The Subgingival Microbial Flora During Pregnancy: a Proposed Interaction Between Endogenous Steroids and Bacteroides Melaninogenicus.

Abstract

The subgingival bacterial flora from 2 gingival sites was cultured and characterized monthly in twenty periodontitis-free women during pregnancy and again post-partum. Monthly plaque samples were also cultured in eleven age and disease matched non-pregnant women. Plaque was processed anaerobically on selective and non-selective media and the predominant colony types were characterized. A portion of each plaque sample was tested for bacterial uptake of C('14)-estradiol and C('14)-progesterone. Plasma levels of estrogens and progesterone were measured four times in each subject. The number of gingival bleeding sites, the Gingival Index and the Plaque Index were determined at each sampling period. In the second trimester there was significant increase in gingivitis, the ratio of anaerobic to aerobic bacteria, and the proportional levels of Bacteroides melaninogenicus ss. intermedius. In the third trimester both gingivitis and the levels of B. melaninogenicus ss. intermedius decreased. Plaque uptake of C('14)-steroids increased significantly during pregnancy and paralleled the plaque levels of B. melaninogenicus ss. intermedius. In the second trimester, recovery of B. melaninogenicus ss. intermedius was strongly correlated with plasma levels of estrogens and progesterone. No changes were observed in clinical parameters or the subgingival flora of non-pregnant subjects. The increase in plaque uptake of C('14)-steroids in conjunction with an increase in B. melaninogenicus ss. intermedius in the second trimester suggested the possibility of direct steroid bacterial interaction. Pure cultures of B. melaninogenicus ss. asaccharolyticus took up C('14)-estradiol and C('14)-progesterone in the ranges of 0.00 to 0.71 x 10('-4) (mu)m/(mu)g bacterial protein and 0.41 to 1.76 x 10('-4) (mu)m/(mu)g protein respectively. This uptake was less than that of the intermedius and melaninogenicus subspecies which had mean uptakes of estradiol and progesterone which ranged from 2.15 to 5.48 x 10('-4) (mu)m/(mu)g protein and 2.41 to 4.12 x 10('-4) (mu)m/(mu)g protein respectively. The cell uptake of steroids was found to be temperature dependent. Thin-layer chromatography of the reaction mixtures from steroid uptake studies with B. melaninogenicus ss. intermedius revealed a labelled spot which did not correspond to st and ards for progesterone and estradiol. Structural similarities between vitamin K, an essential growth factor for B. melaninogenicus, and the steroids led to an attempt to substitute progesterone and estradiol for vitamin K in the growth of these organisms. All tested strains of B. melaninogenicus ss. intermedius and ss. assaccharolyticus required menadione for growth and growth of one strain of ss. melaninogenicus was stimulated by menadione. Progesterone and estradiol in one, two and three times the molar concentration of menadione failed to support growth of B. melaninogenicus ss. asaccharolyticus. Both progesterone and estradiol supported growth of intermedius and melaninogenicus subspecies to the same extent as menadione. Estradiol appeared to exhibit substrate inhibition at higher concentrations. Since vitamin K is thought to mediate fumarate reduction to succinate, fumarate was added to the assay for uptake of C('14)-steroids. The addition of fumarate increased the uptake of estradiol and progesterone by all three subspecies. The conversion of fumarate to succinate by B. melaninogenicus in the steroid uptake system was demonstrated by gas-liquid chromatography and was shown to require the presence of steroids. The identification of the potential for endogenous steroids to induce a shift in the normal microflora may provide new insights to the association of periodontal disease with conditions characterized by altered steroid levels.