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To study the cause of nonspecific vaginitis, we analyzed vaginal fluid from normal women and from 53 women with nonspecific vaginitis, using quantitative anaerobic cultures and gas-liquid chromatography for short-chained organic-acid metabolites of the microbial flora. In normal vaginal fluid, lactate was the predominant acid, and the predominant organisms were lactobacillus and streptococcus species (lactate producers). In nonspecific vaginitis, lactate was decreased, whereas succinate, acetate, butyrate, and propionate were increased, the predominant flora included Gardnerella (Haemophilus) vaginalis (acetate producer), and anaerobes, which included bacteroides species (succinate producers) and peptococcus species (butyrate and acetate producers). After metronidazole therapy, symptoms and signs of nonspecific vaginitis cleared, butyrate and propionate disappeared, and lactate and lactate-producing organisms became predominant. We conclude that certain anaerobes act with G. vaginalis as causes of nonspecific vaginitis, and that a high ratio of succinate to lactate in vaginal fluid is a useful indicator in the diagnosis of this condition.

To determine whether bacterial vaginosis (BV), also known as nonspecific vaginitis, could be diagnosed by evaluating a Gram stain of vaginal fluid, we examined samples from 60 women of whom 25 had clinical evidence of BV and 35 had candidal vaginitis or normal examinations. An inverse relationship between the quantity of the Lactobacillus morphotype (large gram-positive rods) and of the Gardnerella morphotype (small gram-variable rods) was noted on Gram stain (P < 0.001). When Gram stain showed a predominance (3 to 4+) of the Lactobacillus morphotype with or without the Gardnerella morphotype, it was interpreted as normal. When Gram stain showed mixed flora consisting of gram-positive, gramnegative, or gram-variable bacteria and the Lactobacillus morphotype was decreased or absent (0 to 2+), the Gram stain was interpreted as consistent with BV. Gram stain was consistent with BV in 25 of 25 women given a clinical diagnosis of BV and in none of 35 women with candidal vaginitis or normal examinations. Duplicate slides prepared from 20 additional specimens of vaginal fluid were stained by two methods and examined by three evaluators. Interevaluator interpretations and intraevaluator interpretations of duplicate slides were in agreement with one another and with the clinical diagnosis .90% of the time. We concluded that a microscopically detectable change in vaginal microflora from the Lactobacillus morphotype, with or without the Gardnerella morphotype (normal), to a mixed flora with few or no Lactobacillus morphotypes (BV) can be used in the diagnosis of BV.

To determine the cause of the acute urethral syndrome, we studied 59 women with dysuria and frequent urination without "significant bacteriuria" (defined as greater than or equal to 10(5) organisms per milliliter), 35 women with typical cystitis and 66 women with no symptoms of urinary-tract infection. Although none of the 59 women with urethral syndrome had greater than 3.4 x 10(4) bacteria per milliliter in either of two successive midstream urine specimens, samples of bladder urine obtained by suprapubic aspiration or catheterization from 24 women contained coliforms, and samples from three contained Staphylococcus saprophyticus; all but one of these 27 women also had pyuria. Of the 32 women with sterile bladder urine, 10 of 16 with pyuria and one of 16 without pyuria were infected with Chlamydia trachomatis (P = 0.002). Chlamydial infection was found in 11 of 42 women with urethral syndrome and pyuria, in three of 66 without symptoms, and in one of 35 with cystitis (P less than 0.01 when the group with urethral syndrome is compared with either of the other groups). Thus, 42 of 59 women with urethral syndrome had abnormal pyuria and 37 of these 42 were infected with coliforms, S. saprophyticus, or C. trachomatis, whereas few women without pyuria had demonstrable infection. Bacteriuria of greater than or equal to 10(5) per milliliter may be an insensitive diagnostic criterion when applied to symptomatic lower-urinary-tract infection.

New selective and differential human blood bilayer agar media with Tween 80 (HBT medium) or without Tween 80 (HB medium), developed for the isolation of Gardnerella (Haemophilus) vaginalis, permitted significantly higher G. vaginalis isolation rates than have been obtained for other media used for this purpose. HB medium consists of a basal layer of Columbia agar base containing colistin and naladixic acid with added amphotericin B and an overlayer of the same composition plus 5% human blood. HBT agar also contains Proteose Peptone No. 3 (Difco Laboratories) and Tween 80 in the basal layer and the overlayer. Both Tween 80 and the bilayer composition enhanced G. vaginalis production of human blood hemolysis, permitting detection of this organism even in the presence of heavy growth of other vaginal flora. The use of HB or HBT medium thus permitted the demonstration that G. vaginalis was present in vaginal fluid from a large percentage (up to 68%) of normal women. However, the concentration of G. vaginalis was found by semiquantitative analysis to be significantly higher in vaginal fluid from women with nonspecific vaginitis than in fluid from normal women.

To assess correlations between the content of the diamines putrescine and cadaverine in vaginal fluid and the clinical manifestations of vaginitis, a rapid procedure for the determination of diamines has been developed. The procedure involves thin-layer chromatography of the dansylated vaginal washings; then the fluorescence of dansyl diamines is compared with that of dansyl alanine. For 520 college women, the result of the test for diamines was positive in 173 (88%) of 197 women with nonspecific vaginitis or vaginitis due to Trichomonas vaginalis and was negative in 291 (90%) of 323 women without nonspecific vaginitis or vaginitis due to T. vaginalis. The diamine content of vaginal fluid specimens after therapy with metronidazole was correlated with the clinical response to treatment.

Vaginal fluid samples from normal college students, college students with bacterial (nonspecific) vaginosis, and sexually transmitted disease clinic patients with bacterial vaginosis, before and after therapy with metronidazole, ampicillin, or amoxicillin, were evaluated by direct Gram stain and culture for the predominant anaerobic and facultative flora. Curved rods were detected by direct Gram stain of vaginal fluid from 31 (51%) of 61 women with bacterial vaginosis and none of 42 normal student controls (P less than 0.001). Curved, gram-variable to gram-negative organisms were recovered from six of these 31 women, seven other women with bacterial vaginosis, and no controls. All 13 isolates were anaerobic, motile, and oxidase-negative, produced succinic acid as their major metabolic product, and hydrolyzed starch. After treatment with ampicillin or amoxicillin (n = 10) or greater than or equal to 2 g of metronidazole (n = 9), no curved motile rods were detected by Gram stain or culture, although the minimal inhibitory concentration of metronidazole was greater than or equal 8 micrograms/ml for 11 of the 13 isolates tested.

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