Non-oral Strains Research Articles

Qualitative Modifications of the Aligners’ Bacterial Flora under the Effect of Tea Tree Essential Oil

Objectives: The aim of our study is to look for the different Gram + and Gram - germs present in the biofilm of thermoplastic appliances and to note the qualitative modifications of the aligners’ bacterial flora after the use of the tea tree essential oil. Materials and Methods: Thirty-four patients were involved in the study. They were divided into two equal groups: a control group using a toothbrush and toothpaste as cleaning means and a test group using the tea tree essential oil as a supplement. Results: The results showed that the bacterial flora of the thermoplastic device is very diverse with a predominance of the genus Staphylococcus. It was isolated in more than half of the devices used by the control group. In terms of species, Enterobacter cloacae complex and Staphylococcus aureus were the most represented. In addition to the bacteria, usually found in the oral cavity, several other non-oral strains were frequently isolated, underscoring the importance of hand hygiene. The tea tree essential oil showed its effectiveness on Staphylococcus, which was significantly less encountered in the test group (p = 0.031). Conclusion: Thermoplastic appliances contain a diverse bacterial flora; Tea tree essential oil can be used for disinfecting purpose.

Peptostreptococcus stomatis sp. nov., isolated from the human oral cavity

Seven strains of anaerobic Gram-positive cocci isolated from human oral sites were subjected to a comprehensive range of phenotypic and genotypic tests. 16S rRNA gene sequence analysis revealed that the strains constituted a homogeneous group that was distinct from species with validly published names, but related to Peptostreptococcus anaerobius. All oral strains tested belonged to this group, whereas all non-oral strains studied were confirmed as P. anaerobius. A novel species, Peptostreptococcus stomatis sp. nov., is proposed to accommodate these oral strains. P. stomatis is weakly saccharolytic and produces acetic, butyric, isobutyric, isovaleric and isocaproic acids as end products of fermentation. The type strain of P. stomatis is W2278T (=DSM 17678T = CCUG 51858T); the G+C content of the DNA of this strain is 36 mol%.

Serological studies of peptostreptococci using an indirect fluorescent antibody test.

Members of the genus Peptostreptococcus are frequent isolates from periodontal lesions. A study was undertaken for determination of the serological relationships of oral and non-oral strains of several species of this genus. Eighty-nine strains of peptostreptococci representing seven species were tested by means of an indirect fluorescent antibody technique (IFA), with rabbit antisera to Peptostreptococcus anaerobius ATCC 27337, Peptostreptococcus micros VPI 2618-A, and Peptostreptococcus productus ATCC 27340. Each antiserum showed positive fluorescence when reacted with homologous cells. When anti-P. micros VPI 2618-A serum was added to suspensions of P. micros ATCC 33270 and clinical isolates of P. micros in the IFA, positive fluorescence was observed to a titer of 1:64 with 26 out of 29 strains. Positive fluorescence was also seen when P. anaerobius VPI 5737 and clinical isolates of P. anaerobius were tested with anti-P. anaerobius ATCC 27337 serum; a titer of at least 1:64 was observed in 10 out of 14 strains. These results support the presence of two serological groups of P. anaerobius, rabbit anti-P. anaerobius ATCC 27337 serum reacting with Group II organisms. No interspecies cross-reactivity was observed except with four strains of Peptostreptococcus magnus, which reacted with several antisera as well as with normal rabbit serum and the saline controls. These results indicate that rabbit antisera and the use of an IFA are useful in the identification of P. anaerobius and P. micros from oral lesions.

Cross-inhibition between black-pigmented Bacteroides species.

Cross-inhibition within the group of black-pigmented Bacteroides, including both oral and non-oral strains, was studied by means of a membrane filter technique. It was found that B. gingivalis possessed the most extended inhibitory capacity among all species tested. B. gingivalis showed inhibitory activity against B. intermedius, B. endodontalis, B. loescheii, and B. melaninogenicus. B. endodontalis was active against some B. intermedius strains. Among the saccharolytic species, some B. melaninogenicus strains were inhibitory for some B. endodontalis strains, some B. gingivalis strains, and some B. intermedius strains. These inhibitory activities observed in vitro may play a role in the colonization of the periodontal pocket.

An electron microscope survey of the surface structures and hydrophobicity of oral and non-oral species of the bacterial genus Bacteroides

Seventeen strains of Bacteroides representing 10 species were examined by negative staining; the majority were from the mouth but a few non-oral strains were included. Seven species had peritrichously-arranged, non-flagellar appendages which could be divided by morphology and ultra-structure into two subgroups, fibrils and fimbriae. Bacteroides asaccharolyticus strains B536 and B537 and Bacteroides gingivalis strains W50, W83, WPH15 and WPH35 had fimbriae with mean width of 4.4 nm and 0.5 to 6.0 μm long depending on the strain. The fimbrial length within each strain also varied. Fibrils were present on two fresh oral isolates of Bacteroides melaninogenicus, Bacteroides intermedius strains T588 and W09, Bacteroides corporis ATCC 33547, Bacteroides oralis ATCC 33269 and Bacteroides buccae ATCC 33574. Fibrils consistently clumped into bundles of variable thickness and formed a fringe around the cell periphery, ranging from 0.27 to 1.2 μm long depending on the strain. Fibril lengths of each strain were uniform. Fibrils had no measurable width and the clumps tapered towards the free ends. Bacteroides loeschii VPI 9085, Bacteroides pentosaceus strains NP333 and Jl and Bacteroides capillosus 29799 had no detectable surface appendages. Fimbriate strains had a layer outside the outer membrane, with a mean thickness of between 17.8 and 28.6 nm. Both fibrillar and fimbriate strains produced many small membranous vesicles budding from the outer membrane. There were two morphological forms of vesicles, ones with either fimbriae or fibrils (species-dependent) and ones with no attached appendages. Of eleven strains tested for cell-surface hydrophobicity by partitioning between hexadecane and buffer, all but one was non-hydrophobic.

The direct immunofluorescence in the presumptive identification of Bacteroides species

Two kits containing fluorescein labeled-polyvalent antisera to Bacteroides fragilis, Bacteroides distasonis, Bacteroides ovatus, Bacteroides thetaiotaomicron, Bacteroides vulgatus (Fluoretec-F) and to Bacteroides melaninogenicus subsp. melaninogenicus and intermedins and to Bacteroides asaccharolyticus (non-oral strains) (Fluoretec-M) were evaluated for the identification of the respective organisms in bacterial cultures and clinical specimens. In both cases immunofluorescent tests were compared to the bacteriological data. The results clearly demonstrated that Fluoretec-F has a specificity 97% and a sensitivity 96%. The corresponding values for Fluoretec-M were 99% and 87.5% respectively, either clinical specimens or pure cultures were examined. The presence of aerobic of facultative anaerobic bacteria and obligate anaerobic bacteria other than those to which antisera were included in the test kits did not seem to influence the test result. No cross-reactivity between Fluoretec-F and -M were observed. Also, no positive fluorescence was obtained when other anaerobic bacteria than Bacteroides spp. were tested. The same was also the case with various aerobic or facultative anaerobic bacteria. The use of Fluoretec-F and -M is considered as a useful tool for the early accurate identification of the Bacteroides species mentioned, which may serve as a guide for an appropriate prompt therapy which is especially needed in serious anaerobic infections.

Morphology and ultrastructure of oral strains of Actinobacillus actinomycetemcomitans and Haemophilus aphrophilus.

Selected human oral and nonoral strains of the genera Actinobacillus and Haemophilus were examined by transmission and scanning electron microscopy. The strains examined were morphologically identical to recognized Actinobacillus actinomycetemcomitans, Haemophilus aphrophilus, and Haemophilus paraphrophilus. By transmission electron microscopy, the cells were typically gram negative in morphology, with several strains possessing some extracellular ruthenium red-staining polymeric material. Numerous vesicular structures, morphologically identical to lipopolysaccharide vesicles, were seen to originate from and be continuous with the surface of the outer membrane. Large numbers of these vesicles were also found in the external environment. Scanning electron microscopic observations revealed that both actinobacilli and haemophili possessed surface projections and an amorphous surface material which connected and covered adjacent cells.

Antigenic studies of oral and nonoral black-pigmented Bacteroides strains.

Antigens of several oral and nonoral strains of Bacteroides asaccharolyticus (proposed classification of oral B. asaccharolyticus, Bacteroides gingivalis), Bacteroides melaninogenicus subsp. intermedius, B. melaninogenicus subsp. melaninogenicus, and B. melaninogenicus subsp. levii were identified in soluble preparations obtained by sonication, autoclaving, and NaOH treatment of whole bacterial cells. The sonicate preparations contained the most complete representation of soluble antigens using antisera to the whole organism in gel precipitation tests. Among strains of B. melaninogenicus subsp. intermedius many common antigens were detected, and no consistent antigenic differences were seen between strains from oral and nonoral sites. None of the antigens of B. melaninogenicus subsp. intermedius reacted with sera raised to several strains of oral or nonoral B. asaccharolyticus, nor did antigens prepared from the latter strains react with antisera to B. melaninogenicus subsp. intermedius. At least one common antigen was shared by strains of B. melaninogenicus subsp. intermedius and strains of B. melaninogenicus subsp. melaninogenicus; however, subspecies-specific antigens were also found. Antigens from and antisera to oral and nonoral strains of B. asaccharolyticus did not react with sera to and antigens from B. melaninogenicus subsp. melaninogenicus. Strains of B. asaccharolyticus isolated from the oral cavity were antigenically distinct from strains of B. asaccharolyticus obtained from nonoral sites and lesions. This lack of cross-reactivity between the oral and nonoral strains of B. asaccharolyticus together with recent findings of marked genetic differences between oral and nonoral strains of B. asaccharolyticus suggest that these groups of organisms may represent different species.

Genetic Heterogeneity in Bacteroides asaccharolyticus (Holdeman and Moore 1970) Finegold and Barnes 1977 (Approved Lists, 1980) and Proposal of Bacteroides gingivalis sp. nov. and Bacteroides macacae (Slots and Genco) comb. nov.

Strains of Bacteroides asaccharolyticus (Holdeman and Moore 1970) Finegold and Barnes 1977 (Approved Lists, 1980) which had been isolated from human mouths, most often from periodontal pockets, showed remarkably little deoxyribonucleic acid base sequence homology with the deoxyribonucleic acid of strains of the same species which were isolated from nonoral sites. In addition, four strains of catalase-positive Bacteroides melaninogenicus subsp. macacae Slots and Genco 1980 isolated from periodontal pockets in monkeys were distinct from B. asaccharolyticus. With deoxyribonucleic acid immobilized on membrane filters and index deoxyribonucleic acid made radioactive by translation repair with [3H] thymidine triphosphate, we found that the deoxyribonucleic acid homology among the three groups could readily be distinguished by their deoxyribonucleic acid base contents. The nonoral strains had 51 to 52 mol% guanine plus cytosine, the human oral strains had 46.5 to 48.5 mol% guanine plus cytosine, and the monkey catalase-positive strains had 42 to 43 mol% guanine plus cytosine in their deoxyribonucleic acids. Some strains which fit the species description of B. asaccharolyticus were not genetically related to the three groups. A new species, Bacteroides gingivalis sp. nov., which comprises those strains that resemble B. asaccharolyticus but have 46.5 to 48.5 mol% guanine plus cytosine, is described. These strains can also be distinguished serologically and by their production of phenylacetic acid. The type strain is Slots’ strain 2561 (=ATCC 33277). Bacteroides melaninogenicus subsp. macacae, which is comprised of the monkey isolates which resemble B. asaccharolyticus but which have a guanine plus cytosine content of 42 to 43 mol% and give a positive catalase reaction, is elevated to species status. Its name is thus Bacteroides macacae (Slots and Genco) comb. nov. The type strain of this species is Slots’ strain 7728-L6C (=ATCC 33141).

Evaluation of Fluoretec-M for detection of oral strains of Bacteroides asaccharolyticus and Bacteroides melaninogenicus.

Fluoretec-M is a polyvalent conjugate used in direct fluorescent-antibody staining for identification of the Bacteroides asaccharolyticus-Bacteroides melaninogenicus group. The Fluoretec-M reagent detected all oral and nonoral test strains of B. melaninogaenicus subsp. intermedius, all test strains of B. melaninogenicus subsp. melaninogenicus, and the nonoral strains of B. asaccharolyticus. However, the Fluoretec-M polyvalent reagent and the monovalent conjugates which constitute Fluoretec-M did not detect the oral strains B. asaccharolyticus. The use of Fluoretec-M can therefore generate false-negative results in studies of specimens from oral cavity and from nonoral sites in which an infection with B. asacacharolyticus of oral origin may have taken place. It is suggested that antibodies reactive with the oral antigenic type of B. asaccharolyticus be included in the preparative procedure of the Fluoretec-M reagent.

Direct hemagglutination technique for differentiating Bacteroides asaccharolyticus oral strains from nonoral strains

A simple and economical method for differentiating Bacteroides asaccharolyticus of oral sources from nonoral sources is described. The present data indicate that oral strains of B. asaccharolyticus strongly agglutinate sheep erythrocytes, whereas isolates from various nonoral sites typically are devoid of hemagglutination activity. The direct hemagglutination test may aid in determining the source of B. asaccharolyticus present in an infection, and thus the procedure has potential value as a means of biotyping.