doi:10.1038/nature10149 Supplementary Discussion 1. Fossil Tooth Preservation Several lines of evidence indicate that diagenetic processes have not substantially altered the Sr isotope ratios of fossil enamel from Sterkfontein and Swartkrans. A previous study on modern rodent enamel from the Sterkfontein Valley and fossil rodent enamel from Sterkfontein Member 4 and Swartkrans Member 1 found that Sr concentrations and Sr isotope ratios were similar for modern and fossil rodents 13. In cases of significant post-mortem chemical alteration, increased concentrations of Sr and reduced variability of Sr isotope ratios would be expected. These results were consistent with an earlier, broader study of trace elements at Sterkfontein, Swartkrans and Makapansgat that showed little evidence for diagenetic Sr in fossil enamel 30. Moreover, even if diagenesis did impact Sr concentrations and isotope ratios of fossils from the Sterkfontein Valley, it would not impact smaller hominins differently than all other taxa. Thus, diagenesis is an implausible explanation for the anomalously high proportion of non-local small hominins observed here. 2. Sex Estimation Using Hominin Teeth For each tooth category we selected the smallest or largest hominin teeth that were available for destructive sampling from the Ditsong National Museum of Natural History (formerly the Transvaal Museum). Despite overlap in molar size between male and female gorillas and chimpanzees, male molars are still significantly larger on average 23. For example, mean chimpanzee maxillary M3 (maxillary and mandibular M3s were the only molars used in this study) occlusal area is 91.1 ± 16.5 mm 2 while female WWW.NATURE.COM/NATURE 1
and male chimpanzee maxillary M3 areas are highly distinct at 84.5 ± 13.3 and 97.7 ± 16.9 mm 2 respectively [t-test, P < 0.001; sexes were unevenly represented in the Mahler 23 dataset so to avoid this potential bias we obtained bootstrapped estimates (20 subsamples per group; 1 x 10 4 iterations) of means and measures of variance; Supplementary Table 1]. Thus, one standard deviation (SD) above the species mean (as we use in this paper to estimate sex of australopith taxa) is nearly two SD above the female mean. Male and female gorilla maxillary M3s are also highly distinct (t-test, P < 0.001) and one SD above the gorilla mean (218.7 ± 35.9 mm 2 ) is more than two SD above the female gorilla mean (197.1 ± 25.3 mm 2 ). Given this and current body size estimates of the South African australopiths which puts their dimorphism between that of chimpanzees and gorillas 22, we believe our sex estimates to be reasonable. There are other possible interpretations of this size difference, although sex is presently the most parsimonious. For instance, it might be argued that more than one species exists within the current P. robustus or A. africanus hypodigms, but that is a minority opinion (but see 31-33 ). Another possibility is that smaller individuals had restricted access to limited resources, and were thus more likely to emigrate in search of better foraging opportunities. It could also be argued that tooth size and landscape use changed in both taxa through time. Such explanations are unnecessarily rococo, however, given known size differences in the dentition of dimorphic hominoids 22,23. 3. Analysis of SK 876 A previous study 19 reported 87 Sr/ 86 Sr for the tooth enamel of SK 876, a mandible of Paranthropus robustus from Swartkrans. SK 876 was very likely a male. For instance, WWW.NATURE.COM/NATURE 2
its M 3 is one of the largest in the hypodigm. Sillen et al. 19 suggested that this individual was non-local because its enamel 87 Sr/ 86 Sr is 0.72143. However, although this value is lower than those of other hominins in the present study, it does fall within the range of Malmani dolomite plants and animals and is therefore not demonstrably non-local. If SK 876 is added to our analysis, it decreases the proportion of non-local Hominidae from 32% to 30% and decreases the proportion of non-local likely males from 17% to 14%. This strengthens the pattern in which females are more likely to be non-local than males. We have not included SK 876 in our primary analysis given the significant methodological differences between our study and that of Sillen et al. 19 and because its inclusion would not significantly alter our results. 4. Landscape Use in Paranthropus robustus and Australopithecus africanus There are no statistically significant differences in the proportions of non-local individuals between Paranthropus robustus and Australopithecus africanus. However, it is worth noting that one large Paranthropus specimen is non-local (SK 4) and another (SK 38) borders on non-local 87 Sr/ 86 Sr values. As discussed above, SK 876 is almost certainly a male and has the lowest 87 Sr/ 86 Sr value of any australopith analyzed to date 19. While these data do not directly indicate differences in ranging behavior or dispersal between Paranthropus and Australopithecus males, they do suggest the idea merits additional study. 5. Differences between Geological Substrates The geological substrates in and around the Sterkfontein Valley depicted in Figure 1 correspond closely to vegetation zones 34. For example, the Malmani dolomite in WWW.NATURE.COM/NATURE 3
which the fossil sites are centered supports Carletonville Dolomite Grassland on undulating plains dissected by rocky ridges. Grass genera including Panicum, Eragrostis, Aristida and Diheteropogon are variably dominant across this landscape with patches of tree cover including Celtis africana, Olea europaea africana, and Rhus pyroides 35. In contrast, Andesite Mountain Bushveld is found on the Hekpoort andesite/basalt to the north and supports an abundance of woody vegetation including Acacia caffra and Acacia karoo 34. Several other vegetation types have been identified across the geological substrates in the area (e.g., Marikana Thornveld, Gauteng Shale Mountain Bushveld, Egoli Granite Grassland). Further studies documenting the resources for primates in these habitats might shed more light on Plio-Pleistocene hominin landscape use. Potential resources found in the dolomites, but not generally present in other geological substrates, include abundant caves. The availability of water on the dolomites compared to other substrates also requires consideration. There is little topographic change between many of the geological substrates, so one can readily walk between the dolomites, shales, andesites, and quartzites. Thus, there is little reason to believe that the hominins were significantly constrained to the dolomites by topography. 6. Home Range Size It is not possible to estimate actual hominin home range size from these data without making several unwarranted assumptions. For instance, if we assume that the home range was a circle centered on Swartkrans cave, then in order to remain completely on dolomite, the individual s ranging from the site could not have exceeded about 3 km (the shortest distance to another geological substrate) for a total area of about 28 km 2. WWW.NATURE.COM/NATURE 4
However, an individual could have ranged more broadly while staying within the dolomite because the dolomite substrate extends for >30 km toward the northeast and southwest (Fig. 1). Therefore, while individuals with Sr isotope values consistent with dolomite may have had home range sizes of less than 28 km 2, we cannot exclude the possibility that their home ranges were much larger. For comparison, savanna baboons (Papio cynocephalus, Papio ursinus, and Papio anubis) have home range sizes from about 4-24 km 2 with an average of 15 km 2 based on 8 study areas and 21 groups of baboons 36. Gorilla (all species) has home range sizes spanning from 3-40 km 2, with most between 10-30 km 2 based on 10 groups 37. Chimpanzee and bonobo (Pan troglodytes and Pan paniscus) home range size varies from 4 to >50 km 2, but is usually between 10-26 km 2, with larger home range sizes found among savanna chimpanzees 38,39. Given the savanna elements in the fossil fauna of Sterkfontein and Swartkrans one might expect the hominins to have ranged over distances similar to, or greater than, those of savanna chimpanzees, especially as bipedalism has been linked to increased energetic efficiency for long distance travel 27. However, both Australopithecus africanus and Paranthropus robustus consumed significant quantities of C 4 vegetation 40. This may have allowed them to meet their energetic demands without ranging widely in search of preferred forest resources as is required for savanna chimpanzees. Ultimately, coupled carbon and strontium isotope data could allow investigation of the potential relationship between hominin landscape use and diet, but there are presently only two specimens for which both carbon and strontium isotope data are available (Sts 72 and SK 876). WWW.NATURE.COM/NATURE 5
Supplementary Figure 1. Comparison of 87 Sr/ 86 Sr measured by solution and laser ablation multi-collector inductively coupled plasma mass spectrometry (MC-ICP- MS). Open circles represent modern rodent enamel from an owl roost at Gladysvale Cave 17 and solid circles represent fossil rodent enamel 1 from Sterkfontein (Member 4) and Swartkrans (Member 1). Laser ablation MC-ICP-MS differed from true solution values by 0.0002 ± 0.0006 (2σ; n = 24) for modern rodent enamel 17 and 0.0005 ± 0.0010 (2σ; n = 19) for fossil rodent enamel 1. Given the broad range of biologically available 87 Sr/ 86 Sr in this region of South Africa (at least from 0.712 0.774), the additional error acquired by using laser ablation MC-ICP-MS rather than solution MC-ICP-MS is negligible for this application. For example, the external error for laser ablation MC-ICP- MS (Supplementary Table 4) is far smaller than most intra-tooth variability within the hominin specimens themselves (shown as whiskers in Fig. 2). Also, none of the nonlocal assignments of modern or fossil rodents would change in the previous studies 13,14 if solution values were used rather than laser values. WWW.NATURE.COM/NATURE 6
Supplementary Figure 2. MD * BL diameters for australopith teeth (MD = mesiodistal, BL = buccolingual; measurements in mm 2 ; Supplementary Table 2). Small open squares represent the mean, boxes indicate ±1 standard deviation from the mean, and whiskers represent minimum and maximum values. Sample descriptive statistics are based on all WWW.NATURE.COM/NATURE 7
measurable teeth recovered from Sterkfontein, Makapansgat, Kromdraai, Swartkrans, and Drimolen. Relative size of the canines does not change appreciably if assessed through MD or BL diameters alone rather than by MD * BL diameter. The only potential exception is SK 94 which has a very large MD diameter but an average BL diameter. WWW.NATURE.COM/NATURE 8
Supplementary Table 1. 87 Sr/ 86 Sr of fossil tooth enamel from Sterkfontein ( Sts and TM specimens) and Swartkrans ( SK and SKX specimens). Individuals are demonstrably non-local if their 87 Sr/ 86 Sr is outside the range of 87 Sr/ 86 Sr for plants and animals collected on the Malmani dolomite (0.72079-0.73393). ++ = above the mean in mesiodistal * buccolingual area (MD * BL) for the hypodigm by greater than 1 standard deviation (SD); + = above the mean by less than or equal to 1 SD; - = below the mean by less than or equal to 1 SD; -- = below the mean by greater than 1 standard deviation; mand. = mandibular, max. = maxillary, C = permanent canine, M3 = permanent third molar, * = samples analyzed by solution MC-ICP-MS rather than laser ablation MC-ICP-MS. Specimen # Taxon 87 Sr/ 86 Sr External error (2σ) # Laser scans Nonlocal? Tooth Size Tooth Sts 51 Australopithecus africanus 0.72897 0.00202 15 -- mand. C Sts 54 Australopithecus africanus 0.73693 0.00269 12 yes - max. M3 TM 1561 Australopithecus africanus 0.73495 0.00075 8 yes - max. M3 Sts 48 Australopithecus africanus 0.73010 0.00055 4 - max. C Sts 72 Australopithecus africanus 0.72908 0.00156 9 - max. M3 Sts 28 Australopithecus africanus 0.72729 0.00045 15 ++ max. M3 TM 1518 Australopithecus africanus 0.72652 0.00023 8 + mand. M3 Sts 3 Australopithecus africanus 0.73273 0.00191 4 ++ mand. C SK 96 Paranthropus robustus 0.74544 0.00215 13 yes -- mand. C SK 835 Paranthropus robustus 0.73235 0.00057 8 - max. M3 SKX 241 Paranthropus robustus 0.74155 0.00567 4 yes -- mand. C SK 92 Paranthropus robustus 0.73013 0.00052 4 - max. C SK 95 Paranthropus robustus 0.73954 0.00321 4 yes -- max. C SK 22 Paranthropus robustus 0.72781 0.00038 13 + mand. M3 SK 836 Paranthropus robustus 0.73099 0.00123 9 ++ max. M3 SK 87 Paranthropus robustus 0.73100 0.00080 4 ++ mand. C SK 4 Paranthropus robustus 0.73654 0.00209 4 yes ++ max. C SK 94 Paranthropus robustus 0.73031 0.00103 4 + mand. C SK 38 Paranthropus robustus 0.73397 0.00108 4 ++ max. C WWW.NATURE.COM/NATURE 9
Specimen # Taxon 87 Sr/ 86 Sr External error (2σ) # Laser scans SKX 5285 Raphicerus sp. 0.72691 1 Nonlocal? SK 12363 Raphicerus sp. 0.74508 0.00224 3 yes SKX 14150 Raphicerus sp. 0.72886 0.00052 3 SKX 8494a Raphicerus sp. 0.73098 0.00028 6 SKX 8535a Raphicerus sp. 0.72724 0.00032 6 SK 457 Papio robinsoni 0.73024 0.00040 4 SK 598 Papio robinsoni 0.73085 0.00056 4 SK 623 Papio robinsoni 0.73327 0.00076 4 SK 500 Papio robinsoni 0.73107 0.00022 3 SK 521 Theropithecus sp. 0.72786 0.00130 3 SK 2162 Theropithecus sp. 0.73586 0.00144 3 yes SK 581 Theropithecus sp. 0.72583 0.00030 2 SK 5946 Connochaetes sp. 0.73450 0.00212 4 yes SKX 671 Connochaetes sp. 0.72659 0.00226 11 SKX 14769a Connochaetes sp. 0.72546 0.00096 10 SKX 9411a Connochaetes sp. 0.72580 0.00010 2 SK 2626 Equus capensis 0.73058 0.00072 3 Sts 2580 Damaliscus/Parmularius 0.71966 0.00140 4 yes Sts 2560 Hippotragus gigas 0.73002 0.00042 2 Sts 2524 Hippotragus gigas 0.72737 0.00122 2 Sts 1996 Antidorcas recki 0.73121 0.00083 10 Sts 2546 Antidorcas recki 0.72557 0.00024 2 Sts 2076 Antidorcas recki 0.73091 1 SKX 8455a Antidorcas marsupialis 0.728451 * SKX 4878 Antidorcas marsupialis 0.722543 * SK 2304 Tragelaphus sp. 0.73248 0.00040 6 SK 2576 Tragelaphus sp. 0.73206 0.00256 3 SK 4076 Procavia antigua 0.73845 0.00042 3 yes SK 182 Procavia antigua 0.72534 1 SK 161 Procavia antigua 0.73110 1 Sts 108 Procavia antigua 0.72372 1 Sts 107 Procavia antigua 0.72416 1 Sts-Procavia b Procavia sp. 0.72232 0.00052 2 Sts-Procavia g Procavia sp. 0.72234 0.00240 2 Tooth Size Tooth WWW.NATURE.COM/NATURE 10
Specimen # Taxon 87 Sr/ 86 Sr External error (2σ) # Laser scans Nonlocal? Tooth Size Tooth SKX 653 Procavia transvaalensis 0.72635 0.00032 5 SKX 3170 Procavia transvaalensis 0.727938 1 SKX 1373 Procavia transvaalensis 0.727855 1 SK 199 Procavia transvaalensis 0.728447 1 WWW.NATURE.COM/NATURE 11
Supplementary Table 2. MD * BL measurements of australopith teeth (MD = mesiodistal; BL = buccolingual). Mand. = mandibular, max. = maxillary, C = canine, M3 = third molar. Specimen # Taxon Tooth MD diameter (mm) BL diameter (mm) MD * BL (mm 2 ) Sts 3 A. africanus mand. C 10.5 12.1 127.1 Sts 28 A. africanus max. M3 15.2 17.9 272.1 Sts 48 A. africanus max. C 9.2 9.5 87.4 Sts 51 A. africanus mand. C 8.5 9.2 78.2 Sts 54 A. africanus max. M3 13 14.9 193.7 Sts 72 A. africanus max. M3 12.4 14.7 182.3 TM 1518 A. africanus mand. M3 16.7 14.8 247.2 TM 1561 A. africanus max. M3 13.6 15.1 205.4 SK 4 P. robustus max. C 8.9 10.3 91.7 SK 22 P. robustus mand. M3 18.2 14.7 267.5 SK 38 P. robustus max. C 9 10 90 SK 87 P. robustus mand. C 7.8 9.2 71.8 SK 92 P. robustus max. C 8.1 8.7 70.5 SK 94 P. robustus mand. C 8.3 8 66.4 SK 95 P. robustus max. C 8.1 8.4 68 SK 96 P. robustus mand. C 6.9 7.2 49.7 SK 835 P. robustus max. M3 13.9 16.6 230.7 SK 836 P. robustus max. M3 16.1 18.1 291.4 SKX 241 P. robustus mand. C 7.1 7.4 52.5 WWW.NATURE.COM/NATURE 12
Supplementary Table 3. Biologically available 87 Sr/ 86 Sr in 11 geological zones as determined from plants. A sample of modern animals (n = 19) from the Malmani dolomite had 87 Sr/ 86 Sr ranging from 0.720 0.733, in close agreement with the range of individual plants. Nested ANOVA using ranked data shows that 87 Sr/ 86 Sr ratios differ significantly across geological substrates (random effect Geology: F 10,123 = 102.329, P < 0.001); post hoc comparisons (Unequal N HSD) show that all but one (Daspoort quartzite) geological substrate differ from the Malmani dolomite (P < 0.001). There are also differences across sites within the same geological type (nested random effect Site: F 10,123 = 102.329, P < 0.001), but significant contrasts arise between only two of three sites on Archaean granite-gneiss, and between two of 10 sites on dolomite. Consequently, variation within geological substrates accounted for only a small portion of the total variance observed (variance components analysis, 12% of variance explained), whereas the between substrate variation accounts for 80% of the variance. These results strongly support the use of strontium isotope analysis as an indicator of landscape use across geological substrates. There are additional geological substrates in the region (Fig. 1), but their inclusion here would not alter our finding that a disproportionate number of small hominins are non-local. Geology # Sites Total # plants 87 Sr/ 86 Sr (range of individual plants) 87 Sr/ 86 Sr (mean of sites ± 1σ) Malmani dolomite 10 59 0.72079-0.73393 0.72864 ± 0.00305 Archaean granite-gneiss 3 18 0.72383-0.73815 0.73033 ± 0.00434 Daspoort quartzite 1 4 0.72507 0.72802 0.72663 Orange Grove quartzite/shale 2 4 0.73234 0.73904 0.73591 ± 0.00281 Magaliesberg quartzite 3 10 0.73328-0.74620 0.73842 ± 0.00356 Hospital Hill formation 1 11 0.73387-0.74030 0.73758 Black Reef quartzite 3 4 0.73489 0.75165 0.74110 ± 0.00796 Bushveld gabbro 1 10 0.71233-0.71416 0.71308 Hekpoort andesite/basalt 3 9 0.74068-0.75414 0.74548 ± 0.00578 Witwatersrand quartzite 2 10 0.74466-0.77375 0.75356 ± 0.00563 Timeball Hill shale 2 15 0.74776-0.75807 0.75326 ± 0.00330 WWW.NATURE.COM/NATURE 13
Supplementary Table 4. Modern rodent teeth used as bracketing standards during laser ablation MC-ICP-MS. Specimen 87 Sr/ 86 Sr (solution) 87 Sr/ 86 Sr (laser) laser external error (2σ) # laser scans # laser sessions 26-r1 0.737850 0.73782 0.00075 55 26 26-to10z 0.729718 0.72989 0.00028 72 32 WWW.NATURE.COM/NATURE 14
Supplementary Table 5. Typical operating conditions for laser ablation MC-ICP-MS analyses (with Edwards E2 M80 Big80 80 L min -1 rotary pump) at the Africa Earth Observatory Network (AEON) EarthLAB Facility at the University of Cape Town, South Africa. Forward power 1300 W Reflected power <5 W Cones Nickel (NuPlasma Type B) Accelerating voltage 4000 V Analyzer vacuum 3-5 x 10-9 mbar Argon gas flows Coolant 13.00 L.min -1 Auxiliary 0.90 L.min -1 Mix 0.80 L.min -1 NewWave UP 213 Sweep gas He Flow rate 0.50±0.05 L.min -1 Cleaning Run Spot size 250µm Line raster length 750µm Translation rate 50µm.s -1 Pulse frequency 10 Hz Pulse energy ~0.25 mj Energy density ~0.5 J.cm -2 Analysis Run Spot size 200µm Line raster length 750µm Translation rate 5µm.s -1 Pulse frequency 20 Hz Pulse energy ~1.35 mj Energy density ~4.35 J.cm -2 Data collection Gas background 30 s Sample 180 s Integration 0.2 s WWW.NATURE.COM/NATURE 15
Supplementary Table 6. Operation parameters for MC-ICP-MS solution analyses on the ThermoFisher Neptune used in this study at the Max Planck Institute for Evolutionary Anthropology in Leipzig, Germany. Forward power 1200 W Reflected power <4 W Cones Sample Skimmer Nickel Nickel (X-cone) Argon gas flows Coolant Auxiliary Sample gas 15 L/min 0.8 L/min 1.17 L/min Interface cones Ni Mass resolution Low (400) Lens settings Optimized for maximum signal intensity Nebulizer Elemental Scientific Inc., Microflow 100 µl/min, Perfluoroalkoxy (PFA) Sensitivity on 88 Sr 50 V/ppm Cup configuration L4 ( 82 Kr); L3 ( 83 Kr); L2 ( 84 Sr); L1 ( 85 Rb); Ax ( 86 Sr); H1 ( 87 Sr); H2 ( 88 Sr) Data collection 1 block, 50 cycles, 2 s integrations WWW.NATURE.COM/NATURE 16
Supplementary Table 7. Bootstrapped estimates (1 x 10 4 iterations) of means and measures of variance (SD = standard deviation) of maxillary M3 occlusal areas in chimpanzees (Pan troglodytes) and gorillas (Gorilla gorilla) of both sexes (data from 23 ). CI = confidence interval. Species Sex Mean SD Mean + 1 SD Mean + 2 SD -95% CI +95% CI Chimpanzee Both 91.1 16.5 107.6 124.1 85.0 97.6 F 84.5 13.3 97.8 111.1 79.2 90.1 M 97.7 16.9 114.5 131.4 91.0 104.7 Gorilla Both 218.7 35.9 254.6 290.5 206.9 231.1 F 197.1 25.3 222.4 247.8 187.4 207.0 M 240.4 31.3 271.7 303.0 227.6 254.0 WWW.NATURE.COM/NATURE 17
Supplementary References 30. Sponheimer, M. & Lee-Thorp, J. A. Enamel diagenesis at South African Australopith sites: Implications for paleoecological reconstruction with trace elements. Geochim. Cosmochim. Acta 70, 1644-1654 (2006). 31. Clarke, R. J. Latest information on Sterkfontein s Australopithecus skeleton and a new look at Australopithecus. S. Afr. J. Sci. 104, 443-449 (2008). 32. Kimbel, W. H. & White, T. D. in Evolutionary History of the Robust Australopithecines (ed Grine, F. E.) 175-192 (Aldine de Gruyter, NY, 1988). 33. Grine, F. E. in Evolutionary History of the Robust Australopithecines (ed Grine, F. E.) 223-243 (Aldine de Gruyter, NY, 1988). 34. Mucina, L. & Rutherford, M. C. The vegetation of South Africa, Lesotho and Swaziland. Strelitzia 19. (South African National Biodiversity Institute, Pretoria, 2006). 35. Siebert, F. & Siebert, S. J. Dolomitic vegetation of the Sterkfontein Caves World Heritage Site and its importance in the conservation of Rocky Highveld Grassland. Koedoe 48, 17-31 (2005). 36. Melnich, D.J. & Pearl, M. C. in Primate Societies (eds Smuts et al) (1987).in Primate Societies (eds Smuts, B. B. Cheney, D. L. Seyfarth, R. M. Wrangham, R. W. & Struhsaker, T. T.) 250-266 (Univ. of Chicago Press, Chicago, 1987). 37. Robbins, M. M. in Primates in Perspective (eds Cambell, C. J., Fuentes, A., Mackinnon, K. C., Panger, M. & Bearder, S. Kk ) 305-321 (Oxford University Press, New York, 2007). 38. Stumpf, R. in Primates in Perspective (eds Cambell, C. J., Fuentes, A., Mackinnon, K. C., Panger, M. & Bearder, S. K.) 321-344 (Oxford University Press, NY, 2007). 39. Pruetz, J. D. & Bertolani, P. Chimpanzee (Pan troglodytes verus) behavioral responses to stresses associated with living in a savanna-mosaic environment: implications for hominin adaptations to open habitats. PaleoAnthropology 2009, 252 262 doi:10.4207/pa.2009.art33 (2009). 40. Sponheimer, M. et al. Hominins, sedges, and termites: new carbon isotope data from the Sterkfontein Valley and Kruger National Park. J. Hum. Evol. 48, 301-312 (2005). WWW.NATURE.COM/NATURE 18