Prevalence and Genotyping of Human Papillomavirus among Women in the Departments of Niari and Bouenza, Republic of the Congo

Human papillomaviruses (HPV) are at the origin of many mucosal, benign or malignant skin pathologies. Some so-called high-risk genotypes, especially types 16 and 18, are involved in the genesis of cervical cancer, one of the most common cancers in women. To determine the prevalence of HPV infections and to identify oncogenic genotypes circulating in the departments of Niari and Bouenza. A total of 207 cervical samples collected between 2019 and 2020 were studied. Cervico vaginal smears were performed on all samples for cytological analysis, and HPV DNA detection and genotyping were performed on GeneXpert. The average age of the women who participated in this study was 43.67 ± 12.31 years in Niari served between socio-demographic characteristics and genotyping (p > 0.05). On the other hand, HPV-HR prevalence varied significantly according to cervical cytology (p = 0.001). The results obtained show the predominance of HPV 16 in women in these two departments of Congo.


Introduction
Cervical cancer is the most common cancer in women in developing countries. It is estimated to be responsible for nearly 260,000 deaths per year, about 80% of which are in developing countries [1]. In sub-Saharan Africa, it accounts for 22.2% of all cancer cases in women [2], and is the leading cause of death from female cancer in Africa behind breast cancer [3]. However, persistent infection with certain oncogenic papillomavirus (HPV) types is firmly established as the necessary cause of most precancerous and cancerous epithelial lesions of the cervix and a variable fraction of neoplastic lesions of the vulva, vagina, anus, penis and oropharynx [4].
Indeed, these HPV infections are among the most prevalent in the world [5], with a global incidence of more than 30 million new cases per year. Thus, HPV infection is one of the most common sexually transmitted viral infections [6].
In Congo, about 214 deaths from cervical cancer occur per year and are diagnosed each year (estimates for 2020) [7]. Indeed, cervical cancer ranks as the 1 st cause of death among women who die from cancer in Congo and is the 3 rd leading cause of cancer death in women aged 15 to 44 years [7], and age-adjusted incidence and mortality rates are estimated at 25.2 and 13.0 per 100,000 women, respectively [8].
In addition, the local context shows a lack of scientific information on the molecular epidemiology of HPV infections and its complications in the departments of Niari and Bouenza. This information is crucial in order to provide virological data in the category of women with normal cytology after a cervico-vaginal smear (FCV) examination, as well as in those of women with precancerous and cancerous lesions of the cervix [5], in order to be able to develop effective future strategies for the prevention and control of cervical cancer in Congo.
Thus, this study aimed to determine the prevalence of HPV infections in the departments of Niari and Bouenza, and to identify the oncogenic genotypes present. Journal of Biosciences and Medicines riod from April 2019 to December 2020. The women included in this study were recruited from Dolisie and Nkayi General Hospitals. A total of 207 women had a Pap smear.
Cytological tests were motivated by certain gynecological problems such as bleeding, vaginal discharge, pelvic pain, pruritus or infertility exploration. Eligible patients included all women who came to the laboratory at these general hospitals for a pap smear, regardless of their medical history during the designated recruitment period. Most of them have never had a medical history regarding cervical cancer or its precancerous lesions. We excluded women who were physically or mentally unable to undergo an interview, women who were pregnant or undergoing known treatment for cervical lesions, or who had refused to participate in the study. The study protocol had been approved by the Health Sciences Research Ethics Board (HSRB): 258/MRSIT/IRSSA/CERSSA.

Collection of Cervical Cells
During a cervical speculum examination, samples were taken using a cytobrush of the cervix. After scraping, the cells were suspended in special collection vials containing 10 mL of BD SurePath solution TM and stored at −20˚C. These samples were used for cytological and molecular analyses.

Cytological Study
After sampling in the different departments, the samples were sent to the pathology laboratory of the Loandjili General Hospital (HGL) in Pointe-Noire for the cytopathological study. The examination was performed by a pathologist.

HPV Detection and Genotyping
All HPV tests were performed on cervical cell samples stored in a SurePath liquid medium at the Loandjili Hospital laboratory in Pointe Noire. Identification of HPV types was performed by real-time PCR using geneXpert technology (CEPHEID), USA from the Xpert ® HPV kit.
The Xpert HPV Assay test is an automated test for the qualitative detection

Statistical Analyses
Statistical significance between HPV genotypes, cytological status and sociodemographic characteristics was assessed by the exact Fisher test or the Khie-2 test using Epi-Info version 7 software (https://www.cdc.gov/epiinfo/fra/fr_pc.html).
Meaning was defined as p < 0.05 for all tests.

Socio-Demographic Characteristics
A total of 207 cervical samples were analyzed, including 128 from the department of Niari and 79 from the department of Bouenza. The average age of women was 43.67 ± 12.31 years and 38.29 ± 10.52 years respectively in the departments of Niari and Bouenza with extremes ranging from 18 to 80 years. Socio-demographic characteristics are shown in Table 1. The main local risk factors related to HPV infection studied in women in these departments were: age, age of first sexual intercourse, number of sexual partners and gestures, taking oral contraceptives, smoking. No significant association was observed between socio-demographic characteristics and genotyping in our study population (p > 0.05).

Cytological Profile
The cytological profile by department ( Table 2)

Genotyping of HPV-HR
A total of 79 of the 207 women in our study tested positive for one or more HPV-HR types. The genotypic distribution by department is shown in Table 3.
In the department of Niari 48 (37.5%) women tested positive for HPV, and in the department of Bouenza 31 (39.24%) women tested positive for HPV. Multiple infections ranged from two to more than three strains of HPV.

HPV Infection and Socio-Demographic Characteristics
In the department of Niari 43.9% of women with high-risk HPV infection were between 30 and 40 years old. Of the 87.5% of women who had their first sexual intercourse before the age of 18, 37.5% had an HPV-HR infection. Women who had more than 5 sexual partners had 43.75% HPV-HR carrying. HPV-HR infection was observed in 37.07% of women who had more than 5 pregnancies compared to 42.85% in those who had less than 5 pregnancies.    No statistically significant differences were observed between socio-demographic characteristics and HPV-HR genotyping (p > 0.05). All results are reported in Table 4.
In the department of Bouenza, it was observed a portage of HPV-HR infection in 39.47% of cases in 30 -40 year olds. 38.77% of HPV-HR infection was observed in women who had their first sexual intercourse before the age of 18.
Women who had more than 5 sexual partners reported an infection 39.47% while those who had more than 5 pregnancies were infected only to the rate of 36.58%.
No statistically significant association was observed between socio-demographic characteristics and HPV genotyping (p > 0.05). All the results are reported in Table 5.  Table 6 shows the prevalence of HPV-HR infection according to the cytological profile in the department of Niari.

Discussion
Epidemiological data on HPV strain genotyping and cervical cancer remain  [12] respectively. On the other hand, our average age for the department of Bouenza is also in agreement with that of Reesink-Peters et al. (2004) who obtained an average age population of 37 years in the Netherlands [13]. These similarities in the average age of the study population could be explained by the fact that the dynamics of the Congolese population have remained the same for the last 5 years.
In the departments of Niari and Bouenza, 87.5% and 62.02% of women had their first sexual intercourse before the age of 18, respectively. These results are  years in India [16]. The precocity of the age of the first sexual intercourse seems to be explained by the culture and the sociology of the Congolese population which is very modern, young and very Westernized in mentality.
In the present study, 25% of women in the department of Niari and 48.1% of that of Bouenza had more than 5 sexual partners. These results can be superimposed on that of Carol Chelimo [17]. Regardless of the department, more than 51% of the women had at least 5 pregnancies. This result is close to that of Olivier et al. (2005) who obtained 59.6% in an epidemiological and cytopathological profile study of dysplastic lesions of the cervix in Congo-Kinshasa [18].
The modernization of manners and the precocity of the age of the first sexual intercourse are as many factors explaining the multiplicity of sexual partners.
However, the level of poverty in the population also plays a major role in this sense.  [21]. While cervical cancer is the 2nd cancer in Congolese women, only 12.35% of our study population had a cytological abnormality. This relatively low rate could be explained by the young nature of our population which is able thanks to a strong local immunity to spontaneously eliminate the virus [22].

P. C. Nganga et al. Journal of Biosciences and Medicines
The prevalence of HPV-HR infection was 37.5% and 39.24% in the departments of Niari and Bouenza respectively. These results are similar to those of BOUMBA et al. (2015) and LOUBANOU et al. (2020) which reported prevalences of 41.1% and 41.5% respectively [23] [24]. On the other hand, they do not agree with Gordana Kovacevic's study in Serbia which was 51.8% [25], and are also much higher than those described in the majority of European studies which are between 11.3% and 18.3% [26]. Our HPV prevalence observed in the two departments, although low compared to certain African countries, show higher proportions than the African average. The types of HPV-HR observed in this study allow us to confirm the importance of screening in these populations studied.
The prevalence of HPV infection in women without cervical lesions was 85.41% and 80.64% respectively in Niari and Bouenza. These HPV-HR frequencies in women without cervical lesions were well above the global average (10.4%), higher than the average in the sub-Saharan Africa region (22.1%) and finally higher than the Boumba et al. study (2015) (23.5%) [23] [27].
Indeed, the exposure of our study population to many risk factors for HPV infection and risky sexual behaviors could explain this high rate of HPV infection observed in these two departments. These results also show the lack of knowledge of HPV infection in our population, which is rarely detected. The lack of a national screening program and awareness campaigns could also explain this lack of knowledge and high rates of HPV-HR in this category of women. On the other hand, 14.54% and 19.34% of women with HPV-HR infection had at least one cytological abnormality in the departments of Niari and Bouenza respectively. These results in agreement with the majority of studies in the literature prove the lack of information and awareness among populations about this pathology and suggest that women who consult are potentially at risk because they do so for the most part only as a result of a gynecological problem.
In both departments, HPV 16 was the most prevalent genotype, 45.82% in Niari and 48.38% in Bouenza. These results are in agreement with the study by LOUBANOU et al. (2020) which placed HPV 16 in first place, and other types of HPV in second place [24]. They also agree with the study by BOUMBA et al.
(2015) on the distribution of HPV whose genotypes HPV 16 (20.6%) and HPV 33 (14.3%) were the most prevalent [23]. Although these results allowed us to obtain the prevalence of HPV 16 isolation genotypes, the GeneXpert technique used in this study did not allow us to determine the prevalence of other HPV types, due to the fact that this typing is pooled.
High-risk oncogenic HPV genotypes have been observed in low-grade, highgrade and invasive cervical cancers. These results are consistent with the study by N. El Gnaoui et al. [28].
These high prevalences confirm how aggressive HPV 16 is compared to other types and reinforce the idea of establishing better epidemiological monitoring of the populations of these two departments in order to prevent the persistence and progression of these HPV-HR to cervical cancer [23] [24]. Journal of Biosciences and Medicines It should be noted that this study, despite its limitations related to sample size, presents a pragmatic situational analysis of the molecular epidemiology of this virus in the two departments concerned.

Conclusion
The present study made it possible to define the cytological profile of precancerous and cancerous lesions of the cervix in the departments of Niari and Bouenza. Our results showed that despite the absence of cytological lesions, more than 50% of HPV-HR were identified in women with normal cytology with HPV type 16 as the most prevalent. This work is a reminder of the importance of the urgent establishment of a national program to fight against this cancer and HPV infections in Congo. However, future studies are needed to map HPV-HR throughout the national territory.