ARTICLE https://doi.org/10.1038/s41467-020-15907-4 OPEN Latitudinal gradient in dairy production with the introduction of farming in Atlantic Europe Miriam Cubas 1,2,3 ✉, Alexandre Lucquin 1, Harry K. Robson1, André Carlo Colonese1,4,5, Pablo Arias6, Bruno Aubry7, Cyrille Billard8, Denis Jan9, Mariana Diniz10, Ricardo Fernandes11,12,13, Ramón Fábregas Valcarce 14, Cécile Germain-Vallée9, Laurent Juhel15, Arturo de Lombera-Hermida14, Cyril Marcigny 16, Sylvain Mazet 7, Grégor Marchand 17, César Neves 10, Roberto Ontañón-Peredo 18, Xose Pedro Rodríguez-Álvarez19,20, Teresa Simões21, João Zilhão 10,22,23 & Oliver E. Craig 1 1234567890():,; The introduction of farming had far-reaching impacts on health, social structure and demo- graphy. Although the spread of domesticated plants and animals has been extensively tracked, it is unclear how these nascent economies developed within different environmental and cultural settings. Using molecular and isotopic analysis of lipids from pottery, here we investigate the foods prepared by the earliest farming communities of the European Atlantic seaboard. Surprisingly, we find an absence of aquatic foods, including in ceramics from coastal sites, except in the Western Baltic where this tradition continued from indigenous ceramic using hunter-gatherer-fishers. The frequency of dairy products in pottery increased as farming was progressively introduced along a northerly latitudinal gradient. This finding implies that early farming communities needed time to adapt their economic practices before expanding into more northerly areas. Latitudinal differences in the scale of dairy production might also have influenced the evolution of adult lactase persistence across Europe. 1 BioArCh, Department of Archaeology, University of York, Wentworth Way, Heslington, York YO10 5DD, UK. 2 Departamento de Historia, Universidad de Oviedo, C/Amparo Pedregal s/n, E-33011 Oviedo, Spain. 3 Sociedad de Ciencias Aranzadi, Zorroagagaina 11, E-20014 Donostia-San Sebastian, Spain. 4 Department of Prehistory, Edifici B Facultat de Filosofia i Lletres, Universitat Autònoma de Barcelona, Carrer de la Fortuna, Bellaterra, E-08193 Barcelona, Spain. 5 Institute of Environmental Science and Technology (ICTA), Edifici Z, Universitat Autònoma de Barcelona, Carrer de les columns, Bellaterra, E-08193 Barcelona, Spain. 6 Instituto Internacional de Investigaciones Prehistóricas de Cantabria, Universidad de Cantabria-Gobierno de Cantabria, Avd de los Castros s/n, E-39005 Santander, Spain. 7 INRAP Centre Archéologique du Grand Quevilly, 30 boulevard de Verdun, 76120 Le Grand Quevilly, France. 8 DRAC du Département Normandie Service Régional de l’Archéologie, 13 bis, rue Saint-Ouen, 14052 CAEN cedex 4, France. 9 Service Archéologie du Conseil Départemental du Calvados, 36 rue Fred Scamaroni, 14000 Caen, France. 10 Centro de Arqueologia da Universidade de Lisboa -UNIARQ- Alameda da Universidade, 1600-214 Lisbon, Portugal. 11 Department of Archaeology, Max Planck Institute for the Science of Human History, 07745 Jena, Germany. 12 School of Archaeology, University of Oxford, 1 South Parks Road, Oxford OX1 3TG, UK. 13 Faculty of Arts, Masaryk University, Arne Nováka 1, 602 00 Brno-střed, Czech Republic. 14 Facultad de Geografía e Historia, University of Santiago de Compostela, Praza Universidade, 1, E-15782 Santiago de Compostela, Spain. 15 INRAP, Centre Archéologique de Cesson- Sévigné, 37 rue du Bignon CS 67737, 35577 Cesson-Sévigné cedex, France. 16 INRAP, Centre Archéologique de Bourguébus, Boulevard de l’Europe, 14540 Bourguébus, France. 17 Centre de Recherche en Archéologie Archéosciences Histoire, UMR 6566 CNRS - CReAAH, Campus Beaulieu - Bât 24 - 25. 263 avenue du Général Leclerc - CS 74 205, 35042 RENNES Cedex, France. 18 Museo de Prehistoria y Arqueología de Cantabria y Cuevas Prehistóricas de Cantabria-Instituto Internacional de Investigaciones Prehistóricas de Cantabria, Ruiz de Alda, 19, E-39009 Santander, Spain. 19 IPHES, Institut Català de Paleoecologia Humana i Evolució Social, C/ Marcel.lí Domingo s/n- Campus Sescelades URV (Edifici W3), E-43007 Tarragona, Spain. 20 Área de Prehistoria, Universitat Rovira i Virgili (URV), Avinguda de Catalunya 35, E-43002 Tarragona, Spain. 21 Museu Arqueológico de São Miguel de Odrinhas. Av. Prof. Dr. D. Fernando de Almeida, São Miguel de Odrinhas, São João das Lampas, 2705-739 Sintra, Portugal. 22 SERP (Seminari d’Estudis i Recerques Prehistòriques; SGR2017-00011), Departament d’Història i Arqueologia, Facultat de Geografia i Història, Universitat de Barcelona, c/ Montalegre 6, E-08001 Barcelona, Spain. 23 ICREA, Catalan Institution for Research and Advanced Studies, Passeig Lluís Companys 23, E-08010 Barcelona, Spain. ✉email: miriam.cubas@york.ac.uk NATURE COMMUNICATIONS | (2020)11:2036 | https://doi.org/10.1038/s41467-020-15907-4 | www.nature.com/naturecommunications 1 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-15907-4 T he motivations for the introduction of farming in Europe areas Neolithic settlement clearly avoided Mesolithic territories and the nature of the earliest farming communities are key although for much of the region the degree of farmer/forager topics in European prehistory. Traditionally this issue has interaction is debated29. The DNA evidence points to some lim- been often reduced to polarised hypotheses of demic diffusion ited admixture between foragers and farmers although precisely versus acculturation to describe processes applicable to the whole when and where this occurred is still difficult to discern6,30. of Europe. More recently archaeologists have stressed the need to A second question is the degree to which early agro-pastoral consider regional variations as well as putting forward other more economies varied along the Atlantic European margin. The Neo- complex models, e.g. refs. 1–3. Our understanding of agricultural lithisation process of this region followed different rhythms and origins in Europe has also been reinvigorated by DNA analysis of dynamics related to both maritime and continental influences, e.g. human remains. These studies broadly support a Near Eastern refs. 27–29,31,32. In reality, the expansion of farming was relatively origin for Europe’s earliest farming communities directly asso- drawn out; pottery and domesticates appear some 1500 years later ciated with the beginning of food production, e.g. refs. 4–6. in the most northerly regions of the Atlantic Europe compared with Genetic studies, however, offer little detail regarding the social or the south. Thus, regional adaptations driven by local environmental economic drivers that led to the inception of the Neolithic, and factors and through interaction with different cultural groups may currently there are insufficient data to track demographic change be expected in the intervening periods. Potential economic differ- throughout all regions. Consequently, the regional development ences are partly indicated by the composition of faunal assemblages, of early agro-pastoral economies is less well understood. Indeed, which generally show a greater quantity of cattle remains relative to economic transformation may well have been independent of ovicaprines from the south to the north19. However, the scale of demographic changes and correlated instead to geographic and dairying by early farmers across Atlantic Europe compared with climatic factors as well as the types of interactions pioneer other regions of Europe is so far unknown. farmers had with various indigenous hunter–gatherer–fishers as To tackle these questions, here we provide new evidence from they moved in and around their territories. the organic residue analysis of 246 pottery sherds from 24 Early The widespread application of scientific methods, such as the Neolithic sites situated between Portugal and Normandy as well stable isotope analysis of human remains and organic residue as the Western Baltic (Supplementary Note 1). At each site, analysis of pottery, is beginning to highlight the varied nature of pottery representing only the initial phases of the Neolithic Europe’s earliest farming communities. A surprising finding is expansion were carefully chosen to create a representative corpus that dairy production, once thought to have developed much later often from small and fragmented ceramic assemblages. We in the Neolithic7, was a component of even the earliest Neolithic combine these data with previously published data from 39 economies and may even have been one of the motivations for archaeological sites15,16,18,20,33,34 to create a supra-regional per- ruminant domestication8. Although milk derived lipids have now spective on pottery use and animal exploitation by the earliest been identified in Early Neolithic pottery from Southern9,10, farmers (ca. 5500–3500 cal BC) (Fig. 1a, b). Together these data Central11–14 and Northern Europe15–18, the scale and intensity of cover regions with variable densities of Mesolithic sites, and a dairying in relation to meat production is still unknown. Indeed, latitudinal range that encompasses the entire Atlantic region from cumulatively such studies are beginning to highlight regional Portugal to Northern Scotland and the Western Baltic. Our patterns of variation in early animal husbandry9,10,12,19, provid- results reveal an increased frequency of dairy products in Early ing a new understanding of how early farmers adapted to a range Neolithic pottery assemblages correlating with their date and of environmental and cultural settings. latitude. One region that has received relatively little attention with respect to organic residue analysis is the Atlantic coast of Europe, with studies so far confined to Britain and Ireland18,20. Compared Results with other areas of Europe, this geographical unit has a particu- Molecular and isotopic results. A total of 246 pottery sherds larly high density of Late Mesolithic sites21, notably in Brittany, were analysed according to the well-established analytical pro- Denmark, along the Cantabrian coast of Spain and the Tagus and cedures as described in ‘Methods’. Interpretable amounts of lipids Sado estuaries of Portugal. Fish and shellfish were heavily (i.e. >5 µg g−135,36) were obtained from 234 samples (95% of the exploited in these highly productive marine and estuarine eco- total) (Supplementary Table 1). Excellent lipid preservation is in tones during the Late Mesolithic period, immediately prior to the accordance with previous studies from the British Isles, Ireland arrival of farming, e.g. refs. 22,23. How and why farming became and Northern Europe15,18,20,37 and contrasts with studies fully established along the Atlantic coastlines and estuaries undertaken in the Mediterranean and South-eastern Europe9,10. replacing hunter–gatherer–fisher subsistence practices has been a Molecular and isotopic data obtained from the acidified methanol source of much debate. One might expect that fish, shellfish or and solvent extraction procedures are summarised in Supple- marine mammals were processed in the earliest Neolithic pottery mentary Data 1 and 2. reflecting continuity in economic practices particularly at sites The distribution of fatty acids recorded in pottery from the located in areas close to where previous hunter–gatherer–fisher Iberian Peninsula and French sites, with relatively high amounts activity is recorded. However, paleodietary reconstructions of of C18:0, cholesterol and the occasional presence of mid-chain coastal Early Neolithic skeletal remains using stable isotope ana- ketones (C33, C35) and triacylglycerols (C42–C54), is consistent lysis appear to refute this hypothesis and show little evidence for with degraded animal fats in the majority of cases (Supplemen- the continued consumption of aquatic derived protein with the tary Data 1). By measuring the stable carbon isotope (δ13C) onset of the Neolithic23,24. Despite this, human remains from this values of the two main saturated fatty acids (C16:0 and C18:0), we period are extremely scarce in the coastal regions of the Iberian were able to distinguish the source of these animal fats with Peninsula and France, and this approach lacks the resolution to greater certainty. This robust approach has been widely used for rule out marine foods entirely25. There is isotopic evidence from identifying fats derived from a wide range of sources and is based an Early Neolithic site in the Northern Isles of Britain for sporadic on physiological differences in fatty acid biosynthesis between consumption of marine resources to supplement diets based lar- tissues37–39. As well as absolute ranges (Fig. 2), ruminant adipose, gely on domesticated plants and animals26. In some cases, ruminant dairy and porcine fats are distinguished according to Mesolithic coastal communities co-existed with neighbouring differences in the carbon isotope values between the two main farming villages27 resulting in technological exchange28, in other fatty acids (Δ13C = δ13C18:0 − δ13C16:0; Fig. 2 in ref. 37). In total, 2 NATURE COMMUNICATIONS | (2020)11:2036 | https://doi.org/10.1038/s41467-020-15907-4 | www.nature.com/naturecommunications NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-15907-4 ARTICLE a Estimated Neolithic arrival b Estimated arrival date (cal BC) Date S.E.M. (cal BC) 5300 300 35 41 42 5150 32-34 36 52-57 38-40 43 30-31 44-47 5000 55 55 29 37 48-49 250 28 27 4850 24-26 50-51 20 21 22-23 58-61 4700 19 50 4550 50 11-13 18 200 14-17 4400 10 Latitude 4250 45 4100 45 150 3950 8 9 7 3800 40 6 3650 40 1 1 100 N N 2 3500 3-5 1 3350 0 300 km 0 300 km 35 3200 35 50 –10 –5 0 5 10 –10 –5 0 5 10 c Estimated Δ13 C (‰) d Estimated Δ13 C (‰) S.E.M. 0.0 0.30 North England, Scotland and 0.28 Ireland Mean:– 4.58 –0.5 Std Dev: 1.13 0.26 55 55 –1.0 0.24 Western Baltic –1.5 0.22 Mean:–2.54 South England Std Dev: 1.53 0.20 Mean: –3.54 –2.0 50 50 Std Dev: 1.66 0.18 France & Channel Islands –2.5 Latitude 0.16 Mean: –2.94 North Spain Std Dev: 1.31 –3.0 0.14 Mean: –1.83 45 Std Dev: 1.65 45 0.12 –3.5 0.10 –4.0 0.08 40 40 0.06 –4.5 N N 0.04 Central-Southern Portugal –5.0 Mean: –1.10 0.02 Std Dev: 1.25 0 300 km –5.5 0 300 km 35 35 0.00 –10 –5 0 5 10 –10 –5 0 5 10 Longitude Longitude Fig. 1 Map of Atlantic Europe showing estimated dates for the Neolithic dispersal and fatty acid carbon isotope compositions from Early Neolithic pottery. a Output from SpreadR model showing estimated earliest arrival of the Early Neolithic and location of archaeological sites included in this paper (Supplementary Data 3). b Standard error of the mean for the arrival dates shown in a. c Output of AverageR model showing spatial estimates of Δ13C values of fatty acids based on the analysis of 647 individual potsherds (Supplementary Data 2). Estimates are limited to areas where the standard error of the mean is <0.30‰. More negative Δ13C values (i.e. ≤3.3‰) are typically associated with dairy fats, those between −3.3 and −1.0‰ are typical of ruminant fats and values ≥ −1‰ are typical of non-ruminant fats38. Mean and standard deviations of Δ13C values by region are shown (Supplementary Data 3 provides these data for each site). d Standard error of the mean for the estimated Δ13C values shown in c. Source data are provided as a Source Data file. 91.5% (n = 225) of the ceramic samples from Iberia, France and −0.67; p = 7.8312E−75) along the Western Atlantic coast from the Western Baltic provided sufficient lipids for analysis by GC- the Iberian Peninsula to Scotland with a greater proportion of C-IRMS. These data were combined with existing datasets from potsherds from higher latitudes containing dairy products Britain, Ireland and published data from the Western Bal- (Fig. 2c–e). Together, these data reveal different cultural and/or tic15,16,18,20,33,34, creating a dataset of 647 samples (Supplemen- ecological scenarios for pottery use. Stable carbon isotope values tary Data 2), which were then compared with modern authentic typical of marine oils were only observed in Early Neolithic reference samples of animal fats and oils (Supplementary Table 2, pottery from the Western Baltic (Fig. 2f) corresponding to the Fig. 2). presence of aquatic lipid biomarkers in these vessels16. In this region, there is a degree of continuation in pottery use from the Latitudinal gradient. Interestingly, the proportion of dairy and preceding hunter–gatherer–fishers of the ceramic Late Mesolithic ruminant carcass fats varies considerably between Early Neolithic Ertebølle culture16,33. In contrast, there was no evidence for assemblages across Atlantic Europe (Fig. 2a–f). There is a sig- aquatic biomarkers in any of the analysed vessels from sites nificant latitudinal gradient in the Δ13C values (Spearman = located on the Atlantic seaboard (Supplementary Data 1), despite NATURE COMMUNICATIONS | (2020)11:2036 | https://doi.org/10.1038/s41467-020-15907-4 | www.nature.com/naturecommunications 3 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-15907-4 δ 13C16:0(‰) δ 13C16:0(‰) −40 −35 −30 −25 −20 −15 −10 −40 −35 −30 −25 −20 −15 −10 −15 −15 a b Marine oils Marine oils −20 −20 δ13C18:0(‰) Porcine adipose fats Porcine adipose fats −25 −25 −30 −30 Ruminant Ruminant adipose fats adipose fats −35 −35 Ruminant dairy fats Ruminant dairy fats −40 −40 −40 −35 −30 −25 −20 −15 −10 −40 −35 −30 −25 −20 −15 −10 −15 −15 c d Marine oils −20 Marine oils −20 δ13C18:0(‰) Porcine adipose fats Porcine adipose fats −25 −25 −30 −30 Ruminant Ruminant adipose fats adipose fats −35 −35 Ruminant dairy fats Ruminant dairy fats −40 −40 −40 −35 −30 −25 −20 −15 −10 −40 −35 −30 −25 −20 −15 −10 −15 −15 e f Marine oils −20 Marine oils −20 δ13C18:0(‰) Porcine adipose fats Porcine adipose fats −25 −25 −30 −30 Ruminant Ruminant adipose fats adipose fats −35 −35 Ruminant dairy fats Ruminant dairy fats −40 −40 Fig. 2δ13C values of C16:0 and C18:0 n-alkanoic acids obtained from Early Neolithic pottery from the Atlantic coast of Europe and the Western Baltic (n = 647) (Supplementary Data 2). a Central-southern Portugal. b Northern Spain. c France and the Channel Islands. d Southern England. e Northern England, Ireland and Scotland. f Western Baltic. The 68% confidence ellipses are based on modern European authentic reference fats and oils (ruminant adipose fats, ruminant dairy fats, porcine adipose fats and marine oils) (Supplementary Table 2). Note that several of the vessels from the Western Baltic fall outside of the ellipses and as such a mixture of resources, including marine oils with ruminant adipose/dairy fats is envisaged. Source data are provided as a Source Data file. the use of highly sensitive protocols for their detection, and the may arise due to variability in the fatty acid content and isotope fact that such compounds have been reported in much older values of potential contributing food sources. The model was samples from a wide range of environments, e.g. refs. 12,16,40. applied to the median isotope values from each region (Supple- Apart from one sample from the site of Lesmurdie Road in mentary Table 3), excluding the Western Baltic where the data Scotland18, no aquatic products were found in Early Neolithic are bimodally distributed between marine and terrestrial values. pottery from Britain and Ireland using similar approaches18,20. This approach highlights a high degree of equifinality in inferring This result either points to significantly reduced fishing and pottery use based on the isotope data alone and it is often not shellfish gathering at the start of the Neolithic or, given the possible to definitively exclude any of the potential source fats. occasional finds of fish remains and mollusc shell at some Neo- Nevertheless, the model output (Fig. 3) shows that there is a clear lithic sites, e.g. refs. 18,25,41,42 that aquatic resources were pro- increase in the proportion of lipids derived from dairy compared cessed in other ways. with meat northwards across the study transect area, confirming the pattern observed from consideration of the Δ13C values alone. Bayesian mixing model. In order to highlight the potential effects of the mixing of different foodstuffs on the fatty acid stable Discussion isotope values, a Bayesian mixing model was deployed43. This Although we could not detect marine resources in any of the approach seeks to examine any biases in the interpretation that ceramic vessels with the exception of those from the Western 4 NATURE COMMUNICATIONS | (2020)11:2036 | https://doi.org/10.1038/s41467-020-15907-4 | www.nature.com/naturecommunications NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-15907-4 ARTICLE 100 a 100 b 100 c 80 80 80 % lipid by weight 60 60 60 40 40 40 20 20 20 0 0 0 Marine oils Ruminant Ruminant Porcine Marine oils Ruminant Ruminant Porcine Marine oils Ruminant Ruminant Porcine adipose dairy fats adipose adipose dairy fats adipose adipose dairy fats adipose fats fats fats 100 100 d e 80 80 % lipid by weight 60 60 40 40 20 20 0 0 Marine oils Ruminant Ruminant Porcine Marine oils Ruminant Ruminant Porcine adipose dairy fats adipose adipose dairy fats adipose fats fats Fig. 3 Bayesian modelled estimated proportions of lipids from different animal products in Early Neolithic pottery from different regions of Atlantic Europe (n = 563). a Central-southern Portugal. b Northern Spain. c France and the Channel Islands. d. Southern England. e Northern England, Ireland and Scotland. The output of the model provides estimations of % lipid by weight (y-axis) based on the median δ13C values of C16:0 and C18:0 n-alkanoic acids extracted from pots from each region with an uncertainty of 0.5‰ (Supplementary Table 3 and Supplementary Data 2). The boxes represent a 68% credible interval while the whiskers represent a 95% credible interval. The horizontal continuous line indicates the mean while the horizontal discontinuous line indicates the median. Source data are provided as a Source Data file. Baltic, our results show considerable geographic differences in the Along the Atlantic coast of Central and Southern Portugal, use of ceramic artefacts along the Atlantic coast of Europe. The farming arrived with the spread of ovicaprine based economies of results also provide valuable insights into animal management the Impressa–Cardial wares from the Mediterranean through the strategies for their primary and secondary products, which is straits of Gibraltar27,32 although other routes have been proposed significant given the poor preservation of faunal remains, largely for Northern Portugal and Galicia46. The analyses of Early due to the prevailing acidic soils in these regions (Supplementary Neolithic pottery assemblages from the Iberian sites point to Table 4). Where available, Neolithic faunal data show that ovi- economies oriented toward meat or mixed meat and milk pro- caprines tend to dominate Iberian assemblages, whereas cattle duction (Fig. 2a, b). Similarly, low frequencies of dairy lipids are were more important in Northern France, the UK and Denmark, observed in both Atlantic and Mediterranean Cardial pottery10 e.g. refs. 8,19,41,44. Based on the increased proportion of dairy supporting a mixed meat/milk economy during this early phase. residues associated with pottery from higher latitudes, we deduce Along the northern coast of Spain, farming and pottery were that intensive dairying is closely linked with cattle-based econo- introduced through the Ebro valley or across the Western Pyr- mies, while sheep and goats were exploited for both their meat enees 500 years later, ca. 5000–4500 cal BC29. The increased and milk, at least in the initial phases of the Neolithic. A similar frequency of dairy residues in this region may reflect a secondary association between cattle and dairying has been reported for the Neolithisation front derived from Mediterranean populations Early Neolithic of South-eastern Europe and the Near East9 and (Epicardial) who had developed more intensified dairying may have been important for the initial expansion of farming practices. beyond the Mediterranean climate zone45. In Atlantic France, pottery was obtained from sites north of the Although we compare Early Neolithic assemblages across Loire river (Brittany and Normandy) dating to ca. 5300–4900 cal Atlantic Europe, defined by the first centuries following BC. Here, there is a clear technological and stylistic influence of the appearance of pottery and domesticated animals, the final Rubané pottery that developed earlier in the Paris basin, sampling transect spans ca. 1500 years and follows the dis- which was linked with the cattle-based economies of the Central persal of farming, which appears ca. 5400/5300 cal BC in the European Neolithic47. At these sites, pottery was used to process south and ca. 3500 cal BC in the north (Fig. 1a, b). Therefore, both ruminant milk and meat (Fig. 2c), which is consistent with the frequency of dairy residues in pottery is also correlated the Early Neolithic cattle remains from Northern France and with the date that domesticates were introduced to any Central Europe that have mortality profiles indicative of a mixed given area. meat/milk economy48. Ruminant carcass fats were also identified NATURE COMMUNICATIONS | (2020)11:2036 | https://doi.org/10.1038/s41467-020-15907-4 | www.nature.com/naturecommunications 5 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-15907-4 in a small selection of ‘La Hoguette’ type potsherds from Alizay farmers who migrated to these new territories, the environments and Fontenay-le-Marmion, usually associated with indigenous they moved into and the response of local foragers. Economic foragers49, although their function could not be discriminated adaptations were needed before higher latitudes could be used for from other Early Neolithic pots from the region. food production, resulting in hiatuses in the Neolithic expansion. In Western Britain and Ireland, a coastal Atlantic origin for the Only in the Western Baltic, where a tradition of pottery use by Neolithic (ca. 4000 cal BC) has long been argued for on cultural Mesolithic hunter–gatherer–fishers was already established, were grounds31, and is supported by new genetic evidence6, although marine resources detected in Early Neolithic pottery. In the other the precise origin and influence of the various Neolithic cultures locations, there is little evidence that the exploitation of coastal of Northern France to the development of the British Neolithic is and estuarine environments had any influence on pottery use in debated, and there is little evidence for direct cultural transmis- the subsequent Early Neolithic. However, the frequency of dairy sion50. Indeed, the spread of farming was ‘delayed’ for at least half versus other terrestrial animal fats in pottery seems to be strongly a millennium following its arrival in adjacent regions on the influenced by latitude. Even in Britain and Ireland, where Early European continental mainland. Over 80% of the Early Neolithic Neolithic sites in the south and the north have similar dates, dairy pottery from Britain and Ireland were found to contain dairy fats were more frequent at higher latitude sites perhaps high- fats18, twice that observed in North-western France and there is lighting the importance of local environmental conditions or evidence for increased use of pottery for this purpose from nutritional requirements. Although Early Neolithic populations Southern England to Scotland and Ireland (Figs. 1c, d and 2d, e). in Western Europe were largely lactose intolerant62,63, variation Its importance is further supported by recent findings of cattle in the scale of dairying observed across the Atlantic transect may milk proteins in the dental calculus of Early Neolithic individuals have created a latitudinal gradient in selection pressure for adult from the UK51. lactase persistence (LP). A hypothesis supported by the high While the ecological limits on cereal production are often selection pressure for LP in North-western Europe inferred from explicitly linked to the dispersal of farming52, dairying also has its modern distribution64, although subsequent large-scale epi- specific requirements in terms of water availability, pasture sodes of migration, particularly in the Bronze Age63, may also quality and forage provision53 and requires considerable exper- have influenced the LP distribution. tise, especially in herd management. The reproductive timing of both cattle and sheep would have been further environmentally Methods constrained54, requiring cultural and biological adaptation to suit Sample information. Pottery sherds were selected from archaeological sites from local conditions. Isotope analysis of cattle from Bercy in Northern Portugal, Spain and France dated ca. 5500 and 3500 cal BC and the Western Baltic France shows that the extension of their birthing season and their margin, dated between ca. 3950 and 3300 cal BC (Fig. 1a, b). AMS radiocarbon (14C) dates attribute these sites to the Early Neolithic, i.e. contemporary with the early weaning to suit more intensified dairying was established at earliest introduction of domesticated animals and plants. Descriptions of individual least by the start of the 4th millennium cal BC55,56, corresponding sites and samples are provided in Supplementary Note 1. Representative number of to the arrival of cattle into Britain. Milk and dairy products samples were taken related to the total number of sherds or ceramic vessels brought nutritional benefits, such as sources of fat and vitamin D. (Supplementary Table 1). In total, 246 samples were collected according to their morphological and decorative variability and their spatial distribution at each site. The latter may have been particularly critical for populations moving to higher latitudes where less of this vitamin is produced in vivo due to the reduced exposure to sunlight57. Dairying may Lipid extraction and analytical protocol. Lipids were extracted by direct trans- esterification from 246 pottery sherds using an acidified methanol protocol35,65. also have been particularly important to farming populations Briefly, methanol (4 mL) was added to 1 g of pottery powder and sonicated for 15 struggling to establish cereal agriculture as they expanded into min. The suspension was acidified with sulfuric acid (H2SO4, 800 µL) and then new territories sub-optimal for this purpose. Indeed, the available heated for 4 h at 70 °C. Extraction of lipids was performed using n-hexane (3 × 4 archaeobotanical evidence shows that the degree of arable farm- mL). For quantification of lipids present in the resulting acid/methanol extracts (AEs), an internal standard (10 µL of hexatriacontane C36:0) was added to all of the ing in Britain varied considerably both locally and regionally samples. Most of the samples from Iberia and France (185/224) were also extracted during the Neolithic period58, and in some regions it may even using DCM:MeOH (2:1, 3 × 2 mL) using established protocols, e.g. ref. 66. The have failed following its initial introduction59. resulting total lipid extracts (TLEs) were dried under N2 and derivatized with N,O- Along the Western Baltic coastlines of Northern Germany, bis(trimethylsilyl)trifluoroacetamide (BSTFA) at 70 °C for 1 h. An additional Denmark and Southern Sweden, the first evidence for domes- internal standard (10 µL of hexatriacontane C36:0) was added to all samples prior to analysis. ticated animals and plants appears at ca. 4000 cal BC associated with the emergence of the Early Neolithic Funnel Beaker culture (TRB), e.g. ref. 60. At the same time there is a change in material Gas chromatography-flame ion detector. GC-FID was carried out on AEs using an Agilent 7890S gas chromatograph (Agilent Technologies, Cheadle, Cheshire, culture, notably from Late Mesolithic Ertebølle to TRB pottery, UK). Samples were re-dissolved in hexane and 1 µL was injected into the GC at but unlike other areas of Europe the exploitation of wild terres- 300 °C with a splitless injector, using helium as carrier gas (2 mL min−1). The GC trial game and fishing continued to be economically significant16. column was a polymide coated fused-silica DB-1 (15 m × 320 μm × 0.1 μm; J&W Nevertheless, even at these coastal sites, dairy products feature Scientific, Folsom, CA, USA). The GC oven was set at 100 °C for 2 min, then increased by 20 °C min−1 until 325 °C, where it was held for 3 min. among the commodities present in these earliest TRB ceramics. Previous studies suggest that they were often processed sepa- rately, particularly in small beakers, flasks and bowls61. The Gas chromatography-mass spectrometry. Analysis (AEs and TLEs) was carried out on an Agilent 7890A series chromatograph connected to an Agilent 5975 Inert variable use of pottery at coastal TRB sites encompassing both XL mass detector (Agilent technologies, Cheadle, Cheshire, UK). Samples were aquatic, dairy and other terrestrial resources may well be a con- injected with at splitless injector at 300 °C (1 µL) using helium as the carrier gas sequence of the interaction of farmers and indigenous foragers. (constant flow, 3 mL min−1). The spectra scanning window was between 50 and Further genomic analysis is needed to clarify the nature of such 800 m/z with a MS ionisation energy of 70 eV. A DB-5MS (50%-phenyl)- methylpolysiloxane column (30 m × 0.250 mm × 0.25 µm; J&W Scientific, Folsom, interactions but, here at least, indigenous and well-established CA, USA) was used. The temperature program was 5 °C for 2 min, 10 °C min−1 culinary practices seemed to have persisted well into the until 325 °C, followed by an isothermal hold for 15 min. Neolithic. In order to identify ω-(o-alkylphenyl) alkanoic acids and isoprenoid fatty Overall, our study shows that when the ‘Neolithic’ arrived at acids67,68, and to calculate the ratio of phytanic acid diastereomers69, analysis of the AEs (n = 241) was performed on a DB-23 (50%-cyanopropyl)-methylpolysiloxane different regions along the Atlantic coast there were different column (60 m × 0.250 mm × 0.25 µm; J&W Scientific, Folsom, CA, USA). Briefly, regional responses. We suggest these responses were influenced samples were re-dissolved in hexane and 1 µL was injected with a splitless injector both by the different economic and cultural traditions of the at 300 °C. Helium was used as the carrier gas with a constant flow of 3 mL min−1. 6 NATURE COMMUNICATIONS | (2020)11:2036 | https://doi.org/10.1038/s41467-020-15907-4 | www.nature.com/naturecommunications NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-15907-4 ARTICLE The ionisation energy of the MS was 70 eV and spectra were obtained in SIM mode εij ~ N(0, σε): residual error for individual j in site i. (74, 87, 88, 101, 105, 171, 213, 262, 290, 312, 318, 326, 346 m/z)40. The temperature SpreadR employs an extreme quantile approach by replacing the minima or first profile was 50 °C for 2 min, 10 °C min−1 until 100 °C, 4 °C min−1 to 140 °C, arrival with a 0.5%—quantile76. This quantile was estimated using Bayesian 0.5 °C min−1 to 160 °C, 20 °C min−1 to 250 °C, with an isothermal hold for 10 min. quantile regression using a spline smoother77. Estimated uncertainty, expressed as High temperature gas chromatography-mass spectrometry was performed on standard error of the mean, should be viewed as a low uncertainty estimate which the TLEs (n = 189) using the same apparatus and conditions as above. Mass may underestimate the true uncertainty77. spectra were obtained by scanning between 50 and 1000 m/z and analysis was performed with a HT-DB-1 100% dimethylpolysiloxane column (15 m × 0.320 Reporting summary. Further information on research design is available in mm × 0.1 μm; J&W Scientific, Folsom, CA, USA). The injector was maintained at the Nature Research Reporting Summary linked to this article. 350 °C. The temperature program was 50 °C for 2 min, 10 °C min−1 to 350 °C followed by an isothermal hold for 15 min. Data availability Gas chromatography-combustion isotope ratio mass spectrometry. Com- The authors declare that all data supporting this research are available within the paper, pound specific isotope analysis was undertaken on the AEs of 225 samples. Stable its Supplementary Information and Supplementary Data files. Source data underlying carbon isotope values of methyl palmitate (C16:0) and methyl stearate (C18:0), Figs. 1, 2 and 3 are provided as a Source Data file. EUROVOL dataset has been used in derived from precursor fatty acids, were measured by GC-C-IRMS, following the creation of Fig. 1 a, b (https://discovery.ucl.ac.uk/id/eprint/1469811/) and USDA existing procedures38. The instrumentation consisted of an Agilent 7890B series Food Composition Databases (https://ndb.nal.usda.gov/ndb/) have been used to obtain GC (Agilent Technologies, Santa Clara, CA, USA) linked by an Isoprime GC5 concentration of C16:0 and C18:0 n-alkanoic acids in the different products interface (Isoprime Cheadle, UK) to an Isoprime 100 (Isoprime, Cheadle, UK) and (Supplementary Table 2). to an Agilent 5975C inert mass spectrometer detector (MSD). Samples were re- dissolved and 1 µL was injected into DB-5MS ultra-inert fused-silica column (60 m × 0.250 mm × 0.25 µm, J&W Scientific, Folsom, CA, USA). The temperature Received: 18 September 2019; Accepted: 31 March 2020; program was 50 °C for 0.5 min, 25 °C min−1 to 175 °C, 8 °C min−1 to 325 °C, isothermal hold for 20 min. The carrier gas used was ultra-high purity grade helium (3 mL min−1). The gas flow eluting from the column was split into two streams. One was directed to the MSD for compound identification, while the other was directed through the CuO furnace tube at 850 °C to convert all the carbon species to CO2. Ion intensities (44, 45 and 46 m/z) of eluted products were recorded and References the corresponding 13C/12C ratios were computed. 1. Zvelebil, M. & Rowley-Conwy, P. Transition to farming in Northern Europe: a Data analysis (IonVantage and IonOS software; Isoprime, Cheadle, UK) made hunter gatherer perspective. Nor. Archaeol. Rev. 17, 104–128 (1984). comparisons between samples and a standard reference gas (CO2) of known 2. Gronenborn, D. in Going Over: The Mesolithic-Neolithic Transition in North- isotopic composition. The results are expressed in per mill (‰) relative to an West Europe (eds. Whittle, A. & Cummings, V.) 73–98 (Oxford University international standard, VPDB. 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Reprints and permission information is available at http://www.nature.com/reprints Author contributions M.C. and O.E.C. designed the research; M.C., A.L., H.K.R. and A.C.C. performed Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in the research; M.C., A.L., H.K.R., R.F. and O.E.C. analysed the data; M.C., A.L., H.K.R., published maps and institutional affiliations. A.C.C., R.F. and O.E.C. wrote the paper. P.A., B.A., C.B., D.J., M.D., R.F.V., C.G.V., L.J., A.L.H., C.M., S.M., G.M., C.N, R.O.P., X.P.R.A., T.S. and J.Z. provided samples and contextual information. All the authors were involved in reviewing the manuscript. 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