Host‐associated genomic differentiation in congeneric butterflies: now you see it,now you do not

Ecotypic variation among populations may become associated with widespread genomic differentiation, but theory predicts that this should happen only under particular conditions of gene flow, selection and population size. In closely related species, we might expect the strength of host‐associated genomic differentiation (HAD) to be correlated with the degree of phenotypic differentiation in host‐adaptive traits. Using microsatellite and Amplified Fragment Length Polymorphism (AFLP) markers, and controlling for isolation by distance between populations, we sought HAD in two congeneric species of butterflies with different degrees of host plant specialization. Prior work on Euphydryas editha had shown strong interpopulation differentiation in host‐adapted traits, resulting in incipient reproductive isolation among host‐associated ecotypes. We show here that Euphydryas aurinia had much weaker host‐associated phenotypic differentiation. Contrary to our expectations, we detected HAD in Euphydryas aurinia, but not in E. editha. Even within an E. aurinia population that fed on both hosts, we found weak but significant sympatric HAD that persisted in samples taken 9 years apart. The finding of significantly stronger HAD in the system with less phenotypic differentiation may seem paradoxical. Our findings can be explained by multiple factors, ranging from differences in dispersal or effective population size, to spatial variation in genomic or phenotypic traits and to structure induced by past histories of host‐adapted populations. Other infrequently measured factors, such as differences in recombination rates, may also play a role. Our result adds to recent work as a further caution against assumptions of simple relationships between genomic and adaptive phenotypic differentiation.     

Convergence beyond flower morphology? Reproductive biology of hummingbird‐pollinated plants in the Brazilian Cerrado

Convergent reproductive traits in non‐related plants may be the result of similar environmental conditions and/or specialised interactions with pollinators. Here, we documented the pollination and reproductive biology of Bionia coriacea (Fabaceae), Esterhazya splendida (Orobanchaceae) and Ananas ananassoides (Bromeliaceae) as case studies in the context of hummingbird pollination in Cerrado, the Neotropical savanna of Central South America. We combined our results with a survey of hummingbird pollination studies in the region to investigate the recently suggested association of hummingbird pollination and self‐compatibility. Plant species studied here differed in their specialisation for ornithophily, from more generalist A. ananassoides to somewhat specialist B. coriacea and E. splendida. This continuum of specialisation in floral traits also translated into floral visitor composition. Amazilia fimbriata was the most frequent pollinator for all species, and the differences in floral display and nectar energy availability among plant species affect hummingbirds' behaviour. Most of the hummingbird‐pollinated Cerrado plants (60.0%, n = 20), including those studied here, were self‐incompatible, in contrast to other biomes in the Neotropics. Association to more generalist, often territorial, hummingbirds, and resulting reduced pollen flow in open savanna areas may explain predominance of self‐incompatibility. But it is possible that mating system is more associated with the predominance of woody hummingbird plants in the Cerrado plant assemblage than to the pollination system itself.     

Selection on outlier loci and their association with adaptive phenotypes in Littorina saxatilis contact zones

A fundamental issue in speciation research is to evaluate phenotypic variation and the genomics driving the evolution of reproductive isolation between sister taxa. Above all, hybrid zones are excellent study systems for researchers to examine the association of genetic differentiation, phenotypic variation and the strength of selection. We investigated two contact zones in the marine gastropod Littorina saxatilis and utilized landmark‐based geometric morphometric analysis together with amplified fragment length polymorphism (AFLP) markers to assess phenotypic and genomic divergence between ecotypes under divergent selection. From genetic markers, we calculated the cline width, linkage disequilibrium and the average effective selection on a locus. Additionally, we conducted an association analysis linking the outlier loci and phenotypic variation between ecotypes and show that a proportion of outlier loci are associated with key adaptive phenotypic traits.     

Molecular evolution of the sex peptide network in Drosophila

Successful reproduction depends on interactions between numerous proteins beyond those involved directly in gamete fusion. Although such reproductive proteins evolve in response to sexual selection pressures, how networks of interacting proteins arise and evolve as reproductive phenotypes change remains an open question. Here, we investigated the molecular evolution of the ‘sex peptide network’ of Drosophila melanogaster, a functionally well‐characterized reproductive protein network. In this species, the peptide hormone sex peptide (SP) and its interacting proteins cause major changes in female physiology and behaviour after mating. In contrast, females of more distantly related Drosophila species do not respond to SP. In spite of these phenotypic differences, we detected orthologs of all network proteins across 22 diverse Drosophila species and found evidence that most orthologs likely function in reproduction throughout the genus. Within SP‐responsive species, we detected the recurrent, adaptive evolution of several network proteins, consistent with sexual selection acting to continually refine network function. We also found some evidence for adaptive evolution of several proteins along two specific phylogenetic lineages that correspond with increased expression of the SP receptor in female reproductive tracts or increased sperm length, respectively. Finally, we used gene expression profiling to examine the likely degree of functional conservation of the paralogs of an SP network protein that arose via gene duplication. Our results suggest a dynamic history for the SP network in which network members arose before the onset of robust SP‐mediated responses and then were shaped by both purifying and positive selection.     

Transcribed microsatellite allele lengths are often correlated with gene expression in natural sunflower populations

Microsatellites are common in genomes of most eukaryotic species. Due to their high mutability, an adaptive role for microsatellites has been considered. However, little is known concerning the contribution of microsatellites towards phenotypic variation. We used populations of the common sunflower (Helianthus annuus) at two latitudes to quantify the effect of microsatellite allele length on phenotype at the level of gene expression. We conducted a common garden experiment with seed collected from sunflower populations in Kansas and Oklahoma followed by an RNA‐Seq experiment on 95 individuals. The effect of microsatellite allele length on gene expression was assessed across 3,325 microsatellites that could be consistently scored. Our study revealed 479 microsatellites at which allele length significantly correlates with gene expression (eSTRs). When irregular allele sizes not conforming to the motif length were removed, the number of eSTRs rose to 2,379. The percentage of variation in gene expression explained by eSTRs ranged from 1%–86% when controlling for population and allele‐by‐population interaction effects at the 479 eSTRs. Of these eSTRs, 70.4% are in untranslated regions (UTRs). A gene ontology (GO) analysis revealed that eSTRs are significantly enriched for GO terms associated with cis‐ and trans‐regulatory processes. Our findings suggest that a substantial number of transcribed microsatellites can influence gene expression.     

Polymorphism at the Clock gene predicts phenology of long‐distance migration in birds

Dissecting phenotypic variance in life history traits into its genetic and environmental components is at the focus of evolutionary studies and of pivotal importance to identify the mechanisms and predict the consequences of human‐driven environmental change. The timing of recurrent life history events (phenology) is under strong selection, but the study of the genes that control potential environmental canalization in phenological traits is at its infancy. Candidate genes for circadian behaviour entrained by photoperiod have been screened as potential controllers of phenological variation of breeding and moult in birds, with inconsistent results. Despite photoperiodic control of migration is well established, no study has reported on migration phenology in relation to polymorphism at candidate genes in birds. We analysed variation in spring migration dates within four trans‐Saharan migratory species (Luscinia megarhynchos; Ficedula hypoleuca; Anthus trivialis; Saxicola rubetra) at a Mediterranean island in relation to Clock and Adcyap1 polymorphism. Individuals with larger number of glutamine residues in the poly‐Q region of Clock gene migrated significantly later in one or, respectively, two species depending on sex and whether the within‐individual mean length or the length of the longer Clock allele was considered. The results hinted at dominance of the longer Clock allele. No significant evidence for migration date to covary with Adcyap1 polymorphism emerged. This is the first evidence that migration phenology is associated with Clock in birds. This finding is important for evolutionary studies of migration and sheds light on the mechanisms that drive bird phenological changes and population trends in response to climate change.     

Combining next‐generation sequencing and progeny testing for rapid identification of induced recessive and dominant mutations in maize M2 individuals

Molecular identification of mutant alleles responsible for certain phenotypic alterations is a central goal of genetic analyses. In this study we describe a rapid procedure suitable for the identification of induced recessive and dominant mutations applied to two Zea mays mutants expressing a dwarf and a pale green phenotype, respectively, which were obtained through pollen ethyl methanesulfonate (EMS) mutagenesis. First, without prior backcrossing, induced mutations (single nucleotide polymorphisms, SNPs) segregating in a (M₂) family derived from a heterozygous (M₁) parent were identified using whole‐genome shotgun (WGS) sequencing of a small number of (M₂) individuals with mutant and wild‐type phenotypes. Second, the state of zygosity of the mutation causing the phenotype was determined for each sequenced individual by phenotypic segregation analysis of the self‐pollinated (M₃) offspring. Finally, we filtered for segregating EMS‐induced SNPs whose state of zygosity matched the determined state of zygosity of the mutant locus in each sequenced (M₂) individuals. Through this procedure, combining sequencing of individuals and Mendelian inheritance, three and four SNPs in linkage passed our zygosity filter for the homozygous dwarf and heterozygous pale green mutation, respectively. The dwarf mutation was found to be allelic to the an1 locus and caused by an insertion in the largest exon of the AN1 gene. The pale green mutation affected the nuclear W2 gene and was caused by a non‐synonymous amino acid exchange in encoded chloroplast DNA polymerase with a predicted deleterious effect. This coincided with lower cpDNA levels in pale green plants.     

The genetic basis of drought tolerance in the high oil crop Sesamum indicum

Unlike most of the important food crops, sesame can survive drought but severe and repeated drought episodes, especially occurring during the reproductive stage, significantly curtail the productivity of this high oil crop. Genome‐wide association study was conducted for traits related to drought tolerance using 400 diverse sesame accessions, including landraces and modern cultivars. Ten stable QTLs explaining more than 40% of the phenotypic variation and located on four linkage groups were significantly associated with drought tolerance related traits. Accessions from the tropical area harboured higher numbers of drought tolerance alleles at the peak loci and were found to be more tolerant than those from the northern area, indicating a long‐term genetic adaptation to drought‐prone environments. We found that sesame has already fixed important alleles conferring survival to drought which may explain its relative high drought tolerance. However, most of the alleles crucial for productivity and yield maintenance under drought conditions are far from been fixed. This study also revealed that pyramiding the favourable alleles observed at the peak loci is of high potential for enhancing drought tolerance in sesame. In addition, our results highlighted two important pleiotropic QTLs harbouring known and unreported drought tolerance genes such as SiABI4, SiTTM3, SiGOLS1, SiNIMIN1 and SiSAM. By integrating candidate gene association study, gene expression and transgenic experiments, we demonstrated that SiSAM confers drought tolerance by modulating polyamine levels and ROS homeostasis, and a missense mutation in the coding region partly contributes to the natural variation of drought tolerance in sesame.     

Association between host wing morphology polymorphism and Wolbachia infection in Vollenhovia emeryi (Hymenoptera: Myrmicinae)

Many eusocial insects, including ants, show complex colony structures, distributions, and reproductive strategies. In the ant Vollenhovia emeryi Wheeler (Hymenoptera: Myrmicinae), queens and males are produced clonally, while sterile workers arise sexually, unlike other ant species and Hymenopteran insects in general. Furthermore, there is a wing length polymorphism in the queen caste. Despite its evolutionary remarkable traits, little is known about the population structure of this ant species, which may provide insight into its unique reproductive mode and polymorphic traits. We performed in‐depth analyses of ant populations from Korea, Japan, and North America using three mitochondrial genes (COI, COII, and Cytb). The long‐winged (L) morph is predominant in Korean populations, and the short‐winged (S) morph is very rare. Interestingly, all L morphs were infected with Wolbachia, while all Korean S morphs lacked Wolbachia, demonstrating a association between a symbiont and a phenotypic trait. A phylogenetic analysis revealed that the S morph is derived from the L morph. We propose that the S morph is associated with potential resistance to Wolbachia infection and that Wolbachia infection does not influence clonal reproduction (as is the case in other ant species).     

Variation in floral morphology and plant reproductive success in four Ipomoea species (Convolvulaceae) with contrasting breeding systems

• This study tested the hypothesis that self‐compatibility would be associated with floral traits that facilitate autonomous self‐pollination to ensure reproduction under low pollinator visitation. In a comparison of two pairs of Ipomoea species with contrasting breeding systems, we predicted that self‐compatible (SC) species would have smaller, less variable flowers, reduced herkogamy, lower pollinator visitation and higher reproductive success than their self‐incompatible (SI) congeners.
• We studied sympatric species pairs, I. hederacea (SC)– I. mitchellae (SI) and I. purpurea (SC)–I. indica (SI), in Mexico, over two years. We quantified variation in floral traits and nectar production, documented pollinator visitation, and determined natural fruit and seed set. Hand‐pollination and bagging experiments were conducted to determine potential for autonomous self‐pollination and apomixis.
• Self‐compatible Ipomoea species had smaller flowers and lower nectar production than SI species; however, floral variation and integration did not vary according to breeding system. Bees were primary pollinators of all species, but visitation rates were seven times lower in SC than SI species. SC species had a high capacity for autonomous self‐pollination due to reduced herkogamy at the highest anther levels. Self‐compatible species had two to six times higher fruit set than SI species.
• Results generally support the hypothesis that self‐compatibility and autonomous self‐pollination ensure reproduction under low pollinator visitation. However, high variation in morphological traits of SC Ipomoea species suggests they maintain variation through outcrossing. Furthermore, reduced herkogamy was associated with high potential for autonomous self‐pollination, providing a reproductive advantage that possibly underlies transitions to self‐compatibility in Ipomoea.

     

Quantifying natural seasonal variation in mutation parameters with mutation accumulation lines

Mutations create novel genetic variants, but their contribution to variation in fitness and other phenotypes may depend on environmental conditions. Furthermore, natural environments may be highly heterogeneous. We assessed phenotypes associated with survival and reproductive success in over 30,000 plants representing 100 mutation accumulation lines of Arabidopsis thaliana across four temporal environments at a single field site. In each of the four assays, environmental variance was substantially larger than mutational variance. For some traits, whether mutational variance was significantly varied between seasons. The founder genotype had mean trait values near the mean of the distribution of the mutation accumulation lines in all field experiments. New mutations also contributed more phenotypic variation than would be predicted, given phenotypic and sequence‐level divergence among natural populations of A. thaliana. The combination of large environmental variance with a mean effect of mutation near zero suggests that mutations could contribute substantially to standing genetic variation.     

Genome‐wide association study for reproductive traits in a Large White pig population

Using the PorcineSNP80 BeadChip, we performed a genome‐wide association study for seven reproductive traits, including total number born, number born alive, litter birth weight, average birth weight, gestation length, age at first service and age at first farrowing, in a population of 1207 Large White pigs. In total, we detected 12 genome‐wide significant and 41 suggestive significant SNPs associated with six reproductive traits. The proportion of phenotypic variance explained by all significant SNPs for each trait ranged from 4.46% (number born alive) to 11.49% (gestation length). Among them, 29 significant SNPs were located within known QTL regions for swine reproductive traits, such as corpus luteum number, stillborn number and litter size, of which one QTL region associated with litter size contained the ALGA0098819 SNP for total number born. Subsequently, we found that 376 functional genes contained or were near these significant SNPs. Of these, 14 genes—BHLHA15, OCM2, IL1B2, GCK, SMAD2, HABP2, PAQR5, GRB10, PRELID2, DMKN, GPI, GPIHBP1, ADCY2 and ACVR2B—were considered important candidates for swine reproductive traits based on their critical roles in embryonic development, energy metabolism and growth development. Our findings contribute to the understanding of the genetic mechanisms for reproductive traits and could have a positive effect on pig breeding programs.     

Genomic pedigree reconstruction identifies predictors of mating and reproductive success in an invasive vertebrate

The persistence of an invasive species is influenced by its reproductive ecology, and a successful control program must operate on this premise. However, the reproductive ecology of invasive species may be enigmatic due to factors that also limit their management, such as cryptic coloration and behavior. We explored the mating and reproductive ecology of the invasive Brown Treesnake (BTS: Boiga irregularis) by reconstructing a multigenerational genomic pedigree based on 654 single nucleotide polymorphisms for a geographically closed population established in 2004 on Guam (N = 426). The pedigree allowed annual estimates of individual mating and reproductive success to be inferred for snakes in the study population over a 14‐year period. We then employed generalized linear mixed models to gauge how well phenotypic and genomic data could predict sex‐specific annual mating and reproductive success. Average snout–vent length (SVL), average body condition index (BCI), and trappability were significantly related to annual mating success for males, with average SVL also related to annual mating success for females. Male and female annual reproductive success was positively affected by SVL, BCI, and trappability. Surprisingly, the degree to which individuals were inbred had no effect on annual mating or reproductive success. When juxtaposed with current control methods, these results indicate that baited traps, a common interdiction tool, may target fecund BTS in some regards but not others. Our study emphasizes the importance of reproductive ecology as a focus for improving BTS control and promotes genomic pedigree reconstruction for such an endeavor in this invasive species and others.     

Evolution of novel mimicry rings facilitated by adaptive introgression in tropical butterflies

Understanding the genetic basis of phenotypic variation and the mechanisms involved in the evolution of adaptive novelty, especially in adaptive radiations, is a major goal in evolutionary biology. Here, we used whole‐genome sequence data to investigate the origin of the yellow hindwing bar in the Heliconius cydno radiation. We found modular variation associated with hindwing phenotype in two narrow noncoding regions upstream and downstream of the cortex gene, which was recently identified as a pigmentation pattern controller in multiple species of Heliconius. Genetic variation at each of these modules suggests an independent control of the dorsal and ventral hindwing patterning, with the upstream module associated with the ventral phenotype and the downstream module with the dorsal one. Furthermore, we detected introgression between H. cydno and its closely related species Heliconius melpomene in these modules, likely allowing both species to participate in novel mimicry rings. In sum, our findings support the role of regulatory modularity coupled with adaptive introgression as an elegant mechanism by which novel phenotypic combinations can evolve and fuel an adaptive radiation.     

Genome‐wide association analysis for quantitative trait loci influencing Warner–Bratzler shear force in five taurine cattle breeds

We performed a genome‐wide association study for Warner–Bratzler shear force (WBSF), a measure of meat tenderness, by genotyping 3360 animals from five breeds with 54 790 BovineSNP50 and 96 putative single‐nucleotide polymorphisms (SNPs) within μ‐calpain [HUGO nomenclature calpain 1, (mu/I) large subunit; CAPN1] and calpastatin (CAST). Within‐ and across‐breed analyses estimated SNP allele substitution effects (ASEs) by genomic best linear unbiased prediction (GBLUP) and variance components by restricted maximum likelihood under an animal model incorporating a genomic relationship matrix. GBLUP estimates of ASEs from the across‐breed analysis were moderately correlated (0.31–0.66) with those from the individual within‐breed analyses, indicating that prediction equations for molecular estimates of breeding value developed from across‐breed analyses should be effective for genomic selection within breeds. We identified 79 genomic regions associated with WBSF in at least three breeds, but only eight were detected in all five breeds, suggesting that the within‐breed analyses were underpowered, that different quantitative trait loci (QTL) underlie variation between breeds or that the BovineSNP50 SNP density is insufficient to detect common QTL among breeds. In the across‐breed analysis, CAPN1 was followed by CAST as the most strongly associated WBSF QTL genome‐wide, and associations with both were detected in all five breeds. We show that none of the four commercialized CAST and CAPN1 SNP diagnostics are causal for associations with WBSF, and we putatively fine‐map the CAPN1 causal mutation to a 4581‐bp region. We estimate that variation in CAST and CAPN1 explains 1.02 and 1.85% of the phenotypic variation in WBSF respectively.     

Multidisciplinary approaches for species delimitation in Sisyrinchium (Iridaceae)

A remarkable morphological similarity in natural populations of different groups of Sisyrinchium spp. (Iridaceae) makes classification of some species difficult. The present paper focused on two morphological categories of S. sellowianum (MC‐I and MC‐II) with distinct morphological traits. Morphological, cytogenetic, reproductive biology and genetic studies were performed, aiming to elucidate and better characterize such variation. The basic chromosome number established for the species was x = 9 and diploid and tetraploid populations were observed for MC‐I, whereas MC‐II showed only diploid populations. Different pollen morphologies were recognized in each morphological category. Based on pollination experiments, MC‐I is out‐crossing, whereas MC‐II is selfing. The populations were highly differentiated (F_ST = 0.46, θ^B = 0.62). In MC‐I, observed variation was greater within populations (69%) than among populations (31%), whereas in MC‐II, 61% of the variation was among populations and 39% within populations. This study evidenced that the two morphological categories correspond to distinct and reproductively isolated species and that floral traits reflect their breeding system, with allogamous plants in MC‐I and autogamous in MC‐II. Furthermore, MC‐II is unveiled as S. platycaule, a neglected taxon of Sisyrinchium in Brazil. The tools applied in this study were substantially able to check the identity of the two morphological categories as two different species and present the variation encountered in the taxa.