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Frequency and coinfection between genotypes of human papillomavirus in a population of asymptomatic women in northern Peru



Describe the prevalence of HPV genotypes via PCR and DNA sequencing in 397 women who attended to the gynecological outpatient center in the Hospital Regional Docente de Cajamarca from March to September 2017.


A positive PCR result for HPV was observed in 121 cervical samples. A high-risk genotype was found in 63.6% (77/121) of patients, a probably oncogenic type in 23.1% (28/121) and a low-risk type in 7.4%. Among the high-risk genotypes, HPV-31 was the most common one present in 20% (21/77), followed by HPV-16 in 11.4% (12/77). Coinfections between two or more genotypes were observed in 12 cases.


Cervical cancer is the second leading cause of female cancer mortality in Peru accounting for more than 1700 deaths annually [1]. Human papillomavirus (HPV) is the main responsible for this cancer and one of the most common sexually transmitted infection in Peru [2, 3]. HPV is a DNA virus from the papillomavirus family with over 170 types completely sequenced. Although the association between these genotypes and their risk of malignancy varies, its presence in 99.7% of cervical cancers has been recognized as core to the development of this neoplasia [4, 5].

HPV genotypes identified in women with cervical cancer are traditionally classified based on their associated risk of malignancy [5, 6]. Thus, 13 genotypes have been classified as “high risk” as they are detected in more than 90% of women with cervical cancer, with HPV 16 accounting for approximately 50% of cases, followed by HPV 18 in 20% [6,7,8,9]. Furthermore, in 2009 the International Agency for Research on Cancer (IARC) further classified other 12 genotypes as “probably oncogenic” and 2 as “low risk” based on their limited evidence in humans for cervical cancer [9].

After over 30 years of screening services and with cervical cancer declared a national priority in 2000, HPV infections remains a major concern in Peru especially affecting underserved areas where surveillance is limited [3, 10,11,12]. The main objective of this study was to describe the prevalence of the HPV oncogenic genotypes among women from Cajamarca, Peru as well as to provide preliminary reports of their potential risk factors.

Main text


Patients and study design

A consecutive cross-sectional study was conducted in the Hospital Regional Docente de Cajamarca, Peru. Women attending the gynecological outpatient center who had a history of at least 1 sexual encounter were studied from March to September 2017. Patients were excluded if there was evidence of pregnancy, severe gynecological bleeding, hysterectomy or previous history of HPV-related disease including cancer, warts and other cutaneous manifestations.

This study has been approved by two independent Ethics Committees: Hospital Regional Docente de Cajamarca and Universidad Peruana de Ciencias Aplicadas. Once the outpatient consultation ended, the attending physicians informed the patients about the study purpose and asked for written consent before enrollment. After a standardized questionnaire was completed by the physician a cervical sample was collected for further analysis. The questionnaire includes the following data: age, marital status, age of first sexual intercourse, number of sexual partners, sexual partners in the last 6 months, extramarital sexual relations, history of sexually transmitted infection (STI), abortions, Number of births, use of condoms, last pap, personal and family history of cervical cancer, use of sex toys, history of sexual abuse and date of the Last Papanicolaou test.

Sample collection and preservation

Cervical cell samples from the ectocervix and endocervix were collected from each woman using a cytobrush for preservation in a tube containing phosphate buffered saline (pH 8.6). The samples were then stored at − 4 °C and sent to the Molecular Biology Laboratory at Universidad Peruana de Ciencias Aplicadas. Once samples arrived at the laboratory, the cytobrushes were discarded and the tubes were vortexed and centrifuged to pellet the cells, which were resuspended in 1 mL of phosphate buffered saline. Three aliquots of each fresh specimen were stored at − 20 °C until testing.

HPV DNA extraction, amplification and genotype sequencing

Viral genomic DNA was extracted from a total volume of 200 μL of the sample by the guanidinium thiocyanate extraction method [13] and the purified material was re-suspended in a final volume of 30 μL deionized water. Samples were electrophoresed on a 1% agarose gel to check the quality of the DNA.

Human papillomavirus amplification was done using the primers and conditions described by Lurchachaiwong et al. [14]. PCR products were analyzed on 2% agarose gel stained with.

ethidium bromide and bands were detected by UV transillumination (Kodack Logic 1500, USA). Positive specimens were confirmed by direct sequencing serving as the gold standard (Macrogen-Korea).

The HPV genotypes were categorized into three groups: high, probably oncogenic and low risk based on the IARC classification [9] (Additional file 1: Table S1).

Statistical analysis

Quantitative variables were described as frequencies and percentages for each group using the GraphPad Prism3 statistical (Graph Pad Sofware Inc., San Diego, USA).


A total of 397 women were studied from March to September 2017. Most patients were between 36 and 45 years old (35%) followed closely by the age group 26–35 years old (30.2%); 51 patients were under 25 years old (12.8%) with only two patients under 18 years old (Table 1).

Table 1 Human papillomavirus infection in women from Cajamarca, Peru

Human papillomavirus DNA was amplified in 121 of our patient’s samples and all of them were successfully sequenced for genotype identification. A high-risk genotype was found in 63.6% (77/121) of patients, a probably oncogenic type in 23.1% (28/121) and a low-risk type in 7.4%; other genotypes were detected in 19.83%. All sequenced genotypes are shown in Table 2.

Table 2 Prevalence of HPV genotypes detected in women from Cajamarca, Peru

Coinfections between two or more genotypes were observed in 12 cases. The most common coinfections were between HPV types 39–45–68, 40–43–91 and 31–91 each of them in two cases (Additional file 1: Table S2).

Demographic and other potential risk factors for HPV infections were also registered from each patient. Most women were married/cohabiting, had one sexual partner, maintaining sexual relationships in the last 6 months. However, only 35.5% of them uses condoms and the use of sex toys was uncommon. Extramarital affairs were observed in 3.3% of patients and there were 31 cases with a history of sexual abuse. Most patients were multiparous with three or more births (36.8%) (Table 3).

Table 3 Demographics and characteristics among women with HPV

Among women who were infected by a high-risk HPV genotype, similar characteristics were observed with certain exceptions. In this group, it was slightly more common to observe 2-lifetime sexual partners and the majority were sexually active (82.1%). Although uncommon, 53.8% of extramarital affairs were observed in this group. Additionally, 19 of these women have never had a Pap smear test for cervical cancer screening (Table 3).


Cervical cancer is the most common female cancer in Peruvian women between 15 and 44 years-old, with about 4700 new cases diagnosed annually and an estimated mortality of 24.6 per 100,000 women [3, 10]. There is clear evidence that HPV is the main responsible for cervical cancer and prevalence studies of the oncogenic genotypes are encourage especially in low-income communities were surveillance reports are still limited [3, 10,11,12, 15].

In our study population of 397 women, a total of 121 samples were positive for HPV via PCR, with a high-risk genotype present in 77 samples and a probably oncogenic genotype in 28. In the high-risk group, the most common genotypes were HPV-31 (20%) followed by HPV-16 (11.4%). These results differ from a previous investigation our research team conducted in Cajamarca between 2010 and 2012, in which the HPV-16 was the most common genotype in 38.5%, followed by the HPV-39 in 9.6% [15].

Another investigation in 465 women, without the diagnosis of cervical cancer, from Lima, Peru reported the genotypes HPV-16 (23.8%) and HPV-6 (11.9%) [16] as the most common ones. Furthermore, a study conducted in an Amazonian region of Peru showed a higher frequency of high-risk HPV among females from an urban population in Iquitos compared to a native Amazonian population. However, the genotypes distribution in both population were different. In the urban population, HPV-16 was the most common type in 58.5% followed by HPV-18 and HPV-31. On the contrary, in the Amazonian native community, HPV-16 was uncommon, as other unique HPV types were more frequently observed in these patients such as HPV-39, HPV-71, and HPV-96 [17]. Thus, HPV genotypes distribution can widely vary between two different close regions from the same country and changes in genotype prevalence are expected, with studies reporting an HPV-16 prevalence significant decrease in the last years after the introduction of the vaccine [18,19,20].

Multiple genotypes can be detected in patients with cervical cancer suggesting that coinfections might be a potential risk factor for carcinogenesis. A recently published study from 2017, reported a 4.1-fold higher risk of developing invasive cervical carcinoma in subjects infected with any HPV genotype, excluding HPV-16, in association with HPV-18 [21]. Thus, the risk of cervical cancer is genotype specific, but it might be increased with certain genotype interactions. In our study, only one case of HPV-16 coinfection was observed, but coinfections with a high-risk type occurred in 5 samples, including a triple coinfection between HPV-39, HPV-45, and HPV-68.

Genital HPV infections are spread by unprotected penetrative intercourse or close skin-to-skin physician contact [22, 23]. In our population, most patients did not use condoms (64.5%), and this increased risk practice was observed in all our HPV-positive patients (56.2%), high-risk HPV positive (54.5%) and probably oncogenic HPV positive women (57.1%). Fomite contact or vaginally inserted sex toys can potentially spread HPV as the virus can be detected up to 24 h after standard cleaning, but the evidence is not definitive. The use of sexual toys was very uncommon in our patients (2%), with a positive HPV sample in 4 of the 8 cases.

The risk of HPV infection in women is directly related to the number of male sex partners. Furthermore, as with other sexually transmitted diseases, sex with a new partner is a stronger risk factor than sex with long-term partners [24,25,26,27,28]. In our study, females with 3 or more partners represented 25.6% of HPV-positive patients. Additionally, 4 cases of HPV were observed in females who had 2 or more partners in the last 6 months; although most of our patients with HPV (80.2%) were sexually active with 1 partner in the last 6 months. Similar results were observed in our previous investigation in which the groups of women with a history of 2 sexual partners and the group with 3 or more partners represented 25% and 17.3% of the total HPV positive cases [15].

In 2011, Almonte et al. conducted a research in women attending cervical cancer screening in the Peruvian Amazon. In this study, early age at first sexual intercourse and more than 5 sexual partners were risk factors for having HPV infection. More interestingly, high parity, no schooling and the lack of a good-quality screening with an adequate follow-up were the main risk factors for high-grade cervical disease [29]. Even though, assessing a risk for cervical disease was not part of our objectives, we observed that 36.8% of our population had 3 or more births and a similar tendency was reported among HPV positive women.

Human papillomavirus DNA testing is increasingly being used as it improves the sensitivity for detection of cervical cancer precursors when used in combination with cervical cytology. However, using molecular testing can also decrease specificity resulting in potential unnecessary referrals for colposcopy [30]. Additionally, HPV PCR is not recommended in women under 30 years old since the HPV may clear up spontaneously in younger women [31, 32]. Thus, the Papanicolaou test is recommended as the first screening method with a co-testing HPV PCR only in women ≥ 30 years old. In our population, 24.7% (19/77) of our HPV high-risk group never had a pap smear; however, most of these women (73.7%) were under 30 years old.


Despite national efforts for cervical cancer screening and prevention, HPV infections resulting in invasive cervical cancer remains a major health issue in Peru. Additionally, in underserved areas such as Cajamarca HPV genotype surveillance is limited. Contrary to previous reports in which HPV-16 was predominant, an increase of HPV-31 have been observed which is now the most common high-risk genotype in our study population.


The present study had one important limitations. We designed the study for the detection and genotypification of HPV, thus our results are only for prevalence reporting and the implications of these genotypes or the potential risk for cancer in our patients is unknown. Second, due financial limitations and our study design it was impossible to compare the isolated genotypes with pap smear results as they were not available in most of our patients.



polymerase chain reaction


deoxibonucleic acid


base pairs


human papillomavirus


phosphate buffered saline


  1. Aguilar A, Pinto J, Araujo J, Fajardo W, Bravo L, Pinillos L, et al. Control of cervical cancer in Peru: current barriers and challenges for the future. Mol Clin Oncol. 2016;5(2):241–5.

    Article  PubMed  PubMed Central  Google Scholar 

  2. Santos C, Munoz N, Klug S, Almonte M, Guerrero I, Alvarez M, et al. HPV types and cofactors causing cervical cancer in Peru. Br J Cancer. 2001;85(7):966–71.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  3. Bruni L, Barrionuevo-Rosas L, Albero G, Serrano B, Mena M, Gómez D, et al. ICO/IARC information centre on HPV and cancer (HPV information centre). Human papillomavirus and related diseases in Peru. Summary Report 27 July 2017. Accessed 27 July 2017.

  4. Walboomers J, Jacobs M, Manos M, Bosch F, Kummer J, Shah K, et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J Pathol. 1999;189(1):12.

    Article  PubMed  CAS  Google Scholar 

  5. Senapati R, Nayak B, Kar S, Dwibedi B. HPV genotypes co-infections associated with cervical carcinoma: special focus on phylogenetically related and non-vaccine targeted genotypes. PLoS ONE. 2017;12(11):e0187844.

    Article  PubMed  PubMed Central  Google Scholar 

  6. Jentschke M, Soergel P, Hillemanns P. Importance of HPV genotyping for the screening, therapy and management of cervical neoplasias. Geburtshilfe Frauenheilkd. 2012;72(6):507–12.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  7. Bernard E, Pons-Salort M, Fayre M, Heard I, Delaqocque-Asagneau E, Didier G, et al. Comparing human papillomavirus prevalences in women with normal cytology or invasive cervical cancer to rank genotypes according to their oncogenic potential: a meta-analysis of observational studies. BMC Infect Dis. 2013;13:373.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Muñoz N, Bosch F, de SanJose S, Herrero R, Castellsague X, Shah K, et al. Epidemiologic classification of human papillomavirus types associated with cervical cancer. N Engl J Med. 2003;348(6):518–27.

    Article  PubMed  Google Scholar 

  9. Bouvard V, Baan R, Straif K, Grosse Y, Secretan B, El Ghissassi F, et al. A review of human carcinogens–Part B: biological agents. Lancet Oncol. 2009;10(4):321–2.

    Article  PubMed  Google Scholar 

  10. Pan American Health Organization (PAHO). Cervical cancer prevention in Peru: lessons learned from the TATI demonstration project. Washington, D.C. [Cited on May 26, 2018]. Accessed 22 May 2018.

  11. Almonte M, Ferreccio C, Winkler J, Cuzick J, Tsu V, Robles S, et al. Cervical screening by visual inspection, HPV testing, liquid-based and conventional cytology in Amazonian Peru. Int J Cancer. 2007;121(4):796–802.

    Article  PubMed  CAS  Google Scholar 

  12. Levinson K, Abuelo C, Salmeron J, Chyung E, Zou J, Belison S, et al. The Peru Cervical Cancer Prevention Study (PERCAPS): the technology to make screening accessible. Gynecol Oncol. 2013;129(2):318–23.

    Article  PubMed  PubMed Central  Google Scholar 

  13. Camps M, Vilella A, Marcos MA, Letang E, Muñoz J, Salvadó E, et al. Incidence of respiratory viruses among travelers with a febrile syndrome returning from tropical and subtropical areas. J Med Virol. 2008;80(4):711–5.

    Article  PubMed  CAS  Google Scholar 

  14. Lurchachaiwong W, Junyangdikul P, Payungporn S, Sampatanukul P, Chansaenroj J, Tresukosol D, et al. Human papillomavirus genotypes among infected Thai women with different cytological findings by analysis of E1 genes. New Microbiol. 2011;34(2):147–56.

    PubMed  CAS  Google Scholar 

  15. Silva-Caso W, Olivera-Irazabal M, Leon-Alvarez P, Del Valle LJ, Diaz-Estacio S, Vargas M, et al. Identification of human papillomavirus as a preventive strategy for cervical cancer in asymptomatic women in the Peruvian Andes. Asian Pac J Trop Med. 2014;7S1:S121–6.

    Article  PubMed  Google Scholar 

  16. Sullcahuaman-Allende Y, del Castro-Mujica MC, Mejia C, Castaneda C, Castillo M, Dolores-Cerna K, et al. Demographic characteristics of human papillomavirus detected by PCR-RFLP in peruvian women. Send to Rev Peru Med Exp Salud Publica. 2015;32(3):509–14 (Article in Spanish).

    Article  Google Scholar 

  17. Martorell M, Garcia-Garcia J, Gomez-Cabrero D, Del Aguila A. Comparison of the prevalence and distribution of human papillomavirus infection and cervical lesions between urban and native habitants of an Amazonian region of Peru. Genet Mol Res. 2012;11(3):2099–106.

    Article  PubMed  CAS  Google Scholar 

  18. Dong L, Hu S, Zhang Q, Feng R, Zhang L, Zhao X, et al. Changes in genotype prevalence of human papillomavirus over 10-year follow-up of a cervical cancer screening cohort. Zhonghua Liu Xing Bing Xue Za Zhi. 2017;38(1):20–5.

    PubMed  CAS  Google Scholar 

  19. Mesher D, Soldan K, Howell-Jones R, Panwar K, Manyenga P, Jit M, et al. Reduction in HPV 16/18 prevalence in sexually active young women following the introduction of HPV immunisation in England. Vaccine. 2013;32(1):26–32.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  20. Mesher D, Panwar K, Thomas S, Beddows S, Soldan K. Continuing reductions in HPV 16/18 in a population with high coverage of bivalent HPV vaccination in England: an ongoing cross-sectional study. BMJ Open. 2016;6(2):e009915.

    Article  PubMed  PubMed Central  Google Scholar 

  21. Senapati R, Nayak B, Kumar-Kar S, Dwibedi B. HPV genotypes co-infections associated with cervical carcinoma: special focus on phylogenetically related and non-vaccine targeted genotypes. PLoS ONE. 2017;12(11):e0187844.

    Article  PubMed  PubMed Central  Google Scholar 

  22. Burchell A, Winer R, de SanJose S, Franco E. Chapter 6: Epidemiology and transmission dynamics of genital HPV infection. Vaccine. 2006; 24 Suppl 3:S3/52–61.

  23. Palefsky J. Cutaneous and genital HPV-associated lesions in HIV-infected patients. Clin Dermatol. 1997;15(3):439.

    Article  PubMed  CAS  Google Scholar 

  24. Anderson T, Schick V, Herbenick D, Dodge B, Fortenberry J. A study of human papillomavirus on vaginally inserted sex toys, before and after cleaning, among women who have sex with women and men. Sex Transm Infect. 2014;90(7):529–31.

    Article  PubMed  CAS  Google Scholar 

  25. Tarkowski T, Koumans E, Sawyer M, Pierce A, Black C, Papp J, et al. Epidemiology of human papillomavirus infection and abnormal cytologic test results in an urban adolescent population. J Infect Dis. 2004;189(1):46.

    Article  PubMed  Google Scholar 

  26. Peyton C, Gravitt P, Hunt W, Hundley R, Zhao M, Apple R, et al. Determinants of genital human papillomavirus detection in a US population. J Infect Dis. 2001;183(11):1554.

    Article  PubMed  CAS  Google Scholar 

  27. Karlsson R, Jonsson M, Edlund K, Evander M, Gustavsson A, Bodén E, et al. Lifetime number of partners as the only independent risk factor for human papillomavirus infection: a population-based study. Sex Transm Dis. 1995;22(2):119.

    Article  PubMed  CAS  Google Scholar 

  28. Moscicki A, Hills N, Shiboski S, Powell K, Jay N, Hanson E, et al. Risks for incident human papillomavirus infection and low-grade squamous intraepithelial lesion development in young females. JAMA. 2001;285(23):2995.

    Article  PubMed  CAS  Google Scholar 

  29. Almonte M, Ferreccio C, Gonzales M, Delgado J, Buckley C, Luciani S, et al. Risk factors for high-risk human papillomavirus infection and cofactors for high-grade cervical disease in Peru. Int J Gynecol Cancer. 2011;21(9):1654–63.

    Article  PubMed  Google Scholar 

  30. Schiffman M, Wentzensen N, Wacholder S, Kinney W, Cage J, Castle P. Human papillomavirus testing in the prevention of cervical cancer. J Natl Cancer Inst. 2011;103(5):368–83.

    Article  PubMed  PubMed Central  Google Scholar 

  31. Meijer C, Berkhof J, Castle P, Hesselink A, Franco E, Ronco G, et al. Guidelines for human papillomavirus DNA test requirements for primary cervical cancer screening in women of 30 years and older. Int J Cancer. 2009;124(3):516–20.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  32. Leinonen M, Nieminen P, Kotaniemi-Talonen L, Malila N, Tarkkanen J, Laurila P, et al. Age-specific evaluation of primary human papillomavirus screening vs conventional cytology in a randomized setting. Natl Cancer Inst. 2009;101(23):1612.

    Article  CAS  Google Scholar 

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Authors’ contributions

JdVM, LPB, PRP and LBG designed the study protocol. JdVM and MAAL: responsible for obtaining funding and laboratory work supervision. CPR, MAAL, LPB and PRP performed the PCR for HPV and sequencing. LBG, LPV, WSC, PW, JBM and JAA was responsible for the clinical assessment, samples collection and database completion. MAAL and LEC was responsables for data analysis. LPB, PRP, JdVM and PW drafted the manuscript. All authors critically revised the manuscript for intellectual content. All authors read and approved the final manuscript.


Gynecology health personnel from Hospital Regional Docente de Cajamarca, Peru.

Competing interests

The authors declare that they have no competing interests.

Availability of data and materials

Abstraction format used in the study and dataset are available and accessible from the corresponding author upon request. Data available in the link:

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Not applicable.

Ethics approval and consent to participate

This study has been approved by two independent Ethics Committees from Hospital Regional Docente de Cajamarca and Universidad Peruana de Ciencias Aplicadas. All samples were analyzed after a written informed consent was signed.


This work was supported by 4th research incentive of the Universidad Peruana de Ciencias Aplicadas (Grant: UPC-EXP-02-2017). Lima, Peru.

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Correspondence to Juana del Valle-Mendoza.

Additional file

Additional file 1: Table S1.

Human papillomavirus types and oncogenic potential. Table S2. Demographics and characteristics among women with HPV.

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Ponce-Benavente, L., Rejas-Pinelo, P., Aguilar-luis, M.A. et al. Frequency and coinfection between genotypes of human papillomavirus in a population of asymptomatic women in northern Peru. BMC Res Notes 11, 530 (2018).

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