# Getting to know ancient dogs through a genomic approach

Mahaut Goor, Blaschikoff Ludmilla, Guimarães Silvia, Serra Octávio, Simões Fernanda, Detry Cleia, Bustamante-Álvarez Macarena, Brehard Stéphanie, Radu Valentin, Boroneant Adina, et al.

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## ► To cite this version:

Mahaut Goor, Blaschikoff Ludmilla, Guimarães Silvia, Serra Octávio, Simões Fernanda, et al. Getting to know ancient dogs through a genomic approach. 10th ICAZ AGPM (International Council for Archaeozoology- Archaeozoology, Genetics, Proteomics and Morphometrics working group), Oct 2023, Munich (Allemagne), Germany. **hal-04740753**

HAL Id: hal-04740753

https://hal.science/hal-04740753v1

Submitted on 17 Oct 2024

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# GETTING TO KNOW ANCIENT DOGS THROUGH A GENOMIC APPROACH

10th ICAZ APGM - 10th-13th october 2023

Mahaut Goor, Ludmilla Blaschikoff, Silvia Guimarães, Octávio Serra, Fernanda Simões, Cleia Detry, Macarena Bustamante-Álvarez, Stéphanie Brehard, Adrian Bălăşescu, Hossein Davoudi, Marjan Mashkour, Valentin Radu, Adina Boroneant, Francisco J. Heras Mora, Regis Debruyne, Christophe Hitte, Catarina Ginja, Ana Elisabete Pires, Morgane Ollivier

contacts: mahautgoor@gmail.com;
morgane.ollivier@univ-rennes.fr;
ana.elisabete.pires@gmail.com

Studying ancient dogs' genomes offers valuable information about past populations, genetic ancestries, origins, and evolution. As humans shifted from hunter-gatherer to agropastoral societies during Neolithic, *Canis lupus familiaris* is the only domestic species that accompanied humans throughout these changes, and is therefore an excellent model to study the impact of human cultural shifts on their immediate environment.

**AIM:** This research explores the origins and diversity of Western Eurasian dogs from SouthWestern Europe to the Near & Middle-East dated from Palaeolithic (>33,000yrs BP) to the Middle Ages (750 AD).

**Sampling:** In order to study dogs' genetic diversity and their spread throughout Europe, a total of 240 dogs and wolves' mitochondrial genomes were obtained through Whole Genome Sequencing and a target-enrichment assay and applying specific bioinformatic pipelines. Among those newly-generated mitogenomes, **151** have a coverage **>90%**, with **a depth of >5X**.

A nuclear DNA enrichment assay targeting 4,003 specific Single Nucleotide Polymorphism (SNPs) [5] was performed on 174 samples. **In 74 samples, >2% of the total SNPs array with >3X coverage were present.**

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Mitochondrial DNA sequences are used to assess relationships intra & interspecies, mainly through the study of haplogroups, which are specific clusters of diversity. To our dataset, we added other genomes from present-day dogs [4][12][13][14][15][16] [17] and other canids as well as from ancient dogs [1][2][16][18][19][20][22]. Below is presented a preliminary Maximum Likelihood phylogenetic tree (Iqtree).

<figure><img src="image_4.png" /><figcaption>Preliminary cladogram (ML tree) of 466 ancient and present-day mitochondrial dog genomes</figcaption></figure>

The substitution model is TN+I+I+R4 (model Finder). All monophyletic clades were collapsed if they were from the same location (eg: Romania). Sample's nomenclature as: ID_Sites_Country_Hg/Sp_Date(direct or indirect) in BP.

**Purple** = **Wolves**, **Red**=**dog HgA**, **Blue**=**HgB**, **Yellow**=**HgC**, **Green**=**HgD**

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Map made using Qgis 3.32.3.

## SPECIFIC MARKERS UNDER EVOLUTION ?

We present preliminary results about 19 SNP markers associated with morphology and function. The study of such markers can ultimately help addressing relations between the evolution of morphological features of ancient specimens, evidence of adaptation under new environmental conditions and/or selection events. In dogs there are an increased prevalence of large-effect loci, nevertheless morphological features are often influenced by several loci and interpretation of molecular data should be cautious.

We were able to retrieve these markers for 62 ancient samples with a minimum coverage of 5X

### BODY SIZE

**IGSF1 gene:** Presence of derived allele (homozygous form) associated with an increase in body size-by-weight [6]. We demonstrate the first appearance of this variant around 6,575 BP in a dog from Romania from unknown Hg (no mitogenome data).

Derived alleles in GHR-1[11], HMGA2 [11], IGF1_15_41221438 [21] and IGF1_15_41229597 [21] genes are all associated with small body size in present-day dogs.

**GHR-1 gene:** earliest presence of the derived allele around 7,000 BP detected in a sample from Azerbaijan (HgB).

In contrast, the **reference allele** in homozygosity is associated with a large body size. Osteometric and odontometric data from a mandible (length, ~16cm) and a M1 tooth supports this finding for a Roman dog from Merida, Spain (HgA).

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**HMGA2**, IGF1_15_41221438 and IGF1_15_41229597 markers: Detected in Chalcolithic Romanian dogs (HgD ~6,300 BP) excavated in neighboring sites (Hârsova_tell and Popina-Bordusani). This result is in accordance with osteometric data showing that these specimens had a reduced body size (eg: length of the mandibule for sample 1831: 9.8cm).

### COAT COLOR

**ASIP [25] - TYRP - MITF genes:**

Presence of derived alleles as early as Neolithic Romania (HgA ~6,575 BP) and Greece (HgD ~6544 BP) showing apparent diversity in those genes, that can imply coat color variations.

### FUR

**FGF5 gene [24]:** we notice the first appearance of derived allele associated with long-hair trait in Israel around 1,600 BP as well as variant related with less hair shedding (MC5R) in the same dog (HgA).

**Hypothesis:** The ancient dogs bearing derived variants mostly belong to Hg A and D and could have been introduced in Europe following the Neolithic wave from the Near and Middle East, we hypothesize that these variants could have originated in those regions before being disseminated.

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Figure made using Google Sheets (pink = REF/REF, light red = REF/ALT, dark red = ALT/ALT, white = missing data)

<figure><img src="image_9.png" /><figcaption>Dogs genotypes for 19 targeted nuclear variants associated with morphology and function</figcaption></figure>

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Map made using mapchart.net and Google Sheets

Separation of countries are made according to the timing of the neolithic diffusion between countries

Mitochondrial and morphological data provide clues about the emergence and diffusion of some genomic variants in past dog populations. Ultimately, it will provide insights into human-canid relationships throughout chrono-cultural periods.

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## NEXT STEPS ?

* Recover radiocarbon dating for all crucial samples

* Refine the phylogenetic tree using Bayesien approach

* Test our hypothesis of different scenarios of phylogeographic diffusion using models and demo-genetic analysis

* Test for admixture between contemporaneous, ancient dogs and wolves populations in each studied regions to highlight possible links between phenotypes and their origin

[1]: Bergström, A et al., 2016, Science (352) [3]: Gopalakrishnan et al., Current Biology (28), 2018 [4]: Lindblad-Toh K et al., 2005, Nature - Kim KS et al., 1998, Mol Phylogen Evol [5]: Manin et al., 2023, submitted [6]: Plassais J., et al., 2019, Nature [7]: Skoglund et al. 2015 [8]: WOOF project [9]: Ollivier et al., 2018 [10]: Cadieu et al. 2009 [11]: Rimbault M, et al., 2013, Genome Res [12]: Björnfeldt et al., 2006 [13]: Webb Allard et al., 2009, J Forensic Sci [14]: Pang JF, et al., 2009, Mol Biol Evol [15]: Baranowska I, et al. 2009, Pgen [16]: Thalman O, et al. 2013, Science [17]: Strakova A, et al. 2016, Elife [18]: Ameen et al., 2019, Biological Science (286) [19]: Bergström et al., 2022, Nature 607 [20]: Botigué L, et al., 2017, Nat Commun 8 [21]: Sutter et al., 2007, Science et Gray et al. 2010, BMC Biol [22]: Leathlobair et al., 2018, Science 361 [23]: Mikkel-Holger S. Sinding et al., 2020, Science368, [24]: Dierks et al., 2013, Epub [25]: Berryere TG, et al. 2005, Genome

This project was supported by an ERASMUS+ grant, the "Société Française de Bioinformatique", CROC (CNRS-MITI) and Woof Projects - POCI-01-0145-FEDER-029545 & PTDC/HAR-ARQ/29545/2017 (Portugal)

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