Quaternary International 398 (2016) 129e135

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Quaternary International
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Dental calculus reveals potential respiratory irritants and ingestion of

essential plant-based nutrients at Lower Palaeolithic Qesem Cave
Israel
Karen Hardy a, *, Anita Radini b, c, Stephen Buckley b, Rachel Sarig d, g, Les Copeland e,
Avi Gopher f, Ran Barkai f
a
ICREA (Catalan Institution for Research and Advanced Studies), Departament de Prehisto ria, Facultat de Filosofia i Lletres,
Universitat Autonoma de Barcelona, Spain
b
BioArCh, University of York, UK
c
University of Leicester Archaeological Services (ULAS), School of Archaeology and Ancient History, University of Leicester, UK
d
Dan David Center for Human Evolution and Biohistory, the Steinhardt Museum of Natural History and National Research Center, Tel Aviv University, Israel
e
Faculty of Agriculture and Environment, University of Sydney, Australia
f
Department of Archaeology and Near Eastern Cultures, Tel Aviv University, Israel
g
The Department of Orthodontics, the Maurice and Gabriela Goldschleger School of Dental Medicine, Tel Aviv University, Israel

a r t i c l e i n f o a b s t r a c t

Article history: Reconstructing detailed aspects of the lives of Lower Palaeolithic hominins, who lived during the Middle
Available online 18 June 2015 Pleistocene, is challenging due to the restricted nature of the surviving evidence, predominantly animal
bones and stone tools. Qesem Cave, Israel (420e200 ka) is a site that has produced evidence for a wealth
Keywords: of innovative features including controlled use of fire, represented by a repeatedly used hearth.
Lower Palaeolithic Numerous charred bone and stone tools as well as wood ash have been found throughout the ten metres
Diet
of archaeological deposits. Here, we describe the presence of a range of potentially inhaled, and ingested,
Plants
materials extracted from samples of dental calculus from the Qesem Cave hominins. These finds offer an
Respiratory irritants
Dental calculus
insight into the environment in and around the cave, while micro-charcoal highlights the need for smoke
management in enclosed environments. Plant fibres and a phytolith may be evidence of oral hygiene
activities or of using the teeth to work raw materials. Starch granules and chemical compounds provide a
direct link to ingested plant food items. This extends the evidence for consumption of plant foods
containing essential nutrients including polyunsaturated fatty acids and carbohydrates, into the Lower
Palaeolithic. Together, these results represent a significant breakthrough towards a better understanding
of Middle Pleistocene dietary breadth and highlight some of the challenges facing the adoption of the
habitual use of fire for cooking by the Qesem Cave hominins, as well as offering an insight into their
ecological knowledge and technological adaptability.
© 2015 Elsevier Ltd and INQUA. All rights reserved.

1. Introduction economic and cultural development of the human species, even-

The Middle Pleistocene was a period of major biological and
behavioural change in human evolution (Nowell and White, 2010).
Qesem Cave is a karst chamber cave in Israel (420,000e200,000 BP)
(Gopher et al., 2010; Mercier, 2013) that has been in excavation
since 2001 (Fig. 1). Its finds add weight to the notion that the Late
Lower Palaeolithic period was crucial in terms of biological,
* Corresponding author.
E-mail address: khardy@icrea.cat (K. Hardy).
tually leading to the appearance of anatomically modern humans as
well as Neanderthals. At Qesem Cave, evidence for innovative
behaviour includes development of a new mode of adaptation,
possibly triggered by the disappearance of elephants, which led to
an increasing need to hunt the abundant medium-sized ungulates
(Stiner et al., 2009; Ben-Dor et al., 2011; Hershkovitz et al., 2011;
Barkai and Gopher, 2013; Blasco et al., 2014; Shahack-Gross et al.,
2014). This in turn led to a requirement for higher efficiency in
processing food (Ben-Dor et al., 2011; Barkai and Gopher, 2013), to
maximize dietary yield of both animal and plant based calories
(Groopman et al., 2015). The use of fire inside Qesem Cave, recorded

http://dx.doi.org/10.1016/j.quaint.2015.04.033
1040-6182/© 2015 Elsevier Ltd and INQUA. All rights reserved.
130 K. Hardy et al. / Quaternary International 398 (2016) 129e135
Fig. 1. Qesem Cave.
from 400,000 years ago (Karkanas et al., 2007; Mercier, 2013), and
the repeated use of a central hearth from 300,000 years ago
(Gopher et al., 2010; Mercier, 2013; Blasco et al., 2014; Shahack-
Gross et al., 2014), may be linked to this. The Amudian industry
that dominates the cave's lithic assemblages is a distinct blade-
dominated late Lower Palaeolithic Acheulo-Yabrudian Cultural
Complex (AYCC) industry (Gopher et al., 2005; Barkai and Gopher,
2013). It post dates the Acheulian industry and predates the Mid-
dle Palaeolithic Mousterian traditions.
Eight hominin teeth from Qesem Cave have been studied thus
far (Hershkovitz et al., 2011) and additional teeth are presented in
Hershkovitz et al., 2016). Here, we present evidence for potentially
inhaled and ingested material in dental calculus extracted from
three of these teeth. Dental calculus is formed by bacteria and
calcium phosphate salts which combine to create calculus. Recent
studies have shown that dental calculus can act as a store for
inhaled and ingested material, with evidence for chemical com-
pounds surviving at least into the Middle Palaeolithic (Hardy et al.,
2012). Though the Qesem Cave samples were extremely small, we
have been able to provide the earliest evidence of exposure to
potential respiratory irritants, including micro-charcoal, pollen and
evidence for fungal spores, in human history. Our results also
extend the survival of chemical compounds into the Lower Palae-
olithic and offer the first direct evidence that the diet of these
hominins included plants, incorporating both carbohydrates and
the essential polyunsaturated fatty linoleic and linolenic acids.
2. Materials and methods
Most archaeological human remains at Tel Aviv University,
Israel, are stored in individual closed boxes; in the case of the in-
dividual teeth, these were stored individually in closed plastic bags,
within the closed boxes. Three dental calculus samples from three
teeth were identified and extracted. Tooth G/9b 520e525, was
found in the mid-area of the cave, while the teeth G/22 685e690
and G/22 705e710 were found in the same location in the cave, but
at slightly different elevations. It is possible they may be from the
same individual, as their age of death estimates are similar and the
interproximal facets on the teeth articulate well together. However,
this cannot be unequivocally confirmed (Hershkovitz et al., 2011).
Extraction took place inside a specially constructed box, covered in
aluminium paper. As soon as the samples were removed, the paper
was folded and inserted individually into Eppendorf tubes; they
remained in these tubes until their extraction for analysis. Ideally,
each sample of dental calculus should be split into two for different
analyses to be conducted on each piece; however, here, the largest
sample weighed only 0.36 mg and was too small to split. The largest
piece was therefore selected for chemical analysis using the dual
methods of sequential thermal desorption-gas chromatogra-
phyemass spectrometry (TD-GC-MS) and pyrolysis-gas chroma-
tography mass spectrometry (Py-GC-MS). Optical microscopy, for
morphological examination, was conducted on the two smaller
pieces [S1].
2.1. Chemical analysis
TD/Py-GC-MS was performed on a CDS Pyroprobe 2000 via a
CDS1500 valved interface (320
C), to a HewlettePackard 5890
Series II GC fitted with a split injector (280
C) interfaced to a Trio
1000 mass spectrometer (electron voltage 70 eV, filament current
220 mA, source temperature 200
C, multiplier voltage 450 V,
interface temperature 300
C). The acquisition was controlled by
Windows based MasSpecII32 Data System, in full scan mode
(35e650 amu). The sample was weighed into a quartz tube with
glass wool plugs. The tube was placed into a pyroprobe platinum
heating coil and sealed into the valved interface. The sample was
 K. Hardy et al. / Quaternary International 398 (2016) 129e135
thermally desorbed at 310
C for 10 s in open split mode at 30 mls/
minute. The GC temperature program and data acquisition were
commenced at the same time. Separation was performed on a fused
silica capillary column (30 m  0.25 mm i.d) coated with 0.25 um
5% phenyl methyl silicone (DB-5). Initially the GC was held at 35
C
for 5 min and then temperature programmed from 35
C to 320
C
at 6
C min and held at final temperature for 15 min (total time
67.5 min), with He as the carrier gas (constant flow 1 ml/min, initial
pressure of 45 kPa, split at 30 mls/min). The run was repeated with
the same sample being pyrolysed at 610
C for 10 s with the split
open. Peaks were identified on the basis of both their mass spectra
(NIST Mass Spectral Database and additional data referenced
below), and relative retention times (relative retention indices
(RRIs)).
2.2. Optical analysis
Samples G/9b 520e525 and G/22 705e710 were degraded for
microscopic microfossil analysis. Samples for optical microscopy
were removed in the laboratory in Barcelona, weighed, and then
the two smaller samples were transferred directly into new tubes
for processing. The larger sample was taken to York for chemical
analysis. Samples were degraded, mounted and analysed on the
same day, to reduce all possible contamination or destruction of the
sample. The laboratory in Barcelona is cleaned weekly, and
contamination control samples are also evaluated at regular in-
tervals. Two control samples of sediment were also taken from the
cave deposits and examined. Other than small flecks of charcoal,
which appeared of a similar size to those found in the samples, no
other organic microfossils were found in the sediment samples.
Additional laboratory procedures to avoid contamination of mod-
ern and ancient origin followed established methods fully
described elsewhere (Hardy et al., 2009; Warinner et al., 2014). The
calculus samples were suspended in 1.5 ml of 0.6 M HCl. After
15 min, the samples were centrifuged at room temperature at
13,000 rpm for 15 min and the pellet washed in distilled water. All
pellet material was transferred onto microscope slides, in 50%
glycerol in water. The slides were sealed and left for 12 h. The first
microscopic analysis was conducted on an Olympus IX 71 inverted
microscope using magnifications between 50 and 200, while
imaging was conducted using a Colour View camera and Cell D
imaging system. All visible material was photographed. Some
pieces of debris had adhering minute flecks of calculus that had not
degraded. As the dental calculus had not dissolved completely, a
second microscopic analysis was conducted at a later date. In
Table 1
Results of dental calculus samples analysed from Qesem Cave.
Sample Tooth Analysis Age
numbers
C1 G/9b 520 Upper right Optical >300,000
e525 canine. microscopy
C1G/22 685 PM21 35 TD, Py GCeMS >300,000 probably 300
e690 Lower left e400,000
canine
P3 G/22 705 PM2 L Lower P3 Optical >300,000 probably 300
e710 microscopy e400,000
131
Fig. 2. Reconstructed total ion chromatogram of the pyrogram (pyrolysis profile)
(610
C for 10 s) of sample G22 685e690, tooth PM21 35 (0.36 mg), after thermal
desorption (310
C for 10 s). Peak identities (‘x’ indicates carbon chain length): filled
squares, Cx indicates n-1-alkenes; filled circles, Cx indicates n-alkanes. Also shown is
carbon dioxide (CO2).
addition to the contamination controls we had in place, this also
confirms the integrity of the microfossils as we were able to witness
the adhering calculus material degrade when we injected further
HCl into the microscope slides. Once the flecks of calculus were
located, a part of the sealant was removed and HCl (8% solution)
was injected into the slide to allow the calculus matrix to dissolve
in controlled conditions. The second suite of analyses was con-
ducted using a Zeiss compound microscope with magnifications up
to 800. All visible material was recorded and imaged, although
the injection of HCl into the slides generated some small bubbles
that are visible in the pictures.
3. Results
Results of the chemical analysis are consistent with high
abundances of linoleic (C18:2) and linolenic acids (C18:3) character-
istic of plant oils that are most concentrated in seeds (Fig. 2, Table 1,
[S1]). The pyrolysis-GC-MS trace was dominated by C8 to C16 n-1-
alkenes and n-alkanes, typical of unsaturated and saturated acyl
lipids (Buckley et al., 1999). Notably, the results display a bimodal
distribution with maxima at C11/C12 and C14. Although the rela-
tionship between the bound/biopolymer-derived n-1-alkenes and
n-alkanes and the fatty acids derived from the acyl lipids in the
original fat or oil is complex (Asomaning et al., 2014)[S1], particu-
larly where diagenetic processes may be involved, this bimodal
distribution of hydrocarbons is indicative of a source rich in linoleic
Cultural Finds environmental
context
Yabrudian 2 pollen grains (Pinus spp.)
Plant fibres, one smooth/sinuate long cell phytolith
10 starch granules,
Micro-charcoal,
1 fungal spore
Mineral grit
Amudian Fat/oil (acyl lipid) high in linoleic and linolenic acids (e.g. seeds fats such as
Pinus spp., walnuts)
Amudian 1 pollen grain (Pinus spp.)
Plant fibres,
Micro-charcoal,
Lepidoptera (butterfly/moth) wing scale
2 fungal spores
Mineral grit
132 K. Hardy et al. / Quaternary International 398 (2016) 129e135
Fig. 3. Microfossils extracted from dental calculus. A ¼ starch granules (G/9b 520e525), B ¼ microcharcoal (G/22 705e710), C ¼ fungi, type 1 spore (G/9b 520e525), D ¼ fungi, type
3 spore (G/22 705e710), E ¼ fungi, type 2 spore (G/9b 520e525), F ¼ pine pollen grain (G/9b 520e525), G ¼ Lepidoptera (moth and butterfly) (G/22 705e710), wing scale, H ¼ long
smooth long cell phytolith (G/9b 520e525), note fragments of calculus still adhering to it.
and linolenic acids (Niimura et al., 2003). Ten intact starch granules
were found in sample G/9b 520e525 (Fig. 3, Table 1, [S1]).
Fragments of micro-charcoal (5e60 mm) and soot, which is
present in the form of ‘black dots’ consisting of the carbon skeleton
of organic debris, were recovered as were three pollen grains,
identified as pine (Pinus sp.) (Moore et al., 1991). In two cases, very
small fragments of dental calculus were still attached to the grains
which had evidence of some deterioration and distortion, most
likely as a result of diagenetic processes. Three fungal spores from
three different species were also recovered (Fig. 3), although closer
identification is not currently possible [S1]. Long plant fibres were
found in both samples. A single, long cell phytolith of the type
found in leaves and stems of many monocotyledon and some
dicotyledon plants (Albert et al., 2007) was also found [S1]. A wing
scale of either a butterfly or moth (Lepidoptera) was recovered from
sample G/22 705e710. The wing scale was lamellar, with a narrow,
oval shape and the characteristic pedicel clearly visible. Small cal-
culus particles were still attached to it. Both the fungal spores and
insect remains included the presence of chitin, the main structural
polysaccharide in these species.
4. Discussion
Despite the extreme age of the Qesem dental calculus samples
and the limited amount of material available for study, the findings
shed light on hitherto unknown aspects of Middle Pleistocene
hominin life. We have also demonstrated the survival of a wide
range of material items of debris and chemical compounds well
into the Lower Palaeolithic and we have highlighted the value of
dental calculus as a source of biographical detail deep into the
human past.
4.1. Environment
The presence of small micro-charcoal fragments in the calculus
is striking. Particles up to 70 mm habitually enter the mouth during
oral breathing (Se et al., 2010) and while these fragments could also
result from ingestion of char adhering to roasted food, their size
suggests they result from accidental inhalation (Pope III and
Dockery, 2006). These findings, combined with the sedimentary
and micromorphological evidence from the cave, are indicative of
fire and suggest a smoky atmosphere inside the cave. The three
pollen grains recovered from the calculus suggest that pine species
were in the environment near the cave.
Reconstructing non-dietary aspects of the life led by Lower
Palaeolithic hominins is challenging; however, the evidence for
airborne contamination provides a fascinating new insight into
living conditions both inside and outside the cave during the
Middle Pleistocene. The control of fire is considered to be a crucial
driver in the development of the human lineage in social as well as
physical terms (Wiessner, 2014). At Qesem, evidence for combus-
tion events in different places inside the cave was found in the
lower part of the sequence. A repeatedly used hearth, dated to
approximately 300,000 BP was found in the centre of the cave
(Shahack-Gross et al., 2014). Evidence for blade production and
animal processing found around this hearth (Karkanas et al., 2007),
suggests it had become a focal point, undoubtedly also generating
social interaction. As the hearth was placed inside the cave, the
need to deal with the atmospheric pollution caused by smoke was
 K. Hardy et al. / Quaternary International 398 (2016) 129e135
necessary. Open fires, and the smoke they generate, have many
benefits including warmth, cooking, insect control, light, food and
fuel drying, and food flavouring (Bruce et al., 2000). The apparent
advantages of placing the central hearth well inside the cave, which
may be linked to an increased intensity of cooking, and as a focus
for flint knapping and animal processing (Mercier, 2013; Blasco
et al., 2014; Shahack-Gross et al., 2014) and presumably social
interaction, suggests a need for smoke management. A smoky at-
mosphere can be an irritant and can at times cause serious health
problems; a minimum amount of smoke management would
therefore have been necessary before fire could be enjoyed as a
focus for social interaction, rather than simply an uncomfortable
necessity. Smoke can cause reactive coughing and eye irritation in
addition to potentially more serious lung problems, in particular
among children (Smith et al., 2000). Acute respiratory infection,
which has been largely correlated with internal use of biomass fuel
(wood, dung and plant (crop) residue) is still the primary cause of
death for children under five in developing countries (Bruce et al.,
2000; Smith et al., 2000) while low birth weight and infant mor-
tality has been linked to the use of wood fuel by their mothers.
Whether the Qesem Cave hominins suffered any medical reactions
caused by environmental airborne material is unknown; however,
still recently, internal exposure to biomass fuel was thought to
contribute around 4% of the global burden of disease through
‘disability adjusted life-years lost’ (Smith et al., 2000). For the very
small population groups in the Lower Palaeolithic, to have persis-
tent chronic illness undermining reproduction and hunting effi-
ciency could have been disastrous. For successful use of fire to
develop in an internal location, in a constructive way that
permitted a population to thrive, as well as being potentially useful
for social and functional purposes, the health risks, particularly to
the young, and the unpleasant immediate side effects of a smoky
atmosphere must have been small.
The location of the hearth inside Qesem Cave (Fig. 1) is unlikely
to have been accidental. The hearth was large (4 m2) (Shahack-
Gross et al., 2014), and cave features such as the location, direc-
tion and size of cave openings and the inner topography and fea-
tures of the cave are likely to have been factors in selecting its
location. Reconstruction of the cave structure and topography,
based on excavation and micromorphology (Karkanas et al., 2007),
have led to identification of a relatively large opening in the
southwest side of the cave and a possible chimney in the northern
part of the cave, as well a large quantity of piled stones, 4e5 m wide
and about 2 m high and in the form of a convex dome to the
southwest of the hearth. This pile, which may be the result of
accumulation linked to a roof collapse (Frumkin et al., 2009), is
thought to have been in place prior to the establishment of the
hearth. A stone lining around the northwest part of the hearth (see
Shahack-Gross et al., 2014), may have served as a small windbreak
for draughts coming from the chimney.
The fungal spores present in the dental calculus could have
come from airborne mould spores from either inside or outside
the cave, or from particles of ingested food. Fungal spores,
whatever their source, are potentially harmful to humans
(Sorenson, 1999). Taken together, the fungal spores, pollen and
micro-charcoal provide an interesting perspective, in suggesting
that items considered as potential allergens today were present
in the cave environment.
4.2. Diet
Reconstructing Lower Palaeolithic diet can offer an improved
perspective on human adaptation and may feed into a better un-
derstanding of modern human dietary physiology (Lindeberg,
133
2009). Dietary reconstructions are based largely on animal bones
(Stiner et al., 2009; Blasco et al., 2014), dental pathology
(Constantino et al., 2010), microwear patterns on teeth (Pe rez-
Perez et al., 2003) and stone tools (Lemorini et al., 2006), and sta-
ble isotope analysis (Schoeninger, 2014). The focus has largely been
on animal products; however evidence from tooth microwear
patterns from two European Lower Palaeolithic sites, suggests a
highly abrasive diet including seeds, nuts, roots and bulbs (Pe rez-
Perez et al., 2003) while carbonized remains of seeds and nuts
from the site of Gesher Benot Ya'aqov, Jordan Valley, may indicate
their exploitation 780,000 years ago (Goren-Inbar et al., 2002).
Linoleic and alpha-linolenic acids are polyunsaturated fatty
acids that are not synthesized by animals and must be obtained
from dietary sources in sufficient quantity for the body to function
(Ellie and Rolfes, 2008). The clearly detectable presence of these
essential polyunsaturated fatty acids in the dental calculus from
Qesem Cave is therefore highly significant. These compounds are
abundant in the seeds of edible plant species such as pistachia and
linseed; however, seeds/nuts from Pinus halepensis fit particularly
well with the evidence and these would have been readily available
in the environment around Qesem Cave.
Starch occurs as insoluble, semi-crystalline granules in most
plant tissues, but is found in largest amounts in seeds and
storage organs. The size and form of the starch granules found
in the dental calculus suggest they represent items of food
rather than transient leaf starch granules, which are small and
disc-like due to their synthesis in the plant in diurnal cycles of
accumulation and breakdown. Two starch granules extracted
from one of the calculus samples fit together in a way that is
suggestive of compound granules, which can occur from the
fusion of granules that develop simultaneously within a single
amyloplast in storage tissues such as seeds and tubers (Smith,
2012). Starch granules in plant storage tissues accumulate over
a long period and can differ in size and morphology between
botanical sources and seeds, tubers and nuts (Bule on et al.,
1998).
Together, the identification of starch granules and specific
chemical compounds in the dental calculus samples from Qesem
Cave, point to deliberate ingestion of essential dietary components,
most likely in the form of concentrated sources such as seeds or
nuts. The phytolith and plant fibres found in the calculus could be
the result of non-dietary activities including raw material pro-
cessing, oral hygiene (Ungar et al., 2001), or the remains of food.
Their presence may coincide with the microwear pattern on the
buccal surface of the teeth which is highly variable and comprises
striations of different lengths, depths, orientations and intensities,
suggestive of chewing on a wide range of items (see Sarig et al.,
2016).
It is unlikely that the Homo lineage could have evolved suc-
cessfully without a diet that ensured not only survival, but also the
ability to thrive, including successful reproduction. Starchy plant
foods provide the energy needed to support an enlarged brain and
the metabolic functions of other human tissues and these are far
more efficient when cooked (Wrangham, 2007, 2009; Hardy et al.,
2015). While it may be possible to survive without carbohydrates
and have a diet focused heavily on animal products, through the
process of gluconeogenesis which synthesises glucose from non-
carbohydrate sources, it is energetically more expensive (Hardy
et al., 2015). A very high fat intake will prevent protein toxicity
and provide energy for metabolism, but it comes at the cost of high
levels of ketones in the blood, which can compromise reproductive
function (Kim and Felig, 1972) and can affect the efficiency of the
brain which has an essential requirement for glucose in order to
function (Hardy et al., 2015).
134 K. Hardy et al. / Quaternary International 398 (2016) 129e135
5. Conclusion
There is little doubt that the lives and knowledge base of Lower
Palaeolithic hominins were more complex than our understanding
of archaeological remains can currently testify to. The evidence
from the Qesem Cave dental calculus samples provides confirma-
tion of a population that clearly made deliberate use of local,
nutritional, plant resources. The selection of a useful range of plant
foods requires a degree of ecological knowledge which, to date, has
not been attributed to early Homo species. However, higher pri-
mates such as chimpanzees and gorillas, habitually incorporate a
wide range of plant foods into their diet and are able to alter their
food selection according to their health (Hardy et al., 2013). It is
thought that the ecological knowledge which this ability is based
on developed over very long time periods, beginning well before
the Pan/Homo Last Common Ancestor (Huffman, 2003). Among
Australopithecine species, stable isotope analyses suggest a focus
first on C3 plants with a switch to C4 plants from 3.5 mya in certain
cases (Lee-Thorp et al., 2012). There is no reason to suggest that the
Homo lineage would have lost their knowledge of edible plants
before the onset of agriculture and evidence for the consumption of
plant materials has also been demonstrated among the more recent
Neanderthal populations (Henry et al., 2011; Hardy et al., 2012). It
seems likely that the ecological knowledge base was gradually
expanded, perhaps culminating in use of the very broad range of
plant species for which there is evidence in the Late Upper Palae-
olithic (Kislev et al., 1992, 2002; Weiss et al., 2004). When viewed
from this perspective, it is unsurprising that the Lower Palaeolithic
hominins from Qesem Cave were able to ensure a diet that fulfilled
their physiological requirements.
Overall, these results provide a perspective on the use of plant
resources by the occupants of Qesem Cave, that suggests an
ecological knowledge sufficiently developed to permit the selection
of physiologically essential plant foods. The successful use of in-
ternal hearths suggests a population that was capable of reason and
adaptability.
Acknowledgments
Qesem Cave excavations are supported by the Israel Science
Foundation, CARE Archaeological Foundation, Leakey Foundation,
Wenner-Gren Foundation, Thyssen Foundation and the Dan David
Foundation. The Spanish Ministry of Science and Innovation (grant
number HAR2012-35376) supported the dental calculus project.
Sample processing was conducted in the Archaeobotany Labora-
tory, Universitat Autonoma de Barcelona, Spain. Optical microscopy
was conducted in the Archaeobotany Laboratory, Universitat
Auto noma de Barcelona, in the Ancient Starch Analysis Laboratory
at the University of Leicester, and at BioArCh, University of York, UK.
The chemical analysis was conducted at Pharos Research, UK. Prof. J.
Garti and Prof. S. Lev-Yadun are thanked for their assistance in
identifying some elements in the calculus.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.quaint.2015.04.033.
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