Strain heterogeneity in a non-pathogenic fungus highlights factors contributing to virulence.

Fungal pathogens exhibit extensive strain heterogeneity, including variation in virulence. Whether closely related non-pathogenic species also exhibit strain heterogeneity remains unknown. Here, we comprehensively characterized the pathogenic potentials (i.e., the ability to cause morbidity and mortality) of 16 diverse strains of Aspergillus fischeri, a non-pathogenic close relative of the major pathogen Aspergillus fumigatus. In vitro immune response assays and in vivo virulence assays using a mouse model of pulmonary aspergillosis showed that A. fischeri strains varied widely in their pathogenic potential. Furthermore, pangenome analyses suggest that A. fischeri genomic and phenotypic diversity is even greater. Genomic, transcriptomic, and metabolomic profiling identified several pathways and secondary metabolites associated with variation in virulence. Notably, strain virulence was associated with the simultaneous presence of the secondary metabolites hexadehydroastechrome and gliotoxin. We submit that examining the pathogenic potentials of non-pathogenic close relatives is key for understanding the origins of fungal pathogenicity.

Archaic Introgression Shaped Human Circadian Traits.

When the ancestors of modern Eurasians migrated out of Africa and interbred with Eurasian archaic hominins, namely, Neanderthals and Denisovans, DNA of archaic ancestry integrated into the genomes of anatomically modern humans. This process potentially accelerated adaptation to Eurasian environmental factors, including reduced ultraviolet radiation and increased variation in seasonal dynamics. However, whether these groups differed substantially in circadian biology and whether archaic introgression adaptively contributed to human chronotypes remain unknown. Here, we traced the evolution of chronotype based on genomes from archaic hominins and present-day humans. First, we inferred differences in circadian gene sequences, splicing, and regulation between archaic hominins and modern humans. We identified 28 circadian genes containing variants with potential to alter splicing in archaics (e.g., CLOCK, PER2, RORB, and RORC) and 16 circadian genes likely divergently regulated between present-day humans and archaic hominins, including RORA. These differences suggest the potential for introgression to modify circadian gene expression. Testing this hypothesis, we found that introgressed variants are enriched among expression quantitative trait loci for circadian genes. Supporting the functional relevance of these regulatory effects, we found that many introgressed alleles have associations with chronotype. Strikingly, the strongest introgressed effects on chronotype increase morningness, consistent with adaptations to high latitude in other species. Finally, we identified several circadian loci with evidence of adaptive introgression or latitudinal clines in allele frequency. These findings identify differences in circadian gene regulation between modern humans and archaic hominins and support the contribution of introgression via coordinated effects on variation in human chronotype.

Machine learning reveals the diversity of human 3D chromatin contact patterns.

Understanding variation in chromatin contact patterns across human populations is critical for interpreting non-coding variants and their ultimate effects on gene expression and phenotypes. However, experimental determination of chromatin contacts at a population-scale is prohibitively expensive. To overcome this challenge, we develop and validate a machine learning method to quantify the diversity 3D chromatin contacts at 2 kilobase resolution from genome sequence alone. We then apply this approach to thousands of diverse modern humans and the inferred human-archaic hominin ancestral genome. While patterns of 3D contact divergence genome-wide are qualitatively similar to patterns of sequence divergence, we find that 3D divergence in local 1-megabase genomic windows does not follow sequence divergence. In particular, we identify 392 windows with significantly greater 3D divergence than expected from sequence. Moreover, 26% of genomic windows have rare 3D contact variation observed in a small number of individuals. Using in silico mutagenesis we find that most sequence changes to do not result in changes to 3D chromatin contacts. However in windows with substantial 3D divergence, just one or a few variants can lead to divergent 3D chromatin contacts without the individuals carrying those variants having high sequence divergence. In summary, inferring 3D chromatin contact maps across human populations reveals diverse contact patterns. We anticipate that these genetically diverse maps of 3D chromatin contact will provide a reference for future work on the function and evolution of 3D chromatin contact variation across human populations.

The immune deficiency and c-Jun N-terminal kinase pathways drive the functional integration of the immune and circulatory systems of mosquitoes.

The immune and circulatory systems of animals are functionally integrated. In mammals, the spleen and lymph nodes filter and destroy microbes circulating in the blood and lymph, respectively. In insects, immune cells that surround the heart valves (ostia), called periostial haemocytes, destroy pathogens in the areas of the body that experience the swiftest haemolymph (blood) flow. An infection recruits additional periostial haemocytes, amplifying heart-associated immune responses. Although the structural mechanics of periostial haemocyte aggregation have been defined, the genetic factors that regulate this process remain less understood. Here, we conducted RNA sequencing in the African malaria mosquito, Anopheles gambiae, and discovered that an infection upregulates multiple components of the immune deficiency (IMD) and c-Jun N-terminal kinase (JNK) pathways in the heart with periostial haemocytes. This upregulation is greater in the heart with periostial haemocytes than in the circulating haemocytes or the entire abdomen. RNA interference-based knockdown then showed that the IMD and JNK pathways drive periostial haemocyte aggregation and alter phagocytosis and melanization on the heart, thereby demonstrating that these pathways regulate the functional integration between the immune and circulatory systems. Understanding how insects fight infection lays the foundation for novel strategies that could protect beneficial insects and harm detrimental ones.

Genomic and Molecular Identification of Genes Contributing to the Caspofungin Paradoxical Effect in Aspergillus fumigatus.

Aspergillus fumigatus is a deadly opportunistic fungal pathogen responsible for ~100,000 annual deaths. Azoles are the first line antifungal agent used against A. fumigatus, but azole resistance has rapidly evolved making treatment challenging. Caspofungin is an important second-line therapy against invasive pulmonary aspergillosis, a severe A. fumigatus infection. Caspofungin functions by inhibiting ß-1,3-glucan synthesis, a primary and essential component of the fungal cell wall. A phenomenon termed the caspofungin paradoxical effect (CPE) has been observed in several fungal species where at higher concentrations of caspofungin, chitin replaces ß-1,3-glucan, morphology returns to normal, and growth rate increases. CPE appears to occur in vivo, and it is therefore clinically important to better understand the genetic contributors to CPE. We applied genomewide association (GWA) analysis and molecular genetics to identify and validate candidate genes involved in CPE. We quantified CPE across 67 clinical isolates and conducted three independent GWA analyses to identify genetic variants associated with CPE. We identified 48 single nucleotide polymorphisms (SNPs) associated with CPE. We used a CRISPR/Cas9 approach to generate gene deletion mutants for seven genes harboring candidate SNPs. Two null mutants, ΔAfu3g13230 and ΔAfu4g07080 (dscP), resulted in reduced basal growth rate and a loss of CPE. We further characterized the dscP phosphatase-null mutant and observed a significant reduction in conidia production and extremely high sensitivity to caspofungin at both low and high concentrations. Collectively, our work reveals the contribution of Afu3g13230 and dscP in CPE and sheds new light on the complex genetic interactions governing this phenotype. IMPORTANCE This is one of the first studies to apply genomewide association (GWA) analysis to identify genes involved in an Aspergillus fumigatus phenotype. A. fumigatus is an opportunistic fungal pathogen that causes hundreds of thousands of infections and ~100,000 deaths each year, and antifungal resistance has rapidly evolved in this species. A phenomenon called the caspofungin paradoxical effect (CPE) occurs in some isolates, where high concentrations of the drug lead to increased growth rate. There is clinical relevance in understanding the genetic basis of this phenotype, since caspofungin concentrations could lead to unintended adverse clinical outcomes in certain cases. Using GWA analysis, we identified several interesting candidate polymorphisms and genes and then generated gene deletion mutants to determine whether these genes were important for CPE. Two of these mutant strains (ΔAfu3g13230 and ΔAfu4g07080/ΔdscP) displayed a loss of the CPE. This study sheds light on the genes involved in clinically important phenotype CPE.

A novel, likely pathogenic variant in UBTF-related neurodegeneration with brain atrophy is associated with a severe divergent neurodevelopmental phenotype.

BACKGROUND: A de novo, pathogenic, missense variant in UBTF, c.628G>A p.Glu210Lys, has been described as the cause of an emerging neurodegenerative disorder, Childhood-Onset Neurodegeneration with Brain Atrophy (CONDBA). The p.Glu210Lys alteration yields a positively charged stretch of three lysine residues. Functional studies confirmed this change results in a stronger interaction with negatively charged DNA and gain-of-function activity when compared to the wild-type sequence. The CONDBA phenotype reported in association with p.Glu210Lys consists of normal early-neurodevelopment followed by progressive motor, cognitive, and behavioral regression in early-to-middle childhood. METHODS AND RESULTS: The current proband presented at 9 months of age with baseline developmental delay and more extensive neuroradiological findings, including pontine hypoplasia, thalamic volume loss and signal abnormality, and hypomyelination. Like the recurrent CONDBA p.Glu210Lys variant, this novel variant, c.608A>G p.(Gln203Arg) lies within the highly conserved second HMG-box homology domain and involves the replacement of the wild-type residue with a positively charged residue, arginine. Computational structural modeling demonstrates that this amino acid substitution potentiates the interaction between UBTF and DNA, likely resulting in a gain-of-function effect for the UBTF protein, UBF. CONCLUSION: Here we present a new divergent phenotype associated with a novel, likely pathogenic, missense variant at a different position in the UBTF gene, c.608A>G p.(Gln203Arg).

Personalized structural biology reveals the molecular mechanisms underlying heterogeneous epileptic phenotypes caused by de novo KCNC2 variants.

Whole-exome sequencing (WES) in the clinic has identified several rare monogenic developmental and epileptic encephalopathies (DEE) caused by ion channel variants. However, WES often fails to provide actionable insight for rare diseases, such as DEEs, due to the challenges of interpreting variants of unknown significance (VUS). Here, we describe a "personalized structural biology" (PSB) approach that leverages recent innovations in the analysis of protein 3D structures to address this challenge. We illustrate this approach in an Undiagnosed Diseases Network (UDN) individual with DEE symptoms and a de novo VUS in KCNC2 (p.V469L), the Kv3.2 voltage-gated potassium channel. A nearby KCNC2 variant (p.V471L) was recently suggested to cause DEE-like phenotypes. Computational structural modeling suggests that both affect protein function. However, despite their proximity, the p.V469L variant is likely to sterically block the channel pore, while the p.V471L variant is likely to stabilize the open state. Biochemical and electrophysiological analyses demonstrate heterogeneous loss-of-function and gain-of-function effects, as well as differential response to 4-aminopyridine treatment. Molecular dynamics simulations illustrate that the pore of the p.V469L variant is more constricted, increasing the energetic barrier for K+ permeation, whereas the p.V471L variant stabilizes the open conformation. Our results implicate variants in KCNC2 as causative for DEE and guide the interpretation of a UDN individual. They further delineate the molecular basis for the heterogeneous clinical phenotypes resulting from two proximal pathogenic variants. This demonstrates how the PSB approach can provide an analytical framework for individualized hypothesis-driven interpretation of protein-coding VUS.

Comparative Genomics Reveals a Single Nucleotide Deletion in pksP That Results in White-Spore Phenotype in Natural Variants of Aspergillus fumigatus.

Aspergillus fumigatus is a potentially deadly opportunistic human pathogen. A. fumigatus has evolved a variety of mechanisms to evade detection by the immune system. For example, the conidium surface is covered in a layer of 1,8-dihydroxynaphthalene (DHN) melanin which masks the antigen macrophages use for recognition. DHN melanin also protects conidia from ultraviolet radiation and gives A. fumigatus conidia their characteristic green-grayish color. Here, we conducted genomic analysis of two closely related white-spore natural variants of A. fumigatus in comparison to two closely related green-spore isolates to identify a genetic basis of the white-spore phenotype. Illumina whole-genome resequencing data of the four isolates was used to identify variants that were shared in the white-spore isolates and different from both the green-spore isolates and the Af293 reference genome (which is also a green-spore isolate). We identified 4,279 single nucleotide variants and 1,785 insertion/deletions fitting this pattern. Among these, we identified 64 variants predicted to be high impact, loss-of-function mutations. One of these variants is a single nucleotide deletion that results in a frameshift in pksP (Afu2g17600), the core biosynthetic gene in the DHN melanin encoding gene cluster. The frameshift mutation in the white-spore isolates leads to a truncated protein in which a phosphopantetheine attachment site (PP-binding domain) is interrupted and an additional PP-binding domain and a thioesterase domain are omitted. Growth rate analysis of white-spore and green-spore isolates at 37°C and 48°C revealed that white-spore isolates are thermosensitive. Growth rate of A. fumigatus Af293 and a pksP null mutant in the Af293 background suggests pksP is not directly involved in the thermosensitivity phenotype. Further, our study identified a mutation in a gene (Afu4g04740) associated with thermal sensitivity in yeasts which could also be responsible for the thermosensitivity of the white-spore mutants. Overall, we used comparative genomics to identify the mutation and protein alterations responsible for the white-spore phenotype of environmental isolates of A. fumigatus.

Transcriptome profiles of Anopheles gambiae harboring natural low-level Plasmodium infection reveal adaptive advantages for the mosquito.

Anopheline mosquitoes are the sole vectors for the Plasmodium pathogens responsible for malaria, which is among the oldest and most devastating of human diseases. The continuing global impact of malaria reflects the evolutionary success of a complex vector-pathogen relationship that accordingly has been the long-term focus of both debate and study. An open question in the biology of malaria transmission is the impact of naturally occurring low-level Plasmodium infections of the vector on the mosquito's health and longevity as well as critical behaviors such as host-preference/seeking. To begin to answer this, we have completed a comparative RNAseq-based transcriptome profile study examining the effect of biologically salient, salivary gland transmission-stage Plasmodium infection on the molecular physiology of Anopheles gambiae s.s. head, sensory appendages, and salivary glands. When compared with their uninfected counterparts, Plasmodium infected mosquitoes exhibit increased transcript abundance of genes associated with olfactory acuity as well as a range of synergistic processes that align with increased fitness based on both anti-aging and reproductive advantages. Taken together, these data argue against the long-held paradigm that malaria infection is pathogenic for anophelines and, instead suggests there are biological and evolutionary advantages for the mosquito that drive the preservation of its high vectorial capacity.

Quantifying the contribution of Neanderthal introgression to the heritability of complex traits.

Eurasians have ~2% Neanderthal ancestry, but we lack a comprehensive understanding of the genome-wide influence of Neanderthal introgression on modern human diseases and traits. Here, we quantify the contribution of introgressed alleles to the heritability of more than 400 diverse traits. We show that genomic regions in which detectable Neanderthal ancestry remains are depleted of heritability for all traits considered, except those related to skin and hair. Introgressed variants themselves are also depleted for contributions to the heritability of most traits. However, introgressed variants shared across multiple Neanderthal populations are enriched for heritability and have consistent directions of effect on several traits with potential relevance to human adaptation to non-African environments, including hair and skin traits, autoimmunity, chronotype, bone density, lung capacity, and menopause age. Integrating our results, we propose a model in which selection against introgressed functional variation was the dominant trend (especially for cognitive traits); however, for a few traits, introgressed variants provided beneficial variation via uni-directional (e.g., lightening skin color) or bi-directional (e.g., modulating immune response) effects.

Neanderthal introgression reintroduced functional ancestral alleles lost in Eurasian populations.

Neanderthal ancestry remains across modern Eurasian genomes and introgressed sequences influence diverse phenotypes. Here, we demonstrate that introgressed sequences reintroduced thousands of ancestral alleles that were lost in Eurasian populations before introgression. Our simulations and variant effect predictions argue that these reintroduced alleles (RAs) are more likely to be tolerated by modern humans than are introgressed Neanderthal-derived alleles (NDAs) due to their distinct evolutionary histories. Consistent with this, we show enrichment for RAs and depletion for NDAs on introgressed haplotypes with expression quantitative trait loci (eQTL) and phenotype associations. Analysis of available cross-population eQTLs and massively parallel reporter assay data show that RAs commonly influence gene expression independent of linked NDAs. We further validate these independent effects for one RA in vitro. Finally, we demonstrate that NDAs are depleted for regulatory activity compared to RAs, while RAs have activity levels similar to non-introgressed variants. In summary, our study reveals that Neanderthal introgression reintroduced thousands of lost ancestral variants with gene regulatory activity and that these RAs were more tolerated than NDAs. Thus, RAs and their distinct evolutionary histories must be considered when evaluating the effects of introgression.

Polar bear evolution is marked by rapid changes in gene copy number in response to dietary shift.

Polar bear (Ursus maritimus) and brown bear (Ursus arctos) are recently diverged species that inhabit vastly differing habitats. Thus, analysis of the polar bear and brown bear genomes represents a unique opportunity to investigate the evolutionary mechanisms and genetic underpinnings of rapid ecological adaptation in mammals. Copy number (CN) differences in genomic regions between closely related species can underlie adaptive phenotypes and this form of genetic variation has not been explored in the context of polar bear evolution. Here, we analyzed the CN profiles of 17 polar bears, 9 brown bears, and 2 black bears (Ursus americanus). We identified an average of 318 genes per individual that showed evidence of CN variation (CNV). Nearly 200 genes displayed species-specific CN differences between polar bear and brown bear species. Principal component analysis of gene CN provides strong evidence that CNV evolved rapidly in the polar bear lineage and mainly resulted in CN loss. Olfactory receptors composed 47% of CN differentiated genes, with the majority of these genes being at lower CN in the polar bear. Additionally, we found significantly fewer copies of several genes involved in fatty acid metabolism as well as AMY1B, the salivary amylase-encoding gene in the polar bear. These results suggest that natural selection shaped patterns of CNV in response to the transition from an omnivorous to primarily carnivorous diet during polar bear evolution. Our analyses of CNV shed light on the genomic underpinnings of ecological adaptation during polar bear evolution.

Maintaining Biological Cultures and Measuring Gene Expression in Aphis nerii: A Non-model System for Plant-insect Interactions.

Aphids are excellent experimental models for a variety of biological questions ranging from the evolution of symbioses and the development of polyphenisms to questions surrounding insect's interactions with their host plants. Genomic resources are available for several aphid species, and with advances in the next-generation sequencing, transcriptomic studies are being extended to non-model organisms that lack genomes. Furthermore, aphid cultures can be collected from the field and reared in the laboratory for the use in organismal and molecular experiments to bridge the gap between ecological and genetic studies. Last, many aphids can be maintained in the laboratory on their preferred host plants in perpetual, parthenogenic life cycles allowing for comparisons of asexually reproducing genotypes. Aphis nerii, the milkweed-oleander aphid, provides one such model to study insect interactions with toxic plants using both organismal and molecular experiments. Methods for the generation and maintenance of the plant and aphid cultures in the greenhouse and laboratory, DNA and RNA extractions, microsatellite analysis, de novo transcriptome assembly and annotation, transcriptome differential expression analysis, and qPCR verification of differentially expressed genes are outlined and discussed here.

Transcriptional profile and differential fitness in a specialist milkweed insect across host plants varying in toxicity.

Interactions between plants and herbivorous insects have been models for theories of specialization and co-evolution for over a century. Phytochemicals govern many aspects of these interactions and have fostered the evolution of adaptations by insects to tolerate or even specialize on plant defensive chemistry. While genomic approaches are providing new insights into the genes and mechanisms insect specialists employ to tolerate plant secondary metabolites, open questions remain about the evolution and conservation of insect counterdefences, how insects respond to the diversity defences mounted by their host plants, and the costs and benefits of resistance and tolerance to plant defences in natural ecological communities. Using a milkweed-specialist aphid (Aphis nerii) model, we test the effects of host plant species with increased toxicity, likely driven primarily by increased secondary metabolites, on aphid life history traits and whole-body gene expression. We show that more toxic plant species have a negative effect on aphid development and lifetime fecundity. When feeding on more toxic host plants with higher levels of secondary metabolites, aphids regulate a narrow, targeted set of genes, including those involved in canonical detoxification processes (e.g., cytochrome P450s, hydrolases, UDP-glucuronosyltransferases and ABC transporters). These results indicate that A. nerii marshal a variety of metabolic detoxification mechanisms to circumvent milkweed toxicity and facilitate host plant specialization, yet, despite these detoxification mechanisms, aphids experience reduced fitness when feeding on more toxic host plants. Disentangling how specialist insects respond to challenging host plants is a pivotal step in understanding the evolution of specialized diet breadths.

Comprehensive RNA profiling of villous trophoblast and decidua basalis in pregnancies complicated by preterm birth following intra-amniotic infection.

INTRODUCTION: We performed RNA sequencing with the primary goal of discovering key placental villous trophoblast (VT) and decidua basalis (DB) transcripts differentially expressed in intra-amniotic infection (IAI)-induced preterm birth (PTB). METHODS: RNA was extracted from 15 paired VT and DB specimens delivered of women with: 1) spontaneous PTB in the setting of amniocentesis-proven IAI and histological chorioamnionitis (n = 5); 2) spontaneous idiopathic PTB (iPTB, n = 5); and 3) physiologic term pregnancy (n = 5). RNA sequencing was performed using the Illumina HiSeq 2500 platform, and a spectrum of computational tools was used for gene prioritization and pathway analyses. RESULTS: In the VT specimens, 128 unique long transcripts and 7 mature microRNAs differed significantly between pregnancies complicated by IAI relative to iPTB (FDR<0.1). The up-regulated transcripts included many characteristic of myeloblast-derived cells, and bioinformatic analyses revealed enrichment for multiple pathways associated with acute inflammation. In an expanded cohort including additional IAI and iPTB specimens, the expression of three proteins (cathepsin S, lysozyme, and hexokinase 3) and two microRNAs (miR-133a and miR-223) was validated using immunohistochemistry and quantitative PCR, respectively. In the DB specimens, only 11 long transcripts and no microRNAs differed significantly between IAI cases and iPTB controls (FDR<0.1). Comparison of the VT and DB specimens in each clinical scenario revealed signatures distinguishing these placental regions. DISCUSSION: IAI is associated with a transcriptional signature consistent with acute inflammation in the villous trophoblast. The present findings illuminate novel signaling pathways involved in IAI, and suggest putative therapeutic targets and potential biomarkers associated with this condition.

Disease vectors in the era of next generation sequencing.

Almost 20 % of all infectious human diseases are vector borne and, together, are responsible for over one million deaths per annum. Over the past decade, the decreasing costs of massively parallel sequencing technologies have facilitated the agnostic interrogation of insect vector genomes, giving medical entomologists access to an ever-expanding volume of high-quality genomic and transcriptomic data. In this review, we highlight how genomics resources have provided new insights into the physiology, behavior, and evolution of human disease vectors within the context of the global health landscape.

Comparing human and macaque placental transcriptomes to disentangle preterm birth pathology from gestational age effects.

INTRODUCTION: A major issue in the transcriptomic study of spontaneous preterm birth (sPTB) in humans is the inability to collect healthy control tissue at the same gestational age (GA) to compare with pathologic preterm tissue. Thus, gene expression differences identified after the standard comparison of sPTB and term tissues necessarily reflect differences in both sPTB pathology and GA. One potential solution is to use GA-matched controls from a closely related species to tease apart genes that are dysregulated during sPTB from genes that are expressed differently as a result of GA effects. METHODS: To disentangle genes whose expression levels are associated with sPTB pathology from those linked to GA, we compared RNA sequencing data from human preterm placentas, human term placentas, and rhesus macaque placentas at 80% completed gestation (serving as healthy non-human primate GA-matched controls). We first compared sPTB and term human placental transcriptomes to identify significantly differentially expressed genes. We then overlaid the results of the comparison between human sPTB and macaque placental transcriptomes to identify sPTB-specific candidates. Finally, we overlaid the results of the comparison between human term and macaque placental transcriptomes to identify GA-specific candidates. RESULTS: Examination of relative expression for all human genes with macaque orthologs identified 267 candidate genes that were significantly differentially expressed between preterm and term human placentas. 29 genes were identified as sPTB-specific candidates and 37 as GA-specific candidates. Altogether, the 267 differentially expressed genes were significantly enriched for a variety of developmental, metabolic, reproductive, immune, and inflammatory functions. Although there were no notable differences between the functions of the 29 sPTB-specific and 37 GA-specific candidate genes, many of these candidates have been previously shown to be dysregulated in diverse pregnancy-associated pathologies. DISCUSSION: By comparing human sPTB and term transcriptomes with GA-matched control transcriptomes from a closely related species, this study disentangled the confounding effects of sPTB pathology and GA, leading to the identification of 29 promising sPTB-specific candidate genes and 37 genes potentially related to GA effects. The apparent similarity in functions of the sPTB and GA candidates may suggest that the effects of sPTB and GA do not correspond to biologically distinct processes. Alternatively, it may reflect the poor state of knowledge of the transcriptional landscape underlying placental development and disease.

The origin and genetic differentiation of the socially parasitic aphid Tamalia inquilinus.

Social and brood parasitisms are nonconsumptive forms of parasitism involving the exploitation of the colonies or nests of a host. Such parasites are often related to their hosts and may evolve in various ecological contexts, causing evolutionary constraints and opportunities for both parasites and their hosts. In extreme cases, patterns of diversification between social parasites and their hosts can be coupled, such that diversity of one is correlated with or even shapes the diversity of the other. Aphids in the genus Tamalia induce galls on North American manzanita (Arctostaphylos) and related shrubs (Arbutoideae) and are parasitized by nongalling social parasites or inquilines in the same genus. We used RNA sequencing to identify and generate new gene sequences for Tamalia and performed maximum-likelihood, Bayesian and phylogeographic analyses to reconstruct the origins and patterns of diversity and host-associated differentiation in the genus. Our results indicate that the Tamalia inquilines are monophyletic and closely related to their gall-forming hosts on Arctostaphylos, supporting a previously proposed scenario for origins of these parasitic aphids. Unexpectedly, population structure and host-plant-associated differentiation were greater in the non-gall-inducing parasites than in their gall-inducing hosts. RNA-seq indicated contrasting patterns of gene expression between host aphids and parasites, and perhaps functional differences in host-plant relationships. Our results suggest a mode of speciation in which host plants drive within-guild diversification in insect hosts and their parasites. Shared host plants may be sufficient to promote the ecological diversification of a network of phytophagous insects and their parasites, as exemplified by Tamalia aphids.

The genomics of microbial domestication in the fermented food environment.

Shortly after the agricultural revolution, the domestication of bacteria, yeasts, and molds, played an essential role in enhancing the stability, quality, flavor, and texture of food products. These domestication events were probably the result of human food production practices that entailed the continual recycling of isolated microbial communities in the presence of abundant agricultural food sources. We suggest that within these novel agrarian food niches the metabolic requirements of those microbes became regular and predictable resulting in rapid genomic specialization through such mechanisms as pseudogenization, genome decay, interspecific hybridization, gene duplication, and horizontal gene transfer. The ultimate result was domesticated strains of microorganisms with enhanced fermentative capacities.

Divergent and conserved elements comprise the chemoreceptive repertoire of the nonblood-feeding mosquito Toxorhynchites amboinensis.

Many mosquito species serve as vectors of diseases such as malaria and yellow fever, wherein pathogen transmission is tightly associated with the reproductive requirement of taking vertebrate blood meals. Toxorhynchites is one of only three known mosquito genera that does not host-seek and initiates egg development in the absence of a blood-derived protein bolus. These remarkable differences make Toxorhynchites an attractive comparative reference for understanding mosquito chemosensation as it pertains to host-seeking. We performed deep transcriptome profiling of adult female Toxorhynchites amboinensis bodies, antennae and maxillary palps, and identified 25,084 protein-coding "genes" in the de novo assembly. Phylogenomic analysis of 4,266 single-copy "genes" from T. amboinensis, Aedes aegypti, Anopheles gambiae, and Culex quinquefasciatus robustly supported Ae. aegypti as the closest relative of T. amboinensis, with the two species diverged approximately 40 Ma. We identified a large number of T. amboinensis chemosensory "genes," the majority of which have orthologs in other mosquitoes. Finally, cross-species expression analyses indicated that patterns of chemoreceptor transcript abundance were very similar for chemoreceptors that are conserved between T. amboinensis and Ae. aegypti, whereas T. amboinensis appeared deficient in the variety of expressed, lineage-specific chemoreceptors. Our transcriptome assembly of T. amboinensis represents the first comprehensive genomic resource for a nonblood-feeding mosquito and establishes a foundation for future comparative studies of blood-feeding and nonblood-feeding mosquitoes. We hypothesize that chemosensory genes that display discrete patterns of evolution and abundance between T. amboinensis and blood-feeding mosquitoes are likely to play critical roles in host-seeking and hence the vectorial capacity.