The most characteristic traits that distinguish Abiotrophia strains from other streptococci are nutrient requirements, satellitism and pyrrolidonyl arylamidase activity. The presence of Abiotrophia strains was studied in early dental plaque formed on sucrose-containing and sucrose-free (glucose-containing) diets. The isolates were from 0- to 3-day dental plaque formed on the buccal surface of a lower premolar in six subjects. Identification of Abiotrophia strains was based on their pyrrolidonyl arylamidase activity. They were among the predominant cultivable microorganisms isolated from diluted suspensions of early dental plaque formed on both diets. Using biochemical tests without testing for pyrrolidonyl arylamidase activity, the Abiotrophia strains would be biochemically identified as Streptococcus mitis or unidentified streptococci, and many isolates first classified as unidentified streptococci were Abiotrophia strains. Generally, identifications using partial 16S rRNA sequences confirmed the identifications obtained biochemically. However, Abiotrophia elegans and Gemella strains were biochemically identified as Abiotrophia adiacens. Abiotrophia strains produce hydrogen sulfide, which may influence the metabolism and ecology of dental plaque and act as a virulence factor in periodontal disease. They are not able to grow on tryptic soy agar. In the present study, some S. mitis strains were not able to grow on mitis salivarius agar, and some Abiotrophia strains were able to grow on this medium. These observations indicate that the use of these media for estimation of total viable counts and number of streptococci introduces systematic error in studies of microorganisms in dental plaque.

THE BACTERIOLOGICAL COLONIZATION OF HEALING periodontal defects was investigated after treatment with guided tissue regeneration using expanded polytetrafluoroethylene membranes together with local metronidazole gel (25%, 250 mg/g). Twelve patients, each with one pair of comparable defects, had the test defect treated with the membrane plus metronidazole gel and the control defect treated with the membrane alone. Thirty weeks after removal of the membrane, the median gain in probing attachment level as a percentage of the initial defect depth was 92% for the test defects and 50% for the control defects (P = 0.001). The median number of cultivable bacteria decreased from 1.2 X 10(6) at the presurgical examination to 3.0 X 10(5) at the one week examination in the test group (P = 0.02), whereas an increase was observed in the control group. Similarly, a lower median proportion of black-pigmented Gram-negative anaerobic rods was observed one week postsurgically in the test group (0.004%) compared to the control group (3.5%) (P = 0.02). Two weeks after membrane insertion, and at all following examinations, no microbiological differences between test and control group were observed. Consequently, the influence of the metronidazole gel on the treatment result appears to have been confined to the initial regeneration phase. Despite the good clinical results in the test group, all membranes from both test and control pockets were heavily colonized with bacteria at the time of removal. To ensure maximal periodontal regeneration with formation of bone, future research in this area should concentrate on reducing the microbial colonization of the wound area.

The Actinobacillus actinomycetemcomitans population consists of a large number of clones among which the ubiquitous leukotoxin gene operon appears very homogeneous. Population genetic analyses performed by multilocus enzyme electrophoresis together with DNA fingerprinting and analyses of genomic DNA restriction fragment length polymorphisms (RFLP) on 97 strains isolated over a period of 45 years revealed that each of the serotypes a, b, c, d and e comprise genetically isolated subpopulations and that successful horizontal transfer of genomic DNA between strains of different serotypes appears to be extremely rare in vivo. In contrast, recombination between strains of the same serotype in general appears to take place in nature. The results provide evidence that non-serotypeable strains are serotype antigen-deficient variants originating from strains of the known serotypes. Serotype b and c strains may contain transmittable DNA sequences not found in strains of the other serotypes.

The microorganisms associated with mandibular third molar pericoronitis were investigated using direct microscopy and anaerobic culture method. The pericoronal pouch was sampled with paper points in A) 8 patients without mandibular third molar pericoronitis and B) 6 patients with mandibular third molar pericoronitis. Under the microscope, the microflora was found to be a complex mixture comprising gram-positive and gram-negative cocci, rods and filaments (including fusiform and curved rods), motile rods and spirochetes. Significantly higher proportions of motile, gram-negative rods were found in group B than in group A. The predominant cultivable microflora of 9 samples: A (4) and B (5) comprised several species of facultative and obligate anaerobic bacteria, namely Peptostreptococcus, Streptococcus, Actinomyces, Eubacterium, Propionibacterium, Veillonella, Porphyromonas, Prevotella, Bacteroides, Fusobacterium, Campylobacter, Staphylococcus, Stomatococcus, Lactobacillus, Neisseria, Capnocytophaga, Haemophilus, Selenomonas and Centipeda species. The microflora in pericoronitis appeared similar to that of diseased periodontal pockets.

The microorganisms associated with mandibular third molar pericoronitis were investigated using direct microscopy and anaerobic culture method. The pericoronal pouch was sampled with paper points in A) 8 patients without mandibular third molar pericoronitis and B) 6 patients with mandibular third molar pericoronitis. Under the microscope, the microflora was found to be a complex mixture comprising gram-positive and gram-negative cocci, rods and filaments (including fusiform and curved rods), motile rods and spirochetes. Significantly higher proportions of motile, gram-negative rods were found in group B than in group A. The predominant cultivable microflora of 9 samples: A (4) and B (5) comprised several species of facultative and obligate anaerobic bacteria, namely Peptostreptococcus, Streptococcus, Actinomyces, Eubacterium, Propionibacterium, Veillonella, Porphyromonas, Prevotella, Bacteroides, Fusobacterium, Campylobacter, Staphylococcus, Stomatococcus, Lactobacillus, Neisseria, Capnocytophaga, Haemophilus, Selenomonas and Centipeda species. The microflora in pericoronitis appeared similar to that of diseased periodontal pockets.

Plaque from the fitting surface of upper full dentures in 8 patients with denture‐induced stomatitis was sampled with curettes for study. To characterize the predominant cultivable flora, 1249 isolates (138–196 from each sample) were sub‐cultured from anaerobic roll tubes. Streptococci constituted 17–76% of the isolates in each sample (median 29%) and were identified asS. mitior, S. milleri, S. mutans, S. salivarius, andS. sanguis.Staphylococci (mainlyS. aureus) made up 2–15%, median 6%. Gram‐positive rods constituted 5–72% (median 45%), mainlyLactobacillusspp. (0–72%, median 19%) andActinomycesspp. (0–43%, median 9%). Gram‐negative cocci made up 0–28% (median 15%) and Gram‐negative rods only 0–1.8% (median 0%). Yeast viable counts on Sabouraud agar corresponded to 0–1.7% (median 0.27%) of the total viable counts. Stomatitis‐inducing denture plaque has a similar complex and variable, mainly Gram‐positive, bacterial flora as that of denture plaque in subjects with healthy oral mucosa. Yeasts and lactobacilli are, however, in greater evidence in stomatitis.

During a 5 day period without oral hygiene, 4 groups each consisting of 3 subjects with healthy gingivae, rinsed their mouths with solutions of tetracycline, vancomycin, polymyxin B or distilled water. Gingival plaque accumulated rapidly in the group rinsing with water, while tetracycline, and to a lesser degree vancomycin and polymyxin B inhibited plaque formation. Clinical gingivitis was not observed in any of the participants, but gingival exudate and leukocyte emigration were noted and seen to increase during the experimental period, especially in the water group. Rinsing with antibiotics inhibited these signs of subclinical inflammation to varying degrees. The bacterial composition of the gingival plaque changed in the water group during the experimental period towards a higher percentage of gram‐negative bacteria, following the pattern observed in previous investigations. Tetracycline markedly reduced the numbers of gingival plaque organisms, while polymyxin B favored a proliferation of gram‐positive cocci and short rods and depressed gram‐negative bacteria. Rinsing with vancomycin resulted in a pronounced shift towards an almost pure gram‐negative plaque flora. It is concluded that local administration of antibiotics with a limited spectrum may be useful for the study of the pathogenicity of the different components of the gingival microbial flora.

Three years ago, the very successful First European Workshop in Oral Microbiology took place in The Netherlands at the initiative of the oral microbiology group at the University of Nijmegen. That meeting was unusual because of the isolated location of the venue, modest facilities, and low cost of the accommodation, as well as the absence of any formal presentations, working papers, and slide projectors. Competent conveners initiated informal discussions centered around preselected topics, and many participants made brief contributions using the blackboard and an overhead projector. It was decided to repeat the experiment in three years' time in Denmark. The second European Workshop in Oral Microbiology was organised by Professor Mogens Kilian and other staff members of the Department of Oral Biology, Royal Dental College, Aarhus (Ellen V.G. Frandsen, Jesper Reinholdt, Henry Bleeg, Eva A. Strandgaard, and Dorthe Eggertsen). It took place on April 30-May 2, 1987, at Ldgumkloster Refugium, a cultural center for studies, recreation, and small conferences, built in connection with an old Cistercian monastery church in a peaceful village near the German border. The workshop was sponsored by Blumdller A/S, Bie & Bernsten A/S, and Munksgaard International Publishers Ltd. It gathered some seventy participants from eleven European countries (plus one "strayed" Australian), nearly all active in oral microbiology research. Using the same format as the first workshop, the discussions were quite spontaneous. Several participants had, however, prepared mini-contributions, some wanting to share new data or discuss ideas, methods, or interpretation, others inspired by the conveners to introduce the topics discussed at the various sessions. The following is a highly subjective summary of some of the issues, since no formal conclusion or agreement was arrived at. The topic of the first three-hour session was "Development, stability, and definition of the normal oral flora", with P.D. Marsh as the convener. Acquisition of oral flora in children occurs by salivary transmission from the mother and other persons, and, in two quoted cases, even from house dogs. The selective retention of bacteria transmitted depends on several mechanisms of attachment and adhesion, of which the relative importance in vivo is largely unknown. The major role of saliva as a source of nutrients was stressed, emphasizing glycoprotein as a source of nitrogen and carbohydrate. Several host defense mechanisms contribute to regulation of the normal flora, both specific (immunoglobulins) and non-specific (mucins, lysozyme, lactoferrin, peroxidase, polymorphs). Their combined action has not been much studied, but they surely do not eliminate the normal flora. The normal (resident, indigenous) oral flora consists of those organisms which have the mouth as their primary habitat. They establish a more-or-less permanent residence at particular surfaces, in one or more of several oral habitats, with some organisms being dependent on teeth or inflamed gingiva for colonization. Since some 200 species have been identified, it has become clear that many of them are only sometimes detectable in some persons, and in some sites. Examples of such members of the normal flora are Streptococcus mutans, Bacteroides gingivalis, Actinobacillus actinomycetemcomitans, and Candida albicans. The second topic was "Microbial interactions in the oral flora", with H. Donoghue as convener. The discussion started with the role for plaque growth and physiology of co-aggregation of pairs of bacterial species, then touched upon in vitro studies of consortia of bacteria, and in vivo cross-inhibition between species of black-pigmenting Bacteroides. Experiments on oral implantation of Streptococcus mutans and S. sanguis (the latter more easily implanted than the first) were reviewed, and the prospect of raising colonization resistance to "oral pathogens" mentioned. In experimental infections in animals following injection of plaque, pathogenic synergy between species is well-documented. The questions raised included: (1) whether pathogenic synergy exists in periodontal disease?; (2) which species initiate the process, and is there a succession of pathogens?; and (3) what is the definition of a pathogen in mixed infections? In "Newly described oral species", convened by J.M. Hardie, the need to know exactly with what organisms we are working was emphasized. Our taxonomic basis is constantly developing. New species are described, and some old ones have to be renamed. The pigmented and non-pigmented Bacteroides species were reviewed. Several new genera (such as Mitsuokella dentalis) are being separated from the genus Bacteroides. Actinobacillus actinomycetemcomitans belongs neither in the Actinobacillus nor in the Haemophilus genus, and a new genus will have to be formally proposed to include A.a, H. aphrophilusl paraphrophilus, and H. segnis. The taxonomy of oral streptococci is also under revision. Several oral Treponema species have been described, while large spirochetes and those with many endoflagella are still not cultivable. Also, the Eubacterium, Wolinella, and Capnocytophaga genera were discussed. The two topics on the second day were "The pool of virulence factors in plaque" [convener, M. Kilian (J. Carlsson had sent regrets)], and "Methods for the study of virulence factors", convened by B. Guggenheim. Most of the discussion centered on periodontal diseases, with questions such as, what virulence factors are active in vivo, and can we detect them in vitro? Bacterial hydrolytic enzymes can degrade a wide range of biologically and structurally important proteins. These and similar tissue-derived enzymes have great pathogenic potential. However, so far their demonstration in periodontal pockets has not proved useful in the prediction of disease activity. Bacterial perturbation of mechanisms regulating the periodontal inflammatory reaction and immune system seems to be of major importance but presents a complex situation: Neutrophils are inhibited; immunoglobulins are degraded; and bacteria can activate as well as inhibit both the complement system and antibody production (bacteria which themselves may be harmful or protective). In some instances, virulence factors could be dependent on phage infection of the bacteria. In vivo virulence factors may be neutralized by antibodies. Similar factors from different bacteria are, however, antigenically distinct, so that the host will have to produce new protective antibodies to each one as they appear. Although most studies of virulence factors have so far concentrated on a few "periodontal pathogens", many different species contribute to the pool of virulence factors involved in periodontal breakdown. In addition, these bacteria are dependent on other plaque species to produce an environment in which they can grow, e.g., by providing attachment, nutrients, and reduced Eh. The workshop certainly gave insight into the present status and inspiration for future work in oral microbiology. It is hoped that a similar meeting will be held in England in 1990.

Degradation of immunoglobulin A1 (IgAl) has previously been demonstrated in suspected periodontal pathogens. The present study revealed that a considerable proportion of the microbial flora in periodontal pockets of patients with juvenile periodontitis (median 29%) and rapidly progressive periodontitis (median 27%) was capable of degrading IgAl. Four different types of degradation occurred: Complete degradation of IgAl; extensive degradation leaving the Fc part of the molecule intact; traditional IgAl protease activity yielding intact Fc and Fab fragments, and removal of carbohydrate side chains on the IgAl molecule. Apart from species already known to degrade IgAl. IgAl protease was, for the first time, demonstrated in strains of Veillonella spp. The ability to cleave off carbohydrate side chains was a feature of a wide variety of bacteria belonging to the species Streptococcus sanguis, S. mitior, S. milleri, Veillonella spp., Actinomyces naeslundii. A. viscosus. Arachnia propionica , and Bacterionema matruchotii . The ability of subgingivally colonizing bacteria to degrade IgA may be a factor contributing to aggravation and perpetuation of the inflammatory reaction in the periodontal tissues.

In the absence of toothbrushing, the gingival crevice is colonized by a complex indigenous microflora causing gingivitis, a non-specific inflammation. Subgingival plaque may develop by downgrowth into the inflamed pocket of those micro-organisms from supragingival plaque which can multiply there. By direct microscopy, increased proportions of motile rods and spirochetes have been found in diseased pockets. Cultures on selective media have demonstrated increased prevalence of various gram-negative rods. Cultures on non-selective media have revealed the complexity and variability of the subgingival microflora, comprising more than 200 species. Destructive periodontitis is the result of subgingival colonization, which is favored by such ecological changes as plaque accumulation, gingivitis, and gingival exudate. These changes increase the numbers of micro-organisms and alter their proportions, but no single species appears in active sites which is not also commonly present in inactive sites. The subgingival micro-organisms have several virulence factors which promote colonization of the pockets, destroy host defense mechanisms, and provoke inflammation. It appears that different combinations of indigenous bacteria, rather than just a single species, can produce the pathogenic potential necessary to cause progression from gingivitis to destructive periodontitis.

A suitable method for determining the usual diet of an individual is a prerequisite for dietary counseling in caries prevention. The purpose of this study was to develop a method that was able to obtain relevant information on the dietary habits of an individual and to test the validity of this method. During a series of interviews a dietary history interview was therefore developed with special emphasis on dietary factors known to be important in caries etiology, such as frequency of eating and frequency and duration of sugar intake at meals and in-between meals. The validity of the method with regard to caries-related factors was tested by interviewing 49 caries-active and 55 caries-inactive 14-yr-old Danish schoolchildren. It was possible to detect a higher frequency of food intake and a more frequent and long-lasting use of sugar in the caries-active group. No difference was found in the consumption of sticky sugars between the two groups. With the present method it should be possible to pinpoint dietary problems in caries-active persons, so that dietary advice can be given in quantitative terms.

Some recent findings concerning microbial colonization of smooth surfaces of teeth, gingival crevices, occlusal fissures and removable dentures are reviewed considering the many ecologic factors of importance for the oral microbial communities. The oral microbiota is extremely complex comprising at least 200 taxa. In spite of interindividual and site-to-site variations, each oral habitat has a characteristic microbiota ranging from Gram-positive, facultatively anaerobic cocci and rods in occlusal fissures to predominance of Gram-negative, strictly anaerobic rods and spirochetes in deep periodontal pockets.