Journal of Evolution 74 (2014) 96e113

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The late Early Pleistocene human dental remains from Uadi Aalad and Mulhuli-Amo (Buia), Eritrean Danakil: Macromorphology and microstructure

Clément Zanolli a,*, Luca Bondioli b, Alfredo Coppa c, Christopher M. Dean d, Priscilla Bayle e, Francesca Candilio c, Silvia Capuani f, Diego Dreossi g, Ivana Fiore c, David W. Frayer h, Yosief Libsekal i, Lucia Mancini g, Lorenzo Rook j, Tsegai Medin Tekle i,k, Claudio Tuniz a,c,l, Roberto Macchiarelli m,n a Multidisciplinary Laboratory, The ‘Abdus Salam’ International Centre for Theoretical Physics, Trieste, Italy b Museo Nazionale Preistorico Etnografico ‘Luigi Pigorini’, Rome, Italy c Dipartimento di Biologia Ambientale, Università di Roma ‘La Sapienza’, Rome, Italy d Department of Cell and Developmental Biology, University College London, UK e UMR 5199 PACEA, Université Bordeaux 1, France f CNR-IPCF, Dipartimento di Fisica, Università di Roma ‘La Sapienza’, Rome, Italy g Elettra-Sincrotrone Trieste S.C.p.A., SYRMEP Group, Basovizza, Italy h Department of Anthropology, University of Kansas, Lawrence, USA i National Museum of Eritrea, Asmara, Eritrea j Dipartimento di Scienze della Terra, Università di Firenze, Italy k Institut Catala de Paleoecologia Humana i Evolució Social, Universitat Rovira i Virgili, Tarragona, Spain l Centre for Archaeological Science, University of Wollongong, Australia m Département de Préhistoire, UMR 7194, MNHN, Paris, France n Département Géosciences, Université de Poitiers, France article info abstract

Article history: Fieldwork performed during the last 15 years in various Early Pleistocene East African sites has signifi- Received 10 November 2013 cantly enlarged the fossil record of sensu lato (s.l.). Additional evidence comes from the Accepted 22 April 2014 Danakil Depression of Eritrea, where over 200 late Early to early Middle Pleistocene sites have been Available online 19 May 2014 identified within a w1000 m-thick sedimentary succession outcropping in the Dandiero Rift Basin, near Buia. Along with an adult cranium (UA 31), which displays a blend of H. erectus-like and derived morpho- Keywords: architectural features and three pelvic remains, two isolated permanent incisors (UA 222 and UA 369) Buia have also been recovered from the 1 Ma (millions of years ago) Homo-bearing outcrop of Uadi Aalad. Homo erectus/ergaster East Africa Since 2010, our surveys have expanded to the nearby (4.7 km) site of Mulhuli-Amo (MA). This is a Teeth fossiliferous area that has been preliminarily surveyed because of its exceptional concentration of External morphology stone tools. So far, the site has yielded 10 human remains, including the unworn crown of a Internal structure lower permanent molar (MA 93). Using diverse analytical tools (including high resolution mCT and mMRI), we analysed the external and internal macromorphology and microstructure of the three specimens, and whenever possible compared the results with similar evidence from early Homo, H. erectus s.l., H. antecessor, H. heidelbergensis (from North Africa), and modern . We also assessed the UA 369 lower incisor from Uadi Aalad for root completion timing and showed that it compares well with data for root apex closure in modern human populations. Ó 2014 Elsevier Ltd. All rights reserved.

Introduction

Geological, paleontological and paleoanthropological field * Corresponding author. research has been carried out in the Dandiero Rift Basin, northern E-mail address: [email protected] (C. Zanolli). Danakil Depression of Eritrea since 1994. This research has led to http://dx.doi.org/10.1016/j.jhevol.2014.04.005 0047-2484/Ó 2014 Elsevier Ltd. All rights reserved. C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 97 the discovery of over 200 late Early to early Middle Pleistocene sites The nearly 15 m-thick sedimentary succession of Mulhuli-Amo with widespread evidence of vertebrate faunal remains and lithic consists of deltaic and fluvial sediments. The lower interval (ca. 5 artefacts within the nearly 1000 m-thick fluvio-deltaic-lacustrine m) consists of a sandy Gilbert-type delta deposit. The middle in- sedimentary succession outcropping 100 km south of Massawa terval (5 m) is made of pedogenized muddy deposits with isolated (Abbate et al., 1998, 2004; Ghinassi et al., 2009; review in; Rook sandy fluvial channels responsible for the transport and accumu- et al., 2012). Thus far, in an area covering about 40 km2, cranial, lation of bone remains and stone tools in their basal parts. Finally, dental and postcranial human remains have been discovered in two the 5 m-thick upper interval of the succession consists of fluvial localities near the Buia village: Uadi (Wadi) Aalad and Mulhuli-Amo channelized gravelly sand capped with pedogenized mud. Even if (Coppa et al., 2012). most fossil remains from this site are spread on the eroded surface, The continental basin fill (the so-called Maebele Synthem) most of the bones and Acheulean artefacts occur in situ at the base consists, from bottom to top, of six lithostratigraphic units: fluvial of the third interval. Bukra sand and gravel, fluvio-deltaic Alat Formation, fluvial Wara The human fossil-bearing levels of Uadi Aalad and Mulhuli-Amo sand and gravel, lacustrine Goreya Formation; fluvio-deltaic Aro are remarkably similar in sedimentary facies and depositional sand; and alluvial fan Addai fanglomerate (Abbate et al., 2004; history (cf. Ghinassi et al., 2009; Rook et al., 2012), and are Papini et al., 2014). The Homo-bearing deposits belong to the up- considered to sample the same stratigraphic horizon belonging to per part of the 70e100 m-thick fluvio-deltaic Alat Formation the Alat. Besides the presence of reptile remains including croco- (Ghinassi et al., 2009). diles (Crocodylus niloticus), turtles (Pelusios sinuatus), and monitor The upper part of the Alat Formation hosts the top of the Jar- lizards (Varanus niloticus), the vertebrate assemblage at Mulhuli- amillo subchron (0.99 Ma), which occurs about 10 m below the Amo consists of the same mammal taxa represented at Uadi transition to the overlying Wara sand and gravel. The Matuyama- Aalad (i.e., Elephas, Hippopotamus, Kolpochoerus, Kobus, Pelorovis). Brunhes boundary (0.78 Ma) is located about 5 m above the base A total of 10 cranial and dental human remains, likely from two of the Aro sand unit (Albianelli and Napoleone, 2004). Confirmation adults and one immature individual, have been collected so far at of the magnetostratigraphically-based chronological setting comes Mulhuli-Amo. They consist of an isolated frontal fragment bearing a from fission track dating (Bigazzi et al., 2004) and mammal bio- thick right torus (MA 14) and eight parietal fragments (MA 64 and chronology (Ferretti et al., 2003; Martínez-Navarro et al., 2004, MA 88aef) associated with a temporal bone fragment (MA 89), all 2010; Rook et al., 2010, 2012). from a single adult cranium showing structural (thickness distri- All human remains discovered so far occur in the upper part of bution) and architectural features (proportions, curvature) closely the Alat Formation, where the transition from a deltaic to alluvial fitting the morphology represented by UA 31. Finally, the assem- setting is characterized by high frequency, potentially millennial- blage includes an isolated permanent lower molar crown (MA 93) scale, lake-level oscillations (units DL5, FL2a and FL2b in Ghinassi (Coppa et al., 2014). et al., 2009). Specifically, the concentration of fossil bones at the Compared with the Southeast Asian and Indonesian hypodigm base of the FL2b was associated with an increase in fluvial discharge (Indriati, 2004; Kaifu et al., 2005), the human dental assemblage that caused winnowing of the immediately underlying fossil- from chronologically well-constrained Early Pleistocene African bearing fluvio-deltaic deposits. As a whole, both the sedimentary sites (e.g., McDougall et al., 2012) is qualitatively and quantitatively record documenting the evolution of fluvio-deltaic and lacustrine relatively rich until about 1.4 Ma (millions of years ago) (cf. Wood, systems (Ghinassi et al., 2009; Rook et al., 2012), and the vertebrate 1991), while evidence from the later Early Pleistocene is scanty faunal assemblages predominated by taxa known for their water (Schwartz and Tattersall, 2003; Suwa et al., 2007; Brink et al., 2012). dependence (Delfino et al., 2004; Martínez-Navarro et al., 2004), The material we examine here thus enlarges this sample and con- clearly indicate a relatively open paleoenvironmental scenario tributes to partially fill the gap still existing between the dental characterized by the presence of moist grassed habitats adjacent to records available so far for African H. erectus sensu lato (s.l.) on one persistent water (Rook et al., 2012). hand, and on the other hand. First identified in the Uadi Aalad (UA) site, the 5e6 m-thick Building on the announcement (Abbate et al., 1998) and pre- Homo-bearing layer produced a virtually complete adult cranium liminary general description of the two incisors from Uadi Aalad, preserving the face (UA 31), two permanent incisors (UA 222 and collected in 1995 and 1997, respectively (Macchiarelli et al., 2004a), UA 369), and three pelvic portions (UA 173, UA 405, UA 466) and the molar crown from Mulhuli Amo, collected in 2011, here we (Abbate et al., 1998; Macchiarelli et al., 2004a; Bondioli et al., 2006). provide details of their external and internal morphology. More Compared with the Indonesian and Chinese Homo erectus sensu specifically, we compare the structure of the three Eritrean speci- stricto (s.s.) sample (review in Antón, 2003), as well as with African mens sampling H. erectus/ergaster with some earlier and later hu- specimens such as OH 9 and, to a lesser extent, KNM-ER 3733 (Baab, man taxa, particularly with the evidence from the early Middle 2008; Rightmire, 2013) and to the chronogeographically close Pleistocene North African sample of Tighenif, using a variety of calvaria from Daka, Middle Awash (Gilbert and Asfaw, 2008), UA 31 investigative approaches and analytical tools granting high reso- displays a blend of H. erectus/ergaster-like and derived morpho- lution access to their microstructure (Zanolli and Mazurier, 2013). architectural features more commonly found in Middle Pleisto- In doing so, we reveal for the first time the primitive versus derived cene specimens. These features include a high positioning of the nature of some tooth structural features in African H. erectus s.l. maximum parietal breadth, weak parietal keeling along the near the end of the Early Pleistocene and investigate their evolu- midline, slight parasagittal flattening, and from sub-vertical to tionary polarity at a macroregional scale. slightly downwards converging parietal walls, documenting extensive variation in late Early Pleistocene East African Homo Materials and methods (Macchiarelli et al., 2004a). Since 2010, the systematic survey of a fossiliferous area near to The three fossil teeth from the Buia area represent an upper left the exceptionally preserved A006 site (the so-called ‘handaxes lateral permanent incisor (UA 222) and a lower left central per- esplanade’) previously reported for its extensive concentration of manent incisor (UA 369) from the Homo site of Uadi Aalad (Abbate Oldowan and Acheulean lithic tools (Martini et al., 2004), has led to et al., 1998; Macchiarelli et al., 2004a), and a lower left M1/M2 the discovery of new fossil human remains at the Mulhuli-Amo (MA) crown (MA 93) from Mulhuli-Amo (Coppa et al., 2012, 2014; Zanolli locality, 4.7 km south of the UA Homo site (Coppa et al., 2012, 2014). et al., 2013). The incisors preserve the crown and root, while the 98 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 molar is represented only by the crown. This material is perma- of each specimen was produced using transparent epoxy resin nently stored at the Geo-Paleontological Laboratory of the National (Hardrock 554, Phase Inc.). Positive transparent araldite replicas Museum of Eritrea. were analysed using a stereomicroscope and the patterns of microstriations evaluated by scanning electron microscopy (SEM) Outer morphology (EVO 60 Leitz GmbH). For the latter analysis, the replicas were sputter coated with gold by means of an Emitech K550X system. The degree of occlusal wear was assessed according to Smith However, in contrast with the two incisors, MA 93 did not provide (1984), with description of the nonmetric crown topography any observable details. following Scott and Turner (1997), based on Arizona State Univer- sity Dental Anthropology System (ASUDAS) scores (Turner et al., High resolution magnetic resonance micro-imaging and root growth 1991). Maximum mesiodistal (MD) and buccolingual (BL) crown assessment diameters (in mm) were measured on the original specimens with Mitutoyo Digimatic calipers (to the nearest 0.1 mm) and then on To assess non-destructively the growth-related dentine micro- the unsmoothed virtual surfaces generated after segmentation of structures (daily von Ebner and long-period Andresen lines) in the microtomographic record of each specimen (see below). Since subfossil and fossil teeth (Bondioli et al., 2013), we developed a new the MD dimension of both incisors is affected by interproximal magnetic resonance microimaging (mMRI) analytical approach, wear, we limited the comparative analysis to the BL diameter. For based on previous successful high resolution MRI imaging of linear measurements, the Eritrean specimens were variably modern human teeth (Weiger et al., 2012). Indeed, techniques in compared with a number of fossil and recent samples (Table 1). magnetic resonance imaging (MRI) have advanced rapidly in the past two decades (Haack et al., 1999), and can now be applied in place of more destructive methods. Magnetic resonance imaging Crown microwear utilizes the magnetic moment exhibited by many naturally occur- ring nuclei, with the proton in the hydrogen nucleus being of Microwear texture analysis was attempted on the three speci- particular importance. Protons are highly abundant in all tissues mens following the standard procedures described by Mahoney and have high magnetic resonance sensitivity. Since the composi- (2006). The teeth were first cleaned with cotton soaked in tion of most biological tissue is dominated by water, the differences alcohol and molds of their crown surfaces were obtained using high in the water environment of adjacent tissue structures can be resolution silicone impression material (Zhermack Elite HD). A cast manipulated by MRI to effectively visualize tissue structure. More recently, magnetic resonance microimaging (mMRI) methods have Table 1 been developed and refined (Callaghan, 1995) to provide images Fossil and recent comparative samples used for crown linear measurements. characterized by a spatial resolution of approximately 100 mmor Samples Tooth References smaller. They have been particularly useful for investigating porous systems (Strange et al., 1993; Allen et al., 1997) and bones (De Santis H. habilis-rudolfensis UI2 Wood, 1991 fi m from East Africa (HHR) LI1 Stynder et al., 2001 et al., 2010; Wehrli, 2013). Speci cally, MRI techniques are able to LMs Wood, 1991; Leakey et al., 2012 retrieve high resolution information about the density, the mean African H. erectus/ergaster from UI2 Wood, 1991 diameter, and the distribution of microfeatures in porous systems. Kenya and Ethiopia (HEA) LI1 Stynder et al., 2001 Indeed, when the pores of a porous system, like the dentine, are LMs Wood, 1991; Suwa et al., 2007 filled with a liquid containing protons (usually water), it is possible H. erectus from Georgia (HEG) UI2 Martinón-Torres et al., 2008 LI1 Martinón-Torres et al., 2008 to obtain indirect information about the microstructural charac- LMs Martinón-Torres et al., 2008; teristics of this material from the signal of the liquid entrapped in Lordkipanidze et al., 2013 the microstructures. Currently, MRI is rarely used in dental diag- H. erectus from Java (HEJ) UI2 Grine and Franzen, 1994; nosis and research mainly because of the very short transverse Zanolli, 2013 LI1 Stynder et al., 2001 relaxation times of the mineralized tooth tissues. LMs Wood, 1991; Widianto, 1993; We performed a preliminary validation test on two perfectly Kaifu et al., 2005; Zanolli, 2013 preserved Neolithic permanent teeth (an upper incisor and a H. erectus from China (HEC) UI2 Wood, 1991 canine), after comparatively assessing their root microstructure LI1 Wood, 1991 (dentine tubules and Andresen lines) by means of mCT, mMRI and LMs Wood, 1991 H. antecessor from Atapuerca UI2 Bermúdez de Castro et al., 1999 histology (Bondioli et al., 2013). We applied this technique to Gran Dolina (HA) LI1 Bermúdez de Castro et al., 1999 visualize the dentine microanatomy and tentatively assess the LMs Bermúdez de Castro et al., 1999 growth pattern in UA 369. To do so, this well preserved lower left North African H. heidelbergensis from UI2 Martinón-Torres et al., 2008 central permanent incisor was immersed in a thin glass walled tube Tighenif and Rabat (HHNA) LI1 Stynder et al., 2001 LMs Wood, 1991 used to contain samples in nuclear magnetic resonance spectros- European H. heidelbergensis from UI2 Martinón-Torres et al., 2012 copy, which had been filled with distilled water at the temperature Atapuerca Sima de LI1 Martinón-Torres et al., 2012 of 18 C. Images of UA 369 were obtained by using Multi Slice Multi los Huesos (HHE) LMs Martinón-Torres et al., 2012 Echo (MSME) imaging sequences with repetition times (TR) equal Neanderthals (NEA) UI2 Voisin et al., 2012 to 380.0 ms, echo times (TE) equal to 3.5 ms, field of view (FOV) LI1 Voisin et al., 2012 2 LMs Voisin et al., 2012 equal to 14 14 mm , matrix 256 256, and number of scans Near Eastern early H. sapiens UI2 Vandermeersch, 1981 averaged (ns) equal to 1024. A total of 11 slices were selected of slice from Qafzeh (NEEHS) LI1 Vandermeersch, 1981 thickness (STH) equal to 200 mm to investigate the central zone of LMs Vandermeersch, 1981 UA 369 dentine. The in-plane resolution in each image slice was European early UI2 Frayer, 1977 m 2 humans (EUPH) LI1 Hillson et al., 2010 55 55 m . LMs Frayer, 1977 All measurements were performed on a Bruker 9.4T Avance-400 Recent (Holocene) humans (RH) UI2 Frayer, 1977 system located at the Physics Department of the ‘La Sapienza’ LI1 Frayer, 1977 University of Rome, operating with a micro-imaging probe (12 mm LMs Frayer, 1977 internal diameter bore) and equipped with a gradient unit C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 99

Table 2 Microtomographic-based inner structure Linear, surface, and volumetric variables used for assessing internal structural variation and molar crown tissue proportions. The three specimens were detailed using X-ray micro- Variables Definitions tomography (mCT) at the SYRMEP beamline (crown portion of the c The enamel area assessed on a buccolingual virtual tooth) and at the Tomolab station (whole tooth) of the Elettra section (mm2) Sincrotrone Trieste laboratory (Basovizza, Italy). The measurements b The dentine area under the enamel surface on a performed on the whole tooth were carried out according to the 2 buccolingual virtual section (mm ) following parameters: 100 kV voltage, 80 mA current, 4.0 s expo- e The enamel-dentine junction (EDJ) length on a buccolingual virtual section (mm) sition time per projection, and a projection each 0.25 , for the in- AET (¼c/e) The bi-dimensional average enamel thickness (mm) cisors; 130 kV, 61 mA, 4.5 s, and 0.15 for MA 93. Their final volumes RET (¼100* The scale-free, bi-dimensional relative enamel were reconstructed with an isotropic voxel size of 10.0 and 8.3 mm, AET/(b1/2)) thickness (Martin, 1985) respectively. Using Amira v.5.3 (Visualization Sciences Group Inc.) Ve The volume of the enamel cap (mm3) Vcd The volume of the coronal dentine (mm3) and ImageJ (Schneider et al., 2012), a semi-automatic, threshold- Vcp The volume of the coronal pulp (mm3) based segmentation was carried out following the half-maximum Vcdp The volume of the coronal dentine, including the height method (HMH; Spoor et al., 1993) and the region of inter- 3 coronal aspect of the pulp chamber (mm ) est thresholding protocol (ROI-Tb; Fajardo et al., 2002) taking Vc The total crown volume, including enamel, dentine, and repeated measurements on different slices of the virtual stack pulp (mm3) SEDJ The EDJ surface (mm2) (Coleman and Colbert, 2007). Vcdp/Vc The percent of coronal volume that is dentine and pulp Besides the pulp chamber volume of the incisors, 14 linear, (¼100*Vcdp/Vc) (%) surface, and volumetric variables describing internal structural ¼ 3D AET ( Ve/SEDJ) The three-dimensional average enamel thickness (mm) variation and tissue proportions were digitally measured (or 3D RET (¼100*3D The scale-free three-dimensional relative enamel AET/(Vcdp1/3)) thickness (see Kono, 2004; Olejniczak et al., 2008a) derived) on MA 93 (Table 2). Intra- and inter-observer tests for measurement accuracy run by two observers revealed differences less than 4%, which is similar to previous analyses (e.g., Macchiarelli characterized by a maximum gradient strength of 1200 mT/m, and et al., 2008; Bondioli et al., 2010; Zanolli et al., 2012). a rise time of 100 ms. Observations on the pulp cavity of the two incisors from Uadi Root extension rate was calculated from the mMRI images ac- Aalad were compared with similar evidence from H. heidelbergensis cording to the method detailed in Macchiarelli et al. (2006), Dean from Tighenif (isolated lower I1; Zanolli, 2011; Zanolli and (2009), and Dean and Cole (2013), which can be applied to both Mazurier, 2013), Neanderthals (Ehringsdorf G3 [upper I2] and fossil and extant human teeth. Continuous data were collected on Regourdou 1 [lower I1]; Macchiarelli et al., 2013; NESPOS Database, the lingual and labial aspects along almost the entire root length of 2013); and recent humans (original data). UA 369. After outlining the main visible growth incremental lines on To assess site-specific variation in their radicular dentine the image of this specimen, dentine tubules were traced from the thickness, ad hoc imaging techniques were used to virtually un- root surface just beneath the last formed enamel at the cervix (point roll the roots of the two incisors (15e85% portion of the total root A) to a second point 200 mm along the dentine tubules into root length) and to generate their morphometric maps (Bayle, 2008; dentine (point B). This thickness of dentine forms at a rate of 2.23e Bondioli et al., 2010; Bayle et al., 2011; Macchiarelli et al., 2.50 mm/day and takes ca. 80e90 days to form (Dean, 1998, 1999, 2013). For comparative purposes, we also considered the 2012; Dean and Cole, 2013). A line parallel with the orientation of following specimens: the upper I2 Javanese H. erectus specimen the growth markers was drawn from the end (point B) of the 200 mm MI92.2 (HEJ: Zanolli, 2011); the North African H. heidelbergensis segment back to the root surface (point C). The distance between isolated lower I1 from Tighenif (HHNA: Zanolli, 2011; Zanolli and point A and point C represents the length that the root had extended Mazurier, 2013); the two specimens Ehringsdorf G3 in ca. 80e90 days. An extension rate was calculated in mm/day for (upper I2) and Regourdou 1 (lower I1) (NEA: Macchiarelli et al., this distance along the root. This procedure was then repeated such 2013; NESPOS Database, 2013); and two recent human incisors that point C became point A in each calculation along the root length. (EH: original data). In H. erectus from Java (S7-37), direct measurements of daily In all three Eritrean specimens, enamel thickness topographic dentine increments in an upper M1 equalled 1.43 mm per day distribution was rendered through a 3D map generated using a over the first 50 mm, 2.46 mm per day between 50 and 100 mm, chromatic scale where thickness increases from thin (dark blue) to and 2.71 mm per day between 100 and 200 mm from the cement- thick (red) (Macchiarelli et al., 2008, 2013; Bayle et al., 2011). For dentine junction (CDJ) (Dean et al., 2001; Dean, 2009). The overall comparative purposes, similar cartographies have been generated mean daily rate over 200 mm of dentine formation from the CDJ for a selected number of specimens representing H. erectus from (point A to point B) was 2.4 mm per day in this upper M1, but no Sangiran (MI92.2 [upper I2] and NG0802.3 [lower M2]; Zanolli, equivalent data exist for anterior teeth in African H. erectus. 2011, 2013); North African H. heidelbergensis (isolated lower I1 Previously, a rate of 2.5 mm per day has been used to calculate 80 and Tighenif 2 [lower M1 and M2]; Zanolli, 2011; Zanolli and days formation for a 200 mm thickness of newly formed root Mazurier, 2013); Neanderthals ( D122 [upper I2], Regour- dentine in modern human teeth. However, Smith et al. (2007) dou 1 [lower I1], La Chaise de Vouthon S5 [lower M1], Krapina D10 have argued that in some teeth of Pan troglodytes dentine for- [lower M2]; Macchiarelli et al., 2013; NESPOS Database, 2013); and mation rates average closer to 2 mm per day at the cervix. Dean recent humans of European origin (original data). (2009) and Dean and Cole (2013) also observed rates of The results of the 2e3D virtual analyses performed on MA 93 w2.0 mm per day in some modern human anterior teeth. Because were variably compared with a number of fossil and recent samples it remains unclear what the exact range of daily dentine forma- (Table 3). tion rates is likely to have been in UA 369, we have adopted a Nonmetric features revealed by virtual imaging at the EDJ of MA cautious approach and have used both 80 and 90 day formation 93 were scored according to Skinner et al. (2008a) for the accessory times (equivalent to 2.2e2.5 mm per day) to provide what we feel cusps, Bailey et al. (2011) and Martínez de Pinillos et al. (2014) for is the most likely range of estimates for the time taken to form the trigonid crest pattern (see also Macchiarelli et al., 2006), and consecutive lengths of root along the CDJ. Martinón-Torres et al. (2014) for the talonid crest expression. 100 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113

Table 3 Fossil and recent comparative samples used for assessing 2e3D molar crown tissue proportions.

Specimens/samples References

H. habilis-rudolfensis from East Africa (HHR) Smith et al., 2012 South African early Homo (SAEH) Smith et al., 2012 Javanese H. erectus (HEJ) Zanolli, 2011 North African H. heidelbergensis from Tighenif (HHNA) Zanolli, 2011; Zanolli and Mazurier, 2013 Neanderthals (NEA) Olejniczak et al., 2008a; Kupczik and Hublin, 2010; Smith et al., 2012; Macchiarelli et al., 2013; NESPOS Database, 2013 North African Aterians (AFMH) Kupczik and Hublin, 2010 African-Near Eastern anatomically modern fossil humans Smith et al., 2012 (ANEFMH) from Morocco (Dar es-Soltane, Grotte des Contrebandiers, ), South Africa (Die Kelders Cave, Equus Cave) and Israel (Qafzeh) The European early Upper Paleolithic specimen Bayle et al., 2010 from Lagar Velho (EUPH) Recent humans (RH) Olejniczak et al., 2008a; Smith et al., 2012; original data

Geometric morphometric analyses its labial and, to a minor extent, lingual crown aspects revealed a number of fine striations covering the entire surfaces. The Comparative geometric morphometric analyses of the MA 93 scratches, which can be interpreted as microwear traces (cf. EDJ were performed on the unsmoothed reconstructed virtual Ungar et al., 2008; Krueger and Ungar, 2010), are mostly vertical surfaces by placing a total of seven landmarks on the apex of the or slightly oblique (85e90 range), sub-parallel and very thin (2e protoconid, metaconid, entoconid and hypoconid, and at each 3 mm) (Fig. 2A). However, some larger furrows show the intermediate lowest point between two dentine horns along the characteristics of trampling damage, with broad entry, dentine marginal ridge, except between the two distal horns (see rectangular section, and narrow end (Fiore et al., 2004). Zanolli et al., 2012). Because of the variable presence of the The uncorrected maximum MD crown diameter is 6.7 mm, hypoconulid, this latter cusp and the distal marginal ridge were while the BL diameter is 7.2 mm (contra Macchiarelli et al., 2004a; not considered (Zanolli and Mazurier, 2013). By using the package see Table 5). The root, which is complete with a closed apex dis- Morpho 0.24.1 (Schlager, 2013) for R v.3.0.1 (R Development Core playing a thin cementum deposit on virtual sections, exhibits lon- Team, 2013), we performed a generalized Procrustes analysis gitudinal grooves, the distal one being deeper. From the lowest (GPA) and a between-group principal component analysis enamel cervical point to its apex, root length is 18.8 mm. (bgPCA) based on the Procrustes shape coordinates (Mitteroecker The enamel shows no evidence of linear enamel hypoplasia, and Bookstein, 2011). For the specific purposes of this analysis, hypocalcifications or macro-defects, and no calculus or patholog- MA 93 was compared with a number of fossil and recent samples ical lesions are noticeable. While part of its crown has been (Table 4). removed by extensive wear, based on the preserved enamel UA 222 lacks shovelling (grade 0e1), an interruption groove and a distinct Results tuberculum dentale, but does show labial convexity (Fig. 1A) and also possesses a longitudinal lingual groove that is slightly deviated Description of the specimens mesially (cf. Martinón-Torres et al., 2012), reaching the wavy rim of the preserved occlusal-lingual margin. UA 222 (Uadi Aalad) This upper left lateral permanent incisor Microtomographic-based virtual sections reveal relatively thin (ULI2) from an adult individual is complete from crown to root enamel (maximum 0.66 mm). The EDJ perfectly mirrors the outer (Fig. 1A). Macroscopically, there is some evidence of weathering, morphology. Despite a slight collar constriction from its occlusal root etching and sediment scratching. From the mCT record, it also aspect to mid-root level, the pulp cavity shows a slightly labiolin- appears that the pulp cavity is filled with homogeneous fine- gually compressed ovoid outline. Conversely, its apical portion is grained sediment. The tooth shows advanced occlusal wear (stage mesiodistally flattened, with marked labiolingual expansion 4), with functional enamel loss and extensive emerging dentine (Fig. 3A). Its total volume is 30.7 mm3. The standardized morpho- creating a slightly labiolingual to mesiodistal oriented bevel. metric map of its virtually unrolled root reveals an area of abso- Interproximal wear is more marked distally, where the lutely thicker dentine (maximum 3.3 mm) on the labial aspect near facet almost reaches the CEJ. While the occlusal surface is the cervix, while thinner dentine is deposited on two vertical strips smooth, with rare pits and few microstriations, SEM analysis of running along the mesial and distal aspects (Fig. 4A).

Table 4 Fossil and recent comparative samples used for in the geometric morphometric analyses of the molar EDJ.

Samples Sites Lower M1 Lower M2 References

Javanese H. erectusa Sangiran NG0802.3 Zanolli, 2011 North African H. heidelbergensisa Tighenif Tighenif 2 Zanolli and Mazurier, 2013 Neanderthals Krapina KRD77, KRD79, KRD80, KRD1, KRD6, KRD10, NESPOS Database, 2013 KRD81, KRD105 KRD86, KRD104, KRD107 La Chaise-de-Vouthon S5, S14e7, S49, BDJ4C9 Regourdou Regourdou 1 Ehringsdorf Ehringsdorf I Recent humans 14 17 Original data

a Added a posteriori in the bgPCA analysis. C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 101

Figure 1. The upper left I2 UA 222 (A) and the lower left I1 UA 369 (B) from Uadi Aalad, and the lower left M1/M2 MA 93 from Mulhuli-Amo (C). For each specimen, the images show: the original (in lingual and mesial views for UA 222 and UA 369, in occlusal and buccal views for MA 93); a mesiodistal (left) and a labiolingual (right) virtual section for UA 222 and UA 369, and two virtual sections for MA 93 respectively passing through the mesial (upper) and the buccal cusps (lower); the microtomographic-based reconstruction of (from the left): the outer surface (the enamel in red), the dentine (yellow), the occlusal view (for the two incisors) and, uniquely for MA 93, the pulp cavity (cyan) (same views as the originals). b, buccal; l, lingual; m, mesial; o, occlusal. Scale bars, 1 cm. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

UA 369 (Uadi Aalad) This specimen represents a complete lower The uncorrected maximum mesiodistal diameter corresponds left central incisor (LLI1) from an adult. Serial virtual slices reveal a to 5.2 mm and the labiolingual diameter to 6.7 mm (Table 5). pulp chamber filled almost completely by a matrix, which occa- Similar to UA 222, no linear enamel hypoplasia, hypocalcifications, sionally causes contrast attenuation with the dentine (Fig. 1B). UA macro-defects, calculus deposits or pathological lesions are 369 also exhibits advanced occlusal wear (stage 4; Smith, 1984), noticeable on this specimen, and the preserved portion of the with a major mesiodistal to labiolingual orientation resulting in a crown shows no developed relief on its lingual and labial aspects, or weakly concave occlusal surface whose enamel rim is thicker shovelling. labially (0.9 mm versus 0.4 mm lingually). Interproximal wear is Measured on virtual sections, maximum enamel radial thick- more marked mesially. Under the stereomicroscope, the occlusal ness reaches 0.8 mm. Internally, the dentine crown surface casts surface is smooth, with rare pits and few microstriations. The the outer morphology. Also in this lower incisor the upper portion SEM imaging shows a microwear pattern similar to that seen in of the pulp cavity, whose total volume corresponds to 15.9 mm3, UA 222 (Fig. 2B). Indeed, mastication-related pits and scratches, shows a labiolingually compressed ovoid outline, followed by a as well as sediment-induced striae and furrows, are present on marked mid-root labiolingual expansion and a mesiodistally flat- both labial and lingual crown aspects, the former having tened apical root morphology (Fig. 3B). undergone some etching-like process that obscures part of the Well-developed longitudinal grooves run along both the mesial features, particularly near the occlusal border. The majority of the and distal radicular aspects, whose maximum length reaches vertical, sub-parallel and thin striae (2e3 mm) correspond to 16.3 mm. As revealed by its profile (Fig. 4B), thicker root dentine in microwear traces (Teaford, 2007; Ungar et al., 2008; Hlusko et al., this lower incisor appears on the cervical half of the labial and 2013), while few longer (1e2 mm), slightly thicker (3e10 mm) lingual aspects (maximum 2.9 mm), while the thinnest values are and randomly oblique scratches more likely result from the found along the apical third of the mesial and distal faces. abrasive action of variably sized sediment particles. Labially, close For magnetic resonance microimaging-based growth assess- to the CEJ, there are four long and relatively deep furrows of ment, a total of 20 consecutive measurements were possible on taphonomic origin, with smooth walls and rounded bottom. both lingual and buccal aspects of the root in UA 369 (Fig. 5A), but Although the general microfeature pattern does not differ from additional measurements became too difficult close to the root UA 222, some of the scratches occurring on both faces indicate apex beyond 12.5 mm on the buccal aspect, and 13.5 mm on the that UA 369 may have experienced slightly different taphonomic lingual aspect of the root. The overall mean buccal extension rate micro-dynamics. (n ¼ 20 estimates) was 8.0 mm per day (range 3.0e10.8 mm per day) 102 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113

specimen (Dean et al., 2001: Table 1). These estimates of crown height and crown formation time allow the plots for UA 369 to be visualized alongside the modern human sample and show that there is little effect (compared with Fig 5B) on how rates of root formation fall within the modern sample. Both plots (Fig. 5B and C) show that root formation in UA 369 could be reconstructed over approximately five years. MA 93 (Mulhuli-Amo) This is a nearly unworn lower left first or second permanent molar crown (LLM1/M2) of a juvenile individual (Fig. 1C). The buccal and lingual surfaces are slightly etched from roots and sediment, resulting in a coarse enamel texture. On the distolingual surface, a shiny area of better preserved enamel mimics an interproximal facet, but its lingually displaced position and its broad extension (larger than the mesial interproximal facet) support a taphonomic, non-functional origin. While some shallow furrows are visible on the lingual aspect, the presence of linear enamel hypoplasia cannot be confirmed here. Conversely, some hints of widely spaced perikymata are still visible in the centre of the distal face, where the enamel remains least worn. In this position, there is indeed a weak expression of enamel hypoplasia, supporting a similar conclusion for the lingual aspect. The hypoplasia would have occurred during crown formation. The rather squat crown reaches 6.1 mm in height at the meta- conid and 6.2 mm at the protoconid. It shows a rectangular outline with a length (MD) of 13.9 mm and a breadth (BL) of 11.9 mm (Table 5). A slightly polished wear facet on the protoconid and two smaller facets on the entoconid and on the hypoconulid indicate that the tooth had begun functional occlusion, but in the earliest stage (stage 1). A small, flat and ovoid interproximal mesial contact facet (2.9 2.3 mm) is present, but there is no distal facet. The mesial wall converges cervically, while the opposite is true buccally, the lingual and distal walls being relatively vertical. The sub- horizontal cervico-enamel line is almost completely preserved and the fracture plane in the inferior part appears to follow the incremental mineralizing front, which suggests that the tooth was at the crown complete stage of formation. There are five main cusps, including a medium-large hypo- conulid (C5; stage 4), plus a small tuberculum intermedium (C7; stage 2), but no tuberculum sextum (C6). Relative cusp size is Figure 2. SEM micrographs of the labial aspect of UA 222 (A) and UA 369 (B). Mostly protoconid > metaconid > hypoconid > entoconid > C5 > C7. The vertically-oriented thin microfeatures are visible on the central portion of the UA 222 occlusal grooves connect in a þ fissure pattern. Compared with the crown (A), where the two larger and deeper furrows crossing near the centre of the incised distal marginal ridge, the mesial margin is not crossed by image result from sediment abrasion. Numerous thin microfeatures are also present on the centre-mesial area spot sampling the UA 369 crown (B), together with few oblique the central groove and is proportionally thicker and higher, forming sediment traces and an etching damage on the upper right corner of the image. Scale a deep elongated mesial fovea (stage 3). Immediately behind the bars, 100 mm. fovea, an uninterrupted sharp mid-trigonid crest (score 1A) links the two mesial cusps. Two distal crests depart from the protoconid and the mean lingual extension rate was 7.8 mm per day (range 4.9e and the metaconid, respectively. They converge in the trigonid 10.0 mm per day). In both roots there was an early peak in root basin, but are interrupted in the centre by the main groove. Because extension rate (10.0 and 10.8 mm per day, respectively) between they do not connect, the latter can be interpreted as an incomplete 1.32 and 1.48 years post enamel completion. distal crest. Between the mid-trigonid crest and the distal trigonid- These results (Fig. 5B) are presented first as cumulative plots of like feature, a short crest originating from the metaconid angles increasing lingual and buccal root length against root formation towards the occlusal basin. No expression of a deflecting wrinkle is time compared with data for a sample of 44 modern human incisor found. The protostylid is absent and the intercuspal groove is tooth roots taken from Dean and Cole (2013). A second plot (Fig. 5C) relatively shallow. puts root growth into context with crown growth for illustrative At the EDJ level, MA 93 exhibits a modestly elevated topography, purposes. Root formation in UA 369 was compared with data for with relatively low but sharp dentine horns systematically under- crown growth in the same modern human sample (Dean and Cole, lying the blunter outer cusps, even at the level of the metaconulid- 2013). It was assumed that the unworn crown height in UA 369 type C7 (Skinner et al., 2008a), which is located on the distal would have been close to that of other unworn lower incisors shoulder of the metaconid dentine horn and is slightly better attributed to H. erectus s.l., a good example of which is the lower expressed than at the outer surface. While on the occlusal surface central incisor of KNM-WT 15000 that measures 9.75 mm. Using the mid-trigonid crest is complete but slightly dipping at the this tooth to provide an estimate of the crown formation time in UA sagittal sulcus level, its EDJ expression is continuous and of regular 369 suggests that it would have taken close to 2.83 years to com- height (grade 3; Bailey et al., 2011), originating slightly mesial from plete crown formation based on a nine day periodicity between the metaconid dentine horn and joining the protoconid dentine adjacent long-period striae of Retzius in the enamel of the Eritrean middle lobe segment (middle-mesial configuration). Together with C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 103

Table 5 Crown dimensions (MD and BL diameters, in mm) of the two permanent incisors from Uadi Aalad (UA 222 and 369) and the permanent molar from Mulhuli-Amo (MA 93) compared with some Pleistocene and recent human specimens/samples.

Specimens/samples UI2 LI1 LM1 LM2

BL BL MD BL MD BL

UA 222 7.2a UA 369 6.7 MA 93 13.9 11.9 13.9 11.9 HHR Mean (s.d.) 6.6 6.8 14.1 0.9 12.2 0.9 15.2 1.2 13.5 1.0 Range 6.0e7.9 6.6e7.0 12.8e15.2 10.6e13.2 14.0e18.3 11.7e15.1 n 4219201818 HEA Mean (s.d.) 8.3 6.6 12.8 0.7 11.3 0.9 13.4 0.9 12.1 0.7 Range 8.2e8.5 6.3e6.8 11.9e14.2 10.6e12.8 12.2e15.2 11.4e13.1 n 328888 HEG Mean 6.9 6.7 13.3 11.8 13.0 11.5 Range e 6.0e7.3 13.0e13.9 11.2e12.6 12.3e14.2 10.7e13.1 n 145555 HEJ Mean (s.d.) 7.3 6.2 12.8 0.9 12.2 0.8 12.4 1.4 11.6 1.3 Range 7.0e7.7 5.8e6.5 11.2e14.8 10.8e14.2 10.1e13.8 9.5e13.2 n 3214141515 HEC Mean (s.d.) 8.1 6.4 0.4 12.6 1.1 11.9 0.9 12.3 0.5 11.8 0.5 Range 8.0e8.2 5.8e6.8 9.9e14.1 10.1e12.8 11.3e13.1 11.1e12.7 n 37121288 HA Mean 8.2 7.6 12.2 11.8 13.5 12.0 n 111111 HHNA Mean 8.5 6.6 13.2 12.7 13.1 13.0 Range ee12.3e14.0 12.5e13.0 12.3e14.0 12.2e13.5 n 113333 HHE Mean (s.d.) 7.7 0.3 6.5 0.3 11.2 0.3 10.4 0.5 11.0 0.2 10.2 0.7 Range 7.3e8.3 5.9e7.2 10.3e12.2 9.5e11.6 9.4e12.2 8.4e11.5 n 27 29 41 40 41 41 NEA Mean (s.d.) 8.7 0.6 7.6 0.4 11.9 0.9 11.2 0.7 12.2 0.9 11.3 0.7 Range 7.8e9.9 6.9e8.2 10.8e13.6 9.7e12.9 10.5e14.0 9.8e12.5 n 22 14 22 22 25 25 NEEHS Mean (s.d.) 7.7 0.5 7.0 11.4 11.7 11.4 11.7 Range 7.0e8.1 6.6e7.3 10.7e12.0 11.3e12.4 10.7e12.0 11.3e12.4 n 735555 EUPH Mean (s.d.) 6.8 0.5 6.3 0.4 11.6 0.9 11.0 0.6 11.3 1.0 10.8 0.8 Range 6.0e7.4 5.5e7.1 10.0e13.0 10.0e12.0 9.5e12.8 9.8e12.0 n 10 20 26 27 22 22 RH Mean (s.d.) 6.5 0.5 6.1 0.4 11.3 0.7 11.0 0.5 10.7 0.7 10.7 0.6 Range 5.4e7.2 4.6e6.9 9.7e12.9 9.2e12.5 9.3e12.3 8.8e11.8 n 61 71 116 118 103 103

a A misprint occurred in the first description of the incisor UA 222 (Macchiarelli et al., 2004a: Table 1) and the correct value is reported here. The standard deviation (s.d.) is provided only for the samples with n 7. EUPH: European early Upper Paleolithic humans (Frayer, 1977 [upper I2 and lower Ms]; Hillson et al., 2010 [lower I1]); HA: H. antecessor from Atapuerca Gran Dolina (Bermúdez de Castro et al., 1999); HEA: African H. erectus/ergaster from Kenya and Ethiopia (Wood, 1991 [upper I2 and lower Ms]; Stynder et al., 2001 [lower I1]; Suwa et al., 2007 [lower Ms]); HEC: H. erectus from China (Wood, 1991); HEG: H. erectus from Georgia (Martinón-Torres et al., 2008 [lower I1, upper I2 and lower Ms]; Lordkipanidze et al., 2013 [lower Ms]); HEJ: H. erectus from Java (Wood, 1991 [lower Ms]; Widianto, 1993 [lower Ms]; Grine and Franzen, 1994 [upper I2]; Stynder et al., 2001 [lower I1]; Kaifu et al., 2005 [lower Ms]; Zanolli, 2013 [upper I2 and lower Ms]); HHE: European H. heidelbergensis from Atapuerca Sima de los Huesos (Martinón-Torres et al., 2012); HHNA: North African H. heidelbergensis from Tighenif and Rabat (Wood, 1991 [lower Ms]; Stynder et al., 2001 [lower I1]; Martinón-Torres et al., 2008 [upper I2]); HHR: H. habilis-rudolfensis from East Africa (Wood, 1991 [upper I2 and lower Ms]; Stynder et al., 2001 [lower I1]; Leakey et al., 2012 [lower Ms]); MA 93: Mulhuli-Amo; NEA: Neanderthals (Voisin et al., 2012); NEEHS: Near Eastern early H. sapiens from Qafzeh (Vandermeersch, 1981); RH: recent humans (Frayer, 1977). the small accessory ridge originating from the metaconid, the two the pulp cavity roof level, while the five main cusps are well rep- enamel distal crests departing from the protoconid and the meta- resented and show that the roof formation was completed (Fig. 1C), conid and converging towards the centre of the trigonid basin there is no relief corresponding to the C7 and to the crests running without joining each other are sharply expressed at the EDJ level. in the occlusal basin. This trigonid crest pattern is similar to the type 10 pattern recently defined by Martínez de Pinillos et al. (2014). A 4-like type talonid Comparative analyses and discussion crest is also visible, but it slightly differs from the original classifi- cation of this trait provided by Martinón-Torres et al. (2014),asit UA 222 (ULI2) and UA 369 (LLI1) displays only a distal buccal segment of the trigonid crest shifting distally towards the centre of the occlusal basin, along with the External morphology Compared with the most common condition expression of two short mesial and distal crest segments running characterizing the African and Eurasian Early-Middle Pleistocene down from the entoconid and hypoconid dentine horns. In addi- dental record (Wood, 1991; Brown and Walker, 1993; Grine and tion, a faint, short accessory ridge set mesially to the entoconid Franzen, 1994; Bermúdez de Castro et al., 1999; Martinón-Torres obliquely runs from the marginal ridge towards the central basin et al., 2007, 2008, 2012; Bailey and Hublin, 2013; Liu et al., 2013), (cf. Martinón-Torres et al., 2014). However, while at the outer sur- including eastern African specimens such as KNM-ER 808 and face the anterior fovea is restricted to a buccolingual groove, a large KNM-WT 15000 (Wood, 1991; Brown and Walker, 1993), the depression occupies the mesial quarter of the corresponding EDJ preserved crowns of both Eritrean incisors are featureless. Only occlusal basin (Fig. 1C). In inferior view, MA 93 exhibits a well UA 222 displays a longitudinal lingual groove and a degree of restricted, mature pulp chamber and thick broken dentine walls. At labial convexity comparable with that seen in some upper 104 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113

Figure 3. Virtual rendering of the pulp cavity of UA 222 (A) and UA 369 (B). UA 222 is compared with the Neanderthal upper I2 Ehringsdorf G3 (NEA; NESPOS Database, 2013), while UA 369 is compared with the lower I1s of H. heidelbergensis from Tighenif (HHNA; Zanolli and Mazurier, 2013) and the Neanderthal Regourdou 1 (NEA; Macchiarelli et al., 2013). The morphology virtually extracted from two recent human incisors (RH; original data) is also shown. d, distal; l, lingual; lb, labial; m, mesial. Scale bar, 5 mm.

incisors from Atapuerca Sima de los Huesos (e.g., the UI1s AT-165 the morphology displayed by the Early Pleistocene lower I2 from and 554; Martinón-Torres et al., 2012). While advanced occlusal ’Ubeidiya, Israel (Belmaker et al., 2002). The Near Eastern specimen wear has partially erased their original morphology, notably in (UB 335) shows relatively small crown size, lack of a lingual the case of UA 222, both specimens lack any trace of either tuberculum, probable absence of the mesial and distal marginal lingual finger-like extensions or a distinct basal eminence as seen ridges, and marked mesial and distal longitudinal root grooves in some Eurasian later Middle Pleistocene upper incisors (Belmaker et al., 2002), a pattern which better fits the character- (Hershkovitz et al., 2011; Martinón-Torres et al., 2012; Liu et al., istics of the two incisors from Buia rather than those displayed by 2013). On the other hand, the lack of an interruption groove on KNM-WT 15000 (Brown and Walker, 1993). the maxillary incisor fits the figures reported for H. erectus s.l. For its buccolingual crown diameter, UA 222 closely approxi- (Manni et al., 2007; see also; Bailey, 2006). mates the estimates available for Javanese H. erectus (Grine and For their general outline and outer gross features, as well as for Franzen, 1994; Zanolli, 2013) but, with the exception of Homo the elongated root with two shallow but proportionally large lon- habilis-rudolfensis from East Africa and Georgian H. erectus, this gitudinal grooves, the incisors from Uadi Aalad globally resemble tooth is systematically smaller compared with the Early-Middle C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 105

Figure 4. Standardized morphometric maps of the virtually unrolled tooth root (15e85% of the total root length) of UA 222 (A) and UA 369 (B). Dentine topographic variation is rendered by a thickness-related chromatic scale increasing from dark blue (0) to red (1). UA 222 is compared with similar evidence from the H. erectus UI2 MI92.2 from Sangiran (HEJ, represented only by the root portion closer to the cervix; Zanolli, 2011, 2013) and the Neanderthal specimen Ehringsdorf G3 (NEA; NESPOS Database, 2013), while UA 369 is compared with the lower I1s of H. heidelbergensis from Tighenif (HHNA; Zanolli and Mazurier, 2013) and the Neanderthal Regourdou 1 (NEA; Macchiarelli et al., 2013). The maps illustrating the condition of two recent human incisors (RH; original data) are also shown. d: distal; l: lingual; lb: labial; m: mesial. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Pleistocene specimens/samples included in the present study morphology and lack of shallowness and orientation from the (notably, the African H. erectus/ergaster sample from eastern Africa; typically labial scratches found on some Neanderthal and pre- Martinón-Torres et al., 2008), and only exceeds the European early Neanderthal incisors and canines, which have been referred to Upper Paleolithic and the recent human average values (Table 5). nonalimentary/manipulative behaviours (Lozano et al., 2009; For the same variable, the lower incisor UA 369 exceeds the esti- Frayer et al., 2010, 2011; Volpato et al., 2012). These marks mates of the H. erectus samples from Java and China (Stynder et al., represent either diet-related microstriations (Ungar et al., 2006; 2001), but fits those of African and Georgian H. erectus (Stynder Teaford, 2007) and/or are the result of taphonomic processes. et al., 2001; Martinón-Torres et al., 2008), H. habilis-rudolfensis Nonetheless, besides the presence in both specimens of randomly (Stynder et al., 2001), and both African and European samples oriented scratches probably resulting from the abrasive action of representing H. heidelbergensis, the highest BL values being dis- variably sized sediment particles, their labial surfaces reveal a played by (Stynder et al., 2001; Martinón-Torres qualitatively and quantitatively similar microtextural pattern et al., 2012) and Neanderthals (Table 5). consisting of thin (2e3 mm), sub-parallel, mostly vertical (or Crown microwear The microfeatures observed on the labial and slightly oblique) fine striations. While microwear analyses of Plio- lingual crown aspects of both Eritrean incisors (Fig. 2) differ in their Pleistocene hominins, including early Homo and H. erectus s.l.,

Figure 5. (A) UA 369 (lower left I1). mMRI labiolingual slice (pixel size, 58.6 mm; slice thickness, 200 mm) through the centre of the root. Scale bar, 500 mm. (B) Root length (in mm) is plotted against root formation time (in years). The grey filled circles represent 44 modern human incisor roots. The blue filled circles represent the buccal and lingual plots for UA 369 calculated to be 80 days apart (2.5 mm per day over 200 mm of dentine formation). The red filled triangles represent the labial and lingual plots for UA 369 calculated to be 90 days apart (2.3 mm per day over 200 mm of dentine formation). All roots are plotted as if zero root height occurred at zero formation time. (C) Tooth length (cumulative crown and root height in mm) for 44 modern human incisors (open black circles) is plotted against tooth formation time (in years). For some modern humans continuous data are available for crown growth but for others the beginning of root growth is placed at the time of crown completion and the height of the crown at the end of enamel formation. The blue filled circles represent the buccal and lingual plots for UA 369 calculated to be 80 days apart (2.5 mm per day over 200 mm of dentine formation). The red filled triangles represent the labial and lingual plots for UA 369 calculated to be 90 days apart (2.3 mm per day over 200 mm of dentine formation). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.) 106 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113

Figure 6. Enamel thickness cartographies of UA 222 (A) and UA 369 (B), both in labial (lb) and lingual (l) views, and of MA 93 (C), in occlusal view. Topographic variation is rendered by a tooth-specific thickness-related pseudo-colour scale ranging from thinner dark-blue to thicker red. Isolated white areas correspond to complete enamel removal following occlusal wear. UA 222 is compared with similar evidence from the H. erectus upper I2 MI92.2 from Sangiran (HEJ; Zanolli, 2011, 2013) and the Neanderthal specimen Krapina D122 (NEA; NESPOS Database, 2013). UA 369 is compared with the lower I1s of H. heidelbergensis from Tighenif (HHNA; Zanolli and Mazurier, 2013) and the Neanderthal Regourdou 1 (NEA; Macchiarelli et al., 2013). MA 93 is compared with the lower M1s of H. heidelbergensis Tighenif 2 (HHNA; Zanolli and Mazurier, 2013) and the Neanderthal S5 from La Chaise de Vouthon (NESPOS Database, 2013) and to the lower M2s of H. erectus NG0802.3 from Sangiran (Zanolli, 2011, 2013), H. heidelbergensis Tighenif 2 (HHNA; Zanolli and Mazurier, 2013) and the Neanderthal Krapina D10 (NESPOS Database, 2013). For each tooth type, the map illustrating the condition of a recent human tooth (RH; original data) is also shown. Independently from their original side, all teeth are shown as left. Scale bar, 1 cm. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.) have concentrated on cheek teeth (e.g., Ungar et al., 2006, 2012; 2010) and the preliminary data from this study do not suggest Pontzer et al., 2011; Ungar, 2011; Ungar and Sponheimer, 2011), it otherwise. The average rates of root growth in UA 369 also appear has been suggested that there was a probable decrease in incisor to fall within those known for modern humans, although it remains use in African H. erectus compared with H. habilis and Homo to be seen if the pattern and the timing of the root growth spurts rudolfensis, either because of a change in diet (broader spectrum; were similar or different in H. erectus s.l. The potential to gather Ungar and Sponheimer, 2011) and/or increased extraoral food data for larger samples of fossil teeth non-destructively using this processing (Ungar, 2011). Accordingly, the evidence from the technique now means questions such as this can be addressed. Eritrean teeth better supports the model of an anisotropic surface Internal structure As revealed by the virtual cartographies illus- whose textural microfeatures indicate a moderate degree of trating enamel distribution in a selected number of tooth crowns complexity resulting from the consumption of relatively soft and sampling Indonesian H. erectus, North African H. heidelbergensis, tough foods, rather than that of an isotropic-like microwear Neanderthals and recent humans (Fig. 6A, B), UA 222 and UA 369 pattern made by a more complex blend of pits and striations share a relatively thin enamel, notably on the lingual aspect. indicating the consumption of hard and brittle food (Ungar and However, while UA 222 apparently exhibits the thinnest labial Sponheimer, 2011). and lingual enamel in the comparative sample of fossil and recent Root growth The results presented here demonstrate that high human upper permanent incisors available to us, UA 369 more resolution MRI micro-imaging clearly has the potential to enable closely approximates the Neanderthal condition (Macchiarelli growth data to be retrieved non-destructively from fossil tooth et al., 2013). roots. Only a w5 year span of root growth could be reconstructed The upper incisor UA 222 presents a peculiar pulp morphology in UA 369, as growth increments in the apical portion of the root resulting in a slightly labiolingually compressed ovoid outline fol- became indistinct. However, if the period between birth and the lowed by a markedly mesiodistally flattened root canal, from the initiation of incisor mineralization (w0.25 years), the crown crown ceiling to the mid-root level (Fig. 3A). In comparison, the formation time (w2.83 years), and the missing apical formation Neanderthal upper I2 from Ehringsdorf, for example, displays a time were each added to the w5 year root formation time much higher volume (59.8 versus 30.7 mm3 in UA 222) and a documented here, the expectation is that the chronological age of regularly-shaped ovoid cavity all along the root (Fig. 3A). While incisor root completion in UA 369 would have been morphologically closer to UA 222, the extant human data available approximately eight to 10 years, as it is in modern humans. This to us (mean volume 17.2 mm3) do not provide evidence of any implies that longer roots in H. erectus s.l. might, at least partly, be accentuated mid-root mesiodistal flattening. formed at the expense of slightly shorter crown formation times. The pulp cavity of UA 369 is similar to UA 222, but with an Alternatively, as in some great ape tooth roots, a second rise in accentuated mesiodistal flattening starting just below the crown root extension rates in the apical portion of the root might also and involving the entire root portion, but notably its mid-region have contributed to longer roots forming in approximately the (Fig. 3B). In this respect, UA 369 nears the morphology of the same time period as modern humans. Neanderthal specimen Regourdou 1 (volume 15.6 mm3; It has previously been suggested that apical closure of the Macchiarelli et al., 2013; see also Le Cabec et al., 2013), but also the anterior teeth in African H. erectus (KNM-WT 15000) may have most common pattern displayed by the recent human lower in- occurred at similar chronological ages to modern humans (Dean, cisors available in our sample (mean volume 11.0 mm3)(Fig. 3B). A C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 107 mesiodistally flattened radicular canal compatible with the outline presence of a C7, better expressed at the EDJ than at the outer of the Eritrean lower incisor also characterizes the lower I2 spec- enamel surface, is compatible with the high frequency of this trait imen UB 355 from ’Ubeidiya (Belmaker et al., 2002) and the observed in H. erectus (Manni et al., 2007; Bailey and Hublin, 2013). recently reported Early Pleistocene lower I2 from Atapuerca Sima Interestingly, in addition to its specific features, MA 93 closely re- del Elefante, Spain (Prado-Simón et al., 2012). Conversely, the early sembles the LM1 KGA10-1 from Konso (Suwa et al., 2007), the two Middle Pleistocene isolated lower I1 from Tighenif (volume sharing a rectangular occlusal outline, the presence of a C7, the lack 15.8 mm3) exhibits a spatula-like shape at crown level turning into of protostylid (which is, conversely, quite a common feature in a regularly-shaped canal with an ovoid cross-section along the H. erectus s.l.; Manni et al., 2007), and low and nearly parallel lateral entire radicular portion (Zanolli and Mazurier, 2013)(Fig. 3B). crown walls. To a lesser extent, the same is true for the lower M2 of Topographic variation in root dentine thickness of both Eritrean the same specimen, even if the latter shows a slight buccolingual incisors has been rendered through a morphometric map gener- tapering of the talonid resulting into a more rounded occlusal ated by virtually unzipping and then unrolling 15e85% of the total outline (Suwa et al., 2007). root length (Fig. 4). Even if the significance (genetic versus func- Regardless of its serial position, MA 93 exhibits an absolutely tional) of the variation in root dentine thickness is not yet fully mesiodistally extended crown (Table 5). If MA 93 is an M2, its MD understood (Bayle et al., 2012) and few specimens have been dimension nears the averages of African H. erectus s.l. (Wood, 1991; detailed so far (Bayle, 2008; Bondioli et al., 2010; Bayle et al., 2011; Suwa et al., 2007) and of the Dmanisi sample (Martinón-Torres Macchiarelli et al., 2013), we note some differences among the et al., 2008; Lordkipanidze et al., 2013), as well as the single value cartographies of the fossil and recent specimens considered in this available so far for H. antecessor (Bermúdez de Castro et al., 1999), study. More specifically, while in UA 222 a unique thicker area is but exceeds those of all remaining Early to Middle Pleistocene found on the labial aspect near the CEJ, in the maps representing comparative samples except H. habilis-rudolfensis, the latter again Indonesian H. erectus, Neanderthals and recent humans, vertically displaying the greatest anteroposterior crown diameter (Leakey extended thicker dentine is present both labially and lingually. et al., 2012). Under this scenario, while the breadth of MA 93 Furthermore, at least in the Neanderthal and modern human roots, does not deviate from the estimates of most among the samples the regions where absolutely and relatively thinner dentine is considered in this analysis (notably, from African, Georgian, Chi- found are much less spread and confined to the lower portion of the nese, and Javanese H. erectus; Wood, 1991; Suwa et al., 2007; map (Fig. 4A). Conversely, UA 369 root topography globally overlaps Martinón-Torres et al., 2008; Lordkipanidze et al., 2013; Zanolli, the pattern of dentine thickness distribution expressed by the 2013), it is lower than measured in H. habilis-rudolfensis and North African H. heidelbergensis and Neanderthal representatives North African H. heidelbergensis. The Eritrean molar also exceeds used in this study, while the modern human cartography locally estimates from European H. heidelbergensis (Table 5). shows relatively thinner dentine (Fig. 4B). Internal structure Comparative tooth crown tissue proportions in MA 93 (LLM1/M2) MA 93 and some fossil and recent human specimens/samples are given in Table 6 (2D estimates) and Table 7 (3D). The comparative External morphology The comparison between the occlusal record available for these kind of structural variables remains morphology of MA 93 and earlier East African lower molars sam- quantitatively scanty in paleoanthropology and the extent of pling H. erectus/ergaster (e.g., KNM-ER 730, 820, 992, KNM-WT intra-taxic/population variation is still virtually unknown, but 15000), as well as with the geographically closer specimens from regardless of whether assigned to M1 or M2, the Eritrean Konso (Suwa et al., 2007) and Melka Kunture (Condemi, 2007; specimen exhibits rather low 2D values, notably for the enamel Morgan et al., 2012) in Ethiopia, reveals a common general area (c). However, for dentine area (b) and enamel-dentine pattern. However, while the presence of five main cusps junction length (e), the specimen fits the estimates from two separated by a Y-shaped groove is found almost ubiquitously in South African early Homo M1s (Smith et al., 2012), and is also Early Pleistocene African lower M1s (Wood, 1991; Bermúdez de near those from an isolated Javanese H. erectus M2 crown for the Castro et al., 1999; Martinón-Torres et al., 2007, 2008), MA 93 variables c, e, and AET (Zanolli, 2011)(Table 6). shows a cusp organization resulting in a þ fissure pattern more The number of comparative observations reported so far for 3D commonly found in the Early Pleistocene lower M2s (Gómez- inner tooth structural organization is even more limited, notably Robles et al., 2014). While not very common in the African record, for the LM1s (Table 7), the most investigated fossil humans being the cruciform groove pattern is relatively frequent in Indonesian the Neanderthals (Macchiarelli et al., 2006, 2007, 2008, 2013; H. erectus (Zanolli, 2013) and in Middle Pleistocene Eurasian Olejniczak et al., 2008a; Kupczik and Hublin, 2010). However, lower M2s and M3s (Bermúdez de Castro et al., 1999; Martinón- while for the enamel (Ve), dentine (Vcd), and pulp (Vcp) volumes Torres et al., 2007, 2012; Bailey and Hublin, 2013). the Eritrean crown falls within the range of variation known for Cusp size sequence in MA 93, with a larger protoconid than the Neanderthals and recent humans, as an M2 it differs from the fig- metaconid and a medium-large hypoconulid, fits the average ures expressed by the single representatives available to us of pattern found in Early Pleistocene Afro-European lower M1s and Indonesian H. erectus (Zanolli, 2011) and H. heidelbergensis from M2s (e.g., KNM-ER 820 and 992 (Wood, 1991), ATD6-5 (Bermúdez Tighenif (Zanolli and Mazurier, 2013), as well as from the later de Castro et al., 1999)). In addition, MA 93 exhibits a well- North African Aterians (Kupczik and Hublin, 2010). developed and complete mid-trigonid crest, which even if found A different picture is provided by the comparison for the percent in Georgian H. erectus and frequent in H. erectus s.s., is generally ratio Vcdp/Vc, which represents the coronal portion that is dentine absent or interrupted by the central groove in African H. erectus/ and pulp, a directly comparable, size-independent parameter ergaster specimens like KNM-ER 820, 992 and KNM-WT 15000 rather finely describing internal crown structural organization. In (Wood, 1991; Brown and Walker,1993; Martinón-Torres et al., 2007, this case and specific comparative context, the structurally most 2008; Bailey and Hublin, 2013; Zanolli, 2013). Unlike most Afro- distant signature with respect to MA 93 is represented by the European H. erectus lower molars, like D211 (Martinón-Torres virtually unworn H. erectus lower M2 from Java used in this study et al., 2008), KGA10-1 (Suwa et al., 2007), KNM-ER 820 and KNM- (specimen NG0802.3; Zanolli, 2013). This specimen shows the WT 15000 (Wood, 1991; Brown and Walker, 1993), the Eritrean lowest ratio of any Pleistocene specimen reported so far and con- molar lacks any expression of a deflecting wrinkle, while the trasts with the higher figures provided by our specimen sampling 108 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113

Table 6 et al., 2008a; Smith et al., 2012; Macchiarelli et al., 2013)(Fig. 6C). 2D-based linear and surface structural variables (crown tissue proportions) assessed Noteworthy, for enamel thickness MA 93 falls near the lower limits in MA 93 and compared with some Pleistocene and recent human specimens/ of the modern human molar variation range. samples. At the EDJ, MA 93 exhibits some non-metric features similar to 2 2 2 Specimens/ c (mm ) b (mm ) e (mm ) AET (mm) RET the condition recently reported for the lower M1 of the early samples Middle Pleistocene adult left hemi-mandible Tighenif 2, (Zanolli MA 93 18.6 38.2 18.6 1.0 16.2 and Mazurier, 2013), particularly the presence of a metaconulid- Lower M1 type C7 (while the lower M2 of the Algerian specimen displays HHR (1) 30.7 47.0 20.9 1.5 21.5 SAEH (2) Mean 33.8 37.0 19.2 1.8 29.0 an interconulid-type C7). However, in the Eritrean molar the Range 28.7e29.3 complete mid-trigonid crest expresses a middle-mesial configura- NEA (13) Mean 21.6 43.0 21.6 1.0 15.5 tion of grade 3, absent in the North African fossil assemblage e e Range 0.9 1.2 12.7 20.5 sampling H. heidelbergensis but recorded with a relatively high ANEFMH (6) Mean 26.1 43.9 22.0 1.2 18.0 Range 1.0e1.5 15.2e23.3 frequency in Neanderthals and, more rarely, in recent humans RH (58) Mean 22.0 40.5 20.4 1.1 17.0 (Bailey et al., 2011). Range 0.8e1.4 11.8e22.6 When compared to the condition revealed at the EDJ level by Lower M2 some fossil human lower M1s and M2s (the proportionally higher HHR (1) 34.6 43.4 20.1 1.7 26.2 dentine horns and the rounder, more buccolingually extended HEJ (1) 21.2 31.2 17.4 1.2 21.7 HHNA (1) 25.2 47.7 21.1 1.2 17.3 occlusal basin displayed by the H. erectus lower M2 from Sangiran; NEA (6) Mean 21.1 44.6 20.5 1.0 15.5 Zanolli, 2011), MA 93 shows absolutely and relatively low dentine Range 1.0e1.1 13.9e16.5 horns (Fig. 7). ANEFMH (4) Mean 27.9 49.2 21.8 1.3 18.3 Range 1.2e1.4 16.3e20.2 RH (47) Mean 22.2 34.4 18.5 1.2 20.5 Geometric morphometric analyses Range 0.9e1.6 14.8e27.7

ANEFMH: pooled African-Near Eastern anatomically modern fossil humans from The results of the between-group principal component analysis Morocco, South Africa and Israel (Smith et al., 2012); HEJ: Javanese H. erectus from Sangiran (Zanolli, 2011, 2013); HHNA: North African H. heidelbergensis from Tigh- (bgPCA) based on the EDJ Procrustes shape coordinates of the lower enif (Zanolli and Mazurier, 2013); HHR: H. habilis-rudolfensis from East Africa (Smith M2s of the MA 93 Eritrean specimen, of a Javanese H. erectus and of et al., 2012); NEA: Neanderthals (Smith et al., 2012); RH: recent humans (Smith a North African H. heidelbergensis, compared to structural evidence et al., 2012); SAEH: South African early Homo (Smith et al., 2012). from lower M1s and LM2s in two samples of Neanderthal (n ¼ 17) and recent human (n ¼ 31), are shown in Fig. 8. The analysis of the North African H. heidelbergensis and the typical Neanderthal between-group principal component scores in reference to the pattern (Olejniczak et al., 2008a)(Table 7). centroid size (Mitteroecker and Bookstein, 2011) suggests that Whether an M1 or M2, the crown from Mulhuli-Amo has there is a weak allometric signal in the group separation, present intermediate-thick enamel (Martin, 1985) with comparable 2e3D only for the bgPC1 (R2 ¼ 0.098). As expected, the bgPCA allows a AET and RET values (Tables 6 and 7) systematically below the es- rather good discrimination between Neanderthals and recent timates from any Early and Middle Pleistocene specimen/sample humans (along the bgPC1), as well as between lower M1s and M2s known to us, notably African early Homo (Smith et al., 2012) and (along the bgPC2). In this context, not only is MA 93 clearly set apart Javanese H. erectus (Zanolli, 2011). To a lesser extent, enamel from both comparative samples, but also from the two late Earlye thickness is below the values for early anatomically modern early Middle Pleistocene Javanese and Algerian specimens included humans from Africa (Smith et al., 2012) and Europe (Bayle et al., in the analysis. 2010). The closest fits in our comparative analysis are an only A relatively low EDJ topography, a feature characterizing the slightly thicker early Middle Pleistocene lower M2 from Tighenif shapes on the extreme negative portion of the bgPC2 axis where (Zanolli and Mazurier, 2013) and the Neanderthals (Olejniczak MA 93 is located (Fig. 8), is reminiscent of the ancestral hominin

Table 7 3D-based linear, surface, and volumetric structural variables (crown tissue proportions) assessed in MA 93 and compared with some Pleistocene and recent human specimens/ samples.

Ve (mm3) Vcd (mm3) Vcp (mm3) Vcdp (mm3) Vc (mm3) SEDJ (mm2) Vcdp/Vc (%) 3D AET (mm) 3D RET

MA 93 261.4 316.8 15.4 332.2 593.5 256.1 56.0 1.0 14.7 Lower M1 NEA (17) Mean 255.7 375.4 15.2 366.4 622.2 238.0 58.87 1.1 15.6 Range 189.8e350.4 275.3e490.2 3.7e37.0 281.3e510.8 490.1e831.8 129.3e334.4 52.0e63.6 0.8e1.6 11.8e24.0 EUPH (1) Mean 283.6 260.3 37.6 298.0 581.6 222.5 51.2 1.3 19.1 RH (13) Mean 245.9 283.1 14.7 288.1 534.0 224.9 53.7 1.1 16.7 Range 204.2e349.9 226.9e392.4 2.7e34.5 186.8e420.8 399.4e770.7 165.7e271.1 46.8e57.3 0.9e1.4 14.5e22.4 Lower M2 HEJ (1) Mean 240.8 217.2 1.9 219.2 460.0 181.1 47.7 1.3 22.1 HHNA (1) Mean 373.1 480.2 22.3 502.4 875.5 312.5 57.4 1.2 15.0 NEA (11) Mean 276.0 353.2 13.7 366.9 642.6 252.3 57.1 1.1 15.4 Range 169.1e397.4 242.2e493.7 1.5e28.2 250.0e519.3 436.1e895.7 182.5e345.6 51.0e64.2 0.8e1.3 11.9e20.9 AFMH (3) Mean 343.0 399.0 8.4 407.4 760.3 265.9 53.4 1.3 17.6 Range 306.7e371.2 351.1e469.1 3.6e17.1 354.7e486.2 665.0e859.2 236.7e304.1 50.4e56.6 1.2e1.4 14.7e19.9 RH (26) Mean 237.2 238.0 13.6 249.5 486.7 195.7 51.0 1.3 20.2 Range 136.4e361.9 167.9e396.5 0.8e35.5 135.3e426.0 306.6e770.0 95.4e285.4 42.7e57.3 0.7e2.3 12.6e40.7

AFMH: North African Aterians (Kupczik and Hublin, 2010); EUPH: the European early Upper Paleolithic human from Lagar Velho (Bayle et al., 2010); HEJ: Javanese H. erectus from Sangiran (Zanolli, 2011, 2013); HHNA: North African H. heidelbergensis from Tighenif (Zanolli and Mazurier, 2013); NEA: Neanderthals (Olejniczak et al., 2008a; Kupczik and Hublin, 2010; Macchiarelli et al., 2013; NESPOS Database, 2013); RH: recent humans (Olejniczak et al., 2008a; and original data). C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 109

Figure 7. Virtual rendering of MA 93 in occlusal and buccal views (A). Following each projection, it is shown (from the left) the outer crown (red), the dentine (gold) with the enamel in semi-transparency (grey), and the distinct topography of the EDJ (yellow). In (B), MA 93 is compared with the lower M1s of H. heidelbergensis Tighenif 2 (HHNA; Zanolli and Mazurier, 2013), the Neanderthal S5 from La Chaise de Vouthon (NEA; NESPOS, 2013) and the European early Upper Paleolithic Lagar Velho 1 (EUPH; Bayle et al., 2010), and in (C) with the lower M2s of H. erectus NG0802.3 from Sangiran (Zanolli, 2011, 2013), H. heidelbergensis Tighenif 2 (HHNA; Zanolli and Mazurier, 2013) and the Neanderthal Regourdou 1(Macchiarelli et al., 2013). The maps illustrating the condition of a recent human lower M1 and M2 (RH; original data) are also shown. Independently from their original side, all crowns are shown as left. Scale bar, 1 cm. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.) condition (Macchiarelli et al., 2004b; Skinner et al., 2008a,b; Concluding remarks Olejniczak et al., 2008b; Braga et al., 2010). However, as revealed by histological or virtual sections of some specimens from East The discovery of ca. 1 Ma human remains in the area of Buia, Africa (Beynon and Wood, 1986), H. antecessor from Gran Dolina Danakil Depression of Eritrea, provides new insights about the (Bermúdez de Castro et al., 2010) and Asian H. erectus (Dean et al., variation, geographic extent and evolutionary trends in 2001; Smith et al., 2009), low dentine horns are rather common in H. erectus/ergaster near the end of the Early Pleistocene, a period Early and Middle Pleistocene Homo compared with the more still poorly sampled in the East African fossil record (Gilbert and elevated and sharper derived morphology commonly seen in Late Asfaw, 2008). The geographic setting of the Dandiero Rift Basin Pleistocene and recent humans (Olejniczak et al., 2008a; Skinner suggests the region may have recurrently played the role of a et al., 2008a, 2010). Nonetheless, besides the Eritrean specimen, buffer zone in human dispersal towards North Africa and the the condition typical of Early Pleistocene African Homo,ifany,is Near East during various phases of the Pleistocene (Abbate and still unreported in a 3D perspective. Sagri, 2012). 110 C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113

MA 93 crown from Mulhuli-Amo is a cruciform fissure. While observed at Dmanisi (Martinón-Torres et al., 2008) and also present in Indonesian H. erectus (Martinón-Torres et al., 2008; Zanolli, 2013), this pattern is more common in Middle Pleistocene mandibular molars (Martinón-Torres et al., 2007; Gómez-Robles et al., 2014). However, it should be noted that, at least for the late Early and Middle Pleistocene African fossil record, the time-related variation patterns in morphological dental trait distribution have not yet undergone the scrutiny of theoretical evolutionary models, such as ‘random walk’ (i.e., Brownian motion) type (Hunt, 2007; see; Gómez-Robles et al., 2014). Dimensionally, the Eritrean molar has a mesiodistal diameter of 13.9 mm, intermediate between the lower M1 (13.5 mm) and the lower M2 (14.4 mm) of the KGA10-1 partial mandible from Konso, Ethiopia (Suwa et al., 2007). Conversely, its buccolingual diameter is small compared with the same teeth (11.9 mm versus 12.8 and 13.0 mm, respectively). Its mesiodistal size contrasts with the short postcanine maxillary tooth row displayed by the UA 31 cranium from the nearby, quasi-contemporaneous site of Uadi Aalad. This maxilla has buccolingually, not mesiodistally, expanded molar sockets (Abbate et al., 1998; Macchiarelli et al., 2004a) and its proportions would better fitashortermandib- ular dental arcade than seen in other H. erectus/ergaster African Figure 8. Between-group principal component analysis (bgPCA) of the Procrustes specimens (e.g., Wood, 1991; Brown and Walker, 1993; Suwa et al., shape coordinates of the EDJs of MA 93 and the LM2 specimens NG0802 representing 2007; Wood and Leakey, 2011). Given the striking morphological H. erectus from Sangiran (HEJ; Zanolli, 2011, 2013) and Tighenif 2 sampling North Af- rican H. heidelbergensis (HHNA; Zanolli and Mazurier, 2013) compared with two lower similarity between UA 31 and the newly discovered (but still M1 and lower M2 samples consisting of 17 Neanderthals (NEA) and 31 recent humans unreported) cranial remains from Mulhuli Amo (Coppa et al., (RH), respectively. See the text for details on the composition of the Neanderthal and 2014), the extent of anteroposterior crown development dis- recent human samples. The size of the circles is directly correlated with the centroid played by MA 93 suggests a certain degree of (likely sex-related) size. dentognathic structural variation in Africa around 1 Ma. In this regard, it is also interesting to note that the virtually contempo- Compared with the condition more typical of African and raneous upper M1 specimen COR 2011 from Cornelia-Uitzoek, Eurasian Early and Middle Pleistocene anterior teeth (e.g., Wood, South Africa, shows a large MD diameter (13.8 mm; Brink et al., 1991; Manni et al., 2007; Martinón-Torres et al., 2007, 2012; 2012), which again would fit a rather longer maxilla than Bailey and Hublin, 2013; Bermúdez de Castro and Martinón- measured in UA 31. As appropriately noted by Brink et al. (2012), Torres, 2013), while occlusally worn, the incisors from Uadi Aalad African Middle Pleistocene molar crowns tend to show larger BL apparently lack any developed expression of accessory features diameters compared with their European counterparts and, with (i.e., distinct tubercles, ridges, or cusp-like features expressed in the few exceptions, almost invariably smaller MD lengths compared cingular region of the lingual surface). For its buccolingual crown with early Homo. diameter, the lower central incisor UA 369 falls within the range of The fossiliferous area of Buia and the early Middle Pleistocene variation of most Early and Middle Pleistocene samples, while the site of Tighenif, Algeria, are nearly 4550 km and likely less than 300 upper lateral incisor is relatively small compared with early African kyr apart (Geraads et al., 1986). Recently re-attributed to Homo, but still comparable with the Javanese H. erectus estimates H. heidelbergensis (Mounier et al., 2009; but see; Antón, 2013) and (Zanolli, 2013). Enamel thickness variation is no longer considered assessed for their inner structural morphology (Zanolli et al., 2010; a useful trait to assess intertaxic evolutionary relationships (Smith Zanolli and Mazurier, 2013), the teeth from Tighenif share a number et al., 2012). Nonetheless, it is noteworthy that the Eritrean incisors of features with the Eritrean specimens. With respect to the UA 369 share relatively thin enamel, absolutely thinner than measured in lower I1, at the external surface these include the BL diameter and the early Middle Pleistocene North African assemblage of Tighenif the lack of enamel superstructures. Conversely, the isolated North (Zanolli and Mazurier, 2013). They also show a topographic pattern African lower I1 differs because of its relatively thicker enamel on the labial aspect closer to Neanderthals than to the modern (notably, on the lingual aspect) and radicular dentine (notably, human condition. In this respect, it would be interesting to labially), and mostly for its spatula-shaped (versus a labiolingually compare the Eritrean specimens to the structural signature virtu- compressed outline) pulp cavity at crown level, which turns into a ally extracted from the lower I2s of ’Ubeidiya (Belmaker et al., 2002) regularly-shaped (versus an ovoid-shaped) cross-section along the and the ca. 1.3 Ma old ATE9-1 partial mandible of Sima del Elefante, entire radicular portion. Compared with the lower M1s and M2s of Spain (Carbonell et al., 2008; Bermúdez de Castro et al., 2011). The Tighenif, the MA 93 crown from Mulhuli-Amo mostly shares a latter shares with UA 369 some morphological details of the medium-large hypoconulid, presence of a small tuberculum inter- radicular canal (Prado-Simón et al., 2012). Conversely, additional medium (because of occlusal wear, only visible at the EDJ level on methodological research is needed before appropriately inter- Tighenif 2, but not present in Tighenif 1), an occlusal groove pattern preting the differences in radicular dentine thickness distribution, (with the lower M2 of Tighenif 2, but not with the lower M1), a even if the fossil specimens detailed in this study share a pattern percentage of coronal volume that is dentine and pulp (Vcdp/Vc), characterized by more accentuated contrasts (notably, near the CEJ) and similarly thickened enamel (2e3D RET values). However, the than recorded in modern human anterior teeth (cf. Bayle et al., Eritrean and Algerian molars differ in crown shape (more bucco- 2011, 2012). lingually expanded in Tighenif), mid-trigonid crest pattern Together with the lack of a deflecting wrinkle and protostylid, a expressed at the EDJ (of middleemiddle type in Tighenif), and de- peculiar feature on the outer occlusal surface of the lower M1/M2 tails of the EDJ conformation (rather low dentine horns in MA 93), C. Zanolli et al. / Journal of Human Evolution 74 (2014) 96e113 111 with Tighenif’s lower M2s more closely approaching the modern Homo cranium from the Danakil (Afar) Depression of Eritrea. Nature 393, 458e human condition (Zanolli and Mazurier, 2013). 460. Abbate, E., Beraky, W., Bruni, P., Falorni, P., Papini, M., Sagri, M., Simret, G., As a whole, while quantitatively limited, comparative evidence Tewolde, M.T., 2004. 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In this 170. respect, while 2e3D virtual tooth imaging potentially discloses Antón, S.C., 2013. Homo erectus and related taxa. In: Begun, D.R. (Ed.), A Companion new promising perspectives in inter-taxic comparisons (e.g., Smith to Paleoanthropology. Blackwell Publishing Ltd., Pondicherry, pp. 497e516. Baab, K.L., 2008. The taxonomic implications of cranial shape variation in Homo et al., 2012; Liu et al., 2013; Macchiarelli et al., 2013), we admit that erectus. J. Hum. Evol. 54, 827e847. the current paucity of similar evidence from more representative Bailey, S.E., 2006. Beyond shovel-shaped incisors: Neandertal dental morphology in Early and Middle Pleistocene African assemblages still limits any a comparative context. Period. Biol. 108, 253e267. “ ” reliable conclusion about the variation patterns and evolutionary Bailey, S.E., Hublin, J.J., 2013. What does it mean to be dentally modern ? In: Scott, G.R., Irish, J.D. (Eds.), Anthropological Perspectives on Tooth Morphology, trends in H. erectus/ergaster tooth endostructural morphology. This Genetics, Evolution, Variation. Cambridge University Press, Cambridge, in turn reduces the chances of providing conclusive, dentally-based pp. 222e249. support to the suggested speciation event occurring in Africa dur- Bailey, S.E., Skinner, M.M., Hublin, J.J., 2011. What lies beneath? An evaluation of lower molar trigonid crest patterns based on both dentine and enamel ing the early Middle Pleistocene (Rightmire, 2013). This emphasises expression. Am. J. Phys. Anthropol. 145, 505e518. the importance of continually developing methods, especially Bayle, P., 2008. Analyses quantitatives par imagerie à haute résolution des sé- those that are non-desctructive, to extract more information out of quences de maturation dentaire et des proportions des tissus des dents déciduales chez les Néanderthaliens et les Hommes modernes. Ph.D. Disser- the existing fossil record (Wood and Leakey, 2011). tation, Université Toulouse III Paul Sabatier. Bayle, P., Macchiarelli, R., Trinkaus, E., Duarte, C., Mazurier, A., Zilhão, J., 2010. Dental maturational pattern and dental tissue proportions in the early Upper Paleolithic Acknowledgements child from , . Proc. Natl. Acad. Sci. 107,1338e1342. Bayle, P., Bondioli, L., Macchiarelli, R., Mazurier, A., Puymerail, L., Volpato, V., Zanolli, C., 2011. Three-dimensional imaging and quantitative characterization The Buia Project (Eritrean-Italian Danakil Expedition: Anthropo- of human fossil remains. Examples from the NESPOS database. In: Archaeological and Geo-Paleontological Research Project), an Macchiarelli, R., Weniger, G.-C. (Eds.), Pleistocene Databases. Acquisition, Stor- ongoing international project developed in agreement and collabo- ing, Sharing. Wissenschaftliche Schriften des Neanderthal Museums 4, Mett- mann, pp. 29e46. rationwiththeEritrean NationalMuseum of Asmara and the Northern Bayle, P., Bondioli, L., Dean, C., Macchiarelli, R., 2012. Unrolled roots: topographic Red Sea Regional Museum of Massawa, is granted by the Italian variation of the dentine thickness in Neandertal and modern human anterior Ministry for Education and Research (COFIN and PRIN national pro- teeth. Proc. Eur. Soc. Study Hum. Evol. 1, 36 (abstract). Belmaker, M., Tchernov, E., Condemi, S., Bar-Yosef, O., 2002. New evidence for jects), the Italian Ministry for Foreign Affairs (DGPCC-V; grants ARC- hominid presence in the Lower Pleistocene of the Southern Levant. J. Hum. Evol. 000387/2011, ARC-000504/2012, ARC-000853/2013), the University 43, 43e56. ‘La Sapienza’ of Rome (‘Grandi Scavi Archeologici’ project; grant Bermúdez de Castro, J.M., Martinón-Torres, M., 2013. A new model for the evolution of the human Pleistocene populations of Europe. Quatern. Int. 295, 102e112. B88C13003900005 and B88C13000440005). During the last years, Bermúdez de Castro, J.M., Rosas, A., Nicolás, M.E., 1999. Dental remains from our field work has also benefited from the support provided by the Atapuerca-TD6 (Gran Dolina site, Burgos, Spain). J. Hum. Evol. 37, 523e566. Italian National Research Council, the University of Florence, the Bermúdez de Castro, J.M., Martinón-Torres, M., Prado, L., Gómez-Robles, A., Rosell, J., López-Polín, L., Arsuaga, J.L., Carbonell, E., 2010. New immature hominin fossil Leakey Foundation, the National Geographic Society, the Spanish from European Lower Pleistocene shows the earliest evidence of a modern Ministry of Science and Education, the National Museum of Prehistory human dental development pattern. Proc. Natl. Acad. Sci. 107, 11739e11744. and Ethnography of Rome, the University of Poitiers, the French CNRS Bermúdez de Castro, J.M., Martinón-Torres, M., Gómez-Robles, A., Prado-Simón, L., fi Martín-Francés, L., Lapresa, M., Olejniczak, A.J., Carbonell, E., 2011. Early Pleis- (MNHN Paris). For scienti c collaboration, we acknowledge E. Abbate tocene human mandible from Sima del Elefante (TE) cave site in Sierra de (Florence), M.G. Belcastro (Bologna), P. Bruni (Florence), G. Carnevale Atapuerca (Spain): a comparative morphological study. J. Hum. Evol. 61, 12e25. (Turin), M. Delfino (Turin), M. Ghinassi (Pavia), B. Martínez-Navarro Beynon, A.D., Wood, B.A., 1986. Variations in enamel thickness and structure in East e (Tarragona), A. Mazurier (Poitiers), P. O’Higgins (York), O. Oms (Bar- African hominids. Am. J. Phys. Anthropol. 70, 177 193. Bigazzi, G., Balestrieri, M.L., Norelli, P., Oddone, M., 2004. Fission-track dating of a celona), M. Papini (Florence), M. Pavia (Turin), M. Sagri (Florence). We tephra layer in the Alat formation of the Dandero Group (Danakil Depression, are also grateful to F. Bernardini (Trieste), M. Chech (Paris), F. Genchi Eritrea). Riv. Ital. Paleontol. Strat. 110, 45e49. (Bologna), A. Keset (Massawa), N. Kiflemariam (Massawa), F. Romag- Bondioli, L., Coppa, A., Frayer, D.W., Libsekal, Y., Rook, L., Macchiarelli, R., 2006. A one million year old human pubic symphysis. J. Hum. Evol. 50, 479e483. noli (Florence), D. Tesfay (Massawa), A. Todero (Bologna), P. Ungar Bondioli, L., Bayle, P., Dean, C., Mazurier, A., Puymerail, L., Ruff, C., Stock, J.T., (Fayetteville), V. Vannata (Rome), and A. Zerai (Asmara). Very useful Volpato, V., Zanolli, C., Macchiarelli, R., 2010. Morphometric maps of long bone and competent comments were provided by one referee when com- shafts and dental roots for imaging topographic thickness variation. Am. J. Phys. Anthropol. 142, 328e334. menting on the comparative evidence from Atapuerca. For detailed Bondioli, L., Capuani, S., Coppa, A., Dean, C.M., Macchiarelli, R., Mancini, L., stratigraphic information about the Mulhuli-Amo site, we especially Zanolli, C., 2013. Dentine growth patterns in human fossil teeth assessed by thank M. Ghinassi and M. Papini. The specimen MA 93 was collected in high resolution magnetic resonance micro-imaging. Proc. Europ. Soc. Stud. Hum. Evol. 2, 50 (abstract). 2011 by the Saho guide Hussein Umar. Finally, we recognize that Braga, J., Thackeray, J.F., Subsol, G., Kahn, J.L., Maret, D., Treil, J., Beck, A., 2010. The without the generous help and warm hospitality of the people from enamel-dentine junction in the postcanine dentition of afri- the Buia village, this work would simply have not been possible. canus: intra-individual metameric and antimeric variation. J. Anat. 216, 62e79. Brink, J.S., Herries, A.I.R., Moggi-Cecchi, J., Gowlett, J.A.J., Bousman, C.B., Hancox, J.P., Grün, R., Eisenmann, V., Adams, J.W., Rossouw, L., 2012. 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