Validation of a simplified grading of Gram stained vaginal smears for use in genitourinary medicine clinics

Abstract

Objectives: To validate a simplified grading scheme for Gram stained smears of vaginal fluid for the diagnosis of bacterial vaginosis (BV) against the accepted "gold" standard of Amsel's composite criteria. Methods: Women attending genitourinary medicine (GUM) clinics, as part of a multicentre study, were diagnosed as having BV if three or more of the following criteria were present; homogeneous discharge, elevated vaginal pH, production of amines, and presence of "clue" cells. Women with less than three of the criteria were considered as normal. Simultaneously, smears were made of vaginal fluid and Gram stained and then assessed qualitatively as normal (grade I), intermediate (grade II), or consistent with BV (grade III). Two new grades were used, grade 0, epithelial cells only with no bacteria, and grade IV, Gram positive cocci only. Results: BV was diagnosed in 83/162 patient visits using the composite criteria, the remainder being regarded as normal. The majority of patients with BV had a smear assessed as grade III (80/83, 96%) and the majority of normal women had a smear assessed as grade I (normal, 48/79, 61%), giving a high sensitivity (97.5%), specificity (96%), and predictive value for a positive (94.1%) and negative (96%) test, kappa index = 0.91. Smears assessed as grade II were found predominantly (12/13) among patients diagnosed as normal, with less than three of the composite criteria. Grades 0 and IV were both only found among normal women. Conclusion: This simplified assessment of Gram stained smears can be used as an alternative to Amsel's criteria and is more applicable for use in busy GUM clinics. Bacterial vaginosis (BV) is a clinical entity that is characterised by a change in vaginal ecology where the normal flora of lactobacillus morphotypes is replaced by a mixed microbial flora consisting of anaerobes and Gardnerella vaginalis.1 The original description in 1955 by Gardner and Dukes 2 remains an accurate description of the presenting clinical features of a malodorous, thin homogeneous vaginal discharge. However, a variety of methods were used for diagnosis until 1984 when a consensus was reached to define the diagnosis of BV using the composite criteria described by Amsel et al. 3 These included a thin homogeneous discharge, elevated vaginal pH above 4.5, release of amines on the addition of 10% potassium hydroxide to vaginal fluid, and the presence of "clue" cells, of which three need to be present for the diagnosis of BV.An alternative method of diagnosis that has been used extensively, particularly in research studies, is the grading (or scoring) of the microbial flora in Gram stained smears of vaginal fluid. This method reflects both the change in vaginal ecology and the strong microbial associations, and was first described by Spiegel et al 4 in 1983. This initial report divided smears into those with normal lactobacillus morphotype flora and those with mixed flora consistent with BV. The method was modified by Nugent et al 5 to include an intermediate category that demonstrated...

Treatment of Bacterial Vaginosis in Early Pregnancy and its Effect on Spontaneous Preterm Delivery and Preterm Premature Rupture of Membranes

“…The vaginal swab sample was transferred to a microscope slide, air dried, and transported to the gynaecological clinic in Skaraborgs Hospital where it was microscopically examined using a phase contrast microscope with a magnification of 400x, according to the modified Hay/Ison criteria for the diagnosis of BV [9]. The Hay/ Ison criteria were developed using Gram-stained smears by oil immersion at a magnification of 1000x; however, because this criteria assesses the type of flora and does not indicate the number of individual bacteria, which is achieved using Nugent's criteria, it is possible to use these criteria with non-stained smears at lower magnification [5].…”

Comparison between Nugent's and Hay/Ison scoring criteria for the diagnosis of Bacterial Vaginosis in WASP prepared vaginal samples

“…Therefore, in 2002, Hay and Ison described an easier score, which consists of the characterization of the vaginal flora into three different categories: Grade I (normal), Grade II (intermediate) and Grade III (BV) [11]. The assessment of the Research Paper Antonucci, Mirandola, Fontana…”

Normal flora and bacterial vaginosis in pregnancy: an overview

“…Instead of grading and allocating a score to individual bacterial species, the Ison and Hay grading system assigns a grade to a mixed group of bacteria, depending on the numerical contribution of individual morphotypes (Ison and Hay, 2002). Where only Lactobacillus morphotypes (normal flora) are present, a smear will be graded as grade I. Grade II comprises intermediate flora, which include reduced Lactobacillus morphotypes with diverse bacterial morphotypes, whereas grade III (BV) contains mixed bacterial morphotypes with few or no Lactobacillus morphotypes (Ison and Hay, 2002).…”

“…Grade 0 will be smears that contain epithelial cells with no bacteria and in which case antibacterial agents in the vagina might be present (Ison and Hay, 2002). Grade IV contains epithelial cells enclosed in Grampositive cocci only (Ison and Hay, 2002 were categorized as grade Ib and specimens containing both L. crispatus and other lactobacilli were categorized as grade Iab (Verhelst et al, 2005). The grade I-like category contains short Gram-positive rods that are unevenly shaped with curved ends and may appear as Chinese letters (diphtheroid cell types) (Verhelst et al, 2005).…”

“…Where only Lactobacillus morphotypes (normal flora) are present, a smear will be graded as grade I. Grade II comprises intermediate flora, which include reduced Lactobacillus morphotypes with diverse bacterial morphotypes, whereas grade III (BV) contains mixed bacterial morphotypes with few or no Lactobacillus morphotypes (Ison and Hay, 2002). Grade 0 will be smears that contain epithelial cells with no bacteria and in which case antibacterial agents in the vagina might be present (Ison and Hay, 2002). Grade IV contains epithelial cells enclosed in Grampositive cocci only (Ison and Hay, 2002 were categorized as grade Ib and specimens containing both L. crispatus and other lactobacilli were categorized as grade Iab (Verhelst et al, 2005).…”