Accepted Manuscript

Title: PULP PATHOSIS ASSOCIATED WITH ANCIENT

MAYA DENTAL INLAYS

Authors: M. Ramı́rez-Salomón, E. Vega-Lizama, P.

Quintana-Owen, A. Cucina, V. Tiesler

PII: S0003-9969(18)30512-0
DOI: https://doi.org/10.1016/j.archoralbio.2018.08.008
Reference: AOB 4230

To appear in: Archives of Oral Biology

Received date: 29-12-2017

Revised date: 9-8-2018
Accepted date: 17-8-2018

Please cite this article as: Ramı́rez-Salomón M, Vega-Lizama E, Quintana-

Owen P, Cucina A, Tiesler V, PULP PATHOSIS ASSOCIATED WITH
ANCIENT MAYA DENTAL INLAYS, Archives of Oral Biology (2018),
https://doi.org/10.1016/j.archoralbio.2018.08.008

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PULP PATHOSIS ASSOCIATED WITH ANCIENT MAYA DENTAL INLAYS.

Running title: Pulp pathosis in ancient Maya inlays

M. Ramírez-Salomóna*, E. Vega-Lizamaa, P. Quintana-Owenb, A. Cucinac, and V. Tieslerc

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a
School of Dentistry, Universidad Autónoma de Yucatán. Calle 61a No. 492ª por Av.

Itzáes. C.P. 97000, Mérida, Yucatán, México. marcoramirezsalomon@gmail.com,

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elmy7@hotmail.com

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b
CINVESTAV (Center for Advanced Research and Studies) National Polytechnic

Institute, IPN, Mérida Unit, Av. Tecnológico Km. 4.5, Plan de Ayala, 97118, Mérida,

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Yucatán, Mexico. pquint@mda.cinvestav.mx
c N
School of Anthropological Sciences, Universidad Autónoma de Yucatán, Km. 1 Carretera
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Merida-Tizimin, 97305, Mérida, Yucatán, Mexico. acucina@yahoo.com,
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vtiesler@yahoo.com
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Corresponding author:
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Marco Ramírez-Salomón, Calle 22 No. 266 por 31 y 33, Fracc. Monterreal, C.P. 97133.

Mérida, Yucatán, México. Tel +52 999 955 1741. email:

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marcoramirezsalomon@gmail.com
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Highlights:

 Pre-Hispanic inlaid teeth manifested low rates of pulp pathologies.

 Interventions in teeth with inlays caused mild reactive manifestations in the

pulp

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  Individuals´ age at death is not correlated to calcification

Abstract

Objective: The ancient Maya used to practice dental inlays as part of the cultural

traditions. Most of those inlays remain in place after more than one thousand years. The

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purpose of this paper is to investigate the incidence of pulp pathosis associated with

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ancient Maya dental inlays to assess the impact that such common practice had on the

population’s oral health. Design: We scored 193 anterior inlaid teeth from 107 pre-

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Hispanic Maya dentitions studied at three archaeological storage facilities (Universidad

Autonoma de Yucatan, Harvard University, Atlas of Guatemala Project). Two hundred

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eleven untreated frontal teeth of pre-Hispanic Mayan collections were used as controls.
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We performed macroscopic, radiographic and microscopic analyses to assess the
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frequency of caries, pulp calcifications, internal root resorption (IRR), and periapical
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lesions (PALs). Results: In the inlaid teeth, the frequencies of pulp calcifications, IRR,

caries and PALs were 59.8%, 2.2%, 18.5% and 19.2%, respectively. Compared with
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untreated teeth, inlaid specimens exhibited greater susceptibility to caries, pulp

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calcifications, IRR and PALs than untreated teeth (pulp calcifications: 44.5%, IRR: 0%,

caries: 1.4%, and PAL: 1.9%). Age-at-death did not have any significant influence on
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susceptibility to pulp calcifications, IRR, caries or PALs. Conclusions: We noted relatively

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low pulp irritation and a low frequency of carious infections, IRR and PALs in Mayan inlaid

teeth. However, these levels exceeded the frequencies of untreated teeth from the same
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area and time period. We follow that the cements used by the pre-Hispanic Maya to fix the

inlays into their sockets provided excellent sealing characteristics on average.

Keywords: Pulp pathosis, dental decoration, dental restorations, dental cement, oral

health, ancient Maya.

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Introduction

Injuries of teeth can trigger irritation and damage of the pulp, in the long term can

lead to pulp pathosis and necrosis, reabsorption of dentin, apical periodontitis, and

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eventually affect the individual’s health and life expectancy (Weisengreen 1986; Manzo-

Palacios, Méndez-Silva, Hernández-Carrillo, Salvatierra-Cortéz, Vázquez 2005). Any

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restorative intervention carried out without the proper technique and without long-lasting

sealing materials that can resist against adverse conditions (like in the oral cavity) is

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usually destined to a short lifespan (Anusavice 2011).. The main reason for failure of a

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restorative procedure is the insurgence of a carious lesion, which in this case is known as

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secondary caries. It is defined as an infectious lesion associated with a previously existing

restoration (Mjör 1985; Mjor, Kidd, Toffenetti, Mjör 1992; Kidd, Joyston-Bechal, Beighton
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1994; Toffenetti 2000; Mjör 2005).
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Although most dental procedures were not performed in ancient times for strictly
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therapeutic purposes, many societies practiced complex dental works and apparently
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knew how to prevent secondary dental damage (Romero-Molina 1958; Fastlicht 1971).

The pre-Hispanic Maya, who once settled the areas of what now is the southeastern
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sector of Mexico, Guatemala, Belize, Honduras and El Salvador, practiced dental

modification and decoration for non-therapeutic purposes until beyond the European
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conquest. Besides cutting and abrading dental crowns, drilling and incrustation was

popular (Romero-Molina 1958; Tiesler-Blos 2001). Dental inlays consisted of drilling frontal
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teeth (incisors and canines and on rare occasions also premolars) and then sealing into

their labial surface one or more semi-precious stones (Figure 1a). This was the most

complex of all autochthonous dental procedures because it required creating a cavity in

the crown’s labial surface into which a tiny stone piece was precisely accommodated and

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cemented (Mata-Amado 1998; Ramírez-Salomon, Tiesler, Oliva-Arias, Mata-Amado 2003;

Figure 1b). During the Maya Classic period (A.D. 250–900), approximately 20% of the

adult population sported inlaid dentitions (Tiesler 1999; Tiesler-Blos 2001).

Drilling into the enamel and dentine to insert an extraneous object mimics modern

odontological practices of filling carious lesions. In both cases the sealing materials are

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associated with risks of developing secondary caries. It is noteworthy, however, that the

few existing studies of dental inlays practiced by the pre-Hispanic Maya indicate low rates

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of secondary caries (Tiesler 1999; Tiesler-Blos 2001). Still more astounding is the fact that

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the majority of ancient dental inlays still rest in the tooth matrix of the individual over a

thousand years later (Tiesler 1999; Tiesler-Blos 2001).

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The stone inlays typically filled the artificial tooth cavity and were held in place with
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a white cement (Figure 1c). Tiesler Blos (2001; Ramírez-Salomón 2016; Tiesler, Cucina,
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Ramírez-Salomón 2017) recently documented additional amorphous fillings in culturally
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produced cavities. Because such fills have not been systematically studied it is unknown

whether the internal geometry of the void is similar to voids containing stone tessellates or
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if these fillings served as replacements once the lithic inlay fell out (Mata-Amado 1995;
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Mata-Amado 1998; Ramírez-Salomón et al. 2003).

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[Figure 1 here]
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The technicalities of drilling tooth cavities has been investigated and reproduced in

a series of experiments (Mata-Amado 1998; Ramírez-Salomón et al. 2003). These authors

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proposed that the process must be completed in 2–4 phases: after an initial enamel notch,

a lithic drill point perforated the enamel and dentine with the help of abrasive sand until the

final cavitation was achieved. The walls and the bottom of the hollow were flattened and

incrusted with a cemented stone tessellate. This sequence can be synthesized into four

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concise steps: 1. initial notching, 2. penetration, 3. shaping of the hollow, and 4. inlay.

Regarding the first step, Mata-Amado suggests that a guiding platform was used to

position the instrument during the initial cutting process (Mata-Amado 1995; Mata-Amado

1998). Additional analyses with scanning electron microscopy (SEM) of the inlay cavity

surfaces (Ramírez-Salomón et al. 2003) have revealed homogeneous concentric grooves

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representing the marks left by the instrument. The pre-Hispanic samples under scrutiny

showed that the drilling typically passed through the enamel completely and penetrated

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well into the dentine.

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Beyond technical procedures, many questions remain regarding ancient Maya

dental interventions, in particular with regards to the materials used and their possible

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applications in modern dentistry. However, this is not the goal of this paper, and the
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material properties of the sealings will be discussed in future publications. Here, we
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explore the degree of secondary pulpar damage produced by this practice.
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There is currently little empirical information as to why pre-Hispanic dental

procedures were seemingly so successful. The aim of this study is to assess the frequency
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of pulp reactions and pathosis associated with the ancient Maya dental inlays. This
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comparison sheds light on the level of the clinical efficiency of ancient Maya inlays and

related cement.
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Materials and Methods

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We studied 193 inlaid anterior teeth from 107 individuals recovered from pre-
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Hispanic Maya dentitions from three international collections. The Universidad Autónoma

de Yucatán collection (UADY) includes specimens from the pre-Hispanic sites of Xcambó,

Yaxuná, Calakmul, Caucel, Chunchucmil, Noh Bec, and Chaac. The collection curated at

the Peabody Museum includes series from Copan, Seibal, Barton Ramie, Baking Pot, and

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Holmul. The dentitions stored at the Archaeological Atlas of Guatemala include specimens

from Ixtontón, Yaltutú, Ixcol, Ix Ek, Ixcoxol, Curucuitz, Ixkun, Sacul, and the Calzada

Mopan conglomerate. In addition, a control sample of 211 anterior teeth from 31

individuals with no inlaid dentitions was included (Table 1). In order to limit unnecessary

manipulation of the skeletal remains, individual data on sex and age-at-death were kindly

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provided by the curators of the respective collections and were obtained from the archives

in which the collections were housed.

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[Table 1 here]

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The teeth were subjected to macroscopic, microscopic, and radiographic analysis.
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The information was recorded in forms and integrated into a database created in
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FileMaker® Pro 9.0 software (File Maker Inc. Santa Clara, US) for subsequent statistical
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analysis.
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Macroscopic Analyses. For general scrutiny of the specimens, we used a 5x eye loupe
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magnifier and white light. We obtained the measurements using a digital Vernier caliper

(Hangzhou Maxwell Tools Co., Ltd., China). The macroscopic analysis was performed to
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record the presence of the inlay in place, of filling/sealing material, cracking or fractures in

the enamel, to assess the position of the inlay from both the incisal and cervical edges, its
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material and shape, occlusal wear, and to detect the presence of secondary carious

lesions associated to, and located around the edges of the inlay. No other forms of caries
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were recorded. On the contrary, in the control group caries were recorded for comparative

purposes only when they affected the buccal surface of the teeth. Carious lesions

occurring in the control sample are not classified as secondary caries as the control group

was not associated with dental inlays.

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Optical Microscopy Analyses. For systematic microscopic scrutiny, we used a

stereomicroscope EZ4 HD Leica™ (Leica Microsystems GmbH, Germany) at 8x and 20x

magnification. At 8X we analyzed the dental crown from proximal and occlusal angles to

gain better information on presence, shape, color, surface and other characteristics of the

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inlay, as well as to confirm the presence of carious destruction of the dental tissue (Figure

2), cracklings and fractures. At 20X, alongside the features described above, we analyzed

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the crown to inspect the interface between the inlay and the dentine, and whereas the

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inlay was no longer in place, to assess the inner walls and bottom floor of the artificial

cavitation, the striations produced by the drill point, any material still adhering to the

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cavitation walls as well as any evidence of perforation into the pulpal chamber.
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A
[Figure 2 here]
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Radiographic Analyses. In compliance with safety requirements (established by Harvard

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University), the operator completed radiographic device initial safety training. Then, a
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sensor adapter for digital radiography equipment and a safe container for archaeological

pieces were designed. Radiographic records were obtained both in the inlaid and the
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control sample using a portable dental x-ray system (Port X II Dental, Genoray Co. Ltd.,®

Korea) at 60 kV and 2 mA. In each tooth, both ortoradial and proximal radiographs were
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obtained from a standard distance of 15 cm with each dental piece fixed on a stable

support. Image processing was performed using Owandy Krystal® radiography system
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digital X- Easy (Owandy SA, Champs sur Marne, France).

The radiographic analysis was performed to assess the extent of pulpal reaction in

response to the mechanical stress exerted by the drilling, exposure of dentine to the

external environment, the morphology of the pulpal chamber, internal root reabsorption,

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and degree of calcification of the pulpal chamber. In those cases where the inlay was still

in place, radiography also allowed any perforations of dentine to be detected and caries-

induced destruction of the inner dental tissues.

Last, periapical lesions were assessed in those cases when the tooth was firmly anchored

into its own socket. They were taken into consideration as the failure of the anthropogenic

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intervention only when they were associated to an inlaid tooth, and in the absence of other

causative factors, like cervical caries or traumas. Due to poor preservation of the skeletal

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material, lesions of the apical end of the root could be scored in a limited number of cases.

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Criteria to assess pulpal reaction

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Resting on the radiographic analysis, we developed a visual criterion to quantify

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calcification of the pulpal chamber in relation to age at death that could be used to assess
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the extent of “aggression” to the pulpal chamber by the drilling process. Based on the
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control group, which did not undergo anthropogenic interventions of the buccal surface, it

represents a comparative “physiological” pattern of calcification by age class to which

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contrast anomalous patterns (i.e., those produced by anomalous mechanical stress like
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drilling).

In first place, the 211 control teeth were organized by age class: 1) individuals up to 20
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years; 2) 20.5 to 30 years; 3) 30.5 to 40 years; 4) 40.5 to 50 years; 5) individuals older

than 50 years of age. Then, all control teeth were radiographed from a clinical and
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proximal angle in order to have a clear picture of the extent of pulpal calcification by age.

The pulpal space was divided into pulpal chamber and root canal. The height and width of
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the chamber, and the width of the canal were recorded; the height of the chamber was

also divided into incisal, middle and cervical thirds. The chamber height and width, and the

root canal width, both in their clinical and proximal views, were recorded as 1) wide; 2)

intermediate; and 3) completely calcified (Figure 3).

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The radiographic images were visually analyzed and assessed by the senior author and by

one of the coauthors (EVL). In case of disagreement in the assessment of pulpal

calcification, a third root-canal specialist from the School of Dentistry was asked to assess

the radiographs. This occurred in 14 teeth with calcification among the sample that

presented inlay (92.7% agreement), and in 10 teeth showing calcification from the control

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group (95.3% agreement). Once a final assessment was reached, an average value of

calcification was assigned to each dental piece.

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[Figure 3 here]

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Results
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Table 2 lists the absolute values and percentage frequencies of calcification and internal
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root resorption (IRR) and the presence of periapical lesions (PALs) and caries in both

inlayed teeth and in the control sample.

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[Table 2 here]
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Calcifications. Of the 193 inlaid Maya teeth, 113 (59.8%) exhibited radiographic evidence

of pulp calcification according to the calcification index established in this work, and 76
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teeth (40.2%) did not manifest any calcification (under the previously described

parameters). Four teeth could not be evaluated for calcification due to their poor
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preservation. In the control group, 94 teeth (44.5%) showed calcifications, and 117 teeth

(55.5%) did not display any signs of calcification. A Fisher’s exact test comparing pulp

calcification between the inlaid teeth and the control sample indicated a statistically

significant difference (P<0.05)

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Internal Root Resorption. Internal root resorption could be assessed in 181 teeth with

inlays (no information could be obtained for an additional 12 specimens). Internal root

resorption was detected in 4 inlaid teeth (4/181 = 2.2%). In the control group no teeth

showed any sign of Internal root resorption. Fisher’s exact test of this comparison revealed

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a statistically significant difference (P<0.05).

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Decay. Thirty-eight teeth with intentional dental drilling of the buccal surface manifested

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carious lesions (38/193; 19.7%) (Table 3). In the control group, only 3 teeth (1.4%)

exhibited carious lesions in their buccal surface. The difference was statistically significant

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(P<0.05) according to Fisher’s exact test. The majority of the caries associated with inlays
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were detected in teeth that had lost the stone (26/38) or whose filling was of unidentified
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origin (11/38). Only one tooth with a stone inlay still in the tooth manifested a carious
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lesion. Overall, the number of teeth that had lost the inlay corresponds to 80

(80/193=41.5%).
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[Table 3 here]
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Periapical Lesions. Periapical lesions could be recorded only in 52 inlaid teeth, which

were still associated with their socket. The frequency of periapical lesions was 19.2% (10
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teeth). It must be stressed that those 10 teeth affected by a periapical lesion did not

manifest any sign of trauma, fracture or other caries unrelated to the inlay that might have
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been the lesion’s causative factor. In the control group, only 4 teeth (1.9%) exhibited a

periapical lesion. Fisher’s exact test revealed a statistically significant difference between

the two populations (P<0.05).

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Age Comparison. The comparative analysis between age at death and calcification of the

pulpal chamber and root canal could be performed on a sample of 102 individuals with

inlaid teeth for which age at death was available. The three categories of calcification

index were correlated to age at death. A Pearson’s correlation returned a r2 value of 0.059

(p=0.1453), indicating a lack of relationship between calcification and age. A further

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ANOVA test was performed between age at death distribution in the three groups. Table

4a shows the descriptive statistics and results of the ANOVA test among the three

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independent variables. As we can appreciate, average age at death is 36.9 years in the

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group showing a wide pulpal chamber, 40.3 years in the intermediate group, and 44.2

years in the group showing extreme reduction/obliteration of the pulpal chamber. However,

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despite the pattern of increased mean age at death among the three different degrees of
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calcification, the ANOVA test returned results that did not reach the alpha 0.05 significance
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thresholds (F=2.67, 2 and 100 d.f., p=0.074).
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For the relationship between age at death and the presence of both caries and periapical

lesions, the sample of inlaid individuals was reorganized into two age groups (Group 1 - 17
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to 30 years of age, and Group 2 - 30+ years of age) in order to handle a binomial
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distribution for the three variables (Table 4b). These two age groups were chosen in order

to handle sample size sufficiently large to be representative. Similarly, to the above-

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mentioned analyses, also the results of a Fisher’s exact test between individuals of the two

age groups and the presence or absence of caries and periapical defects returned no
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statistically significant results (caries p=0.79; periapical defects p=0.692).

All the above indicates that age at death did not have a significant influence on
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susceptibility to pulp calcification, and infectious lesions. However, we cannot rule out that

the lack of a clear pattern between the degree of calcification and age at death can also be

due to the age of the individual when the inlaid was placed. We have no means to infer the

length of time that passed between the inlaid insertion and the moment the individual died.

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[Table 4a, b here]

Discussion

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Limited sample sizes for collective analysis have led to a scarcity of systematic

statistical scrutiny regarding the health consequences of intentional dental procedures

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performed among ancient human populations. This study was conceived therefore by

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combining the information obtained from the skeletal series scored at the Universidad

Autónoma de Yucatán, Harvard University, and the Archaeological Atlas of Guatemala.

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Considered jointly, these samples offer a number and diversity of samples that is
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otherwise difficult to achieve. This research yields results that will help us better
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understand these ancient dental procedures and their impact on the health of individuals.
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As noted above, these dental treatments were related to ritual traditions and/or

aesthetic purposes, but they could have had health consequences for their subjects. As
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documented here and in other studies, the interventions could alter pulp defenses and
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therefore reduce health status by penetrating the protecting barriers of the teeth.

Nevertheless, at least 60% of the adult Maya population underwent some type of dental
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intervention (grinding or inlay) during the first millennium A.D., potentially jeopardizing the

collective dental health status of this group (Mjör 1985; Tiesler 1999; Mjor & Toffenetti
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2000; Versiani, Sousa-Neto, Pecora 2011).

We searched for evidence of pulpal irritation or injury focused on three parameters

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that denote a dental pulp reaction to harmful stimuli: calcifications, Internal root resorption,

and periapical lesions. Our results showed that 2% of teeth exhibit Internal root resorption,

19% of teeth exhibited periapical lesions, and 60% of teeth exhibited calcifications in the

form of tertiary dentine apposition. Tertiary dentin is a reactive dentine that forms in places

12
after being stimulated by irritative conditions (Kuttler 1959; Stanley, White, McCray 1966;

Hargreaves & Goodis 2002). These foci of dentinal deposits are known as calcifications.

When the harmful stimuli are low in intensity, they generate reactionary tertiary dentin

(Hargreaves & Goodis 2002).

We found that roughly 40.2% of inlaid teeth were free of apparent calcification,

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which seems to indicate a general low pulp reaction. Since this is the immediate

physiological response to aggression, our results support the assumption of a generally

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low level of irritation that was produced either by the mechanical drilling procedure or by

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the potential reaction induced by the materials used to fix the inlay into the tooth applying

to both the operative procedure and the materials used. However, a statistical comparison

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with unaltered teeth, using Fisher’s exact test, did reveal significant differences (p=0.002)
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between the two groups. Put in context with undrilled teeth, this result demonstrates that
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pulp calcification was in part triggered by the drilling (60% versus 44.5% in the control
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sample). Such intentional mechanical stress clearly sums up to other (non-intentional)

stimuli in the inlayed individuals.

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An Internal root resorption frequency of 2.2% is close to the estimated average in

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the contemporary population (0.01–1%), according to the endodontic literature (Haapasalo

& Endal 2006; Patel, Ricucci, Durak, Tay 2010; Gabor, Tam, Shen, Haapasalo 2012).
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Most studies of the frequency of Internal root resorption have been based on radiographic

analyses, although some studies used SEM or Cone Beam Computer Tomography
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technologies (Haapasalo & Endal 2006; Patel et al. 2010; Versiani et al. 2011; Gabor et al.

2012). A micro tomography (μCT) study of six upper front teeth (Versiani et al. 2011)
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revealed that 16.7% of teeth had calcifications (1 out of 6) and that 50% of teeth had

Internal root resorption. Our findings are inconsistent with these results, however. The

Versiani (2011) sample consisted of only 6 teeth, a small sample that can be easily

affected by random factors. It is also roughly 30 times smaller than the sample of teeth that

13
we studied. In addition, the teeth studied by Versiani (2011) were donated by an unknown

collector, which makes the sample decontextualized and its origin unclear. Other globally

published statistics are based on optical analyses of radiographs, validating the

radiographic analysis presented in this study and enabling comparisons with most

contemporary Internal root resorption studies (Haapasalo & Endal 2006; Patel et al. 2010;

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Gabor et al. 2012).

Our Fisher’s exact test analysis comparing individuals in the control sample and the

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inlaid sample in relation to Internal root resorption achieved the 0.05 threshold of

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significance (p=0.044), which allowed us to conclude that individuals with inlaid teeth

exhibited greater susceptibility to Internal root resorption. For Internal root resorption to

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occur, some initiation factors are required (e.g., trauma or pulp inflammation). However,
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scholars agree that the process of reabsorption requires that the pulp tissue in the area be
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vital and that the pulp have some degree of necrosis (Haapasalo & Endal 2006). Following
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this logic, our results indicate that individuals without intervention did not exhibit high

frequencies of Internal root resorption because they were not subject to factors that
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caused resorptive pulp reactions. Our findings also support the assumption that
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interventions were made in living individuals and the teeth were stored in the mouth for

longer periods of use, which allowed resorption to progress long enough to become
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macroscopically visible before the person died (Haapasalo & Endal 2006). However, it

must be noted that despite the inlayed sample being significantly different from the control
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sample, only 4 out of 181 teeth with inlays manifested Internal root resorption. This finding

suggests that the drilling process and the permanence of the inlays did not represent an
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overwhelming aggression on the pulp.

The frequency of periapical lesions corresponded to 19.2%. Only 52 teeth were still

attached to the periapical bone, which is where pulp related bone destruction can be

detected. We again performed Fisher’s exact test analysis to compare inlayed individuals

14
and the control sample in terms of the presence of periapical lesions. The significant

difference (p=0.0001) highlights that individuals with inlays exhibited a greater

susceptibility to periapical lesions than untreated individuals. This result again suggests

that the dental interventions were performed in living individuals and held in the mouth

long enough to generate the entire chain of events that would lead to the creation of a

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periapical lesion.

The frequency of caries was 19.7%, which was consistent with the frequency of

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periapical lesions. In fact, of the 10 dental sockets that manifested a periapical lesion,

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seven were inlayed and presented a carious lesion (for the remaining three cases, we

could infer only indirectly that the corresponding tooth once presented an inlay, but we

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could not assess whether it also had a carious lesion due to the lack of dental structure).
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Considering that the ancient Maya did not perform dental work for therapeutic purposes,
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carious lesions are supposed to lead to pulp necrosis and periapical lesions, which makes
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the relation between the frequencies of caries and periapical lesions consistent with the

natural evolution of a dental infectious disease.

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Patterns of caries in inlayed teeth can be compared with modern secondary

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cavities after restoration. The current definition of this type of caries, according to the

International Dental Federation, is "the presence of carious lesion in the margins of an

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existing restoration" (Mjör 1985; Kidd et al. 1992; Kidd et al. 1994; Mjor & Toffenetti 2000;

Mjör 2005), and the current frequency of secondary caries in Latin America is 52%
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(Gómez-Chamba 2011). A rate of caries of 18.5% in pre-Hispanic Maya inlays compares

favorably with the rate of current secondary caries.

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Fisher’s exact test conducted on individuals with and without inlays in terms of

caries revealed a significant difference (p=0.0001), indicating that inlays represented a

mechanical alteration that increased susceptibility to cavities, as expected. Eleven

specimens (out of 16) whose drilling had been filled with some unknown material

15
presented carious lesions; although we have not been able to recognize the nature of this

material, it was seemingly a filling that lacked sealing and antiseptic properties. Instead, of

the remaining 27 carious lesions, 96.3% of them were found in prepared cavities with

missing inlays (26 out of 27), and 3.7% (N=1) were found in one jadeite inlay (Table 2).

These results imply that no caries was found in interventions with black stone inlays

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(formerly known as pyrites). Jadeite is inert and does not release compounds, and pyrite, a

material frequently used for inlays, could have had some protective reactive effect on the

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dentin wall. Overall, however, results indicate that the presence of a stone filling,

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regardless of its nature, granted protection to the tooth.

As discussed, Fisher’s exact test revealed that inlays increased the susceptibility of

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individuals to periapical lesions (p=0.0001). It is important to note that in non-treated
N
individuals the presence of these bone lesions was limited to cases in which the dental
A
pulp was exposed to the oral cavity for reasons unrelated to the process (dental fractures,
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trauma, cervical caries, etc).

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[Figure 3 here]
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We noted low-grade and well-tolerated pulp irritation (Figure 3). The comparative
EP

analysis of the frequency of pulp pathosis among age groups revealed that age had no

significant effect on the diseases studied. This finding can be explained by the dental
CC

intervention not responding to a natural and physiological process, thereby causing a mild

irritation that stimulates calcification and other processes. In order to determine differences
A

in pulp dimensions between teeth without mechanical interventions, it will likely be

necessary to compare individuals with age differences of up to 10 years. When the

interventions are present, differences in calcifications are clearly observable in shorter

periods of time.

16
Conclusion

This study of pulp reactions relies on the largest sample of Maya dental inlays

scored systematically for this purpose to date (N=193). We can conclude that the

PT
frequency of pulpal changes reveal that interventions were performed in living individuals

and that the pulps remained vital for long enough to exhibit the reactions. However, a low

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grade of pulp stimulation was also observed. Although the rate of carious lesions was

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higher than that in the control sample, since the buccal surface of anterior teeth is not the

most suitable surface for bacteria to adhere, only one out of five teeth that had undergone

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intentional intervention had eventually suffered from carious lesions. Moreover, more than
N
half of the inlaid specimens still presented the semi-precious stone in place after more
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than one thousand years. The reduced amount of caries and the persistence of inlays in
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their teeth indicate that the cement used to keep the stone in place was apparently very

effective. Future studies will address the issue of the chemical composition of such
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cement.
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conflict of interests
The authors declare that there is no conflict of interest
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Funding
This research was supported by Consejo Nacional de Ciencia y Tecnologia (CONACYT), Mexico
[project number CB-2010-1 n. 152105 to V.T.]
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Ethical Approval
This work was evaluated and accepted by the Bioethics Committee of the Faculty of Medicine of the
Autonomous University of Yucatan. All the inherent requirements were met, obtaining the
authorization of this committee to continue the project without restrictions.
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Acknowledgements.

This paper derives from the senior author’s doctoral dissertation. We express our

sincere gratitude to Michele Morgan, Olivia Herschensohn, Jane Lyden Rousseau,

17
and Tiffany Lee of the Peabody Museum of Harvard University; our sincere thanks

go to Julio Chi and Enrique Reyes of the Autonomous University of Yucatán; to

Mara Reyes of the Archaeological Atlas of Guatemala (IDAEH); and to Oscar R.

Bolanos from Temple University for their invaluable contributions to this research.

PT
Last but not least, we are thankful to Jose Luis Villamil and Fernando Aguilar,

former and current Deans of the School of Dentistry (UADY), respectively, and to

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Hector Rubio, chairman of the PhD program, for facilitating the investigation. This

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research was supported by Consejo Nacional de Ciencia y Tecnologia

(CONACYT), Mexico [project number CB-2010-1 n. 152105 to V.T.]

U
N
A
M
D
TE
EP
CC
A

18
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Figure legends

Figure 1. (a) Different pre-Hispanic forms of dental interventions (Atlas

Arqueólogico de Guatemala, 109-F35), (b) Close adaptation of the jadeite inlays

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(Peabody Museum 13-F6), and (c) White paste surrounding the black stone inlay

and paste filling cavities (Atlas Arqueólogico de Guatemala, 188A-F31) (photos by

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Marco Ramirez-Salomon; permission to publish granted by Atlas Arqueólogico de

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Guatemala and Peabody Museum, Harvard University).

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Figure 2. Optical microscopic image of a carious lesion associated to an inlay

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(Atlas Arqueológico de Guatemala, 017-F39) (photo by Marco Ramirez-Salomon;
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permission to publish granted by Atlas Arqueólogico de Guatemala).
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Figure 3. Proximal radiographic images of the author’s calcification index, which

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consists of three classification levels: (a) large, (b) medium, and (c) and narrow or
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completely calcified (photos by Marco Ramirez-Salomon).

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Table 1. Number of inlaid and control teeth by tooth type, and by age class.
Inlaid Maxilla Mandible Control Maxilla Mandible
teeth1
I1 52 10 I1 47 9
I2 45 5 I2 50 7
C 60 10 C 68 10
P1 10 1 P1 15 5

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Age <20 20.5-30.0 30.5- 40.5- 50.5+
classes 2 40.0 50.0
Inlaid 4 39 10 13 39

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teeth
Control 35 37 53 34 47
1 The total number of individuals that provided at least one inlaid tooth is 107.

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2 Age at death was not reported for 88 teeth with inlays and 49 control teeth.

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Table 2. Absolute and percentage values of pulp calcification, IRR, PALs, and caries in
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inlaid and control teeth (secondary caries in inlaid teeth and buccal caries in control teeth).
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Presence Absence
Pulp calcification Inlaid teeth 113 (59.8%) 76 (40.2%)
Controls 94 (44.5%) 117 (55%)
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Internal root resorption Inlaid teeth 4 (2.2%) 177 (98%)

Controls 0 (0%) 211 (100%)
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Periapical lesion Inlaid teeth 10 (19.2%) 42 (80.8%)

Controls 4 (1.9%) 207 (98.1%)
Caries Inlaid teeth 38 (19.7%) 155 (80.3%)
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Controls 3 (1.4%) 208 (98.6%)

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Table 3 New. Absolute values of caries in the Maya inlay works.
Caries presence in Maya inlayed teeth
No caries Caries
Missing inlay 54 26
Jadeite, black or other
87 1
stone
Paste fillings 9 0

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Unidentified material 5 11
Total 155 38

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Table 4a. Relationship between age at death and pulp calcification,

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Correlation* r2 = 0.059 p= 0.1453
N Mean s.d.

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Wide 41 36.9 14.84
Intermediate 26 40.3 11.98
Narrow/Obliterated 35 44.2 N 13.58
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ANOVA** F=2.67 2, 100 d.f. p=0.074
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* correlation was performed on 102 individual data;

** ANOVA was calculated based on the descriptive statistics of the three
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categorical groups
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Table 4b. Relationship between age groups and presence of both caries and periapical
lesions.
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No lesion Lesion
Periapical lesion Group 1 20 3
Group 2 17 4
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Fisher´s exact test p=0.692

Caries Group 1 34 6
Group 2 50 12
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Fisher´s exact test p=0.790

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Table 5 – Distribution of, and association among lesions (numbers in parenthesis indicate
the total number of teeth available for analysis)
Caries Periapical Internal Root Calcification
Lesions Resorption
38 (193) 10 (52) 4 (181) 113 (189)
Teeth with caries (N=38) Total
x x x x 0
x x x 3

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x x x 1
x x 3
x x 2

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x x 14
x 15
Teeth without caries (N=155) Total

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(N=155)
x x x 0
x x 2

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x x 1
x x 0
x
x
N 2
0
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x 92
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