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ABSTRACT: Two tandemly aligned glucosyltransferase (GTF) genes whose gene products are responsible for water-soluble glucan synthesis were isolated from Streptococcus dentirousetti NUM1303 and sequenced. One of the GTF genes of S. dentirousetti consisted of a 4110 bp open reading frame (ORF) that encoded for a 1369 amino acid protein and was revealed to be a S. sobrinus gtfS homolog. The percent similarity of amino acid sequences of the GTF-S from S. dentirousetti compared to those from S. sobrinus was 99%. In addition, a putative gtfT was found in tandem in the downstream region of the S. dentirousetti gtfS. The gtfT of S. dentirousetti consisted of a 4527 bp ORF encoding for 1508 amino acids. The similarity of amino acid sequences of the GTF-T from S. dentirousetti and S. sobrinus was 94%. Phylogenetic analysis based on amino acid sequences from other related streptococcal GTFs suggested that both GTF-S and GTF-T of S. dentirousetti are closely related to S. sobrinus.
Microbiology and Immunology 05/2013; 57(5):386-90. · 1.30 Impact Factor
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ABSTRACT: Four Gram-positive, catalase-negative, coccoid-shaped isolates which were obtained from donkey oral cavities formed two distinct clonal groups when characterized by phenotypic and phylogenetic studies. From the results of biochemical tests, the organisms were tentatively identified as a streptococcal species. Comparative 16S rRNA gene sequencing studies confirmed the organisms to be members of the genus Streptococcus. Two of the isolates were related most closely to Streptococcus ursoris with 95.6% similarity based on the 16S rRNA gene and to Streptococcus ratti with 92.0% similarity based on the 60 kDa heat-shock protein gene (groEL). The other two isolates, however, were related to Streptococcus criceti with 95.0% and 89.0% similarities based on the 16S rRNA and groEL genes, respectively. From both phylogenetic and phenotypic evidence, the four isolates formed two distinct clonal groups and are suggested to represent a novel species of the genus Streptococcus. The names proposed for these organisms are Streptococcus orisasini sp. nov. ( type strain NUM 1801(T) = JCM 17942(T) = DMS 25193(T)) and Streptococcus dentasini sp. nov. (type strain NUM 1808(T) = JCM 17943(T) = DMS 25137(T)).
INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY 01/2013; · 2.11 Impact Factor
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ABSTRACT: Streptococcus mutans has been considered the principal etiological agent of dental caries in humans. S. mutans can secrete three kinds of glucosyltransferases (GTFs). One of these, GTF-B, which synthesizes water-insoluble glucans from sucrose, has been considered to be one of the most important factors of cariogenic dental plaque formation. Therefore, determination of whether GTF-B is present in plaque and saliva samples may contribute to the evaluation of individual virulence potential (caries risk). The aim of this study was to develop a feasible enzyme-linked immunosorbent assay (ELISA) for the routine quantification of GTF-B in plaque-derived cultures and clinical samples, and to apply this assay to an epidemiological study. To determine the presence of GTF-B in plaque samples, a sandwich-ELISA was devised, consisting of mouse monoclonal and rabbit polyclonal antibodies against GTF-B and a horseradish peroxidase-conjugated anti-rabbit antibody. The developed ELISA allowed for quantification of the amounts of purified GTF-B with satisfactory sensitivity and specificity; this method was not affected by other components such as plaque and saliva. Plaque samples from healthy volunteers were examined using this ELISA method and microbial analysis to apply the assay to an epidemiological study. A correlation was observed between the amount of extracted GTF-B and S. mutans levels as determined by ELISA and cultivated with Mitis Salivarius Bacitracin agar plates derived from plaque samples, although there were some exceptions. In this regard, this ELISA system has the advantage of estimating both the individual numbers of S. mutans and the productivity of GTF-B, namely, the cariogenic potential of S. mutans simultaneously. These results indicate that this ELISA method is a useful tool for the diagnosis of caries risk.
Hybridoma (2005) 06/2012; 31(3):176-9. · 0.42 Impact Factor
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ABSTRACT: A Gram-negative, rod-shaped, non-spore forming and non-motile bacterium, designated strain NUM 1720(T) , was isolated from the oral cavity of bears. Based on 16S rRNA gene sequence similarity, strain NUM 1720(T) was shown to be related to Gibbsiella quercinecans (99.4%). The gyrB and rpoB gene sequences of strain NUM 1720T showed 98.0% and 98.2% similarity with those of G. quercinecans. The DNA-DNA hybridization value of strain NUM 1720(T) with G. quercinecans was 63.8%. The G + C content of the genomic DNA of the isolates was 55.0 mol%. Fatty acid analysis data supported the affiliation of strain NUM 1720(T) to the genus Gibbsiella. The major menaquinone and ubiquinone were MK-8 and Q-8, respectively. Strain NUM 1720(T) can be differed from G. quercinecans by the reactions to acetoin, inositol and D-arabinose. Strain NUM 1720(T) therefore represents a novel species, for which the name Gibbsiella dentisursi sp. nov. is proposed, with type strain NUM 1720(T) (= JCM 17201(T) = DSM 23818(T)).
Microbiology and Immunology 04/2012; 56(8):506-12. · 1.30 Impact Factor
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ABSTRACT: Oral immunotherapy is potentially useful for the treatment of allergic diseases. We previously demonstrated that allergen-induced airway inflammation and immunoglobulin E (IgE) production in mice were suppressed by oral administration of high-dose transgenic (Tg) rice seeds (approximately 50 g/kg/day) expressing a T cell epitope of Dermatophagoides pteronyssinus group 1 allergen (Der p 1). However, this amount of Tg rice seeds was not realistic in our daily life. In this study, allergen-induced airway inflammation and IgE production following oral immunotherapy with a realistic (lowest) dose of Tg rice seeds were investigated.
Mice orally administered with Tg or non-Tg rice seeds at approximately 5 g/kg/day for 1 week were immunized with recombinant Der p 1, and then challenged with the corresponding allergen. The infiltration of inflammatory cells into the airways and the levels of allergen-specific serum IgE were examined.
Low-dose oral administration of Tg rice seeds significantly inhibited the allergen-induced infiltration of eosinophils and lymphocytes into the airways, but allergen-specific IgE synthesis was not changed.
Low-dose oral immunotherapy with Tg rice seeds could suppress allergen-induced airway inflammation through mechanisms other than the downregulation of IgE synthesis.
International Archives of Allergy and Immunology 01/2012; 158 Suppl 1:66-9. · 2.40 Impact Factor
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ABSTRACT: Celiac disease (CD) is a chronic immune-mediated intestinal inflammatory disorder afflicting genetically susceptible individuals triggered by the consumption of dietary cereals with high gluten content. As with many other organ-specific autoimmune diseases, the dominant tissue-destructive inflammation in CD is T cell-mediated. The proinflammatory cytokine IL-15 that is overexpressed in the intestinal epithelium of CD patients has emerged as a pivotal element that orchestrates intestinal inflammation and T cell-mediated autoimmune tissue destruction. Although no animal model exists that recapitulates the full spectrum of CD pathophysiology, we have previously reported that transgenic mice that overexpress human IL-15 in enterocytes (T3(b)-hlL-15 Tg) display many of the T cell-mediated pathologic features seen in CD. Extending these observations, we now report that T3(b)-hlL-15 Tg mice in addition to recapitulating T cell-mediated effects also display autoantibodies including those against tissue transglutaminase 2 and extensive lamina propria plasmacytosis, all of which are characteristic of CD, thereby reflecting the possibility that locally expressed IL-15 drives both T and B cell pathologic effects seen in CD. More importantly, these findings support the validity and utility of T3(b)-hlL-15 Tg mice as a reasonable model to investigate not only tissue-destructive pathologic processes in CD, but also to explore novel therapeutic modalities for the treatment of this disease.
Journal of Clinical Immunology 09/2011; 31(6):1038-44. · 3.08 Impact Factor
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Kazuya Suzuki,
Osamu Kaminuma,
Lijun Yang,
Toshiro Takai,
Akio Mori,
Makiko Umezu-Goto,
Takayuki Ohtomo,
Yasushi Ohmachi,
Yuko Noda,
Sakiko Hirose,
Ko Okumura,
Hideoki Ogawa, Kazuko Takada,
Masatomo Hirasawa,
Takachika Hiroi,
Fumio Takaiwa
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ABSTRACT: This study tested the feasibility of oral immunotherapy for bronchial asthma using a newly developed subunit vaccine in which a fragment (p45-145) of mite allergen (Der p 1) containing immunodominant human and mouse T cell epitopes was encapsulated in endoplasmic reticulum-derived protein bodies of transgenic (Tg) rice seed. Allergen-specific serum immunoglobulin responses, T cell proliferation, Th1/Th2 cytokine production, airway inflammatory cell infiltration, bronchial hyper-responsiveness (BHR) and lung histology were investigated in allergen-immunized and -challenged mice. Prophylactic oral vaccination with the Tg rice seeds clearly reduced the serum levels of allergen-specific IgE and IgG. Allergen-induced CD4(+) T cell proliferation and production of Th2 cytokines in vitro, infiltration of eosinophils, neutrophils and mononuclear cells into the airways and BHR were also inhibited by oral vaccination. The effects of the vaccine were antigen-specific immune response because the levels of specific IgE and IgG in mice immunized with Der f 2 or ovalbumin were not significantly suppressed by oral vaccination with the Der p 1 expressing Tg rice. Thus, the vaccine does not induce nonspecific bystander suppression, which has been a problem with many oral tolerance regimens. These results suggest that our novel vaccine strategy is a promising approach for allergen-specific oral immunotherapy against allergic diseases including bronchial asthma.
Plant Biotechnology Journal 03/2011; 9(9):982-90. · 5.44 Impact Factor
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ABSTRACT: Streptococcus mutans and Streptococcus sobrinus are considered to be important bacterial species in the initiation of human dental caries. Therefore, the establishment of a reliable genotyping method to distinguish S. mutans from S. sobrinus is of central importance.
We assessed the usefulness of repetitive extragenic palindromic polymerase chain reaction (rep-PCR) using ERIC primer banding patterns in differentiating S. mutans and S. sobrinus.
Five S. mutans and two S. sobrinus prototype strains and 50 clinical isolates (38 S. mutans serotype c, 4 S. sobrinus serotype d, and 8 S. sobrinus serotype g) were examined. The banding patterns of amplicons generated were compared among the prototype strains and clinical isolates, to find common bands that distinguish S. mutans and S. sobrinus.
Multiple banding patterns were seen with all strains tested. The representative strains of S. mutans tested revealed six unique, strong bands at 2,000 bp, 1,700 bp, 1,400 bp, 1,100 bp, 850 bp, and 250 bp, whereas S. sobrinus had seven strong bands at 2,000 bp, 1,800 bp, 1,100 bp, 900 bp, 800 bp, 600 bp, and 550 bp. The band at 1,100 bp was the only band that was observed in both S. mutans and S. sobrinus. Furthermore, most clinical S. mutans isolates revealed identical banding patterns. All S. mutans had amplicons at 1,700 bp, 850 bp, and 250 bp, whereas those of S. sobrinus were at 1,100 bp, 900 bp, and 800 bp.
These results indicate that using rep-PCR with the ERIC primers can distinguish between S. mutans and S. sobrinus.
Journal of Oral Microbiology 01/2011; 3.
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ABSTRACT: Three Gram-positive, catalase-negative, coccus-shaped organisms were isolated from the oral cavities of bears. The isolates were tentatively identified as a streptococcal species based on the results of biochemical tests. Comparative 16S rRNA gene sequencing studies confirmed that the organisms were members of the genus Streptococcus, but they did not correspond to any recognized species of the genus. The nearest phylogenetic relative of the new isolates was Streptococcus ratti ATCC 19645(T) (98.6 %), however, DNA-DNA hybridization analysis showed that the isolates displayed less than 15 % DNA-DNA relatedness with the type strain of S. ratti. Colonies of the novel strains grown on mitis salivarius agar showed an extracellular polysaccharide-producing colony morphology. Based on phenotypic and phylogenetic evidence, it is proposed that the novel isolates are classified in the genus Streptococcus as Streptococcus ursoris sp. nov. The type strain of S. ursoris is NUM 1615(T) (=JCM 16316(T)=DSM 22768(T)).
INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY 02/2010; 61(Pt 1):40-4. · 2.11 Impact Factor
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ABSTRACT: Four strains (NUM 1903(T), NUM 1904, NUM 1912 and NUM 1925) that were obligately anaerobic, pigmented, Gram-negative-staining rods were isolated from the oral cavity of donkeys. These strains were analysed using the Rapid ID 32A, API 20A and API ZYM systems, by DNA-DNA hybridization with other related species and by 16S rRNA gene sequencing. 16S rRNA gene sequence analysis showed that each of the new isolates was a member of the genus Prevotella and related to Prevotella multiformis PPPA21(T), showing about 93 % sequence similarity. Based on phylogenetic and phenotypic evidence, it is proposed that the four strains are representatives of a novel species, for which the name Prevotella dentasini sp. nov. is proposed. The type strain is NUM 1903(T) (=JCM 15908(T)=DSM 22229(T)).
International journal of systematic and evolutionary microbiology 09/2009; 60(Pt 7):1637-9. · 2.27 Impact Factor
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ABSTRACT: Four Gram-stain-positive, catalase-negative, coccoid-shaped isolates were obtained from the oral cavities of wild boars and characterized by phenotypic and phylogenetic studies. On the results of biochemical tests, the organisms were tentatively identified as a streptococcal species. Comparative 16S rRNA gene sequencing studies confirmed that the organisms are members of the genus Streptococcus, with Streptococcus equi subsp. equi ATCC 33398(T) as their closest phylogenetic relative (94.7 % similarity). DNA-DNA hybridization analysis showed that the isolates displayed less than 10 % relatedness to Streptococcus equi subsp. equi DSM 20561(T). From the phylogenetic and phenotypic evidence, the four isolates represent a novel species of the genus Streptococcus, for which the name Streptococcus dentapri sp. nov. (type strain NUM 1529(T) =JCM 15752(T) =DSM 21999(T)) is proposed.
International journal of systematic and evolutionary microbiology 09/2009; 60(Pt 4):820-3. · 2.27 Impact Factor
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ABSTRACT: Nucleotide sequences of water-insoluble glucan-producing glucosyltransferase (gtf) genes of new mutans streptococci isolated from pig oral cavity, Streptococcus orisuis JCM14035, and of Streptococcus criceti HS-6 were determined. The gtf gene of S. orisuis JCM14035 consisted of a 4,401 bp ORF encoding for a 1,466 amino acids, and was revealed to belong to the gtfI group. The percent homology of amino acid sequence of the GTF-I from S. orisuis and S. criceti are 95.0%, however, this score ranges from 77.0% to 78.0% when compared to Streptococcus sobrinus 6715. The deduced N-terminal amino acid sequence was considered responsible for the secretion of GTF-I in S. orisuis JCM14035 and S. criceti HS-6 with high similarity to known GTF proteins from other streptococci. In addition, two other conserved regions, i.e., N-terminal putative catalytic-site and C-terminal glucan binding domain, were also found in GTF-Is of S. orisuis JCM14035 and S. criceti HS-6. Phylogenetic analysis suggested that S. orisuis JCM14035 and S. criceti HS-6, closely related to each other, resemble S. sobrinus and S. downei based on the amino acid sequences of the GTFs.
The Journal of Microbiology 05/2008; 46(2):202-8. · 1.10 Impact Factor
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ABSTRACT: Seventy-three strains of streptococci were isolated from the bat oral cavity. The colonies of strains grown on mitis salivarius agar were similar in morphology to those of mutans-like streptococci. The novel strains were analysed biochemically using the Rapid ID32 Strep microsystem and were subjected to DNA-DNA hybridization with other oral streptococci and to 16S rRNA gene sequence analysis. Based on phylogenetic and phenotypic evidence, it is proposed that these isolates be classified as Streptococcus dentirousetti sp. nov. The type strain of Streptococcus dentirousetti sp. nov. is NUM 1303(T) (=JCM 14596(T)=DSM 18963(T)).
International journal of systematic and evolutionary microbiology 02/2008; 58(Pt 1):160-3. · 2.27 Impact Factor
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ABSTRACT: Gram-positive streptococcal mutans-like strains, but with clearly different colony formation than S. orisuis on Mitis Salivarius agar, were isolated from the pig oral cavity and identified by 16S rRNA sequencing, G+C content, DNA-DNA homology and extensive biochemical and serological testing. The phenotypic data showed that the strains were similar to S. orisuis except for susceptibility to bacitracin. DNA-DNA homology between the isolates and S. orisuis was 72~81%. However, serological data showed that they have a different sero-specific antigen from S. orisuis and other mutans streptococci. A new serotype, designated p, strains are classified in a serovar of S. orisuis, one of mutans streptococci.
Microbiology and Immunology 02/2008; 52(2):64-8. · 1.30 Impact Factor
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ABSTRACT: Five bacterial strains, designated as NUM 1001(T), NUM 1002, NUM 1003, NUM 1004 and NUM 1005, were isolated from the oral cavities of pigs. Colonies grown on mitis salivarius agar were similar in morphology to those of mutans streptococci. The novel isolates were analysed biochemically using the Rapid ID 32 Strep microsystem, subjected to DNA-DNA hybridization with oral streptococci and had their 16S rRNA genes sequenced. On the basis of the phylogenetic and phenotypic evidence obtained, the strains represent a novel species of the genus Streptococcus, for which the name Streptococcus orisuis sp. nov. is proposed. The type strain is NUM 1001(T) (=JCM 14035(T)=DSM 18307(T)).
International journal of systematic and evolutionary microbiology 07/2007; 57(Pt 6):1272-5. · 2.27 Impact Factor
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ABSTRACT: An evaluation on the applicability of Mitis Salivarius agar (MS) medium, commonly used for the detection of oral streptococci in human and animals, to dog specimens and the development of a new selective medium for isolating streptococci from the canine oral cavity are described. Oral samples from dogs were cultured on MS medium under anaerobic conditions. The predominant facultative anaerobic bacteria on MS plates were gram-negative rods. Selectivity of streptococci on MS medium was 21.2%. A new selective medium, designated MS-CAN-AE, was developed for the isolation of streptococci from the canine oral cavity. The average growth recovery of laboratory and clinically isolated strains of streptococci on MS-CAN-AE medium was 84.1% of that on MS medium. Gram-positive rods and gram-negative rods and cocci rarely grew on the MS-CAN-AE. The selectivity of MS-CAN-AE was 95.0% for clinical samples. MS-CAN-AE medium will be helpful for investigations of streptococci in the canine oral cavity.
Journal of Microbiological Methods 10/2006; 66(3):460-5. · 2.09 Impact Factor
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ABSTRACT: A simple and rapid method for Porphyromonas gingivalis detection in clinical samples has been developed using polymerase chain reaction (PCR) and an immunochromatographic assay (ICA) with a lateral-flow device (strip) to detect species-specific 16S rRNA genes.
The PCR used a pair of primer sets labeled with fluorescein isothiocyanate (FITC) or biotin at each 5' terminus. The strip used a nitrocellulose membrane containing streptavidin conjugated to gold particles and anti-FITC line.
PCR and ICA detected as few as 1 and 10 cells of P. gingivalis, respectively. ICA required 5 to 10 minutes more than the initial PCR. The amplifications were not observed in other oral black-pigmented bacteria at concentrations of 10(6) colony forming unit (CFU). The ICA strips showed bands at more than 10(4) CFU/ml equivalents in clinical samples from periodontitis.
A diagnostic assay based on PCR-ICA was developed for the detection of P. gingivalis, and results were obtained visually in 3 hours. PCR-ICA will be a valuable tool for the rapid detection of target bacteria by chair side.
Journal of Periodontology 05/2005; 76(4):508-12. · 2.60 Impact Factor
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ABSTRACT: The susceptibility of Candida albicans to catechin under varying pH conditions and the synergism of the combination of catechin and antimycotics were evaluated.Method: Antifungal activity was determined by broth dilution and calculation of cfu.
The antifungal activity of catechin was pH dependent. The concentration of epigallocatechin gallate (EGCg) causing 90% growth inhibition of tested strains of C. albicans was 2000 mg/L at pH 6.0, 500-1000 mg/L at pH 6.5 and 15.6-250 mg/L at pH 7.0. Among catechins, pyrogallol catechin showed stronger antifungal activity against C. albicans than catechol catechin. The addition of 6.25-25 or 3.12-12.5 mg/L EGCg to amphotericin B 0.125 or 0.25 mg/L (below MIC) at pH 7.0 resulted in enhancement, respectively, of the antifungal effect of amphotericin B against amphotericin B-susceptible or -resistant C. albicans. Combined treatment with 3.12-12.5 mg/L EGCg plus amphotericin B 0.5 mg/L (below MIC) markedly decreased the growth of amphotericin B-resistant C. albicans. When fluconazole-susceptible C. albicans was treated with 25-50 mg/L EGCg and fluconazole 0.125-0.25 mg/L (below MIC), its growth was inhibited by 93.0%-99.4% compared with its growth in the presence of fluconazole alone. The combined use of 12.5 mg/L EGCg and fluconazole 10-50 mg/L (below MIC) inhibited the growth of fluconazole-resistant C. albicans by 98.5%-99.7%.
These results indicate that EGCg enhances the antifungal effect of amphotericin B or fluconazole against antimycotic-susceptible and -resistant C. albicans. Combined treatment with catechin allows the use of lower doses of antimycotics and induces multiple antifungal effects. It is hoped that this may help to avoid the side effects of antimycotics.
Journal of Antimicrobial Chemotherapy 03/2004; 53(2):225-9. · 5.07 Impact Factor
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ABSTRACT: Microbial flora and gingival conditions were compared between a group of patients with phenytoin-induced gingival hyperplasia as a test group, a control group of patients who were administered phenytoin without gingival hyperplasia and a blank group who took no phenytoin and no gingival hyperplasia in mentally retarded patients.
Subgingival plaque samples were collected from a PHT-induced overgrown gingival pocket and microbiological experiments were performed by culture and PCR methods.
The predominant genera in total cultivable bacteria from subgingival plaque samples were streptococcus and actinomyces with recovery ranges of 37.6-42.1% and 23.4-25.5% of total bacteria, respectively, in all groups. The test group showed a significantly higher level of obligate Gram-negative rods than the control and blank groups. Black-pigmented obligate anaerobic Gram-negative rods were detected in 10.3% of total cultivable bacteria in the test group. The black-pigmented rods were predominantly Prevotella intermedia in the test group and Prevotella nigrescens in the control and blank groups. Porphyromonas gingivalis and Porphyromonas endodontalis were also detected in the test group with small values.
These results suggested that black-pigmented rods, particularly P. intermedia, could be habitable in the environment of gingival hyperplasia.
Journal of Periodontal Research 11/2003; 38(5):477-81. · 1.69 Impact Factor
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ABSTRACT: The hemolysin from Prevotella intermedia was partially purified from culture supernatant and then characterized. The hemolysin produced a clear beta-hemolytic zone on a blood agar plate. Hemolytic activity was 2.5-fold greater in culture supernatant compared to that cell-associated. The isolation and purification procedure involved ammonium sulfate and polyethylene glycol precipitations and ion-exchange chromatographies on DEAE-Sephacel and CM-Sepharose. The activity of this hemolysin was stimulated by reductants such as cysteine, dithiothreitol, glutathione etc., and was lost upon oxidation. Trypsin or heat treatment resulted in complete inhibition of hemolytic activity. Ca(2+), Mg(2+) and EDTA did not affect the activity. The optimal pH of this hemolysin was 7.5.
FEMS Immunology & Medical Microbiology 02/2003; 35(1):43-7. · 2.44 Impact Factor
