In the current context of a global human-mediated environmental crisis, understanding which genes and
mechanisms are responsible for adaptation to different climates will enable predictions on how organisms will respond to a rapidly changing world. This is particularly important for the
honey bee, a key-stone species for ecosystem functioning and economy, which is facing increasing pressures from the effects of intensified land use, climate change, and the spread of pests and
pathogens. The aim of this work is searching for signatures of selection along the genome of 87 individuals using two different allele-environment association approaches.JC-G and DH are supported by Fundação para a Ciência e Tecnologia (FCT) through the
scholarships SFRH/BD/68682/2010 and SFRH/BD/84195/2012, respectively. This research
was funded by FCT and COMPETE/QREN/EU through the project PTDC/BIABEC/
099640/2008 and BiodivERsA-FACCE2014-91. Bioinformatic analyses were performed
using resources at the Uppsala Multidisciplinary Center for Advanced Computational
Science (UPPMAX)info:eu-repo/semantics/publishedVersio

Chávez-Galarza, Julio

Costa, Filipe Oliveira

Henriques, Dora

Neves, Cátia

Pinto, M. Alice

Rufino, José

Wallberg, Andreas

Webster, Matthew T.

Neves, Cátia J.

Biblioteca Digital do IPB

			Conference on Conservation Genomics 		PROGRAMME 		3 – 6 May 2016 | Campus de Vairão, Portugal | CIBIO-InBIO 			   Conference on Conservation Genomics Posters P01 An Vanden Broeck | Dispersal, gene flow and sibship analysis of Phengaris (Maculinea) alcon: implications for conservation P02 Ana Lino | Genetic consequences of habitat loss and fragmentation on vertebrates P03 Anne Andersen | Phylogeography of the large blue butterfly (Maculinea arion) P04 Bárbara Santos | Metabarcoding reveals trophic relations between the giant wall gecko, Tarentola gigas and other endemic species in an Integral Reserve in the Atlantic Ocean (Raso islet, Cape Verde) P05 Barbora Zemanová | National Animal Genetic Bank in the Czech Republic – infrastructure for conservation genetics and genomics research P06 Begoña Martinez-Cruz | Population genomics and the demographic history of Imperial Eagles P07 Catarina Silva | Insights into the evolution of phenotypic traits in house sparrows P08 Daniel Kleinman-Ruiz | Genomic tools for the conservation of Iberian lynx: A novel and curated set of genome-wide SNPs  P09 David Stanković | Detection of olm (Proteus anguinus) environmental DNA in karst groundwater P10 Dora Henriques | Searching for signatures of selection in the Iberian honey bee (Apis mellifera iberiensis) using allele-environment association approaches P11 Marketa Zimova | Interspecific competition, hybridization, replacement and adaption: a case study using whole genome sequencing in hares from Scotland P12 Federica Costantini | RAD genotyping to evaluate inter and intra specific levels of genetic structuring in Mediterranean biogenic reefs species P13 Filipa Martins | Towards Next-generation Biodiversity Monitoring: Improving Freshwater Quality Assessment using DNA Metabarcoding P14 Guy Colling | Changes at different altitudinal levels in the metabolic fingerprint of Arnica montana L. leaf extracts by 1H NMR-based metabolomics P15 Joana Paupério | InBIO Barcoding Initiative: moth reference database using Next Generation Sequencing P16 João Pedro Marques | A guide to reconstruct “true” de novo transcriptomes P17 João Queirós | Genome-wide associations identify novel candidate loci associated with genetic susceptibility to tuberculosis in wild boar P18 Johan Michaux | Using Next Generation Sequencing to characterize species diets: a study case with faeces of wild boar in the Spanish Pyrenees P19 Laura Daco | Climate change, plasticity and genetic variation in Anthyllis vulneraria P20 Lua Lopez Perez | The attack of the clones P21 Mafalda Costa | Phylogeography and population genetic structure of the European polecat (Mustela putorius) P22 Manuel Curto | A new approach of whole mitogenome sequencing and its application for phylogenetic analysis of East African white-eyes (Aves, Zosteropidae) P23 Manuel Curto | Mitogenomics using amplicon sequencing with Illumina MiSeq P24 Margarida Gonçalves | Population genomics across the distribution range of the sable antelope  0510152025303540ObtainedExpectedIn the current context of a global human-mediatedenvironmental crisis, understanding which genes andmechanisms are responsible for adaptation to different climateswill enable predictions on how organisms will respond to arapidly changing world. This is particularly important for thehoney bee, a key-stone species for ecosystem functioning andeconomy, which is facing increasing pressures from the effects ofintensified land use, climate change, and the spread of pests andpathogens.Dora Henriques1,2, Andreas Wallberg3, Julio Chávez-Galarza1,2, Cátia Neves1, José Rufino4,5 Filipe O. Costa2, Matthew T. Webster3,M. Alice Pinto1,4SEARCHING FOR SIGNATURES OF SELECTION IN THE IBERIAN HONEY BEE (APIS MELLIFERAIBERIENSIS) USING ALLELE-ENVIRONMENT ASSOCIATION APPROACHES1Centro de Investigação de Montanha (CIMO), Instituto Politécnico de Bragança, Campus de Santa Apolónia, Apartado 1172, 5301-855 Bragança, Portugal, 2CBMA, Centrode Biologia Molecular e Ambiental, Universidade do Minho, Campus de Gualtar, 4710-057 Braga, Portugal3Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden4 Instituto Politécnico de Bragança, 5301-857 Bragança, Portugal.5 Laboratório de Instrumentação e Física Experimental de Partículas, Campus de Gualtar, 4710-057 Braga, Portugal.INTRODUCTIONRESULTS AND DISCUSSIONOBJECTIVESThe aim of this work issearching for signatures ofselection along the genomeof 87 individuals using twodifferent allele-environment associationapproaches.METHODSGenome sequencingIllumina technology was used to sequence the whole genomes of 87 Iberian honey beescollected across three north-south transects in the Iberian Peninsula (Fig.1).The reads were mapped against the reference genome Amel_4.5 using BWA.Several steps of quality control were performed:Identify and mark PCR duplicates: PicardIdentify regions of poorly and inconsistently mapped reads: GATK45SNP calling: FreeBayesRemove SNPs with: quality below 50; depth below 123 or higher then 3000 present in less then37 allelesIn total the dataset is composed by 1,289,449 SNPs.Environmental dataThe environmental data were obtained from the Climatic Research Unit (www.cru.uea.ac.uk)and ArcGIS was used to manipulate the dataSearching for signals of selectionThe allele-environmental association methods LFMM and Samβada were used to search signalsof selection along the genome.The database PANTHER v.8.0 (protein annotation through evolutionary relationships)classification system (http://www.pantherdb.org/) was used to examine if adaptively evolvingloci were enriched for specific functional annotation .Figure 1: Map representing the distribution of  precipitation in May. The red spots indicates the samples position of the  individuals used in this study.OverlapingLFMM uniqueSamβadauniquelong 113 0 564lat 350 35 64prec1 143 21 105prec5 308 288 10prec8 79 272 58tmin1 57 19 11tmin6 5 2 4cld4 108 89 23cld7 8 26 3rh1 16 17 3rh3 19 92 14rh6 6 17 14ins4 20 48 28land 11 21 0Table 1. Number of overlapping SNPs detectedby the allele-environment approaches andnumber of genes detected just by Samβada(P-value < 0.0001) and LFMM (FDR<0,05) for eachenvironmental variable.Figure 2: Genome-wide distribution of significancevalues –log10(q-value) obtained by the allele-environment approach LFMM for the variables withhigher number of outliers a) longitude b) latitude c)precipitation in May c) precipitation in August. The redlines indicate FDR values of 0.05, 0.02 and 0.011 2 3 4 5 6 7 8 9 10 11 12 133 14 15 16Chromosomesa)c)b)c) A total of 1,416 (FDR<0.05), 360 (FDR<0.02),and 220 SNPs (FDR<0.01) were identified bythe LFMM approach (Fig. 2). Samβada detected a total of 2,145 SNPs witha P-value < 0.0001). Of the 1,416 outlier's SNPs detected byLFMM 814 were simultaneously detected bySamβada (Table 1) and these were located inor near 302 genes . The environmentalvariables that showed a greater number ofassociations with the 814 SNPs were long(113), lat (350), prec1 (143), prec5 (308), andcld4 (108) (Table1). The climate-driven candidate gene with moreSNPs associated is GB49881 (37), most ofthem (28) associated with prec1. GB49881 isonly 1,864 bp away from another candidateGB49882, which contained 15 SNPs. The homolog gene GB49882 in Drosophilamelanogaster encodes the protein SHAW,that is a member of shaker family of voltage-gated K+ channels. This protein is widelyexpressed in the nervous system, morespecifically in clock neurons and helps toregulate resting membrane potential incentral neuron. Parisky et al. 2008 haveshown that this gene plays an important rulein the regulation of the sleep circuit orcircadian clock in in Drosophila melanogaster. Of a total of 14 GO categories weresignificantly over-represented (threshold of P-value<0.05) (Fig. 3) We have an over-representation of the termsreceptor binding and receptor activity for themolecular function that are crucial for theorganism recognize the extracellularmessages and translate these signals intocellular response .JC-G and DH are supported by Fundação para a Ciência e Tecnologia (FCT) through thescholarships SFRH/BD/68682/2010 and SFRH/BD/84195/2012, respectively. This researchwas funded by FCT and COMPETE/QREN/EU through the project PTDC/BIA-BEC/099640/2008 and BiodivERsA-FACCE2014-91. Bioinformatic analyses were performedusing resources at the Uppsala Multidisciplinary Center for Advanced ComputationalScience (UPPMAX)Acknowledgements .We found nine genes related withinsecticide response, GB54460 waspreviously described by Chávez-Galarza etal. (2013) and Harpur et al. (2014). Fiveout nine genes are associated withlongitude. These could be due differencesin agriculture practice between thecountries.ReferencesChavez-Galarza J, Henriques D, Johnston JS, et al. (2013) Signatures of selection in the Iberian honey bee(Apis mellifera iberiensis) revealed by a genome scan analysis of single nucleotide polymorphisms. Mol Ecol 22, 5890-5907.Harpur BA, Kent CF, Molodtsova D, et al. (2014) Population genomics of the honey bee reveals strong signatures of positive selection on worker traits. Proc Natl Acad Sci U S A 111, 2614-2619Parisky KM, Agosto J, Pulver SR, et al. (2008) PDF cells are a GABA-responsive wake-promoting component of the Drosophila sleep circuit. Neuron 60, 672-682.*Figure 3: Enrichment of GO-Terms  related with biological and molecular functions (the molecular functions are marked by *).. Abbreviations : syst (system), dev(development), met (metabolic), proc (process), org (organism), reg (regulation), bio (biological). Some of the biological functions over-represented are important for thecellular response such as steroidmetabolism process, cell adhesion,neurological system process, nervoussystem development.Note: prec (precipitation), tmin (minimum temperature), cld (cloud cover)rh (relative humidity), ins (insolation)  and land (Land Cover) numerals 1 to 12 on front of each variable specify the month*

Searching for signatures of selection in the Iberian honey bee (Apis mellifera iberiensis) using allele-environment association approaches

In the current context of a global human-mediated environmental crisis, understanding which genes and mechanisms are responsible for adaptation to different climates will enable predictions on how organisms will respond to a rapidly changing world. This is particularly important for the honey bee, a key-stone species for ecosystem functioning and economy, which is facing increasing pressures from the effects of intensified land use, climate change, and the spread of pests and pathogens. In this study we used Illumina technology to sequence the whole genomes of 86 Iberian honey bees, collected across three longitudinal transects in the Iberian Peninsula. Then, we searched for signatures of selection
using two different methods (LFMM and Samβada), which combine genetic and non-genetic (environmental) data. The LFMM approach identified 2193, 449 and 275 SNPs for FDR values of 0.05,
0.02, and 0.01, respectively. The environmental variables associated with the highest number of SNP outliers were precipitation, latitude and longitude. The LFMM findings were concordant with those obtained with Samβada. Gene ontology analyses show that clusters with higher score are related with
immune system development and sensory organ development for the outlier genes associated with longitude. Precipitation is associated with genes related with biological regulation and
sequence-specific DNA binding. This represents the most comprehensive study of positive selection that integrates genetic information, and environmental variables performed so far in Iberian honeybees.info:eu-repo/semantics/publishedVersio

    Conference on Conservation Genomics  3 – 6 May 2016 | Campus de Vairão, Portugal | CIBIO-InBIO  ABSTRACT BOOK  List of Communications ConGenomics 2016 Conference on Conservation Genomics | 3 – 6 May 2016 | ESF - CIBIO-InBIO  - U Porto | Campus de Vairão | Vairão, Portugal 11 of 122 P04 Bárbara Santos | Metabarcoding reveals trophic relations between the giant wall gecko, Tarentola gigas and other endemic species in an Integral Reserve in the Atlantic Ocean (Raso islet, Cape Verde) P05 Barbora Zemanová | National Animal Genetic Bank in the Czech Republic – infrastructure for conservation genetics and genomics research P06 Begoña Martinez-Cruz | Population genomics and the demographic history of Imperial Eagles P07 Catarina Silva | Insights into the evolution of phenotypic traits in house sparrows P08 Daniel Kleinman-Ruiz | Genomic tools for the conservation of Iberian lynx: A novel and curated set of genome-wide SNPs  P09 David Stanković | Detection of olm (Proteus anguinus) environmental DNA in karst groundwater P10 Dora Henriques | Searching for signatures of selection in the Iberian honey bee (Apis mellifera iberiensis) using allele-environment association approaches P11 Marketa Zimova | Interspecific competition, hybridization, replacement and adaption: a case study using high throughput sequencing in hares from Scotland P12 Federica Costantini | RAD genotyping to evaluate inter and intra specific levels of genetic structuring in Mediterranean biogenic reefs species P13 Filipa Martins | Towards Next-generation Biodiversity Monitoring: Improving Freshwater Quality Assessment using DNA Metabarcoding P14 Guy Colling | Changes at different altitudinal levels in the metabolic fingerprint of Arnica montana L. leaf extracts by 1H NMR-based metabolomics P15 Joana Paupério | InBIO Barcoding Initiative: moth reference database using Next Generation Sequencing P16 João Pedro Marques | A guide to reconstruct “true” de novo transcriptomes P17 João Queirós | Genome-wide associations identify novel candidate loci associated with genetic susceptibility to tuberculosis in wild boar P18 Johan Michaux | Using Next Generation Sequencing to characterize species diets: a study case with faeces of wild boar in the Spanish Pyrenees P19 Laura Daco | Climate change, plasticity and genetic variation in Anthyllis vulneraria Posters ConGenomics 2016 Conference on Conservation Genomics | 3 – 6 May 2016 | ESF - CIBIO-InBIO  - U Porto | Campus de Vairão | Vairão, Portugal 81 of 122  P10   Dora Henriques Searching for signatures of selection in the Iberian honey bee (Apis mellifera iberiensis) using allele-environment association approaches Dora Henriques1,2, Andreas Wallberg3, Julio Chávez-Galarza1,2, Cátia Neves1, José Rufino4,5, Filipe O Costa2, Matthew T Webster3 and M Alice Pinto1,4  (1) Centro de Investigação de Montanha (CIMO), Instituto Politécnico de Bragança, Campus de Santa Apolónia, Apartado 1172, 5301-855 Bragança, Portugal, dorasmh@gmail.com jchavez@ipb.pt, catia.jose7@gmail.com, apinto@ipb.pt (2) CBMA, Centro de Biologia Molecular e Ambiental, Universidade do Minho, Campus de Gualtar, 4710-057 Braga, Portugal (3) Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden (4) Instituto Politécnico de Bragança, 5301-857 Bragança, Portugal. Laboratório de Instrumentação e Física Experimental de Partículas, Campus de Gualtar, 4710-057 Braga, Portugal. dorasmh@gmail.com  In the current context of a global human-mediated environmental crisis, understanding which genes and mechanisms are responsible for adaptation to different climates will enable predictions on how organisms will respond to a rapidly changing world. This is particularly important for the honey bee, a key-stone species for ecosystem functioning and economy, which is facing increasing pressures from the effects of intensified land use, climate change, and the spread of pests and pathogens. In this study we used Illumina technology to sequence the whole genomes of 86 Iberian honey bees, collected across three longitudinal transects in the Iberian Peninsula. Then, we searched for signatures of selection using two different methods (LFMM and Samβada), which combine genetic and non-genetic (environmental) data. The LFMM approach identified 2193, 449 and 275 SNPs for FDR values of 0.05, 0.02, and 0.01, respectively. The environmental variables associated with the highest number of SNP outliers were precipitation, latitude and longitude. The LFMM findings were concordant with those obtained with Samβada. Gene ontology analyses show that clusters with higher score are related with immune system development and sensory organ development for the outlier genes associated with longitude. Precipitation is associated with genes related with biological regulation and sequence-specific DNA binding. This represents the most comprehensive study of positive selection that integrates genetic information, and environmental variables performed so far in Iberian honeybees. 

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Searching for signatures of selection in the Iberian honey bee (Apis mellifera iberiensis) using allele-environment association approaches

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