Africa, and subsequently colonized much of the world following the establishment of global trade routes. However, populations of this mosquito on the islands in the southwestern Indian Ocean (SWIO), where the species occurs with its nearest relatives referred to as the Aegypti Group, have received little study. We re-evaluated the evolutionary history of Ae. aegypti and these relatives, using three data sets: nucleotide sequence data, 18,489 SNPs and 12 microsatellites. We found that: (a) the Aegypti Group diverged 16 MYA (95% HPD: 7-28 MYA) from its nearest African/Asian ancestor; (b) SWIO populations of Ae. aegypti are basal to continental African populations; (c) after diverging 7 MYA (95% HPD: 4-15 MYA) from its nearest formally described relative (Ae. mascarensis), Ae. aegypti moved to continental Africa less than 85,000 years ago, where it recently (<1,000 years ago) split into two recognized subspecies Ae. aegypti formosus and a human commensal, Ae. aegypti aegypti; (d) the Madagascar samples form a clade more distant from all other Ae. aegypti than the named species Ae. mascarensis, implying that Madagascar may harbour a new cryptic species; and (e) there is evidence of introgression between Ae. mascarensis and Ae. aegypti on Réunion, and between the two subspecies elsewhere in the SWIO, a likely consequence of recent introductions of domestic Ae. aegypti aegypti from Asia.

Evolutionary dynamics of host specialization in wood-decay fungi.

BACKGROUND: The majority of wood decomposing fungi are mushroom-forming Agaricomycetes, which exhibit two main modes of plant cell wall decomposition: white rot, in which all plant cell wall components are degraded, including lignin, and brown rot, in which lignin is modified but not appreciably removed. Previous studies suggested that brown rot fungi tend to be specialists of gymnosperm hosts and that brown rot promotes gymnosperm specialization. However, these hypotheses were based on analyses of limited datasets of Agaricomycetes. Overcoming this limitation, we used a phylogeny with 1157 species integrating available sequences, assembled decay mode characters from the literature, and coded host specialization using the newly developed R package, rusda. RESULTS: We found that most brown rot fungi are generalists or gymnosperm specialists, whereas most white rot fungi are angiosperm specialists. A six-state model of the evolution of host specialization revealed high transition rates between generalism and specialization in both decay modes. However, while white rot lineages switched most frequently to angiosperm specialists, brown rot lineages switched most frequently to generalism. A time-calibrated phylogeny revealed that Agaricomycetes is older than the flowering plants but many of the large clades originated after the diversification of the angiosperms in the Cretaceous. CONCLUSIONS: Our results challenge the current view that brown rot fungi are primarily gymnosperm specialists and reveal intensive white rot specialization to angiosperm hosts. We thus suggest that brown rot associated convergent loss of lignocellulose degrading enzymes was correlated with host generalism, rather than gymnosperm specialism. A likelihood model of host specialization evolution together with a time-calibrated phylogeny further suggests that the rise of the angiosperms opened a new mega-niche for wood-decay fungi, which was exploited particularly well by white rot lineages.

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.

Simulated rRNA/DNA Ratios Show Potential To Misclassify Active Populations as Dormant.

The use of rRNA/DNA ratios derived from surveys of rRNA sequences in RNA and DNA extracts is an appealing but poorly validated approach to infer the activity status of environmental microbes. To improve the interpretation of rRNA/DNA ratios, we performed simulations to investigate the effects of community structure, rRNA amplification, and sampling depth on the accuracy of rRNA/DNA ratios in classifying bacterial populations as "active" or "dormant." Community structure was an insignificant factor. In contrast, the extent of rRNA amplification that occurs as cells transition from dormant to growing had a significant effect (P < 0.0001) on classification accuracy, with misclassification errors ranging from 16 to 28%, depending on the rRNA amplification model. The error rate increased to 47% when communities included a mixture of rRNA amplification models, but most of the inflated error was false negatives (i.e., active populations misclassified as dormant). Sampling depth also affected error rates (P < 0.001). Inadequate sampling depth produced various artifacts that are characteristic of rRNA/DNA ratios generated from real communities. These data show important constraints on the use of rRNA/DNA ratios to infer activity status. Whereas classification of populations as active based on rRNA/DNA ratios appears generally valid, classification of populations as dormant is potentially far less accurate.IMPORTANCE The rRNA/DNA ratio approach is appealing because it extracts an extra layer of information from high-throughput DNA sequencing data, offering a means to determine not only the seedbank of taxa present in communities but also the subset of taxa that are metabolically active. This study provides crucial insights into the use of rRNA/DNA ratios to infer the activity status of microbial taxa in complex communities. Our study shows that the approach may not be as robust as previously supposed, particularly in complex communities composed of populations employing different growth strategies, and identifies factors that inflate the erroneous classification of active populations as dormant.

Copy number variation contributes to cryptic genetic variation in outbreak lineages of Cryptococcus gattii from the North American Pacific Northwest.

BACKGROUND: Copy number variants (CNVs) are a class of structural variants (SVs) and are defined as fragments of DNA that are present at variable copy number in comparison with a reference genome. Recent advances in bioinformatics methodologies and sequencing technologies have enabled the high-resolution quantification of genome-wide CNVs. In pathogenic fungi SVs have been shown to alter gene expression, influence host specificity, and drive fungicide resistance, but little attention has focused specifically on CNVs. Using publicly available sequencing data, we identified 90 isolates across 212 Cryptococcus gattii genomes that belong to the VGII subgroups responsible for the recent deadly outbreaks in the North American Pacific Northwest. We generated CNV profiles for each sample to investigate the prevalence and function of CNV in C. gattii. RESULTS: We identified eight genetic clusters among publicly available Illumina whole genome sequence data from 212 C. gattii isolates through population structure analysis. Three clusters represent the VGIIa, VGIIb, and VGIIc subgroups from the North American Pacific Northwest. CNV was bioinformatically predicted and affected ~300-400 Kilobases (Kb) of the C. gattii VGII subgroup genomes. Sixty-seven loci, encompassing 58 genes, showed highly divergent patterns of copy number variation between VGII subgroups. Analysis of PFam domains within divergent CN variable genes revealed enrichment of protein domains associated with transport, cell wall organization and external encapsulating structure. CONCLUSIONS: CNVs may contribute to pathological and phenotypic differences observed between the C. gattii VGIIa, VGIIb, and VGIIc subpopulations. Genes overlapping with population differentiated CNVs were enriched for several virulence related functional terms. These results uncover novel candidate genes to examine the genetic and functional underpinnings of C. gattii pathogenicity.

Robustness in population-structure and demographic-inference results derived from the Aedes aegypti genotyping chip and whole-genome sequencing data.

The mosquito Aedes aegypti is the primary vector of many human arboviruses such as dengue, yellow fever, chikungunya, and Zika, which affect millions of people worldwide. Population genetic studies on this mosquito have been important in understanding its invasion pathways and success as a vector of human disease. The Axiom aegypti1 SNP chip was developed from a sample of geographically diverse A. aegypti populations to facilitate genomic studies on this species. We evaluate the utility of the Axiom aegypti1 SNP chip for population genetics and compare it with a low-depth shotgun sequencing approach using mosquitoes from the native (Africa) and invasive ranges (outside Africa). These analyses indicate that results from the SNP chip are highly reproducible and have a higher sensitivity to capture alternative alleles than a low-coverage whole-genome sequencing approach. Although the SNP chip suffers from ascertainment bias, results from population structure, ancestry, demographic, and phylogenetic analyses using the SNP chip were congruent with those derived from low-coverage whole-genome sequencing, and consistent with previous reports on Africa and outside Africa populations using microsatellites. More importantly, we identified a subset of SNPs that can be reliably used to generate merged databases, opening the door to combined analyses. We conclude that the Axiom aegypti1 SNP chip is a convenient, more accurate, low-cost alternative to low-depth whole-genome sequencing for population genetic studies of A. aegypti that do not rely on full allelic frequency spectra. Whole-genome sequencing and SNP chip data can be easily merged, extending the usefulness of both approaches.

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.

Phylogenetic resolution of the fly superfamily Ephydroidea-Molecular systematics of the enigmatic and diverse relatives of Drosophilidae.

The schizophoran superfamily Ephydroidea (Diptera: Cyclorrhapha) includes eight families, ranging from the well-known vinegar flies (Drosophilidae) and shore flies (Ephydridae), to several small, relatively unusual groups, the phylogenetic placement of which has been particularly challenging for systematists. An extraordinary diversity in life histories, feeding habits and morphology are a hallmark of fly biology, and the Ephydroidea are no exception. Extreme specialization can lead to "orphaned" taxa with no clear evidence for their phylogenetic position. To resolve relationships among a diverse sample of Ephydroidea, including the highly modified flies in the families Braulidae and Mormotomyiidae, we conducted phylogenomic sampling. Using exon capture from Anchored Hybrid Enrichment and transcriptomics to obtain 320 orthologous nuclear genes sampled for 32 species of Ephydroidea and 11 outgroups, we evaluate a new phylogenetic hypothesis for representatives of the superfamily. These data strongly support monophyly of Ephydroidea with Ephydridae as an early branching radiation and the placement of Mormotomyiidae as a family-level lineage sister to all remaining families. We confirm placement of Cryptochetidae as sister taxon to a large clade containing both Drosophilidae and Braulidae-the latter a family of honeybee ectoparasites. Our results reaffirm that sampling of both taxa and characters is critical in hyperdiverse clades and that these factors have a major influence on phylogenomic reconstruction of the history of the schizophoran fly radiation.

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.

From e-voucher to genomic data: Preserving archive specimens as demonstrated with medically important mosquitoes (Diptera: Culicidae) and kissing bugs (Hemiptera: Reduviidae).

Scientific collections such as the U.S. National Museum (USNM) are critical to filling knowledge gaps in molecular systematics studies. The global taxonomic impediment has resulted in a reduction of expert taxonomists generating new collections of rare or understudied taxa and these large historic collections may be the only reliable source of material for some taxa. Integrated systematics studies using both morphological examinations and DNA sequencing are often required for resolving many taxonomic issues but as DNA methods often require partial or complete destruction of a sample, there are many factors to consider before implementing destructive sampling of specimens within scientific collections. We present a methodology for the use of archive specimens that includes two crucial phases: 1) thoroughly documenting specimens destined for destructive sampling-a process called electronic vouchering, and 2) the pipeline used for whole genome sequencing of archived specimens, from extraction of genomic DNA to assembly of putative genomes with basic annotation. The process is presented for eleven specimens from two different insect subfamilies of medical importance to humans: Anophelinae (Diptera: Culicidae)-mosquitoes and Triatominae (Hemiptera: Reduviidae)-kissing bugs. Assembly of whole mitochondrial genome sequences of all 11 specimens along with the results of an ortholog search and BLAST against the NCBI nucleotide database are also presented.

The Prediapause Stage of Aedes japonicus japonicus and the Evolution of Embryonic Diapause in Aedini.

The genus Aedes is well known for its desiccation-resistant eggs, which frequently serve as an overwintering mechanism through diapause. Despite this, relatively little is known about the diapause and overwintering biology of most Aedes species including Aedes japonicus japonicus, an invasive mosquito in the United States. The importance of this mosquito in disease systems like La Crosse virus remain uncertain. Embryonic diapause is used by Ae. j. japonicus to survive temperate winters, and the persistence of this species in the Appalachian region is a result of overwintering, which has important implications for the transmission of this virus to humans. The objective of this study was to identify the prediapause stage, or the stage sensitive to environmental cues needed to induce diapause in this mosquito. By exposing each Ae. j. japonicus life stage independently to short-day photoperiods, we determined that the adult maternal life stage is the prediapause stage. Using the most recent phylogeny and prior literature on the prediapause stages in the genus Aedes, we were able to infer the evolutionary history of the prediapause stages of Aedes mosquitoes that overwinter or aestivate as eggs. This initial ancestral state reconstruction allowed us to hypothesize that Aedini mosquitoes that undergo obligate diapause may have evolved from those utilizing the embryonic prediapause stage, and that the ancestral prediapause state of Aedini appears to be maternally controlled.

Genetic diversity of laboratory strains and implications for research: The case of Aedes aegypti.

The yellow fever mosquito (Aedes aegypti), is the primary vector of dengue, Zika, and chikungunya fever, among other arboviral diseases. It is also a popular laboratory model in vector biology due to its ease of rearing and manipulation in the lab. Established laboratory strains have been used worldwide in thousands of studies for decades. Laboratory evolution of reference strains and contamination among strains are potential severe problems that could dramatically change experimental outcomes and thus is a concern in vector biology. We analyzed laboratory and field colonies of Ae. aegypti and an Ae. aegypti-derived cell line (Aag2) using 12 microsatellites and ~20,000 SNPs to determine the extent of divergence among laboratory strains and relationships to their wild relatives. We found that 1) laboratory populations are less genetically variable than their field counterparts; 2) colonies bearing the same name obtained from different laboratories may be highly divergent; 3) present genetic composition of the LVP strain used as the genome reference is incompatible with its presumed origin; 4) we document changes in two wild caught colonies over ~16 generations of colonization; and 5) the Aag2 Ae. aegypti cell line has experienced minimal genetic changes within and across laboratories. These results illustrate the degree of variability within and among strains of Ae. aegypti, with implications for cross-study comparisons, and highlight the need of a common mosquito repository and the implementation of strain validation tools.

First Record of Mansonia dyari From Saint Croix, United States Virgin Islands.

The first report of Mansonia dyari on Saint Croix, United States Virgin Islands (USVI), is confirmed. Adult and larval specimens were collected in 2018 and 2019 through adult surveillance and larval collections. Specimens were identified by microscopic methods, and a representative specimen was confirmed by DNA sequencing (mitochondrial cytochrome c oxidase subunit I). Morphological features are reviewed and compared with Mansonia flaveola, a species previously reported in the USVI. Notes are provided on the locations, collection methods, and mosquito associates found with Ma. dyari in the USVI.

Population genomics of Culiseta melanura, the principal vector of Eastern equine encephalitis virus in the United States.

BACKGROUND: Eastern Equine Encephalitis (EEE) (Togaviridae, Alphavirus) is a highly pathogenic mosquito-borne arbovirus that circulates in an enzootic cycle involving Culiseta melanura mosquitoes and wild Passeriformes birds in freshwater swamp habitats. Recently, the northeastern United States has experienced an intensification of virus activity with increased human involvement and northward expansion into new regions. In addition to its principal role in enzootic transmission of EEE virus among avian hosts, recent studies on the blood-feeding behavior of Cs. melanura throughout its geographic range suggest that this mosquito may also be involved in epizootic / epidemic transmission to equines and humans in certain locales. Variations in blood feeding behavior may be a function of host availability, environmental factors, and/or underlying genetic differences among regional populations. Despite the importance of Cs. melanura in transmission and maintenance of EEE virus, the genetics of this species remains largely unexplored. METHODOLOGY AND PRINCIPLE FINDINGS: To investigate the occurrence of genetic variation in Cs. melanura, the genome of this mosquito vector was sequenced resulting in a draft genome assembly of 1.28 gigabases with a contig N50 of 93.36 kilobases. Populations of Cs. melanura from 10 EEE virus foci in the eastern North America were genotyped with double-digest RAD-seq. Following alignment of reads to the reference genome, variant calling, and filtering, 40,384 SNPs were retained for downstream analyses. Subsequent analyses revealed genetic differentiation between northern and southern populations of this mosquito species. Moreover, limited fine-scale population structure was detected throughout northeastern North America, suggesting local differentiation of populations but also a history of ancestral polymorphism or contemporary gene flow. Additionally, a genetically distinct cluster was identified predominantly at two northern sites. CONCLUSION AND SIGNIFICANCE: This study elucidates the first evidence of fine-scale population structure in Cs. melanura throughout its eastern range and detects evidence of gene flow between populations in northeastern North America. This investigation provides the groundwork for examining the consequences of genetic variations in the populations of this mosquito species that could influence vector-host interactions and the risk of human and equine infection with EEE virus.

Structural and wetting properties of nature's finest silks (order Embioptera).

Insects from the order Embioptera (webspinners) spin silk fibres which are less than 200 nm in diameter. In this work, we characterized and compared the diameters of single silk fibres from nine species-Antipaluria urichi, Pararhagadochir trinitatis, Saussurembia calypso, Diradius vandykei, Aposthonia ceylonica, Haploembia solieri, H. tarsalis, Oligotoma nigra and O. saundersii. Silk from seven of these species have not been previously quantified. Our studies cover five of the 10 named taxonomic families and represent about one third of the known taxonomic family-level diversity in the order Embioptera. Naturally spun silk varied in diameter from 43.6 ± 1.7 nm for D. vandykei to 122.4 ± 3.2 nm for An. urichi. Mean fibre diameter did not correlate with adult female body length. Fibre diameter is more similar in closely related species than in more distantly related species. Field observations indicated that silk appears shiny and smooth when exposed to rainwater. We therefore measured contact angles to learn more about interactions between silk and water. Higher contact angles were measured for silks with wider fibre diameter and higher quantity of hydrophobic amino acids. High static contact angles (ranging up to 122° ± 3° for An. urichi) indicated that silken sheets spun by four arboreal, webspinner species were hydrophobic. A second contact angle measurement made on a previously wetted patch of silk resulted in a lower contact angle (average difference was greater than 27°) for all four species. Our studies suggest that silk fibres which had been previously exposed to water exhibited irreversible changes in hydrophobicity and water adhesion properties. Our results are in alignment with the 'super-pinning' site hypothesis by Yarger and co-workers to describe the hydrophobic, yet water adhesive, properties exhibited by webspinner silk fibres. The physical and chemical insights gained here may inform the synthesis and development of smaller diameter silk fibres with unique water adhesion properties.

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.

The First Evidence of Nanism in Ixodes (Ixodes) scapularis (Acari: Ixodidae), Found Parasitizing a Human Host.

Ixodes scapularis Say 1821, the primary vector of several human pathogens in the northeastern and upper Midwestern United States, has considerable genetic and morphological variation throughout its range. Recently, developmental or teratological abnormalities have been observed in this species for the first time, further complicating morphological identification. Here, we report the first evidence of nanism (dwarfism) in I. scapularis, found parasitizing a human host. We used molecular methods and scanning electron microscopy to identify the specimen. Morphological identification confirmed that the specimen is substantially smaller, approximately half the size, than a typical I. scapularis female. Here we discuss the recent reports of teratological abnormalities in I. scapularis, particularly from the Hudson River valley region of the northeastern United States, and highlight the need for additional studies of teratology in this important species and its potential implications in disease transmission.

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.