Furthermore, studies of fertility and sex ratio require the ability to sex-type degraded tissue (e.g. unhatched eggs). However, allelic dropout is common when attempting to PCR-amplify large products from degraded samples and dropout causes errors in sex typing. We therefore used the recently released data from multiple sequenced avian genomes to develop new primer sets, including some that amplify small PCR products (<150 bp) on both the W and Z chromosomes to enable sex typing of degraded samples. These markers are of utility for sex-typing a wide range of species and a combination of the ZW and Z markers developed can be used to identify rearrangements on the Z chromosome.

# Habitat segregation in an allopolyploid and its parent species in Cardamine
(51686)

Reiko Akiyama, Masaomi Hatakeyama, Rie Shimizu-Inatsugi, Xiangchao Gan, Miltos Tsiantis, Jun Sese, Kentaro K. Shimizu.

Institute of Evolutionary Biology and Envi environmental Studies, University of Zurich; Functional Genomic Center Zurich; Department of Comparative Development and Genetics, Max Planck Institute for Plant Breeding Research; Computational Biology Research Center (CBRC), AIST, Japan.

Whole genome duplication (polyploidy) is pervasive in plants. Polyploids are generally expected to have intermediate and broader environmental distributions than diploids. However, empirical data on environment of polyploids and its diploid parents in field are limited. We quantified habitat environment of diploids C. hirsuta and C. amara and the allopolyploid C. flexuosa originated from C. hirsuta and C. amara in their native area in Switzerland in order to examine whether habitat environment differs among species. The diploids C. hirsuta and C. amara segregated in that C. hirsuta occurred in dry, bright, and nutrient-rich habitat in contrast to C. amara, while the allopolyploid C. flexuosa occurred in intermediate and fluctuating environment. These findings provided evidence that the parent species occurred in extreme environments, while the allopolyploid occurs in intermediate environment. They also suggest that the C. amara, C. hirsuta, and C. flexuosa serve as a useful system to study the molecular basis of adaptive significance of allopolyploidy in wild plants.

# Is hybridization impacting the diversification rates of land plant polyploids ?
(51697)

Nils Arrigo, Michael S. Barker.

Unil; University of Arizona.

We assess the evolutionary significance of hybridization by comparing the extinction and speciation rates of species with a hybrid ancestry to those of non-hybrids. We focus on land
plants, and more particularly on polyploids because their high prevalence allows the comparison of allopolyploids (i.e. stabilized hybrids) to autopolyploids (i.e. non-hybrids). The study is based on 38 comprehensive genus-level phylogenies (built from publicly available DNA sequences), and on over 3,500 species for which we reviewed the polyploidy origin based on the current literature. Using this survey, we first show that allo- and autopolyploids are equally abundant in the majority of the inspected genera. State-specific diversification rates (i.e. the polyploidization, speciation and extinction rates of diploids, allo- and autopolyploids) are then estimated using the Multiple State Speciation and Extinction model (FitzJohn et al., 2009). We show that the extinction rates of allopolyploids are significantly lower than those of the autopolyploids. No differences are however observed in speciation rates, resulting in a significantly increased net diversification rate for the allopolyploids. These results are discussed in light of the classic hypotheses about the importance of hybridization in evolution.

# Mesopolyploid evolution in the Australian and New Zealand crucifers (51887)

Terezie Mandáková, Milan Pouch, Klára Harmanová, Martin A. Lysak.

CEITEC - Central European Institute of Technology, Masaryk University.

We showed previously that three endemic Australian crucifer species (Brassicaceae) have undergone a whole-genome duplication (AUS-WGD) followed by lineage-specific diploidization, generating some of the lowest chromosome numbers known for crucifers (Mandáková et al. 2010, Plant Cell). The New Zealand genus Pachycladon has undergone either the AUS-WGD or an independent WGD followed by less extensive fractionation towards n = 10 genomes (Mandáková et al. 2010, BMC Plant Biol.).

To further elucidate the origin and fate of Australian/New Zealand mesopolyploid genomes, we analyzed 12 species (9 genera) with variable chromosome numbers (n = 4, 5, 6, 7) by comparative chromosome painting (CCP) and gene sequencing. We concluded that:

(i) all Australian members of the Microlepidieae tribe share the AUS-WGD, which most likely spurred the diversification of the group on the Australian continent,

(ii) cytogenetic signatures and multi-gene phylogenies suggest that the allopolyploid ancestral genome (n = 15) was formed through an inter-tribal hybridization (Crucihimalayeae x Smelowskieae/Descurainieae) and a long-distance dispersal from Asia,

(iii) the AUS-WGD was followed by independent and massive genome diploidizations towards the extant diversity of quasi-diploid genomes (n = 4 - 7),

(iv) the New Zealand Pachycladon was formed by an independent and younger WGD event.
# Multiple patterns of genome evolution in the polyploid-rich genus Cardamine
(51944)

**Terezie Mandakova**, Aleš Kovařík, Judita Zozomová-Lihová, Rie Shimizu-Inatsugi, Kentaro K. Shimizu, Klaus Mummenhoff, Karol Marhold, Martin A. Lysak.

CEITEC, Masaryk University; Institute of Biophysics, AS CR, Brno, Czech Republic; Institute of Botany, SAS, Bratislava, Slovakia; Institute of Evolutionary Biology and Environmental Studies, University of Zurich, Switzerland; University Osnabrück, Germany.

Cardamine (bittercress) is one of the largest Brassicaceae genera (200 spp.). Most Cardamine species are polyploids with chromosome numbers up to n = 128. Due to the feasibility of chromosome painting, we documented both recurrent and deviating patterns of genome evolution in Cardamine polyploids of contrasting age:

(i) some tetraploid (2n = 32) populations of C. pratensis are on the way to decrease their chromosome number by "chromosome fusions" (2n = 30, 28).

(ii) hybridization and polyploidization is ongoing – we reconstruct the origin of the triploid hybrid C. x insueta (2n = 24, RRA) through hybridization between C. amara (AA) and C. rivularis (RR) ca. 100 years ago. Hybridization involving C. x insueta and hypotetraploid C. pratensis (2n = 30, PPPP) resulted in the origin of the hypohexaploid C. x schulzii (2n = 46, PPPPRA). This shows how a semifertile triploid hybrid can promote the origin of trigenomic allopolyploids.

(iii) we elucidate the origin of several European and Asian tetra- to octoploid species through hybridization events involving three diploid species.

(iv) some taxa are high polyploids - we demonstrate that C. bulbifera (2n = 96) is a largely sterile dodecaploid.

(v) several species, including watercress and horseradish, are shown to be autopolyploids.

Poster session B - POL 300

# Multiplex amplicon sequencing for genotyping and phylogenetic inference in polyploid wild wheats (51996)

**Stella Huynh**, François Felber, Christian Parisod.

University of Neuchâtel; Musée et jardins botaniques cantonaux de Lausanne.

Inferring the evolutionary history of polyploid species remains a major challenge. Recent theoretical advances have dramatically improved inference of reticulation and incomplete lineage sorting from genome-wide data (e.g. using Bayesian statistical modeling such as BEAST and PADRE). Obtaining good quality data from complex polyploid genome however currently remains a bottleneck. Capturing polymorphisms within and among homeologous genes is indeed crucial for such phylogenetic work. Here, we report on a multilocus dataset of
50 low-copy loci in hundreds of individuals from four diploid and four polyploid wild wheat species (Aegilops spp.). Loci of 400bp, encompassing polymorphic introns, are PCR amplified and individually tagged for multiplexed paired-end sequencing with Illumina MiSeq in one step. This work thus contributes to progress of methodologies offering high quality data to decipher the evolutionary history of polyploid species.

# Does genome duplication results in reproductive isolation in tetraploid-octoploid complexes? The case study of Gladiolus communis (52038)

**Mariana Castro, João Loureiro, Brian Husband, Sílvia Castro.**

*CFE, Centre for Functional Ecology; Department of Integrative Biology, University of Guelph.*

Polyploidization is an important mechanism of sympatric speciation in flowering plants. Under random mating, the establishment of new polyploids is restrict by strong positive frequency-dependent selection. Consequently, the establishment and spread of cytotypes will only occur when assortative mating is promoted. Our main objective was to assess breeding barriers operating between tetraploids and octoploids of Gladiolus communis at a contact zone in Western Iberian Peninsula. Spatial segregation, phenological isolation and crossing ability were investigated in 108 natural populations and common garden experiments. We observed that G. communis has a complex contact zone. There were two dominant cytotypes occurring in pure-ploidy populations (4x and 8x, 43.5% and 38.9%, respectively); mixed-ploidy (13.0%) and hexaploid populations (4.6%) were also detected, suggesting that both cytotypes can co-occur and produce viable hexaploid populations. Within mixed-ploidy populations each cytotype appeared to show different micro-habitat distributions. Also, overlap in flowering phenology was observed suggesting that pollen can be freely exchanged between cytotypes. Controlled pollination experiments revealed that, despite fitness being lower in inter-cytotype crosses than within-cytotypes, both produced viable offspring. Overall, our results indicate low reproductive isolation between cytotypes which could constrain polyploid speciation and promote a complex and dynamic contact zone.

# Fertility restoration during polyploid speciation in Mimulus (52173)

**Sofie Meeus, Mario Vallejo-Marín.**

*University of Stirling.*

Interspecific hybridisation between evolutionary divergent parental genomes produces hybrids which often are sterile due to genic incompatibilities, erroneous meiotic pairing and/or odd chromosome number establishing a reproductive barrier among diverged species. However, when accompanied by polyploidisation (“allopolypleidy”), sterile hybrids may give
rise to an entirely new euploid species which is reproductively isolated from its parental
species. However, even though polyploidisation leads to even chromosome numbers which is
essential for chromosomal pairing during meiosis, and thus fertility, new polyploids have to
face the challenges of chromosome segregation brought about the additional set(s) of
homologous chromosomes that lead to multivalent formation. Current research indicates that
genome restructuring occurs rapidly during the earliest generations of polyploid speciation.
However, much remains unknown about both the mechanisms responsible for and the rate at
which fertility is restored after polyploidisation. To investigate the rate and mechanisms of
fertility restoration during polyploid speciation, three subsequent generations of a newly
discovered allopolyplloid from the UK, Mimulus peregrinus, are experimentally generated
from different synthetic sterile triploid hybrid lines using colchicine. Fertility measured as
pollen viability is linked to early-generation chromosomal rearrangements using FISH to
assess the rate of fertility restoration in new allopolyloids.

Reference-guided de novo assembly: improved genome of a non-model plant
species (52224)

Heidi Lischer, Damianos Melidis, Masaomi Hatakeyama, Kentaro Shimizu.

URPP Evolution in Action, University of Zurich; Institute of Evolutionary Biology and Environmental Studies, University of Zurich; Swiss Institute of Bioinformatics, Lausanne; Functional Genomics Center Zurich.

The development of next-generation sequencing has made it possible to sequence whole
gnomes at a relatively low cost. However, de novo assembly of genomes still remains
challenging due to short read length, repetitive regions, missing data, polymorphisms and
sequencing errors. We implement a reference-guided assembly approach, which uses an
available reference sequence from a close relative in order to improve the genome assembly
outcome. The main idea is to first map reads against a reference genome to reduce the
complexity of de novo assembly within continuous covered regions. In a further step, reads
with no similarity to the reference are integrated. We apply this approach in the assembly of
the A. halleri genome and simulated reads. The genus Arabidopsis is an important model to
study environmental adaptation and polyploid speciation. With our reference-guided de novo
assembly strategy the number of scaffolds is reduced and the N50 highly increased compared
to a de novo assembly approach. The improved genome will facilitate the study of polyploidy
speciation at the genetic level, as this requires high quality genomes of the parental species.
We show that the de novo assembly of a genome can be improved by integrating information
of a related genome.

Polyploidy, morphology, and systemic fungal endophyte frequencies of
Festuca rubra L. in a broad scale geographic distribution (52316)
**Serdar Dirihan**, Marjo Helander, Irma Saloniemi, Henry Väre, Pedro E. Gundel, Kari Saikkonen.

*Section of Ecology, Department of Biology, University of Turku, 20014 Turku, Finland; Natural Resources Institute Finland (LUKE) Itäinen Pitkäkatu 3 20520 Turku, Finland; Division of Genetics and Physiology, Department of Biology, University of Turku, 20014 Turku, Finland; Botanical Museum, Finnish Museum of Natural History, University of Helsinki, 00014, Finland; IFEVA - CONICET - Faculty of Agronomy, Buenos Aires University (UBA), Buenos Aires, Argentina.*

We conducted studies of ploidy levels and presence of systemic fungal endophyte infection in the grass Festuca rubra L. across latitudes from Spain to northernmost Finland and Greenland. First we implemented a time- and cost-efficient flow cytometry procedure which is based on to use a plate-based analysis with a fully automated software tools. Using this procedure we analyzed 29 F. rubra populations. Most of the plants were hexaploids (2n=42; 83%), but also tetraploid (2n=28; 10%) and octoploid (2n=56; 7%) plants were detected. While ploidy levels seemed to be positively correlated with latitude, we could not detect explicit geographic gradients. The endophyte infection frequencies of plants varied from 0% to 81%. Our results suggest that variation in infection frequencies and ploidy levels might partly reflect Festuca distribution after the glaciation period, and be partly the consequences of local selective forces. We also analyzed morphological characters of 17 F. rubra populations from a broad latitudinal gradient across Europe. We found correlations among some morphological measurements and ploidy; e.g. the correlation between basal leaf width and plant height was strong for tetraploids and octoploids, but not for hexaploids. Our results suggest that selection favored similar morphology in each study site in spite of different ploidy levels.

# Temporal and spatial context of phylogeographic structure in the hexaploid KwaZulu-Natal Yellowfish (Labeobarbus natalensis) (52414)

**Connor Stobie**, Kerry Reid, Carel Oosthuizen, Paulette Bloomer.

*University of Pretoria.*

Labeobarbus natalensis (the KwaZulu-Natal Yellowfish) has a narrow distribution and limited dispersal capabilities, making it an ideal model species for phylogeographic analysis. However, the genus has undergone historical genome duplication events and is hexaploid. This complicates many traditional genetic analyses based on nuclear DNA. A previous study by our group revealed the pattern of mitochondrial DNA (mtDNA) diversity across the species' distribution, with several genetically divergent lineages restricted to different rivers. In the present study we explore the use of several different nuclear molecular markers in this polyploid species – microsatellites, exons, introns and a Next-Generation Sequencing (NGS) technology, Restriction-site Associated DNA sequencing (RAD-seq). Analyses of these nuclear DNA regions will also enable testing as to whether L. natalensis is functionally a hexaploid or a diploid despite its physical karyotype. Here we present some of the successful markers from each of the four categories and explore their relevance in identifying the patterns and processes affecting L. natalensis diversity. We focus primarily on the results
obtained from RAD-seq, which has been completed for 23 individuals. Future directions
include exploring transcriptomics in L. natalensis as a potentially superior method for analysis
in polyploid species.

# Interploid reproductive isolation variation within the Campanula rotundifolia polyploid complex (52509)

**Brittany Sutherland**, Laura Galloway.

*University of Virginia.*

Whole genome duplication often confers reproductive isolation (RI) between polyploids and their diploid progenitors. Strong or complete RI is documented in numerous diploid-tetraploid systems. Similar RI levels are often assumed in complexes comprising higher-order polyploids. However, evidence for strong RI among higher-order polyploids is sparse, and sometimes does not demonstrate high RI. We investigate the pattern and strength of interploid RI in Campanula rotundifolia s.l., a polyploid complex comprising diploids, tetraploids, and hexaploids. Tetraploid populations from throughout the geographic range were reciprocally crossed with diploid and hexaploid test populations. Maternal and F1 characters were then evaluated and compared to intraploid crosses to assess RI. RI was considerably lower among tetraploid-hexaploid crosses. However RI occurred at different life history stages for diploid and hexaploid hybrids. Diploid-tetraploid hybrids had extremely low germination while germination of tetraploid-hexaploid hybrids was indistinguishable from intraploid controls. Tetraploid-hexaploid hybrids experienced lower fertility while surviving diploid-tetraploid hybrids often had high fertility. Interestingly, these fertile diploid-tetraploid offspring were often even-ploidy instead of the expected triploid, suggesting that meiotic irregularity may be a mechanism for successful interploid reproduction. This study suggests RI estimates based on diploid-tetraploid systems may not apply to higher-order polyploid complexes.

# Sweet vernal grass in Europe: The origin of polyploid genomes in one genus (52511)

**Zuzana Chumová**, Pavel Trávníček, Eliška Záveská, Terezie Mandáková.

*Department of Botany, Faculty of Sciences, Charles University in Prague, 128 01, Czech Republic; Institute of Botany, Academy of Sciences of the Czech Republic, Průhonice, 252 43, Czech Republic; CEITEC - Central European Institute of Technology, Masaryk University, Brno, 601 77, Czech Republic.*

The genus Anthoxanthum (sweet vernal grass, Poaceae) represents a taxonomically intricate polyploid complex with large phenotypic variation and still poorly resolved evolutionary relationships. Our study was aimed at elucidating i) phylogenetic relationships among all
European taxa belonging to Anthoxanthum odoratum complex and ii) the evolutionary history of polyploids.

In order to get insight into the geographic distribution of ploidy levels and assess the taxonomic value of genome size data, flow cytometry was used. To answer the phylogenetic and evolutionary questions, single-copy nuclear loci (GBSSI), ITS and cpDNA sequencing was used together with FISH and GISH analyses.

Eight taxonomic groups that partly corresponded to traditionally recognized species (A. odoratum 2x and 4x, A. alpinum 2x and 4x, A. aristatum, A. ovatum, A. gracile, A. amarum) were delimited on the basis of genome size values and phenotypic variation. While our data supported the merger of A. aristatum and A. ovatum, eastern Mediterranean populations traditionally referred to as diploid A. odoratum were shown to be cytologically distinct and may represent a new taxon. Autopolyploid origin was suggested for 4x A. alpinum, on the contrary, 4x A. odoratum is an allopolyploid based on the molecular and GISH analyses.

Poster session B - POL 300

**Whole genome duplication does not show consistent response in fish and plant lineages fate. (52552)**

**Sacha Laurent, Nicolas Salamin, Marc Robinson-Rechavi.**

*Departement of Ecology and Evolution, University of Lausanne; Swiss Institute of Bioinformatics.*

Impact of whole genome duplications upon species fate has been debated since the discovery of their pervasiveness in the tree of life. Stemming from this and the apparent increase in species number in clades emerging from paleopolyploidy, many hypothesis have been proposed to explain those patterns. Neutral patterns such as reciprocal gene loss could theoretically lead to such observation, although this mode of action as been recently discussed and cast down. Adaptive processes, such as neofunctionalization of one of the duplicate genes, could also favor the apparition of new species. Data in favor of or against an increase in diversity following paleopolyploidy have been alternatively presented in the past few years. Using recent methods for estimating diversification of lineages, we analyse already published but also new phylogenies of species belonging to various plants and fishes group. Overall we do not find a consistent signal, indicating diverging responses in lineages fate after whole genome duplication.

Poster session B - POL 300

**The role of ploidy in host resistance: an experimental test of triploid and tetraploid snails (52615)**

**Christina Jenkins, Mark Dybdahl, Scott Nuismer.**

*Washington State University; Washington State University; University of Idaho.*
The evolution of sexual reproduction is widely believed to be driven by host-parasite coevolution. Meanwhile, elevated ploidy is expected to alter host-parasite coevolution through increased heterozygosity and dosage from the expression of duplicated genes. Interestingly, many asexual populations exhibit higher ploidy compared to their sexual ancestors. Yet, there are gaps in our understanding of the confounding effects of ploidy variation on the maintenance of sex. If resistance to parasites is based on self recognition, then the increased heterozygosity associated with polyploidy could decrease host resistance. However, if resistance is based on a quantitative model of infection, then higher host ploidy should lead to an increase in immune molecules that neutralize parasites. Here, we experimentally inoculated triploid and tetraploid asexual hosts to explicitly address if an increase in ploidy changes an individual's resistance. We used the snail, Potamopyrgus antipodarum, and its coevolved parasite, Microphallus. We tested hosts from five different populations with parasites from three different sources. We found that there was no significant difference in infection rate in triploids and tetraploids across all populations suggesting that host resistance is not driven by the quantitative model of infection.

# Terra incognita of contemporary plant biosystematics: untangling the sources of complexity in Ranunculus subg. Batrachium (52638)

**Jan Prančl**, Zdeněk Kaplan, Petr Koutecký, Pavel Trávníček.

*Institution of Botany of the CAS; Department of Botany, Faculty of Science, University of South Bohemia.*

Water crowfoots (Ranunculus subg. Batrachium) can be considered as a perfect model group for the study of polyploidy and hybridization in aquatic environment, comprising of several polyploid lineages of different origin, showing frequent intraspecific ploidy variation, frequently hybridizing across ploidy levels, and also possessing specific morfological adaptations reflecting the evolution (e.g. heterophyly). On the other hand, they belong between the most intricate European angiosperms. We applied flow cytometry on the extensive dataset (ca. 2700 Batrachium individuals from >600 populations) from the Czech Republic (for the detailed point of view), 23 other European and 3 non-European countries (for the broad point of view). Flow cytometry was proved to be an efficient method for determination of Batrachium species and also revealed numerous cases of recent hybridization, including introgression. Preliminary results of direct sequencing (ITS, cpDNA) has already revealed a number of remarkable findings, such as parallel evolution of traits in unrelated taxa and intraspecific cryptic variation. A gene flow between large-flowered and small-flowered species is probably unequal; we detected „chloroplast capture“ phenomenon. The situation is particularly complex in running waters, where even sterile hybrids are fixed via clonal propagation.
# Deep genetic splits in a diploid-polyploid Arabidopsis arenosa species complex (52642)

**Filip Kolar**, Eliska Zaveska, Gabriela Fuxova, Lucie Hyklova, Magdalena Lucanova, Atsushi J. Nagano, Hiroshi Kudoh, Stanislav Spaniel, Karol Marhold.

*Natural History Museum, University of Oslo; Institute of Botany, University of Innsbruck; Department of Botany, Faculty of Science, Charles University in Prague; Institute of Botany, Academy of Sciences of the Czech Republic, Průhonice; Center for Ecological Research, Kyoto University; Institute of Botany, Slovak Academy of Sciences, Bratislava.*

Knowledge on eco-geographical distribution of genetic variation within and across ploidy levels is a necessary prerequisite for any experimental or molecular study of plant systems that underwent genome duplication. The Arabidopsis arenosa group, an intricate diploid-autotetraploid complex, still remains largely neglected with respect to the evolutionary history and distribution of genetic variation across its range (spanning through most of the Europe). We conducted a large population-level sampling across its whole range and evaluated patterns of cytotype and genetic diversity of approx. 140 populations of A. arenosa using flow cytometry, RAD sequencing and nuclear microsatellites. We detected a largely parapatric distribution of the diploid (SE Europe) and tetraploid cytotype (NW Europe) that form two contact zones – narrow one in Slovenia and a wide mosaic-like one in South-Eastern and Western Carpathians. The diploid populations split into five genetically very distinct groups that are geographically segregated but do not reflect the most obvious morphological differentiation between lowland vs. alpine areas. The presence of marked ploidy-level, morphological, ecological and hitherto largely neglected genetic variation highlights A. arenosa as very promising model for addressing various evolutionary questions on origins and consequences of plant genome duplication.

Poster session B - POL 300

# A haploid fungus that appears like a polyploid. (52697)

**Pawel Rosikiewicz**, Tania Wyss, Frederic Masclaux, Ian Sanders.

*University of Lausanne.*

Rhizophagus irregularis is an arbuscular mycorrhizal fungus (AMF) used commercially to increase yields of crop plants, such as rice and wheat. Despite its potential to increase food production, AMF genetics and genomics has not been well studied. This is partially due to their unusual genetic structure. Each individual AMF carries multiple genetically different haploid nuclei. Therefore, we have to consider AMF as similar to polyploids. Different nuclei are transferred to the offspring in multinucleate spores. Moreover, different AMF can fuse, giving rise to progeny that are genetically and phenotypically distinct from the parents. I will present how I produced new crossed offspring of genetically different parental AMF. These crossed lines were genotyped with next generation sequencing (NGS). Then, NGS data were analyzed with the combination of methods typical for analysis of genetic data from polyploid organisms and cancer cell lines. My results showed that a process of nuclear exchange in AMF was not random and one genome was always dominant. The non-random
nature of AMF genetic exchange presents an opportunity to predict future genotypes of AMF crosses and produce novel AMF with known benefits to plant hosts.

# Growth differences between diploid and tetraploid cytotypes in a mixed ploidy population of the Patagonian steppe plant Hypochaeris incana (Asteraceae) (52711)

Karin Tremetsberger, Estrella Urtubey, Katharina Lapin, Christiane König.

*Institute of Botany, BOKU, Vienna, Austria; Instituto de Botánica Darwinion, San Isidro, Argentina; Division of Systematic and Evolutionary Botany, University of Vienna, Austria.*

Hypochaeris incana grows in the Patagonian steppe of southern South America. It is a self-incompatible, perennial herb with leaves in rosettes. A phylogeographic study suggests that H. incana reacted to Pleistocene climate changes through in-situ persistence. Today, tetraploid northern populations are thriving in isolation from each other, whereas diploid southern populations have recently expanded. Moreover, the data suggest that diploids have repeatedly been replaced by tetraploids in the northern Patagonian region. Here, we evaluate vegetative and reproductive growth parameters of diploids and tetraploids in a mixed ploidy population in northern Patagonia. For competitive exclusion to occur, we expect that - in the absence of habitat segregation - tetraploids are vegetatively and/or reproductively more vigorous than diploids. Whereas diploids do better with respect to growth (number of rosettes) and reproduction (number of ovules/head, fruit set), tetraploids have the ability to form large clones via underground stolons, with genets up to 90 m distant, a feature not encountered in diploids. Moreover, tetraploids might have better ability to withstand drought and show delayed phenological development in comparison to diploids. Thus, altered growth parameters of tetraploids compared to their diploid progenitors could ultimately lead to the replacement of diploids in local populations.

# Phylogenetic and biogeographical patterns of allopolyploid speciation in an intertidal fucoid seaweed assemblage (52902)

Joao Neiva, Licinia Gouveia, Cristina Paulino, Nelson Coelho, Gareth Pearson, Ester Serrao.

Algarve University.

The development and application of new molecular tools has contributed to a renewed interest on the roles of hybridization and polyploidy in evolution. These processes are associated in allopolyploid speciation, best known from plants and a few vertebrate lineages. New models beyond traditional plant systems may provide new insights underlying the factors contributing to allopolyploid establishment and evolutionary success. We investigated the evolutionary history of the intertidal seaweed Hesperophycus/Pelvetiopsis complex by screening a diverse
panel of populations of Hesperophycus californicus and Pelvetiopsis limitata senso lato for a
range of different nuclear/organelle sequencing/typing markers. Four well defined species
were recovered, including the taxonomically dubious P. arborescens and a new undescribed
species. Gene paraphyly, cyto-nuclear conflict, fixed heterozygosity and allele states
supported the allopolyploid nature of two of these entities and the male contribution of P.
arborescens for both. This latter species exhibited an extremely reduced geographical
distribution that did not overlap with the most recent (and possibly still expanding)
allopolyploid taxa. Conversely, the older allopolyploid is presently the most widespread taxa,
and its maternal ancestor is presumed extinct. Our results provide evidence for ecological
divergence allowing allopolyploids to locally co-exist with diploid parental species, and show
that in the long-term they can even outperform them.

Poster session B - POL 300

The evolution of the ability to learn (52092)

Machteld Verzijden, Jessica Abbott, Volker Loeschcke.

Aarhus University; Lund University.

The ability to learn is important because it allows animals to cope with unpredictable environments. Animals can learn from experience to guide their future actions, and do so in many contexts, such as food finding, nesting and choosing a mate. Species differ in their ability to learn, which indicates that this trait can evolve. However, we have a very limited understanding of how learning can evolve, and lack knowledge in particular, for learning in the context of mate choice. I will present data from an on-going study addressing how the ability to learn to choose a mate can evolve, by using laboratory populations of a fruit fly species as a model to mimic evolution experimentally. In particular, in this study males are selected on their ability to learn to associate female virginity with an arbitrary trait.

Poster session B - POL 300

The regulation of evolvability through Hsp90 expression (52097)

Robert Peuss, Hendrik Eggert, Sophie A.O. Armitage, **Joachim Kurtz**.

University of Münster, Institute for Evolution and Biodiversity.

The balance between robustness and evolvability is a long-standing question in evolution. It has been suggested that heat shock protein 90 (Hsp90), a chaperone, serves as a capacitor for evolution, since reduced availability of Hsp90 releases cryptic genetic variation. Hsp90 down-regulation might moreover serve to increase novel genetic variation, since it also is involved in transposon silencing in the germline. We addressed the question of whether Hsp90 expression, and thus evolvability, might be regulated upon demand. Specifically, we tested whether Hsp90 shows reduced transcription under stressful environmental conditions where increased variation, leading to increased evolvability, might have fitness advantages. We
simulated such a situation by placing untreated (focal) Tribolium castaneum flour beetles together with wounded conspecifics. This treatment exposed the focal beetles to an environmental condition that indicates a stressful environment without directly stressing the focal individuals themselves, which might have unwanted direct effects on their stress response. Using RT-qPCR, we found a consistent reduction in the Hsp90 expression of focal beetles. Our study for the first time finds indirect down-regulation of a key gene involved in evolvability, and poses the exciting question of whether evolvability might be regulated through the use of information derived from the social environment.

## The effect of worker mediated environmental stress on colony fitness in common black ant Formica fusca (52125)

Siiri Fuchs, Liselotte Sundström, Dalial Freitak.

University of Helsinki, Finland; Tvärminne Zoological Station, University of Helsinki, Finland; Centre of Excellence in Biological Interactions, University of Jyväskylä, Finland.

Organisms need to adapt to environmental changes, such as variation in pathogen pressure, in order to survive. Adaptation can occur by developmental plasticity during the life of an individual or via inherited traits across generations. Adaptation entails alterations in life-history strategies and, hence, trade-offs between essential functions such as growth, disease resistance, and reproduction. Social insects, such as ants, form colonies with high population density, which favours the spread of pathogens. To fend off epidemics social insects have developed behaviours that protect them collectively, among others the spread of immune factors by a mechanism similar to vaccination. We investigated how the common black ant Formica fusca workers deal with environmental stress (bacterium and oxidative stress) and how this affects the brood they help to raise. Our study shows that workers mediate stress, which can be seen as changes in brood development, survival and size. We also see differences in gene expression depending on diet, caste and age. This study shows how organisms respond to varying environmental conditions and use different strategies to adapt and gain fitness.

## Effects of nest microclimate on maternal effects in a small passerine bird (52264)

Josefa Bleu, Claudy Haussy, Clotilde Biard.

Institute of Ecology and Environmental Sciences - Paris.

Maternal effects are a source of phenotypic plasticity. They can have extremely large effects, affecting the morphology, physiology and/or behaviour of the offspring. In particular, they allow a plasticity that may play an important role in shaping responses to rapid environmental
changes, such as those generated by climate change. Yet, maternal effects are seldom studied in the context of the adaptation to climate change. Previous studies have shown the sensitivity of small passerine birds to climate change in Europe. These effects may be driven by changes in pre-natal maternal investment, as it is crucial for offspring development and success. The aim of this study is to test this hypothesis by investigating how pre-natal maternal effects change in response to nest micro-temperature during egg laying. More precisely, we will compare reproductive investment between control and experimentally heated nests in a great tit population. In particular, clutch size, hatching and fledging success and egg characteristics such as egg mass, lysozyme and carotenoid content will be compared. These results will allow a better understanding of the mechanisms involved in the response to global change which is primordial to better predict responses of natural populations.

# Inconsistency of whole genome transcription responses to diet in Drosophila (52292)

**Jelle Zandveld**, Joost van den Heuvel, Erik van den Akker, Bas Zwaan.

*Wageningen University; Newcastle University; Leiden University Medical Centre; Technische Universiteit Delft.*

In Drosophila, the response of whole genome transcription to diet is extremely plastic. On dietary restriction, a reduced food intake without malnutrition that extends lifespan and reduces fecundity, many whole genome transcription studies are performed. We performed a meta-analysis between studies using a highly similar protocol to test the consistency of the transcriptome responses to dietary restriction. Striking inconsistent patterns between studies were found for the up/down regulation on the gene level, the associated GO-terms and gene expression top-hits. The addition of another dataset that includes egg laying data suggests that differential egg laying responses with age may account for these inconsistencies. We recommend that availability of phenotypic data is essential for the right interpretation of an organism's whole genome transcription pattern to diet in particular, but potentially to many environmental interventions in general.

# Trans-generational effects of Diuron herbicide on the methylome of the Pacific Oyster Crassostrea gigas (52314)

**Rodolfo Rondon**, Evelyne Bachère, Farida Akcha, Guillaume Mitta, Caroline Montagnani, Céline Cosseau, Christoph Grunau.

*Laboratoire d'Interaction Hôte Pathogènes Environnement (IHPE), CNRS, UMR5244; Centre Atlantique, Laboratoire Ecotoxicologie.*
Environmentally induced epigenetic transgenerational effects involve the transmission of phenotypic changes to the offspring. It is well established that various environmental stressors including chemical pollutants can modulate the epigenome of individuals. The pacific Oyster Crassostrea gigas is the main oyster species exploited worldwide. It currently suffers from devastating summer mortality outbreaks. It is well established that this mass mortalities are the result of a complex pathosystem including both biotic and abiotic factors among which toxic chemicals are major actors. It has been previously demonstrated that the parental exposure of C. gigas to Diuron has consequences on the phenotypes of the offspring. Altogether, these data suggest that Diuron could act on the epigenome of the parental germ cells and subsequently impact the transcriptionnal profile and phenotype of the subsequent generation. To examine these questions, we established the transcriptional profile and the methylome of C. gigas offspring of diuron exposed genitors. We provide evidence that differences exist between spats produced from exposed or non exposed parents both in the transcriptome and methylome. These differences occur in specific loci and we will discuss the molecular mechanisms that link methylome modifications to changes in life history traits of C. gigas.

# Parasite-induced changes in host phenotype and gene expression after infection (52319)

**Barbara Feldmeyer**, Johanna Mazur, Hannes Lerp, Harald Binder, Susanne Foitzik.

Mainz University; University Medical Center Mainz.

Parasite infections, next to their detrimental consequence for host fitness, can influence host organisms on several levels from physiology, morphology, and life history to behavior. Parasite-induced changes in the host can either be attributed to host defenses, they can be by-products of infection, or the result of parasite manipulation. In social insects, not only the infected individual itself, but the whole colony is affected by the parasites presence and has to suffer the consequences as community. In the ant Temnothorax nylanderi, individuals parasitized by a cestode, show a suite of phenotypic changes, including smaller body size, decreased activity and increased survival. Moreover, parasitized colonies show lower aggression levels than healthy colonies. To understand the proximate mechanisms of parasite-induced behavioral changes, we compared brain gene expression patterns between workers from infected and healthy colonies. We report on candidate genes and associated functional categories involved in the parasite-induced phenotypic alterations. The most striking finding is a large scale down regulation of muscle-related genes in infected workers, fitting well to their inactive phenotype and increased survival. These altered host phenotypes may in turn benefit the parasite through an increased transmission probability to its final host.

# Maternally-derived thyroid hormones in avian eggs: variation and function (52362)
**Suvi Ruuskanen**, Ton G.G. Groothuis, Veerle M. Darras, Sonja Schaper, Marcel E. Visser.

*University of Turku; University of Groningen; KU Leuven; Netherlands Institute of Ecology.*

Maternal effects via eggs are a powerful way to influence offspring phenotype and fitness, thus strong selection is expected. For example, avian maternal steroid hormone levels in egg yolk vary systematically with environmental factors, are heritable and affect offspring fitness-related traits. However although data from mammals and fish suggests a prominent role for maternal thyroid hormones (TH) on offspring growth and survival, sources of avian egg TH variation and effects on offspring have so far been largely neglected in ecological studies. We studied variation in yolk TH concentrations (thyroxine, T4 and triiodothyronine, T3) in wild great tits (Parus major) and estimated heritability using siblings in captivity. We further experimentally elevated yolk TH in a wild population to study effects on fitness-related traits. Yolk T4 concentration correlated negatively with ambient temperature and positively with laydate. There was significant variation in yolk TH among females. Yolk T4 increased with layorder within clutches. However, experimental yolk TH elevation had no effect on skeletal or body mass growth, metabolic rate or hatching or fledging success. To understand more of the potential function of maternally-derived TH, an experimental yolk TH reduction should be conducted and effects of yolk TH on pre-hatching development should be studied.

# Adaptive Integrative Shape Variations in Phenotypic Plasticity (52431)

**Kinya Nishimura.**

*Graduate School of Fisheries Sciences, Hokka.*

Stability and plasticity are antithesis essential ability of organism to cope with the environments. “Phenotypic plasticity” is an important conceptual issue in evolutionary biology. Generally, even though morphogenesis makes a stabilized constitutive morphology, some organisms were gifted morphological plasticity with cell proliferation and tissue restructuring in various environmental circumstances.

The Hokkaido salamander (Hynobius retardatus) is in the list of organisms having ability of phenotypic plasticity. Larvae of H. retardatus change morphology to deal with predators and prey. The emerging morphologies can be classified into some distinctive morph categories establishing the polyphenism. We can perceive differences among typical individuals belonging in each category. However, discrimination by perception is dubious, and application of a conventional criterion is also unreliable. Previous studies of the morphological plasticity cannot touch on various important biological issues such as individual variation within each morph category, integrative shape variations and the natural selection acting on the variations.

I adopted the geometric morphometric analysis to the larvae of H. retardatus that changed morphological development in experimentally manipulated trophic interaction environmental settings. I clarified adaptive integrative shape variations among individuals within each morph category that suggests developmental shape module integration evolved by natural selection.
# Negative public information in mate-choice copying helps the spread of a novel trait (52462)

**Susana Varela**, Mauro Santos, Margarida Matos.

CE3C - Faculdade de Ciências, Universidade de Lisboa; GBE - Universitat Autonòma de Barcelona, Spain.

Numerous field and laboratory experiments have shown that many species have the capacity for social learning, including mate-choice decisions that can be influenced by witnessing the mating decisions of others. This phenomenon, known as mate-choice copying, is a type of non-genetic information that can increase the behavioural plasticity of the copying individuals, and cause genetic changes across generations in the genotype of the copied individuals. Here we develop a numerical model of mate-choice copying that follows the population genetics tradition, consisting in tracking allele frequencies in a population over time under scenarios with and without copying. In contrast to previous evolutionary models, we consider both positive and negative social information because many mating systems are driven by males in pursuit of a mate, and female refusal of copulation may provide negative social information. The inclusion of negative social information to mate-choice copying helps the spread of a novel trait, even if female innate mate-choice preference is biased towards the common male-type. We argue that the presence or absence of copying might simply mirror the associated cost-benefit relationship of the mating system of a given species, and suggest how to test this prediction.

# Memory and learning in gene-regulation networks (52464)

**Kostas Kouvaris**, Jeff Clune, Loizos Kounios, Markus Brede, Richard Watson.

University of Southampton; University of Wyoming.

The ability of a single genotype to control the development of multiple distinct phenotypes, e.g. differentiation of cell types or multicellular dimorphism, is important for understanding the environmental sensitivity of development, epigenetic inheritance of phenotype states, phenotypic variability under mutation and the evolution of evolvability. Recent work (Pavlicev et al. 2010, Watson et al. 2014) shows that the constraints and biases of development, e.g. as controlled by the structure and organisation of gene regulation networks, can be shaped by natural selection to 'record' the pattern of functional dependencies imposed by past selection, as per Rupert Reidl's suggestion (1978). This enables a single gene-network (given random perturbations to embryonic expression levels) to (re)produce multiple distinct phenotypes that have been selected for in past environments. Moreover, we show that principles of memory, learning and generalisation, well understood in the context of connectionist (neural network) models of memory and learning, can be used to characterise the capabilities and limitations of such 'developmental memory' in gene networks. We also demonstrate how conditions that avoid 'over-fitting' in machine learning, such as the
application of a connection cost that reduces model complexity, can be used to predict conditions that enhance phenotypic flexibility in evolved gene networks.

# The role of nucleic acid methylating enzymes in the red flour beetle Tribolium castaneum (52484)

**Nora Stolte**, Hendrik Eggert, Julia Ebeling, Maike Diddens-de Buhr, Frank Lyko, Joachim Kurtz.

*Institute of Evolution and Biodiversity, Westfälische Wilhelms-Universität Münster; German Cancer Research Centre, Heidelberg.*

Epigenetic regulation enables rapid phenotypic adaptation of organisms to environmental changes. Cytosine methylation of DNA and certain RNAs is an important epigenetic mechanism, which is generated by an evolutionarily conserved family of enzymes, so called DNA methyltransferases (DNMTs). The ancestral DNMT toolkit consists of three enzymes DNMT1, 2 and 3, however, the evolution of cytosine methylation systems has generated great variation in the sets of Dnmt genes between species. For example, while Drosophila lacks a functional DNA methylation system, honeybees possess all three DNMTs, and DNA methylation is crucial for caste determination.

We investigated the status of a functional DNA methylation system in the Red Flour Beetle Tribolium castaneum, which possesses Dnmt1 and Dnmt2 but lacks Dnmt3. In the present study, we show that DNMT1 is essential for early embryonic development, although we were not able to detect functional levels of DNA methylation via whole genome bisulfit sequencing. Parental RNAi knock-down of Dnmt1 in the mothers caused a developmental arrest in the offspring embryos.

This, to our knowledge, first evidence of a functional role of DNMT1 during embryogenesis in T. castaneum, calls for further studies investigating additional functions of this enzyme and its role beyond DNA methylation.

# Ephemeral vs persistent triploids: two epigenetic modus operandi (52522)

**France Beauregard**, Christelle Leung, Bernard Angers.

*Université de Montréal.*

Phenotypic variation is generally a prerequisite to occupy different niches. Clones can only rely on epigenetic processes to regulate the expression of their genes and produce different phenotypes. Consequently, allopolyploids are benefited by a wider range of alleles they can express for a given gene. Multiple clonal lineages with different ploidy levels are found in gynogenetic hybrid complexes Ambystoma spp. and Chrosomus eos-neogaeus, both widespread in North America. In Chrosomus fish, only diploid reproduce clonally while
triploids are constantly formed de novo. In contrast, triploids not only perpetuate clonally in
Ambystoma salamanders, but represent the most common biotype. The objective of this
project is to compare the effects of additional haplome on the epigenetic variation in these
hybrid complexes. When genetic and environmental effects are taken into account, the third
genome has little effect on the epigenetic profiles in fish Chrosomus eos-neogaeus while it is
more important than the environmental effect for salamanders Ambystoma. These results
suggest that the additional genome may provide a niche diversification from a unique diploid
hybrid in the Ambystoma complex.

Life history consequences of dietary mismatches in Drosophila melanogaster
(52550)

Elizabeth Duxbury, Tracey Chapman.

University of East Anglia.

Changes in nutritional environment can lead to evolved and plastic responses in life history
traits such as lifespan and reproductive rate. Using Drosophila melanogaster, we first
investigated the effect on life history traits of changes in the quality (high or low protein
content) of developmental (larval) diet versus adult diets within a single generation. We
compared the fitness of individuals maintained on mismatched diets (i.e. low quality larval
followed by high quality adult diet, or vice versa) with those on constant diets (low or high
quality larval and adult diets throughout). We measured larval traits, age-specific survival and
age-specific reproductive output. The results showed that, contrary to predictions from
‘Thrifty Phenotype’ theory, individuals maintained on mismatched diets did not suffer fitness
costs in comparison to those maintained on constant diets. We next investigated evolutionary
responses of life histories to diet manipulations, using replicated populations experimentally
evolved on different dietary regimes. The evolutionary treatments were populations that had
an evolutionary history of food supplied at either regular or unpredictable intervals (the latter
creating periods of feast and famine). I discuss the results in the context of the evolution of
lifespan, of responses to dietary restriction and of life history trade-offs.

Plasticity and evolutionary divergence in ovariole number in Drosophila
(52563)

Cláudia Mendes, Élio Sucena, Christen Mirth.

Instituto Gulbenkian de Ciência, Oeiras, Portugal; Universidade de Lisboa, Faculdade de Ciências, Departamento de Biologia Animal, Lisboa, Portugal.

Developmental plasticity, the ability of an organism to adjust its developmental trajectory in
response to environmental variation, may facilitate success in novel environments and
promote evolutionary diversification. Here, we sought to understand the contribution of developmental plasticity to the evolution of morphological diversity by comparing the underlying developmental mechanisms of the plastic response and of the evolutionary divergence in ovariole number, an important reproductive trait in Drosophila. Our analysis in Drosophila melanogaster revealed that ovariole number is severely reduced when early third instar larvae are malnourished. This reduction in ovariole number is a result of a slower rate of ovary growth and a delayed onset of terminal filament (TF) differentiation, the first step in ovariole morphogenesis. Interestingly, we found that similar developmental mechanisms are likely responsible for the considerable differences in ovariole number between two geographical races of Drosophila mojavensis. These desert-adapted populations utilize distinct cacti for all stages of their life cycles. Thus, our results suggest that environmental and genetic variation can affect ovariole number through similar developmental mechanisms, supporting the view that developmental plasticity may play an important role in the evolution of morphological diversity.

# Phenotypic plasticity and adaptive potential for climate change in mountain birch – analysis of mature siblings in tree-line common gardens (52573)

Irma Saloniemi, Marjo Helander, Kari Saikkonen.

University of Turku; University of Turku; The Natural Resources Institute Finland.

Simultaneous studies of genetic variation and plasticity are demanding in natural populations, especially in long-lived organisms like trees. Mountain birch (B. pubescens spp. czerepanovii) is a tree-line species that occurs above the Arctic Circle in Northern Scandinavia, but previous studies suggest that it has high level of phenotypic and genotypic variation in fitness and morphology related traits. Our aim is to estimate the adaptive potential for the species as temperatures increase in high latitudes. We base our study on 25-40 years old siblings that grow in 4 common gardens in natural, but locally variable northern environments, and one garden in southern Finland. Each of the twenty sib-families originates from seeds of a single tree of northern origin. Each family is divided between the 3-5 gardens, and the gardens are divided in 3-6 blocks with 16 progeny per family in each of the blocks. Performance of these tree families was intensively monitored during the first decade of their growth, and we continue the measurements on the adult trees. Phenotypic plasticity may explain high variation, but it may hinder adaptation by increased phenotypic variation. On the other hand, it may also restrict too fast responses, or even accelerate adaptation

# Reproductive tactics and sperm competition related to oxidative stress in the Seba's short tailed bat, Carollia perspicillata. (52758)

Nicolas Fasel, Charlotte Wesseling, Fabrice Helfenstein, Heinz Richner.
Securing privileged access to mates is costly, and hence may lead to the evolution of alternative reproductive tactics (ARTs), such as sneaking copulations with lower pre-copulatory costs but higher sperm competition. It predicts a stronger investment by sneakers in sperm traits that increase fertilization efficiency. In order to investigate the role of oxidative stress as a potential mediator of different reproductive physiologies associated with ARTs, we assessed ejaculate and blood redox profile of male Seba's short tailed bat (Carollia perspicillata). We found a significant increase in oxidative solicitation (proportion of oxidized glutathione) in the blood of harem males, indicating a cost, without however leading to more damage (lipid peroxidation). Sperm quality (velocity and longevity) was significantly higher in bachelor males, but did not correlate with the ejaculate redox profile. The important ingestion of dietary antioxidant may efficiently protect both somatic and reproductive functions in this frugivorous bat species, since an important endogenous antioxidant (superoxide dismutase) did not correlate with social status or sperm competition level. It suggests that a privileged access to mates implies costs for harem holders, and intense sperm competition for bachelors leads to adaptions in sperm traits.

Context-dependent plastic response to the presence of predator cues in a
widespread newt species (52844)

Zoltán Tóth.

Lendület Evolutionary Ecology Research Group, Plant Protection Institute, HAS.

Previous research on phenotypic plasticity mostly focused on developmental plasticity (i.e. different developmental trajectories triggered by environmental cues) in amphibian model systems, so we have limited information about how anticipatory parental behaviours and maternal effects interact and jointly contribute to individuals’ reproductive success. In this study I examined behavioural and maternal responses of gravid female smooth newts, Lissotriton vulgaris, to the presence of chemical cues originating from invertebrate predators. I found no indication for substantial oviposition preference among individuals, as many of them deposited eggs both into the ‘predator-cue’ and the ‘clear-water’ environment. However, these latter females were also found to adjust the size of the laid eggs to the perceived environment: egg size decreased with time during the reproductive season in the absence of predator cues, but not when such cues were present. Besides, females differed in the extent of egg-wrapping between ponds of origin, but individuals did not adjust their wrapping behaviour to the perceived environment. According to these results, female smooth newts have the potential to exhibit activational plasticity in the form of a maternal effect when chemical cues from predators are present, with which individuals may improve their offspring’s fitness in risky natal environments.
**Environment modifies equilibrium proportions of alternative reproductive strategies in the bulb mite. (52850)**

**Anna Maria Skrzynecka**, Jacek Radwan.

Jagiellonian University, Institute of Environmental Sciences; Adam Mickiewicz University,
Institute of Environmental Biology(Poznan,PL).

The bulb mite (Rhizoglyphus robini) expresses alternative reproductive phenotypes (ARPs). Aggressive fighters, possessing modified pair of legs used as a weapon, may coexist in populations with unarmed, benign scramblers. Male morph in this species is heritable, but given no evidence of frequency-dependence in earlier work on this system, it is not clear what maintains genetic variance underlying ARPs. If the polymorphism is balanced, proportion of fighters should come back to equilibrium value after being disturbed. We used artificial selection to alter fighters' proportions to 50% or 94%, and then for 14 generations we traced how the proportion changes in 5 replicate populations. To investigate if equilibrium point is affected by environmental conditions, we ran this protocols under two temperatures: 24 and 18°C. We found that under both temperatures, proportions evolved to stable values regardless of initial proportion of morphs, indicating that the polymorphism is balanced. However, the equilibrium proportion differed significantly between the temperatures (76% at 24°C, and 98% at 18°C), indicating that morph success can be affected by environment. Finally, we found that fighters' proportion was also affected by maternal effects. Overall, the proportion of morphs seems to be shaped by interaction between historical and current environmental state.

**Epigenetic modifications in moth sexual communication signals (52903)**

**Rik Lievers**, Peter Kuperus, Dennis van Veldhuizen, Astrid T. Groot.

IBED.

The evolutionary diversification of moth sexual communication signals is puzzling as it is believed to be under stabilizing selection, which limits evolutionary change. Yet, several moth species show significant intraspecific variation in the pheromone composition. The observed variation can in large part be explained by genetic variation, however, the sexual communication signal has also been shown to by plastic1. Moth sexual signals can be modulated depending on the prevailing olfactory cues present in the environment. This suggests that behavioral differentiation may precede genetic differentiation in the sexual signals of moths. The molecular mechanism behind this phenotypic plasticity is poorly understood. Epigenetic modifications, such as DNA methylation, are an important candidate. We investigate whether DNA methylation is involved in regulating heritable and plastic variation in the sexual communication signals of noctuid moths. Our first results show that females indeed vary in their methylation patterns, that these methylation patterns seem heritable, and that DNA methylation could potentially contribute to plasticity in sex pheromone composition.
Seasonality maintains alternative life history phenotypes (51633)

Sami Kivelä, Panu Välimäki, Karl Gotthard.

Stockholm University; University of Oulu.

Many organisms express discrete alternative phenotypes (polyphenisms) in relation to predictable environmental variation across generations, seasonal polyphenisms in insect morphology being well-known examples of this. However, the evolution of alternative life-history phenotypes, which are also common, remains poorly understood. We analyse the evolution of alternative life histories in seasonal environments by using temperate insects as a model system. Temperate insects provide an excellent system to study alternative life history phenotypes as they typically express alternative developmental pathways of diapause and direct development that have life-history correlates. We develop a methodologically novel and holistic simulation model and optimize development time, growth rate, body size, reproductive effort, and adult life span simultaneously in both developmental pathways. The model predicts that direct development should be associated with shorter development time (duration of growth) and adult life span, higher growth rate and reproductive effort, smaller body size as well as lower fecundity compared to the diapause pathway, because the two generations divide the available time unequally. These predictions are consistent with many empirical data. In conclusion, the analysis shows that seasonality alone can explain the evolution of alternative life histories.

Live fast, die young? – Trans-generational effects of parental food stress in a
butterfly (51665)

Luisa Woestmann, Marjo Saastamoinen.

Metapopulation Research Group, University of Helsinki.

Environmental conditions experienced by parents are potentially important factors that shape the offspring's phenotype and are known as trans-generational effects. Poor environmental conditions often have negative effects on offspring via low provision of resources to the eggs, resulting in poor quality offspring with reduced fitness. Maternal effects are well-studied in many organisms, whereas paternal effects have received less attention. There are, however, important effects driven also by the father. Male butterflies, for example, transfer spermatophores during mating that comprise sperm and accessory gland secretions, which contain nutrients that are contributed to the female and can increase her fecundity and life span. In this study, we investigated the effect of food deprivation of males and females during
the reproductive stage in the Glanville fritillary butterfly (Melitaea cinxia) on life-history traits including fecundity as well as possible trans-generational effects by both parents. Food deprivation of mothers resulted in a direct effect on female reproduction leading to reduced lifetime reproductive success, whereas stressed fathers sired more offspring, possibly due to higher investment into their spermatophores.

# Grandparental immune priming in the sex-role reversed pipefish Syngnathus typhle (51849)

Anne Beemelmanns, Olivia Roth.

Helmholtz Centre for Ocean Research Kiel (GEOMAR).

Trans-generational immune priming (TGIP), the transfer of non-genetic information about parental pathogenic experience to the progeny boosts the offspring immune response and permits a more efficient reaction against parasite genotypes. While TGIP was discussed as a short-term effect, selection could favor TGIP that goes across generations in case the parasite assemblage is stable and costs are limited.

We experimentally assessed the evolution of grandparental-TGIP in the sex-role reversed pipefish Syngnathus typhle on life history and gene-expression of F2-offspring.

We found strong grandparental effects with additive, sex-specific contribution to different pathways of the immune system and clear involvement of epigenetic effects. While genes of the adaptive immune system and complement component system were affected by grandmaternal effects, genes of the innate immune system revealed grandpaternal effects.

DNA-methylation and especially histone-acetylation genes were influenced by the grandfathers demonstrating that epigenetic regulation mechanisms are essential for transferring non-genetic information over more than one generation down the paternal line.

While adaptive if similar parasitic assemblage is met, grandparental-TGIP involves costs displayed as reduced growth of F2-offspring.

We conclude that the transfer of phenotypic experiences does not cease after one generation: additive grandparental effects may seriously affect the phenotype of their grandchildren.

# An evolutionary analysis of dizygotic twinning in humans (51914)

Richard Smock, **Wade Hazel**, Joseph Tomkins.

DePauw University; DePauw University; University of Western Australia.

Dizygotic twinning results from double ovulation and is a heritable trait that occurs in all human populations at low, but variable rates. Females producing twins typically are older, taller, have larger body mass indices and higher lifetime reproductive success, relative to females that do not produce twins. We develop an analytical model that compares the fitness
of four reproductive strategies: (1) never producing twins, (2) producing twins at a low but constant rate, (3) producing twins conditionally at a rate dependent on age and quality, and (4) double ovulating conditionally, but carrying only a single offspring to term. When we parameterized the model using estimates of demographic variables from a natural population, we found that when either egg quality declines with female age, or females vary in their optimal interbirth interval, the strategy with the greatest fitness is conditional double ovulation, but carrying only a single offspring to term. These results suggest that dizygotic twinning is a byproduct of adaptive phenotypic plasticity in ovulation rate.

# Genotype and phenotype changes over generations on a multinucleated fungus (52016)

**Ivan Mateus, Fred Masclaux, Tania Wyss, Ian Sanders.**

*University of Lausanne.*

Arbuscular mycorrhizal fungi (AMF) are one of the most important endo-symbionts of plants. They provide mineral nutrients to the plant in exchange of photosynthesized carbon. These fungi have been considered as ancient asexual organisms that reproduce clonally by producing multinucleate spores. Recently whole-genome sequencing of in-vitro cultures of the model AMF species Rhizophagus irregularis revealed little genetic polymorphism among coexisting nuclei. However, single-spore cultures segregated from parental cultures differentially alter rice growth, suggesting significant polymorphism between the single-spore cultures.

In addition R. irregularis present hyphae without divisions. This feature has as consequence that nuclei coexist within the same hyphae. From an evolutionary point of view, this feature challenges the definition of an individual. Therefore, we studied how the genetic variation is partitioned among different spores over different generations and its consequences on the phenotype of the descendants. The results of this study 1) give more insights about how an asexual organism could exist for million of years without a sexual cycle and 2) help to understand at what level selection acts on R. irregularis.

# Predator-induced plasticity of tadpoles: a search for developmental windows (52040)

**Günter Gollmann, Barbara Fischer, Christine Mayer, Philipp Mitteroecker.**

*University of Vienna, Department of Theoretical Biology; University of Oslo, Department of Biosciences.*
Many organisms vary in their plasticity over their lifetime and the induction of plastic traits is sometimes limited to certain developmental windows of an organism's lifetime. In a mesocosm experiment with tadpoles of the agile frog, Rana dalmatina, we varied the timing of introduction of caged dragonfly larvae. The tadpoles were photographed in lateral view at weekly intervals, to study growth as well as shape variation assessed by geometric morphometrics. In the presence of predators, tadpoles reduced growth and developed higher tail fins. These effects on growth rates and body shape were stronger in tadpoles first confronted with predators in the second week, than in those exposed to caged predators from the onset of the experiment (a few days after hatching). Our results indicate that there exist windows of predator-induced plasticity in Rana dalmatina tadpoles.

# Evolutionary medicine and intrinsically vulnerable facultative adaptations (52044)

Randolph Nesse.

*Arizona State University.*

A major focus for evolutionary medicine has been how natural selection shapes mechanisms that regulate facultative responses, especially defensive responses such as vomiting or the fight-flight response. A signal detection analysis reveals that optimal systems give rise to many false alarms, a phenomenon described as the Smoke Detector Principle. However, some systems enter runaway positive feedback loops, such as in panic disorder. This presentation offers mathematical models describing the characteristics of systems that are intrinsically vulnerable because their thresholds adjust adaptively to increased sensitivity on repeated arousal. This phenomenon is likely to provide explanations for vulnerability to anxiety disorders and chronic pain. The tradeoffs involved are likely to constrain selection for mechanisms that adjust thresholds or response intensity as a function of experience in a particular environment.

# Annual and seasonal variability in leaf traits of Iris variegata genotypes growing in contrasting light conditions (52088)

Uros Zivkovic, Stevan Avramov, Natasa Barisic Klisaric, Danijela Miljkovic, Ljiljana Tubic, Danijela Misic, Branislav Siler, Aleksej Tarasjev.

*Institute for Biological Research "Sinisa Stankovic".*

During their evolution plants developed a set of mechanisms to adapt to the varying environmental conditions, resulting in diversified phenotypes. We were interested in the response of plants towards changing environmental parameters, to light in particular. For this study we selected genotypes of Iris variegata that experienced different light
conditions in their habitats: a) genotypes situated along the top and slope of a dune, where it coexisted with annual and perennial herbs and low shrubs; b) genotypes inhabited the woodland understories.

We grew I.variegata genotypes in two experimental light treatments (high and low) and tested
differences between spring, summer and autumn as well as between two successive years. The
plants responses were analysed at different levels of biological information and it was
revealed that dissimilar light conditions markedly affected all aspects of plant anatomy and
physiology. Repeated measures model ANOVA detected a significant effect of year, habitat
and genotype in most analysed traits. Models of changes during growing seasons were
treatment- and year-specific.

We also present results from an on-going study which aims to assess occurrence patterns of
phenotypic plasticity based on system-level analysis of the same samples that includes
metabolic profiling approach combined with robust statistical analysis.

Evolutionary and ecological patterns of global DNA cytosine methylation in angiosperm plants (51693)

**Monica Medrano**, Conchita Alonso, Ricardo Pérez, Azucena Canto, Víctor Parra-Tabla, Carlos M. Herrera.

Estación Biológica de Doñana, CSIC, Sevilla, Spain; Instituto de Investigaciones Químicas, CIC-Cartuja, CSIC-US, Sevilla, Spain; Centro de Investigación Científica de Yucatán, A.C., CONACYT, Mérida, México; Departamento de Ecología Tropical, Universidad Autónoma de Yucatán, Mérida, México.

DNA cytosine methylation is an ubiquitous epigenetic mechanism involved in many
important functions, including regulation of gene expression and maintenance of genomic
integrity. In plants, intraspecific variation in cytosine methylation has been related to genetic
and ecological factors. Genome-wide cytosine methylation is also known to vary substantially
between the few species that have been thoroughly studied, but little is known on interspecific
patterns, or the degree to which differences between species in methylation levels are related
to evolutionary history, ecological settings and/or biogeography. DNA methylation has been
implicated in plant macroevolution through effects on speciation and appearance of
evolutionary novelty, although these suggestions still lack compelling empirical support. We
will contribute to fill this gap by analyzing global DNA cytosine methylation levels obtained
by HPLC in a large sample of phylogenetically diverse angiosperms (ca. 100 families) from a
broad range of habitat types on two continents. The following three questions will be
addressed: (i) is interspecific variation in DNA methylation levels phylogenetically
structured?; (ii) are there discernible regional or ecological patterns in genome methylation
levels?; and (iii) what are the relative importances of phylogeny, ecology and biogeography in
determining plant community-wide levels of cytosine methylation?
# Fine-scale population epigenetic structure in relation to gastrointestinal parasite load in red grouse (Lagopus lagopus scotica) (51779)

**Marius Wenzel, Stuart Piertney.**

*University of Aberdeen; University of Aberdeen.*

Cytosine methylation patterns may be altered by environmental stresses and consequentially affect phenotypic plasticity and adaptation. Parasites are potent stressors, but there is little understanding of the epigenetic consequences of long-term parasite insult. Here, we estimate epigenetic diversity and differentiation among 21 red grouse (Lagopus lagopus scotica) populations in north-east Scotland and test for association of gastrointestinal parasite load (caecal nematode Trichostrongylus tenuis) with hepatic genome-wide and locus-specific methylation states assayed through methylation-sensitive AFLP. The populations differed significantly in genome-wide methylation levels and were also significantly epigenetically (FSC = 0.0227; P < 0.001) and genetically (FSC = 0.0058; P < 0.001) differentiated. However, parasite load was not associated with either genome-wide methylation levels or epigenetic differentiation. Instead, eight epilocus-specific methylation states were disproportionately differentiated and 35 epilocus-specific methylation states were significantly positively and negatively associated with parasite load. Following Sanger sequencing, genome mapping and GENEONTOLOGY (GO) annotation, some of these epiloci were linked to genes involved in regulation of cell cycle, signalling, metabolism, immune system and notably rRNA methylation, histone acetylation and small RNAs. These findings demonstrate a locus-specific epigenetic signature of parasite load in populations of a wild bird and suggest intriguing physiological effects of parasite-associated cytosine methylation.

Poster session B - POL 300

# Epigenetic signatures of parental effects in soil mites (51911)

**Stuart Piertney, Marianne Mugabo, Tim Benton.**

*University of Aberdeen; University of Leeds.*

Variation among individuals in their life-history phenotype is influenced by past environmental conditions communicated across generations via parental effects. The extent to which such parental effects influences epigenetic variation remains underexplored, despite offering a mechanism through which phenotypic plasticity can be generated.

Here we describe a study that examines interrelationships between parental effects and cytosine methylation in the soil mite Sancassania berlesei. Multiple microcosm populations of soil mite were established from a single natural population that differ in their parental environment by manipulating the age, quality and sex ratio of the adults that comprise the different breeding populations. The age and size at maturity, fecundity, egg size, egg protein content and survival of offspring was followed for the subsequent four generations. The patterns of genome-wide DNA methylation were assayed using a modified version of RAD-genotyping that involves parallel restrictions with methylation sensitive and insensitive
isoschizomers. Comparison of the levels of DNA methylation between different parental backgrounds provides estimates of the extent to which environment affects epigenetic signatures and how it is correlated with phenotype. Comparison between parental backgrounds highlights whether epigenetic variation is heritable. In both cases epialleles are examined using gene ontology and pathway analyses to highlight any functional linkages with phenotype.

# Contribution of epigenetic mechanisms to phenotypic plasticity in Neurospora crassa (52183)

Ilkka Kronholm, Jouni Laakso, Tarmo Ketola.

*University of Jyväskylä; University of Helsinki.*

Organisms respond to environmental changes by altering their development to produce a different phenotype and by maintaining physiological homeostasis on the face of fluctuating environments, a phenomenon called phenotypic plasticity. Plasticity can be achieved by regulation of gene expression. These gene expression patterns can also be directed and maintained by epigenetic mechanisms, chemical modifications of DNA or associated proteins.

Altering epigenetic patterns can have effects on phenotypic plasticity but few examples exists so far. Thus, the generality of this phenomenon remains unknown. We investigated to what extent epigenetic mechanisms contribute to phenotypic plasticity in the filamentous fungus Neurospora crassa. We used knockout mutants for different epigenetic mechanisms, such as DNA methylation, histone modifications, histone deacetylation, and RNA interference. To quantify phenotypic plasticity we measured reaction norms for the different mutants in different environments, such as NaCl stress, oxidative stress, pH and sugar concentration.

We show to what extent and which epigenetic mechanisms contribute to phenotypic plasticity in different environments in Neurospora. These results can resolve if epigenetic mechanisms are responsible for phenotypic plasticity and the ability to cope with adverse environmental conditions. Furthermore, they will help us to understand if and how epigenetics interacts with evolutionary adaptation via phenotypic plasticity.

# DNA methylation patterns, CpG o/e ratio and Non-Gaussian distributions : a pan-species study (52309)

Benoit Aliaga, Jan Bulla, David Duval, Christoph Grunau.

*Laboratoire Interactions Hôtes-Pathogènes-Environnements (IHPE) UMR5244 CNRS/University of Perpignan; University of Bergen.*
Epigenetic mechanisms contribute to generate phenotypic variability and could therefore be involved in evolution. The most studied of these mechanisms is DNA methylation i.e. the addition of a methyl group to cytosines leading to 5-methyl-cytosine (5mC). Methylation is predominantly (but not exclusively) located in CpG dinucleotides. Whereas in vertebrates and plants, DNA methylation is known to be involved in control of gene expression, its role in invertebrates is far less understood. Being one of the most analysed epigenetic information carriers does not mean that DNA methylation was exhaustively studied in many species. Its analysis remains expensive and needs expertise. Since 5mC is spontaneously deaminated to thymine, CpG underrepresentation can be used as a proxy to estimate 5mC levels and distribution patterns. Our work has consisted in predicting the methylation status in transcriptomes for 365 species from CpG observed to expected ratios (CpG o/e). We developed new statistical methods based on different Gaussian and non Gaussian distributions. Best results were obtained with a non Gaussian distribution for more than 80% of the studied species. We will present this new method that allows prediction of DNA methylation using mRNA data alone.

# Impact of coastal pollution on epigenetics and reproductive fitness of marine snail Hexplex trunculus (52365)

Maja Šrut, Anamaria Štambuk, Anita Erdelez, Dorotea Polović, Martina Furdek, Melita Peharda, Goran Klobučar.

Department of Biology, Faculty of Science, University of Zagreb, Croatia; Institute of Oceanography and Fisheries, Croatia; Division for Marine and Environmental Research, Ruđer Bošković Institute, Croatia.

Hexplex trunculus is a marine gastropod whose reproductive fitness can be severely affected by very low concentration of antifouling compound trybutil-tin (TBT). TBT has strong xenoandrogen impact in snails, causing development of imposex, e.g. superimposition of male sexual characteristic in females, thereby affecting the fitness of entire populations. TBT is also known as a DNA demethylating agent. The aim of this study was to unravel the interactions between TBT bioaccumulation, reproductive fitness, epigenetic and genetic endpoints in murex snail H. trunculus. Four populations inhabiting polluted sites with intense marine traffic and boat maintenance activity and three inhabiting sites with low anthropogenic impact were sampled in the coastal eastern Adriatic. Higher incidence of imposex was observed in snails inhabiting polluted than the reference sites. Analysis of methylation sensitive amplification polymorphism (MSAP) revealed existence of epigenetic differentiation among populations and lack of genetic population structure. Level of DNA methylation significantly correlated with the imposex level, suggesting epigenetic background of imposex development.

# Epigenetic variation and selection by aphids in Arabidopsis thaliana (52372)
**Klara Kropivšek**, Christian Heichinger, Tobias Züst, Lindsay Turnbull, Bernhard Schmid, Ueli Grossniklaus.

*Institute of Plant Biology & Zurich-Basel Plant Science Center, University of Zurich; Institute of Evolutionary Biology and Environmental Studies, University of Zurich; Department of Ecology and Evolutionary Biology, Cornell University; Department of Plant Sciences, University of Oxford.*

Aphids, plant parasites, exert strong selective pressure that leads to stronger defense responses in plants, such as higher trichome densities (Levin, 1973). Different aphid species led to selection of distinct phenotypes in experimentally evolved populations of Arabidopsis (Züst et al., 2012). Preliminary experiments indicate that this can occur within a given genotype, which suggests a role for epigenetic variation in this adaptive response (Hirsch et al., 2012). We measured trichome densities in 2 accessions of Arabidopsis that were exposed to different aphid species over 5 generations (Züst et al., 2012), and grown in absence of aphids for 2 generations. Both accessions responded with higher trichome densities in one aphid species treatment compared to the other. This finding is in concordance with other data on epigenetic inheritance (Scoville et al., 2011), yet the causal relationship between epigenetic changes and observed phenotypes needs to be elucidated at the molecular and genetic level.
We will analyse the genome-wide distribution of DNA-methylation to identify differentially methylated regions and to identify candidate genes involved in this defense response. We will investigate the function of selected genes by altering their DNA-methylation status using RNA-directed DNA-methylation to establish a causal link between adaptive phenotypic changes and the underlying epigenetic variation.

Poster session B - POL 300

# Heritable methylation patterns in widespread apomictic dandelions (52806)

**Veronica Preite.**

NIOO.

Evidence has been found for stress-induced methylation changes in different plant species, but it is unclear how stable these methylation changes are over multiple generations. We studied heritable methylation patterns triggered by environmental stress in apomictic dandelions, using methylation-sensitive (MS) AFLPs.

We exposed apomictic clone members to control, drought and salicylic acid stress and evaluated the offspring under control conditions for two subsequent generations. We found spontaneous methylation changes that occurred in the control as well as the stressed groups which showed a heritable fraction that was stable across three generations. Additionally, clone members that originated from different geographic sampling locations showed distinctive methylation patterns derived possibly from accumulating methylation changes. In contrast to our expectations, we found no evident stress-induced methylation changes. We also studied natural variation across dandelion populations which revealed a modest fraction of heritable methylation variation that could not be explained by genetic variation. Thus autonomous epigenetic variation potentially adds to the adaptive capacity of apomictic dandelions.
These results suggest that spontaneous rather than stress-induced methylation changes may play a role in building up heritable epigenetic variation that adds to adaptive differences within clonal lineages and across populations.

# DNA Methylation, Epigenetics, and Evolution in Vertebrates (52390)

## Annalisa Varriale.

*Università degli Studi di Napoli Federico II.*

DNA methylation is a key epigenetic modification in the vertebrate genomes known to be involved in biological processes such as regulation of gene expression, DNA structure and control of transposable elements. Despite increasing knowledge about DNA methylation, we still lack a complete understanding of its specific functions and correlation with environment and gene expression in diverse organisms. To understand how global DNA methylation levels changed under environmental influence during vertebrate evolution, we analyzed its distribution pattern along the whole genome in mammals, reptiles and fishes showing that it is correlated with temperature, independently on phylogenetic inheritance. Other studies in mammals and plants have evidenced that environmental stimuli can promote epigenetic changes that, in turn, might generate localized changes in DNA sequence resulting in phenotypic effects. All these observations suggest that environment can affect the epigenome of vertebrates by generating hugely different methylation patterns that could, possibly, reflect in phenotypic differences. We are at the first steps towards the understanding of mechanisms that underlie the role of environment in molding the entire genome over evolutionary times. The next challenge will be to map similarities and differences of DNA methylation in vertebrates and to associate them with environmental adaptation and evolution.

# Social and dietary stress and its implications on the life-history traits in wild rodents (bank vole – Myodes glareolus) through epigenetic changes. (52873)

## Joannes van Cann, Esa Koskela, Mappes Tapio, Sims Angela, Watts Phill.

*University of Jyväskylä; University of Oulu.*

Cyclic mammals can experience different forms of stress such as low food availability and high population densities. Especially for prenatal or early life stress, individuals may incur profound effects on life-history traits. It is theorized that the environment of the mother can lead to predictive adaptive changes in the offspring which can persevere over multiple generations, and that this is regulated by epigenetic mechanisms. In this study pregnant bank voles were exposed to either one or both stressors consisting of a low-protein diet, which mimics an environment with low food availability, and repeated close contact between
pregnant females, which mimics a high population density. Life-history traits of the offspring
were measured after which they were placed in semi-natural enclosures with high or low
population densities, where they were allowed to breed. Preliminary results show that social
stress has a negative effect on the litter size, while food stress has a negative effect on birth
weight. However, exposure to both stressors does not result in a lower birth weight, indicating
a role for terminal investment as an adaptation to stress. These life-history modifications may
indicate persisting epigenetic changes, in turn leading to differential breeding success and
survival in natural rodent populations.

Epigenetics in forest trees - Do methylation patterns in conifers change with environmental conditions? (52912)

Katrin Heer, Kristian Ullrich, Stefan Rensing, Lars Opgenoorth.

*Conservation Biology, University of Marburg; Department of Ecology, University Marburg;*
*Plant Cell Biology, University of Marburg.*

Forest trees are particularly challenged by climate change due to their sessile lifestyle and long generation times. While adaptation based on genetic changes might to be too slow to permit a reaction to rapid environmental changes in long-lived species, epigenetic mechanisms might enable a faster response to a changing environment. In a proof-of-concept study, we compare genome-wide methylation patterns in clone pairs of Norway spruce (Picea abies) of which the ortet and ramet grow under different environmental conditions since 1990. By using SeqCap Epi Kit (NimbleGen) we combine exome capture and bisulfite sequencing and target ~80 Gb in genes that were identified and annotated in the Norway spruce genome project. If we find consistent changes in methylation patterns across clone pairs this would strongly indicate that methylation patterns are involved in the reaction to changing environmental conditions within the lifetime of the trees. First results of this proof-of-concept study and a critical review of the feasibility of this approach will be presented during the conference.

Plasticity and epigenetics between native and invasive Phragmites lineages,
between genotypes, and within genotypes (53023)

Vladimir Douhovnikoff, Amy Spens.

Biology Department, Bowdoin College.

We hypothesize that epigenetic control, a form of plasticity that is transferable across cell
divisions as well as sexual generations, confers dynamic and stable responses to
environmental heterogeneity, characteristics that may be critical for successful plant
invasions. Phragmites provides an exceptional opportunity for the comparison of invasive and
non-invasive lineages within the same species, across a range of environmental conditions. Phragmites is also a facultatively clonal plant making possible the study of replicated identical genotypes. In an environmentally variable field based study in three marshes in Maine, USA we have measured epigenetic variation between clonal ramets, between genotypes, and between conspecific native and invasive lineages. Epigenetic variation was quantified using methylation sensitive amplified fragment length polymorphisms (MS--AFLPs). Results found large and significant differences(p < 0.001) in epigenome between lineages (native vs. invasive) and within genotypes (clonal ramets) but smaller difference between genotypes within lineages (genetically distinct genotypes). This suggests that the native and invasive lineages have distinct epigenomes, most epigenetic variation within a lineage is independent of genetic variation, and clonal ramets have distinct epigenomes in response to local site variation. In our working model, these results would indicate high levels of epigenetic plasticity within genotypes.

## Adaptations in the brains of Neotropical butterflies (51638)

**Stephen Montgomery**, Richard Merrill, Swidbert Ott.

University College London; University of Cambridge; University of Leicester.

Interspecific comparisons of brain structure can inform our functional understanding of brain regions, identify adaptations to species-specific ecologies, and explore what constrains adaptive changes in brain structure and shapes co-evolution between functionally-related structures. The value of such comparisons is enhanced when the species considered have known ecological differences. Lepidoptera have long been a favoured model in evolutionary biology, but to date descriptions of brain anatomy have largely focused on a few commonly used neurobiological model species. Through a combination of inter-specific and intra-specific comparative analyses my research explores how ecological selection pressures shape brain structure across different phylogenetic timescales in mimetic, Neotropical butterflies. Results to date have identified adaptive divergence in sensory neuropil, reflecting diel pattern and habitat preference, and the first description of sexual dimorphism in the antennal lobes of a diurnal butterfly, which is hypothesised to be linked to heightened reliance on pheromone communication in ithomiines. Further analyses concern the evolution and developmental plasticity of the mushroom bodies in Heliconiini. These structures, which are linked to memory and learning, are notably expanded in Heliconius. I am using comparative and experimental analyses to test the hypothesis that this expansion is related to a derived diet and foraging behaviour.

## Large brains enhance female, but not male survival in the guppy (51994)

**Alexander Kotrschal**, Séverine Buechel, Sarah Zala, Alberto Corral, Dustin Penn, Niclas Kolm.
Brain size is remarkably variable in vertebrates. To fully understand how brain size may evolve in natural populations, data on the fitness effects of brain size is required. However, to date such evidence is lacking. Using guppies (Poecilia reticulata) that were artificially selected for large and small relative brain size, we investigated how brain size affects survival. We cohoused mixed groups of 4800 marked small- and large-brained individuals equally distributed in six large semi-natural streams with their natural predator, Crenicichla alta, and monitored survival in weekly censuses until a predefined criterion of 50 % survival was met. Five months later, large-brained females had 13 % higher survival compared to small-brained females while brain size had no discernible effect on male survival. We suggest that large-brained females have a cognitive advantage that allow them to better evade predation, whereas large-brained males are more colorful, which may counteract any potentially beneficial effects of brain size. Our study provides the first experimental evidence that a larger brain size may confer survival benefits under naturalistic conditions and shows that the advantages can be sex-specific. We conclude that such fitness benefits provide an important selective force in the evolution of brain size.

## A mathematical model for brain development (52118)

Mauricio González-Forero, Timm Faulwasser, Laurent Lehmann.

Growing and maintaining a large brain entails energetic costs. A large brain can evolve if these costs are compensated by benefits from correspondingly high cognitive abilities. Benefits are thought to arise from ecological and social pressures, and mostly verbal theories have yielded predictions that have received some support. Here we develop a mathematical model for energy allocation into brain growth. We consider a body composed of brain cells, reproductive cells, and aneural, non-reproductive cells. Brain cells give the body skills to solve problems. Problem solving allows the body to obtain energy from the environment. The energy obtained can be allocated into maintaining and producing one or more of the three cell types. Under standard life-history assumptions, energy allocation maximizes lifetime number of offspring. Performing this maximization, we obtain ontogenetic curves for brain size. The model depends on measurable metabolic parameters, and is thus testable. The model can be extended to include social environmental pressures by letting problem solving depend on social partner's skill. Such an extension may yield discerning predictions among hypotheses for the evolution of brain size.

## Genetic mechanisms of caste differentiation in response to stress imposed by dominant female breeders of naked mole rats. (52303)
**Bitao Qiu**, Qiye Li, Pei Zhang, Zongji Wang, Lijun Jin, Yang Zhou, Vadim N. Gladyshev, Jacobus J Boomsma, Guojie Zhang.

*Centre for Social Evolution, University of Copenhagen; China National GeneBank, BGI-Shenzhen; Centre for GeoGenetics, Natural History Museum of Denmark, University of Copenhagen; School of Bioscience & Bioengineering, South China University of Technology; Division of Genetics, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School.*

Naked mole rats evolved eusocial division of labor by behavioral suppression, but the regulatory mechanisms and evolutionary underpinning of behavioral caste differentiation in mammals are largely unknown. We sequenced transcriptomes and DNA-methylomes of brains and gonads of male and female breeders and subordinates, and mapped these data on the genome. Only 4-5% of the brain genes were differentially expressed between castes, and a significant fraction of them interacted in gene networks with similar functional modules for both sexes. Gonadotropin-releasing hormone (GNRH1) was central in one of these modules, and linked with differentially expressed stress-response-genes, particularly neuropeptide-Y (NPY) and corticotrophin-releasing hormone (CRH). However, ovaries showed large expression differences between female castes with the estrogen synthesis pathway being blocked in subordinate females. Testes showed only modest gene expression differences with sperm-related genes being down-regulated in subordinates. Spermatogenesis in naked mole rats appeared to be degenerated, consistent with lack of sperm competition and complete mate-choice control by the breeding female. Caste differences in brain DNA methylation were also modest, but some of them were likewise associated with a stress response module. Our study strongly reinforces that reproductive suppression of both males and females is maintained by stress imposed by the bullying dominant female.

# Vomeronasal receptor families in the deer mouse Peromyscus maniculatus: Towards an evolutionary analysis (52867)

**Jean-Marc Lassance**, Yoh Isogai, Catherine Dulac, Hopi Hoekstra.

*Harvard University; Howard Hughes Medical Institute.*

Chemoreceptors are the first element in a complex pathway that can ultimately affect behavior -- changes in their biochemical properties or expression pattern have the potential to modulate specific behavioral responses. In particular, the activation of sensory neurons in the VNO is directly associated with changes in behavior, such as aggression, avoidance or mating. To understand the diversity and evolution of pheromone systems in wild populations, we are studying deer mice (genus Peromyscus), which diverged from Mus approximately 25 MYA. Here, we report on the characterization of VNO receptors from the most common and widespread species in North America, P. maniculatus. In total, we identified 150 and 90 putative V1R and V2R genes, respectively, in the Peromyscus genome, fewer than in M. musculus (239 and 120, respectively). While clades previously identified in Mus have representatives in Peromyscus, our phylogenetic reconstructions indicate that most gene duplications took place after the split between the two lineages; several clades show sign of
lineage-specific expansion or contraction. These differences in the chemosensory receptor repertoires likely reflect the unique habitats as well as social and mating behavior of deer mice.

# Bloodmeal size defines ejaculate production and impacts on male mating behaviour in bedbugs (51677)

Oliver Otti, Bettina Kaldun.

*Animal Population Ecology, Animal Ecology I, University of Bayreuth.*

Reproductive costs have been investigated mainly in females, even though males certainly also pay costs. As nutrition provides the basis for the production of biomass and reproductive tissues, variation in food availability might cause variation in sperm and seminal fluid production. We assume that meal size is directly associated with ejaculate size. Here we analysed the production of sperm and seminal fluid within three different feeding regimes with the result that large meals produced significantly more sperm and seminal fluid than smaller meals. While the number of offspring produced after a single mating was not influenced by feeding-status, copulation duration and ejaculate expenditure were shorter respectively smaller under food restriction. Food restricted males maintained mating for a shorter time than fully fed males, suggesting in a multiple mating context males pay a cost of reproduction and a direct effect of feeding on ejaculate production and male mating behaviour. Therefore, environmental variation, e.g. food availability, has a direct effect on variation in male reproductive traits, indicating that natural selection and sexual selection interactively define selection pressure on reproductive traits. As males often modulate their ejaculate size depending on the mating situation, future studies are needed to elucidate whether the amount of ejaculate available might interfere with different mating strategies.

# Parental roles, offspring fitness and gene expression under flexible parenting strategies (51699)

Darren J. Parker, Christopher B. Cunningham, Craig A. Walling, Clare E. Stamper, Megan L. Head, Eileen M. Roy-Zokan, Elizabeth C. McKinney, Michael G. Ritchie, Allen J. Moore.

*University of St. Andrews; University of Georgia; University of Exeter; University of Edinburgh; The Australian National University.*

Parenting in the burying beetle Nicrophorus vespilloides includes direct feeding of offspring and flexibility in both which and how many parents provide care. This flexibility is unusual and raises questions of why both sexes should care. We first examine the consequences of flexibility, and find that offspring do equally well regardless of the sex or number of parents. We used RNA-seq to examine differential gene expression associated with parenting and
found a largely overlapping set of expressed genes in both uniparental and biparental females and in uniparental males. These include genes such as vitellogenin, more commonly associated with reproduction in females, and takeout, associated with sex-specific behaviour. Biparental males did not differentially express many genes associated with parenting. Instead, gene expression was similar to non-caring states. Thus, by combining our behavioural and genetic analyses, our study shows that biparental care involves reduced male care rather than parental specialisation.

## Evolution of communication in socially polymorphic halictid bees (51720)

**Bernadette Wittwer**, Mark A. Elgar, Naomi E. Pierce, Sarah D. Kocher.

*School of BioSciences, University of Melbourne; Department of Organismic and Evolutionary Biology, Museum of Comparative Zoology, Harvard University.*

Communication is a key component for the maintenance and evolution of social behaviour. Studies into the evolution of communication in social insects focus primarily on chemical signals, with remarkably few investigating the functional importance of the receiving apparatus: the antennae. Here, we examine antennal diversity in halictid bees. Within the Halictidae, eusociality has been gained and lost a number of times, making the group ideal for investigating how communication systems are modified as social structure changes. We focus primarily on a socially polymorphic halictid bee, Lasioglossum albipes, where different populations produce either solitary or eusocial nests. An initial screen for genetic differences between behavioural morphs indicated there could be genotypic differences in genes associated with olfaction. Our research combines this behavioural and genomic data with morphological assays of antennal structures to provide insights into the role of communication in the evolution of eusociality.

## Predicting individual behaviour of wild crickets from lab measures of personality? (51873)

**David Fisher**, Adèle James, Rolando Rodríguez-Muñoz, Tom Tregenza.

*University of Exeter, Cornwall, UK; Université François, Tours, Rabelais, France.*

Examining the relevance of “animal personality” involves identifying the eco-evolutionary processes that lead to consistent among-individual behavioural variation. Typically, personality is assayed in captivity and related to important traits such as lifespan or fitness in the wild. However, behaviour is characteristically context specific and personalities in the laboratory may not represent personalities displayed in the wild. We compared laboratory and field measurements of the behaviour of wild field crickets (Gryllus campestris), by repeatedly measuring individual behaviour in the wild and repeatedly capturing the same individuals and
measuring their behaviour in captivity. We focused on three traits: boldness, activity and exploration. For all traits, behaviour in the laboratory did not predict behaviour in the wild. Boldness and activity were repeatable in the laboratory, but in the wild only activity was repeatable, despite measuring over 170 individuals 900+ times for boldness and 4000+ times for activity and exploration. Traits measured in the wild were predominantly influenced by external factors. We explored links between personality measures in both contexts and life-history traits and mating success in the wild. We discuss our results in the context of the Pace of Life Syndrome hypothesis as an explanation for the ubiquity of animal personalities

# Associations between personality and life-history traits differ across blue tit populations (51923)

Gabrielle Dubuc Messier, Anne Charmantier, Denis Réale.

Université du Québec à Montréal; Centre d'écologie fonctionnelle et évolutive, CNRS.

The pace-of-life syndrome hypothesis predicts that populations experiencing contrasted conditions will differ in a suite of behavioural, physiological and life history traits because these traits have coevolved under habitat-specific selective pressures. Such differences in coevolution among traits should thus be reflected by different P (co)variance matrices depending on the population and on the habitat. We compared mean phenotype and P matrices for personality (exploration patterns and handling aggression) and life-history traits (laying date and clutch size) across three blue tit (Cyanistes caeruleus) populations leaving in contrasted habitats. At the metapopulation level, we found a covariance between some behavioural and life history traits that is consistent with the pace-of-life hypothesis. Mean phenotypes and the covariance between traits varied substantially among populations, although habitat did not explain this variation as expected initially. Overall, our results support the pace-of-life hypothesis but they also reveal that we need considerable knowledge on the biology of a species and on the populations' ecological contexts before we can predict the relationships between personality and life history traits.

# Brain size affects the predator response of female guppies (Poecilia reticulata) (52000)

Wouter van der Bijl, Malin Thyselius, Alexander Kotrschal, Niclas Kolm.

Stockholm University.

There exists large variation in both absolute and relative brain size throughout the vertebrate lineages. While there has been a large effort to identify ecological drivers of this variation using comparative analysis, there exists little experimental work. Artificial selection in guppies (Poecilia reticulata) has produced small and large brained individuals. Recent work
on these selection lines has shown that large brained females had better survival when exposed to prolonged predation pressure under semi-natural conditions.

To elucidate the proximate mechanism behind this result we examined the behavioral response of these artificially selected guppies towards a model predator. Large brained females, but not males, spent less time performing predator inspections, an inherently risky behavior. Video tracking revealed that large brained females were further away from the predator while the response to a novel object control (a coffee mug) revealed no brain size differences.

These results suggest that large brained female guppies take less risk when confronted with a predator. They might be able to assess risk from a larger distance due to a superior visual system, or need less sensory information to reach an accurate conclusion. In natural situations, the threat of predation may thus result in selection for large brains.

# Allelic variation at TrpA1 and trpl regulates thermal-mediated behavior in the lab and field in Drosophila (52026)

Subhash Rajpurohit, Justin Walsh, Alan Bergland, Dmitri Petrov, **Paul Schmidt**.

*University of Pennsylvania; Stanford University.*

In genomic screens for SNPs that vary predictably over spatial and temporal gradients in natural populations of D. melanogaster, we identified seasonal/clinal polymorphism in two transient receptor potential ion channel (TRP) genes, TrpA1 and trpl. These ion channels are associated with Ca2+ influx, are light sensitive, and regulate aspects of sensory behavior related to temperature. Temperature varies predictably with season/latitude in eastern North America, generates context-dependency to multiple fitness parameters, and may underlie a series of adaptive life history tradeoffs. We hypothesized that these candidate cation channel polymorphisms were of functional significance and affected organismal response to temperature in the wild. To test this hypothesis, we constructed a replicated series of population cages to evaluate the focal SNPs of interest in a randomized genetic background. After ten generations of recombination, we evaluated the behavior of TrpA1 and trpl alleles using three distinct methodologies in the laboratory and field. The data demonstrate that, for both genes, the allele associated with northern locales and winter season exhibits a pronounced preference for cooler temperatures whereas the allele associated with southern/summer habitats prefers warmer temperatures. These data suggest that naturally occurring, cis-regulatory variation at these genes affects thermal behavior and contributes to climatic adaptation.

# Variation of absorption spectrum and gene expression pattern of red opsins under different light environments and its effect on behavioral spectral sensitivity in guppies, Poecilia reticulata (52171)
**Yusuke Sakai**, Satoshi Kasagi, Shoji Kawamura, Hajime Ohtsuki, Masakado Kawata.

*Department of Ecology and Evolutionary Biology, Graduate School of Life Sciences, Tohoku University; Graduate School of Frontier Sciences, The University of Tokyo.*

Color vision can facilitate finding food, avoiding predators and selecting mates. Color vision is enabled by utility of different spectral types of cone visual pigments which comprise an opsin and a chromophore. The guppy exhibits variation of spectral sensitivity among its four subtypes of long-wavelength sensitive opsin genes (LWS-1, LWS-2, LWS-3 and LWS-4). Additionally, allelic spectral variation is known for LWS-1 (A180 and S180). However, not know is if gene expression level differs among the loci and the alleles and if the spectral and expression variation is associated with variation of behavioral response to varying light spectra. In this study we examined the effect of different LWS-1 genotypes (A180/A180 and S180/S180) and opsin gene expression levels on behavioral response to different light regimes by optomotor response. The A180/A180 fish showed higher sensitivity to lights (546nm) than S180/S180 fish. Furthermore, juveniles reared in different light environments showed variation of gene expression in LWS-1. Individuals reared under orange light showed increased expression levels of LWS-1 and were more sensitive to orange than other lights. These results depict plasticity of opsin gene expression under different lightings and its interaction with opsin absorption spectra and behavioral response under varying light environment.

**Image analysis of weaverbird nests reveals signature weave patterns (52308)**

Ida E Bailey, André Backes, Patrick T Walsh, Kate V Morgan, Simone L Meddle, Susan D Healy, **Eira Ihalainen**.

*University of St Andrews; Universidade Federal de Uberlândia, Faculdade de Computação, Brazil; School of Biological Sciences, University of Edinburgh, Ashworth, King's Buildings, Mayfield Road,; Roslin Institute, Royal School of Veterinary Studies, University of Edinburgh, Easter Bush, EH25 9RG.*

In nature, many animals build structures. Individuality in construction behaviour within species may allow for more flexible use of building sites and materials, a potentially advantageous trait in stochastic environments. While measuring structures at the scale of their gross morphology (e.g. length, volume, weight) is relatively easy at this level of detail may be insufficient for detecting individuality in construction patterns. Capturing individuality in animal-built structures, such as can be done with the structures designed and built by human architects or artists, is therefore more challenging. Here we tested whether computer-aided image texture classification approaches can be used to describe textural variation in the nests of weaverbirds (Ploceus species) in order to attribute nests to the individual weaverbird that built them. We found that such an approach does allow the assignment of a signature to weaverbirds' nests. We suggest that this approach is a useful tool with which to examine individual variation across a range of animal constructions and not just for nests.
# Effects of colony genotype composition on a single brain methylome (52375)

**Oksana Riba-Grognuz**, DeWayne Shoemaker, Laurent Keller.

*University of Lausanne; Swiss Institute of Bioinformatics; USDA-ARS Center for Medical, Agricultural, and Veterinary Entomology.*

Epigenetic mechanisms regulate behavioral plasticity by translating environmental cues into stable transcriptional responses. In eusocial insects the morphological and behavioral phenotypes can be influenced by the genotypic composition of a colony. The red fire ant Solenopsis invicta provides a unique opportunity to study the role of genotypic composition, because they exist in two social forms determined by two variants (SB and Sb) of a social chromosome. Monogyne colonies contain a single queen with SB/SB genotype and exclusively SB/SB workers. By contrast, polygyne colonies contain multiple queens with an SB/Sb genotype and a mixture of SB/SB and SB/Sb workers. Experiments showed that SB/SB workers change their behavior and accept several SB/Sb queens if their colony contains more than 10% SB/Sb workers. To investigate how the genotypic composition of a colony affects patterns of brain DNA methylation and worker behavior we performed RNA-seq and BS-seq on single brains of SB/SB and SB/Sb workers introduced in host colonies of both social forms. Our results reveal the effect of DNA methylation on transcriptional response of single brains and the resulting behavioral response mediated by changes in social environment.

# I am too old for this: ageing great tits (Parus major) innovate less and persist less at a problem-solving task (52382)

**Maria Poujai.**

*University of Lausanne.*

Ageing is known to reduce animals' fitness over time, but few studies have examined its effect on their behavioural traits. Innovative problem-solving is a cognitive trait of particular interest as it is increasingly viewed as a manifestation of animals' ability to adjust to changing environments. Solving a novel problem requires several qualities, including low neophobia, high persistence, and learning. We chose adult wild great tits (Parus major) to investigate age differences in problem-solving performance, neophobia, and persistence. Using 5 years of cross-sectional data, we isolated within-individual behavioural changes by taking into account between-individual differences and demographic effects. Our results revealed that great tits experience a consistent within-individual decline in problem-solving ability. We also witnessed a within-individual decrease in persistence, but not in neophobia. These findings suggest that great tits undergo cognitive ageing over time, a process which appears to be induced by a reduction in persistence. However, persistence did not explain all age-dependent variations in cognitive ability. A number of other factors could underpin this decline, including a lack of motivation, stress management, learning difficulties, and limited motor
diversity. Behavioural consequences of ageing may be disadvantageous for older individuals
and thus be a proximate mechanism of reproductive senescence.

Genetic underpinning of a major evolutionary trait: eusociality in the facultative eusocial sweat bee Halictus rubicundus (52417)

**Antonella SORO**, Ioana LEMNIAN, Markus BÖNN, Ivo GROSSE, Margarita URIBE-LOPEZ, Sophie CARDINAL, Bryan N. DANFORTH, Robert J. PAXTON.

University Halle-Wittenberg, Halle (Saale), Germany; Cornell University, New York, USA; German Centre for Integrative Biodiversity Research (iDiv), Halle-Jena-Leipzig, Germany.

Eusociality is a major evolutionary innovation that has independently arisen multiple times in insects and has led to the ecological dominance of social species, but candidate genes that underpin this trait have not yet been identified. The application of advanced sequencing technologies to facultative eusocial species, in which some individuals are solitary and others are eusocial, is a powerful approach for the identification of such genes. Here, we perform a comparative transcriptome analysis of the facultative eusocial sweat bee Halictus rubicundus, chosen because of phylogeographic evidence for genetic differences between social phenotypes (solitary versus eusocial populations) in North America. Specifically, we analyze the transcriptomes of one solitary population and two eusocial populations of H. rubicundus using 454 and Illumina RNA-seq data, assemble a draft transcriptome of H. rubicundus comprising approximately 48,000 unigenes, and use it to map SNPs between the two eusocial populations and the one solitary population. We find marked genetic differentiation between the two social phenotypes of H. rubicundus, suggesting different demographic histories of the two forms, and point to putative candidate genes underpinning the ‘eusocial transition’ in this species.

Social effects on parental care in the cooperatively breeding Seychelles warbler (52427)

**Hannah Dugdale**, Lotte van Boheemen, Martijn Hammers, Sjouke Kingma, David Richardson, Jan Komdeur, Terry Burke.

University of Sheffield; Monash University; Wageningen University; University of Groningen; University of East Anglia.

How and why social behaviours evolve is a central question in evolutionary biology. The expression of social behaviours may be affected by the behaviour of the interacting individuals, which can have both a genetic and environmental component. For example, in cooperatively breeding species, both dominants and subordinates can raise the offspring of the dominants. The additional care provided by helpers may allow dominants to reduce their
parental care, providing indirect fitness benefits through increased dominant survival. Dominants may also modify their behaviour dependent on the identity of their social partner, which may have a genetic component. We tested the load-lightening hypothesis in the Seychelles warbler, and investigated social (indirect) genetic effects on parental care. We detected load-lightening, with dominants reducing their provisioning rates when they have more helpers, and discuss the fitness benefits. There was little between-individual variance in the amount of provisioning by dominants, but the variance in dominant provisioning was influenced by the identity of their social mate. The social environment therefore plays a strong role in parental care levels in the Seychelles warbler. We discuss the genetic component of these social mate effects, and their role in the evolution of social behaviour in wild systems.

# Are dispersing and non-dispersing individuals on different life paths? Looking at telomere lengths in the Collared flycatcher. (52470)

Charlotte Recapet, Blandine Doligez*, Pierre Bize*, These authors share seniorship.

Département Ecologie et Evolution, Université de Lausanne, Switzerland; Laboratoire Biométrie et Biologie Evolutive UMR CNRS 5558, Université Claude Bernard, Lyon, France; School of Biological Sciences, University of Aberdeen, UK.

In natural populations, trade-offs set the range of possible life histories. Nevertheless, variation in individual strategies is frequently observed within populations. Our previous results in a patchy population of a small passerine, the Collared flycatcher, suggest dispersing and non-dispersing individuals manage differently the trade-off between reproduction and maintenance in the short-term. However the longer-term consequences of such variation need to be examined to test whether dispersing and non-dispersing individuals reach the same lifetime fitness outcome. Because assessing survival and future reproduction prospects in a spatially limited study site can be biased with respect to dispersal, an alternative is to use cumulative, long-term markers of fitness-related traits. As telomere dynamics depend on physiological condition and often predict survival, we first investigate whether telomere length is a good marker of survival and reproduction prospects in our study population. We then test whether dispersing and non-dispersing individuals differ in telomere length and whether such difference depends on reproductive constraints. Based on our previous results, we hypothesise that dispersing birds will invest more in maintenance and exhibit longer telomeres, especially under constrained conditions, than non-dispersing ones. Such differences would be consistent with the integration of dispersal behaviour into a pace-of-life syndrome.

# The role of the aggression neuropeptide tachykinin in caste differentiation of ants (52500)

Jack Howe, Morten Schiøtt, Jacobus J. Boomsma.
Tachykinin is a neuropeptide known to control aggression in a wide range of animals, and appears to control the position of the stimulus response threshold for aggressive behaviour in Drosophila (Asahina et al., 2014).

We investigated whether the conserved gene Tachykinin has been coopted in ants that evolved physically differentiated female castes. We used the leaf-cutting ant Acromyrmex echination as a model because it has a reference genome, facilitating gene expression comparisons between: 1. founding queens (aggressive) and virgin queens (non-aggressive), and 2. large, aggressive foraging workers, and small nursing workers that are generally less aggressive.

We found that relative expression levels of tachykinin and its receptor agreed with hypothesised motivation states of the worker castes and their roles in colony defence, but expression appeared to disagree with aggression shown by founding queens, although this conclusion changed when adjusting for brain-mass differences between castes.

Tachykinin expression may therefore be important for explaining the extent of aggression by workers for different aspects colony defence, but it remains possible that aggression of founding queens is regulated by alternative genes involved in ancestral nest foundation and defence that predate the evolution of eusociality in the Hymenoptera.

# Inbreeding impacts reproductive success but not sperm performance in wild male song sparrows (52572)

**Sylvain Losdat**, Lukas F. Keller, Pirmin Nietlisbach, Peter Arcese, Ryan R. Germain, Jane M. Reid.

University of Aberdeen; University of Zürich; University of Zürich; University of British Columbia; University of British Columbia; University of Aberdeen.

Reproductive success is an extremely variable fitness component resulting from numerous behavioural and physiological processes, yet the genetic and environmental determinants of male reproductive success remain to be elucidated. We used long-term genetic pedigree data from wild song sparrows to demonstrate strong inbreeding depression in major components of male reproductive success. However, understanding the evolution of reproductive strategies and fitness in nature requires that the key life-history and physiological constraints underlying such inbreeding depression is understood. Sperm performance is a major determinant of male reproductive success in species with intense sperm competition. However the prediction that inbreeding depression in sperm performance could underlie inbreeding depression in male reproductive success has not been tested in nature. We therefore quantified inbreeding depression in sperm performance in the focal song sparrow population across three years. Contrary to prediction, sperm swimming velocity, motility, and longevity, which are key components of sperm performance, did not show significant inbreeding depression. These data imply that the severe inbreeding depression observed in male reproductive success may
not stem primarily from reduced sperm performance in inbred males, but instead may stem
from inbreeding depression in other fitness-related traits such as sexually-selected behavioural
traits.

Plasticity of aggression: the effects of current and past predation risk, social environment and sex (52579)

Gábor Herczeg, Nurul Izza Ab Ghani, Juha Merilä.

Behavioural Ecology Group, Department of Systematic Zoology and Ecology, Eötvös Loránd
University; Department of Biology, Faculty of Science, University of Putra Malaysia;
Ecological Genetics Research Unit, Department of Biosciences, University of Helsinki.

Behaviours are plastic traits allowing fast changes and fine-tuning in trait expression in
response to situational demands. Aggression is a behaviour where the right behavioural
decisions can have immediate fitness consequences. Using a factorial common garden
experiment we tested how past and present perceived predation risk, the social environment
and sex affected two components of aggression (viz. hesitation to attack and attack intensity)
in a social fish species, the three-spined stickleback (Gasterosteus aculeatus). Significant
individual differences in both hesitation and aggression suggest that these traits represent
animal personality. The social environment influenced expression of aggression: fish kept in
pairs attacked quicker than fish reared either alone or in large groups. Treatment influences on
attack intensity were more complex. While males were on average more aggressive than
females, males reared alone in the absence of past exposure to predator cues and alone-reared
females in the absence of current predatory cues were more aggressive than the rest of the
groups. The results demonstrate complex underpinnings to plasticity-driven behavioural
variation, and draw attention to the fact that the interpretation of the presence/absence of an
expected behavioural response may be difficult if it is influenced by both past and present
environmental conditions.

The effect of brain size on mate choice and mating competition in guppies
artificially selected for relative brain size (52713)

Alberto Corral-Lopez, Alexander Kotrschal, Niclas Kolm.

Zoology, Stockholm University.

Variation in aspects of brain morphology has been suggested as a plausible mechanism
underlying variation in sexual behavior. However, our knowledge of this potential association
is based almost exclusively on comparative analyses. In this study, we used guppies that had
been artificially selected for large or small relative brain size to experimentally test this
association and bridge the gap between micro- and macroevolutionary studies concerning the
link between brain morphology and sexual behavior. A previous study in these artificially
selected lines demonstrated that relative brain size is unlikely to affect variation in male
sexual behavior in the guppy when individuals interacted in pairs. Therefore, in this study we
extended the scope and investigated the effect of relative brain size on female sexual behavior
by means of a dichotomous choice test where females with different brain size were allowed
to choose between males which also differed in brain size. In addition, we investigated male
and female sexual behavior in a situation that permitted social interactions between multiple
individuals, allowing for direct and indirect competition. We discuss our findings in light of
existing theory on the effect of brain size and cognitive ability on mate choice and complex
social interactions during mating.

# The evolution of “good parent” signals in birds: a meta-analysis (52727)

Gergely Hegyi, Dóra Kötél, Miklós Laczi.

Eötvos Loránd University.

Mate choice is generally costly for the choosy sex, so fitness benefits are necessary for the maintenance of choosiness. Genetic benefits are widely examined and have received overall support from the published literature. On the other hand, direct benefits such as the signaling of parental quality are broadly assumed to be important but their testing is rare and their status in the literature is poorly known. Furthermore, in the case of parental quality signals, there is always a possibility that the observed pattern is modified by the preferential investment of the partner. In a phylogenetically corrected meta-analysis, we collated correlative results from birds concerning parental plumage coloration and the feeding rates of the ornamented parent and its partner. Overall evidence for both parental quality indication and preferential partner investment was weak and this was hardly influenced by the sex of the signaler. However, the correlations of feeding rates and ornamentation in both the ornamented sex and the partner showed a robust phylogenetic effect. Feeding correlations of the partner also differed among plumage color types. Finally, there were characteristic patterns in the relative investment of the two parents, indicating that partner investment cannot be neglected when assessing signals of parental quality.

# Spider aggression reflects within-group conflict, not group adaptation (52747)

Jay Biernaskie, Kevin Foster.

University of Oxford.

Why do populations and groups vary in individual aggressiveness? In some group-living spiders, individuals can be classified as ‘docile’ or ‘aggressive’, and this has become a key example of animal personality. However, it is unclear why groups vary in their composition
of docile and aggressive types. A recent study of spider groups claims that the docile to aggressive ratio is a collective trait that is finely adapted for the good of the group. In contrast, we use simple models of adaptive social behaviour to show that aggressive types are most common in contexts that promote within-group conflict. We conclude that variation in spider aggression is best explained by a careful consideration of between- and within-group selection.

**Personality-related survival and sampling bias in wild cricket nymphs (52751)**

**Petri Niemela**, Niels Dingemanse.

LMU-München; Max Planck Institute for Ornithology.

Studies of adaptive individual behavior ask whether individuals differ in behavior(s) and whether individual-level behavior is under selection. Evidence for adaptive personality is biased towards species where behavioral and life-history information can readily be collected in the wild, e.g. ungulates and birds. Here, we report repeatability for behaviors expressed by a wild insect (the field cricket; Gryllus campestris). We used mark-recapture models to simultaneously estimate personality-related survival and sampling bias, focusing on a life-history phase where all individuals can readily be sampled (nymphal stage). We assayed home range, flight initiation distance (FID) and emergence time after human-induced disturbance; all behaviors were repeatable. FID alone predicted both encounter and daily survival probability: bolder individuals were observed more often but lived less long. Individuals were also somewhat repeatable in the temperature under which they were assayed. Such environment repeatability can bias repeatability in behavior upwards though this was not the case. Our findings therefore demonstrate that personality variation exists in wild insects, and that it is associated with components of fitness (cf. survival). The opposite effects of FID on survival versus encounter probability illustrates the importance of applying specific study designs that control for sampling bias in the estimation of personality-related fitness components.

**Peas in a pod: nestling activity is influenced by social interactions and state but not social hierarchy (52782)**

**Isabel Winney**, Mirre JP Simons, Yu-Hsun Hsu, Julia Schroeder, Shinichi Nakagawa, Terry Burke.

University of Sheffield; University of Otago; Max Planck Institute for Ornithology.

To understand why individuals differ and how this variance is maintained is an important aim of evolutionary biology. These interests have propelled the study of animal personality (individual behavioural consistency). The two main mechanisms proposed for generating
between-individual differences in personality are social interactions (imposing frequency-
dependent selection) and state (stable physical traits). State dependent models are split as to
whether state and behaviour should be positively or negatively correlated. These mechanisms
have been tested a few times in adults but rarely in offspring, yet early life-history can have
long-lasting consequences for an individual's later life-history and phenotype. Therefore, we
pull apart social and state effects on a heritable personality trait in nestling house sparrows on
Lundy Island, UK. We used cross-fostering to separate social from natal brood and maternal
effects, and represented social structure with the within-brood competitive hierarchy. Overall,
nestling activity was unaffected by social hierarchy, but between-individual differences in
behaviour were based on mass (larger nestlings moved more) and general social interactions
(activity was more similar within a social brood). Similarity within broods could be driven by
shared genes, environment, or social interactions. These findings support state-dependent
models that stipulate positive feedback between state and behaviour.

Parent-offspring co-adaptation: generating between-family variation in
behavioural reaction norms for parental provisioning and offspring begging?
(52812)

Peter Korsten, Jan Komdeur.

*Department of Animal Behaviour, Bielefeld University; Behavioural Ecology and Self-organization Group, University of Groningen.*

Recent quantitative genetics theory predicts that coadaptation of the supply of parental care and offspring demand for care can generate between-family variation in parental provisioning and offspring solicitation. Hence, family-level parent-offspring coadaptation may be an important evolutionary source of individual behavioural variation. Key predictions are: (1) there is individual variation in the behavioural reaction norms (BRNs) for parental provisioning and offspring solicitation; (2) these BRNs are correlated between parents and their offspring; (3) cross-fostering creates mismatches between co-adjusted parental and offspring behaviours, leading to fitness costs to parents/offspring. To test these predictions, we carried out a large-scale experiment in a free-living population of blue tits (Cyanistes caeruleus). In two years, we cross-fostered 2/3 of the population's broods, while 1/3 served as controls. We estimated BRNs of individual parents by measuring provisioning rates as a function of short-term (6h) brood size manipulations, which created experimental variation in total offspring demand. We estimated BRNs of individual offspring by measuring begging intensity as a function of experimental variation in food deprivation. Our analyses reveal individual variation in the BRNs for parental provisioning and offspring begging, while we find limited evidence for parent-offspring coadaptation on a family level.

Colony composition, division of labor and fitness in clonal ant societies (52837)
**Yuko Ulrich**, Jonathan Saragosti, Daniel Kronauer.

*The Rockefeller University; The Rockefeller University; The Rockefeller University.*

Testing hypotheses on division of labor requires the ability to manipulate colony composition. In particular, the distribution of individual age and genotype across members of a social group, as well as the size of the group, are expected to affect division of labor and colony performance. However, how each of these factors affects individual and group level behavior is still poorly understood. We present results from a series of experiments manipulating these factors independently from one another in Cerapachys biroi, a queenless, parthenogenetic ant. The unconventional biology of C. biroi affords precise experimental control over 1) individual age and group demography, because of cyclic reproduction leading to discrete age cohorts, 2) individual genotype and the genetic composition of groups, because of clonal reproduction and 3) group size, because colonies of any size can be set up from totipotent workers. We set up replicate colonies of known composition and use automated tracking of individual ants in 100+ experimental colonies in parallel over a month to quantify how colony composition affects division of labor and group fitness. We show, for example, a positive effect of group size on fitness, and link this to differences in individual and group-level behavior.

Poster session B - POL 300

# Evolution of behaviors in response to chronic malnutrition in Drosophila melanogaster (52872)

**Roshan Kumar Vijendravarma**, Sunitha Narasimha, Tadeusz Kawecki.

UNIL.

Adaptations to changing ecological factors impinge on changes in several life-history traits. However, in comparison to the physiological and morphological traits very little is known about how behaviors evolve and mediate such adaptations. We experimentally evolved replicate Drosophila melanogaster populations for over ~170 generations for increased tolerance to chronic larval malnutrition by rearing them on poor larval diet. In comparison, to the control populations reared on standard food the malnutrition tolerant populations evolved several behaviors that mediate adaptation to poor nutrition. The behavioral changes in these evolved populations included: firstly, a 'sitter'-like larval foraging behavior; secondly, an increased propensity for larval predatory cannibalism and finally a reduction in their prepupal tunnel building behavior. In conclusion, these behavioral adaptations evolved in response to nutritional stress, serve two key functions: increased nutritional acquisition and reduction in energy expenditure.

Poster session B - POL 300

# Genetic basis for seasonal and circadian adaptation in Nasonia vitripennis (52875)
Light and temperature are important environmental factors that show daily and seasonal oscillations. In most organisms, physiological rhythms are regulated by an internal clock. Timing of migration, hibernation and reproduction need to be adapted to annual changes in the environment to maximize survival and offspring production. Nasonia vitripennis overcomes the unfavourable season by maternal induction of larval diapause, in the offspring. Populations of N. vitripennis collected along a latitudinal gradient in Europe show a cline in diapause induction, most likely caused by the seasonal fluctuations of day length. Interestingly, allelic variation of the clock gene period follows a similar cline. Clock genes Period (per), Cryptochrome2 (cry2) and Cycle (cyc) are part of a feedback loop, leading to circadian oscillations of mRNA and proteins of these genes. Changes in their cyclical expression pattern might indicate how the circadian clock of N. vitripennis processes seasonal changes. The expression levels of per, cry2 and cyc in two populations from the extremes of the cline were examined using quantitative PCR. We found that expression in the heads of wasps from northern and southern regions is dependent on the applied photoperiod and on latitude of origin. This suggest that clock genes are involved in photoperiodic response and their adaptive evolution.

# Breeding a hybrid line to overcome selection limits for voluntary wheel running in mice (52877)

An ongoing selection experiment for increased voluntary wheel running (a highly repeatable and widely varying behavior in mice) resulted in four high runner (HR) lines that run ~3-fold more revolutions per day than control lines.

One study showed that HR lines differed in amount and rate of response to uniform selection and in number of generations to reach selection limits (Careau et al. 2013). In another study, a cross between two HR lines showed sex-specific heterosis for wheel running (Hannon et al. 2011), suggesting different genetic factors underlying wheel running for the two sexes and in replicate lines, i.e., multiple solutions.

A study on thermoregulatory behavior in mice showed that selection limits in replicate lines for nest building could be overcome with continued selection on hybrid lines (Bult and Lynch 2000). Therefore, we crossed two HR lines and continued selection on this hybrid line concurrent with the parental lines.

**Results:** Both sexes showed heterosis in F1. However, continued selection resulted in sex-specific responses. Male hybrids show potential to break the selection limit, while female
hybrids run intermediate to parental lines. We will examine wheel-running component traits
(mean revolutions/minute and minutes/day) and estimate heritability using an animal model.

Genetic basis of variation in circadian rhythms in moth sexual communication
(52905)

Sabine Haenniger, David Heckel, Astrid Groot.

Max Planck Institute for Chemical Ecology; Max Planck Institute for Chemical Ecology;
University of Amsterdam, IBED.

Many moth species show specific daily activity rhythms in their sexual activities, which
seems to have arisen to minimize communication interference between closely related
species: co-occurring species with overlapping sex pheromone blends show temporal
differentiation in their daily sexual activities. However, the genetic differentiation of this
allochronic separation has barely been examined in any species so far. We investigate
differential timing of sexual communication in the noctuid moth Spodoptera frugiperda that
consists of two strains significantly differing in their timing of sexual activity1,2. We
conducted QTL analysis on the variation in timing in the two strains and found that one
chromosome explains most of the variance3. Interestingly, this chromosome harbours the
clock gene vrille, making this our main candidate gene. Even more interestingly, this same
chromosome is also the QTL for the pheromone variation between the two strains3,
suggesting a possible genetic interaction or hitchhiking between the two traits.

1Pashley et al. 1992. Ann. Entomol. Soc. Am. 85: 400-405.

2Schöfl et al. 2009. J. Evol. Biol. 22: 1447-1459.

3Haenniger et al. 2015. Submitted.

The evolution of technical innovation in primates (52941)

Ana Navarrete, Andrew Whalen, Sally Street, Simon Reader, Kevin Laland.

University of St Andrews; University of Hull; McGill University.

Previous studies on behavior evolution in birds reported that measures of both technical
innovation (e.g. tool using) and non-technical innovation (e.g. novel food discovery) correlate
with brain size. In primates, a correlation between general innovation and brain size has been
corroborated, but no analyses on the differences between technical and non-technical
innovation had been reported before. Using a new dataset on brain measures and behavioral
data from the literature, we found that diverse measures of brain size correlate strongly with
technical, but not non-technical, innovation. Additionally, using causal graph analysis, we
found that direct relationships between technical innovation and brain size, social
transmission and social group size. Diet breadth shares an indirect relationship with
innovation and brain size. We interpret this result as consistent with Wilson's behavioral drive
hypothesis, in which innovation and social learning drive brain evolution, with the additional
roles of social group size and diet breadth.

ANALYSIS OF PREZYGOTIC ISOLATION BETWEEN ROBERTSONIAN
AND STANDARD HOUSE MOUSE (Mus musculus domesticus) IN TUNISIA
(53543)

Said Lamia, Ould Brahim Ikhyarhoum, Chatti Nouredine, **Said Khaled**.

University of Monastir, Tunisia; University of Monastir, Tunisia; University of Monastir, Tunisia; University of Monastir, Tunisia.

In Tunisia, the Robertsonian mice (2n=22) occupy the towns and are exclusively commensal whereas standard ones (2n=40) in these locations are pushed to rural constructions. The scarcity of hybrids of several generations and the lack of large hybrid zone can be explained by postzygotic isolating mechanisms. However, the absence of F1 (2n=31) hybrids between the two races could not be related to such mechanisms and behavioural prezygotic isolating mechanisms are suggested. A caging mice of both chromosomal races was performed to evaluate the spatial distribution of mice according to their karyotypes and their geographical origins and to estimate the nature of real crossbreedings by analyzing the karyotypes of theoffsprings. Our results showed a significant grouping of mice of the same chromosomal karyotype whatever their origin is, a preference for intra-racial couplings in both chromosomal races and a well dominance of Robertsonian mice. Therefore, these prezygotic isolating mechanisms could account for the rarity of F1 hybrids and could strengthen, with postzygotic mating, the chromosomal speciation model in house mouse.

Large variations in HIV-1 viral load explained by shifting-mosaic
metapopulation dynamics (51614)

Katrina Lythgoe, Francois Blanquart, Lornenzo Pellis, Christophe Fraser.

Imperial College London; University of Warwick.

An important question in HIV research is why, without therapy, the time it takes to progress to AIDS varies so dramatically among individuals. Although set-point viral load (SPVL) is the most robust predictor of disease progression, standard well-mixed mathematical models of viral dynamics struggle to capture the orders of magnitude variation in SPVL. Here, we develop a biologically plausible model that captures the metapopulation dynamics of HIV within individuals and show that, under a broad range of parameters, a 'shifting-mosaic'
equilibrium is reached where the whole body's viral population is stable over time, but results from an underlying process of local infection and clearance within T-cell centers. Notably, this model can explain the large differences in SPVL observed among patients, and why evolution does not proceed as predicted from well-mixed models. Ultimately, our model demonstrates that taking into account levels of selection in HIV (sites of replication versus the whole host individual) gives a plausible explanation for how viral genotype/phenotype (infectivity) influences host phenotype (viral load).

# Antiviral drug resistance as an adaptive process: a survey (51751)

## Kristen Irwin, Jeffrey D. Jensen.

### EPFL.

Antiviral drug resistance is a matter of grave clinical importance that, historically, has been investigated mostly from a virological perspective. While the proximate mechanisms of resistance can be readily uncovered using these methods, larger evolutionary trends often remain elusive. Recent interest by population geneticists in studies of antiviral resistance has spurred new metrics for evaluating the demographic history of infection, mutation rate, and selective pressures incurred during viral adaptation to antiviral drug treatment. We review recent advances in the field, with a focus on outstanding questions of clinical significance for a range of drug and viral types. We find that the demographic and selective histories revealed by population genomic inference are integral to assessing the evolution of resistance as it pertains to human health.

# A novel classification for genetic elements in prokaryotic vehicles: inter-cellular mobility, symbiotic relationships, cell-level phenotypes and the levels of selection (51785)

## Matti Jalasvuori.

### University of Jyväskylä.

Prokaryotes harbor a variety of genetic replicators including plasmids, viruses and chromosomes, each having differing effects on the phenotype of the hosting cell. It is possible to classify the replicators of Bacteria and Archaea based on their horizontal transfer potential and the type of relationships, mutualistic, commensal or parasitic, that they have with the host cell vehicle. Horizontal movement of replicators can be either active or passive, reflecting whether or not the replicator encodes means to mediate its own transfer from one cell to another. From the perspective of the cell vehicle, the different types of replicators form a continuum from genuinely mutualistic to completely parasitic replicators. This classification
provides a general framework for dissecting prokaryotic systems into components that induce cell-level phenotypes that may be selected for on various levels.

# Are the effects of coevolution with a host equal for each parasite in a clonal population? (52068)

**Michaela Klösener**, Vera Kühlkamp, Rebecca Schulte, Joy Bose.

*Universität Osnabrück, Behavioural Biology, Barbarastr. 11, 49076 Osnabrück.*

The reciprocal evolutionary effects pathogens and their hosts have on one another are one of the most powerful selective forces in evolution, leading to adaptive genetic changes in two or more species. Especially microparasites are thought to adapt fast. However, these selective forces may have different consequences for individuals within a clonal population and lead to phenotypic or genetic diversification. To get insight into these individual changes we performed a laboratory-based evolution experiment by using different clonal strains of Bacillus thuringiensis as a bacterial microparasite and its nematode host organism Caenorhabditis elegans. This system allows us to observe consequences of adaptation in single as well as in mixed infections. After 10 host generations we tested changes in virulence in whole evolved parasite populations as well as in single clones isolated out of these populations. Our results revealed significant differences between the clones of one population in single as well as in multiple infections of the host. This indicates that during evolution, populations can diversify and population based analyses might neglect new, rare clones although these may be key elements for parasite adaptation.

# Evolutionary conflicts between Staphylococcus aureus during transmission and infection (52302)

Bernadette Young, Jessie Wu, Derrick Crook, **Daniel Wilson**.

*University of Oxford.*

Staphylococcus aureus presents a paradox, because despite its notoriety as a dangerous hospital-associated pathogen, it is a common constituent of the body's natural flora, carried harmlessly in the noses of one in three adults. From the perspective of the pathogen, invasive disease occurs rarely compared to asymptomatic carriage. One explanation is that the body's natural flora could be capable of evolving over time to become more virulent, but that virulent strains are less fit for onward transmission. In previous work, we investigated the evolutionary dynamics of nasal carriage in 16 people, and discovered that in one persistently carried nasal population, an excess of protein-truncating substitutions was associated with the transition to a life-threatening invasive blood stream infection. Here, I exploit a new systematic study of 1000 bacteria from 100 patients with invasive Staphylococcus aureus infections to ask
whether selection pressures during infection conflict with successful transmission, and
whether this could this play an important role in the progression of invasive staphylococcal
disease within the body. Our results reveal the evolutionary pathways differentially associated
with life-threatening infections versus onward transmission, and the repeatability of bacterial
evolution during infection.

# Choose your partner carefully: variation in aphid phenotypes when multiple bacterial symbionts coexist. (52306)

Melanie Smee, Sally Raines, Julia Ferrari.

*University of York.*

Bacterial communities within insects play an integral role in the biology of the host. For aphids, the phenotype is a result of both aphid and bacterial symbiont genotype. Little is known about the interactions between multiple species of facultative symbionts occurring within an individual. Selection will be acting on the individual aphid, but also within the aphid where symbiont species may be competing. In the pea aphid, Acyrthosiphon pisum, symbionts have a wide range of ecological effects, from protection from natural enemies to host plant utilisation. It has become clear that different strains of the same facultative symbiont species vary in their effects. Multiple symbiont species that coexist within the same host therefore allow us to study how a community evolves in response to selection on multiple levels. We used A. pisum individuals infected both singly and doubly with strains of Hamiltonella defensa and the X-type symbiont, to investigate how combinations of two common symbionts affected aphid phenotypes in different ecological situations. We found that the outcome of double infections is strongly dependent on the combination of individual strains of the two symbionts present and on the ecological context, highlighting the importance of genotype x genotype x environment interactions.

# Embryo genetics affects composition of host-associated bacteria in brown trout (Salmo trutta) (52368)

Laetitia Wilkins, Luca Fumagalli, Claus Wedekind.

*University of Lausanne.*

Fish embryos are associated with microbial communities whose composition can significantly
influence embryo development. Recent experiments demonstrated that such microbial effects
depend on host factors, i.e., on additive genetic and maternal environmental effects. We tested
whether host characteristics can in turn influence the composition of symbiont bacterial
communities. Brown trout (Salmo trutta) were sampled from a natural population and their
gametes were used to produce embryos in vitro in a full-factorial breeding design. Embryos
were raised singly, and different concentrations of nutrient broth were added to create different environments. We applied 16S rRNA MiSeq technology to characterize bacterial communities on the embryos. As expected, adding nutrient broth to stimulate microbial growth increased embryo mortality and changed the composition of bacterial communities. There were significant dam and sire effects explaining the variation in bacterial community composition associated with their offspring. With regard to bacterial diversity we only found evidence for significant sire effects. Due to the full-factorial experimental breeding, such sire effects directly reveal additive genetic variance. We therefore conclude that host genetics influences symbiont microbial communities even before hatching.

**Population genomics of the symbiotic fungus Rhizophagus irregularis suggests intercontinental dispersal. (52717)**

**Romain Savary**, Frédéric Masclaux, Tania Wyss Lozano Hoyos, Ian Sanders.

DEE.

The symbiosis formed by arbuscular mycorrhizal fungi (AMF) and the vast majority of vascular plant is known to be present worldwide. This association is important for ecosystem stability and some evidence suggests its role in the colonization of vascular plants on land. However little is known about how Rhizophagus irregularis the model fungus for this clade has colonized most places on earth. Moreover the population genomics tools have not been used to explore such patterns of genetic diversity. This is probably due to the difficulty to isolate and cultivate such obligate symbionts in order to obtain clean DNA. Advances in sequencing technics allow us to study the population of this fungus. We used a novel sequencing technique: Restriction site Associated DNA marker (RAD) to obtain a large number of SNPs markers from 63 isolates of 17 countries. We found that isolates from Europe and North America are genetically very similar, but at the same time, very high local genetic differences can be found. This indicates long distance migration, which is mysterious, as no known dispersal mechanism is known.

**The role of drug interactions and collateral effects in the evolution of antibiotic resistance in Pseudomonas aeruginosa (51662)**

**Camilo Barbosa**, Hinrich Schulenburg, Gunther Jansen.

*Department of Evolutionary Ecology and Genetics, CAU Kiel.*

The crisis of antibiotic resistance has led to the development of different deployment strategies aiming to limit adaptation to drugs. For instance, combination therapy is hypothesized to have a stronger effect than monotherapies while still limiting resistance evolution. In this context, the interaction between drugs in a combination and the availability
of mutations conferring simultaneous resistance to multiple drugs (collateral resistance) seem
to modulate resistance evolution. However, this has not been systematically tested for
opportunistic pathogens like Pseudomonas aeruginosa, where combination therapy is routine.
Here we use experimental evolution to test the long-term effects of the interactive and
collateral effects of antibiotic combinations in P. aeruginosa. We found that despite
influencing the success of a combination in the long-term, the interactive and collateral effects
of combinations are not sufficient to predict the outcome of evolution. Instead we observed
that antibiotics like ciprofloxacin or carbenicillin have a restrictive potential that dramatically
constrain antibiotic resistance evolution in P. aeruginosa. Moreover, by properly combining
these antibiotics with aminoglycosides or cell-wall inhibitors, the restrictive potential can be
strengthened, even to the point of extinction. Our results illustrate how proper identification of
antibiotics showing this restrictive potential could aid the selection of drug combinations for
the treatment of P. aeruginosa.

Genomic coevolution of natural phage communities with antibiotic resistant Pseudomonas (51692)

Robert Hertel, Lisa Stuckenschneider, Heiko Liesegang, Gunther Jansen.

*Genomic and applied microbiology, University of Göttingen; Evolutionary Ecology and Genetics, University of Kiel.*

Over 25000 people die yearly in the EU as a result of infections with antibiotic resistant bacteria, a number unlikely to decrease soon. Because the antibiotic arsenal to treat common infections such as Pseudomonas aeruginosa is depleting fast, we need novel solutions. An old idea that, at least among evolutionary biologists, is going through a revival is phage therapy. This approach promises to circumvent antibiotic resistance evolution by deploying viruses that infect and kill bacteria, and also (co)evolve with hosts that become resistant to viral attack. Although phage-bacteria coevolution has been well-studied phenotypically, a precise, quantitative understanding of the longer-term impact of coevolution on the genomic makeup of a diverse viral community is less understood. It also remains unclear how diverse viral communities interact with host genomes and potential host-associated prophages. In this study we therefore isolated and genomically characterized natural communities of bacteriophages able to infect the Pseudomonas aeruginosa. We coevolved these communities with highly antibiotic resistant Pseudomonas and investigated how the dynamics of the evolving interaction shaped the genomic diversity of viral communities. Our research improves the understanding of mechanisms underlying phage-bacteria coevolution, and may aid development of better, genomically informed phage therapies for the treatment of multi-, extensively and panresistant bugs.

Killing in a team - Cooperation and virulence evolution in an opportunistic pathogen (51702)
**Elisa Granato**, Rolf Kümmerli.

*University of Zurich, Switzerland.*

Virulence in bacterial infections is typically mediated through secreted metabolites that can be cooperatively shared across the bacterial collective within the host. Thus, populations of metabolite producers can potentially be invaded by “cheating” mutants, which no longer contribute but still benefit from the shareable metabolites produced by others. Consequently, conditions favoring cheats should ultimately select for reduced virulence.

Here, we test this hypothesis using the opportunistic pathogen Pseudomonas aeruginosa infecting a nematode host. We predict that: (a) cheats should invade, and therefore lower virulence in unstructured habitats, where metabolites and cells mix readily; (b) the spreading of cheats should be hampered and high virulence preserved in structured habitats, where metabolite sharing occurs more locally among cooperators; (c) higher levels of cooperative metabolite production might be maintained in the presence than in the absence of hosts, because hosts provide a structured haven even in an otherwise unstructured environment.

To test these predictions, we let P. aeruginosa experimentally evolve in the absence or presence of its host in liquid (unstructured) and agarized (structured) medium. Subsequently, we measured the evolved level of virulence and related this measure to the evolved production levels of cooperative virulence factors (toxins, siderophores, rhamnolipids) and cheat frequency.

Poster session B - POL 300

# Quantifying transmission fitness cost of MDR Tuberculosis (51708)

**Julija Pecerska.**

*ETH.*

Nowadays tuberculosis is mistakenly thought by many to be an easily curable and non-prevalent disease. However, only due to HIV/AIDS it takes second place as the greatest killer worldwide. A more pressing problem is that of multidrug-resistant (MDR) tuberculosis - bacteria that are immune to first line drug treatments.

Drug resistance is always a problem for the infected individual due to the difficulty of treatment. If MDR strains are furthermore transmitted between hosts, we may encounter a future epidemic for which no optimal treatment is available. In order to select most efficient treatment strategies, it is necessary to differentiate between highly transmissible and unsuccessful MDR strains.

Using whole genome sequencing data of tuberculosis strains from different hosts, we aim to quantify transmission fitness for the drug sensitive and various MDR strains using a novel phylogenetic inference tool. We present a simulation study showing that for simulated tuberculosis epidemics, we can infer the transmission fitness for the various strains. This will allow us to analyse empirical sequencing data which is currently collected in Georgia.
# Reversibility of phage-induced loss of conjugation among drug-resistant bacteria depends on the type of evolutionarily rescuing mutation (51739)

Ville Ojala, Sari Mattila, Matti Jalasvuori.

*University of Jyväskylä.*

Horizontal gene transfer via conjugative plasmids is an important contributor to the worrying spread of antibiotic resistance in bacterial communities. However, there is also a natural counter-selective force against bacterial conjugation in so-called plasmid-dependent phages which utilize the conjugation machinery to infect and kill plasmid-harboring bacteria. Here, by combining in vitro selection experiments and sequence analyses, we associated phage-induced changes in bacterial ability to conjugate with genetic and phenotypic characteristics. We found that various types of spontaneous mutations can evolutionarily rescue plasmid-harboring bacteria by providing a linked phenotype of phage resistance and reduced or lost potential for horizontal gene transfer. Promisingly, most of the changes in conjugation were irreversible even under prolonged antibiotic selection for restoring the previous phenotype. Only when the loss of conjugation was due to a dynamic mutation, namely a tandem repeat addition, the plasmid rapidly reverted back to full conjugation ability. However, in these cases the susceptibility to the phage also returned. Our results suggest that plasmid-dependent phages could be used to prevent the conjugative spread of antibiotic resistance, with long-lasting effects.

# Interactions between horizontally acquired genes create a fitness cost in Pseudomonas aeruginosa (51910)

Alvaro San Millan, Macarena Toll-Riera, Qin Qi, R Craig MacLean.

*University of Oxford; University of Zurich.*

Horizontal gene transfer (HGT) plays a key role in bacterial evolution, especially with respect to antibiotic resistance. Fitness costs associated with mobile genetic elements (MGEs) are thought to constrain HGT, but our understanding of these costs remains fragmentary, making it difficult to predict HGT. Here we use the interaction between P. aeruginosa and a costly plasmid (pNUK73) to investigate the molecular basis of the cost of HGT. Using RNA-Seq, we show that the acquisition of pNUK73 results in a profound alteration of the transcriptional profile of chromosomal genes, including DNA damage response genes. Mutations that inactivate two genes encoded on chromosomally integrated MGEs recover these fitness costs and transcriptional changes by decreasing the expression of the pNUK73 replication gene. Our study demonstrates that interactions between MGEs can compromise bacterial fitness via altered gene expression, and we argue that conflicts between mobile elements impose a general constraint on evolution by HGT.
# Relationship between siderophore availability and virulence in Pseudomonas aeruginosa (52013)

**Michael Weigert**, Adin Ross-Gillespie, Sam Brown, Rolf Kümmerli.

*University of Zürich; Ludwig Maximilians University Munich; University of Edinburgh.*

The ability of bacteria to establish an infection typically relies on the production of virulence factors. While the relationship between pathogenicity and virulence factor secretion is well established, little is known about the optimal level at which a virulence factor should be expressed. If production level scales linearly with pathogenicity then virulence factor over-production may lead to the premature killing of the host, whereas underproduction may compromise the ability of bacteria to establish an infection. Both effects can lead to reduced bacterial fitness, which should select for intermediate levels of virulence factor production. We tested this hypothesis by experimentally manipulating the availability of pyoverdine – a key virulence factor, produced by Pseudomonas aeruginosa – within a model host, the greater waxmoth larvae. We found that pyoverdine availability indeed scaled linearly with host mortality. Furthermore we showed that within-host bacterial fitness peaked at intermediate levels of pyoverdine availability, supporting our hypothesis that both pyoverdine over- and under-availability compromise bacterial fitness. At the mechanistic level, we found that increased pyoverdine availability lowered pathogen fitness not because it altered bacterial gene expression, but rather because it triggered a stronger host immune response.

# Within-host evolution decreases virulence in an opportunistic bacterial pathogen (52109)

**Lauri Mikonranta**, Johanna Mappes, Jouni Laakso, Tarmo Ketola.

*University of Jyväskylä; University of Helsinki.*

Most pathogens evolve in a close antagonistic relationship with their hosts. The conventional theory proposes that evolution of virulence is highly dependent on the efficiency of direct host-to-host transmission. Opportunistic pathogens, however, are not strictly dependent on the hosts due to their ability to reproduce in free-living environment. Therefore it is likely that conflicting selection pressures for survival and reproduction outside versus within the host, rather than transmission potential, shape the evolution of virulence in opportunists. We tested the role of within-host selection in evolution of virulence by letting a pathogen Serratia marcescens sequentially infect Drosophila melanogaster hosts and then compared the virulence to strains that evolved only in the outside-host environment. We found that the pathogen adapted to both Drosophila melanogaster host and novel outside-host environment, leading to rapid evolutionary changes in the bacterial life-history traits including motility, growth rate, biomass yield and secretion of extracellular proteases. Most significantly, selection within the host led to decreased virulence while the selection lines in the outside-host environment maintained the same level of virulence with ancestral bacteria. This
experimental evidence supports the idea that increased virulence is not an inevitable consequence of within-host adaptation even when the epidemiological restrictions are removed.

# Antibiotic dose specialization in bacterial experimental evolution (52261)

**Noémie Harmand**, Romain Gallet, Guillaume Martin, Thomas Lenormand.

*Centre d'Ecologie Fonctionnelle et Evolutive-UMR 5175; Institut des Sciences de l'Evolution de Montpellier.*

Antibiotic resistance is an adaptive process which poses a worldwide medical challenge. Whereas many complex mechanisms of resistance have been investigated since the 40's, the evolution of resistance has mostly been described in a simplified ecological context: treated versus non-treated environments. Evolutionary ecology studies, however, have often emphasized fitness trades-off between environments along a gradient. Thus, considering only treated and untreated environments highly simplifies an antibiotic dose gradient context. In this context, several adaptive strategies could exist between doses and reveal trades-off along the gradient. This would be crucial to understand the role that low doses play on the evolution of super-resistant mutants and in general the role of dose variation in evolution of resistance. We performed experimental evolution of an Escherichia coli strain at five antibiotic doses to investigate whether specialization occurs in resistance evolution. We measured the relative fitness of mutants in each dose as well as without antibiotics, after short and long term evolution. The results have been interpreted in the light of fitness landscape models and provide new insights in the role of dose gradient for the evolution of resistance.

# The genomic architecture of metabolic evolutionary innovation in Pseudomonas aeruginosa (52271)

**Macarena Toll Riera**, Alvaro San Millan, Wagner Andreas, Craig MacLean.

*Institute of Evolutionary Biology and Environmental Sciences, University of Zurich; Department of Zoology, University of Oxford.*

Novel phenotypes that emerge in evolution often have obscure genetic origins. Here we elucidate these origins by evolving populations of the bacterium P.aeruginosa in 95 different chemical environments distinguished by their carbon substrates. Evolutionary adaptation to some environments requires qualitatively novel traits (innovations), but in other environments mere improvement (optimization) of existing traits suffices. Whole-genome sequencing of evolved clones revealed profound differences in the genetic architecture of these two classes of traits. Innovation was characterized by mutations in regulatory and metabolic genes, and optimization by mutations in sensory and signalling systems. Whereas novel gene duplicates
arose at equal frequency during innovation and optimization, mutations in existing duplicates were much more common during innovation, demonstrating that the divergence of existing gene duplicates is a key driver of evolutionary innovation. Both optimization and innovation were associated with fitness costs, but the cost associated with innovation was almost two-fold greater than the cost of optimization, suggesting that pleiotropic trade-offs constrain metabolic innovation. Pleiotropic trade-offs, however, were reduced in clones carrying mutations in recent duplicates, suggesting a link between duplication, mutational robustness and functional innovation. Our observations suggest that phenotypic innovation and optimization can be caused by different classes of genetic change.

## Evolution of S. aureus during nasal carriage involves cross-lineage transfer of mobile genetic elements (52378)

**Tanya Golubchik**, Antonina Votintseva, Daniel Wilson, Rosalind Harding, Rory Bowden, Sarah Walker, Derrick Crook.

Nuffield Department of Medicine, University of Oxford, Oxford, UK; NIHR Oxford Biomedical Research Centre, Oxford, UK; Wellcome Trust Centre for Human Genetics, Oxford, UK; Department of Zoology, University of Oxford, Oxford, UK.

Staphylococcus aureus is a major cause of serious soft tissue infections and bacteraemia, yet it is also a successful commensal, readily isolated from the nasal flora of around one-third of healthy adults. This state of asymptomatic carriage, which can persist over many months, provides the ideal opportunity for new S. aureus lineages to emerge, particularly where different genotypes can interact and share genetic information. We were interested in investigating how this process occurs in vivo. To maximise the potential for interaction between different lineages, we sequenced 36-72 S. aureus isolates from each of 7 persistent long-term carriers of at least 2 distinct genotypes (as determined by spa typing). Despite 10-20 months of prolonged co-colonisation, homologous recombination appeared rare; genotype identity was preserved, and most chromosomal changes appeared to be due to mutation rather than recombination, as expected for a predominantly clonal organism. However, there was overwhelming evidence of mobile genetic element (MGE) activity, including transfer of conjugative plasmids, and gain/loss of bacteriophage and genomic islands. MGE activity occurred both within and between the distinct lineages present within a host. These findings add to the growing body of evidence suggesting that much S. aureus evolution is due to intense MGE activity.

## The genetic basis of phenotypic switching in Pseudomonas fluorescens SBW25 (52419)

**Gayle Ferguson**, Philippe Remigi, Jenna Gallie, Paul Rainey.
Institute of Natural and Mathematical Sciences, Massey University, Auckland, 0745, New Zealand; New Zealand Institute for Advanced Study, Massey University, Auckland, 0745, New Zealand; Department of Environmental Microbiology, Eawag, 8600 Dübendorf, Switzerland; Department of Environmental Systems Science, ETH Zürich, 8092 Zürich, Switzerland; Max-Planck Institute for Evolutionary Biology, 24306 Plön, Germany.

When challenged with repeated selective cycles through two contrasting environments, the bacterium Pseudomonas fluorescens SBW25 evolved, de novo, the ability to switch repeatedly between two bistable states – one where cells secrete an extracellular colanic acid-like capsule (Cap+), and an ancestral, uncapsulated state (Cap-). Subsequent characterisation showed the switch to be epigenetic, though underpinned by mutation at the start of the pyrimidine biosynthesis pathway. Taking a genetic approach, we sought to identify the ultimate cause of bistability. Exploration of the genotype-phenotype map reveals a complex network of ‘players’ whose activities modulate Cap-/Cap+ switching. Among these are the Gac/Rsm signalling pathway and the stringent response, specifically, RpoD and the nucleotide alarmone ppGpp, recently shown to underpin another bistable phenotype – that of antibiotic ‘persisters’.

Poster session B - POL 300

# Do antibiotic resistance alleles affect adaptation to phages? (52496)

Flor Ines Arias Sanchez

ETH Zurich.

Bacteria evolve in complex environments with a variety of selection pressures, including antibiotics and parasitic viruses (phages); yet the interactions among these different selection pressures are unclear. We are addressing this question by performing fluctuation assays and looking at differences in mutation rates to phage resistance. We use a panel of eight E.coli strains carrying various antibiotic-resistance alleles (including chromosomal mutations and plasmids) against a variety of phages from different taxonomic groups. Our results may have far reaching consequences for the development of antimicrobial strategies. Knowing if AB-resistant pathogens are different in their adaptive response to the presence of viral parasites, not only when compared with AB-susceptible variants but also among other strains with different resistant mechanisms will (a) provide insights into how bacteria adapt to ecologically complex scenarios, and (b) will determine whether the efficacy of phage therapy (using phages to treat bacterial infections) will depend on the evolutionary history of pathogenic bacteria in terms of exposure to antibiotics.

Poster session B - POL 300

# Molecular basis of drug resistance-related fitness costs in Mycobacterium tuberculosis (52695)
**Sebastian Gygli**, Olga Schubert, Sonia Borrell, Andrej Trauner, Julia Feldmann, Xueli Guan, Mireia Coscollá, Ruedi Aebersold, Sébastien Gagneux.

*Swiss Tropical and Public Health Institute, Basel, Switzerland; University of Basel, Basel, Switzerland; Institute of Molecular Systems Biology of the ETH Zürich, Zürich, Switzerland; Systems Biology Graduate School, Zürich, Switzerland.*

The antibiotic rifampicin (RIF) targets the bacterial DNA-dependent RNA polymerase. RIF resistance mutations often carry a fitness cost in Mycobacterium tuberculosis (Mtb) in absence of the drug. Recently, secondary, so-called compensatory mutations have been described, which restore the fitness of RIF resistant Mtb mutants. The molecular basis of drug resistance-associated fitness costs and their compensation is poorly understood.

As RIF targets the information pathway (DNA -> mRNA -> proteins), resistance and compensatory mutations will likely influence the proteome of Mtb. To gain more insight into the molecular basis of drug resistance-related fitness costs, we are analyzing the proteomes of drug resistant and susceptible Mtb strains using a quantitative mass spectrometric approach. First, we will compare the proteome of a susceptible Mtb strain to that of a RIF resistant strain derived from it. Furthermore, we will analyse the effect of a putative compensatory mutation on the proteome of the RIF resistant strain.

We anticipate that the analysis of proteomes of drug resistant Mtb strains will lead to a deeper understanding of how drug resistance and compensatory mutations impact on the fitness, and therefore on the transmission potential of drug-resistant Mtb.

## Testing Discrepancies in Mutation Rate Estimations (52712)

**Huw Richards**, Rok Krašovec, Andrew McBain, Christopher Knight.

*Faculty of Life Sciences, University of Manchester; Manchester Pharmacy School, University of Manchester.*

Mutation rate is a central evolutionary parameter that has been studied intensively for nearly a century. For microbes, a key assay used to estimate mutation rate is the fluctuation test, where phenotypically identified mutant cells, arising in the absence of selection, are counted.

However, published mutation rates estimated by this method vary substantially between studies, even of the same bacterial strains. This can make comparison difficult or impossible.

Here we test the possibility that these issues may not merely reflect limited accuracy or reproducibility in the method but have a biological origin. Specifically, recent findings on mutation rate plasticity (mutation rate variation at a given site in a single genotype) suggest that the density to which bacteria are grown can affect their mutation rate. Final population density may be calculated for the fluctuation tests behind many published mutation rates. We therefore collated a dataset comprising hundreds of mutation rate estimates across tens of studies. With this we assess the dependence of mutation rate on final population density. We find that population density can explain a substantial proportion of variation in mutation rate estimates. This suggests that fluctuation tests may be a better tool for mutation rate estimation than widely assumed.
Evolution of commensal bacteria through experimental transmission (52764)

Anaid Diaz, Olivier Restif.

University of Cambridge.

Intestine microbes are an important component of many organisms. Microbes prevalence among individuals in a population raises questions about what adaptations do microbes have in order to spread and persist. We hypothesise that bacterial transmission, which is often neglected, can change the trajectory of microbial evolution, for instance, by selecting on bacterial colonisation rate inside the host and bacteria attractiveness to the host. We are investigating this hypothesis by using experimental evolution of a lab microcosm: Caenorhabditis elegans as a host and Sallmonella enterica as intestinal bacteria. C. elegans can transmit its enteric microbes to conspecifics through faecal-oral route; we recently showed that bacteria can thrive among worms by this process. We are developing this system further by allowing transmission of enteric bacteria among worms over multiple generations. We are assessing the phenotypic and genetic changes of the evolved bacteria compared to two treatments: ancestral population and passages of random bacteria. We will discuss our results and provide new insights about the importance of host-to-host transmission in the evolution of intestinal bacteria.

Population size and Parallel evolution during the evolution of antibiotic
resistance (52790)

Tom Vogwill, Robyn Phillips, Danna Gifford, Craig Maclean.

Department of Zoology, University of Oxford.

How repeatable is adaptation? There are many striking examples of parallel molecular evolution from clinical pathogens, particularly for genes involved in antibiotic resistance evolution. This is often interpreted as suggesting there are relatively few routes through which adaptation can proceed, and adaptation is therefore highly repeatable. However, the probability of parallel evolution is predicted to be strongly dependent on population size. We tested the role of population size in parallel evolution by experimentally evolving antibiotic resistance in the bacterium Pseudomonas fluorescens at three different population sizes. Following whole-genome sequencing of 93 evolved populations, we find that increasing population size does lead to increased parallelism within several genes. However we also find examples where increasing population size decreased the probability of adaptation. We find that the probability of parallel evolution within a gene is dependent on (i) the definition of parallel evolution; (ii) how strongly beneficial a gene is; (iii) whether the gene is a primary or secondary mechanism of adaption. Overall, we conclude that in some cases parallel evolution may be due as much to demography as it to a restricted adaptive landscape.
# Photonic Crystals Cause Active Colour Change in Chameleons (52909)

**Michel Milinkovitch**, Jérémie Teyssier, Suzanne Saenko, Dirk van der Marel.

*University of Geneva, www.lanevol.org.*

Many chameleons, and panther chameleons in particular, have the remarkable ability to exhibit complex and rapid colour changes during social interactions such as male contests or courtship. It is generally interpreted that these changes are due to dispersion/aggregation of pigment-containing organelles within dermal chromatophores. Here, combining microscopy, videography, photometry and photonic band-gap modelling, we show that chameleons shift colour through active tuning of a lattice of guanine nanocrystals within a superficial thick layer of dermal iridophores. In addition, we show that a deeper population of iridophores with larger crystals reflects a substantial proportion of the sunlight power, especially in the near-infrared range. The organisation of iridophores into two superposed layers constitutes an evolutionary novelty for chameleons that allows some species to combine efficient camouflage with spectacular display and passive thermal protection.

# How does the ocellated lizard (Timon lepidus) develop its color pattern?
(51747)

**Sophie Montandon**, Liana Manukyan, Michel Milinkovitch.

*University of Geneva.*

Animal color patterns are of wide adaptive value for camouflage and display. Although mutational analyses have shed some light on the mechanisms underlying Turing pattern formation in the zebrafish, virtually nothing is known on how reptiles achieve their tremendous diversity of colors and color patterns. Like fishes, reptiles exhibit three types of color-producing cells: i) melanophores, that produce melanin, ii) xanthophores/erythrophores that synthetize pteridines or store carotenoids and iii) iridophores that contain photonic crystals generating structural colors such as green, blue or white. To study color pattern formation in reptiles, we use the ocellated lizard (Timon lepidus), which dorsal scales are white or brown in hatchlings and change to green, blue or black in adults. To understand how color changes occur, we used TEM to identify the skin cells involved in colors production. To follow the color state of each dorsal scale, we reconstructed the high-resolution 3D geometry and color texture of specific individuals at regular time intervals. We identified the color of more than 5000 scales per animal and follow their state from newborn to adulthood. We used this data to statistically analyze how scales are changing colors with time. Finally, we mathematically modeled how color pattern is evolving through the life of an animal.
# Not just a colour shift: insect melanism and fecundity (52103)

**Panu Välimäki**, Sami M. Kivelä, Netta Keret, Markku Orell, Toomas Tammaru.

*University of Oulu; University of Stockholm; University of Tartu.*

The life history theory is grounded upon trade-offs that hinder trait evolution; an increase in fitness due to a change in one trait is opposed by fitness costs due to a concomitant change in another trait. To study the role of trade-offs in maintaining larval polymorphism in insects, we applied artificial selection and quantitative genetic methods to a geometrid moth with discrete melanic and non-melanic larval morphs. Larval coloration appeared a heritable trait that responded to artificial selection in a predictable way, proving the genetic basis of the phenotypes. Larval development time is a key determinant of survival in temperate insect populations. There was, however, no genetic correlation between larval coloration and development time that would constrain phenotypic evolution. Instead, melanism was genetically linked to a small body size at maturation. The reduced body size was not due to melanisation process itself because non-melanic larvae from the melanic selection lines pupated at a smaller size than melanic larvae from the non-melanic selection lines. We conclude that directional selection on melanism affects an underlying costly trait or function, which is adaptive under certain conditions (e.g. under a risk of desiccation or pathogen/parasitoid attack), but traded-off against body size, and thus fecundity.

# Understanding the demography of divergence and the genetics of adaptation in White Sands lizards (52116)

**Stefan LAURENT**, Susanne PFEIFER, Matthieu FOLL, Matthew SETTLES, Sam HUNTER, Christine PARENT, Evelien Diepeveen, Jeffrey D. Jensen, Erika Bree Rosenblum.

*EPFL; University of Vienna; IARC, WHO; University of Idaho; University of California.*

The White Sands National Monument in New Mexico (USA) is a large geological formation of white gypsum dunes that accumulated following the end of the last Ice Age. The light soil color presented an evolutionary challenge for terrestrial animals relying on cryptic coloration to escape visually-hunting avian predators. In the lizard species Aspidoscelis inornata and Sceloporus undulatus, populations living in the white sands environment evolved a lighter dorsal color than populations living on the darker soils of the surrounding Chihuahuan Desert. The genetic basis of light dorsal color has been linked to non-synonymous mutations in the melanocortin-1 receptor (Mc1r) gene in both species but with opposite dominance patterns. Here we present new data describing genetic variation at the Mc1r locus and a large number of neutral loci across the genome in light and dark populations of both species. We show that patterns of variation are consistent with a strong selective sweep at the Mc1r locus in A. inornata but not in S. undulatus, which indicates that different histories are responsible for a similar adaptive phenotype. Furthermore, we infer the neutral demographic history of white
sands populations using a recently developed maximum likelihood framework and use these
results to compare the divergence process between light and dark populations.

# Differential regulation of melanin-related genes contributes to plumage coloration differences between morphs of the dark-eyed junco: a common-garden experiment (52681)

**Etienne Kornobis**, Paolo Ribeca, Mark Peterson, Mikus Abolins-Abols, Ellen Ketterson, Borja Milá.

*National Museum of National Sciences, CSIC, Madrid, SPAIN; The Pirbright Institute, Pirbright, Woking, UK; Dept. of Biology, Indiana University, Bloomington, USA.*

The dark-eyed junco (Junco hyemalis) represents a case of rapid diversification and is composed of at least six geographically structured, melanic morphotypes distributed across North America. Increased sexual dimorphism in the most recently diverged morphs is consistent with the role of sexual selection in driving fast plumage divergence. To understand the genetic basis of melanic coloration in the junco radiation, we carried out a controlled experiment using individuals from two highly distinct morphs (Oregon and slate-colored juncos) housed under common-garden conditions. We used RNA-seq to generate transcripts from growing feathers in four differently-colored body parts per morph, and examined the relative roles of genetic variants vs. differential regulation for pigmentation-related transcripts. We observed significant differential regulation of 161 transcripts among tissues from different morphs, five of which correspond to genes involved in melanin metabolism and storage (ASIP, TYR, TYRP1, OCA2 and PMEL). Moreover, we found differential regulation in other genes potentially involved in pigmentation, such as solute carrier genes and HOX genes. Overall, we find a relationship between the extent and pattern of eumelanin and pheomelanin deposition and differences in the regulation of a few genes, whereas SNP variants do not appear to be closely associated with color phenotypes.

# Host specificity in ticks (51575)

**A Raoul Van Oosten**, Erik Matthysen, Dieter JA Heylen, Kurt Jordaens, Thierry Backeljau.

*Evolutionary Ecology Group, University of Antwerp; Joint Experimental Molecular Unit, Entomology Section, Royal Museum for Central Africa, Tervuren, BE; Royal Belgian Institute of Natural Sciences—OD Taxonomy and Phylogeny, Brussels, Belgium.*

Ticks are a numerous group of hematophagous ectoparasites, and under natural circumstances most species infest one or only very few hosts. It is, however, largely unknown whether this is due to host specialisation or simply because ticks do not come into contact with other hosts. Therefore we investigate population dynamics and host specificity in the endophilic tick
Ixodes arboricola, whose life cycle is limited to tree holes and therefore infests only cavity-
nesting birds such as great and blue tits.

Our studies indicated that the life cycle of I. arboricola is strongly associated to that of its
hosts (Heylen et al, 2014, Parasitology) and that therefore its populations are genetically
differentiated even at small spatial scales (Van Oosten et al, 2014, Heredity). In two recent
studies, we found no differences in feeding performance of this tick among several cavity-
nesting birds but lower feeding performance on an unnatural host, while such differences
were absent in the generalist tick Ixodes ricinus. This provides evidence that ticks are host-
specialised and not just constrained by habitat (Van Oosten et al, 2014, Parasitol Res; Van
Oosten et al, in prep).

Poster session B - GEN 2000

# Effects of abiotic environment on the impact of a manipulative parasite on his host (51772)

Sophie Labaude, Frank Cézilly, Xavier Tercier, Thierry Rigaud.

University of Burgundy.

Many parasites are known to manipulate the behaviour of their intermediate hosts, supposedly
in a way to increase their probability of transmission to their definitive host. This
manipulation has been shown to vary in intensity at the intra-specific level. Considering the
important effect of trophically-transmitted parasites on the modulation of trophic chains,
understanding the variations of manipulation intensity and other infection parameters could
lead to a better understanding of the dynamic of an ecosystem as a whole.

I tested the effects of two abiotic parameters, temperature and quality of food, on the success of infestation and the intensity of parasite manipulation. I conducted experimental infestations using the acanthocephalan Pomphorhynchus leavis and its intermediate host, the amphipod Gammarus pulex. We followed the development of parasites and the survival of hosts, and conducted metabolism and behaviour measures on infected and control hosts.

Despite a positive effect on hosts survival and parasites development, food level did not affect
the intensity of manipulation. In addition, temperature was a strong determinant parameter for
the speed of development of parasites. Those results suggest that variations in abiotic
environment could have several effects on host-parasite interactions, leading to modifications
at the scale of the ecosystem.

Poster session B - GEN 2000

# Resistance evolution within a spatial framework (51773)

Layla Höckerstedt, Anna-Liisa Laine.

University of Helsinki; University of Helsinki.