Literature review > Issue 8 > Review on Larsson et al. 

The Nugent classification system [1] is often used for diagnosing bacterial vaginosis (BV) using Gram stained smears of vaginal fluid. In this system, individuals are classified as normal, intermediate or BV depending on the number of different bacterial morphotypes counted in one or more microscopic fields using 1000x (oil immersion) magnification. The authors point out, however, that the actual field diameter (and, hence, area) varies from microscope to microscope and that no standard field size is specified in the Nugent system. Clearly, the field size can influence the Nugent score since more bacteria will typically be seen on a large field compared to a small field. This lack of standardization could conceivably make it difficult to compare the Nugent score readings when different microscopes are used.

To address this problem, the authors developed a straightforward approach for scaling the cutpoints in the Nugent scoring system according to the microscope field area and propose that the Zeiss FL30 (field diameter 0.145 mm) be used as the standard. Using this revised system, they reanalyzed data from two studies of BV. In both studies only a few samples originally classified as normal or BV were reclassified. However, among samples originally classified as intermediate, 24% and 39% were reclassified to BV or normal, respectively. The majority were reclassified as normal, likely because the field size of the microscopes used in these studies was larger than that of the Zeiss FL30, thus allowing for more opportunity to visualize lactobacilli morphotypes. The revised Nugent classification was found to have good agreement (kappas of 0.88 and 0.90) with the Ison/Hay [2,3] classification of the same slides.

Clearly, improved standardization of any diagnostic technique is welcome, and it is gratifying that the more one standardizes the reading field and ensures that a representative area is reviewed, the more ‘definitive’ the diagnosis becomes. In this case, fewer samples were classified as intermediate. These results have obvious implications for the use of Gram stain to define vaginal flora in both screening and therapeutic studies. In addition, the implications of these findings should be reconsidered in light of new developments in molecular identification of the bacteria associated with BV. In particular, although some of these newly described BV-associated bacteria are not cultivatable and thus cannot be definitively characterized by Gram stain, they strongly resemble Mobiluncus morphotypes when visualized by fluorescent in situ hybridization techniques. Thus, the implications for the performance of Nugent criteria for defining BV will need to be reassessed using the presence of BV-associated bacteria as defined not only by traditional culture but by molecular detection methods as well.

In the meantime, the use of the standardized Nugent scoring system described by Larsson et al. should help to standardize readings reported by different investigators using different microscopes, and should help to standardize results across studies of BV epidemiology and therapeutic trials.

References:

1. Nugent RP, Krohn MA, Hillier SH. Reliability of diagnosing bacterial vaginosis is improved by a standardized method of Gram stain interpretation. J Clin Microbiol 1991;29:297–301.

2. Hay PE, Lamont RF, Taylor-Robinson D, et al. Abnormal bacterial colonization of the genital tract and subsequent preterm delivery and late miscarriage. BMJ 1994;308:295–8.

3. Ison CA, Hay PE. Validation of a simplified grading of Gram stained vaginal smears for use in genitourinary medicine clinics. Sex Transm Infect 2002;78:413–15.

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