Abstract The Arroyo Seco 2 site contains a rich archaeological record, exceptional for South America, to explain the expansion of Homo sapiens into the Americas and their interaction with extinct Pleistocene mammals. The following paper provides a detailed overview of material remains found in the earliest cultural episodes at this multi-component site, dated between ca. 12,170 14C yrs B.P. (ca. 14,064 cal yrs B.P.) and 11,180 14C yrs B.P. (ca. 13,068 cal yrs B.P.). Evidence of early occupations includes the presence of lithic tools, a concentration of Pleistocene species remains, human-induced fractured animal bones, and a selection of skeletal parts of extinct fauna. The occurrence of hunter-gatherers in the Southern Cone at ca. 14,000 cal yrs B.P. is added to the growing list of American sites that indicate a human occupation earlier than the Clovis dispersal episode, but posterior to the onset of the deglaciation of the Last Glacial Maximum (LGM) in the North America.

Citation: Politis GG, Gutiérrez MA, Rafuse DJ, Blasi A (2016) The Arrival of Homo sapiens into the Southern Cone at 14,000 Years Ago. PLoS ONE 11(9): e0162870. https://doi.org/10.1371/journal.pone.0162870 Editor: Michael D. Petraglia, Max Planck Institute for the Science of Human History, GERMANY Received: May 4, 2016; Accepted: August 30, 2016; Published: September 28, 2016 Copyright: © 2016 Politis et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability: All relevant data are within the paper and its Supporting Information files. Funding: Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT) (grants PICT-2010-1415, PICT-2013-0199). Competing interests: The authors have declared that no competing interests exist.

Introduction The current data from southern South America suggests Homo sapiens expanded into the Americas during a period earlier than the Clovis hunters of North America (older than ca. 11,500 14C yrs B.P.) [1]. Currently, Monte Verde II is considered by most archaeologists as a prime example of this pre-Clovis occupation [2,3]. New data from the Pampas region of Argentina, support the association of extinct Pleistocene fauna and cultural remains at the Arroyo Seco 2 site (AS2), dated to 12,170 14C yrs B.P. (13,814–14,147 cal yrs B.P.) [4]. The AS2 site is located just outside the city of Tres Arroyos, in the Pampa region of Argentina. It is an open-air archaeological site situated on a low lying knoll between a small temporary lake and a shallow creek (38°21'38" S, 60°14'39" W) (Fig 1 and S1 File). The AS2 site is one of three archaeological sites in the locality, and was discovered and test pitted by archaeology amateurs from Tres Arroyos in the early 1970s. The first systematic fieldwork began in 1977, including the excavation of primary burials, by Alberto Rex González [4,5]. From 1979 to the most recent excavations in 2015, a total of 77 units (~314 m2) were opened in the AS2 site, including shovel tests and 3 long trenches (S1 Fig). PPT PowerPoint slide

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larger image TIFF original image Download: Fig 1. The Arroyo Seco 2 site. (A) Geographic location of the AS2 site. (B) Digital Elevation Model (DEM) of the knoll and location of the excavation units (S1 Fig). (C) Photograph of central excavation units and trench. https://doi.org/10.1371/journal.pone.0162870.g001 The main objective of this paper is to present and evaluate the information regarding the earliest human occupation of the site (early to late Holocene occupations will not be discussed here). As a result of intensive biological, geological, and cultural processes, the AS2 site has a low stratigraphic resolution, as opposed to a high resolution which refers to strong preservation of artifacts and integrity of their spatial configurations [6]. The deposits of the sedimentary sequence are affected by dynamic process during or after their accumulation. These are subject to a variety of changes related to biotic and abiotic factors which affected original sedimentological characteristics and the contained archeological material. In spite of its low resolution, the AS2 site has significant archaeological characteristics for understanding aspects of the historical and evolutionary trajectories of hunter-gatherer societies. First, the site presents an ample temporal scale of human occupation from ca. 12,170 14C yrs B.P. to the 19th century. This extensive chronological dimension in a relatively short stratigraphic sequence (~2 m) of loessial sediments assigned to the La Postrera Formation [7] (Fig 2) has been one of the main causes of its low archaeological resolution. Second, there exists a high diversity of archaeological materials which provide a broad spectrum for detailed analysis (lithic, bone, ceramic, etc.) [4]. Third, the site presents an exceptionally varied and abundant number of human burials (50 individuals and counting), dated between 7805 ± 85 14C yrs B.P. and 4487 ± 45 14C yrs B.P. (n = 25 dates) [8]. PPT PowerPoint slide

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larger image TIFF original image Download: Fig 2. Generalized stratigraphic profile of the AS2 site. https://doi.org/10.1371/journal.pone.0162870.g002 With more than 30 yrs of excavation and interdisciplinary investigation at the AS2 site, it is not the lack of research or material remains, but the intrinsic characteristics of the site that make it a low resolution assemblage. In this sense, we present all the information in the most objective way, with both the certainties and uncertainties. The following paper will evaluate and discusses the general interpretations of the Pleistocene occupation at the AS2 site based on the AMS radiocarbon (14C) dates and the intensive re-analysis of the archeological material and site stratigraphy. Identifying the initial evidence of human occupation has been one of the central objectives of investigation at the site. This site has the potential to discuss two major subjects in American archeology: the early human expansion, and the interaction between hunter-gatherers and extinct Pleistocene fauna.

Geology The inter-sierra plains are developed in the southeast of the province of Buenos Aires, in the “Positive Bonaerense” [9] between the foothills of the Ventania and Tandilia Hill ranges (see Fig 1). Previously cited by Frenguelli [10] as a morphological sub-area called the "Pampa Inteserrana” (inter-sierra pampas) [11], this plain presents maximum heights of 200 masl, which gradually descends east towards the Atlantic Ocean. From a geomorphological point of view, the area where the archaeological site AS2 is located corresponds to the Acretional Loess Plateau Unit. The landscape is represented by an undulating relict plain, composed of continental Pliocene-Late Miocene Sub-cycle deposits which are capped by a thick calcareous duricrust [11]. During the late Pleistocene-Holocene Sub-cycle, loess deposits formed a continuous mantle across the Pampa Interserrana and the aeolian sands developed dunes in areas close to the sea [11]. While in low or depressed areas, fluvial-lacustrine deposits accumulated. The sedimentological analyses at the AS2 site have allowed the identification of four stratigraphic units, in concordance with the profile previously described by Fidalgo et al. [5] and Gentile [7], defined from the base to the top of the sedimentary profile as Z, S, Y, and X (Fig 3 and S2 File). PPT PowerPoint slide

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larger image TIFF original image Download: Fig 3. Geological context of the Arroyo Seco 2 site. (A) Stratigraphic profile from excavation Unit 70 (south wall). (B) Calcium carbonate (CaCO 3 ) and organic material percentages. https://doi.org/10.1371/journal.pone.0162870.g003 Unit Z: This unit lies unconformably on the calcrete crust developed in the diamictite (lithified sedimentary rock) consolidated sediments of the Late Miocene-Pliocene Sub-cycle [11]. Thickness of the Unit Z varies between 0.4 and 0.7 m. It has a pinkish gray loess color (5 YR7/2), composed of sandy silt. The sand content is below 50% of the total sample. The mean (Mz) value of 4.7 phi units is similar in both samples collected (bottom and top of the unit). The organic matter content is low; as such, this unit has the highest values of calcium carbonate for the entire stratigraphic sequence. The presence of abundant authigenic phosphate micro-nodules was recognized under the polarizing microscope. Powdery autigenic calcium carbonate is presented like a caliche (chalky zone). Towards the top of this unit, a layer between 0.12 and 0.17 m was identified with the highest concentration of carbonate, which makes coloration of this part of the column a bit clearer (see Fig 3). Strong cementation between grains does not exist, so the material has a loose consistency. Unit S: This unit is recognized as a sandy-brown (7.5 YR5/2) loess deposit with varying thickness between 0.2 to 0.25 m. It is composed of sandy silt with very fine sand modal fractions and a mean coefficient (Mz) value of 4.55 phi units. The sand fraction percentage is less than 50% of the total sample. At the macroscopic scale, there are traits of incipient pedogenesis. This unit also presents features of postdepositational precipitation of calcium carbonate, represented by abundant soft to indurate nodules which form a nodular area (nodular zone) with abundant rhizoliths in the upper part of the unit. Its limit with the overlying Unit Y is sharp and wavy. Trench excavations around the site (see Fig 1 and S1 File) revealed that the development of the Unit S is restricted to the middle and upper sections of the knoll. Unit Y: This unit is a homogeneous deposit of brown sandy loess (7.5 YR5/2) with variable thickness between 0.3 and 0.4 m. It consists of a silty sand deposit. The sand fraction percentage exceeds 50% of the total sample. No authigenic precipitates of calcium carbonate were found in the mass. The mean coefficient (Mz) value recorded was 4.12 phi units. Fresh volcanic glass shards are also present, but the increment of metamorphic lithic clasts (quartzite) minimizes the percentage. This unit passes in transition to the higher Unit X through a gradual and irregular contact. Unit X: This unit is represented by a dark brown (7.5 YR3/2) to dark grayish brown (2.5 YR4/2) loess, represented by a silty sand deposit. The mean coefficient (Mz) has a 3.98 phi value. This unit is affected by present-day pedogenic processes. The compositional and mineralogical analysis of the four stratigraphic units allowed defining a volcano-pyroclastic mineralogical association characterized by abundant volcanic glass shards and plagioclases (twinning and zoned). Volcanic and metamorphic lithic clasts are also present, with quartz as a minor component. Among the most abundant heavy mineral suites, 1–2% has been recognized as opaques, amphiboles, orthopyroxenes, clinopyroxenes and micas. The features defined here correspond to those of other loess units studied in the inter-sierra plains [12,13]. The major differences found between the units of the AS2 stratigraphy is in the relative frequency of volcanic glass (dense shards and pumice) clasts, from freshly to slightly altered states. The relative abundance of this component is higher in the Units Z and X. The stratigraphic sequence is composed of late Pleistocene Holocene fine aeolian deposits, represented by a loess mantle. It covers an undulating paleorelief composed of consolidated sediments (grouped under the collective name “Pampean Sediments”) of the Late Miocene-Pliocene Sub-cycle [11]. The sedimentary sequence is made up of four loess units, each one representing aeolian episodes separated by discontinuities. In three cases, erosive characteristics were found. In the case of the contact between the Units Z and S, it could only be defined as a stratigraphic unconformity, given the effects of meteoric diagenesis which has engaged these two units, masking the previous depositional features. The Units Y and X present a higher sand fraction content (> 50%) and are defined as loessial sands; while the Units S and Z are referred to as sandy loess, following the proposal of Zárate and Blasi [12,13]. The stratigraphic units were classified, based on the particle size of these deposits and the mineralogical association, mainly by the abundance of volcanic glass. Thus, Units Z and X can be classified as a vitroclastic sandy loess, and a vitroclastic loessial sand respectfully. Unit Y can be classified as a loessial sand, and Unit S as a sandy loess unit. Additionally, postdepositational reorganization by pedogenesis, and calcium carbonate precipitation by meteoric diagenesis, were observed affecting at least two of the stratigraphic units (Z and S). The first pedogenetic process is the result of moments of stability in the landscape, under wetter climate conditions, which favors the development of vegetation cover and a pedogenetical reorganization of the sediment. The second diagenetic process is more related to fluctuations in the paleo-water table, the groundwater chemical composition, the water depth from the surface, and the generated meteoric diagenesis produced at the expense of evaporation "per ascensum" (not pedogenic carbonates) [14–17]. The formation of certain morphologies of carbonates associated with pedogenic processes is not excluded. The precipitation of calcium carbonate in specific sectors of the stratigraphic sequence and under its different states (chalky, nodular, rhizolith zones) is reflected in the calcarerous profile. The carbonate related to the phreatic zone (chalky) and the vadose zone (nodular, rhizolith zones) can be differentiated in this profile. The first and second aeolian episodes (Z and S) likely occurred during the arid phases of the Late Pleistocene coinciding with the Last Glacial Maximum period. The last depositional event (Units S) took place after erosive episodes of unknown intensity and duration. The deposit of Unit S is possibly related to the deflated material of the Unit Z mixed with other local dust-storm sediments. Later, the existence of a stability interval is presumed based on the incipient pedogenetic reorganization of Unit S, which would have taken place during the Late Pleistocene-Early Holocene under more humid conditions [18]. The third aeolian episode occurred at the Early Holocene, with the accumulation of Unit Y; which according to its mineralogy could be related to accumulations of local sands by deflation of nearby source areas. A marked reduction in the aeolian activity is shown at the regional scale for this time period [18]. Furthermore, a rise in regional groundwater level would have also occurred, related to the marine ingression during the Holocene which peaked at 6000 yrs B.P. [19]. Around 5000–4500 yrs B.P., the influence of an arid climate [20] led to the precipitation of a chalky authigenic calcium carbonate in the pheatic zone, and nodular and rizoliths in the vadose zone [14]. The meteoric diagenesis, jointly affected the Units Z and S. The present shallow depth semiconfined aquifer in this area flows at 1.7 to 2 m under the ground surface. It is confined by low-permeability layers conformed of calcrete duricrust and indurated Pampean Sediments bellow the loess mantel. This substrate acts as an aquitard and as a regional source of alkaline surface and groundwater. According to the chemical characteristic of the present aquifer, it may produce dissolving and re-precipitation of the previous authigenic carbonates through time. It is possible that the calcium carbonate precipitated during the Middle Holocene in the Units Z and S are partially dissolved or re-precipitated as an effect of the current fluctuations in the phreatic. Finally, the Unit X is related to aeolian deposits connected to Late Holocene arid climate conditions, and the reworking by current agricultural activity.

Discussion Considering the data from radiocarbon dates (see Table 1), Pleistocene fauna death occurred in the AS2 site between ca. 12,200 and 7300 14C yrs B.P. Within this range, at least four different dates-of-death occurred: (1) Megatherium and Equus, ca. 12,170 14C yrs B.P. (13,975–14,152 cal yrs B.P.); (2) Toxodon, ca. 11,750 14C yrs B.P. (13,473–13,594 cal yrs B.P.); (3) Equus and Hippidion, ca. 11,182 14C yrs B.P. (13,035–13,100 cal yrs BP); and (4) Eutatus, ca. 7388 14C yrs B.P. (8175–8208 cal yrs B.P.). In order to discuss the human signal in the lower part of the stratigraphic Unit Y and S, where the narrowest association between Pleistocene fauna and lithic artifacts are observed, four lines of evidence need to be deliberated: 1) the spatial and stratigraphic association of archaeological material; 2) the selection of extinct fauna skeletal parts; 3) the concentration of Pleistocene species in a restricted area with positive landscape features; and 4) the human processing features in extinct fauna bones. 1) Spatial and stratigraphic association of archaeological material. Dated extinct Pleistocene bones associated with a variety of lithic artifacts and debris are scattered in the stratigraphic units Y and S, from 0.72 to 1.3 m below the surface. However; despite the close association of large sized flaked lithic artifacts, including a lutita and a rhyolite tool, with a concentration of lower limb bones of Megatherium, Hippidion, Equus, and Camelidae cf. Hemiauchenia (see above), no discrete horizons or paleo-surfaces have been detected (see Fig 4). The radiocarbon dates from extinct fauna in the lower part of the stratigraphic units Y and S demonstrate an important chronological dispersion; however the delimited horizontal and vertical spatial association in specific sectors of the site supports a close relation between Pleistocene fauna and lithic artifacts. What is important to make clear is that like any multi-component site with an ample chronological range and short stratigraphic sequence with low sedimentation rate, there is a high grade of complexity in its formation. While the action of variable and constant post-deposit processes has been studied in detail from the beginning of the systematic research at the site [40], understanding the formation processes of the archaeological assemblage in the site has been one of the central interests in the research projects and an important part of current study. Given the spatial and temporal resolution of the human burials, as well as the pre and post occupations related with faunal processing, the site represents a type of cumulative palimpsest (sensu Bailey [47]); however, one with discrete and rapid episodes of anthropogenic burials. In other words, the site represents successive episodes of occupations, superimposed one on top of the other without much loss of material, but with low resolution of spatial organization. The formation of the palimpsest is recurrent in the deposit of archaeological materials [48,49]; and in general, the palimpsest sites can offer very useful information regarding the formation processes [47], and eventually an estimation of the magnitude of the post-deposit disturbance can be made. With this in mind, the significance of the site, nor its importance in addressing some of the central topics of the historical trajectory and evolution of the Pampean indigenous populations, should not be underestimated [50]. 2) Selection of extinct fauna skeletal parts. Bone quantification shows a higher representation of the appendicular skeletal parts over the axial skeleton. Very few axial skeletal parts from identified species correspond to fragments of skull, vertebra, acetabulum and rib. This assemblage is comparable with deposit and transport strategies of hunter-gatherers as shown in ethnographic studies. For example, the Efe and Lese groups hunt elephants cooperatively in the northeast of the Ituri Rainforest in Africa. While there is a variable pattern in terms of the amount of bone transported to the campsite, some bone elements are never processed (and thus never transported): skull, mandible, scapula and innominate bone [51]. Another interesting ethnographic case is the Hadza from the savannahs of Africa, which is environmentally closer to the Pampa grasslands during the end of the Pleistocene, in terms of the diversity of fauna and size of prey. This ethnographic group transports the entire carcass from the kill site to the residential campsite, with the exception of the larger sized animals (giraffe, and elephant) [52]. Both types of methods used to transport carcasses can generate sites in where one of the principal activities was the processing of carcasses; deposits comparable to the Pleistocene bone assemblage at AS2 site. While it is possible that bone mineral density has influenced the representation of skeletal parts [53], it does not appear to be an important factor in the differential bone survival. There is a remarkable under-representation or even absence of certain bone elements with high density values such as teeth and cranial parts like tympanic annulus and occipital condyle. Most of the larger size megamammals, such as the Megatherium would have been hunted or scavenged close to the site, and its parts transported later. Given the body mass of this species (between 4 and 5 tons) [30], it would have been extremely difficult to transport the entire carcass and even challenging to transport complete hindquarters weighing between 600 and 750 kg, and forequarters weighing between 250 and 300 kg. Taking into consideration these values, the best hypothesis is that the Megatherium was hunted or scavenged near the site, the skeleton was butchered into smaller parts, and these units were then transported to their current location at the site. The larger bones were transported with portions of meat already removed, and the bone may have been used for other purposes such as bone quarrying [54,55]. The under-representation of some anatomical skeletal elements supports the human action, through the selection of determined parts of the carcass based on the size of the prey, as one of the principal agents in the formation of the Pleistocene fauna deposit. The anatomical representation of megamammals appears to be the result of differential transport of items based on yield of meat or bone. In this sense, the low frequency of dental material appears to be the consequence of discarding of the skulls at the hunting/scavenging site. 3) Concentration of Pleistocene species in restricted areas, with positive landscape features. In the ca. 100 m2 of the main excavation area, at the upper sector of the knoll, 11 taxa classifications have been assigned to Pleistocene fauna. During the temporal lapse between ca. 12,170 14C yrs B.P. and 11,180 14C yrs B.P. there is evidence of Megatherium, Toxodon, two extinct horse, and probably Glossotherium bone deposit. The probabilities that the Pleistocene fauna deposit are not of anthropic origin and that the site functioned as a place of natural death, a “bone trap” of Pleistocene species, are very low. Besides the fact that the bone assemblage appears to be the result of differential transport, the deposit is located on a positive landscape feature which is currently ~2.6 m above the level of the lake floor (see Fig 1). The reconstruction of the paleotopography indicates that this difference would have been even greater during the Late Pleistocene, a period when the shallow lake was active: between 4 and 5 m, and that a greater degree in the slope of the knoll would have existed [7]. The characteristic of the landscape does not favor the hypothesis that the knoll functioned as a type of “bone trap” [56]; which, in general, are found in depressions of the landscape like rivers and streams [57] or other bodies of water [58]. Furthermore, the limited vertical dimension of the extinct Pleistocene bone in the AS2 site assemblage suggests that the accumulation is discontinuous. In other words, if megamammal bones were being deposited naturally at the site, one would expect to find more bone specimens in the lower stratigraphic Unit Z, which formed during the Late Pleistocene. According to the regional geological record, the formation of the stratigraphic Unit Z may have started ca. 19,000 to 20,000 years BP [7] during which time the maximum diversity of mammals is recorded in Pampas [59]. Another aspect to consider is how, at a regional scale, these types of loess knolls do not behave as natural accumulators of bone. While there are currently no systematic regional paleontological studies which discuss this, local observations suggest that the bone concentration at the AS2 site is very high, and the variety of species represented is greater than any natural accumulations of bones usually found in the aeolian La Postrera Formation deposits in the Pampean region [60]. Therefore, the density and diversity of the Pleistocene bone remains in AS2, favors the action of hunter-gatherers as the principal accumulators of the deposit. 4) Human processing features in extinct fauna bones. The AS2 bone assemblage is highly fractured, the majority of which are classified as dry fractures. The frequency of these dry fractures is a consequence of various taphonomic processes which over long periods of time breaks down bone and cause fragmentation [40]. The identification of specimens with fresh fractures implies dynamic (human-hammerstone) force on the bone; although static (carnivore-bite) force cannot be discarded. However, while large carnivores with bit force similar to hyena and lions existed towards the end of the Pleistocene in southern South America [61], there is minimal evidence to suggest their interaction in the AS2 site. In reference to medium sized carnivores such as the puma (Puma concolor), further experiments are needed to confirm if this predator was in fact capable of producing similar sized notches. Therefore, while some ambiguity in fractured bone will always exist, and normally will require a more direct type of evidence (i.e, cut marks) to ratify the human origin, the information presented here helps to reduce much of this ambiguity and sustain the anthropic fracturing of the fresh fractured specimens. Moreover, one specimen of Equus neogeus bone (a proximal right radius fragment) dated at 12,170 ± 45 14C yrs B.P. presents strong evidence of anthropic fracturing with associated technological traits (see Fig 8). Finally, the absence of cut marks is likely related to the extensive natural modifications (weathering and chemical deposits) which mask or destroy this type of evidence [40]. On the other hand, the lack of marks may be the consequence of a butchering strategy of megamammals, since the amount of soft tissue would require little contact with the bones cortical surface. As demonstrated in actualistic studies, large quantities of meat from large sized prey can be filleted without making contact with the bone [62,63]. Use-wear analysis in the associated lithic artifacts in the AS2 assemblage is consistent with this hypothesis; as only three artifacts present evidence of bone contact [22].

Conclusions While each one of the four lines of evidence resumed and discussed previously could be explained in some way without a human intervention (see for example the discussion in Haynes [64]); the combination of the four lines of evidence suggests that the hypothesis more parsimony (Occam's razor) is that which situates the human action as a central causal factor, however not exclusive, in the formation of the extinct fauna bone assemblage and in its association with lithic artifacts. The information discussed here supports the following model of cultural formation for the early deposits at the AS2 site. The earliest human signal is situated in ca. 12,170 14C yrs B.P. (14,064 cal yrs B.P.) In this time period, occupants hunted/scavenged an extinct horse (Equus neogeus) and giant ground sloth (Megatherium americanum), probably along the border of the temporary lake (or another body of water) located near the site. The butchering began with the processing of the hind limbs in this plausible nearby location; which, when separated in smaller anatomical units (field-butchered units) were transported to the top part of the knoll where the processing was later finished. This area would have functioned as a short-term campsite/carcass processing site (sensus Fisher Jr.[51]). There, some anatomical units were disarticulated and the larger bones were broken using large stone tools. Some smaller artifacts were also used to cut and process the skins. The hunting/scavenging events of the early hunter-gatherers at the AS2 site were likely repeated several times. Temporal campsites were installed in the area for the butchering of Equus and Hippidon at ca. 11,180 14C yrs B.P. Given that these are smaller bodied animals (< 500 kg), the carcasses entered into the site more complete, and the anatomical units included elements like rib, teeth, and a piece of pelvis. During this period, other species of megafauna (Toxodon, Hemiauchenia and Glossotherium) were transported to the site, although the evidence of human agency is still vague for these taxa. Between these events, the AS2 site was likely visited by carnivores which scavenged some of the bone remains, though evidence for this activity is minimal. In conclusion, while there is a high degree of complexity in the formation of the AS2 site, the first evidence of human occupations dated between ca. 12,170 14C yrs B.P. and 11,118 14C yrs B.P. (14,064 and 13,068 cal yrs B.P.) are interpreted as a succession of transitory extinct mammal processing campsites; which formed a type of palimpsest and consequently cannot be differentiated among each other. The hunting of Pleistocene fauna in America is a heavily debated topic [65–68]. Generally speaking, in North America there is a wide agreement concerning the human predation of mammoth (Mammuthus spp.), mastodon (Mamut americanum) and bison (Bison spp.). Less conclusive is the exploitation of American horse (Equus sp.) [69], extinct camelid (Camelops sp.) [70] and the giant ground sloth (Magalonix jeffersonii) [71]. For South America, the exploitation of Pleistocene fauna has been proposed for mastodon [72], American horse (Hippidion saldiasi, Equus) [72–74], giant ground sloth Megatherium americanum [75], Doedicurus clavicaudatus [76] and possibly Hemiauchenia sp. [77] and Eutatus seguini [23]. With respect to Megatherium, AS2 is added to a list of sites which include the Campo Laborde site, in Argentina [75], and tentatively also to the El Vano site, in Venezuela [78] where Eremotherium rusconii remains were found. In the case of the two species of extinct horse, the evidence of AS2 adds to the wealth of important data that supports the human consumption of horse in various South American sites [72,73]. Finally, like other early sites, the first signal of human occupation in AS2, dated in ca. 12,170 14C yrs B.P. supports that Clovis was not the first human population in America (for a recent review see Madsen [2]). The second human signal in AS2, dated in 11,200 14C yrs B.P. is contemporary with the initial occupations of Clovis [1,79]. However, nor AS2, or other sites from Argentina and the rest of the Southern Cone [80–82] have ages various millennium older than Clovis, and despite recent claims [83], there is not yet a robust human signal firmly dated older than 13,000 14C yrs B.P. The evidence presented here supports a hypothesis of Homo sapiens dispersion in the Southern Cone before Clovis but after the Last Glacial Maximum (LGM). The AS2 site, along with other sites from Patagonia (such as Piedra Museo, Cueva Casa del Minero, and Cueva Cerro Tres Tetas, see discussion in Steele and Politis [25]), supports the idea of a human occupation previous to the transmission of the fish-tail projectile point technology; which in the Southern Cone is dated between 11,000 14C yrs B.P. and 10,000 14C yrs B.P. [82,84,85]. The absence of these projectile points in the AS2 site is in agreement with the chronology for this type of technology in the Southern Cone. The age of the AS2 site is consistent with this model, and that of a first pulse of entering of the peopling in America between 17–15 ka cal B.P. and posterior to the onset of the deglaciation of the Last Glacial Maximum (LGM) [86]. In this sense, the arrival of Homo sapiens into the Southern Cone at 14,000 years ago represents the last step in the expansion of modern humans throughout the world and the final continental colonization.

Acknowledgments The authors would like to acknowledge the work of numerous researchers who have participated in the investigations at the Arroyo Seco 2 site; in particular, for the research presented here we thank Marcela Leipus, Mónica Salemme, James Steele, and Thomas Stafford. Also to the municipality of Tres Arroyos and the staff at the Municipio de Tres Arroyos y Mueso Municipal “José A. Mulazzi”. Excavations were conducted as part of different research projects at the Instituto de Investigaciones Arqueológicas y Paleontológicas del Cuaternario Pampeano (INCUAPA-CONICET); the Agencia Nacional de Promoción Científica y Tecnológica, and the Facultad de Ciencias Sociales, Universidad Nacional del Centro de la Provincia de Buenos Aires (UNICEN).

Author Contributions Conceptualization: GGP MAG DJR AB. Data curation: GGP MAG DJR AB. Formal analysis: GGP MAG DJR AB. Funding acquisition: GGP MAG DJR AB. Investigation: GGP MAG DJR AB. Methodology: GGP MAG DJR AB. Project administration: GGP MAG DJR AB. Resources: GGP MAG DJR AB. Software: GGP MAG DJR AB. Supervision: GGP MAG DJR AB. Validation: GGP MAG DJR AB. Visualization: GGP MAG DJR AB. Writing – original draft: GGP MAG DJR AB. Writing – review & editing: GGP MAG DJR AB.