Tropical medicine rounds

Multifocal epithelial hyperplasia in a community in the Mayan

area of Mexico
Maria R. González-Losa1, MD, PhD, Rosa E. Suarez-Allén1, LTS,
Jaqueline Canul-Canche1, MS, Laura Conde-Ferráez1, PhD, and Nixma Eljure-Lopez2, MD

1
Laboratorio de Virologı́a, Centro de
Investigaciones Regionales,
Universidad Autónoma de Yucatán
(Virology Laboratory, Regional
Research Center, Autonomous
University of Yucatan), Mérida,
Yucatan, Mexico, and 2Centro
Dermatológico, Secretaria de Salud de
Yucatán, (Dermatology Center, Health
Services Yucatan) Mérida, Yucatan,
Mexico
Correspondence
Dr Maria R. González-Losa, MD, PhD
Centro de Investigaciones Regionales
‘‘Dr Hideyo Noguchi’’
Avenida Itzaez y Calle 59 No. 490
Centro
CP 97000 Mérida,
Yucatan
Mexico
E-mail: glosa@uady.mx
Abstract
Background Multifocal epithelial hyperplasia is a pathology of the oral mucosa which has
been reported in diverse ethnic groups. Human papillomavirus (HPV) types 13 and 32 DNA
has been detected in these lesions. The aims of this paper are to describe the epidemio-
logical and clinical characteristics of an outbreak in a rural community in the Mayan area of
Mexico and to identify a possible route of transmission through saliva.
Methods A cross-sectional study was conducted in Chemax (Yucatan, Mexico). Clinical
and epidemiological data were obtained through direct interviews. Samples of oral cells
and saliva were taken. HPV 13 and 32 were identified by polymerase chain reaction using
specific primers.
Results A total of 57 patients were studied, of whom 79.1% were aged <15 years, 38.6%
were male, and 61.3% were female. The duration of lesions ranged from one month to
50 years. Lesions were located on the lips, jugal mucosa, and more frequently, the tongue.
HPV 13 was found in all the patients and HPV 32 in none. A total of 42 saliva samples
were positive for HPV 13.
Conclusions Human papillomavirus type 13 is involved in multifocal epithelial hyperplasia
among the Mexican Mayan population. The presence of HPV 13 in cells from saliva, com-
bined with poor hygiene behaviors, may explain the familial distribution of the pathology.

Conflicts of interest: None.

geographical regions from which reports of the disease are

Introduction
Multifocal epithelial hyperplasia (MEH), also known as
Heck’s disease, is a pathology of the oral mucosa. The
disease is characterized by single or multiple well-defined
lesions, which range in size from 1 mm to 5 mm. The
lesions are nodules and papules which frequently coalesce
and have the same color as the oral mucosa. These lesions
are asymptomatic and can be found on the mucosa of the
lower and upper lips, tongue, and hard palate. The
lesions grow slowly, persist for many years, and tend to
regress spontaneously.1
Multifocal epithelial hyperplasia is more frequent in chil-
dren; it mainly occurs during the first two decades of life
and is not common in adults.2 Its frequency is equal in both
sexes, although some authors have reported a higher fre-
quency in females.2 The disease has been reported in
diverse ethnic groups. In 1965, Archard described multiple
lesions in the oral mucosa in Indians of New Mexico and
coined the term ‘‘focal epithelial hyperplasia’’, as reported
304 by González-Lopez.3 Latin America represents one of the
more common. Estrada reported the presence of multiple
lesions in the oral tissues of Caramatas and Katios Indians
from Colombia4 and MEH has also been reported in Vene-
zuela, Brazil, and Mexico.5–11
Several factors have been associated with the develop-
ment of MEH, including chronic irritation caused by
smoking, the presence of Galvanic (direct) current, vita-
min A deficiency, and household conditions such as over-
crowding.2
Most patients have at least one close relative with simi-
lar lesions, which suggests that the disease may have a
genetic predisposition.2
Molecular biology techniques such as hybridization and
polymerase chain reaction (PCR) have revealed the
presence of human papillomavirus (HPV) genotypes 13
and 32 in the lesions. Genotype 13 was the first of these
to be described and is more frequently found in this
pathology.11,12
The aims of this paper are to describe the epidemiologi-
cal and clinical characteristics of an outbreak of MEH in

International Journal of Dermatology 2011, 50, 304–309 ª 2011 The International Society of Dermatology
González-Losa et al.
a rural community in the Mayan area of Mexico and to
identify a possible route of transmission through saliva.
Materials and Methods
Study population
A cross-sectional study was conducted in Chemax, Yuca-
tan, Mexico. Chemax is a rural community in the Mayan
area of Mexico, located in the east of Yucatan State, and
has a population of around 12,764 inhabitants. More
than 80% of the inhabitants of Chemax speak the Mayan
language, and their principal economic income is
obtained from agriculture and farming.13 All participants
were patients who voluntarily sought medical care during
2005 and 2006. A total of 57 subjects with a clinical
diagnosis of MEH were included in the study.
Specimen and data collection
14
Oral cells from lesions were collected by oral brushing.
Cell samples were agitated in 5 ml of phosphate-buffered
saline (PBS) with antibiotics (penicillin 500 U/ml, strepto-
mycin 500 lg/ml, gentamicin 4 mg/ml). All participants
were asked to carry out oral hygiene procedures with
purified water and then to provide a saliva sample in a
sterile bottle. Both the cell and saliva samples were kept
at 4 C during transportation to the virology laboratory
at the Regional Research Center.
Clinical and epidemiological data were obtained
through direct interviews with patients or, in the case of
child subjects, their mothers. Interviews were carried out
by an experienced social worker.
Oral brushing cells and saliva were pelleted by centrifu-
gation and resuspended in 1 ml of PBS and stored at
)70 C until further analysis. Cell suspensions were
heated at 95 C for 10 minutes prior to PCR.15
The quality of the sample was assessed by PCR using
b-globin primers GH20 and PCO4 to confirm the pres-
ence of adequate DNA.
Human papillomavirus types 13 and 32 detection
The presence of HPV 13 DNA in the cells was determined
by PCR with specific primers that amplified a 240-bp frag-
ment of L1 gene. The PCR was performed in 50 ll reaction
mix containing 1 · PCR buffer, 3 mM MgCl2, 200 lm
dNTPs, 50 pmol of each primer, and 1.0 unit of Taq poly-
merase. Thermal cycling conditions were: 94 C for four
minutes, followed by 38 cycles of 94 C for 30 seconds,
50 C for 30 seconds and 72 C for 30 seconds, and a final
extension of 72 C for 10 minutes.11
A sample previously typified as HPV 13 by restriction
fragment length polymorphism16 was used as a positive
control. A PCR mix with water in place of DNA was
used as a negative control.
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Multifocal epithelial hyperplasia in Mexico Tropical medicine rounds 305
The presence of HPV 32 DNA in the cells was deter-
mined by PCR with specific primers that amplified 963 bp
of E2 gene. The PCR was performed in 100 ll of reaction
mix containing 1 · PCR buffer, 3 mM MgCl2, 200 lm
each of dATP, dGTP, dCTP and dUTP, 50 pmol of each
primer, and 1.0 unit of Taq polymerase. Thermal cycling
conditions were: 94 C for five minutes, followed by 35
cycles of 94 C for 60 seconds, 42 C for 60 seconds and
72 C for 60 seconds, and a final extension at 72 C for
10 minutes.17 A plasmid containing the complete HPV 32
genome (kindly provided by Dr M. Favre, Pasteur Institute,
Paris, France) was used as a positive control for the PCR
reaction.18 A PCR mix with water in place of DNA was
used as a negative control. Amplicons were electropho-
resced on an 8% polyacrylamide gel and silver-stained.
Results
Description of the population and clinical findings
Although we studied samples from 57 individuals, we
were able to obtain clinical and epidemiological data for
only 44 of these because it was not possible to conduct
interviews with 13 of the subjects.
The mean age of the patients was 12.9 years (range:
3–71 years); 61.0% were aged <10 years, 18.1% were
aged 11–15 years, 9.0% were aged 16–20 years, 4.5%
were aged 26–30 years, and 6.8% were aged >30 years.
Distribution by gender showed that 38.6% were male
and 61.3% were female.
Analysis of household characteristics showed that 84%
of subjects lived in homes with only one or two bed-
rooms, 72% lived in homes in which at least five people
shared the same room, and 34% lived in homes without
sanitary facilities. None of the patients smoked or lived
with smokers.
Eleven patients (25%) reported that no other member
of their family had the same lesions; 10 (22.7%) reported
that some family members (brothers, mothers, uncles,
aunts) who were not included in the study had lesions.
The remaining 23 (52.2%) patients came from four fami-
lies with two affected members per family, one family
with three affected members, and one family with 12
affected members.
The minimum reported duration of a lesion was one
month, and the maximum was 50 years. However, 40.9% of
patients were unable to report the duration of their lesions.
Lesions were located on the lips, buccal mucosa, and,
more frequently, the tongue. All the patients had multiple
lesions (Fig. 1).
Human papillomavirus identification
All cell samples from lesions were positive in b-globin
PCR reaction. Human papillomavirus type 13 DNA was
International Journal of Dermatology 2011, 50, 304–309
306 Tropical medicine rounds Multifocal epithelial hyperplasia in Mexico
(a) (b)
Figure 1 Multifocal epithelial hyperplasia. (a) A 39-year-old woman displays multiple papules on the tongue mucosa.
(b) An 8-year-old girl exhibits papules on the lower lip. (c) A 4-year-old boy shows papules on the upper lip.
present in all the samples, but HPV 32 was not detected
with the specific PCR. Of the 48 saliva samples collected,
42 (87.5%) were positive for b-globin in exfoliated cells,
and six (12.5%) were negative and were therefore
excluded from further analysis. All b-globin-positive sam-
ples were positive for HPV 13.
Discussion
The aims of this paper are to describe the epidemiological
and clinical characteristics of an outbreak of MEH in a
rural community in the Mayan area of Mexico and to
identify a possible route of transmission through saliva.
Herein, we report for the first time an outbreak involv-
ing 57 cases in southeast Mexico. Previously, this condi-
tion was described by González-Lopez3 as occurring in
diverse groups in Mexico. Rojas19 first reported clinical
cases in 1971, and Paz-Bueso et al. 20 described patholog-
ical findings from lesions in one patient in 1986. Two
other studies have shown low prevalences of the disease.
Sedano21 reported a prevalence of 0.05% amongst
32,022 children from different states in Mexico, and
González-Lopez3 studied two communities from Mexico
State, one including 2,854 mestizo (mixed race) children
and the other including 426 native Indian Mazahua
infants, and identified 30 (1%) and 31 (7%) cases, respec-
tively.
A retrospective study carried out by Ledesma-Montes
et al. analyzed 1,000 dermatology patient files in Manuel
Gea Gonzalez Hospital in Mexico City and found nine
cases (0.9%) with a clinical diagnosis of MEH.4
In terms of geographical regions, the highest numbers
of reports of MEH come from Latin America. Cases of
MEH have been identified in Guatemala,22 Peru,23 Vene-
zuela,2 Colombia12, and Brazil.8 One outstanding study
International Journal of Dermatology 2011, 50, 304–309
González-Losa et al.
(c)
performed in Peru included the entire school-age popula-
tion within an administrative district (3,877 children) and
identified 1,465 cases of MEH, which represents the high-
est prevalence (37.8%) reported worldwide.23
This work included 57 cases from a single community,
which is one of the highest numbers of cases covered in a
single study to date and which represents the highest prev-
alence found in Mexico. To our knowledge, this work
reports the highest number of cases confirmed with molec-
ular methods worldwide. At present, it is widely accepted
that MEH is caused by papillomavirus, specifically types
13 and 32. In this study, we used specific PCR primers to
identify both HPV types; genotype 13 was found in all
patients, and genotype 32 was found in none.
Only two previous studies carried out in Mexico used
viral identification. The first of these, performed in 1987,
identified HPV 13 in seven patients.24 More recently, in
2004, another study reporting 22 cases identified HPV 13
in 17 patients, HPV 32 in three, and neither genotype in
two patients.25
Our findings are in agreement with these studies and
thereby support the suggestion that, in the Mexican popu-
lation, genotype 13 is the causal agent of MEH. Very few
studies worldwide have included molecular diagnosis.
However, HPV 13 has been found in Colombia,11,12 in
Spain in an Ecuadorian child,26 and in Norway.27
Human papillomavirus type 32 was identified for the
first time in Greenland and in Denmark19 and has been
found in patients from Germany, Israel, and South
Africa.28–30
Multifocal epithelial hyperplasia has been supposed to
show a genetic predisposition, mainly because it has been
found in specific ethnic groups; however, this pathology
has also been described in groups with diverse genetic
backgrounds. Moreover, very few of the studies available
ª 2011 The International Society of Dermatology
González-Losa et al.
support this hypothesis. In 2004, García-Corona et al.
reported that allele HLA-DRB1*0404 is significantly
more frequently found in MEH patients in Mexico, com-
pared with healthy individuals from the same ethnic
group.25
In addition, MEH has been considered to represent a
familial disease by several authors.2,3,31 In our study, we
found 23 patients belonging to only four families, in one
of which 12 members were affected.
In order to evaluate the possibility of HPV 13 transmis-
sion through saliva, we examined saliva samples collected
from patients and found that the virus was present in the
cells contained in the saliva. As far as we know, this is
the first time such a finding has been reported. The pres-
ence of the virus in saliva, along with the habit of sharing
dishes and cutlery in combination with poor hygiene con-
ditions, may explain why this pathology is preferentially
found in rural areas in which sanitary facilities are lack-
ing or suboptimal and hygiene habits are poor. We found,
for example, that although purified water is available for
children in schools in the community, the children all
share the same cup. This and other similar behaviors
represent a potential source of transmission of diverse
diseases. It is also important to consider the possibility of
transmission by close contact such as that in kissing.
Furthermore, it was noticeable that most patients pre-
sented dental decay, which is directly related to poor oral
hygiene habits.
Multifocal epithelial hyperplasia is a pathology that is
usually found in children and only occasionally in adults
because it generally resolves spontaneously early in life.
In this work, we found seven (12.2%) patients aged
>18 years, including one 71-year-old patient. None of
these were able to recall the exact duration of their
lesions, but they all agreed that it had been very lengthy.
This indicates that the virus was present in this popula-
tion long before the children in the present study devel-
oped the lesions, which raises questions about why the
adults had not eliminated the virus and whether this out-
break was associated with environmental changes or with
recent changes in nutrition that make children more sus-
ceptible.
Vitamin A deficiency has been proposed as a factor
associated with the development of MEH.2 In this work,
we intended to identify the presence of such deficiencies
through interviews concerning dietary information,
validated by the Faculty of Nutrition at the Universidad
Autónoma de Yucatán. However, many of the patients or
their mothers were unable to provide enough information
(they replied that they did not remember), and therefore
it was not possible to consider this variable in our analy-
ses. Given the important role played by vitamin A in the
maintenance of the epithelia, its mechanisms of action in
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Multifocal epithelial hyperplasia in Mexico Tropical medicine rounds 307
MEH must be properly determined before oral supple-
ments can be proposed as a therapy coadjutant.
Multifocal epithelial hyperplasia is considered a benign
and mild disease. However, confluent and multiple lesions
lead to cosmetic problems, mainly in adolescents and
young adults who seek medical care in such circum-
stances. Although HPV 13 is a non-oncogenic virus, it
has been postulated that chronic inflammation derived
from chronic trauma can cause malignant cell transforma-
tion.32 Recently, a case was published in which the
trauma produced by an overdenture caused squamous cell
carcinoma to develop.33 In MEH, the lesions are usually
exposed to continual injury as a result of mastication,
which may cause malignant transformation in the long
term. However, this is very rare.
Therefore, this pathology should not be underesti-
mated, and any doctor or dentist who identifies it should
refer the patient for treatment. Many different treatment
modalities have been performed worldwide, including sur-
gery, cryosurgery, laser procedures, systemic interferon-a,
and topic interferon-b.34 Based on our results, we con-
sider MEH to be an infectious disease caused by HPV,
which can be transmitted through the common use of
objects contaminated with saliva, and therefore families
from communities in which hygiene conditions are poor
and access to medical and dental services is limited are
more susceptible to this disease. In the Mayan community
in southeast Mexico, HPV type 13 is the causal agent of
this pathology.
Acknowledgments
This project was supported by the Secretaria de Salud de
Yucatán and the Universidad Autónoma de Yucatán. We
thank Zoila M. Jiménez Pacheco for her collaboration
with the photographs and Janet Colli Pech for technical
support.
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