Increased numbers of vaginal PMNs have been associated with increased risk of PTB [4,5]. A number of studies have assessed the vaginal PMN count using vaginal smears, but these have been clinician-taken samples. Could self-taken samples be used for assessing vaginal PMNs? This is the second question that Culhane and colleagues sought to answer in their study. They counted the mean number of leukocytes on the same vaginal smears taken from three different vaginal sites. They found a significantly higher leukocyte count from the introital smear compared to the mid-vagina and vaginal fornix smears with the mean values being 6.5, 2.8 and 2.5 respectively (p=0.001). They suggest that due to the significant variation of the number of leukocytes at different sites in the vagina, self-taken smears are not appropriate for measuring vaginal leukocytes counts.

Assessing PMN counts on smears of the urethra and cervix by microscopy is known to be subject to inconsistencies due to field selection, and preparation techniques such as the thickness and spread of the smear, which affect the cellular density [9]. Assessment of PMN counts on vaginal smears is no different, so the development of a more reliable method of quantifying PMN counts would be useful.

Ramsey and colleagues attempted this by assessing the polymorphonuclear leukocyte to epithelial cell ratio (PMN/EPI ratio) on vaginal smears taken from the upper vagina, at 20-25 weeks gestation, to see if this ratio predicted preterm birth. They performed a nested case-control study as part of a larger prospective longitudinal cohort of pregnant women. The population was mainly African-American women attending an antenatal clinic at the University of Alabama.

Eighty-three women with a spontaneous preterm birth at less than 35 weeks gestation were compared with a control group of 108 randomly selected women who delivered at term. The mean PMN count was similar in the cases (13 + 20 cells/field) and in the controls (10 + 14 cells/field) [p=0.17]. The mean PMN/EPI ratio was significantly higher among the cases (3.4 + 6.0) than among the controls (1.8 + 2.4) (p = 0.01). PMN/EPI ratios greater than the 95th percentile value based on control values (7.0/field) were significantly associated with spontaneous PTB (odds ratio = 3.8, 95% CI 1.3-11.2; p = 0.009). In a logistic regression model controlling for the presence of BV, a PMN/EPI ratio greater than 7.0/field remained significantly associated with spontaneous preterm birth (odds ratio = 3.8, 95% CI 1.3-11.2; p = 0.01). The intraobserver coefficient of variation for PMN counts was 65.3% and the use of PMN/EPI ratio improved this to 38.6%.

The high specificity of the PMN/EPI ratio (95.4%) means that a negative test would be good at identifying women at low risk of PTB, but neither test had good sensitivity at predicting women at high risk of PTB. The authors concluded that the PMN/EPI ratio provided internal standardization of variation in slide cellular density and was significantly associated with subsequent spontaneous preterm birth at less than 35 weeks gestation.

Both studies suggest that vaginal flora and leukocyte counts can be assessed on the same vaginal smear. The Culhane study indicates that BV diagnosed by Nugent score does not vary by site of vaginal sample collection, but the vaginal leukocyte count does. Consequently they suggest that self-taken vaginal smears are appropriate for assessing vaginal flora but are not appropriate for measuring vaginal leukocyte counts. Ramsey and colleagues highlight the potential inconsistencies in counting PMNs on vaginal smears. They measured the ratio of PMNs to epithelial cells and showed improved reliability and prediction of women at risk of PTB. It would be interesting to know whether the significant difference in leukocyte count at different parts of the vagina remains if the PMN/EPI ratio is used rather than absolute leukocyte counts.

Further research is needed to help understand the clinical significance of higher numbers of leukocytes in the vagina of pregnant women but evaluating a Gram stained vaginal smear for both BV and leukocytes is a simple and cheap test. Currently it is suggested that the use of a combination of markers may improve the sensitivity and specificity of predicting preterm birth [10,11]. The Gram-stained vaginal smear, assessed for both vaginal flora and leukocyte count or ratio, may well have a part to play in this.

References:

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