African Journal of Reproductive Health Women's Health and Action Research Centre ISSN: 1118-4841 Vol. 13, Num. 1, 2009, pp. 37-45

African Journal of Reproductive Health, Vol. 13, No. 1, March, 2009, pp. 37-45

Original Research Article

Atiene S. Sagay¹ , Godwin E. Imade¹, Viola Onwuliri² , Daniel Z. Egah³, Matthew J. Grigg⁴, Jonah Musa¹, Tom D. Thacher⁵, James O. Adisa⁶, Malcolm Potts⁷ and Roger V. Short⁶

¹Departments of Obstetrics and Gynaecology, ²Biochemistry, ³Medical Microbiology, Faculty of Medical Sciences, University of Jos, Jos, Plateau State, Nigeria;
⁴ Faculty of Medical Sciences, Dentistry and Health Sciences, University of Melbourne, Melbourne, Australia;
⁶Dept of Pathology, ⁵Family Medicine, Jos University Teaching Hospital, Jos, Nigeria;
⁷ School of Public Health, University of California, Berkeley, USA.
*For correspondence: Atiene S. Sagay FWACS, FRCOG, Dept. of Obstetrics and Gynaecology, Faculty of Medical Sciences, University of Jos, Nigeria E-mail: atsagay58@yahoo.com Tel: 2348034519740

Code Number: rh09005

ABSTRACT

Vaginal douche products have been associated with cervical cancer. We examined female sex workers (FSWs) in Nigeria who douche with lemon or lime juice and compared the findings with that of nonusers. We obtained Pap smears and performed colposcopy of the vulva, vagina and cervix. A total of 374 FSWs comprising 81 Lemon users (LUs) and 293 non lemon users (NLUs) were examined. Their mean age was 27.8 ± 6.7 (range 16-63) years. At colposcopy, 17(4.5%) had genital warts [LUs 5(6.2%); NLUs 12(4.1%); p=0.43], 61(16.3%) had suspected squamous intraepithelial lesions (SILs) [LUs 17(21.0%); NLUs 44(15.0%); p=0.20] and 65(17.4%) had other findings. Pap smear cytology showed that 87(24.6%) had SILs [LUs 26(33.3%); NLUs 61(22.1%); p=0.03]. Lemon/lime use was associated with cervical dysplasia after controlling for HIV status (Adjusted OR=1.8; 95% CI, 1.0-3.0). Our data suggests an association between the practice of douching with citrus juice and cervical dysplasia (Afr J Reprod Health 2009; 13[1]:37-45).

RĖSUMĖ

Les produits de la douche vaginale ont été liés au cancer du col. Nous avons étudié les prostituées au Nigéria qui se douchent en se servant du jus de limon et nous les avons comparées à celles qui ne s’en servent pas. Nous avons collecté le papanicolaou et nous avons fait la colposcopie de la vulve, du vagin et du col. Au total, nous avons étudié 374 prostituées qui comprennent 81 qui se servent du citron et 293 qui ne se servent pas du citron. Leur âge moyen s’est élevé à 27,8 ± 6, 7 (entre 16 et 63 ans). La colposcopie a montré que 17(4,5%) avaient des verrues génitales [celles qui se servent du citron = 5(6,2%) ; celles qui ne s’en servent pas = 12(4,1%) ; p=0,43], 61(16,3% avaient des lésions intraepitheliales squameuses suspectes (LIS) [celles qui se servent du citron = 17(21,0%) ; celles qui ne s’en servent pas = 44(15,0%) ; p = 0,20] et 65 (17,4%) avaient d’autres résultats. La cytologie du papanicolaou a montré que 87(24,6%) avaient LIS [Celles qui servent du citron = 26(33,3%) ; celles qui ne s’en servent pas = 61(22,19%) ; p = 0,03]. L’utilisation du citron/du jus de limon a été liée à la dysplasie cervicale après avoir contrôlé pour l’état du VIH (OR yiste = 1,8 ; 95% CI, 1,0 – 3,0). Nos données indiquent un lien entre la pratique de se doucher avec le jus de limon et la dysplasie cervicale (Afr J Reprod Health 2009; 13[1]:37-45).

KEYWORDS: CIN, Cytology, Douching, Lime, Nigeria

Introduction

Vaginal douching before or after sexual intercourse is a widespread practice among women of diverse cultures worldwide, and commercial antiseptics, soap with water and salty water are commonly used agents.^1-3 Substances like Nonoxynol-9 (N-9), Krest bitter lemon (soft drink) and lime juice have been demonstrated to have antimicrobial and spermicidal properties and have been considered as candidates for the prevention of pregnancy and sexually transmitted infections.^4-6 Vaginal douching with lemon or lime juice is a common practice among female sex workers (FSWs) in northern Nigeria.⁷ Recent pre-clinical research compared the cytotoxicity of lemon and lime juice to that of the spermicide (N-9). When tested on cervical explant tissue, lemon and lime juices caused damage to cells comparable to that of N-9.⁸ In sexually abstinent volunteers, douching with lime juice caused dose-dependent epithelial damage to the cervix and vagina.⁹ Primate studies have, however, shown that daily intravaginal administration of undiluted lime juice (pH 2.5) for one month does not appear to damage the cervical or vaginal epithelium.¹⁰ Frequent vaginal douching can modify vaginal flora and increase the risk of cervical infections.¹¹ Our earlier publication showed no association between the prevalence of HIV and other STIs and sex-related vaginal douching with citrus juice.¹² Lysol and other tarbased vaginal douche products were voluntarily removed from the US market over 30 years ago because several studies showed significant association with cervical cancer.¹³ The impact of sexrelated vaginal douching with lemon or lime juice on cervical dysplasia, which is sexually acquired, has not been determined. We sought to determine if there was any association between a history of vaginal douching with lemon/lime and cervical dysplasia among FSWs in northern Nigeria.

Methods

Advocacy and mobilization

Jos is an urban centre in north-central Nigeria with a population of about one million people. We held separate meetings with brothel managers and representatives of FSWs in the Jos metropolis. The purpose of the study was explained at these meetings in order to seek their cooperation. The FSWs expressed concerns about confidentiality and the financial repercussions of time commitment in public health facilities and so a private health centre with adequate facilities was chosen as the study site in preference to a public health centre.

Study Design and Clinical Methods

Douching with citrus juice is a sexrelated practice, therefore, we recruited consenting brothel-based FSWs as subjects for this study. We conducted a cross-sectional observational study of consecutive volunteers. The health talks, pre-and post-test counseling, gynae-cological examination, colposcopy, genital sample collection and laboratory testing were all done at Solat Women Hospital, Jos, Nigeria.

A written informed consent was obtained from each subject and a structured questionnaire was administered by a trained nursecounselor. Personal bio-data, information about lemon/lime usage and duration of sex work were obtained. A gynaecologist who was unaware of the subject’s responses obtained a Pap smear and conducted colposcopic examination of the cervix, vagina and vulva, before and after application of 3% acetic acid. A Pap smear was obtained as previously described¹⁴ and colposcopy examination was conducted using a Carl Zeiss colposcope NAG2, West Germany. Pap smear slides were examined by a trained cytologist and reported using the Bethesda terminology.¹⁵ At colposcopy, the findings were graded as warts (warty lesion on cervix, vagina and/or vulva), cervicitis and suspected squamous intraepithelial lesions (SILs). Biopsy specimens were taken of suspected SILs. Women with detectable abnormalities were offered appropriate treatment and follow up free of cost. Ethical approval for this study was granted by the Ethical Review Board of the Jos University Teaching Hospital.

Statistical Analysis

Data entry and analysis were done using Epi Info version 3.3.2 (CDC, Atlanta Georgia, USA). Associations between cervical dysplasia and use of lemon/lime variables were tested using odds ratios (OR) with 95% confidence intervals (CI) and the Chi square (X² ) test. Multivariate logistic regression was used to assess the independent effect of lemon/lime use on the risk of cervical dysplasia adjusted for HIV status. A P-value less than 0.05 was considered significant.

Results

Of the general study population of 398 subjects who consented to participate in this study and completed questionnaires, 24 did not return for a Pap smear and colposcopy (they were menstruating at the time of the initial evaluation). Only 1.5% (6/398) of these women had previously had a Pap smear. A total of 374 FSWs comprising 81 Lemon users (LUs) and 293 non-lemon users (NLUs) were recruited and examined. The mean age was 27.8 ± 6.7 (range 16-63) years and 94% of the subjects were Christians and 5.5% Moslems. About half (49%) of the subjects were single while 47% were either divorced or widowed and 3.5% were married. Only 7% were illiterate. Age (P=0.56) and education (P=0.76) were unrelated to lemon/lime use. The duration of sex work (P=0.004) correlated positively with lime use. The characteristics of all subjects who participated in this study are outlined in Table 1.

Lime use (90.4%) was more prevalent than lemon use (14.2%) and the principal reason for douching was to prevent infection (75% of subjects). Among LUs, 12% applied lemon/lime juice only before sex while 54% applied it only after sex. About half (49%) of LUs reported discomfort or pain with use. Colposcopy showed 17(4.5%) subjects with genital warts [LUs 5(6.2%); NLUs 12(4.1%); p=0.43], 61(16.3%) with suspected SILs [LUs 17(21.0%); NLUs 44(15.0%); p=0.20] and 65(17.4%) had findings suggestive of chronic cervicitis (Table 3). We did not find vulval or vaginal abrasions at colposcopy. Cytology of Pap smears showed that 87(24.6%) had SILs [LUs 26(33.3%); NLUs 61(22.1%); p=0.03]. Lemon/lime use was associated with cervical dysplasia after controlling for HIV status (Adjusted odds ratio=1.76; 95% CI, 1.0-3.0). In our recent report¹² , HIV prevalence for LUs and NLUs were 48.8% and 48.2% respectively (OR 1.0; 95% CI 0.6-1.2, p=0.9427). Abnormal cytology was not associated with increasing duration of lemon/lime use (P= 0.20 for trend).

Discussion

The main reason why FSWs in this study used citrus juice for sex was to prevent infection. The concentration of citrus juice used, method of application and frequency of use could not be standardized because of obvious limitations in the study design warranted by ethical barriers. However, the following findings deserve careful consideration.

Cytology of Pap smears showed that lemon/lime use was associated with cervical dysplasia (SILs) after controlling for HIV status. On the one hand, the prevalence of SILs among FSWs in this study (87/354; 24.6%) was higher than previously reported for women in the same city (7.7%).¹⁶ On the other hand; there is a suggestion that the practice of douching with citrus juice may be a risk factor for cervical dysplasia. To the best of our knowledge, this association has never previously been reported and it deserves further evaluation in communities where this practice is common. It is also noteworthy that although Pap smear services were available in the local tertiary health facility, only 1.5% of the FSWs had previously had a Pap test. The absence of a population-based cervical screening program in Nigeria is putting the lives of many women at risk. In 1997, Zhang and colleagues¹⁷ reported the results of a meta-analysis of 6 studies of vaginal douching and cervical cancer. Among women who douched at least once per week, the pooled adjusted relative risk was 1.86 (95% CI = 1.29-2.68). Of note, this statistically significant difference was for vaginal douching with any product. Cervical human papilloma virus (HPV) infection is widely accepted as a precursor to dysplasia but in this study, we found no difference in the prevalence of genital warts between LUs and NLUs. The aetiology of cervical cancer is multifactorial. Lysol and other tar-based vaginal douche products were voluntarily removed from the USA market over 30 years ago because several studies showed significant association with cervical cancer. ¹³

It has been demonstrated that even a 20% concentration of either lemon or lime juice had the effect of inhibiting Lactobacilli⁸ and that frequent vaginal douching interferes with the normal vaginal flora thereby predisposing to cervical infections and pelvic inflammatory disease (PID).^2,11,18 This study showed no difference in prevalence of colposcopically detected evidence of chronic cervicitis between lemon/lime users and non users. In our earlier report on this cohort of FSWs, we showed that sex-related douching with lemon or lime juice was not a risk factor for HIV and other sexually transmissible infections.¹² There was however an increased prevalence of bacterial vaginosis although this did not reach statistical significance. ¹²

When tested on cervical explant tissue (cells obtained from routine hysterectomies that have been kept alive in lab cultures), results indicated that the lemon and lime juices caused damage to cells comparable to that of N-9.⁸ Douching with lime juice caused dosedependent epithelial damage to the cervix and vagina in sexually abstinent women volunteers ⁹ but among FSWs who used lime douches for sex, our study found no colposcopically detectable evidence of abrasions or bruises. Whether alkaline semen plays a role in modulating the impact of lemon/lime juice on the epithelium of the genital tract remains a matter of conjecture.

Our study had several limitations. We relied on self reporting by subjects and have no means of verifying claims or controlling for other factors like degree of dilution of lime juice, timing of douching and condom use. We were unable to control for frequency of condom use, timing of douching and degree of citrus juice dilution. Because of the cross-sectional nature of the study, we cannot be confident that use of lime juice preceded the development of cervical dysplasia. Confounding by association may have arisen if use of lime juice is associated with other practices that increase the risk of cervical dysplasia.

There is a widespread habit of vaginal douching among FSWs in Africa and citrus juice is among the frequently used agents. ^2,3,7 Our findings however suggest that sex related douching with citrus juice does not appear to damage genital tract epithelium nor is it associated with increased risk of cervical infections or genital warts. However, further studies to explore the association between douching with lime juice and cervical dysplasia are warranted in communities where this practice is common.

Acknowledgements

We are grateful to the women who participated in this study; and the research staff involved in the field work, counselling and testing of biological samples. We are thankful to the Medical Director and Staff of Solat Women Hospital, Jos for their cooperation and contribution. The strong support and encouragement received from Brian Haill are appreciated. This study was supported by public donation to the Mary Magdalene project, www.aids.net.au.

References

1. Wang B, Li X, Stanton B, Yang H, Fang X, Zhao R, Dong B, Zhou Y, Liu W, Liang S. Vaginal douching, condom use and sexually transmitted infections among Chinese female sex workers. Sex Transm Dis, 2005; 32:696-702.
2. Gresenuet G, Kreiss JK, Chapko MK, Hillier SL, Weiss NS. HIV infection and vaginal douching in central Africa. AIDS, 1997; 11:101-6.
3. Fonck K, Kaul R, Keli F, Bwayo JJ, Ngugi EN, Moses S, Temmerman M. Sexually transmitted infections and vaginal douching in a population of female sex workers in Nairobi, Kenya. Sex Transm Infect, 2001; 77:271-5.
4. Rustomjee R, Abdool Karim Q, Abdool Karim SS, Laga M, Stein Z. Phase I trial of nonoxynol-9 film among sex workers in South Africa. AIDS, 1999; 13:1511-5.
5. Nwoha PU. The immobilization of all spermatozoa in vitro by bitter lemon drink and the effect of alkaline pH. Contraception, 1992; 46:537-42.
6. Clarke GN, McCoombe SG, Short RV. Sperm immobilizing properties of lemon juice. Fertil Steril, 2006; 85:1529-30.
7. Imade GE, Sagay AS, Onwuliri VA, Egah DZ, Potts M and Short RV. Use of lemon or lime juice douches in women in Jos Nigeria. Sexual Health, 2005; 2: 237-9.
8. Lackman-Smith C, Snyder B, Luckenbaugh K. In vitro assessment of efficacy and cytotoxicity of natural substances proposed as vaginal microbicides against sexual transmission of HIV. Poster presented at 13th Conference on Retroviruses and Opportunistic Infections, Paper 894, February 6, 2006.
9. Why women should not use lemon or lime juice as a microbicide. Global Campaign for Microbicides 2006, PATH. http://www.nswp. org/pdf/LEMON-060624.PDF
10. Short R.V. New ways of preventing HIV infection: thinking simply, simply thinking. Phil Trans R Soc B, 2006; 361(1469): 811-20
11. Scholes D, Stergachis A, Ichikawa LE, Heidrich FE, Holmes KK, Stamm WE. Vaginal douching as a risk factor for cervical Chlamydia trachomatis infection. Obstet Gynecol, 1998; 91:993-7.
12. Imade G, Sagay A, Egah D, Onwuliri V, Grigg M, Egbodo C, Thacher T, Potts M, and Short, R Prevalence of HIV and other sexually transmissible infections in relation to lemon or lime juice douching among female sex workers in Jos, Nigeria. Sexual Health, 2008; 5: 55–60.
13. Harry W. Haverkos Multifactorial Etiology of Cervical Cancer: A Hypothesis Med Gen Med, 2005; 7(4): 57.
14. Boon ME, Suurmeiyer AJH. Papanicolaou Staining Method. In Pap smear, 3^rd Edition. Taylor Francis UK. 1998. 257-266.
15. NCWI. The 1988 Bethesda system for reporting cervical / vaginal cytologic diagnosis. JAMA, 1989; 262: 931-4.
16. Obafunwa J.O., Sagay A.S., Otubu J.A.M. Prevalence of Cervical Intraepithelial Neoplasia in Jos, Nigeria. Tropical Journal of Obstetrics and Gynaecology, 1991; 9(2):16-17.
17. Zhang J, Thomas AG, Leybovich E. Vaginal douching and adverse health effects: A metaanalysis. Am J Public Health. 1997; 87:1207–1211.
18. Scholes D, Daling JR, Stergachis A, Weiss NS, Wang SP, Grayston JT. Vaginal douching as a risk factor for acute pelvic inflammatory disease. Obstet Gynecol. 1993 81(4):601-6.

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