Recent evidence suggests that smoking affects the composition of the disease-associated subgingival biofilm, yet little is known about its effects during the formation of this biofilm. The present investigation was undertaken to examine the contributions of smoking to the composition and proinflammatory characteristics of the biofilm during de novo plaque formation. Marginal and subgingival plaque and gingival crevicular fluid samples were collected from 15 current smokers and from 15 individuals who had never smoked (nonsmokers) following 1, 2, 4, and 7 days of undisturbed plaque formation. 16S rRNA gene cloning and sequencing were used for bacterial identification, and multiplex bead-based flow cytometry was used to quantify the levels of 27 immune mediators. Smokers demonstrated a highly diverse, relatively unstable initial colonization of both marginal and subgingival biofilms, with lower niche saturation than that seen in nonsmokers. Periodontal pathogens belonging to the genera Fusobacterium , Cardiobacterium , Synergistes , and Selenomonas , as well as respiratory pathogens belonging to the genera Haemophilus and Pseudomonas , colonized the early biofilms of smokers and continued to persist over the observation period, suggesting that smoking favors early acquisition and colonization of pathogens in oral biofilms. Smokers also demonstrated an early proinflammatory response to this colonization, which persisted over 7 days. Further, a positive correlation between proinflammatory cytokine levels and commensal bacteria was observed in smokers but not in nonsmokers. Taken together, the data suggest that smoking influences both the composition of the nascent biofilm and the host response to this colonization.

Oral infections have a strong ethnic predilection; suggesting that ethnicity is a critical determinant of oral microbial colonization. Dental plaque and saliva samples from 192 subjects belonging to four major ethnicities in the United States were analyzed using terminal restriction fragment length polymorphism (t-RFLP) and 16S pyrosequencing. Ethnicity-specific clustering of microbial communities was apparent in saliva and subgingival biofilms, and a machine-learning classifier was capable of identifying an individual’s ethnicity from subgingival microbial signatures. The classifier identified African Americans with a 100% sensitivity and 74% specificity and Caucasians with a 50% sensitivity and 91% specificity. The data demonstrates a significant association between ethnic affiliation and the composition of the oral microbiome; to the extent that these microbial signatures appear to be capable of discriminating between ethnicities.

Although smoking and diabetes have been established as the only two risk factors for periodontitis, their individual and synergistic impacts on the periodontal microbiome are not well studied. The present investigation analyzed 2.7 million 16S sequences from 175 non-smoking normoglycemic individuals (controls), smokers, diabetics and diabetic smokers with periodontitis as well as periodontally healthy controls, smokers and diabetics to assess subgingival bacterial biodiversity and co-occurrence patterns. The microbial signatures of periodontally healthy smokers, but not diabetics, were highly aligned with the disease-associated microbiomes of their respective cohorts. Diabetics were dominated by species belonging to Fusobacterium, Parvimonas, Peptostreptococcus, Gemella, Streptococcus, Leptotrichia, Filifactor, Veillonella, TM7 and Terrahemophilus. These microbiomes exhibited significant clustering based on HbA1c levels (pre-diabetic (10%). Smokers with periodontitis evidenced a robust core microbiome (species identified in at least 80% of individuals) dominated by anaerobes, with inter-individual differences attributable largely to the ‘rare biosphere’. Diabetics and diabetic smokers, on the other hand, were microbially heterogeneous and enriched for facultative species. In smokers, microbial co-occurrence networks were sparse and predominantly congeneric, while robust inter-generic networks were observed in diabetics and diabetic smokers. Smoking and hyperglycemia impact the subgingival microbiome in distinct ways, and when these perturbations intersect, their synergistic effect is greater than what would be expected from the sum of each effect separately. Thus, this study underscores the importance of early intervention strategies in maintaining health-compatible microbiomes in high-risk individuals, as well as the need to personalize these interventions based on the environmental perturbation.

Background: Changes in clinical profiles, microbial succession, and immune mediator fluctuations have all been separately examined during onset and resolution of experimental gingivitis in smokers. However, because both the bacterial challenge and the host response contribute to periodontal disease, the purpose of this investigation is to simultaneously examine clinical, bacterial, and immune changes that occur during the onset and resolution of disease in smokers.Methods: Experimental gingivitis was induced in 15 smokers for 21 days, followed by treatment with a sonic toothbrush for 21 days. Marginal and subgingival plaque and gingival crevicular fluid samples were collected at baseline; after 7, 14, and 21 days of undisturbed plaque formation; and 21 days after reinstitution of brushing. 16S cloning and sequencing was used for bacterial quantification, and multiplexed bead‐based flow cytometry was used to quantify the levels of 27 immune mediators.Results: Onset of clinical gingivitis was preceded by significant changes in the marginal and subgingival biofilms, with a decrease in the abundance of early colonizers, namely, Streptococcus, Veillonella, and Pseudomonas, and an increase in levels of periodontopathogens, such as Treponema, Selenomonas, Parvimonas, Dialister, and Campylobacter. This was accompanied by a decrease in anti‐inflammatory, chemokine, and T‐helper 2 (Th2) responses and altered Th1/Th2 ratios. Although the bacterial communities continued to shift in the same direction after onset of clinical gingivitis and returned to baseline levels after resolution of disease, the anti‐inflammatory, chemokine, and Th2 profiles demonstrated an increase from day 14 that continued even after clinical health was evident.Conclusion: Both marginal and subgingival biofilms in smokers are characterized by early acquisition of pathogenic organisms, which elicit a sustained host response that persists even after removal of the bacterial challenge.