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PubMed2000

Scanning electron microscopy and fluorescent in situ hybridization of experimental Brachyspira (Serpulina) pilosicoli infection in growing pigs.

Two groups of six 8-week-old pigs were challenged with 1x10(9) cfu Brachyspira (Serpulina) pilosicoli or Serpulina intermedia daily for 3 consecutive days to study the pathology of porcine colonic spirochetosis by scanning electron microscopy (SEM) and fluorescent in situ hybridization (FISH) with oligonucleotide probes targeting ribosomal RNA specific for B. pilosicoli and the genus Brachyspira/Serpulina. Six pigs served as noninoculated controls. The animals were euthanatized successively between postinoculation days 14 and 24. B. pilosicoli was reisolated in feces from all of the inoculated pigs; however, only two pigs developed transient watery diarrhea. S. intermedia was reisolated from four of the inoculated pigs, but clinical signs were not observed. Gross examination of the B. pilosicoli-infected pigs revealed dilated large intestines with a hyperemic mucosa, whereas the large intestines of the S. intermedia-inoculated pigs and the control pigs appeared normal. SEM examination of B. pilosicoli-infected pigs revealed degenerated epithelial cells and spirochetal colonization of the colonic mucosa in four pigs. By FISH, B. pilosicoli cells were found colonizing and invading the surface epithelium and the crypts in all the pigs. Spirochetal crypt colonization markedly exceeded the occurrence of spirochetes on the mucosal surface. SEM examination of S. intermedia-inoculated pigs revealed no abnormalities, and Serpulina cells were detected only sporadically in the otherwise normal-appearing mucosa of four pigs by FISH. The results provide further evidence that B. pilosicoli is associated with colitis in pigs, although the gross lesions are mild. The spirochete is capable of colonizing the large intestine, inducing mucosal damage, invasion of the crypt and surface epithelium, and focal infiltration of the lamina propria. In addition, the study shows the applicability of FISH for specific identification of B. pilosicoli in formalin-fixed tissue.

Animals↗
PubMed2001

Naturally occurring spirochetes in the colonic mucosa of raccoons (Procyon lotor).

The large intestines of 21 raccoons (Procyon lotor; 11 wild caught, 10 laboratory confined) were examined for the presence of intestinal spirochetes. Light microscopy of sections stained with hematoxylin and eosin and Warthin-Starry stain showed the presence of spiral shaped organisms deep within the lumina of intestinal glands at the ileocolonic junction of 16 raccoons (76% prevalence). All laboratory-confined, group-housed raccoons harbored the organisms, but only 6/11 (55% prevalence) live-trapped raccoons were positive for these spirochetes. The organisms were free in the glandular lumina, and there were no microscopic lesions. Two types of spirochetes were identified in the colonic glands: a slender spirochete 10-13 microm in length, 0.3 microm in diameter, and possessing long, thin tapered ends and a larger, regularly waved spiral organism (0.5 microm in diameter). The slender spirochete did not resemble any of the known spirochete genera and failed to grow on medium used to propagate oral treponemes and members of the genus Brachyspira.

Animals↗
PubMed1983

The effect of supragingival plaque removal on anaerobic bacteria deep periodontal pockets.

A study was made to determine if the numbers of subgingival anaerobes in deep periodontal pockets can be controlled by removal of only supragingival plaque. The study was based on the premises that the subgingival flora is dependent on the supragingival plaque for its source of organisms as well as for its perpetuation. Daily professional removal of only supragingival plaque produced a statistically significant reduction per sample in subgingival facultative and obligatory anaerobes.

Adult↗
PubMed2006

Improvement in motion efficiency of the spirochete Brachyspira pilosicoli in viscous environments.

Spirochetes are unique among swimming bacteria in terms of their lack of external flagella. They actively move in viscous environments, and, surprisingly, the swimming speed of the spirochete Leptospira interrogans has been reported to increase with viscosity in methylcellulose solutions. Many researchers consider that the presence of a loose, quasi-rigid network formed by linear polymer molecules is related to this strange phenomenon. One of the authors has proposed a theory that expresses this idea mathematically and successfully explains the speed properties of an externally flagellated bacterium in viscous environments. This theory predicts that the ratio of swimming speed to wave frequency (v/f ratio, motion efficiency in a sense) increases with viscosity. In this study, we demonstrated a new method of measuring the swimming speed and wave frequency of spirochetes and the motion characteristics of a swine intestinal spirochete, Brachyspira pilosicoli strain NK1f, measured in viscous environments. Several sets of swimming speed and wave frequency data were simultaneously derived from an animation obtained by our method. The v/f ratio of NK1f displayed a tendency to increase with increasing viscosity, suggesting the validity of the above-mentioned theory. Improvement of motion efficiency is at least one of the factors that maintain spirochete motility in viscous environments.

Animals↗
PubMed1992

Microbiota associated with residual clefts and neighboring teeth in patients with cleft lip, alveolus, and palate.

Twenty patients with residual clefts or pronounced soft tissue grooves, treated for uni- or bilateral cleft lip, alveolus, and palate were included in this study. Ten patients were recalled for dental prophylaxis at regular intervals, 10 patients were not. One microbiologic sample was obtained from the cleft area and two samples from a tooth adjacent to the cleft (sites adjacent and distant to the cleft). Between the recall and the nonrecall group there were notable differences in the presence of anaerobic Gram-negative organisms. Fusobacterium spp., Prevotella melaninogenica, and P. intermedia were more often found in nonrecall patients. While rarely seen in recall patients, spirochetes and motile rods were a common feature of nonrecall patients. The putative periodontal pathogens Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis were not detected in either group. The differences between the recall and the nonrecall groups were more pronounced when the respective samples from teeth were related to each other than when the samples obtained from the clefts were compared. The cleft flora was less complex irrespective of how good maintenance was and resembled the flora of teeth of well-maintained patients. Samples from clefts were never Wolinella positive, and harbored significantly less Capnocytophaga and Actinomyces viscosus than samples from dental sites.

Actinomyces viscosus↗
PubMed1982

Relationship between the percentage of subgingival spirochetes and the severity of periodontal disease.

The purpose of this study was to determine the correlations between various clinical assessments of inflammatory periodontal disease and the percentage of motile bacteria in the subgingival flora of sites representing widely varying states of periodontal disease. Darkfield microscopy was used to determine the percentage of spirochetes and other motile bacteria at two sites from each of 60 volunteers. For each site the following clinical assessments were recorded: Plaque Index (PII), gingival exudate (GE), Gingival Index (GI), bleeding tendency (BT), pocket depth (PD), connective tissue attachment loss (AL) and Periodontal Disease Index (PDI). Statistically significant (P less than 0.001) positive correlations (r) were found between the percentage of subgingival spirochetes and PII (r=0.54), GE (r=0.61), GI (r=0.57), BT (r=0.59), PD (r=0.56), AL (r=0.56) and PDI (r=0.61). Clinically healthy sites harbored much lower percentages of motile bacteria than did clinically disease sites. Most of the observed variation in the percentage of motile bacteria could be accounted for by variations in the percentage of spirochetes. The most significant increases in the relative percentages of subgingival spirochetes occurred when bleeding upon probing was observed as a sign of inflammation and/or when pocket depth and attachment loss exceeded 3 mm.

Adult↗
PubMed1983

Acute necrotizing ulcerative gingivitis. A transmission electron microscope study.

The purpose of this study was to provide additional evidence that in the lesions of acute necrotizing ulcerative gingivitis (ANUG), spirochetes and other microorganisms are capable of invading the nonnecrotic lamina propria of affected gingivae. Biopsies from eight patients with pathognomonic signs of ANUG were obtained under local anesthesia and processed for light and transmission electron microscopy. The results indicated that spirochetes and other bacteria are capable of penetrating the viable connective tissues of the lamina propria. The four zones of the ANUG lesion as first described by Listgarten were observed without difficulty. The maximum depth of bacterial infiltration into the lamina propria ranged from 155 to 400 microns from the nearest epithelial basal lamina. The histologic picture of the ANUG lesion did not always conform to the acute clinical manifestations.

Adolescent↗
PubMed1984

Metronidazole in periodontitis. I. Clinical and bacteriological results after 15 to 30 weeks.

The statistical association of certain anaerobic organisms such as black pigmented bacteroides (BPB) species and spirochetes with clinical signs of active periodontitis, i.e. bleeding upon probing and bone loss, suggests that the lesions may actually reflect a "specific infection" involving these or unidentified species. All the known oral species of BPB and spirochetes are anaerobes which suggests that antimicrobial treatment directed specifically against anaerobes might be effective in periodontal therapy. In this report, the short-term results of metronidazole treatment plus mechanical debridement in patients with extensive periodontal disease and of a double-blind clinical study in which metronidazole plus mechanical debridement is compared to placebo plus mechanical debridement are described. The findings indicate that 1 week of systemic metronidazole can optimize the clinical reduction of pockets and increase the apparent attachment in periodontitis patients who receive concurrent mechanical debridement of their root surfaces. In patients with extensive clinical involvement, metronidazole resulted in a significant reduction in the number of sites exhibiting pocket depths and attachment loss greater than or equal to 7 mm. When metronidazole plus mechanical debridement was compared with placebo plus mechanical debridement in a double-blind study, the metronidazole patients exhibited a significant improvement in those sites initially greater than or equal to 7 mm. The beneficial effect of the metronidazole was associated with a significant and sustained reduction of certain anaerobic organisms such as Bacteroides gingivalis and the large spirochetes. These data indicate that treatment aimed specifically toward the anaerobic component of the plaque flora can be associated with impressive clinical improvements 15 to 30 weeks after the initiation of treatment.

Alveolar Process↗
PubMed1985

Microscopic monitoring of pathogens associated with periodontal diseases. A review.

Microscopic monitoring of the subgingival microbiota to detect pathogens associated with periodontal diseases has been an active area of investigation. Researchers have demonstrated that plaque samples obtained at diseased sites contain more motile bacteria and less coccoid forms than healthy sites. Therapy at diseased locations has resulted in a shift in the microbiota to one that resembles healthy sites. However, the ability to predict disease activity based on morphotype counts still remains unverified. The current literature addressing chairside microscopic assessments as a diagnostic parameter is reviewed and discussed.

Bacteria↗
PubMed1985

Bacterial profiles of subgingival plaques in periodontitis.

In this report over 400 subgingival plaque samples taken from over 110 patients were examined microscopically and culturally for 30 bacterial parameters. The patients could be placed into six disease categories based upon clinical criteria. The bacterial profile of each clinical category was generally distinctive of that category. Periodontal patients who had been successfully treated and maintained had plaques that were populated by significantly higher proportions of Streptococcus sanguis, Actinomyces viscosus, A. odontolyticus and S. mutans and significantly lower proportions of B. gingivalis and spirochetes compared to the five untreated disease categories. The spirochetes were the overwhelming microbial type in the plaques of adult periodontitis (AP) patients, averaging about 45% of the microscopic count. The bacteriological results could not distinguish between ADA Type III and IV periodontitis, suggesting that the same type of infection was occurring in an active site in any AP patient. The patients designated as early onset periodontitis (EOP) differed from the other patients by their relative youth and by their significantly higher proportions of Bacteroides gingivalis and/or B. intermedius. Two types of EOP were recognized in which the most diseased variant was characterized by having an average of 49% spirochetes in the plaque. Four localized juvenile periodontitis (LJP) patients were notable in not having detectable A. actinomycetemcomitans. The data indicate that the various types of periodontitis, with the possible exception of LJP are specific anaerobic infections involving spirochetes and to a lesser extent B. gingivalis and B. intermedius.

Actinomyces↗
PubMed1986

Occurrence of Actinobacillus actinomycetemcomitans and spirochetes in relation to age in localized juvenile periodontitis.

Occurrence of Actinobacillus actinomycetemcomitans (A.a.) and spirochetes was examined in relation to the age of patients suffering from localized juvenile periodontitis (LJP). Twenty LJP patients were divided into three age groups: 14-16, 17-19, and 20-25 years of age. Two sites with radiographic alveolar bone loss and two sites without bone loss were examined in each patient. Probing depth, gingival bleeding after probing, and orthopantomographic alveolar bone loss were recorded. Subgingival bacterial samples were taken from the same four sites for cultivation of A.a., and for dark-field microscopic assessment of spirochetes. A.a. was isolated in the youngest group more frequently than in the two older ones. Spirochetes were detected in the youngest group as well, but were not related to the age of the patients. A.a. occurred as frequently with as without spirochetes, both in pockets exhibiting and not exhibiting bone loss. There was no difference in the spirochete proportions between the A.a.-positive and -negative sites. The occurrence of A.a. was not related to the absence, presence, or proportions of the spirochetes, but, unlike the situation with spirochetes, was related to the age of the LJP patients. Thus the occurrence of A.a. is possibly associated with the activity of the disease.

Actinobacillus↗
PubMed1986

Fluorescent antibody and flagella stains for rapid detection of bacteria at periodontally healthy and diseased sites.

Two rapid methods of identifying bacteria associated with periodontal disease were investigated to determine their diagnostic usefulness in longitudinal or epidemiologic studies. Three nonmotile organisms were identified by fluorescent antibody stains (FA) while percentages of motile bacteria were assessed by counting all spirochetes, flagellated and nonflagellated organisms stained with a simplified silver-plating stain for flagella. Relationships between disease activity and these bacteria from subgingival plaque samples taken at 18 individual sites (12 diseased, 6 healthy) were determined by correlating the quantity of detectable bacteria with the Gingival Index (GI), Plaque Index (PLI) and probing depth (PD). The highest correlations found with the FA stains were between Bacteroides gingivalis and probing depth (rs = 0.85), GI (rs = 0.80) and PLI (rs = 0.80). Bacteroides melaninogenicus and/or Bacteroides intermedius also correlated well with the GI (rs = 0.66), PLI (rs = 0.64), and PD (rs = 0.59), but to a lesser degree than B. gingivalis. Flagella stains showed that spirochetes correlated highly with PD (rs = 0.82), as did the total motile group with PLI (rs = 0.82). Motile bacteria alone were only moderately associated with the clinical parameters measured. The results of this investigation suggest that FA- and flagella-staining methods can be valuable screening tools for the detection of bacterial species and motile organisms in longitudinal or epidemiologic studies.

Actinomyces↗
PubMed1987

Trypsin-like activity in subgingival plaque. A diagnostic marker for spirochetes and periodontal disease?

Taxonomic screening of subgingival plaque organisms with various enzyme assays have shown that Treponema denticola, Bacteroides gingivalis and an unspeciated Capnocytophaga species possess a trypsin-like enzyme (TLE) that can be detected by the hydrolysis of N-benzoyl-DL-arginine-2-naphthylamide (BANA). As these organisms can be considered to be periodontopathic, it was of interest to determine whether this BANA hydrolyzing enzyme could be detected directly in subgingival plaque samples. Subgingival plaque samples were collected from single sites of known pocket depth, and after dispersal by vortexing, aliquots were incubated overnight with BANA and were counted microscopically. The color reactions were developed with fast garnet, read by the eye and classified as positive (red to red-orange), negative (yellow) and questionable. In the BANA-positive plaques, the spirochetes averaged 43% of the microscopic count, whereas in the BANA negative plaques the spirochetes averaged 8% of the microscopic count. The average pocket depth of BANA-positive plaques was 6.7 mm, whereas the average pocket depth of BANA-negative plaques was 4.5 mm. When both of these parameters were combined, the presence of a positive BANA reaction was usually indicative of subgingival plaques containing greater than 34% spirochetes removed from sites that had probing depths of 7 mm or more. Seventy-one per cent of the plaques removed from untreated periodontal patients were BANA-positive, while only 8% of the plaques removed from successfully treated patients seen at maintenance recall visits were BANA-positive.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteroides↗
PubMed1987

Comparison of dark-field microscopy and a flagella stain for monitoring the effect of a Water Pik on bacterial motility.

The purpose of this examiner-blind study was threefold: to compare the microbial counts obtained by two different techniques for assessing bacterial motility, to assess the inter-rater reliability of these two techniques and to evaluate the effect of a water irrigating device (Water Pik) on bacterial motility at 3- and 6-mm probing depths. Subgingival plaque samples were taken from 10 healthy patients having at least two sites that probed greater than 6 mm, (one control, one experimental). Half of the patients were sampled at 3 mm, the other half at 6 mm, both at baseline (Day 0) and at Day 21. Two slides were prepared from each plaque sample, one for dark-field evaluation and one stained with a simplified silver-plating technique for flagella. All slides were read simultaneously by 3 observers, and the per cent motility calculated for spirochetes, motiles and all others. Strong positive inter-rater reliability correlations ranging from r = 0.95 to r = 0.99 were found for both the dark-field and flagella staining techniques. Spirochete counts obtained by both techniques were highly correlated (r = 0.91), whereas counts for motiles resulted in negative correlations between the techniques. Dark-field counts were consistently higher than the flagella stain counts for motile rods. Spirochetes were reduced, but not significantly, after irrigation of both 3-mm and 6-mm sites. Bacterial motility can be evaluated by both dark-field and flagella-staining techniques with a high degree of inter-rater reliability.

Adult↗
PubMed1989

Effects of adjunctive treatment of periodontitis with tetracycline and spiramycin.

The present study was undertaken to compare the efficacy of two antibiotics, spiramycin and tetracycline, with a placebo when used adjunctively with scaling and root planing in the treatment of advanced adult chronic periodontitis. This was a double-blind, parallel, randomized trial with one factor (drug) at three levels. Ninety-six patients (mean age 46 +/- 1) were randomly assigned into one of three groups. All groups were scaled and root planed with each respective group receiving either spiramycin, tetracycline, or a placebo for 2 weeks. Two sites with probing depth of at least 7 mm were evaluated and the following clinical parameters were measured at baseline, 2, 8, 12, and 24 weeks: plaque index, bleeding on probing, crevicular fluid, probing depth, and change in the attachment level. The changes in the subgingival bacteria were monitored also using a differential staining technique. Seventy-nine patients completed the study. At the end of 24 weeks, although all three groups had shown clinical improvement when compared to the baseline data, there were no significant intergroup differences in any of the clinical parameters measured. While the proportion of spirochetes were significantly decreased (P less than 0.05) at 2- and 8-week intervals in both tetracycline and spiramycin groups (26% to 0.04% and 28% to 0.04%, respectively), compared to the placebo group (30% to 7%), only in the spiramycin group was the proportion of spirochetes significantly lower than the placebo group at the 24-week interval (3% and 11%, respectively). At week 24, the proportion of spirochetes in the tetracycline group had rebounded to 7%, which was not significantly different from the placebo group.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗