Preterm birth complicates 12% of all pregnancies in the United States and is one of the leading causes of infant morbidity and mortality.1–2 Evidence indicates that maternal infection and inflammation of the lower and upper genital tract, as well as at sites distant from the pelvis, play a major role in the etiology of preterm birth in some women.3–5
Periodontal disease is a chronic anaerobic inflammatory condition that affects as many as 50% of pregnant women in the United States.6–8 Emerging evidence supports a relationship between periodontal disease and other systemic conditions, including cardiovascular disease and diabetes mellitus.9–11 Recent studies in the United States have demonstrated an association between maternal periodontal disease and multiple adverse pregnancy outcomes, including preterm birth, low birth weight, fetal growth restriction, preeclampsia, and perinatal mortality.6–8,12–16 The mechanisms by which periodontal disease and preterm birth are associated are not clear. It has been hypothesized that in the presence of severe periodontal disease, oral organisms can disseminate hematogenously to target the placenta, membranes, and fetus.17–18 This bacterial challenge may result in increased cytokine expression and precipitate preterm labor.
We sought to determine if women who have a spontaneous preterm birth at less than 32 weeks of gestation are more likely to have periodontal disease than women who have either an indicated preterm birth at less than 32 weeks of gestation or a term birth. In addition, we sought to determine whether selected markers of upper genital tract inflammation were associated with periodontal disease in these women.
MATERIALS AND METHODS
This case-control study was approved by the Institutional Review Board at the University of Alabama at Birmingham. Women delivering at our institution at between 24 0/7 and 31 6/7 weeks of gestation, as well as women following spontaneous vaginal delivery at term (> 37 weeks) who were enrolled in an ongoing study of risk factors for preterm birth at our institution (Perinatal Emphasis Research Center at the University of Alabama at Birmingham) were eligible for the study. The study population for this periodontal disease supplement to Perinatal Emphasis Research Center was a sample of convenience, in that women were recruited only when delivered during daytime business hours on weekdays. Spontaneous preterm birth was defined as delivery following spontaneous preterm labor or spontaneous preterm premature rupture of membranes. Indicated preterm birth was defined as delivery for maternal or fetal indications excluding chorioamnionitis. All patients enrolled in this periodontal disease supplement to the Perinatal Emphasis Research Center provided informed consent before participation.
Study participants underwent a dental examination by a periodontist from the School of Dentistry within 72 hours postpartum. Eighteen of the women had a dental examination performed at 24 weeks of gestation as part of the larger Perinatal Emphasis Research Center observational study, and the dental examination was not repeated at the postpartum visit for these women. The dental examination technique used was a modification of the Community Periodontal and Treatment Index, also known in the United States as the Periodontal Screening and Recording Index.19 Briefly, the oral cavity and dentition were divided into 6 standard regions or sextants. At least 6 areas of each tooth were examined using a periodontal probe. The highest score in millimeters for any tooth in each sextant was recorded for probing depth and clinical attachment level. To perform the dental examination, a 0.5-mm diameter probe is slipped between the gingiva and the tooth until resistance is met by the probe. The distance from the cemento-enamel junction and pocket base was defined as attachment loss. Any evidence of gingival inflammation, defined as bleeding on periodontal probing of any tooth, was recorded for each sextant. Periodontal health was defined as no evidence of attachment loss or gingival inflammation. Periodontal disease was defined as 1) gingival inflammation and no attachment loss (gingivitis); 2) 3–5 mm (mild periodontitis); or 3) > 5 mm (severe periodontal disease) in any one sextant. The extent of periodontal disease was classified using an index. This Extent Index describes the proportion of sextants with a threshold attachment level (ie, the Extent 5 Index is the proportion of sextants with an attachment loss of > 5 mm).20 In reporting the index, the denominator is sextants in any group of women, not an individual score for each woman.
At the time of delivery, the placenta and a sample of neonatal umbilical cord blood were collected using aseptic technique, refrigerated, and processed for culture within 12 hours of delivery. Placental culture included swabs and placental tissue for aerobic and anaerobic culture and culture for Mycoplasma species and Ureaplasma urealyticum obtained using aseptic technique. The swabs were taken between the chorion and amnion in 3 separate sites. The placental tissue was taken just beneath the chorion. All placentas were then placed in formalin and underwent histologic examination by a single pathologist. Umbilical cord blood was cultured for Mycoplasma sp and U urealyticum, and the remaining fetal cord serum was aliquoted and stored at −70°C for batched analysis. Interleukin-6 (IL-6) levels in the cord serum were determined by commercially available enzyme-linked immunosorbent assay (ELISA) kits (R&D Systems, Inc, Minneapolis, MN). The lower limit of sensitivity of the assay was 0.7 pg/mL. The intra-assay and interassay coefficients of variation were 2.6% and 4.5%, respectively. Concentrations of cord IL-6 greater than or equal to the 95th percentile for the combined indicated preterm and term control groups of the larger Perinatal Emphasis Research Center study were considered elevated. The periodontists, laboratory personnel, and pathologist were blinded to delivery outcome.
Maternal demographic, intrapartum, delivery, and postpartum information was recorded by a trained obstetric research nurse. Neonatal outcome data through hospital discharge or death was also recorded. Neonatal systemic inflammatory response syndrome was defined as the presence of negative cerebrospinal fluid and blood cultures plus clinically suspected sepsis and/or a band/band plus polymorphonuclear cell ratio of 0.15 or greater. Composite neonatal morbidity was defined as any one or more of the following complications: neonatal systemic inflammatory response syndrome, intraventricular hemorrhage (grades 3 or 4), periventricular leukomalacia, respiratory distress syndrome, bronchopulmonary dysplasia, necrotizing enterocolitis, and/or neonatal death.
Based on prior studies in our pregnant patient population,6 we estimated the baseline rate of severe periodontal disease to be 20%. Using a 2-sided test of significance with an α level of 0.05 and power of 80%, we estimated needing 45 cases and 45 term controls, assuming a 50% exposure rate in cases versus a 20% exposure rate in controls (an odds ratio of 4.0). However, because spontaneous preterm deliveries accounted for two thirds of the preterm births at our institution, we calculated the sample size for the preterm birth groups using a 2:1 ratio of cases to controls. We estimated needing 57 spontaneous cases versus 38 indicated controls for the same comparison as described for the term control group. We therefore attempted to recruit 57 cases of spontaneous preterm birth, 38 indicated preterm controls, and 45 term controls for our study.
Statistical analyses were performed using SAS 8.2 software (SAS Institute, Inc, Cary, NC). Analyses were performed for cases and the 2 separate control groups, not across all 3 groups. Unadjusted odds ratios (ORs) and their 95% confidence intervals (CIs) were calculated separately for spontaneous cases compared with indicated and term controls. For demographic characteristics, groups were compared using χ2 tests or Fisher exact test, where appropriate. Means of continuous variables were compared using Student t test. Distribution of IL-6 between groups was compared using a Wilcoxon rank sum test. Logistic regression analysis, adjusting for possible confounding factors, was used to calculate adjusted ORs for spontaneous preterm birth.
From October 1998 through May 2001, 139 maternal-infant pairs were enrolled: 59 following a spontaneous preterm birth, 36 following an indicated preterm birth, and 44 following uncomplicated spontaneous labor at term. The indicated preterm births included 31 (86%) for severe preeclampsia and 5 (14%) for nonreassuring fetal status. There were no significant differences in birth group among the 18 women with a periodontal examination at 24 weeks (n = 10 term, n = 6 spontaneous preterm, n = 2 indicated preterm; P = .053). Selected maternal demographics are presented in Table 1. Most of the differences noted among the birth groups reflect the demographics of our patient population. Our local patient population (those receiving prenatal care and delivering in our hospital) is predominantly African American, funded by Medicaid, and younger, whereas maternal transfers (from across the state with an expectant preterm delivery) are more likely to be older, have private insurance, and a higher percentage of white women.
We evaluated the potential association between periodontal disease and spontaneous preterm birth in several ways. The extent or the number of regions (sextants) with mild-to-severe attachment loss is reported using the Extent Index.20 With a threshold attachment loss of more than 3 mm and more than 5 mm, the Extent 3 and 5 Index for each study group are presented in Table 2. Using either threshold for periodontal disease, the spontaneous preterm birth group had significantly more extensive periodontal disease than the term birth group, ie, more areas of the mouth were affected. The spontaneous preterm birth group also had more extensive periodontal disease than the indicated preterm birth group, but the more than 5 mm threshold was not statistically significant.
We also evaluated whether women had evidence of severe disease in at least one area of the mouth in relation to birth group. None of the women had completely healthy gums. Women with a spontaneous preterm birth at less than 32 weeks of gestation had higher rates of severe periodontal disease than both the indicated preterm birth or term birth groups (Table 3). Multivariable analyses were performed, controlling for maternal age, race, education, insurance status, parity, history of a spontaneous preterm birth, and smoking. The unadjusted and adjusted ORs for spontaneous preterm birth in women with periodontal disease compared with women without severe periodontal disease are presented in Table 4. Each regression model included the spontaneous preterm birth group as well as one or both of the control groups. Severe periodontal disease was associated with an almost 3-fold risk for spontaneous preterm birth compared with the combined controls, indicated preterm birth, and term birth (OR 2.6, 95% CI 1.1–6.2). Because periodontal disease may be in the causal pathway in each of a woman's pregnancies, we repeated the logistic regression adjusting for the same factors but excluding history of preterm birth. Again, severe periodontal disease was associated with a 3-fold risk for spontaneous preterm birth (OR 3.4, 95% CI 1.5–7.7; Table 4).
The association between markers of upper genital tract inflammation at the time of delivery and periodontal disease is presented in Table 5. Women with and without severe periodontal disease had similar rates of positive placental cultures, positive umbilical cord cultures for U urealyticum and Mycoplasma species, histologic chorioamnionitis, funisitis, median cord IL-6 level, and an elevated umbilical cord IL-6 level (≥ 95% or 41.2 pg/mL). These findings were true in the overall population as well as within each birth group (Table 5, data for indicated preterm and term birth not shown). Using the number of participants in the overall population with and without periodontal disease (n = 51 and n = 88, respectively), we determined that we had 80% power (α = 0.05) to detect a significant difference between groups (baseline rate of histological chorioamnionitis of 50% in the group with periodontal disease, compared with 25% in the group without periodontal disease). Organisms that are known periodontal pathogens were identified by placental culture in only 5 of the women (2 women with spontaneous preterm birth and 3 women with a term birth) and included Peptostreptococcus micros, Streptococcus constellatus, and Eubacterium species.21
None of the infants delivered at term had any of the adverse neonatal outcomes assessed in this study. Neonatal outcome and presence or absence of severe periodontal disease in the preterm population (spontaneous and indicated) are presented in Table 6. All of the adverse neonatal outcomes evaluated were represented similarly among women with and without periodontal disease.
The results of our study demonstrate an association between severe periodontal disease and spontaneous preterm birth at less than 32 weeks of gestation. Others have shown an association between periodontal disease and preterm birth.6–8,12–16 In a case-control study of 124 women, Offenbacher and colleagues12 found that preterm low birth weight cases had significantly worse periodontal disease than control women with normal birth weight infants. Cases were defined as women with a current or previous history of a birth in which the infant (< 2,500 g) was delivered after preterm labor or premature rupture of membranes. In their multivariable analysis in that study, the OR for periodontal disease in cases was 7.5 (95% CI 1.95–28.8). These investigators also demonstrated a significant association between periodontal disease and preterm birth in a larger prospective study (n = 814) in which 9% of women delivering at less than 37 weeks and 13.6% of women at less than 32 weeks of gestation had moderate-severe periodontal disease, compared with 4.5% of women delivering at term.8 In a large prospective cohort study of 1,313 women at our institution, Jeffcoat and colleagues6 showed that women with severe periodontal disease at 21–24 weeks of gestation had a higher OR for subsequent spontaneous preterm birth at less than 37 weeks (adjusted OR 4.5, 95% CI 2.2–9.2) and less than 32 weeks of gestation (adjusted OR 7.1, 95% CI 1.7–27.4). Our results are consistent with these findings, with the added strength of comparing well-defined cases of spontaneous preterm birth at less than 32 weeks of gestation and 2 control groups: one of preterm birth following maternal or fetal indications and one of term birth.
The mechanisms by which periodontal disease and preterm birth are associated are not clear. It has been hypothesized that, in the presence of severe periodontal disease, cytokines produced by the infected periodontium may appear in the systemic circulation and target the placenta and fetus, precipitating preterm labor. Alternatively, oral organisms can disseminate hematogenously to target the placenta, membranes, and fetus. This infectious challenge may result in increased cytokine expression and precipitate preterm labor.17–18 In the prospective study by Offenbacher and colleagues8 described above, maternal immunoglobulin (Ig)G and fetal IgM directed against known periodontal pathogens were measured at the time of delivery.22 The highest risk for preterm birth was observed among mothers without evidence of a protective IgG response combined with a fetus with an IgM response (OR 10.3, P < .001).22 These data would suggest that when the fetal-placental unit is exposed to the periodontal pathogens in the absence of maternal antibody protection, this may precipitate preterm birth.
Previous reports have documented a consistent relationship between upper genital tract infection/inflammation and preterm birth, particularly early spontaneous preterm birth.3–5 Previous studies have not evaluated the presence of upper genital tract inflammation at the time of delivery and the potential association with periodontal disease. In our study, we could not demonstrate any association between markers of upper genital tract inflammation at the time of birth and moderate-to-severe periodontal disease in cases of spontaneous preterm birth, indicated preterm birth, or term controls. Our data would suggest that the potential mechanism of periodontal disease–associated preterm birth is not hematogenous spread of the periodontal organisms to the placenta with colonization and subsequent inflammation precipitating preterm labor or premature rupture of membranes.
Periodontal pathogenic organisms may be more difficult to culture from the placenta. However, we used meticulous methods and were able to isolate multiple species of anaerobic and Gram-negative organisms, including several known periodontal pathogens. Perhaps the relative lack of growth of periodontal organisms represents sampling error. However, we sampled multiple sites of the chorioamnion, as well as placental tissue, for culture in these women. It is possible that the organisms do spread to the placenta but are not viable and do not colonize the placenta and membranes. Perhaps more sensitive tests, such as DNA tests for periodontal pathogens, may demonstrate that the presence of the organisms and the antigenic challenge of these few bacteria is enough to precipitate preterm labor. Regardless, the inflammation that occurs is not sufficient to be demonstrated by evidence of higher rates of histological chorioamnionitis or elevated umbilical cord levels of the proinflammatory cytokine, IL-6. Another possibility to consider is that periodontal disease and spontaneous preterm birth are not, in fact, directly associated in a cause-and-effect manner. Rather, these 2 conditions may be linked by a third factor that predisposes the pregnant woman to both adverse outcomes: periodontal disease and spontaneous preterm birth.
Finally, previous studies of inflammation and preterm birth have demonstrated an increased risk for neonatal morbidity, especially neurological morbidity, in the setting of intrauterine infection and inflammation.23–28 We did not see an association between periodontal disease and adverse neonatal outcome in the preterm infants in our study. This further suggests that, if the preterm birth associated with periodontal disease is inflammation-mediated, the levels of inflammation are not of the same magnitude as those seen in infection-mediated preterm birth previously studied. Further investigation into the potential etiology of the association between periodontal disease and preterm birth is warranted.
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© 2004 by The American College of Obstetricians and Gynecologists.
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