Prevalence of HPV infection by cervical cytologic status in Brazil.
ABSTRACT To assess the prevalence of human papillomavirus (HPV) infection according to cervical cytologic status in northeastern Brazil; identify other risk factors for low- and high-grade squamous intraepithelial lesions (LSILs and HSILs); and identify the most prevalent HPV genotypes associated with the lesions.
Two cervical smears were collected from 250 women referred for cancer screening, one for cytologic examination and the other to test for the presence of HPV by PCR with genotyping by dot blot hybridization.
There were 110 healthy cervices, 82 LSILs, and 58 HSILs. The overall HPV prevalence was 48%, with higher rates for HSILs, and HPV-16 was the most prevalent type. Age, multiple sexual partners, type of HPV present, smoking, and early onset of sexual activity were risk factors for cervical lesions.
Age, multiple sexual partners, and infection with HPV-16 increased the risk of having LSILs or HSILs. Early onset of sexual activity and smoking only increased the risk of having HSILs.
Article: Esophageal squamous cell cancer in patients with head and neck cancer: Prevalence of human papillomavirus DNA sequences.[show abstract] [hide abstract]
ABSTRACT: An etiologic role for human papillomavirus (HPV) infections in either head and neck (HNC) or esophageal carcinogenesis remains debatable. Patients with head and neck cancer are at high risk for developing a second esophageal squamous cell cancer (ESCC). The aim of our study was to determine whether HPV infections play a role in this multifocal carcinogenesis. Samples from 2 groups of HNC patients were studied: Random esophageal biopsies were collected from the first group of 60 patients who had been screened for asymptomatic ESCC. The second group consisted of 21 patients with pairs of HNC and ESCC. Both the fresh frozen biopsy samples of the first group and the paraffin-embedded specimens of the second group were evaluated for the presence of HPV DNA sequences by PCR amplification, cloning and sequencing. HPV DNA sequences were detected in 66.7% of normal/inflammatory (34/51) and dysplastic and malignant (6/9) esophageal tissues from HNC patients being screened endoscopically. Similarly, in the second group of 21 patients with both HNC and ESCC, HPV DNA sequences were demonstrated in 13 (61.9%) of the HNC biopsies and in 14 (66.7%) of the ESCC biopsies. The prevalence of high-risk-type HPV 16 was low (5/51, 9.8%) in normal/inflammatory esophageal mucosa but higher (10/24, 47.6%) in ESCC. The low-risk HPV 11 was present in 37.3% (19/51) of normal/inflammatory, 66.7% (4/6) of dysplastic and 28.9% (13/45) of the carcinoma samples. The same HPV type was present in only 3/21 pairs of HNC and ESCC samples, suggesting that a clonal expansion from the HNC to a subsequent ESCC, or visa versa, is unlikely. The high prevalence of "low-risk" HPV infections points to the need for studies on possible interactions of these infections with the use of alcohol and tobacco in the pathogenesis of these tumors.International Journal of Cancer 04/2004; 109(2):253-8. · 5.44 Impact Factor
[show abstract] [hide abstract]
ABSTRACT: Clinical and subclinical human papillomavirus (HPV) infections are the most common sexually transmitted infections in the world, and most sexually-active individuals are likely to be exposed to HPV infection during their lifetimes. More than 40 genotypes of HPV infect the epithelial lining of the anogenital tract and other mucosal areas of the body; of these, 13-18 types are considered to be high-oncogenic risk HPV types (HR-HPV). Persistent infection with HR-HPVs is now unequivocally established as a necessary cause of cervical cancer and is likely to be responsible for a substantial proportion of other anogenital neoplasms and upper aero-digestive tract cancers. Low oncogenic risk HPV types (LR-HPV) are also responsible for considerable morbidity as the cause of genital warts. Youth and certain sexual characteristics are key risk factors for HPV acquisition and persistence of HPV infection, but other mediating factors include smoking, oral contraceptive (OC) use, other STIs (e.g. chlamydia, herpes simplex virus), chronic inflammation, immunosuppressive conditions including HIV infection, parity, dietary factors, and polymorphisms in the human leukocyte antigen system. Not surprisingly, these factors are also established or candidate cofactors identified in epidemiologic studies of cervical cancer. HPV transmissibility and molecular events in HPV-induced carcinogenesis have been the focus of recent multidisciplinary epidemiologic studies. This shift in research focus coincides with a shift in cancer prevention techniques towards immunization with HPV vaccines and HPV testing of precancerous lesions.Vaccine 04/2006; 24 Suppl 1:S1-15. · 3.77 Impact Factor
Article: The papillomavirus life cycle.[show abstract] [hide abstract]
ABSTRACT: Papillomaviruses infect epithelial cells, and depend on epithelial differentiation for completion of their life cycle. The expression of viral gene products is closely regulated as the infected basal cell migrates towards the epithelial surface. Expression of E6 and E7 in the lower epithelial layers drives cells into S-phase, which creates an environment that is conducive for viral genome replication and cell proliferation. Genome amplification, which is necessary for the production of infectious virions, is prevented until the levels of viral replication proteins rise, and depends on the co-expression of several viral proteins. Virus capsid proteins are expressed in cells that also express E4 as the infected cell enters the upper epithelial layers. The timing of these events varies depending on the infecting papillomavirus, and in the case of the high-risk human papillomaviruses (HPVs), on the severity of neoplasia. Viruses that are evolutionarily related, such as HPV1 and canine oral papillomavirus (COPV), generally organize their productive cycle in a similar way, despite infecting different hosts and epithelial sites. In some instances, such as following HPV16 infection of the cervix or cottontail rabbit papillomavirus (CRPV) infection of domestic rabbits, papillomaviruses can undergo abortive infections in which the productive cycle of the virus is not completed. As with other DNA tumour viruses, such abortive infections can predispose to cancer.Journal of Clinical Virology 04/2005; 32 Suppl 1:S7-15. · 3.97 Impact Factor
Prevalence of HPV infection by cervical cytologic status in Brazil
José Veríssimo Fernandesa,⁎, Rosely de Vasconcellos Meissnera, Maria Goretti Freire de Carvalhob,
Thales Allyrio Araújo de Medeiros Fernandesc, Paulo Roberto Medeiros de Azevedod, Luisa Lina Villae
aDepartment of Microbiology and Parasitology, Universidade Federal do Rio Grande do Norte, Natal - RN, Brazil
bDepartment of Pathology, Universidade Federal do Rio Grande do Norte, Natal - RN, Brazil
cDepartment of Biomedical Sciences, Universidade do Estado do Rio Grande do Norte, Mossoró - RN, Brazil
dDepartament of Statistics, Universidade Federal do Rio Grande do Norte, Natal - RN, Brazil
eLudwig Institute for Cancer Research, Hospital Alemão Oswaldo Cruz, São Paulo, SP, Brasil
a b s t r a c ta r t i c l ei n f o
Received 9 September 2008
Received in revised form 12 November 2008
Accepted 1 December 2008
Polymerase chain reaction
Objectives: To assess the prevalence of human papillomavirus (HPV) infection according to cervical cytologic
status in northeastern Brazil; identify other risk factors for low- and high-grade squamous intraepithelial
lesions (LSILs and HSILs); and identify the most prevalent HPV genotypes associated with the lesions.
Method: Two cervical smears were collected from 250 women referred for cancer screening, one for
cytologic examination and the other to test for the presence of HPV by PCR with genotyping by dot blot
hybridization. Result: There were 110 healthy cervices, 82 LSILs, and 58 HSILs. The overall HPV prevalence
was 48%, with higher rates for HSILs, and HPV-16 was the most prevalent type. Age, multiple sexual partners,
type of HPV present, smoking, and early onset of sexual activity were risk factors for cervical lesions.
Conclusion: Age, multiple sexual partners, and infection with HPV-16 increased the risk of having LSILs or
HSILs. Early onset of sexual activity and smoking only increased the risk of having HSILs.
© 2008 InternationalFederation ofGynecologyand Obstetrics.Publishedby Elsevier IrelandLtd. Allrights reserved.
Human papillomavirus (HPV) is a large group of epitheliotropic
viruses of more than 120 different types. Of the 40 HPV types infecting
the human body, more than a dozen are associated with probable or
definite oncogenic risk [1,2]. Infection of the cervix is initiated when
infectious particles reach and then bind to the basal layer of the
cervical epithelium, where they enter cells through small tears. It has
been suggested that for maintenance of the infection, the virus has to
infect an epithelial stem cell .
The infection occurs mainly inyoung, sexuallyactive women in the
first years following their first sexual encounters [4,5]. The highest
HPV prevalence is among women younger than 25 years. It decreases
from age group to age group and is lowest among women aged
between 45 and 50 years, but increases in the postmenopausal years
. Most HPV infections clear spontaneously, but cervical lesions can
develop in women who have persistent infection with high-risk types
of HPV . A current epidemiologic challenge is to identify etiologic
cofactors that lead to HPV persistence and progression to neoplastic
lesions, and clarify how these cofactors contribute to carcinogenesis
. The high-risk types of HPV have a higher tendency to cause
persistent infection and, in the presence of other environmental and
host factors, are associated with an increased risk of cervical lesions
that may progress to cervical cancer [8,9].
Various studies of screening testing for cervical cancer were
recently realized in the south and southeast regions of Brazil as part of
the Latin America Screening Study. These studies have analyzed the
relations between the cytologic and histologic abnormalities present
in cervical smears, and between HPV infection and the predictive
factors of these anomalies [10–12], but so far have not included the
Northeast region of Brazil. Moreover, the method used for HPV de-
tection in these studies was Hybrid Capture 2, which does not identify
The present study was conducted to analyze prevalence of HPV
infection in women with healthy and abnormal uterine cervices in the
Northeast region of Brazil, and to identify the HPV types prevalent in
the local population, as well as risk factors for HPV infection.
2. Materials and methods
The cervical samples used were obtained between April 2001 and
June 2002 at Luis Antonio Hospital in Natal, Rio Grande do Norte State,
Northeastern Brazil, from 258 women referred for cancer screening
because of a history of cytologic alterations. The women's age ranged
from 15 to 65 years. All women were informed about the methods and
objectives of the research. All signed an informed consent form and
International Journal of Gynecology and Obstetrics 105 (2009) 21–24
⁎ Corresponding author. Departamento de Microbiologia e Parasitologia, Centro de
Biociências, Universidade Federal do Rio Grande do Norte, Av. Sen. Salgado Filho, S/N,
Campus Universitário, Lagoa Nova, CEP: 59072-970, Natal, RN, Brazil. Fax: +55 84
E-mail address: firstname.lastname@example.org (J.V. Fernandes).
0020-7292/$ – see front matter © 2008 International Federation of Gynecology and Obstetrics. Published by Elsevier Ireland Ltd. All rights reserved.
Contents lists available at ScienceDirect
International Journal of Gynecology and Obstetrics
journal homepage: www.elsevier.com/locate/ijgo
answered a standardized questionnaire about their marital status,
use,smokinghabits,and family historyof cancer.The patients'ethnicity
was defined based on self-reports according to the criterion of the
Instituto Brasileiro de Geografia e Estatística (IBGE) that classifies
ethnicity into 5 categories: white, black, mulatto, yellow, and native. In
this study, the black, mulatto, yellow, and native categories were
combined into a nonwhite category. The study was approved by the
Ethical Committee in Research of Federal University of Rio Grande do
The cervical smears were obtained using a cytobrush for optimal
collection of exfoliated cells. A trained cytopathologist from the
Department of Pathology of Federal University of Rio Grande do
Norte stained the smears and analyzed them according to the
Bethesda classification (available at: http://bethesda2001.cancer.
gov/terminology.html). All abnormal smears were re-examined for
result confirmation. Samples with scant squamous cellularity were
considered unsatisfactory for evaluation and excluded from analysis.
The women with abnormal cytologic results were followed up
according to the guidelines of the National Cancer Program of Brazil.
Based on the results of the cytologic evaluation, the 250 retained
participants were allocated to 1 of 3 groups: a normal cytology group
(n=110); a low-grade squamous intraepithelial lesions (LSIL) group
(n=82); and a high-grade squamous intraepithelial lesions (HSIL)
Another sample of exfoliated cervical cells was collected from each
participant and the collection brush was placed in a tube containing
PBS, vancomycin, and nystatin. The tube was kept at 4 °C until
processing for DNA extraction. The content was centrifuged at 3000 g
for 10 minutes following vigorous tube agitation and brush removal;
the supernatant was removed; and the remaining pellet was
processed for DNA extraction, using rapid isolation of DNA from
mammals, with proteinase K .
To check the qualityof theobtained DNA, aliquotswithabout 30 ng
of DNA were submitted to polymerase chain reaction (PCR) using
the primers PCO3+/PCO4+, specific for the amplification of a 110 bp
fragmentof theβ-globingene .Onlythe 250DNA samplespositive
to β-globin were checked for HPV DNA.
Of the 250 good-quality DNA samples 110 were from participants
in the normal cytology group, 82 from participants in the LSIL group,
and 58 fromparticipantsin the HSIL group. The presence orabsence of
HPV DNA was established using the degenerate primers MY09/M11,
specific for the L1 open reading frame (ORF) of the HPV genome .
The products of PCR were submitted to electrophoresis on 7%
polyacrilamide gel, followed by silver staining . The specimens
were considered positive for HPV DNA if they presented a band of
450 bp. The amplicons were submitted to HPV genotyping by Dot blot
hybridization as described .
To check for associations between sociodemographic character-
istics and cervical lesions, and for differences in the prevalences of
HPV infection in the 3 groups, we used the Pearson χ2test for
comparison of proportions. To identify variables that could be
considered risk factors for cervical lesions we used all the considered
variables in a multivariate logistic regression analysis, with the
dependent variable assigned a value of 1 in the presence of low- or
high-grade intraepithelial cervical lesions and a value of 0 in the
absence of lesions. To check for associations between HPV type and
health status we used univariate logistic regression. All analyses were
performed using the software SPSS, version 13.0 (SPSS, Chicago,
Illinois, USA). P≤0.05 was considered statistically significant.
Statistical analysis by the χ2test showed an association between
age and presence of HSIL (P=0.000) and between ethnicity and
presence of HSIL (P=0.002). Considering separately married and
single women, we did not observe an association between marital
status and the presence of LSIL or HSIL (P=0.09 and P=0.62),
respectively (Table 1).
We observed that most (81.5%) of the women with both a normal
cytology result and HPV infectionwere younger than 34 years, with a
lesser prevalence of HPV infection in the older age groups. Among
women with LSILs, the prevalence of HPV infection increased as they
reached the age of 34 years, decreased as they aged from 35 to
44 years, and increased again after they were 45 years old. Most
(77.8%) of the women with HSILs were between 25 and 54 years old
(Table 1). Differences in HPV prevalence between the normal
cytology group (24.5% of the participants) and the LSIL (58.5%) and
HSIL groups (77.6%) were statistically significant by comparison of
the proportions, revealing that HPV infection increased the risk of
developing both low-grade (P=0.000) and high-grade (P=0.000)
premalignant lesions. Age was associated with the occurrence of
HSIL (P=0.000), as was nonwhite status (P=0.002). We observed
no associations between marital status and the presence of cervical
lesions (Table 1).
The women older than 34 years were at greater risk for LSILs and
those older than 45 years were at greater risk for HSILs. The women
who had their first sexual intercourse between the ages of 14 and
17 years were at greater risk only for HSILs. Having had multiple
sexual partners and being a smoker also increased the risk of having
HSILs. There was no evidence that age at first pregnancy, number of
pregnancies, use of oral contraceptives, and a family history of cancer
increased the risk of either LSIL or HSIL (Table 2).
Cervical HPV infection increased the risk of both LSIL and HSIL.
Fourteen different types of HPV were identified in the study
population, and most had a high oncogenic potential. Infection was
causedby1 typeof virus in 19.0%of thewomen in thenormalcytology
group, 40.2% of the women in the LSIL group, and 67.2% of the women
in the HSIL group, and HPV-16 was the most prevalent type in all 3
groups. The second most common type was HPV-58 in the first 2
groups and HPV-45in the HSILgroup. Infectionwith2 HPV typeswere
detected in the 3 groups, the most frequent coinfection being with
or HPV-45 and HPV-59 in the HSIL group. With the probes we used, it
was not possible to identify the HPV types carried by 2 participants in
the normal cytology group, 1 participant in the LSIL group, and 1
participantin theHSIL group.SingleinfectionbyHPV-16 increased the
Distribution of socio-demographic characteristics, according to health status
VariableNormal cytology group LSIL group HSIL group
with HPV (%)
with HPV (%)
with HPV (%)
Age at study entrya
4 4 (100.0)
aStatistical analysis for age among the groups: Group I×Group II: χ2=5.262,=0.261;
Group I×Group III: χ2=39.606 P=0.000.
bStatistical analysis for ethnicity among the groups: Group I×Group II: χ2=0.030,
P=0.862; Group I×Group III: χ2=9.166, P=0.002.
cStatistical analysis for marital status among the groups: Group I×Group II:
χ2=2.963, P=0.085; Group I×Group III: χ2=0.241, P=0.623.
J.V. Fernandes et al. / International Journal of Gynecology and Obstetrics 105 (2009) 21–24
risk of both LSIL and HSIL, whereas single infection by HPV-58 and
coinfection with HPV-56 and HPV-58 only increased the risk of LSIL
The prevalence of HPV infection in women with normal cytology
Recife, also in Northeast Brazil , it was similar to that estimated for
of meta-analyses [19,20]. This high prevalence of HPV infection in
of women whose sexual activity began before they were 18 years old,
when they were biologically the most vulnerable to this virus. It could
also be attributed to false-negative results, which commonly occur in
cytologic studies [2,21]. Besides, most infections were caused by high-
risk HPVtypes,mainlybyHPV-16,whichhasa greatercapacitythanthe
other types to maintain infection without cytologic alterations.
The overall HPV prevalence among the participants with normal
and less in the 35 to 44 years age group, a finding similar to those
reported for other World Regions . Among women with LSILs, the
Distribution of HPV types according to health status and associated odds ratio obtained by univariate logistic regression model
VariableNormal cytology group(n=110) LSIL group(n=82)HSIL group(n=56)
No. of patients No. of patients OR95% CI No. of patientsOR 95% CI
Abbreviations and note: See Table 2.
Distribution of patients by cytological status according to the considered variables, and associated odds ratio (OR) obtained by multivariate logistic regression model
VariableNormal cytology group(n=110) LSIL group(n=82)HSIL group(n=56)
No. of patientsNo. of patientsOR 95% CI No. of patientsOR95% CI
Age at study entry, y
Age at 1st sexual intercourse, y
Age at 1st pregnancy, y
No. of pregnancies
Oral contraceptive use, y
Multiple sexual partners
Family history of cancer
Abbreviations: CI, confidence interval; HSIL, High-grade lesion; LSIL, low-grade lesion; NA, not applicable; OR, odds ratio.
aDenotes statistical significance.
J.V. Fernandes et al. / International Journal of Gynecology and Obstetrics 105 (2009) 21–24
prevalence was higher in this study than that reported for Porto Alegre,
in Southern Brazil ; it was similar to that estimated for Africa ;
and it was less than that reported for Recife , Brasilia , or
estimated for Central and South America . It was highest among
womenyounger than 34 years, less in the 35 to 44 years age group, and
higher again among women 45 years or older. Most of the womenwith
between age group and HPV prevalence. The overall HPV prevalence in
Alegre , but less than that reported for Recife or Brasilia [18,24].
These differences can be explained not only by variations usually
observed between populations of different geographic regions, but also
sensitivity of the HPV DNA assays used in different studies [2,25].
An association between ethnicity and cervical lesion was observed
only for HSILs, suggesting a trend for lesions to progress faster in
nonwhitewomen. Mostof thewomen in the3 groups were marriedor
had only 1 sexual partner. No associations were observed between
HPV prevalence or development of cervical lesions and marital status.
Compared with the participants with normal cytology results, HPV
prevalence was significantly higher in participants with either lesion
grade. The overall prevalence of HPV-16 in this study was similar to
that estimated for Africa, but higher than that reported for Recife 
or estimated for South America [19,20]. We found an association
between HPV infection and the presence of LSILs among women
infected with HPV-16 and HPV-58 as single infections as well as
among those infected with both HPV-56 and HPV-58. We also found
an association between HPV-16 infection and presence of HSILs.
We found both LSILs and HSILs to be associated with age and
multiple sexual partners. On the other hand, we found only HSIL to be
associated with early age at first sexual intercourse or with smoking,
even when all variables were considered simultaneously in a logistic
history of cancer. These results are concordant with those reported in
studies conducted in São Paulo, Porto Alegre, and Buenos Aires [12,22].
Inthe3 studygroups,themostof thewomentestingpositivefor HPV
were infected with 1 or 2 high-risk types. Among women with normal
than that reported for Recife . These results show that HPV-16 is
high-risk HPV types alsoinfect the same population, thesewomen inour
region are at high risk for HPV infection and, consequently, for cervical
cancer. The distribution of the HPV types in this study was similar to that
described for women in Africa and in Central and South America.
The results of this studylead us toconcludethat high-grade lesions
are associated with age and ethnicity. Multiple lifetime sexual
first sexual intercourse and multiple sexual partners, smoking, and
infection with HPV-16 all increased the risk of having HSILs. The most
prevalent virus type was HPV-16 in the 3 groups, followed by HPV-58
in the normal cytologyand LSIL groups, and HPV-45 in the HSIL group.
Despite similarities in the order of prevalence of some HPV types
56, 57, and 59. By virtue of the existence of multiple HPV types and
variations in their distributions, evenwithin the same geographic region,
more local studies should be conducted to evaluate the cost-benefit and
expected efficacy of a HPV vaccination campaign, if found justified.
This study was supported by funds from the Conselho Nacional
de Desenvolvimento Científico e Tecnológico (National Council of
Scientific and Technological Development [CNPq]) and the Coorde-
nação de Aperfeiçoamento de Pessoal de Nível Superior (Coordination
of Improvement of Higher Education [CAPES]).
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