What Is Normal Vaginal Flora?

Genitourin Med 1997;73:23-28 23 What is normal vaginal flora?

Original J

C F Priestley, B M Jones, J Dhar, Linda Goodwin Genitourin Med: first published as 10.1136/sti.73.1.23 on 1 February 1997. Downloaded from Article

Objective: To observe the composition of the vaginal flora of healthy women over time, and in relation to hormonal changes, sexual activity, and hygiene habits. Design: A longitudinal surveillance of the vaginal flora over an eight week period. Subjects: 26 female health care workers in local genitourinary medicine clinics. Methods: The participants were anonymised. They filled in diary cards daily. Blind vaginal swabs were self-taken two-seven times weekly. A smear was air-dried for later Gram staining. The swabs were also cultured for Candida spp, Gardnerella vaginalis, anaerobes, Mycoplasma hominis and Ureaplasma urealyticum. Results: Of 26 subjects, only four had normal vaginal microbiology throughout. One woman, who was not sexually active, had bacterial vaginosis (BV) throughout and nine (35%) had intermittent BV. Candidiasis was found intermittently in eight women (31%), and eight had normal microscopy. U urealyticum was isolated intermittently in 40% of women with BV, 25% with candida, and 50% with normal microscopy. Many women were symptomatic, but symptoms correlated poorly with microbiological findings. All but two women were sexually active; however, more women with BV were exposed to semen. BV seemed to be related to frequent use of scented soap, and there appeared to be an additive effect of clothing and hygiene factors. Conclusions: Our study raises doubts about what should be regarded as normal vaginal flora. It calls into question the significance of finding BV or U urealyticum on a single occasion in asymptomatic women, or of finding normal flora in symptomatic women. The effect of external factors on the vaginal flora deserve further study. (Genitourin Med 1997;73:23-28) Keywords: vaginal flora; bacterial vaginosis; candidiasis; ureaplasma

Introduction Methods Bacterial vaginosis was first reported in 1955 Volunteer women http://sti.bmj.com/ by Gardner and Dukes,' who described the The study took place over an eight week unique clinical signs and symptoms and the period to cover two menstrual cycles. Subjects distinctive nature of the vaginal discharge were recruited from health care staff of geni- associated with it. They also described a tourinary medicine departments in Sheffield, "new") causative organism, which they named Rotherham, and Barnsley. They were

"Haemophilis vaginalis", subsequently anonymised to encourage participation and on September 30, 2021 by guest. Protected copyright. renamed Gardnerella vaginalis. increase the accuracy of recorded personal Forty years on, while the complexity of the data. Application was made to the local Ethics microbiological picture has been well Committee; however it was felt that formal described,23 the aetiology and pathogenesis of approval for the study was not required as par- the condition remain poorly understood, and ticipation was both voluntary and anonymous. the reasons as to why the normal, healthy lacto- The participants randomly chose numbered bacillus flora is superseded by an overwhelming packs containing swabs, slides and Amies "cocktail" of bacteria are still not known. transport media, and were asked to complete Department of Fortunately, laboratory diagnosis can be readily diary forms to record age, contraceptive Genitourinary Medicine, The Royal made by simple, Gram stain microscopy method used, menstruation, tampon use, Hallamshire Hospital, where "clue cells" and the typical, abnormal coitus, apparel (tights and trousers), bath Sheffield bacterial flora of BV are easily recognised.4 additives, and any symptoms of discharge, irri- C J F Priestley Dhar The experience of some workers suggests tation, or odour. The diary forms were J that factors such as hormonal status, coitus, retained separately until the end of the study. Department of Medical Microbiology, and exposure to semen may play an important The women took blind vaginal swabs from University of Sheffield role in the pathogenesis of BV. We were also themselves two-seven times per week, prior to Medical School interested in the role of clothing and hygiene bathing. They prepared air-dried smears, for B M Jones L Goodwin habits such as the use of certain bath additives, subsequent Gram staining, before placing the Address correspondence to: which are said to precipitate vulvovaginal can- swab into the transport medium for culture. Dr C J F Priestley, didiasis in some women. As there are few doc- These swabs were left at room temperature Department of Genitourinary Medicine, Weymouth and umented studies on these aspects, we designed until the participant could deliver them to the District Hospital, Melcombe a longitudinal study to observe the composi- clinic, and swabs taken on a Friday or Avenue, Weymouth, Dorset DT4 7TB, UK. tion of the vaginal flora in healthy women, in Saturday, in particular, could be left for two to Accepted for publication relation to hormonal changes and external three days before being plated out. For this 8 November 1996 factors. reason we placed greater emphasis on the 24 24Priestley, Jones, Dhar, Goodwin

Gram stained slides than cultures, in particu- further three only completed six or seven lar for the diagnosis of BV. The samples, iden- weeks of the study, but we felt that sufficient tified only by number, were tested "blind" in data were obtained to include them in the the laboratory. We also asked the participants analysis. Overall, the 26 women included in to measure the pH of the secretions, using the analysis took a mean of 3.2 swabs per week Genitourin Med: first published as 10.1136/sti.73.1.23 on 1 February 1997. Downloaded from standard paper strips, prior to preparing the (range 1.7-7.0) for a six to eight week period. slides; however, we became concerned about the accuracy of a reading from a blind vaginal Microbiology swab because of potential contamination from One woman had BV throughout the period of the vulva or cervical mucosa, and pH was not study. G vaginalis, anaerobes, Ureaplasma ure- measured by all the participants. alyticum and Mycoplasma hominis were isolated from the majority of her swabs. She com- Microbiological tests plained of some irritation but not of vaginal Microscopy Gram stained smears were exam- discharge or odour, the typical symptoms of ined for the presence of lactobacilli, "clue BV. BV was found intermittently in nine cells", "gram-variable" organisms, curved rods women (34-6%) in between 10 and 62% and yeasts. The results were recorded as "nor- (mean 27.5%) of swabs taken. G vaginalis was mal", where the flora comprised lactobacilli isolated from 64% and anaerobes from 28% of only; "normal, but with yeasts"; "intermedi- swabs showing BV. ate", with an abnormal BV-type flora but Candida spp were found intermittently in some lactobacilli still present; or "bacterial eight women (308%) in between 14 and 70% vaginosis", where the smears showed "clue (mean 35 5%) of swabs taken. Only eight cells" and masses of gram-variable bacteria, women (30.8%) had normal microscopy and a complete absence of lactobacilli.4 throughout. Ureaplasma urealyticum was isolated inter- Cultures The swabs were cultured for G vagi- mittently in 10 (38.5%) women; four (40%) of nalis, anaerobic organisms, Mycoplasma those with BV, two (25%) of those with can- hominis, Ureaplasma urealyticum, and yeasts. didiasis, and four (50%) of those with normal Plates of Columbia agar (Unipath Ltd, microscopy. Table 1 shows the characteristics Basingstoke, UK) containing 7% horse blood of these women. The presence of U ure- were inoculated and incubated, at 37°C in an alyticum did not appear to be related to sexual atmosphere of 5% CO, in air for 48 hours. G activity or exposure to semen. vaginalis isolates were identified by Gram stain Mycoplasma hominis was only isolated in two and hippurate hydrolysis tests. Yeasts were women-the one with constant BV, in whom confirmed by microscopy only but were it was isolated in 96% of swabs, and one other assumed to be Candida spp. For the isolation with intermittent BV, in whom it was found in of anaerobic organisms, plates of Columbia 3% of swabs taken. ,B-haemolytic streptococci Blood agar, containing kanamycin at 75 ,ug per were isolated from two women, both of whom ml were inoculated and incubated for 48 had Candida spp; in one it was found only on a http://sti.bmj.com/ hours, at 37°C in an anaerobic cabinet (Don single occasion, and in the other it was isolated Whitley Scientific Ltd, UK) containing nitro- from four consecutive swabs (22% of total) gen 80%, hydrogen 10% and carbon dioxide during the second week of the study. 10%. A 50 ,ug metronidazole disc was added Overall, only four out of 26, or 15% of to the inoculated streaks and, if a zone of women, had apparently completely normal

colony inhibition was seen after incubation, flora throughout. on September 30, 2021 by guest. Protected copyright. the presence of "anaerobes" was recorded. There were no significant differences in the Cultures for the isolation of mycoplasma and number of swabs taken per week between the Ureaplasma spp were made by inoculating women with BV, Candida spp, normal plates of "MUD" medium (Unipath Ltd, microscopy, or U urealyticum, and in addition, Basingstoke, UK) directly with the swab, and there were no significant differences in the incubating for two days in the anaerobic cabi- proportions of positive swabs taken on a net. Cultures were examined and species iden- Friday or Saturday with those taken the rest of tified using a x 10 microscope objective. the week. This suggests that the intermittent nature of the microbiological findings was not Analysis Statistical analysis was performed due to a significant loss of sensitivity arising using Epi info, version 6. Results were from the culture methods. analysed using the Fisher exact 2-tailed test for small numbers and the chi square test for larger numbers. Table 1 Characteristics ofwomen with U urealyticum (UU) isolatedfrom vaginal swabs UU-% of Exposure to Results Subject no Microscopy swabs taken semen All female staff working in the three genitouri- 5 normal 6-5 + nary medicine clinics (approximately 45 6 normal 7-7 + 7 nonnal 53-8 women) were invited to participate and 33 8 normal 73-3 were entered into the study. However, four 9 candida 81-0 + women not 16 candida 2-7 did complete the study and three 20 BV 20-0 + did not return their diaries. Of the remaining 22 BV 74-1 + 26 women, two 23 BV 82-4 + three took fewer than swabs 26 BV 893 - per week overall (1.7, 1-8, 1.9 per week) and a What is normal vaginalflora? 25

Table 2 Characteristics ofwomen with BV, candidiasis, and normal microscopy Effect of menstruation Normal Candida BV p value Two women had had hysterectomies, and one Variable (n = 8) (n = 8) (n =10) BVvs normal was post-menopausal. For the remaining Average age range 39.4 35-6 405 - women, there was no difference between the Sexually active 8 7 9 0-91 three groups in the duration of menstruation, Coitus perweek 2-1 1-5 1-4 Genitourin Med: first published as 10.1136/sti.73.1.23 on 1 February 1997. Downloaded from (range) (06-4.5) (0-2 4) (0-2 5) but the women with candidiasis appeared to Exposure to semen 5 (62 5%) 4 (50%) 9 (90%) 0-27 use tampons for a greater proportion of this Menstruating 7 7 9 1-00 Tampon use 6 (75%) 7 (88%) 6 (60%) 0-64 time (table 2). A comparison of the weeks of Tampons > 50% of menstruation 4 (50%) 7 (88%) 4 (40%) 1-00 Symptoms: the menstrual cycle during which BV and Discharge > 2 days per week 1 (13%) 1 (13%) 1 (10%) 1-00 Candida spp were present showed that BV was Irritation > 2 days per week 0 (0%) 1 (13%) 1 (10%) 1-00 more common the first week of the Odour > 2 days per week 0 (0%) 0 (0%) 1 (10%) 1-00 during External factors: cycle, whereas Candida spp were found more Tights > 2 days per week 4 (50%) 6 (75%) 6 (60%) 1 00 frequently during the last week (table Trousers > 2 days per week 8 (100%) 5 (63%) 7 (70%) 0-22 3). Bubble bath > 2 days per week 2 (25%) 4 (50%) 4 (40%) 0-64 Scented soap > 2 days per week 2 (25%) 4 (50%) 9 (90%) 0.009* > 3 of above > 2 days per week 1 (13%) 3 (38%) 7 (70%) 0.02* Relation with symptoms There were no significant differences in these variables between the BV and candida groups, or There was no significant difference between between those with candida and normal microscopy. the groups in the numbers of women with discharge, odour, or irritation for more than two The women were divided for analysis into days per week (table 2). three groups; those with normal flora, candidi- We looked at symptoms in the individual asis, and BV on micrQscopy. There was no sig- subjects (fig 1). In these graphs, the subjects nificant difference in the age groups of the are ranged in ascending order of the frequency women. which abnormal swabs were found, and there was no correlation between the frequency with Effect ofsexual activity which BV, Candida spp, or U urealyticum were All but two were sexually active (table 2). found and the frequency of symptoms. Subject Those with normal flora tended to have inter- 25, with intermittent BV, appears to be symp- course more frequently, but there was no sig- tomatic, but she actually complained of symp- nificant difference between the groups. toms on as many days when the microbiology However, all nine women with intermittent was normal as when it was abnormal. The BV had unprotected sex, compared with 50% same was true for women with candidiasis, of those with candidiasis and 63% of those although three women (subjects 9, 13, and 15) with normal microscopy, although this differ- were symptomatic enough to treat themselves. ence did not reach significance (p = 0.2). The women with normal microscopy had Details of oral sexual contact were not fewer symptoms overall, with one exception. obtained. The woman with constant BV had Subject 6, with normal microscopy, had U ure- never had sex with a man, and had not been alyticum isolated from 8% of swabs taken; its sexually active at all for the previous five years. presence was not related to symptoms. There The numbers were too small to draw any con- was no correlation between the symptoms and http://sti.bmj.com/ clusions about the effects of contraception. the menstrual cycle.

Normal microscopy Bacterial vaginosis 7 7 6 6 a) 5 -h5a) 5 on September 30, 2021 by guest. Protected copyright.

4 '-.4" 4- a) a) 0. 3 . 3 c0 cc 2 1 0 rtl g 1 2 3 4 5 6 7 8 17 18 19 20 21 22 23 24 25 26 Subject no. Subject no. Candida spp. Ureaplasma spp. 7 7 6 6 M Discharge

a 5 E1 Irritation a)'-4 |:D Odour 0. Q 4 a) a) CL 3 0.3 CO 2 0

1 am n.[L-- mon. 1 ll *n 9 10 11 12 13 14 15 16 16 5 6 20 7 8 22 9 23 26 Subject no. Subject no. Figure 1 Symptoms in relation to microbiology of the vagina. i 0x. . . _ _K . . _ .0 . _ . . . .0s 26 Priestley, Jones, Dhar, Goodwin

Figure 2 Longitudinal changes in flora in Subject Subject no. 23 I Normal 23. Abnormal Contraception: coil Intermediate Bacterial vaginosis S Spermatozoa

Age: 35-39 years Genitourin Med: first published as 10.1136/sti.73.1.23 on 1 February 1997. Downloaded from

Symptoms: Discharge m 0 Irritation Odour a Ua Culture: Candida spp. mE 11 11 111D11D1D1E11E1D1 a Ureaplasma spp. so _0 _~ 0 Ill __5 T n _~ Gardnerella spp. mW I I DU-L~ m Anaerobes mW LI D1111w _ _0 [E N BV Microscopy: a AUD I@ s s m FSI 1 lsis s Dn1 U*E LD__ <5 >5 pH n WEDll I_ El El 1 E Coitus DEl El El Menstruating Euilll Elull I~~~~~~~~~~~~~~~~~~~~~~~~|:.'...E1I',,:,S,,,', -..;I.gI 1 2 3 4 5 6 7 8 Week of study

Externalfactors (table 2) There also appeared to be an additive effect; There was no significant difference between women with BV were significantly more likely the three groups in the frequency of wearing to use three or more of these factors for more tights or trousers or using bubble bath, but than two days a week (p = 0 03). more women with BV used scented soap fre- We have encountered patients with BV who quently. This difference was significant are overzealous with washing because they are between those with BV and normal flora (p = aware of an odour. To determine if this could 0.009), but not between those with BV and account for the difference, we looked at the = candida (p 0 9). relationship between odour and the use of http://sti.bmj.com/

Figure 3 Longitudinal changes in vaginalflora in Subject no. 18 I Normal Subject 18. | Abnormal Contraception: POP Intermediate Bacterial vaginosis

Age: 30-34 years I Spermatozoa on September 30, 2021 by guest. Protected copyright. Symptoms: I- - Discharge - Irritation - Odour .

Culture: Ureaplasma spp. DIP ED[EDr I DID DID Gardnerella spp. DII [L] Dii DI L] DII]1 DID1 Anaerobes Zul -_II]D - N BV Microscopy: EW mED DII] - DITlr DIE DIEDT <5 >5 pH _ DIEDI 1 ElIDI 1,IID1 1 _ Coitus El El3 El EDE E H Menstruating LI.IZI, LP E .iI,i..1.. :B- .'°;sf I E :1; s I 1 2 3 4 5 6 7 8 Week of study What is normal vaginalflora? 27

Figure 4 Longitudinal changes in vaginalflora in Subject no. 22 Normal Subject 22. Abnormal Post-menopausal Intermediate Bacterial vaginosis _ Spermatozoa Age->50 Genitourin Med: first published as 10.1136/sti.73.1.23 on 1 February 1997. Downloaded from

Symptoms: Discharge Irritation Odour Culture: Ureaplasma spp. O Gardnerella spp. 1111111111lliL11 0 Anaerobes ll ilill liTTTTlll L N BV Microscopy: 1m _ 1ls IO Coitus E m E E m11 eIs 1 Is 11 1_ ll 1 2 3 4 5 6 7 8 Week of study

scented soap. There was no correlation, sug- study period, accounting for 26% of swabs gesting that in our cohort, more frequent use taken. U urealyticum was isolated during the of scented soap was not due to increased first three weeks of the study; it was not iso- awareness of odour. lated during the next three weeks but returned in weeks seven and eight. This pattern sug- Longitudinal changes in women with intermittent gests that the negative swabs were not false BV negatives. This woman was asymptomatic The microbiological findings of the women throughout. with intermittent BV, together with data from their diaries, were looked at in detail. Subject 23 (fig 2) had BV or intermediate flora on Discussion 32% of swabs taken. Intermediate flora was Few longitudinal studies of the vaginal flora noted on four occasions, and on two of these it have been performed, and this is the most progressed to BV. This appeared to occur in extensive that has been reported to date. In the second half of the menstrual cycle, follow- 1977 Bartlett performed quantitative bacteri- ing periods of frequent sexual activity. The ology on samples collected from five volun- http://sti.bmj.com/ appearances of BV correlated with the pres- teers at intervals throughout the menstrual ence of G vaginalis and anaerobes on culture. cycle.5 He found that the concentrations of U urealyticum was present on the majority of anaerobes were maintained at relatively con- swabs taken. The pH rose, as would be stant levels throughout the menstrual cycle, expected, during menstruation, and also after but that the concentrations of aerobic bacteria

the periods of frequent coitus. It was noted decreased in the premenstrual week. Analysis on September 30, 2021 by guest. Protected copyright. that the periods when BV was found were pre- of sequential specimens showed considerable ceded by a rise in pH. This woman had few variation in the species recovered. In 1980 symptoms, which were not related to abnor- Sautter collected serial specimens from seven malities in the flora. nurses during one menstrual cycle.6 He too Subject 18 (fig 3) had BV during one week of found a cyclical variation, with bacteroides the study (15% of swabs taken). Anaerobes being isolated mainly during the first half of were isolated during this period and also fre- the menstrual cycle. He also found U ure- quently when the microscopy was normal. alyticum constantly in one, and intermittently Again, the period of BV was preceded by a rise in five, of the seven subjects. In 1982 Larsen in pH. Her few symptoms were not correlated cultured the vaginal fluid of 31 women during to the vaginal flora. and after menstruation7; 19 women showed a Subject 22 (fig 4) was post-menopausal; she decrease in the number of anaerobes and 22 a had two episodes of BV at the beginning of the decrease in the number of aerobes after menstruation. In our study, 38% of healthy women had Table 3 Effect of menstrual cycle on the vaginalflora (menstruating women with BVor intermittent BV, 38% intermittent U ure- Candidiasis; n = 16) alyticum, and 31% intermittent candidiasis; Candidiasis (n = 7) Total BV (n = 9) only 15% had apparently completely normal Days of total swabs total swabs flora throughout. Many women had symp- menstrual cycle taken % positive p value taken % positive p value toms, but these were not closely related to the 1 to7 39 44 62 60 microbiological findings. Cultures for Neisseria 8to 14 48 42 63 40 gonorrhoeae and Chlamydia trachomatis were 15 to 21 38 39 0-36 48 52 0.05 not performed as this would have compro- 22 onwards 42 57 46 37 mised the anonymous nature of the study; 28 Priestley, J7ones, Dhar, Goodwin

however, vaginal discharge is an uncommon who have suffered from symptomatic candidia- presentation of these infections, and they are sis, avoiding the known or suspected precipi- unlikely to have been a common cause of the tants. However, this study emphasises the symptoms described. Because of potential multifactorial aetiology of BV, and we feel that delays in plating out the swabs, we placed these factors warrant further investigation. Genitourin Med: first published as 10.1136/sti.73.1.23 on 1 February 1997. Downloaded from more reliance on the microscopy than culture The significance of U urealyticum and M of the specimens, and did not perform quanti- hominis as pathogens has not been established tative cultures. However, microscopy of a without doubt. U urealyticum is an infrequent Gram stained smear has been shown to be cause of urethritis in men'" and it is suggested both sensitive and specific in the diagnosis of that it can be sexually transmitted. In our BV,8 and both our microscopy and culture study its presence was not related to exposure results, using standard diagnostic methods to semen, and indeed it was found most fre- widely used in genitourinary clinics, are in quently in a woman who had never had sex accordance with these previous studies in with a man. Similarly, the significance of the demonstrating the dynamic nature of the fi-haemolytic streptococcus as a vaginal microbial population of the vaginal flora and pathogen is unclear, although it has been asso- its cyclical variation. ciated with an adverse outcome of preg- It has been recognised that BV and candidi- nancy.'2 In our study it was only found asis may resolve spontaneously without treat- sporadically in two women, both of whom also ment9; however, the high frequency with had Candida spp., and it could not be deter- which these conditions may occur intermit- mined whether it was a contaminant or a nor- tently may not have been appreciated until mal constituent of the vaginal flora. now. We have demonstrated that abnormali- ties may occur for only a few consecutive days before resolving spontaneously. This has Conclusion implications for future studies; if swabs are Our study raises doubts about what should be taken less frequently or indeed only on a single regarded as normal vaginal flora. It calls into occasion, episodes of BV or candidiasis may question the significance of finding bacterial be missed, or a single positive swab may be vaginosis, Candida spp., M hominis, U ure- given more emphasis than it deserves. In com- alyticum, or ,B-haemolytic streptococci on a sin- mon with other studies,'0 we found BV more gle occasion in asymptomatic women, as well commonly in women having unprotected sex, as the significance of finding normal flora on a but it was only found consistently in one single occasion in symptomatic women. woman who was not sexually active. This pro- We are grateful to our colleagues in Sheffield, Barnsley, and vides further evidence that BV is not necessar- Rotherham who so readily participated in this study, and to Ms ily sexually transmitted. Pauline Whitaker for her help in preparing the manuscript. We had reservations about the accuracy of the pH reading from a blind vaginal swab, 1 Gardner HL, Dukes CD. Haemophilis vaginalis vaginitis. Am J Obstet Gynecol 1955;69:962-72. http://sti.bmj.com/ because of possible contamination from the 2 Spiegel CA, Amsel R, Eschenbach D, Schoenkneckt R, vulva or cervical mucosa, and it was not mea- Holmes KK. Anaerobic bacteria in non-specific vaginitis. NEngJtMed 1980;303:601-7. sured by all the participants. However, the 3 Spiegel CA, Davick P, Totten PA, Chen KCS, Eschenbach results in these women are presented because DA, Amsel R, Holmes KK. Gardnerella vaginalis and anaerobic bacteria in the etiology of bacterial vaginosis. of their potential significance; it has not yet ScandJ3 InfDis Suppl 1983;40:41-3. been established whether the rise in pH that is 4 Nugent RP, Krohn MA, Hillier SL. Reliability of diagnosing

in bacterial vaginosis is improved by standard method of on September 30, 2021 by guest. Protected copyright. found association with BV is a cause or an Gram stain interpretation. Y Clin Microbiol 1991 ;29: effect. Our finding that the change to the BV 297-301. 5 Bartlett JG, Onderdonk AB, Drude E, Goldstein C, type flora is preceded by a rise in pH would, if Anderka M, Alpert S, McCormack WM. Quantitative confirmed, suggest that the pH increase may bacteriology of the vaginal flora. J Infect Dis 1977;136: 271-7. be a cause rather than an effect of the condi- 6 Sautter RL, Brown WJ. Sequential vaginal cultures from tion. normal young women. 7 Clin Microbiol 1980;11:479-84. 7 Larsen B, Galask RP. Vaginal microbial flora: composition Our results are consistent with the theory and influences of host physiology. Ann Int Med 1982;96: that semen is one of a number of factors that 926-30. can alter the 8 Hay PE, Taylor-Robinson D, Lamont RF. The diagnosis environment of the vagina, possi- of bacterial vaginosis in a gynaecology clinic. BrJ Obstet bly by raising the pH, and trigger off a change Gynaecol 1992;99:63-6. in the flora. 9 Moi H. Prevalence of bacterial vaginosis and its association We looked at some of the external with genital infections, inflammation, and contraceptive factors which could potentially alter the envi- methods in women attending sexually transmitted disease and primary health clinics. Int J STD AIDS 1990;1: ronment of the vagina. BV seemed to be 86-94. related to frequent use of scented soap, and 10 Bowie WR. Urethritis in males. In: Holmes KK, Mardh there appeared to be an PA, Sparling PF, Weisner PJ, eds. Sexually Transmitted additive effect of Diseases. 2nd ed New York: McGraw-Hill, 1989:627-39. clothing and hygiene factors. Obviously no 11 Janier M, Lassau F, Casin I, Grelot T, Scieux C, Zavavo A, firm conclusions can be drawn in view of et al. Male uretritis with and without discharge: a clinical the and microbiological study. Sex Transm Dis 1995;22: small numbers studied, and in addition, the 244-52. results may have been 12 Gibbs RS, Romero R, Hillier SL, Eschenbach DA, Sweet confounded by this RL. A review of premature birth and subclinical infec- highly aware group of women, especially those tion. AmJr Obstet Gynecol 1992;166:1515-28.