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1.
J Biol Chem ; 297(3): 101038, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34343567

RESUMO

Modular protein assembly has been widely reported as a mechanism for constructing allosteric machinery. Recently, a distinctive allosteric system has been identified in a bienzyme assembly comprising a 3-deoxy-d-arabino heptulosonate-7-phosphate synthase (DAH7PS) and chorismate mutase (CM). These enzymes catalyze the first and branch point reactions of aromatic amino acid biosynthesis in the bacterium Prevotella nigrescens (PniDAH7PS), respectively. The interactions between these two distinct catalytic domains support functional interreliance within this bifunctional enzyme. The binding of prephenate, the product of CM-catalyzed reaction, to the CM domain is associated with a striking rearrangement of overall protein conformation that alters the interdomain interactions and allosterically inhibits the DAH7PS activity. Here, we have further investigated the complex allosteric communication demonstrated by this bifunctional enzyme. We observed allosteric activation of CM activity in the presence of all DAH7PS substrates. Using small-angle X-ray scattering (SAXS) experiments, we show that changes in overall protein conformations and dynamics are associated with the presence of different DAH7PS substrates and the allosteric inhibitor prephenate. Furthermore, we have identified an extended interhelix loop located in CM domain, loopC320-F333, as a crucial segment for the interdomain structural and catalytic communications. Our results suggest that the dual-function enzyme PniDAH7PS contains a reciprocal allosteric system between the two enzymatic moieties as a result of this bidirectional interdomain communication. This arrangement allows for a complex feedback and feedforward system for control of pathway flux by connecting the initiation and branch point of aromatic amino acid biosynthesis.


Assuntos
3-Desoxi-7-Fosfo-Heptulonato Sintase/química , 3-Desoxi-7-Fosfo-Heptulonato Sintase/metabolismo , Aminoácidos Aromáticos/biossíntese , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Prevotella nigrescens/metabolismo , 3-Desoxi-7-Fosfo-Heptulonato Sintase/genética , Regulação Alostérica , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Vias Biossintéticas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Prevotella nigrescens/química , Prevotella nigrescens/enzimologia , Prevotella nigrescens/genética , Domínios Proteicos , Espalhamento a Baixo Ângulo , Alinhamento de Sequência
2.
Int J Med Sci ; 18(2): 432-440, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33390812

RESUMO

Prevotella nigrescens is an oral pathogen that is frequently observed in the subgingival plaque of periodontitis patients. Interleukin-1ß (IL-1ß) is known to be involved in the immunopathology of periodontal diseases and has been implicated in the destruction of bone. In this study, we investigated the mechanism of IL-1ß production by P. nigrescens in murine bone marrow-derived dendritic cells (BMDCs). Our results showed that a host receptor, Toll-like receptor 2 (TLR2), but not TLR4 is required for pro-IL-1ß induction and nucleotide-binding oligomerization domain like receptor pyrin domain containing 3 (NLRP3) priming in BMDCs in response to P. nigrescens and activation of the NLRP3 inflammasome is necessary for processing of pro-IL-1ß into mature IL-1ß. In addition, an inhibitor assay revealed that production of reactive oxygen species, P2X7R activity, and release of cathepsin B are involved in IL-1ß production in BMDCs in response to P. nigrescens.


Assuntos
Infecções por Bactérias Gram-Negativas/imunologia , Interleucina-1beta/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Periodontite/imunologia , Prevotella nigrescens/imunologia , Receptor 2 Toll-Like/metabolismo , Animais , Catepsina B/metabolismo , Células Cultivadas , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Infecções por Bactérias Gram-Negativas/microbiologia , Humanos , Inflamassomos/imunologia , Inflamassomos/metabolismo , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Periodontite/microbiologia , Cultura Primária de Células , Espécies Reativas de Oxigênio/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 2 Toll-Like/genética
3.
ACS Appl Bio Mater ; 4(3): 2523-2533, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-35014370

RESUMO

The beneficial effects of Sr- and Mg-doped hydroxyapatite (HAp) on osteoblast proliferation and bone regeneration have been investigated in the past, and the antibacterial ability of Zn ions is well known. However, HAp coatings doped with these three elements via thermal spraying have not yet been investigated. In this study, HAp powder was synthesized at different pH values (4, 6, 8, and 10) and calcined at different temperatures (200, 400, 600, 800, and 1000 °C) to obtain HAp with the highest purity. Subsequently, strontium-, magnesium-, and zinc-doped HAp powders were synthesized at the optimal pH value and calcination temperature. The HAp powder was then coated onto Ti disks using atmospheric plasma spraying (APS) or vapor-induced pore-forming atmospheric plasma spraying (VIPF-APS) techniques at different working currents (350, 400, and 450 A) and spraying distances (10 and 15 cm). X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron microscopy equipped with energy-dispersive spectroscopy were used for material characterization to determine the optimal parameters. With these optimal coating parameters, HAp, Zn-HAp, SrMg-HAp, and ZnSrMg-HAp powders were deposited onto the Ti disks using VIPF-APS and named HAp-Ti, Zn-HAp-Ti, SrMg-HAp-Ti, and ZnSrMg-HAp-Ti, respectively. The in vitro bioactivity of these four groups was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and alkaline phosphatase (ALPase) activity assay. Besides, the antibacterial activities against Prevotella nigrescens, Porphyromonas gingivalis, and Fusobacterium nucleatum were assessed. The results showed that the purity of HAp synthesized at pH 10 and 800 °C was 98.40%. A porous coating without cracks was obtained at a 10 cm spraying distance and 400 A working current using VIPF-APS. SrMg-HAp-Ti and ZnSrMg-HAp-Ti resulted in higher osteoblast proliferation and ALPase activity than the control. Moreover, both Zn-HAp-Ti and ZnSrMg-HAp-Ti exhibited antibacterial activity against the three bacteria. Therefore, ZnSrMg-HAp has potential as a coating for biomedical materials due to its ability to reduce bacterial infection and enhance osseointegration.


Assuntos
Antibacterianos/farmacologia , Atmosfera/química , Materiais Revestidos Biocompatíveis/farmacologia , Fusobacterium nucleatum/efeitos dos fármacos , Porphyromonas gingivalis/efeitos dos fármacos , Prevotella nigrescens/efeitos dos fármacos , Antibacterianos/síntese química , Antibacterianos/química , Materiais Revestidos Biocompatíveis/síntese química , Materiais Revestidos Biocompatíveis/química , Durapatita/química , Durapatita/farmacologia , Magnésio/química , Magnésio/farmacologia , Teste de Materiais , Testes de Sensibilidade Microbiana , Tamanho da Partícula , Porosidade , Estrôncio/química , Estrôncio/farmacologia , Propriedades de Superfície , Zinco/química , Zinco/farmacologia
4.
J Cyst Fibros ; 19(2): 211-218, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31607634

RESUMO

Prevotella spp. are frequently identified in Cystic Fibrosis sputum. This study examined whether infection with Prevotella nigrescens, a frequently identified member of this species, contributes to inflammation in CF bronchial epithelial cells through activation of TLR- and NF-κB signalling pathways. CFBE41o- cells were infected with either P.nigrescens or Pseudomonas aeruginosa and incubated under anaerobic conditions for 4h. P.nigrescens activated TLR2 signalling but not TLR4 signalling while P.aeruginosa activated TLR4 signalling with a lesser effect on TLR2. P.aeruginosa induced significant IκBα phosphorylation 10min post infection with a return to control levels by 30min post infection. A significant induction in nuclear p65 DNA binding was observed at 2h post infection. In contrast, infection with P.nigrescens induced phosphorylation of IκBα 120min post infection, with significant induction in nuclear p65 DNA binding at 4h post infection only. Cytokine gene and protein responses were lower for P.nigrescens compared to P.aeruginosa. This study demonstrates the ability of a clinical P.nigrescens isolate to provoke a delayed NF-κB(p65) driven response through induction in TLR2 signalling and activation of sustained levels of IKKα.


Assuntos
Fibrose Cística , Prevotella nigrescens/fisiologia , Pseudomonas aeruginosa/fisiologia , Mucosa Respiratória , Receptor 4 Toll-Like/metabolismo , Fator de Transcrição RelA/metabolismo , Bactérias Anaeróbias , Células Cultivadas , Fibrose Cística/imunologia , Fibrose Cística/microbiologia , Interações Hospedeiro-Patógeno , Humanos , Inflamação/metabolismo , NF-kappa B/metabolismo , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , Transdução de Sinais , Receptor 2 Toll-Like/metabolismo
5.
Braz. arch. biol. technol ; 63: e20190196, 2020. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1132166

RESUMO

Abstract Endodontic infections result from oral pathogenic bacteria which reach and infect dental pulp, as well as surrounding tissues, through cracks, unrepaired caries and failed caries restorations. This study aims to determine the chemical composition of essential oil from Psidium cattleianum leaves (PC-EO) and to assess its antibacterial activity against endodontic bacteria. Antibacterial activity of PC-EO was evaluated in terms of its minimum inhibitory concentration (MIC) values by the broth microdilution method on 96-well microplates. Bacteria Porphyromonas gingivalis (MIC = 20 µg/mL), Prevotella nigrescens (MIC = 62.5 µg/mL), Fusobacterium nucleatum (MIC = 12.5 µg/mL), Actinomyces naeslundii (MIC = 50 µg/mL), Bacteroides fragilis (MIC = 12.5 µg/mL), Aggregatibacter actinomycetemcomitans (MIC = 6.25 µg/mL) and Peptostreptococcus anaerobius (MIC = 62.5 µg/mL) were evaluated and compared to chlorhexidine dihydrochloride (CDH), the positive control. PC-EO was obtained by hydrodistillation with the use of a Clevenger-type apparatus whereas its chemical composition was analyzed by gas chromatography-flame ionization detection (GC-FID) and gas chromatography-mass spectrometry (GC-MS). Viridiflorol (17.9%), β-caryophyllene (11.8%), 1,8-cineole (10.8%) and β-selinene (8.6%) were the major constituents found in PC-EO, which exhibited high antibacterial activity against all endodontic pathogens under investigation. Therefore, PC-EO, a promising source of bioactive compounds, may provide therapeutic solutions for the field of endodontics.


Assuntos
Óleos Voláteis/farmacologia , Clorexidina/farmacologia , Psidium/química , Antibacterianos/farmacologia , Peptostreptococcus/efeitos dos fármacos , Bacteroides fragilis/efeitos dos fármacos , Actinomyces/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Fusobacterium nucleatum/efeitos dos fármacos , Aggregatibacter actinomycetemcomitans/efeitos dos fármacos , Porphyromonas gingivalis/efeitos dos fármacos , Prevotella nigrescens/efeitos dos fármacos , Cromatografia Gasosa-Espectrometria de Massas
6.
BMC Res Notes ; 12(1): 328, 2019 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-31182149

RESUMO

OBJECTIVE: The aim of the present study is to compare and assess the risk of periodontitis due to the presence of four putative periodontopathic bacteria viz., Eikenella corrodens, Campylobacter rectus, Prevotella intermedia and Prevotella nigrescens. To fulfil the above objective, polymerase Chain reaction using the primers targeting 16S rRNA gene of the bacterial species was performed with the subgingival plaque collected from the permanent first molars of type 1 diabetic children and age matched healthy children. RESULTS: The prevalence of periodontal pathogens in diabetic and healthy children was 6% and 16% for E. corrodens, 18% and 36% for C. rectus, 2% and 2% for P. intermedia, 4% and 0%, for P. nigrescens respectively. Statistically, significant difference was not observed for the prevalence of all the four periodontal pathogens between type 1 diabetic and healthy children (P = 1.00). The results of the present study thus reveal a negative correlation of type I diabetes to periodontitis in association to Eikenella corrodens, Campylobacter rectus, Prevotella intermedia and Prevotella nigrescens.


Assuntos
Campylobacter rectus/genética , Placa Dentária/microbiologia , Diabetes Mellitus Tipo 1/microbiologia , Eikenella corrodens/genética , Periodontite/microbiologia , Prevotella intermedia/genética , Prevotella nigrescens/genética , Adolescente , Técnicas de Tipagem Bacteriana , Campylobacter rectus/classificação , Campylobacter rectus/isolamento & purificação , Estudos de Casos e Controles , Criança , Placa Dentária/complicações , Placa Dentária/diagnóstico , Placa Dentária/patologia , Índice de Placa Dentária , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 1/patologia , Eikenella corrodens/classificação , Eikenella corrodens/isolamento & purificação , Feminino , Humanos , Masculino , Periodontite/complicações , Periodontite/diagnóstico , Periodontite/patologia , Prevotella intermedia/classificação , Prevotella intermedia/isolamento & purificação , Prevotella nigrescens/classificação , Prevotella nigrescens/isolamento & purificação , RNA Ribossômico 16S/genética
7.
J Biol Chem ; 294(13): 4828-4842, 2019 03 29.
Artigo em Inglês | MEDLINE | ID: mdl-30670586

RESUMO

Because of their special organization, multifunctional enzymes play crucial roles in improving the performance of metabolic pathways. For example, the bacterium Prevotella nigrescens contains a distinctive bifunctional protein comprising a 3-deoxy-d-arabino heptulosonate-7-phosphate synthase (DAH7PS), catalyzing the first reaction of the biosynthetic pathway of aromatic amino acids, and a chorismate mutase (CM), functioning at a branch of this pathway leading to the synthesis of tyrosine and phenylalanine. In this study, we characterized this P. nigrescens enzyme and found that its two catalytic activities exhibit substantial hetero-interdependence and that the separation of its two distinct catalytic domains results in a dramatic loss of both DAH7PS and CM activities. The protein displayed a unique dimeric assembly, with dimerization solely via the CM domain. Small angle X-ray scattering (SAXS)-based structural analysis of this protein indicated a DAH7PS-CM hetero-interaction between the DAH7PS and CM domains, unlike the homo-association between DAH7PS domains normally observed for other DAH7PS proteins. This hetero-interaction provides a structural basis for the functional interdependence between the two domains observed here. Moreover, we observed that DAH7PS is allosterically inhibited by prephenate, the product of the CM-catalyzed reaction. This allostery was accompanied by a striking conformational change as observed by SAXS, implying that altering the hetero-domain interaction underpins the allosteric inhibition. We conclude that for this C-terminal CM-linked DAH7PS, catalytic function and allosteric regulation appear to be delivered by a common mechanism, revealing a distinct and efficient evolutionary strategy to utilize the functional advantages of a bifunctional enzyme.


Assuntos
Alquil e Aril Transferases/química , Aminoácidos Aromáticos/biossíntese , Proteínas de Bactérias/química , Prevotella nigrescens/enzimologia , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Regulação Alostérica , Aminoácidos Aromáticos/química , Aminoácidos Aromáticos/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Catálise , Cristalografia por Raios X , Prevotella nigrescens/genética , Domínios Proteicos , Espalhamento a Baixo Ângulo , Difração de Raios X
8.
Skeletal Radiol ; 48(8): 1305-1309, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30683976

RESUMO

Infection of costal cartilage is a rare observation. We report the case of a 43-year-old male patient without relevant history who presented with a progressive painful swelling of the left chest wall since 4 months. Computed tomography (CT) and magnetic resonance imaging (MRI) demonstrated an abscess within the left ninth costal cartilage with surrounding reactive changes. A CT-guided biopsy was performed and the culture of the sample revealed the presence of Prevotella nigrescens. Musculoskeletal infections by Prevotella are rarely described in the literature, Prevotella oralis and Prevotella bivia being the most frequently observed pathogens. These infections usually originate from a hematogenous spread after thoracic surgery or dental procedure. In our patient, conservative treatment was chosen. A clinical improvement was noted after 1-month antibiotherapy, confirmed by short-term and 6-month imaging follow-up showing the complete disappearance of all previously observed abnormalities.


Assuntos
Infecções por Bactérias Gram-Negativas/diagnóstico , Infecções por Bactérias Gram-Negativas/terapia , Prevotella nigrescens , Síndrome de Tietze/diagnóstico , Síndrome de Tietze/microbiologia , Adulto , Humanos , Masculino , Síndrome de Tietze/terapia
9.
Photodiagnosis Photodyn Ther ; 25: 92-102, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30468900

RESUMO

BACKGROUND: The aim of this study was to evaluate the effect of multiple sessions of antimicrobial photodynamic therapy (aPDT) on the treatment of experimental periodontitis (EP) in rats treated with chemotherapy. METHODS: Chemotherapy using 5-fluorouracil (5-FU) consisted of intraperitoneal administration of 60 and 40 mg/kg of 5-FU. 120 rats were subjected to chemotherapy with 5-FU and divided into groups: PT (periodontal treatment); PT+1aPDT (PT and single aPDT session); PT+4aPDT(PT and 4 sessions of aPDT); 1aPDT (single aPDT session); 4aPDT(4 sessions of aPDT). EP was induced in the mandibular molars via ligature placement. The alveolar bone loss (ABL) area in the furcation region was analysed histometrically. Proliferating cell nuclear antigen (PCNA), tartrate-resistant acid phosphatase (TRAP), receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG) and cleaved caspase-3 (CC3) were analysed by immunohistochemistry. Prostaglandin E2 was quantified using an ELISA, tumor necrosis factor (TNF)-α and interleukin (IL)-6 were assessed using a multiplex method. The prevalence of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Prevotella nigrescens, Prevotella intermedia and Fusobacterium nucleatum was assessed using PCR. The data were statistically analysed (P < 0.05). RESULTS: The PT+4aPDT group showed lower ABL than the PT or 1aPDT groups on day 7. Rats treated with aPDT showed a higher number of PCNA-positive cells with reduced immunolabeling of RANKL. Significant reductions in Prevotella nigrescens were observed in the PT+4aPDT group and in Aggregatibacter actinomycetemcomitans for the 1aPDT and 4aPDT groups. CONCLUSION: Repeated sessions of aPDT as an adjunct or alternative therapy were effective at reducing ABL, regulating bone metabolism, and reducing Prevotella nigrescens and Aggregatibacter actinomycetemcomitans.


Assuntos
Fluoruracila/farmacologia , Periodontite/tratamento farmacológico , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/farmacologia , Aggregatibacter actinomycetemcomitans/efeitos dos fármacos , Perda do Osso Alveolar/terapia , Animais , Raspagem Dentária/métodos , Modelos Animais de Doenças , Interleucina-6/biossíntese , Masculino , Doenças Mandibulares , Periodontite/terapia , Prevotella nigrescens/efeitos dos fármacos , Ratos , Fator de Necrose Tumoral alfa/biossíntese
10.
Anaerobe ; 54: 128-135, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30189320

RESUMO

Quorum sensing (QS) signaling regulates the motility, adhesion, and biofilm formation of bacteria, and at the same time activates immune response in eukaryotic organisms. We recently demonstrated that the QS molecule, dihydroxy-2, 3-pentanedione (DPD), and its analogs significantly inhibit estradiol-regulated virulence of Prevotella aurantiaca, one of the four species in the Prevotella intermedia group. Here, we examined the combined effects of estradiol and QS signaling on 1) cytokine response of human gingival keratinocytes (HMK) against whole cell extract (WCE) of P. intermedia, Prevotella nigrescens, and Prevotella pallens, and 2) biofilm formation of these three Prevotella species. All experiments were performed in the presence or absence of estradiol, and with different QS molecules: DPD and its analogs (ethyl-DPD, butyl-DPD, and isobutyl-DPD). Concentrations of interleukin (IL)-1ß, -6, and -8 were determined by the Luminex multiplex immunoassay, biofilm mass was quantitatively evaluated by measuring protein concentration via the Bradford method, and the microtopography of biofilms was assessed by scanning electron microscopy (SEM) imaging. Concentrations of IL-6 and IL-8 were elevated when HMK cells were incubated with estradiol and WCE of P. intermedia and P. nigrescens, but decreased when incubated with estradiol and WCE of P. pallens. Butyl-DPD neutralized the estradiol- and WCE-induced regulation of HMK interleukin expression and, at the same time, inhibited the biofilm formation of P. intermedia and P. nigrescens. SEM micrographs revealed a decrease in biofilm mass after application of butyl-DPD, which was most detectable among the P. intermedia ATCC 25611 and P. nigrescens ATCC 33563 and AHN 8293 strains. In conclusion, butyl-DPD analog is able to neutralize the WCE-induced epithelial cytokine response and, at the same time, to inhibit the biofilm formation of P. intermedia and P. nigrescens.


Assuntos
Infecções por Bacteroidaceae/imunologia , Células Epiteliais/imunologia , Gengiva/imunologia , Interleucina-1beta/imunologia , Interleucina-6/imunologia , Interleucina-8/imunologia , Prevotella/fisiologia , Percepção de Quorum , Infecções por Bacteroidaceae/genética , Infecções por Bacteroidaceae/microbiologia , Biofilmes , Células Epiteliais/microbiologia , Gengiva/microbiologia , Humanos , Interleucina-1beta/genética , Interleucina-6/genética , Interleucina-8/genética , Queratinócitos/imunologia , Queratinócitos/microbiologia , Prevotella/classificação , Prevotella/genética , Prevotella/patogenicidade , Prevotella intermedia/genética , Prevotella intermedia/patogenicidade , Prevotella intermedia/fisiologia , Prevotella nigrescens/genética , Prevotella nigrescens/patogenicidade , Prevotella nigrescens/fisiologia
11.
Anaerobe ; 54: 201-204, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29913204

RESUMO

The accuracy of a phenotypic scheme to recognize periodontal Prevotella intermedia/nigrescens group clinical isolates on primary isolation culture plates was assessed with matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS). A total of 84 fresh subgingival isolates from 23 chronic periodontitis patients were presumptively recognized on anaerobically-incubated enriched Brucella blood agar primary isolation plates as P. intermedia/nigrescens based on their dark-pigmented colony morphology, brick-red autofluorescence under long-wave ultraviolet light, and a negative fluorescence test for lactose production. The presumptive P. intermedia/nigrescens clinical isolates were subjected to MALDI-TOF MS analysis using Bruker MALDI Biotyper analytic software containing mass spectra for P. intermedia and Prevotella nigrescens in its reference library of bacterial protein profiles. Using a ≥1.7 log score agreement threshold, 60 (71.4%) of the presumptive P. intermedia/nigrescens clinical isolates were confirmed as either P. intermedia (25 isolates) or P. nigrescens (35 isolates). All isolates with a <1.7 log score were also identified as P. intermedia or P. nigrescens from the top choice designated on the MALDI Biotyper most likely species identification list. These MALDI-TOF MS findings document the ability of the phenotypic scheme to correctly recognize most periodontal P. intermedia/nigrescens group clinical isolates on primary isolation culture plates.


Assuntos
Técnicas de Tipagem Bacteriana/métodos , Infecções por Bacteroidaceae/microbiologia , Periodontite Crônica/microbiologia , Prevotella intermedia/isolamento & purificação , Prevotella nigrescens/isolamento & purificação , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Adulto , Infecções por Bacteroidaceae/diagnóstico , Periodontite Crônica/diagnóstico , Feminino , Humanos , Masculino , Fenótipo , Prevotella intermedia/química , Prevotella intermedia/genética , Prevotella nigrescens/química , Prevotella nigrescens/genética
12.
Artigo em Inglês | MEDLINE | ID: mdl-28983469

RESUMO

High-throughput sequencing has helped to reveal the close relationship between Prevotella and periodontal disease, but the roles of subspecies diversity and genomic variation within this genus in periodontal diseases still need to be investigated. We performed a comparative genome analysis of 48 Prevotella intermedia and Prevotella nigrescens isolates that from the same cohort of subjects to identify the main drivers of their pathogenicity and adaptation to different environments. The comparisons were done between two species and between disease and health based on pooled sequences. The results showed that both P. intermedia and P. nigrescens have highly dynamic genomes and can take up various exogenous factors through horizontal gene transfer. The major differences between disease-derived and health-derived samples of P. intermedia and P. nigrescens were factors related to genome modification and recombination, indicating that the Prevotella isolates from disease sites may be more capable of genomic reconstruction. We also identified genetic elements specific to each sample, and found that disease groups had more unique virulence factors related to capsule and lipopolysaccharide synthesis, secretion systems, proteinases, and toxins, suggesting that strains from disease sites may have more specific virulence, particularly for P. intermedia. The differentially represented pathways between samples from disease and health were related to energy metabolism, carbohydrate and lipid metabolism, and amino acid metabolism, consistent with data from the whole subgingival microbiome in periodontal disease and health. Disease-derived samples had gained or lost several metabolic genes compared to healthy-derived samples, which could be linked with the difference in virulence performance between diseased and healthy sample groups. Our findings suggest that P. intermedia and P. nigrescens may serve as "crucial substances" in subgingival plaque, which may reflect changes in microbial and environmental dynamics in subgingival microbial ecosystems. This provides insight into the potential of P. intermedia and P. nigrescens as new predictive biomarkers and targets for effective interventions in periodontal disease.


Assuntos
Infecções por Bacteroidaceae/microbiologia , Periodontite/microbiologia , Prevotella intermedia/genética , Prevotella nigrescens/genética , Adulto , Sequência de Bases , Estudos de Coortes , Feminino , Transferência Genética Horizontal , Variação Genética , Genoma Bacteriano , Gengiva/microbiologia , Gengiva/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Prevotella intermedia/isolamento & purificação , Prevotella nigrescens/isolamento & purificação , Fatores de Virulência/genética
13.
J Periodontol ; 88(5): 511-517, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28067105

RESUMO

BACKGROUND: Various antimicrobial agents are widely used in the therapy of oral inflammatory diseases. However, their side effects and the appearance of drug resistance justify research on natural antimicrobial agents to target oral pathogens that are safe for the host. In the present study, antimicrobial properties of mastic extract on commensal and pathogenic oral bacteria, as well as its possible cytotoxic effect toward cells of epithelial and mesenchymal origin, were evaluated and compared with the common antimicrobial agents hydrogen peroxide (H2O2) and chlorhexidine digluconate (CHX). METHODS: Oral and periodontal pathogens (Porphyromonas gingivalis, Streptococcus mutans [Sm], Streptococcus oralis, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, Prevotella intermedia, and Prevotella nigrescens) were treated with different concentrations of mastic extract, 3% H2O2, and 0.2% CHX, and evaluated with an agar diffusion test. The cytotoxic effect of mastic extract was tested on four cell lines of epithelial and mesenchymal origin (HaCaT, SaOS-2, MC3T3-E1, periodontal ligament [PDL] cells) by neutral red and 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide assay. RESULTS: Mastic extract led to significantly (P ≤0.016) increased inhibition of the tested periodontal pathogens compared with H2O2. No effect of mastic extract was observed on Sm. Mastic extract showed beneficial effects on cell viability because viability values of tested cells were significantly (P ≤0.016) lower for cells treated with CHX and H2O2 compared with mastic extract-treated cells after stimulation for 2, 4, and 6 hours. CONCLUSION: The present data demonstrate mastic extract's inhibition of periodontal pathogens, as well as beneficial effects on cell viability, compared with H2О2, suggesting that it could be considered an alternative antibacterial agent in the prevention of periodontal disease.


Assuntos
Anti-Infecciosos/farmacologia , Periodontite/tratamento farmacológico , Fitoterapia/métodos , Pistacia/química , Extratos Vegetais/farmacologia , Animais , Anti-Infecciosos/uso terapêutico , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Fusobacterium nucleatum/efeitos dos fármacos , Humanos , Camundongos , Periodontite/microbiologia , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Caules de Planta/química , Porphyromonas gingivalis/efeitos dos fármacos , Prevotella intermedia/efeitos dos fármacos , Prevotella nigrescens/efeitos dos fármacos , Streptococcus mutans/efeitos dos fármacos , Streptococcus oralis/efeitos dos fármacos
14.
J Biomed Opt ; 21(8): 84003, 2016 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-27552307

RESUMO

A number of anaerobic oral bacteria, notably Prevotellaceae, exhibit red fluorescence when excited by short-wavelength visible light due to their accumulation of porphyrins, particularly protoporphyrin IX. pH affects the fluorescence of abiotic preparations of porphyrins due to transformations in speciation between monomers, higher aggregates, and dimers. To elucidate whether the porphyrin speciation phenomenon could be manifested within a microbiological system, suspensions of Prevotella intermedia and Prevotella nigrescens were examined by fluorescence spectrophotometry while being titrated against NaOH. The initial pH of the samples was <6, which was then raised toward the maximum found within a diseased periodontal pocket, being ∼pH 8.7. The intensity of the fluorescence emissions increased between 600 and 650 nm with increasing pH. Peak fluorescence emissions occurred at 635±1 nm with a second emission peak developing with increasing pH at 622 nm. A linear relationship was demonstrated between pH and the log10 ratio of 635:622 nm excitation fluorescence intensities. These findings suggest that the pH range found within the oral cavity could affect the fluorescence of oral bacteria in vivo, which may in turn have connotations for any clinical diagnoses that may be inferred from dental plaque fluorescence.


Assuntos
Placa Dentária/diagnóstico , Placa Dentária/microbiologia , Prevotella nigrescens/fisiologia , Humanos , Concentração de Íons de Hidrogênio , Porfirinas/química , Prevotella intermedia/fisiologia , Espectrometria de Fluorescência
15.
Anaerobe ; 40: 18-27, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27118478

RESUMO

Oral infections such as periodontitis and tooth decay are the most common diseases of humankind. Oleoresins from different copaifera species display antimicrobial and anti-inflammatory activities. Copaifera reticulata is the commonest tree of this genus and grows abundantly in several Brazilian states, such as Pará, Amazonas, and Ceará. The present study has evaluated the chemical composition and antimicrobial potential of the Copaifera reticulata oleoresin (CRO) against the causative agents of tooth decay and periodontitis and has assessed the CRO cytotoxic potential. Cutting edge analytical techniques (GC-MS and LC-MS) aided the chemical characterization of CRO. Antimicrobial assays included determination of the Minimum Inhibitory Concentration (MIC), determination of the Minimum Bactericidal Concentration (MBC), determination of the Minimum Inhibitory Concentration of Biofilm (MICB50), Time Kill Assay, and Checkerboard Dilution. Conduction of XTT assays on human lung fibroblasts (GM07492-A cells) helped to examine the CRO cytotoxic potential. Chromatographic analyses revealed that the major constituents of CRO were ß-bisabolene, trans-α-bergamotene, ß-selinene, α-selinene, and the terpene acids ent-agathic-15-methyl ester, ent-copalic acid, and ent-polyalthic acid. MIC and MBC results ranged from 6.25 to 200 µg/mL against the tested bacteria. The time-kill assay conducted with CRO at concentrations between 50 and 100 µg/mL showed bactericidal activity against Fusobacterium nucleatum (ATCC 25586) and Streptococcus mitis (ATCC 49456) after 4 h, Prevotella nigrescens (ATCC 33563) after 6 h, Porphyromonas gingivalis (ATCC 33277) and Lactobacillus casei (clinical isolate) after 12 h, and Streptococcus salivarius (ATCC 25975) and Streptococcus mutans (ATCC 25175) after 18 h. The fractional inhibitory concentration indexes (FICIs) revealed antagonistic interaction for Lactobacillus casei (clinical isolate), indifferent effect for Porphyromonas gingivalis (ATCC 33277), Fusobacterium nucleatum (ATCC 25586), Prevotella nigrescens (ATCC 33563), and Streptococcus salivarius (ATCC 25975), and additive effect for Streptococcus mutans (ATCC 25175) and Streptococcus mitis (ATCC 49456). Treatment of GM07492-A cells with CRO demonstrated that concentrations up to 39 µg/mL significantly reduced cell viability as compared to the negative control, being IC50 equal to 51.85 ± 5.4 µg/mL. These results indicated that CRO plays an important part in the search for novel sources of agents that can act against oral pathogens.


Assuntos
Antibacterianos/farmacologia , Fabaceae/química , Extratos Vegetais/farmacologia , Porphyromonas gingivalis/efeitos dos fármacos , Prevotella nigrescens/efeitos dos fármacos , Antibacterianos/química , Antibacterianos/isolamento & purificação , Compostos Bicíclicos com Pontes/isolamento & purificação , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cárie Dentária/microbiologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Humanos , Lactobacillus casei/efeitos dos fármacos , Lactobacillus casei/crescimento & desenvolvimento , Lactobacillus casei/isolamento & purificação , Testes de Sensibilidade Microbiana , Sesquiterpenos Monocíclicos , Periodontite/microbiologia , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Porphyromonas gingivalis/crescimento & desenvolvimento , Porphyromonas gingivalis/isolamento & purificação , Prevotella nigrescens/crescimento & desenvolvimento , Prevotella nigrescens/isolamento & purificação , Sesquiterpenos/isolamento & purificação , Streptococcus mutans/efeitos dos fármacos , Streptococcus mutans/crescimento & desenvolvimento , Streptococcus mutans/isolamento & purificação , Streptococcus salivarius/efeitos dos fármacos , Streptococcus salivarius/crescimento & desenvolvimento , Streptococcus salivarius/isolamento & purificação , Terpenos/isolamento & purificação
16.
Anaerobe ; 39: 91-6, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26996070

RESUMO

Rheumatoid arthritis (RA) and periodontitis (PD) are chronic inflammatory disorders that cause bone loss. PD tends to be more prevalent and severe in RA patients. Previous experimental studies demonstrated that RA triggers alveolar bone loss similarly to PD. The aim of this study was to investigate if arthritis-induced alveolar bone loss is associated with modification in the oral microbiota. Checkerboard DNA-DNA hybridization was employed to analyze forty oral bacterial species in 3 groups of C57BL/6 mice: control (n = 12; without any challenge); Y4 (n = 8; received oral inoculation of Aggregatibacter Actinomycetemcomitans strain FDC Y4) and AIA group (n = 12; chronic antigen-induced arthritis). The results showed that AIA and Y4 group exhibited similar patterns of bone loss. The AIA group exhibited higher counts of most bacterial species analyzed with predominance of Gram-negative species similarly to infection-induced PD. Prevotella nigrescens and Treponema denticola were detected only in the Y4 group whereas Campylobacter showae, Streptococcus mitis and Streptococcus oralis were only found in the AIA group. Counts of Parvimonas micra, Selenomonas Noxia and Veillonella parvula were greater in the AIA group whereas Actinomyces viscosus and Neisseira mucosa were in large proportion in Y4 group. In conclusion, AIA is associated with changes in the composition of the oral microbiota, which might account for the alveolar bone loss observed in AIA mice.


Assuntos
Perda do Osso Alveolar/microbiologia , Processo Alveolar/microbiologia , Artrite Experimental/microbiologia , Maxila/microbiologia , Microbiota/genética , Periodontite/microbiologia , Aggregatibacter actinomycetemcomitans/classificação , Aggregatibacter actinomycetemcomitans/genética , Aggregatibacter actinomycetemcomitans/isolamento & purificação , Perda do Osso Alveolar/patologia , Processo Alveolar/patologia , Animais , Artrite Experimental/patologia , Campylobacter/classificação , Campylobacter/genética , Campylobacter/isolamento & purificação , DNA Bacteriano/genética , Humanos , Masculino , Maxila/patologia , Camundongos , Camundongos Endogâmicos C57BL , Boca/microbiologia , Boca/patologia , Periodontite/patologia , Prevotella nigrescens/classificação , Prevotella nigrescens/genética , Prevotella nigrescens/isolamento & purificação , Streptococcus mitis/classificação , Streptococcus mitis/genética , Streptococcus mitis/isolamento & purificação , Streptococcus oralis/classificação , Streptococcus oralis/genética , Streptococcus oralis/isolamento & purificação , Treponema denticola/classificação , Treponema denticola/genética , Treponema denticola/isolamento & purificação
17.
Nat Prod Res ; 30(10): 1178-81, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26165725

RESUMO

This work investigated the antibacterial activity of 15 commercially available plant-derived essential oils (EOs) against a panel of oral pathogens. The broth microdilution method afforded the minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of the assayed EOs. The EO obtained from Cinnamomum zeylanicum (Lauraceae) (CZ-EO) displayed moderate activity against Fusobacterium nucleatum (MIC and MBC = 125 µg/mL), Actinomyces naeslundii (MIC and MBC = 125 µg/mL), Prevotella nigrescens (MIC and MBC = 125 µg/mL) and Streptococcus mutans (MIC = 200 µg/mL; MBC = 400 µg/mL). (Z)-isosafrole (85.3%) was the main chemical component of this oil. We did not detect cinnamaldehyde, previously described as the major constituent of CZ-EO, in specimens collected in other countries.


Assuntos
Antibacterianos/farmacologia , Cinnamomum zeylanicum/química , Óleos Voláteis/farmacologia , Óleos Vegetais/farmacologia , Actinomyces/efeitos dos fármacos , Fusobacterium nucleatum/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Boca/microbiologia , Prevotella nigrescens/efeitos dos fármacos , Streptococcus mutans/efeitos dos fármacos
18.
Indian J Dent Res ; 26(2): 189-95, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26096116

RESUMO

BACKGROUND: The study was aimed to assess the prevalence of periodontal pathogens namely Tannerella forsythia (T.f), Campylobacter rectus (C.r), Eikenella corrodens (E.c), Porphyromonas gingivalis (P.g), Treponema denticola (T.d), Prevotella nigrescens (P.n) ,Aggregatibacter actinomycetemcomitans (A.a), P.g (fi mA gene) and Prevotella intermedia (P.i), in the subgingival and the atheromatous plaque of patients with coronary artery disease (CAD), and comparing them with the noncardiac subjects thereby indicating the role of periodontal pathogenic bacteria in the progression of atherosclerosis in south Indian population. MATERIALS AND METHODS: 51 cardiac and non cardiac subjects within the age group of 40-80 years,who met the eligibility criteria, were selected and categorized as the experimental and control group respectively. Total number of teeth was recorded, and oral hygiene was evaluated using Plaque Index and Oral Hygiene Index (OHI). Periodontal pocket depth and clinical attachment level were also assessed as a part of periodontal examination. Subgingival plaque samples were collected with the help of with Gracey's curette in both the groups. In experimental group, biopsy was obtained from the atherosclerotic plaque during Coronary artery bypass grafting CABG procedure. Both, subgingival and the coronary atherosclerotic plaque samples were subjected to polymerase chain reaction (PCR) analysis for identification of the periodontal bacteria. STATISTICAL ANALYSIS: Mean, standard deviation and test of significance of quantitative variables such as periodontal parameters were compared between experimental group and control group. Kappa measures of agreement was done to analyze the relationship between the presence/absence of microorganisms in the subgingival and atherosclerotic plaque samples in the experimental group. RESULTS: The mean Plaque Index, Gingival Index, Russel's Periodontal Index, OHI, clinical attachment level, Pocket Depth Index was statistically significant in both the groups. Association of T.f, E.c, C.r, P.g, P.g (fi mA), T.d, P.i, P.n was significantly associated. A.a was absent in the control group, whereas only one patient in the experimental group was positive for the bacteria. Kappa analysis showed the significant association of periodontal bacteria T.f, C.r, P.g, P.g (fi mA), P.i and P.n in subgingival plaque and atherosclerotic plaque of the same patients of the experimental group. CONCLUSION: It is concluded that CAD subjects had higher prevalence of periodontal pathogens in subgingival biofilms as compared to the non cardiac subjects. Further, the number of bacteria was significantly associated between the subgingival and atherosclerotic plaques of the cardiac patients in south Indian population.


Assuntos
Doença da Artéria Coronariana/microbiologia , Placa Aterosclerótica/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Aggregatibacter actinomycetemcomitans/isolamento & purificação , Biópsia , Campylobacter rectus/isolamento & purificação , Estudos de Casos e Controles , Ponte de Artéria Coronária , Doença da Artéria Coronariana/cirurgia , Inquéritos de Saúde Bucal , Progressão da Doença , Eikenella corrodens/isolamento & purificação , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Porphyromonas gingivalis/isolamento & purificação , Prevalência , Prevotella intermedia/isolamento & purificação , Prevotella nigrescens/isolamento & purificação , Tannerella forsythia/isolamento & purificação , Treponema denticola/isolamento & purificação
19.
BMC Oral Health ; 15: 25, 2015 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-25888404

RESUMO

BACKGROUND: Aggressive periodontitis (AgP) is one of the most severe forms of periodontal diseases. In Morocco, Aggregatibacter actinomycetemcomitans has been strongly associated with AgP, however limited knowledge is available about the implication of other periodontal pathogens in this entity. Therefore, the main aim of this study was to evaluate the composition of the subgingival microbiota in Moroccan patients with AgP. METHODS: Subgingival plaque samples were collected from 50 aggressive, 13 localized and 37 generalized periodontitis patients. Samples from 20 chronic periodontitis (ChP) patients were taken as controls. Samples collected from the four deepest periodontal pockets in each patient were pooled in pre-reduced transport fluid and examined by culture. RESULTS: A. actinomycetemcomitans was significantly more frequent (p = 0.004) in generalised AgP compared to ChP, and Porphyromonas gingivalis was less prevalent in localized AgP, when compared with generalized AgP (p = 0.040) or ChP (p = 0.016). Prevotella intermedia, Fusobacterium nucleatum and Tannerella forsythia were also frequently detected in all groups. Mean proportions of A. actinomycetemcomitans were significantly higher in AgP groups, when compared to ChP, and generalized AgP patients harbored significantly higher proportions of P. gingivalis and T. forsythia, when compared to localized AgP or ChP. CONCLUSIONS: A. actinomycetemcomitans, P. gingivalis, T. forsythia, P. intermedia and F. nucleatum were frequently detected in this Moroccan population with AgP. Differences in frequency of detection, counts and proportions of A. actinomycetemcomitans, P. gingivalis and T. forsythia suggests the presence of distinct microbiological profiles for localized AgP, generalized AgP and ChP patients.


Assuntos
Periodontite Agressiva/microbiologia , Bactérias Gram-Negativas/isolamento & purificação , Adolescente , Adulto , Aggregatibacter actinomycetemcomitans/isolamento & purificação , Carga Bacteriana , Campylobacter rectus/isolamento & purificação , Capnocytophaga/isolamento & purificação , Periodontite Crônica/microbiologia , Estudos Transversais , Placa Dentária/microbiologia , Eikenella corrodens/isolamento & purificação , Eubacterium/isolamento & purificação , Feminino , Fusobacterium nucleatum/isolamento & purificação , Humanos , Masculino , Peptostreptococcus/isolamento & purificação , Bolsa Periodontal/microbiologia , Periodontite/microbiologia , Porphyromonas gingivalis/isolamento & purificação , Prevotella intermedia/isolamento & purificação , Prevotella nigrescens/isolamento & purificação , Tannerella forsythia/isolamento & purificação , Adulto Jovem
20.
J Endod ; 41(6): 817-23, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25771141

RESUMO

INTRODUCTION: This study investigated the presence of target bacterial species and the levels of endotoxins in teeth with apical periodontitis. Levels of inflammatory mediators (interleukin [IL]-1ß and tumor necrosis factor [TNF]-α) were determined after macrophage stimulation with endodontic content after different phases of endodontic therapy using different irrigants. METHODS: Thirty primarily infected root canals were randomly assigned into 3 groups according to the irrigant used for root canal preparation (n = 10 per group): GI: 2.5% sodium hypochlorite, GII: 2% chlorhexidine gel, and GIII (control group): saline solution. Root canal samples were taken by using paper points before (s1) and after root canal instrumentation (s2), subsequently to 17% EDTA (s3), after 30 days of intracanal medication (Ca[OH]2 + saline solution) (s4), and before root canal obturation (s5). Polymerase chain reaction (16S recombinant DNA) and limulus amebocyte lysate assay were used for bacterial and endotoxin detection, respectively. Macrophages were stimulated with the root canal contents for IL-1ß/TNF-α measurement using enzyme-linked immunosorbent assay. RESULTS: Porphyromonas gingivalis (17/30), Porphyromonas endodontalis (15/30), and Prevotella nigrescens (11/30) were the most prevalent bacterial species. At s1, endotoxins were detected in 100% of the root canals (median = 32.43 EU/mL). In parallel, substantial amounts of IL-1ß and TNF-α were produced by endodontic content-stimulated macrophages. At s2, a significant reduction in endotoxin levels was observed in all groups, with GI presenting the greatest reduction (P < .05). After a root canal rinse with EDTA (s3), intracanal medication (s4), and before root canal obturation (s5), endotoxin levels reduced without differences between groups (P < .05). IL-1ß and TNF-α release decreased proportionally to the levels of residual endotoxin (P < .05). CONCLUSIONS: Regardless of the use of sodium hypochlorite or CHX, the greatest endotoxin reduction occurs after chemomechanical preparation. Increasing steps of root canal therapy associated with intracanal medication enhances endotoxin reduction, leading to a progressively lower activation of proinflammatory cells such as macrophages.


Assuntos
Lipopolissacarídeos/análise , Ativação de Macrófagos/efeitos dos fármacos , Periodontite Periapical/microbiologia , Periodontite Periapical/terapia , Irrigantes do Canal Radicular/uso terapêutico , Preparo de Canal Radicular/métodos , Clorexidina/uso terapêutico , Humanos , Interleucina-1beta/análise , Ativação de Macrófagos/imunologia , Periodontite Periapical/imunologia , Porphyromonas endodontalis/isolamento & purificação , Porphyromonas gingivalis/isolamento & purificação , Prevotella nigrescens/isolamento & purificação , Hipoclorito de Sódio/uso terapêutico , Fator de Necrose Tumoral alfa/análise
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