Metronidazole in the treatment of localized juvenile periodontitis

Abstract Systemic metronidazole and tetracycline were compared as adjunctive agents in the treatment of localized juvenile periodontitis (LJP). 27 patients with Actinobacillus actinomycetemcomitans-positive (Aa) LJP were treated with scaling and rootplaning, control of oral hygiene and periodontal surgery if indicated. The patients were randomly divided into 3 equal groups: the 1st group had metronidazole 200 mg × 3 × 10 days, the 2nd tetracycline 250 mg × 4 × 12 days, the 3rd group received no medication and served as a control. 6 patients had periodontal surgery. 4 sites with the most advanced bone loss as determined on radiographs were selected in each subject for test sites. Gingival index, gingival bleeding after probing (GB), probing depth (PD), suppuration, and radiographic bone loss were registered, and subgingival Aa was selectively cultured. GB and PD>4 mm were registered in the whole dentition as well. All parameters were monitored at baseline and at 6 and 18 months after treatment. By the end of the study, Aa was suppressed to below detection level at all test sites only in the metronidazole group, at 17/26 sites (4 patients) in the tetracycline group and at 19/26 sites (6 patients) in the control group. Clinically, all groups showed improvement. In conclusion, metronidazole was more effective than tetracycline in the suppression of Aa and the suppression of Aa appeared to produce better clinical results.     

Prognostic criteria for the efficiency of non-surgical periodontal therapy in advanced periodontitis

Abstract The aim of the study was to find out which clinical, radiographic and microbiological variables can be used as prognostic criteria for the efficiency of the commonly used initial treatment protocol comprising scaling, root planing and instruction on oral hygiene in advanced adult periodontitis. 46 patients (mean age 48 years) with untreated, advanced periodontitis volunteered for the study. The clinical examination included recordings of plaque, gingival and calculus indices, probing pocket depths, bleeding and suppuration after probing, probing attachment levels and furcation involvements. Infrabony and furcation lesions were assessed from panoramic radiographs. Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis were cultured from the deepest, most inflamed periodontal pockets, from surface of the tongue and from saliva. 3 months after the completion of non-surgical treatment comprising meticulous scaling and root planing and instruction on oral hygiene, the healing was assessed clinically, and 13 patients were assigned to a maintenance care programme (MC) and 33 to further treatment procedures (FT). Evaluation of the baseline clinical and radiographic data showed a significantly higher %s of 6 mm deep periodontal pockets, surfaces with suppuration, and sites with subgingival calculus, as well as higher numbers of infrabony lesions, in FT-patients than in MC-patients. Subgingival A. actinomycetemcomitans was isolated at baseline in 55% of the FT-patients and in 38% of the MC-patients, and P. gingivalis in 27% and 23%, respectively. A. actinomycetemcomitans was eradicated by non-surgical treatment from only one patient. P. gingivalis was detected in 15% of the patients in both groups after treatment. Combining of the clinical and the microbiological baseline data demonstrated that simultaneous presence of multiple deep periodontal pockets (6 mm at 10% of the sites) and subgingival A. actinomycetemcomitans alone or concomitant with P. gingivalis was significantly more prevalent in the FT-group than in the MC-group. This simultaneous presence anticipated insufficient response to initial non-surgical therapy. The results suggest that a treatment regimen more efficient than the traditional one is needed without delay in the treatment of A. actinomycetemcomitans infected generalized, severe periodontal disease in adults.     

Microbiological and clinical effects of surgical treatment of localized juvenile periodontitis

Since Actinobacillus actinomycetemcomitans appears to be a key etiologic agent in localized juvenile periodontitis, this study determined the effectiveness of different treatment modalities in suppressing A. actinomycetemcomitans in localized juvenile periodontitis lesions. A total of 25 deep periodontal lesions from 7 patients with localized juvenile periodontitis were included in the study. The test periodontal lesions either received scaling and root planing alone, scaling and root planing together with soft tissue curettage, or modified Widman flap surgery. Subgingival A. actinomycetemcomitans were enumerated using selective culturing. Clinical measurements included changes in probing periodontal attachment level, probing periodontal pocket depth, gingival index, plaque index, and digital subtraction of standardized serial radiographs. The microbiological and clinical effects of treatment were monitored over a period of 16 weeks. All periodontal lesions studied demonstrated high numbers of A. actinomycetemcomitans prior to treatment. Scaling and root planing alone did not markedly change the subgingival A. actinomycetemcomitans counts, nor any of the clinical parameters studied. In contrast, soft tissue curettage as well as modified Widman flap surgery suppressed A. actinomycetemcomitans to undetectable levels immediately after therapy in more than 80% of the lesions studied. A total of 5 periodontal lesions exhibited gain of probing periodontal attachment after subgingival curettage or Widman flap treatment; 3 of these sites revealed no detectable A. actinomycetemcomitans, and the remaining 2 sites harbored only low levels of A. actinomycetemcomitans. 5 periodontal lesions which lost probing attachment after treatment all demonstrated high numbers of subgingival A. actinomycetemcomitans. Changes in alveolar bone, assessed by digital subtraction of serial radiographs, correlated with changes in probing periodontal attachment level, confirming the clinical results. The present study revealed a close relationship between post-treatment A. actinomycetemcomitans levels and the clinical response to treatment, which supports the concept that A. actinomycetemcomitans is an important organism in the etiology of localized juvenile periodontitis. This study also showed that a substantial suppression of subgingival A. actinomycetemcomitans cannot be achieved by periodontal scaling and root planing alone, but can be accomplished by surgical removal of periodontal tissues.(ABSTRACT TRUNCATED AT 400 WORDS)     

Subgingival distribution of A. actinomycetemcomitans in periodontitis

Abstract This investigation developed an experimental design that (1) detailed the distribution of A. actinomycetemcomitans in Subgingival plaque related to the level of serum antibody to this pathogen; (2) used broad based subgingival plaque sampling to allow a definition of the distribution of A. actinomycetemcomitans infection in periodontitis patients; (3) described the distribution of A. actinomycetemcomitans serotypes in patients and within sites; and, (4) assessed how this infection impacted upon local clinical symptoms of disease. We noted a significant positive relationship between the level of IgG anti-A. actinomycetemcomitans antibody and the frequency of teeth infected until nearly 13 teeth demonstrated an infection. Furthermore, the results showed a generally negative relationship between the antibody level and the burden of A. actinomycetemcomitans in the infected sites. Interproximal sites associated with first molar teeth were the predominant sites for subgingival colonization; incisors were also frequently infected in this population. The first molar teeth also exhibited the greatest level of A. actinomycetemcomitans while the incisors demonstrated a high level of A. actinomycetemcomitans in individual sites. The results clearly indicated the majority of the sites sampled were colonized by a single serotype of A. actinomycetemcomitans. We detected A. actinomycetemcomitans nearly 2 × times more frequently and a significant increase in the proportion of A. actinomycetemcomitans was found in samples obtained from teeth with bleeding on probing. The results also showed a significant trend for both pocket depth and attachment levels to be related to the presence and proportion of A. actinomycetemcomitans in the subgingival plaque. These findings detail the microbiological, immunological and clinical characteristics of a unique subset of periodontitis patients that appear to exhibit disease associated (caused?) with A. actinomycetemcomitans infection irrespective of clinical categorization. The results support a unique distribution of this microorganism in the subgingival ecology that is related to active host immune responses and clinical presentation of the tooth.     

小檗碱预防和治疗牙周炎的研究进展

小檗碱是一种从黄连等传统中药植物中提取的天然异喹啉类生物碱。小檗碱具有抑菌、抗炎、抗骨吸收、降血糖等多种生物学功效,且副作用小,近年来越来越多的研究报道了其在牙周炎防治中具有潜在的应用前景。本文总结了小檗碱在牙周炎预防和治疗方面的研究进展,旨在为牙周疾病的临床防治工作提供新的思路。研究表明,小檗碱可以通过抑制牙周致病菌生长、减轻牙周组织炎症反应以及调控牙槽骨吸收等抑制牙周炎的发生发展,从而达到预防和治疗牙周炎的效果。但是,牙周炎的发生机制十分复杂,目前的研究仍比较局限,未来需要更多的体内体外研究进一步探讨小檗碱抑制牙周炎发生发展的具体机制,以及进行更多的大样本前瞻性临床研究来确定小檗碱对牙周炎的防治效果。     

Periodontal pathogens and clinical parameters in chronic periodontitis

The use of next generation sequencing and bioinformatics has revealed the complexity and richness of the human oral microbiota. While some species are well known for their periodontal pathogenicity, the molecular‐based approaches for bacterial identification have raised awareness about new putative periodontal pathogens. Although they are found increased in case of periodontitis, there is currently a lack of data on their interrelationship with the periodontal measures. We processed the sequencing data of the subgingival microbiota of 75 patients with hemochromatosis and chronic periodontitis in order to characterize the well‐described and newly identified subgingival periodontal pathogens. We used correlation tests and statistical models to assess the association between the periodontal pathogens and mean pocket depth, and to determine the most relevant bacterial biomarkers of periodontitis severity. Based on correlation test results, nine taxa were selected and included in the statistical models. The multiple linear regression models adjusted for systemic and periodontal clinical variables showed that mean pocket depth was negatively associated with Aggregatibacter and Rothia, and positively associated with Porphyromonas. Furthermore, a bacterial ratio that was previously described as a signature of dysbiosis in periodontitis (%Porphyromonas+%Treponema+%Tannerella)/(%Rothia+%Corynebacterium) was the most significant predictor. In this specific population, we found that the best model in predicting the mean pocket depth was microbial dysbiosis using the dysbiosis ratio taxa formula. While further studies are needed to assess the validity of these results on the general population, such a dysbiosis ratio could be used in the future to monitor the subgingival microbiota.     

Suppression of the periodontopathic microflora in localized juvenile periodontitis by systemic tetracycline

Abstract. Since recent studies have implicated Actinobacillus actinomycetemcomitans in the etiology of localized juvenile periodontitis, this investigation determined the effectiveness of subgingival debridement, topical Betadine Solution®, and systemic tetiacycline in suppressing subgingival A. actinomycetemcomitans and other microorganisms. A total of 20 deep periodontal pockets and 10 normal periodontal sites of 6 localized juvenile periodontitis patients was included in the study. Each patient was treated in 3 stages over a period of 22 weeks, and the result of treatment was monitored for an additional 38 weeks. The first stage of treatment included plaque control, as well as thorough scaling and root planing, composed of at least 6 h of debridement. No concomitant periodontal surgery was performed. In the second stage, Betadine saturated cotton gauze was inserted into the periodontal pockets for 10 min. Stage 3 involved systemic tetracycline therapy (1 g/day) for J4 days. The subgingival microflora was determined at frequent intervals by selective culturing of A. actinomycetemcomitans and Capnocytophaga and by direct microscopic examination. The clinical effect was assessed by measuring changes in probing periodontal attachment level, probing periodontal pocket depth, radiographic alveolar bone mass, and other relevant clinical parameters. Scaling and root planing reduced the total subgingival bacterial counts and the proportions of certain Gram-negative bacteria, but no periodontal pocket became free of A actinomycetemcomitans. Betadine application had little or no effect on the subgingival microflora. In contrast, tetracycline administered via the systemic route suppressed. A actinomycetemcomitans, Capnocytophaga, and spirochetes to low or undetectable levels in all test periodontal pockets. A, actinomycetemcomitans reappeared in 9 of the deep periodontal pockets after the administration of tetracycline. Most of these 9 pockets became free of detectable A. actinomycetemcomitans during the second week of tetracycline administration, whereas pockets which yielded no A. actinomycetemcomitans after tetracycline therapy became free of the organisms during the first week of tetracycline treatment. This data suggests that systemic tetracycline therapy of localized juvenile periodontitis should, as a practical rate, be continued for 3 weeks. Periodontal destruction continued in 4 deep pockets which all showed high posttetracycline A, actinomycetemcomitans counts. All 6 pockets which demonstrated a marked gain in periodontal attachment yielded no cultivable A. actinomycetemcomitans. No association was found between periodontal disease status and subgingival Capnocytophaga, spirochetes or motile rods. The present study indicates that A. actinomycetemcomitans is an important etiologic agent in localized juvenile periodontitis. Also, this study demonstrates that the effectiveness of therapy can be monitored by subgingival A. actinomycetemcomitans counts, and that periodontal A, actinomycetemcomitans infections cannot be resolved by root surface debridement alone but can be cured by systemic tetracycline therapy.     

Transmission of Actinobacillus actinomycetemcomitans in families of adult periodontitis patients

At present Actinobacillus actinomycetemcomitans is regarded as an important microorganism in the etiology of some forms of periodontitis. The purpose of the present investigation was to study the number of Restriction Endonuclease Analysis (REA)-types present in the oral cavity of A. actinomycetem-comitans positive subjects and to study the possibility of transmission of A. actinomycetemcomitans within families of adult periodontitis patients. DNA of A. actinomycetemcomitans isolates was digested with a combination of the restriction endonucleases Pst I and Bam HI, after which the DNA fragments were separated by agarose gelelectrophoresis. To study the number of REA-types, multiple A. actinomycetemcomitans isolates obtained from 8 different sites in the oral cavity of five subjects were typed. The results showed that in most cases only one REA-type is present. In the 13 families investigated in 4 of the 26 children (15%) and in 1 of the 13 spouses (8%) of the adult periodontitis patients an indistinguishable REA-type was found within the families. This suggests that also in the case of adult periodontitis transmission of A. actinomycetemcomitans is possible, but does not seem to occur easily.     

牙周牙髓联合治疗与单纯牙周治疗对重度牙周炎疗效的临床对比研究

目的 探索重度牙周炎牙齿的牙髓治疗时机及牙周牙髓联合治疗对重度牙周炎的疗效。方法 采用多中心研究方法,选择重度牙周炎患牙180颗,随机分为牙周牙髓联合治疗组（试验组）和单纯牙周治疗组（对照组）,各90颗患牙。对照组仅进行牙周治疗;试验组记录牙髓状态,行根管治疗及牙周治疗;两组分别在治疗前和治疗后1、3、6及12个月进行检查,记录临床指标。结果 试验组90颗患牙中,22颗牙髓完全坏死,45颗部分坏死（冠髓坏死或根髓部分坏死）,23颗为活髓。治疗前,对照组和试验组各项临床指标的差异均无统计学意义（P>0.05）;治疗后3个月,试验组和对照组的临床指标开始出现差异（P<0.05）。结论 牙髓活力迟钝的重度牙周炎患牙及时进行完善的牙髓治疗,可控制炎症的发展,有利于牙周组织的愈合。     

用PCR和反杂交法对透明质酸在慢性牙周炎治疗中微生物学作用的研究

目的：观察外源性透明质酸局部应用对慢性牙周炎的微生物学作用。方法：采用口内自身对照研究。选择慢性牙周炎患者20例，受试牙306个，进行常规洁治、刮治和根面平整，实验组龈下辅助给予0．2％透明质酸(HA)凝胶。采用PCR和反杂交方法检测龈下菌斑中A．α．、P．g．、P．i．、B．f．和T．d．等五种牙周致病菌的动态变化。结果：治疗后，对照组和实验组5种牙周致病菌量的下降在统计学上无显著性。结论：未发现HA对慢性牙周炎患者的牙周致病菌在体内有显著抑制作用。     

Periodontitis patient and the spouse: periodontal bacteria before and after treatment

Abstract Suspected periodontal pathogens can be transmitted between spouses. The treatment response may be unsuccessful in periodontitis patients, if the spouse harbors these bacteria. The aim of the present 6-month follow-up study was to clarify whether the microbiological treatment outcome of periodontitis patients is related to the detection of suspected periodontal pathogens in the saliva of the spouse. 10 patients with advanced periodontitis and their spouses were included in the study. The patients received mechanical periodontal treatment and 500 mg metronidazole systemically 2 × a day for 7 days. The presence of visible plaque, gingival bleeding after probing, suppuration, supragingival and subgingival calculus and pocket depths were assessed at baseline and 1 and 6 months after treatment. Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia/Prevotella nigrescens, Campylobacter rectus and Peptostreptococcus micros were cultured from pooled subgingival samples from the patients and from salivary samples from the spouses at corresponding occasions, Periodontal conditions in the patients improved after treatment as determined by the significantly lower values of clinical variables 1 and 6 months after treatment compared to those at baseline. However, the re-emergence of periodontal bacteria after treatment of the patients was not related to the concurrent detection of the respective bacteria in the saliva of the spouses. In this study design, it seemed that the salivary bacterial load in the spouse was of minor importance for the microbiological treatment outcome of the patient.     

Antimicrobials for the treatment of aggressive periodontitis

Aggressive periodontitis is characterized by a considerable attachment loss over a relatively short period of time. It may be the consequence of either the presence of highly aggressive pathogens or a highly susceptible host. In the first case, the use of antimicrobials should be beneficial in the treatment of those patients. However, due to the organization of the micro-organisms as a biofilm, the increasing incidence of allergies and resistance against antimicrobials and their side-effects, there is still controversy about their benefit in the treatment of periodontal disease. This paper discusses indications for the use of antimicrobials, the substances prescribed and the type of application under the conditions of aggressive periodontitis.     

Prevalence of periodontal pathogens in localized and generalized forms of early-onset periodontitis

The primary objectives of this study were to investigate the prevalence of 8 putative periodontal pathogens in subjects with early-onset periodontitis (EOP) and to evaluate the microbial differences between localized and generalized forms of this periodontal disease condition. Thirty-one females and 11 males with a mean age of 30.3 (s.d. 4.0) years were examined. Seventeen subjects had generalized (GEOP) and 25 had localized early-onset periodontitis (LEOP). Subgingival plaque samples were assayed using PCR which provided subject prevalence data for the pathogens; Bacteroides forsythus 78.6%, Treponema denticola 88.1%, Actinobacillus actinomycetemcomitans 19.0%, Porphyromonas gingivalis 16.7%, Prevotella intermedia 40.4%, Prevotella nigrescens 61.9%, Eikenella corrodens 42.3% and Campylobacter rectus 92.8%. Only 3 healthy sites harbored one or more of these periodontal pathogens. Seven of the 8 subjects positive for A. actinomycetemcomitans had LEOP. P. intermedia was present in 58.8% of GEOP compared with 28% of LEOP subjects (p=0.046). At 82.4% of GEOP sites P. nigrescens was present while this bacteria was detected at 52% of LEOP (p=0.044). P. gingivalis was isolated from 22.6% of females but no male subjects (p=0.084). C. rectus was recovered from all female subjects compared to 72.7% of males (p=0.014). A. actinomycetemcomitans (37.5%) and C. rectus (86.5%) were more frequently identified in non-smokers compared to 7.6% and 68.8% of smokers, respectively (p <0.05). Microbial associations coincided with the clinical division of the cases into LEOP and GEOP in 83% of the subjects.     

Specific antibodies against Actinobacillus actinomycetemcomitans in serum and saliva of patients with advanced periodontitis

The aim of the present study was to discover any possible correlation between specific antibodies against Actinobacillus actinomycetemcomitans (A.a.) in serum and saliva. The test group consisted of 38 patients aged 31–68 yr (mean 49) with advanced periodontitis. Twenty-nine subjects aged 23–67 yr, without periodontal destruction, formed a control group with a reference level of specific salivary antibodies against A.a. A subgingival plaque sample for culturing A.a. , a specimen of stimulated whole saliva, and a sample of venous blood were taken from each subject of the test group. Specific IgG and IgA antibodies against A.a. were determined from serum and stimulated whole saliva by means of the ELISA test. Fifteen of the patients (39%) had cultivable A.a. Six of the 15 A.a. culture-positive patients and one of the 29 reference subjects exhibited very high antibody titers against A.a. in saliva. Specific IgG and IgA antibodies in saliva correlated highly significantly with the corresponding antibody values in serum among the patients in the test group. It was concluded that among patients with severe adult periodontitis, the less invasive saliva sample has a diagnostic value equal to that of the serum sample concerning specific antibodies against A.a.     

Impact of smoking on the clinical, microbiological and immunological parameters of adult patients with periodontitis

OBJECTIVES: The aim of the current study was to assess the impact of smoking on the clinical indices, the humoral immune response and the detection frequency of putative periodontal pathogens in patients with periodontitis cross-sectionally and following therapy. MATERIAL AND METHODS: Clinical measurements, subgingival plaque samples, gingival crevicular fluid (GCF) and sera were collected from 40 untreated patients with moderate-to-advanced chronic periodontitis before and after treatment over a period of 6 months. The treatment consisted of the initial therapy of scaling and root planing. Smoking status was self-reported and was confirmed by cotinine enzyme inhibition assay (CEIA). Whole-mouth clinical measurements were recorded with a manual periodontal probe at baseline (BAS) and at 6 months (RAS). Selected-site analyses were performed on the deepest site in each quadrant before and after therapy and clinical indices were recorded with an electronic pressure-sensitive probe. GCF sample volume was quantified using the Periotron 6000. Polymerase chain reaction (PCR) was utilized to determine the presence of Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, Prevotella intermedia, Treponema denticola and Tanerella forsythensis in subgingival plaque. Enzyme-linked immunosorbent assay examined the systemic antibody titres to these bacteria, and thiocyanate disassociation determined the antibody avidity to these organisms. RESULTS: At baseline, smokers showed significantly less gingival inflammation and lower GCF volume compared with non-smokers. After treatment, a compromised clinical outcome was noted for smokers in terms of pocket depth reduction and gain in attachment levels. No significant differences in the detection of putative periodontal pathogens in subgingival plaque existed between smokers and non-smokers. A consistent trend was noted in that smokers had lower sera immunoglobulin G antibody titres to these organisms before and after treatment (statistically significant for A. actinomycetemcomitans). This pattern was less clear when antibody avidities were considered, revealing only small differences, if any, between the two groups of patients. CONCLUSION: Current data indicate that smokers with periodontal disease have a suppressed inflammatory response, a significantly less favourable clinical outcome and seem to have an altered host antibody response to antigenic challenge than non-smokers. In contrast, the subgingival microflora of smokers appears similar to that of non-smokers.     

The effect of periodontal treatment on the salivary bacterial load and early plaque formation

AIM: The purpose of the present study was to investigate in periodontitis patients the relationship between the number of bacteria in the saliva and the amount of de novo plaque formation before and after treatment. METHODS: At baseline, before any treatment was provided, patients rinsed with 10 ml sterile saline. After professional tooth cleaning the patients were instructed to abolish all tooth cleaning procedures for the subsequent 24 h. After this period, the rinsing procedure was repeated and the amount of de novo plaque was assessed. Three months after the initial periodontal therapy was completed the experiment was repeated. The microbiological evaluation of the rinsing samples was carried out by means of phase contrast microscopy and anaerobic culturing. RESULTS: After treatment the amount of de novo plaque was less compared to before treatment, 0.40 and 0.65 respectively. Both before and after treatment more de novo plaque was present at sites with inflammation than at healthy sites. In order to evaluate the contribution of the numbers of salivary bacteria to the amount of de novo plaque formation an analysis was carried out for healthy sites. This analysis included only healthy sites as determined before treatment and the same sites after treatment. The results showed a significant reduction in the de novo plaque formation after treatment (0.49 before and 0.22 after treatment). Phase contrast microscopic evaluation showed that the number of bacteria in the rinsing samples after treatment was less than before treatment. After treatment also a reduction was found in the prevalence of Prevotella intermedia, Tannerella forsythensis and Peptostreptococcus micros in the rinsing samples. CONCLUSION: The present study confirms the observation in the literature that the periodontal condition is of major importance in the rate of de novo plaque formation. In addition, the results suggest that the number of bacteria in the saliva may play a role.     

Discrimination of refractory periodontitis subjects using clinical and laboratory parameters alone and in combination

The purpose of the present investigation was to use baseline clinical and laboratory parameters to distinguish subjects refractory to conventional periodontal therapy. Baseline clinical, microbial and host parameters were compared in 61 successfully-treated and 27 refractory subjects. Refractory subjects showed mean full-mouth attachment level (AL) loss and/or >3 sites with new AL loss >2.5 mm within 1 year after both scaling and root planing and surgery with systemic tetracycline. Successfully-treated subjects showed mean AL gain and no sites with new AL loss >2.5 mm after either regimen. Gingival redness, bleeding on probing, suppuration, supragingival plaque accumulation, pocket depth and AL were measured at 6 sites per tooth in each subject. The levels of 40 subgingival taxa were determined in subgingival plaque samples from up to 28 sites in each subject using checkerboard DNA-DNA hybridization. Serum antibody (Ab) to 85 subgingival species was determined using checkerboard immunoblotting. Levels of serum IgG2 and Gm23 allotype were measured using radial immunodiffusion; FcgammaRIIa and FcgammaRIIIb receptor haplotypes were determined using PCR and allele specific oligonucleotide probes. Odds ratios of a subject being refractory were determined by comparing measured parameters in the 2 subject groups using univariate and multivariate techniques. 17 of 151 clinical, microbial and immunological variables were significant using chi2 analysis after adjusting for multiple comparisons. For example, the odds ratios of a subject being refractory were 12.2, 5.4 and 6.9 if the subject had Ab >50 microg/ml to >9 species; S. constellatus counts >2.4% of the total DNA probe count or >2.1% of sites with AL >6 mm. The 17 significant predictor variables were used in logistic regression and discriminant analyses. Similar variables were selected using both analyses including the number of serum Ab to subgingival species >50 microg/ml, % S. constellatus in plaque samples and % sites with attachment loss >6 mm. In the logistic regression analysis model, the odds ratios associated with >9 species exhibiting >Ab 50 microg/ml, >2.1% of sites with AL >6 mm and >2.4% S. constellatus in plaque were 8.7, 6.8 and 2.4, respectively, after adjusting for other variables in the model. Discriminant analysis using these variables provided sensitivity, specificity, positive and negative predictive values of 0.66, 0.92, 0.80 and 0.85 respectively. Refractory periodontitis subjects could be distinguished using a subset of clinical, microbiological and immunological parameters.