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Background: The purpose of this 6‐week, single‐blinded, randomized clinical trial was to determine if the use of an interproximal brush, with or without a tracking device, is more effective than an oral irrigator in improving interproximal probing depth (PD), clinical attachment level (CAL), plaque index (PI), gingival index (GI), bleeding on probing (BOP), and inflammatory markers. Methods: Seventy‐six patients with Stages III–IV, Grade B periodontitis and a 5–7 mm posterior interproximal PD with BOP were randomized: (1) interproximal brush alone (IB; n = 26), (2) interproximal brush with tracking device (TD; n = 23), (3) oral irrigator (OI; n = 27). Participants used devices once daily for 6 weeks. Clinical measurements (PD, CAL, PI, BOP, GI) and gingival crevicular fluid (GCF) samples were collected at baseline and 6 weeks. Results: All groups showed a significant reduction in PD and CAL (≥1.1 mm, p < 0.0001) and improvement in BOP (≥56%, p < 0.0001) and GI (≥82%, p < 0.001) at the experimental site with no differences among groups. The IB and IB+TD groups showed a significant reduction in PI (≥0.9, p ≤ 0.01). Interleukin (IL)‐1β was reduced in all groups (p = 0.006), but IB+TB more than OI (p ≤ 0.05). IL‐10 was reduced among all groups (p = 0.01), while interferon‐gamma significantly increased (p = 0.01) in all groups. Conclusions: IB and OI improved clinical parameters of PD and CAL and reduced inflammatory markers (BOP, GI, GCF IL‐1β). IB had better interproximal plaque reduction. Tracking did not significantly improve clinical parameters compared with the IB and OI groups, suggesting future modifications are needed. [ABSTRACT FROM AUTHOR]

Background: The objective of this exploratory study was to evaluate inflammatory markers in periodontal maintenance patients from a randomized, double-masked, parallel intervention clinical trial comparing local simvastatin (SIM) to carrier alone following mini-flap access.Methods: Fifty patients with a 6-9-mm inflamed pocket during periodontal maintenance therapy (PMT) were treated with papilla reflection (PR)/root planing and placement of 2.2-mg simvastatin in methylcellulose (SIM/MCL) or methylcellulose alone (MCL). A small piece of interproximal soft tissue was harvested at baseline and 2 weeks postoperatively, gingival crevicular fluid (GCF) obtained at baseline, 2 weeks and 12 months, and bleeding on probing (BOP) and clinical attachment level (CAL) were measured at baseline and 12 months. Pro-inflammatory interleukin (IL)-6 and anti-inflammatory IL-10 gene activation were determined by reverse transcriptase polymerase chain reaction (rt-PCR). GCF IL-1β, IL-6, IL-10, and vascular endothelial growth factor (VEGF-A) were measured with multiplex technology. Comparisons between groups and over time used logistic regression and general estimating equations. Associations between inflammatory markers and 12-month outcomes used Wilcoxon rank sum tests or Pearson correlations.Results: Patients in the SIM group had 4.17 greater odds (p = 0.047) of improved BOP at 12 months. Median IL-6 and VEGF were significantly increased for all patients after 2 weeks of healing (p < 0.0001 and p = 0.03, respectively), while median IL-10 gene activation was increased after 2 weeks in SIM/MCL (NS). Overall, elevated GCF IL-10 at 2 weeks was significantly correlated with improved CAL at 12 months (r = -0.32, p = 0.03).Conclusions: Local SIM/MCL may have anti-inflammatory effects that potentially are associated with improved long-term CAL outcomes. [ABSTRACT FROM AUTHOR]

Background: The purpose of this double-masked, randomized, controlled trial was to determine if the local application of simvastatin (SIM), combined with minimally invasive papilla reflection and root planing (PR/RP), is effective in improving clinical attachment level (CAL), probing depth (PD) reduction, and increasing interproximal bone height (IBH) in persistent 6-9 mm periodontal pockets in patients receiving periodontal maintenance therapy (PMT).Methods: Fifty patients with Stage III, Grade B periodontitis presenting with a 6-9 mm interproximal PD with a history of bleeding on probing (BOP) were included in the study. Experimental [PR/RP+SIM/methylcellulose (MCL); n = 27] and control (PR/RP+MCL; n = 23) therapies were randomly assigned. Root surfaces were accessed via reflection of interproximal papillae, followed by RP assisted with endoscope evaluation, acid etching, and SIM/MCL or MCL application. CAL, PD, BOP, plaque presence, and IBH (using standardized vertical bitewing radiographs) were evaluated at baseline and 12 months. Measurements were compared by group and time using Chi-square, Wilcoxon rank-sum, and t-tests.Results: Both PR/RP+SIM/MCL and PR/RP+MCL, respectively, resulted in improvements in clinical outcomes (CAL: -1.9 ± 0.3 mm, p < 0.0001; -1.0 ± 0.3 mm, p < 0.003; PD: -2.3 mm ± 0.3, p < 0.0001; -1.3 mm ± 0.3, p < 0.0001; BOP: -58.7%; -41.7%, p < 0.05) and stable IBH (-0.2 ± 0.12, -0.4 ± 0.2, p = 0.22) from baseline to 12 months post-therapy. PR/RP+SIM/MCL had more improvement in CAL (p = 0.03), PD (p = 0.007), and BOP (p = 0.047).Conclusions: The addition of SIM/MCL to PR/RP improved CAL, PD, and BOP compared with PR/RP alone in periodontal maintenance patients. [ABSTRACT FROM AUTHOR]

Summary: Large quantities of organic matter are continuously deposited, and (a)biotic gradients intersect in the soil–rhizosphere, where biodegradation contributes to the global cycles of elements. The betaproteobacterial genus Aromatoleum comprises cosmopolitan, facultative denitrifying degradation specialists. Aromatoleum aromaticum. pCyN1 stands out for anaerobically decomposing plant‐derived monoterpenes in addition to monoaromatic hydrocarbons, polar aromatics and aliphatics. The catabolic network's structure and flexibility in A. aromaticum pCyN1 were studied across 34 growth conditions by superimposing proteome profiles onto the manually annotated 4.37 Mbp genome. Strain pCyN1 employs three fundamentally different enzymes for C–H‐bond cleavage at the methyl groups of p‐cymene/4‐ethyltoluene, toluene and p‐cresol respectively. Regulation of degradation modules displayed substrate specificities ranging from narrow (toluene and cyclohexane carboxylate) via medium‐wide (one module shared by p‐cymene, 4‐ethyltoluene, α‐phellandrene, α‐terpinene, γ‐terpinene and limonene) to broad (central benzoyl‐CoA pathway serving 16 aromatic substrates). Remarkably, three variants of ATP‐dependent (class I) benzoyl‐CoA reductase and four different β‐oxidation routes establish a degradation hub that accommodates the substrate diversity. The respiratory system displayed several conspicuous profiles, e.g. the presence of nitrous oxide reductase under oxic and of low‐affinity oxidase under anoxic conditions. Overall, nutritional versatility in conjunction with network regulation endow A. aromaticum pCyN1 with broad adaptability. [ABSTRACT FROM AUTHOR]

Several studies have attempted to understand the origin and evolution of single‐exon genes (SEGs) in eukaryotic organisms, including fishes, but few have examined the functional and evolutionary relationships between SEGs and multiple‐exon gene (MEG) paralogs, in particular the conservation of promoter regions. Given that SEGs originate via the reverse transcription of mRNA from a "parental" MEGs, such comparisons may enable identifying evolutionarily‐related SEG/MEG paralogs, which might fulfill equivalent physiological functions. Here, the relationship of SEG proportion with MEG count, gene density, intron count, and chromosome size was assessed for the genome of the European sea bass, Dicentrarchus labrax. Then, SEGs with an MEG parent were identified, and promoter sequences of SEG/MEG paralogs were compared, to identify highly conserved functional motifs. The results revealed a total count of 1,585 (8.3% of total genes) SEGs in the European sea bass genome, which was correlated with MEG count but not with gene density. The significant correlation of SEG content with the number of MEGs suggests that SEGs were continuously and independently generated over evolutionary time following species divergence through retrotranscription events, followed by tandem duplications. Functional annotation showed that the majority of SEGs are functional, as is evident from their expression in RNA‐seq data used to support homology‐based genome annotation. Differences in 5′UTR and 3′UTR lengths between SEG/MEG paralogs observed in this study may contribute to gene expression divergence between them and therefore lead to the emergence of new SEG functions. The comparison of nonsynonymous to synonymous changes (Ka/Ks) between SEG/MEG parents showed that 74 of them are under positive selection (Ka/Ks > 1; p =.0447). An additional fifteen SEGs with an MEG parent have a common promoter, which implies that they are under the influence of common regulatory networks. [ABSTRACT FROM AUTHOR]

Background: Efficient methods to treat persistent pockets during periodontal maintenance therapy (PMT) require further investigation. The hypothesis of this randomized controlled clinical trial was that local application of enamel matrix derivative (EMD) added to papilla reflection/root preparation (PR/RP) could enhance clinical and inflammatory outcomes, primarily clinical attachment level (CAL).Methods: Fifty PMT patients with generalized stage III-IV, grade B periodontitis presenting with a 6- to 9-mm interproximal PD were randomly allocated to (PR/RP+EMD; n = 24) and control (PR/RP+saline; n = 26) therapies by sex and smoking status. Roots were treated with reflection of interproximal papillae, root planing assisted with endoscope evaluation, and acid etching, followed by EMD or saline application. Probing depth (PD), CAL, plaque index (PI), and interproximal bone height were evaluated at baseline and 12-months post-therapy. Gingival crevicular fluid, bleeding on probing (BOP), and interleukin-1β were tested (ELISA) at baseline, 2 weeks, and 6 and 12 months. Groups were compared over time and between groups with Wilcoxon Rank Sum and t-tests.Results: Both PR/RP+ EMD and PR/RP+S resulted in significant improvements in clinical outcomes (PD and CAL, BOP) from baseline to 12 months. No significant differences were found in clinical or inflammatory outcomes between the experimental and control groups.Conclusions: The addition of EMD to PR/RP does not significantly improve clinical or inflammatory outcomes compared with PR/RP alone during periodontal maintenance therapy. [ABSTRACT FROM AUTHOR]

Background: Loss of alveolar ridge width and height after tooth extraction is well documented, but models to evaluate ridge preservation are neither standardized nor cost-effective. This rat model characterizes the pattern of bone turnover and inflammation after extraction and bone grafting with or without local simvastatin (SIM).Methods: Fifty retired-breeder rats underwent extraction of the maxillary right first molar and standard surgical defect creation under inhalation/local anesthesia. The left side of each animal served as unmanipulated control. Untreated groups (n = 8 to 9 per group) were compared (analysis of variance, t test) at days 0, 7, 14, and 28 for alveolar ridge height and width and for markers of inflammation and bone turnover by microcomputed tomography, histology, and enzyme-linked immunosorbent assay. Seventeen additional specimens had defects grafted with either bone mineralized matrix (BMM) or a BMM+SIM conjugate.Results: Extraction-induced bone loss (BL) was noted on buccal, palatal, and interproximal height (P <0.05) and ridge width (P <0.01). Week 1 inflammation positively correlated with ridge height; thereafter, a more intense inflammatory reaction corresponded to reduction in alveolar bone height and density (r = 0.74; P <0.05; Spearman). BMM+SIM preserved the most interproximal bone height (P <0.01), increased ridge width and bone density (P <0.01), enhanced 7-day prostaglandin E2 (P <0.01), and reduced 28-day inflammation density (P <0.05).Conclusions: The standard defect used in the current study paralleled human postextraction alveolar BL. Defect grafting, especially BMM+SIM, reduced inflammation and preserved bone. [ABSTRACT FROM AUTHOR]

Background: Minocycline microspheres (MMs) are being used to treat residual inflamed periodontal pockets during periodontal maintenance therapy (PMT), but evidence for efficacy from randomized clinical trials is lacking. The purpose of this study is to evaluate the effect of MMs plus scaling and root planing (SRP) on these sites.Methods: Sixty patients with chronic periodontitis on 6-month PMT intervals to be followed for 1 year were randomized (51 completed the study) into two statistically similar groups, SRP + MM (aged 66.8 years) and SRP alone (aged 67 years), to treat a ≥5 mm posterior interproximal pocket during PMT with a history of bleeding on probing (BOP). Group treatments were applied to the site at baseline and 6 months. Clinical attachment levels (CALs; primary outcome), probing depths (PDs), plaque, and BOP also were recorded at baseline and 6 and 12 months. In addition, gingival crevicular fluid was analyzed for an inflammation index ratio of interleukin (IL)-1β/IL-1 receptor antagonist (ra) using enzyme-linked immunosorbent assays.Results: All clinical parameters improved significantly (P <0.005) from baseline in both groups with no differences between groups at any time point. CAL decreased 17% (0.9 ± 0.8 mm) and 13% (0.7 ± 0.9 mm) in SRP + MM and 11% (0.7 ± 1.1 mm) and 21% (1.2 ± 0.9 mm) in SRP at 6 and 12 months, respectively. The odds of having BOP decreased 90% (down to 38% of patients) and 95% (26%) in SRP + MM and 82% (42%) and 82% (41%) in SRP at 6 and 12 months, respectively. IL-1β/IL-1ra decreased a significant 61% (P = 0.009) only in SRP + MM at 6 months.Conclusions: SRP of inflamed moderate pockets during 6-month PMT, with or without MMs, improves CALs, along with PDs and BOP over a 1-year period. The use of MMs did not result in an additional benefit over SRP alone. [ABSTRACT FROM AUTHOR]

Background: Local application of statins has shown potential in preventing and regenerating bone loss associated with experimental periodontitis. This study evaluates the effect of a novel simvastatin (SIM) prodrug (capable of delivering high doses to periodontitis inflammatory lesion and cells) on experimental periodontitis bone loss and inflammation.Methods: Forty mature female Sprague Dawley rats were subjected to ligature-induced experimental periodontitis between maxillary first and second molars (M1-M2). Equal groups were treated with three weekly doses of: 1) prodrug carrier alone (mPEG); 2) 0.5 mg SIM dose equivalent in carrier (SIM/SIM-mPEG); 3) 1.0 mg SIM/SIM-mPEG; 4) 1.5 mg SIM/SIM-mPEG; or 5) ligature alone. Contralateral molars served as unmanipulated controls. Four weeks after initiation of periodontitis, animals were euthanized, the M1-M2 interproximal was evaluated with microcomputed tomography and histology, and data were analyzed with one-way analysis of variance.Results: Ligature alone caused a mean bone loss of 1.01 ± 0.06 mm from the cemento-enamel junction, whereas all doses of SIM/SIM-mPEG reduced bone loss, especially 1.5 mg SIM/SIM-mPEG (0.68 ± 0.05 mm, P <0.001), which was not statistically different from contralateral control (0.47 ± 0.06 mm). A dose of 1.5 mg SIM/SIM-mPEG also reduced percentage of neutrophils compared with carrier alone (2.0% ± 1.0% versus 5.7% ± 1.1%; P <0.05), and increased amount of uninflamed connective tissue in the M1-M2 interproximal area (65.2% ± 3.3% versus 46.3% ± 3.3%; P <0.001). The mPEG carrier alone did not have bone-sparing or anti-inflammatory properties.Conclusion: Multiple local 1.5-mg doses of a macromolecular SIM prodrug decreases amount of experimental periodontitis bone loss and inflammation in rats. [ABSTRACT FROM AUTHOR]

The molecular roles of many RNA-binding proteins in bacterial post-transcriptional gene regulation are not well understood. Approaches combining in vivo UV crosslinking with RNA deep sequencing ( CLIP-seq) have begun to revolutionize the transcriptome-wide mapping of eukaryotic RNA-binding protein target sites. We have applied CLIP-seq to chart the target landscape of two major bacterial post-transcriptional regulators, Hfq and CsrA, in the model pathogen Salmonella Typhimurium. By detecting binding sites at single-nucleotide resolution, we identify RNA preferences and structural constraints of Hfq and CsrA during their interactions with hundreds of cellular transcripts. This reveals 3′-located Rho-independent terminators as a universal motif involved in Hfq- RNA interactions. Additionally, Hfq preferentially binds 5′ to sRNA-target sites in mRNAs, and 3′ to seed sequences in sRNAs, reflecting a simple logic in how Hfq facilitates sRNA- mRNA interactions. Importantly, global knowledge of Hfq sites significantly improves sRNA-target predictions. CsrA binds AUGGA sequences in apical loops and targets many Salmonella virulence mRNAs. Overall, our generic CLIP-seq approach will bring new insights into post-transcriptional gene regulation by RNA-binding proteins in diverse bacterial species. [ABSTRACT FROM AUTHOR]

Background: Numerous studies have documented the clinical outcomes of laser therapy for untreated periodontitis, but very few have reported on lasers treating inflamed pockets during maintenance therapy. The aim of this study is to compare the effectiveness of scaling and root planing (SRP) plus the adjunctive use of diode laser therapy to SRP alone on changes in the clinical parameters of disease and on the gingival crevicular fluid (GCF) inflammatory mediator interleukin-1β (IL-1β) in patients receiving regular periodontal maintenance therapy.Methods: This single-masked and randomized, controlled, prospective study includes 22 patients receiving regular periodontal maintenance therapy who had one or more periodontal sites with a probing depth (PD) ≥ 5 mm with bleeding on probing (BOP). Fifty-six sites were treated with SRP and adjunctive laser therapy (SRP + L). Fifty-eight sites were treated with SRP alone. Clinical parameters, including PD, clinical attachment level (CAL), and BOP, and GCF IL-1β levels were measured immediately before treatment (baseline) and 3 months after treatment.Results: Sites treated with SRP + L and SRP alone resulted in statistically significant reductions in PD and BOP and gains in CAL. These changes were not significantly different between the two therapies. Similarly, differences in GCF IL-1β levels between SRP + L and SRP alone were not statistically significant.Conclusion: In periodontal maintenance patients, SRP + L did not enhance clinical outcomes compared to SRP alone in the treatment of inflamed sites with ≥ 5 mm PD. [ABSTRACT FROM AUTHOR]

Bacteria of the class D ehalococcoidia ( DEH) (phylum C hloroflexi) are widely distributed in the marine subsurface and are especially prevalent in deep marine sediments. Nevertheless, little is known about the specific distributions of DEH subgroups at different sites and depths. This study therefore specifically examined the distributions of DEH through depths of various marine sediment cores by quantitative PCR and pyrosequencing using newly designed DEH 16 S rRNA gene targeting primers. Quantification of DEH showed populations may establish in shallow sediments (i.e. upper centimetres), although as low relative proportions of total Bacteria, yet often became more prevalent in deeper sediments. Pyrosequencing revealed pronounced diversity co-exists within single biogeochemical zones, and that clear and sometimes abrupt shifts in relative proportions of DEH subgroups occur with depth. These shifts indicate varying metabolic properties exist among DEH subgroups. The distributional changes in DEH subgroups with depth may be related to a combination of biogeochemical factors including the availability of electron acceptors such as sulfate, the composition of organic matter and depositional regimes. Collectively, the results suggest DEH exhibit wider metabolic and genomic diversity than previously recognized, and this contributes to their widespread occurrence in the marine subsurface. [ABSTRACT FROM AUTHOR]

The aim of this study was to characterize the transcriptome of a balanced polymorphism, under the regulation of a single gene, for phosphate fertilizer responsiveness/arsenate tolerance in wild grass Holcus lanatus genotypes screened from the same habitat., De novo transcriptome sequencing, RNAseq (RNA sequencing) and single nucleotide polymorphism ( SNP) calling were conducted on RNA extracted from H. lanatus. Roche 454 sequencing data were assembled into c. 22 000 isotigs, and paired-end Illumina reads for phosphorus-starved (P−) and phosphorus-treated ( P+) genovars of tolerant ( T) and nontolerant ( N) phenotypes were mapped to this reference transcriptome., Heatmaps of the gene expression data showed strong clustering of each P+/ P− treated genovar, as well as clustering by N/ T phenotype. Statistical analysis identified 87 isotigs to be significantly differentially expressed between N and T phenotypes and 258 between P+ and P− treated plants. SNPs and transcript expression that systematically differed between N and T phenotypes had regulatory function, namely proteases, kinases and ribonuclear RNA-binding protein and transposable elements., A single gene for arsenate tolerance led to distinct phenotype transcriptomes and SNP profiles, with large differences in upstream post-translational and post-transcriptional regulatory genes rather than in genes directly involved in P nutrition transport and metabolism per se. [ABSTRACT FROM AUTHOR]

Magnetotactic bacteria ( MTB) represent a group of diverse motile prokaryotes that biomineralize magnetosomes, the organelles responsible for magnetotaxis. Magnetosomes consist of intracellular, membrane-bounded, tens-of-nanometre-sized crystals of the magnetic minerals magnetite ( Fe3O4) or greigite ( Fe3S4) and are usually organized as a chain within the cell acting like a compass needle. Most information regarding the biomineralization processes involved in magnetosome formation comes from studies involving Alphaproteobacteria species which biomineralize cuboctahedral and elongated prismatic crystals of magnetite. Many magnetosome genes, the mam genes, identified in these organisms are conserved in all known MTB. Here we present a comparative genomic analysis of magnetotactic Deltaproteobacteria that synthesize bullet-shaped crystals of magnetite and/or greigite. We show that in addition to mam genes, there is a conserved set of genes, designated mad genes, specific to the magnetotactic Deltaproteobacteria, some also being present in Candidatus Magnetobacterium bavaricum of the Nitrospirae phylum, but absent in the magnetotactic Alphaproteobacteria. Our results suggest that the number of genes associated with magnetotaxis in magnetotactic Deltaproteobacteria is larger than previously thought. We also demonstrate that the minimum set of mam genes necessary for magnetosome formation in Magnetospirillum is also conserved in magnetite-producing, magnetotactic Deltaproteobacteria. Some putative novel functions of mad genes are discussed. [ABSTRACT FROM AUTHOR]

The chromosomal translocation t(8;14)(q24;q32) with juxtaposition of MYC to enhancer elements in the immunoglobulin heavy chain (IGH) gene locus is the genetic hallmark of the majority of Burkitt lymphoma and a subset of Diffuse large B‐cell lymphoma patients. Around 3% of adult B‐lineage acute lymphoblastic leukemia (ALL) patients show this aberration. Flow cytometry mostly reveals a “mature B‐ALL” or “Burkitt‐type” ALL immunophenotype. Using long‐distance PCR for t(8;14)/MYC‐IGH fusion, we investigated bone marrow, peripheral blood and a few other samples with suspected Burkitt‐ALL or mature B‐ALL and identified 133 MYC‐IGH‐positive cases. The location of the chromosomal breaks in the IGH joining and the 8 different switch regions was determined using a set of long‐distance PCRs. The chromosomal breakpoints with the adjacent MYC regions on 8q24 were characterized by direct sequencing in 49 cases. The distribution of chromosomal breaks among the IGH joining and switch regions was the following: JH 23.3%, M 21.8%, G1 15.0%, G2 7.5%, G3 3.8%, G4 4.5%, A1 12.8%, A2 3.8%, E 7.5%. Two breakpoint clusters near MYC were delineated. There was no clear correlation between the degree of somatic hypermutation and the chromosomal break locations. Epstein Barr virus was detected in 5 cases (4%). This detailed and extensive molecular analysis illustrates the molecular complexity of the MYC‐IGH translocations and the detected distribution of breakpoints provides additional evidence that this translocation results from failed switch and VDJ recombinations. This study may serve as a model for the analysis of other IGH translocations in B‐cell lymphoma. Highlights: Establishes an extensive map of MYC‐IGH translocations with previously unmatched accuracy.Shows an unexpectedly high frequency of MYC translocations to the Ce switch region.Reveals complex MYC translocations in a switch‐rearranged IGH gene locus.Investigates correlations between somatic hypermutation and IGH and MYC break location.Provides a map of the most frequently mutated amino acids in the MYC protein. [ABSTRACT FROM AUTHOR]

Among the dominant deltaproteobacterial sulfate-reducing bacteria ( SRB), members of the genus Desulfobacula are not only present in (hydrocarbon-rich) marine sediments, but occur also frequently in the anoxic water bodies encountered in marine upwelling areas. Here, we present the 5.2 Mbp genome of Desulfobacula toluolica Tol2, which is the first of an aromatic compound-degrading, marine SRB. The genome has apparently been shaped by viral attacks (e.g. CRISPRs) and its high plasticity is reflected by 163 detected genes related to transposases and integrases, a total of 494 paralogous genes and 24 group II introns. Prediction of the catabolic network of strain Tol2 was refined by differential proteome and metabolite analysis of substrate-adapted cells. Toluene and p-cresol are degraded by separate suites of specific enzymes for initial arylsuccinate formation via addition to fumarate ( p-cresol-specific enzyme HbsA represents a new phylogenetic branch) as well as for subsequent modified β-oxidation of arylsuccinates to the central intermediate benzoyl- CoA. Proteogenomic evidence suggests specific electron transfer ( EtfAB) and membrane proteins to channel electrons from dehydrogenation of both arylsuccinates directly to the membrane redox pool. In contrast to the known anaerobic degradation pathways in other bacteria, strain Tol2 deaminates phenylalanine non-oxidatively to cinnamate by phenylalanine ammonia-lyase and subsequently forms phenylacetate (both metabolites identified in 13C-labelling experiments). Benzoate degradation involves CoA activation, reductive dearomatization by a class II benzoyl- CoA reductase and hydrolytic ring cleavage as found in the obligate anaerobe Geobacter metallireducens GS-15. The catabolic sub-proteomes displayed high substrate specificity, reflecting the genomically predicted complex and fine-tuned regulatory network of strain Tol2. Despite the genetic equipment for a TCA cycle, proteomic evidence supports complete oxidation of acetyl- CoA to CO2 via the Wood- Ljungdahl pathway. Strain Tol2 possesses transmembrane redox complexes similar to that of other Desulfobacteraceae members. The multiple heterodisulfide reductase-like proteins (more than described for Desulfobacterium autotrophicum HRM2) may constitute a multifaceted cytoplasmic electron transfer network. [ABSTRACT FROM AUTHOR]

Background: Simvastatin has been shown to stimulate new bone growth on rat mandibles, but much of the bone is lost over time. The purpose of this study is to evaluate the impact of a locally or systemically applied antiresorptive agent (alendronate) on simvastatin-induced bone formation in and adjacent to a rat periodontal defect. Methods: Fenestration defects were created over mandibular molar roots in 65 mature female Sprague-Dawley rats. Two weeks later, animals were divided into eight groups of eight to nine rats, and three weekly injections around the defect were applied: 1) 0.5 mg simvastatin in ethanol (SIM-EtOH); 2) 0.5 mg simvastatin in alendronate-cyclodextrin conjugate (SIM-ALN-CD); 3) EtOH alone; 4) ALN-CD alone; or 5) no injections. Twenty-four animals were evaluated for new bone width around the defect 21 days after the last injections (short-term) and 41 rats were followed for 48 days (long-term). Three SIM-EtOH groups of long-term rats also were subjected to 2 weeks of daily systemic ALN or saline either during or 3 to 4 weeks after SIM-EtOH injections. Decalcified, hematoxylin-and-eosin-stained cross-sections of the defect area were analyzed for new bone width and groups were compared using mixed-model analyses of variance. Results: All groups showed nearly 100% bone fill, with no differences among the short-term groups. However, in the long-term animals, two-fold to three-fold more new bone width (≤0.004) was seen around the periphery of the defect with the use of systemic ALN after SIM-EtOH injections (0.93 ± 0.12 and 0.78 ± 0.11 mm with early and late systemic ALN, respectively) compared to local SIM/ALN-CD preparations (0.32 ± 0.10 mm) or short-term SIM-EtOH injections (0.35 ± 0.10 mm). No significant new cementum formation or ankylosis was noted. Conclusion: The use of a short course of systemic ALN during the healing period after bone anabolic SIM injections has the potential to enhance local bone augmentation. J Periodontol 2012;83:1463-1471. [ABSTRACT FROM AUTHOR]

The small RNAs associated with the protein Hfq constitute one of the largest classes of post-transcriptional regulators known to date. Most previously investigated members of this class are encoded by conserved free-standing genes. Here, deep sequencing of Hfq-bound transcripts from multiple stages of growth of Salmonella typhimurium revealed a plethora of new small RNA species from within mRNA loci, including DapZ, which overlaps with the 3? region of the biosynthetic gene, dapB. Synthesis of the DapZ small RNA is independent of DapB protein synthesis, and is controlled by HilD, the master regulator of Salmonella invasion genes. DapZ carries a short G/U-rich domain similar to that of the globally acting GcvB small RNA, and uses GcvB-like seed pairing to repress translation of the major ABC transporters, DppA and OppA. This exemplifies double functional output from an mRNA locus by the production of both a protein and an Hfq-dependent trans-acting RNA. Our atlas of Hfq targets suggests that the 3? regions of mRNA genes constitute a rich reservoir that provides the Hfq network with new regulatory small RNAs. [ABSTRACT FROM AUTHOR]

The pathway for anaerobic degradation of 4-methylbenzoate was studied in the denitrifying alphaproteobacterium Magnetospirillum sp. strain pMbN1. Adaptation studies with whole cells indicated substrate-dependent induction of the capacity to degrade 4-methylbenzoate. Differential protein profiling (2D-DIGE) of 4-methylbenzoate- in comparison with benzoate- or succinate-adapted cells revealed the specific abundance increase of substrate-specific protein sets. Their coding genes form distinct clusters on the genome, two of which were assigned to 4-methylbenzoate and one to benzoate degradation. The predicted functions of the gene products agree with a specific 4-methylbenzoyl-CoA degradation pathway in addition to and analogous to the known anaerobic benzoyl-CoA degradation pathway. In vitro benzoyl-CoA and 4-methylbenzoyl-CoA reductase activities revealed the electron donor and ATP-dependent formation of the corresponding conjugated cyclic dienoyl-CoA/4-methyl-dienoyl-CoA products. The 4-methylbenzoyl-CoA reductase activity was induced in the presence of 4-methylbenzoate. In accordance, metabolite analysis of cultures grown with 4-methylbenzoate tentatively identified 4-methylcyclohex-1,5-diene-1-carboxylate. The 4-methylbenzoate induced genes were assigned to code for the putative 4-methylbenzoyl-CoA reductase; their products display pronounced sequence disparity from the conventional class I benzoyl-CoA reductase, which does not accept substituents at the para-position. Identification of 3-methylglutarate together with the formation of specific proteins for ring cleavage and β-oxidation in 4-methylbenzoate-adapted cells suggest conservation of the methyl group along the specific 4-methylbenzoyl-CoA degradation pathway. [ABSTRACT FROM AUTHOR]

Background: To study aldose reductase and the sorbitol pathway in periodontitis and diabetes, rats with experimental periodontitis with or without diabetes were treated with three structurally diverse aldose reductase inhibitors (ARIs). Methods: Periodontitis was induced with three consecutive palatal injections of Porphyromonas gingivalis lipopoly-saccharide (LPS) at 48-hour intervals between the first and second molars on the right side in young, age-matched, streptozotocin-induced rats with and without diabetes 44 days after initiation of diets with and without the ARIs tolrestat, imirestat, and quercetin. As an internal control, phosphate-buffered saline (PBS) was similarly injected on the left side. Twenty-four days after the final injection, all rats were euthanized. Defleshed samples were stained with 5% toluidine blue and palatal digital images were traced to include the enamel crown and exposed root. The root/enamel ratios (to estimate alveolar bone loss) were analyzed with repeated measures analysis of variance. Results: LPS injections resulted in significantly more bone loss versus PBS injections in both the rats with and without diabetes on normal diets (P <0.0001 ). All three ARIs significantly reduced LPS-induced periodontitis in the animals with and without diabetes (P≤0.003) to the level where they were not different from PBS-injected sites in normal diet controls. Conclusion: All ARIs demonstrated efficacy in preventing alveolar bone loss because of periodontitis in both animals with and without diabetes, suggesting a role for the sorbitol pathway and the potential for ARIs to reduce inflammatory responses downstream from aldose reductase. [ABSTRACT FROM AUTHOR]

MicroRNAs have well-established roles in eukaryotic host responses to viruses and extracellular bacterial pathogens. In contrast, microRNA responses to invasive bacteria have remained unknown. Here, we report cell type-dependent microRNA regulations upon infection of mammalian cells with the enteroinvasive pathogen, Salmonella Typhimurium. Murine macrophages strongly upregulate NF-κB associated microRNAs; strikingly, these regulations which are induced by bacterial lipopolysaccharide (LPS) occur and persist regardless of successful host invasion and/or replication, or whether an inflammatory response is mounted, suggesting that microRNAs belong to the first line of anti-bacterial defence. However, a suppression of the global immune regulator miR-155 in endotoxin-tolerant macrophages revealed that microRNA responses also depend on the status of infected cells. This study identifies the let-7 family as the common denominator of Salmonella-regulated microRNAs in macrophages and epithelial cells, and suggests that repression of let-7 relieves cytokine IL-6 and IL-10 mRNAs from negative post-transcriptional control. Our results establish a paradigm of microRNA-mediated feed-forward activation of inflammatory factors when mammalian cells are targeted by bacterial pathogens. [ABSTRACT FROM AUTHOR]

Background: Locally injected simvastatin (SIM) has been shown to induce bone growth in rat models. The purpose of this study is to evaluate the effects of locally injected simvastatin in several human-like clinical situations in a beagle dog model. Methods: Four beagle dogs completed the study and were used in a split-mouth design. Dehiscence defects of 5 x 3 mm were created bilaterally on the lateral aspect of the mandibular second premolar (PM2) mesial roots including removal of root cementum. At the same surgery, porous hydroxyapatite-collagen grafts with resorbable membranes with or without 10-mg SIM were placed buccal to the mandibular first molars (M1). One week later, three weekly local injections of 10-mg SIM in ethanol and contralateral ethanol alone were initiated at three sites through the buccal mucosa: 1) 6 mm apical to the cementoenamel junction (CEJ) of the maxillary fourth premolar (PM4; thin bone over root); 2) 6 mm apical to the CEJ of PM2 (dehiscence defect); and 3) 10 mm distoapical to the CEJ of the maxillary canine (edentulous ridge). Dogs were euthanized 2 months after the final injections. Block sections were harvested and specimens were decalcified and stained with hematoxylin and eosin. Histomorphometry was performed using digitized photographs and analyzed with distribution-free rank tests. Results: Regarding M1, the distance between CEJ and the alveolar crest was significantly more coronal in the SIM group (P= 0.038). Regarding the edentulous ridge, the width of new bone was significantly greater in SIM injection specimens (P= 0.0164). Regarding PM2, buccal bone in the dehiscence defects lacking periosteum was not augmented in the SIM group. Regarding PM4, the total width of bone 5 mm apical to the coronal height of contour (thin buccal bone covering the root) was significantly wider on the SIM side (SIM, 0.63 ± 0.53 mm; contralateral ethanol alone, 0.25 ± 0.19 mm; P= 0.0098). Conclusion: Locally injected SIM has the ability to induce modest amounts of new bone formation in closed injection sites over a periosteal surface. [ABSTRACT FROM AUTHOR]

Summary Microorganisms can degrade saturated hydrocarbons (alkanes) not only under oxic but also under anoxic conditions. Three denitrifying isolates (strains HxN1, OcN1, HdN1) able to grow under anoxic conditions by coupling alkane oxidation to CO2 with NO3- reduction to N2 were compared with respect to their alkane metabolism. Strains HxN1 and OcN1, which are both Betaproteobacteria, utilized n-alkanes from C6 to C8 and C8 to C12 respectively. Both activate alkanes anaerobically in a fumarate-dependent reaction yielding alkylsuccinates, as suggested by present and previous metabolite and gene analyses. However, strain HdN1 was unique in several respects. It belongs to the Gammaproteobacteria and was more versatile towards alkanes, utilizing the range from C6 to C30. Neither analysis of metabolites nor analysis of genes in the complete genome sequence of strain HdN1 hinted at fumarate-dependent alkane activation. Moreover, whereas strains HxN1 and OcN1 grew with alkanes and NO3-, NO2- or N2O added to the medium, strain HdN1 oxidized alkanes only with NO3- or NO2- but not with added N2O; but N2O was readily used for growth with long-chain alcohols or fatty acids. Results suggest that NO2- or a subsequently formed nitrogen compound other than N2O is needed for alkane activation in strain HdN1. From an energetic point of view, nitrogen-oxygen species are generally rather strong oxidants. They may enable enzymatic mechanisms that are not possible under conditions of sulfate reduction or methanogenesis and thus allow a special mode of alkane activation. [ABSTRACT FROM AUTHOR]

In mammalian zygotes, the 5-methyl-cytosine (5mC) content of paternal chromosomes is rapidly changed by a yet unknown but presumably active enzymatic mechanism. Here, we describe the developmental dynamics and parental asymmetries of DNA methylation in relation to the presence of DNA strand breaks, DNA repair markers and a precise timing of zygotic DNA replication. The analysis shows that distinct pre-replicative (active) and replicative (active and passive) phases of DNA demethylation can be observed. These phases of DNA demethylation are concomitant with the appearance of DNA strand breaks and DNA repair markers such as γH2A.X and PARP-1, respectively. The same correlations are found in cloned embryos obtained after somatic cell nuclear transfer. Together, the data suggest that (1) DNA-methylation reprogramming is more complex and extended as anticipated earlier and (2) the DNA demethylation, particularly the rapid loss of 5mC in paternal DNA, is likely to be linked to DNA repair mechanisms. [ABSTRACT FROM AUTHOR]

Except for environmental 16S rRNA gene sequences, no information is available for members of the candidate division OP3. These bacteria appear to thrive in anoxic environments, such as marine sediments, hypersaline deep sea, freshwater lakes, aquifers, flooded paddy soils and methanogenic bioreactors. The 16S rRNA phylogeny suggests that OP3 belongs to the Planctomycetes/ Verrucomicrobia/ Chlamydiae (PVC) superphylum. Metagenomic fosmid libraries were constructed from flooded paddy soil and screened for 16S rRNA gene-containing fragments affiliated with the PVC superphylum. The screening of 63 000 clones resulted in 23 assay-positive fosmids, of which three clones were affiliated with OP3. The 16S rRNA gene sequence divergence between the fragments OP3/1, OP3/2 and OP3/3 ranges from 18% to 25%, indicating that they belong to different OP3 subdivisions. The 23S rRNA phylogeny confirmed the membership of OP3 in the PVC superphylum. Sequencing the OP3 fragments resulted in a total of 105 kb of genomic information and 90 ORFs, of which 47 could be assigned a putative function and 11 were conserved hypothetical. Using BLASTP searches, a high proportion of ORFs had best matches to homologues from Deltaproteobacteria, rather than to those of members of the PVC superphylum. On the fragment OP3/3, a cluster of nine ORFs was predicted to encode the bacterial NADH dehydrogenase I. Given the high proportion of homologues present in deltaproteobacteria and anoxic conditions in the natural environment of OP3 bacteria, the detection of NADH dehydrogenase I may suggest an anaerobic respiration mode. Oligonucleotide frequencies calculated for OP3/1, OP3/2 and OP/3 show high intraphylum correlations. This novel sequence information could therefore be used to identify OP3-related fragments in large metagenomic data sets using marker gene-independent procedures in the future. In addition to the OP3 fragments, a single metagenomic fragment affiliated with the candidate division BRC1 was obtained and analysed. [ABSTRACT FROM AUTHOR]

Background: The analysis of biomarkers in gingival crevicular fluid (GCF) may be helpful in forecasting patient vulnerability to future attachment loss. The purpose of this study is to correlate GCF biomarkers of inflammation and bone resorption with subsequent periodontal attachment and bone loss in a longitudinal trial of a matrix metalloproteinase (MMP) inhibitor. Methods: GCF was collected from two periodontal pockets (mean ± SD: 5.1 ± 1.0 mm) at baseline and annually in postmenopausal females with moderate to advanced periodontitis undergoing periodontal maintenance every 3 to 4 months during a 2-year double-masked, placebo-controlled, randomized clinical trial of subantimicrobial dose doxycycline (SDD; 20 mg two times a day). Subjects were randomized to SDD (n = 64) or a placebo (n = 64). GCF was analyzed for the inflammation markers interleukin (IL)-1β (using enzyme-linked immunosorbent assay), total collagenase activity (using hydrolysis of a synthetic octapeptide), and MMP-8 (using a Western blot) and the bone-resorption marker carboxyterminal telopeptide cross-link fragment of type I collagen (ICTP) (using a radioimmunoassay). Generalized estimating equations were used to associate these biomarkers, categorized into tertiles, with subsequent clinical attachment (using an automated disk probe) or interproximal bone loss (using radiography). Odds ratio (OR) values compared highest to lowest tertile groups. Results: Increases in GCF IL- 1β and MMP-8 during the first year of periodontal maintenance were associated with increased odds of subsequent (year 2) periodontal attachment loss (OR = 1.67; P= 0.01 and OR -- 1.50; P= 0.02, respectively) driven by the placebo group. Elevated baseline ICTP was also associated with increased odds of 1- and 2-year loss of alveolar bone density (OR = 1.98; P= 0.0001) in the placebo group, not the SDD group, and a loss of bone height (OR = 1.38; P= 0.06), again driven by the placebo group. Conclusion: These data support the hypothesis that elevated GCF biomarkers of inflammation and bone resorption from a small number of moderate/deep sites have the potential to identify patients who are vulnerable to progressive periodontitis, and SDD may modify that risk. J Periodontol 2010;81:251-259. [ABSTRACT FROM AUTHOR]

Microbial consortia mediating the anaerobic oxidation of methane with sulfate are composed of methanotrophic Archaea (ANME) and Bacteria related to sulfate-reducing Deltaproteobacteria. Cultured representatives are not available for any of the three ANME clades. Therefore, a metagenomic approach was applied to assess the genetic potential of ANME-1 archaea. In total, 3.4 Mbp sequence information was generated based on metagenomic fosmid libraries constructed directly from a methanotrophic microbial mat in the Black Sea. These sequence data represent, in 30 contigs, about 82–90% of a composite ANME-1 genome. The dataset supports the hypothesis of a reversal of the methanogenesis pathway. Indications for an assimilatory, but not for a dissimilatory sulfate reduction pathway in ANME-1, were found. Draft genome and expression analyses are consistent with acetate and formate as putative electron shuttles. Moreover, the dataset points towards downstream electron-accepting redox components different from the ones known from methanogenic archaea. Whereas catalytic subunits of [NiFe]-hydrogenases are lacking in the dataset, genes for an [FeFe]-hydrogenase homologue were identified, not yet described to be present in methanogenic archaea. Clustered genes annotated as secreted multiheme c-type cytochromes were identified, which have not yet been correlated with methanogenesis-related steps. The genes were shown to be expressed, suggesting direct electron transfer as an additional possible mode to shuttle electrons from ANME-1 to the bacterial sulfate-reducing partner. [ABSTRACT FROM AUTHOR]

The organization of magnetosome genes was analysed in all available complete or partial genomic sequences of magnetotactic bacteria (MTB), including the magnetosome island (MAI) of the magnetotactic marine vibrio strain MV-1 determined in this study. The MAI was found to differ in gene content and organization between Magnetospirillum species and strains MV-1 or MC-1. Although a similar organization of magnetosome genes was found in all MTB, distinct variations in gene order and sequence similarity were uncovered that may account for the observed diversity of biomineralization, cell biology and magnetotaxis found in various MTB. While several magnetosome genes were present in all MTB, others were confined to Magnetospirillum species, indicating that the minimal set of genes required for magnetosome biomineralization might be smaller than previously suggested. A number of novel candidate genes were implicated in magnetosome formation by gene cluster comparison. Based on phylogenetic and compositional evidence we present a model for the evolution of magnetotaxis within the Alphaproteobacteria, which suggests the independent horizontal transfer of magnetosome genes from an unknown ancestor of magnetospirilla into strains MC-1 and MV-1. [ABSTRACT FROM AUTHOR]

Bulk mRNA was used to explore the transcriptional activity of bacterial communities in oxic versus anoxic paddy soil. Two microbial cDNA libraries were constructed from composite samples using semi-randomly primed RT-PCR. cDNAs averaged 500–600 bp in length and were treated as expressed sequence tags (ESTs). Clustering analysis of 805 random cDNAs resulted in 179 and 155 different ESTs for the oxic and anoxic zones respectively. Using an E-value threshold of e−10, a total of 218 different ESTs could be assigned byblastx, while 116 ESTs were predicted novel. Both the proportion and significance of the EST assignments increased with cDNA length. Taxonomic assignment was more powerful in discriminating between the aerobic and anaerobic bacterial communities than functional inference, as most ESTs in both oxygen zones were putative indicators of similar housekeeping functions, in particular ABC-type transporters. A few ESTs were putative indicators for community function in a biogeochemical context, such as β-oxidation of long-chain fatty acids specifically in the oxic zone. Expressed sequence tags assigned to Alpha- and Betaproteobacteria were predominantly found in the oxic zone, while those affiliated with Deltaproteobacteria were more frequently detected in the anoxic zone. At the genus level, multiple assignments to Bradyrhizobium and Geobacter were unique to the oxic and anoxic zones respectively. The phylum-level affiliations of 93 16S rRNA sequences corresponded well with two taxonomically distinct EST patterns. Expressed sequence tags affiliated with Acidobacteria and Chloroflexi were frequently detected in both oxygen zones. In summary, the soil metatranscriptome is accessible for global analysis and such studies have great potential in elucidating the taxonomic and functional status of soil bacterial communities, but study significance depends on the number and length of cDNAs being randomly analysed. [ABSTRACT FROM AUTHOR]

The anaerobic biodegradation of naphthalene, an aromatic hydrocarbon in tar and petroleum, has been repeatedly observed in environments but scarcely in pure cultures. To further explore the relationships and physiology of anaerobic naphthalene-degrading microorganisms, sulfate-reducing bacteria (SRB) were enriched from a Mediterranean sediment with added naphthalene. Two strains (NaphS3, NaphS6) with oval cells were isolated which showed naphthalene-dependent sulfate reduction. According to 16S rRNA gene sequences, both strains were Deltaproteobacteria and closely related to each other and to a previously described naphthalene-degrading sulfate-reducing strain (NaphS2) from a North Sea habitat. Other close relatives were SRB able to degrade alkylbenzenes, and phylotypes enriched anaerobically with benzene. If in adaptation experiments the three naphthalene-grown strains were exposed to 2-methylnaphthalene, this compound was utilized after a pronounced lag phase, indicating that naphthalene did not induce the capacity for 2-methylnaphthalene degradation. Comparative denaturing gel electrophoresis of cells grown with naphthalene or 2-methylnaphthalene revealed a striking protein band which was only present upon growth with the latter substrate. Peptide sequences from this band perfectly matched those of a protein predicted from genomic libraries of the strains. Sequence similarity (50% identity) of the predicted protein to the large subunit of the toluene-activating enzyme (benzylsuccinate synthase) from other anaerobic bacteria indicated that the detected protein is part of an analogous 2-methylnaphthalene-activating enzyme. The absence of this protein in naphthalene-grown cells together with the adaptation experiments as well as isotopic metabolite differentiation upon growth with a mixture of d8-naphthalene and unlabelled 2-methylnaphthalene suggest that the marine strains do not metabolize naphthalene by initial methylation via 2-methylnaphthalene, a previously suggested mechanism. The inability to utilize 1-naphthol or 2-naphthol also excludes these compounds as free intermediates. Results leave open the possibility of naphthalene carboxylation, another previously suggested activation mechanism. [ABSTRACT FROM AUTHOR]

We characterized two overlapping sugar beet ( Beta vulgaris) bacterial artificial chromosome (BAC) clones representing different haplotypes. A total of 254 kbp of the genomic sequence was determined, of which the two BACs share 92 kbp. Eleven of 15 genes discovered in the sequenced interval locate to the overlap region. The haplotypes differ in exons by 1% (nucleotide level) and in non-coding regions by 9% (6% mismatches, 3% gaps; alignable regions only). Large indels or high sequence divergence comprised 11% of either sequence. Of such indels, 68 and 45%, respectively, could be attributed to haplotype-specific integration of transposable elements. We identified novel repeat candidates by comparing the two BAC sequences to a set of genomic sugar beet sequences. Synteny was found with Arabidopsis chromosome 1 (At1), At2 and At4, Medicago chromosome 7, Vitis chromosome 15 and paralogous regions on poplar chromosomes II and XIV. [ABSTRACT FROM AUTHOR]

The complete genome of the bacterium Erwinia tasmaniensis strain Et1/99 consisting of a 3.9 Mb circular chromosome and five plasmids was sequenced . Strain Et1/99 represents an epiphytic plant bacterium related to Erwinia amylovora and E. pyrifoliae, which are responsible for the important plant diseases fire blight and Asian pear shoot blight, respectively. Strain Et1/99 is a non-pathogenic bacterium and is thought to compete with these and other bacteria when occupying the same habitat during initial colonization. Genome analysis revealed tools for colonization, cellular communication and defence modulation, as well as genes coding for the synthesis of levan and a not detected capsular exopolysaccharide. Strain Et1/99 may secrete indole-3-acetic acid to increase availability of nutrients provided on plant surfaces. These nutrients are subsequently accessed and metabolized. Secretion systems include the hypersensitive response type III pathway present in many pathogens. Differences or missing parts within the virulence-related factors distinguish strain Et1/99 from pathogens such as Pectobacterium atrosepticum and the related Erwinia spp. Strain Et1/99 completely lacks the sorbitol operon, which may also affect its inability to invade fire blight host plants. Erwinia amylovora in contrast depends for virulence on utilization of sorbitol, the dominant carbohydrate in rosaceous plants. The presence of other virulence-associated factors in strain Et1/99 indicates the ancestral genomic background of many plant-associated bacteria. [ABSTRACT FROM AUTHOR]

Background: Topical injection of simvastatin in methylcellulose gel was shown to stimulate bone growth and inflammation over mouse calvaria and in rat mandible models. The purpose of these pilot studies was to evaluate the potential of locally injected simvastatin in human-sized periodontal defects. Methods: Chronic periodontal defects were created bilaterally in seven 1-year-old beagle dogs: 3-walled intrabony defects distal of the mandibular second premolar and mesial of the fourth premolar and Class II furcation defects at the buccal furcation of the mandibular first molars. The edentulous space distal to the mandibular canine was left undisturbed. After 16 weeks of healing, defect sites were treated with scaling and root planing, and mandible sides were randomly selected to receive three weekly injections of 0.5 mg simvastatin in 30 µl methylcellulose gel and contralateral gel alone (n = 3) or 2.0 mg simvastatin/methylcellulose gel and contralateral gel alone (n = 4). Two months following drug application, block sections, including teeth and surrounding tissues, and submandibular lymph nodes were obtained for histomorphometric analysis. Results: Two trends were noted in this pilot study: bucal edentulous ridge thickness was 29% greater with simvastatin, 0.5 mg, compared to gel alone (P = 0.0845), and the simvastatin groups had bone-height loss in interproximal intrabony and furcation defects, but the length of new cementum in the interproximal intrabony defects was greater with simvastatin, 0.5 mg (0.35 ± 0.14 mm), compared to gel alone (0.06 ± 0.15 mm; P = 0.069). No new cementum was found in furcations. Conclusions: Multiple injections of simvastatin are not appropriate for the treatment of intrabony or furcation defects. However, this approach shows potential to augment bone thickness in closed alveolar environments. [ABSTRACT FROM AUTHOR]

Background: We recently demonstrated that a 2-year subantimicrobial-dose doxycycline (SDD) regimen (double-masked, placebo-controlled clinical trial) in postmenopausal (PM) women exhibiting mild systemic bone loss (osteopenia) and local bone loss (periodontitis) reduced the progression of periodontal attachment loss (intent-to-treat analysis) and the severity of gingival inflammation and alveolar bone loss (subgroups) without producing antibiotic side effects. We now describe SDD effects on biomarkers of collagen degradation and bone resorption in the gingival crevicular fluid (GCF) of the same vulnerable subjects. Methods: GCF was collected from SDD- and placebo-treated PM subjects (n = 64 each) at the baseline and 1 - and 2-year appointments; the volume was determined; and the samples were analyzed for collagenase activity (using a synthetic peptide as substrate), relative levels of three genetically distinct collagenases (Western blot), a type-1 collagen breakdown product/ bone resorption marker (a carboxyterminal telopeptide cross-link fragment of type I collagen [ICTP]; radioimmunoassay), and interleukin- 1β (enzyme-linked immunosorbent assay). Statistical analyses were performed using generalized estimating equations; primary analyses were intent-to-treat. Results: Collagenase activity was significantly reduced by SDD treatment relative to placebo based on intent-to-treat (P= 0.01). ICTP showed a similar pattern of change during SDD treatment, and GCF collagenase activity and ICTP were positively correlated at all time periods (P<0.001). Matrix metalloproteinase (MMP)-8 accounted for ~80% of total collagenase in GCF, with much less MMP-1 and -13, and SDD reduced the odds of elevated MMP-8 by 60% compared to placebo (P= 0.006). Conclusion: These observations support the therapeutic potential of longterm SDD therapy to reduce periodontal collagen breakdown and alveolar bone resorption in PM women; effects on serum biomarkers of systemic bone loss in these subjects are being analyzed. [ABSTRACT FROM AUTHOR]

Aims: To determine the clinical efficacy of a 2-year continuous sub-antimicrobial dose doxycycline (SDD; 20 mg bid) in post-menopausal, osteopenic, oestrogen-deficient women on periodontal maintenance. Materials and Methods: One-hundred and twenty-eight subjects were randomized to SDD ( n=64) or placebo ( n=64). Clinical measurements were performed at posterior interproximal sites at baseline and every 6 months during this 2-year randomized, double-blind, placebo-controlled clinical trial with adjunctive, no-cost 3–4-month periodontal maintenance. Statistical analyses of secondary outcomes from this clinical trial used Generalized Estimating Equations in primarily intent-to-treat analyses. Results: For the placebo group, 3.4% of the sites showed improvement in clinical attachment levels (CAL) and 2.7% had progressive loss in CAL; for the SDD group, 5.0% of the sites showed an improvement in CAL and 2.2% had progressive loss in CAL. This difference (2.1% of sites) was more favourable in the SDD group than in the placebo [odds ratio (OR)=0.81%, 95% confidence interval (CI): 0.67–0.97, p=0.03] in these well-maintained patients, whereas probing depths, bleeding on probing and supragingival plaque did not differ significantly between groups ( p>0.2). However, in exploratory subgroup analysis of non-smokers, SDD showed reduced bleeding versus placebo (27% versus 33%; p=0.05). In protocol-adherent subjects, the odds of bleeding were 34% lower for SDD ( p=0.05). Conclusions: Analyses of secondary outcomes of this clinical trial indicated that SDD may be of benefit in reducing progressive attachment loss in post-menopausal females; additional research is needed to confirm these findings. Protocol registered at ClinicalTrials.gov. Identifier:NCT00066027 [ABSTRACT FROM AUTHOR]

Aim: Determine the efficacy of 2-year continuous subantimicrobial dose doxycycline (SDD; 20 mg bid) on alveolar bone in post-menopausal osteopenic, oestrogen-deficient women undergoing periodontal maintenance in a 2-year double-blind, placebo-controlled, randomized clinical trial. Material and Methods: One-hundred and twenty-eight subjects randomized to SDD or placebo ( n=64 each). Posterior vertical bite wings taken at baseline, 1 and 2 years for alveolar bone density (ABD), using radiographic absorptiometry (RA) and computer-assisted densitometric image analysis (CADIA), and alveolar bone height (ABH). Statistical analyses utilized generalized estimating equations; primary analyses were intent to treat (ITT). Results are presented as SDD versus placebo. Results: Under ITT, there was no statistically significant effect of SDD on ABD loss (RA: p=0.8; CADIA: p=0.2) or ABH loss ( p=0.2). Most sites (81–95%) were inactive. For subgroup analyses, mean CADIA was higher with SDD for non-smokers ( p=0.05) and baseline probing depths ⩾5 mm ( p=0.003). SDD was associated with 29% lower odds of more progressive ABH loss in women >5 years post-menopausal ( p=0.05) and 36% lower among protocol-adherent subjects ( p=0.03). Conclusions: In post-menopausal osteopenic women with periodontitis, SDD did not differ overall from placebo. Based on exploratory subgroup analyses, additional research is needed to determine the usefulness of SDD in non-smokers, subjects >5 years post-menopausal and in deeper pockets. Protocol registered at ClinicalTrials.gov. Identifier: NCT00066027 [ABSTRACT FROM AUTHOR]

Aim: The aim of this pilot study was to track markers of periodontal inflammation and bone resorption associated with decalcified freeze-dried bone allografts. Material and Methods: Eleven subjects completed standardized treatment of intrabony defects ⩾3 mm with allografts. Gingival crevicular fluid was collected from the defect site and an adjacent interproximal site within the surgical field at baseline, 2, 4, and 8 weeks post-operatively, and analysed for biochemical markers of inflammation/bone resorption. Probing depth, recession, bleeding on probing, plaque, and 6-month radiographic bone height change were measured. Results: Both prostaglandin E2 ( p=0.007) and bone-specific type 1 collagen ( p=0.01) increased in crevicular fluid after 2 weeks in the bone graft sites. Matrix metalloproteinase-9 levels remained constant over time. There were positive correlations between prostaglandin levels during the first 8 weeks and bone height change over 6 months. Conclusions: Periodontal bone grafts stimulate an inflammatory response during the first 2 weeks post-operatively, and the potential negative effects of inhibiting prostaglandins post-operatively should be investigated further. [ABSTRACT FROM AUTHOR]

Background: Based on microbiologic concerns, the safety of a 24-month regimen of subantimicrobial dose doxycycline (SDD; 20 mg twice a day) was evaluated in postmenopausal osteopenic women with periodontitis in a double-blind, placebo-controlled, randomized clinical trial.Methods: Subgingival samples were collected from two sites (probing depth > or = 5 mm) in each of 128 subjects at baseline, with the same sites resampled at the conclusion of the 2-year period. The samples were enumerated on selective and non-selective media and on doxycycline (4 microg/ml) medium. Up to five different colonial morphologies were subcultured from the doxycycline medium, identified to species, and susceptibilities determined to doxycycline and five other antibiotics. Data were analyzed for microbial differences in total colony forming units (CFU), periodontal and opportunistic pathogens, and changes in species and in susceptibilities of isolates recovered on doxycycline medium.Results: There was no significant evidence that changes in total anaerobic counts over the treatment period (P = 0.96) differed between treatment groups. Likewise, periodontal pathogens, opportunistic pathogens, or normal flora did not differ descriptively between groups. Although there was a significant increase (P <0.001) in the total CFU recovered from the 4 microg/ml doxycycline plates at 24 months for SDD versus placebo, the percentage that was clinically resistant to doxycycline (minimal inhibitory concentration [MIC] > or = 16 microg/ml) decreased over the 24-month period in both groups and did not differ between the treatment groups (SDD: 79% to 76%; placebo: 83% to 70%; P = 0.2). There were no significant differences (P >0.28 for each) in the change in cross-resistance between the groups for doxycycline and the other five antibiotics.Conclusions: No antimicrobial effect on the subgingival flora was detected following treatment with SDD for 24 months, relative to baseline or to placebo. The increase in initial resistance (at 4 microg/ml) did not translate into a significant increase in the percent resistant to doxycycline (MIC > or = 16 microg/ml) for patients in the SDD group. [ABSTRACT FROM AUTHOR]

Members of the Bacteroidetes, formerly known as the Cytophaga-Flavobacteria-Bacteroides (CFB) phylum, are among the major taxa of marine heterotrophic bacterioplankton frequently found on macroscopic organic matter particles (marine snow). In addition, they have been shown to also represent a significant part of free-living microbial assemblages in nutrient-rich microenvironments. Their abundance and distribution pattern in combination with enzymatic activity studies has led to the notion that organisms of this group are specialists for degradation of high molecular weight compounds in both the dissolved and particulate fraction of the marine organic matter pool, implying a major role of Bacteroidetes in the marine carbon cycle. Despite their ecological importance, comprehensive molecular data on organisms of this group have been scarce so far. Here we report on the first whole genome analysis of a marine Bacteroidetes representative, ‘ Gramella forsetii’ KT0803. Functional analysis of the predicted proteome disclosed several traits which in joint consideration suggest a clear adaptation of this marine Bacteroidetes representative to the degradation of high molecular weight organic matter, such as a substantial suite of genes encoding hydrolytic enzymes, a predicted preference for polymeric carbon sources and a distinct capability for surface adhesion. [ABSTRACT FROM AUTHOR]

The anaerobic oxidation of methane is a globally significant process which is mediated by consortia of yet uncultivated methanotrophic archaea (ANME) and sulfate-reducing bacteria. In order to gain deeper insights into genome characteristics of the different ANME groups, large-insert genomic libraries were constructed using DNA extracted from a methanotrophic microbial mat growing in the anoxic part of the Black Sea, and from sediments above gas hydrates at the Hydrate Ridge off the coast of Oregon. Analysis of these fosmid libraries with respect to archaeal 16S rRNA gene diversity revealed a single ANME-1b ribotype for the Black Sea libraries, whereas the sequences derived from the Hydrate Ridge library phylogenetically affiliated with the ANME-2a, ANME-2c and ANME-3 group. Genome walking for ANME-1b resulted in a contiguous 155 kb composite genome fragment. The comparison of a set of four genomic fragments belonging to the different ANME groups revealed differences in the rRNA operon structure and the average G+C content, with the ANME-2c contig showing the highest divergence within the set. A detailed analysis of the ANME contigs with respect to genes putatively involved in the anaerobic oxidation of methane led to the identification of: (i) a putative N 5 ,N 10-methenyltetrahydromethanopterin cyclohydrolase gene, (ii) a gene cluster supposedly encoding a novel type of heterodisulfide reductase/dehydrogenase complex and (iii) a gene cluster putatively encoding a new type of CO dehydrogenase/acetyl-CoA synthase enzyme complex. [ABSTRACT FROM AUTHOR]

Background: Simvastatin has been shown to increase bone growth when applied topically to murine bone; however, it causes considerable soft tissue inflammation at high doses (2.2 mg), making future clinical use problematic. This study evaluated the effect of lower simvastatin doses and cyclooxygenase (COX) synthase inhibitors on tissue inflammation and bone growth in rats and gene expression in mice.Methods: Adult female rats were untreated or treated with a single dose of 0.1, 0.5, 1.0, 1.5, or 2.2 mg simvastatin in methylcellulose gel in a polylactic acid membrane (SIM) on the lateral aspect of the mandible. The contralateral mandible side was implanted with methylcellulose gel/polylactic acid membrane alone (GEL), and five rats in each dose pairing were evaluated histomorphometrically after 3, 7, and 24 days. Subsequent rats were similarly treated with 0.5 mg simvastatin (optimal dose) and daily intraperitoneal injections of COX-2 inhibitor (NS-398; 1 mg/kg x 7 days; N = 16), general COX inhibitor (indomethacin; 1 mg/kg x 7 days; N = 16), or no inhibitor (N = 10) and evaluated histomorphometrically after 7 or 24 days by analysis of variance (ANOVA). Gene arrays were also used to evaluate osteogenic gene expression from 0.5 mg simvastatin in murine calvaria (N = 12).Results: There was a 45% increase in bone area with 0.5 mg simvastatin versus gel control (P <0.001; similar to the 2.2-mg dose), and clinical swelling was reduced compared to the high simvastatin dose (P <0.05). The 0.1-mg simvastatin dose failed to stimulate significant bone growth. NS-398 and indomethacin reduced inflammation and bone growth. Simvastatin significantly upregulated procollagen, fibronectin, and matrix metalloproteinase-13 genes.Conclusion: Reducing the simvastatin dose from 2.2 to 0.5 mg reduced inflammation to a more clinically acceptable level without sacrificing bone-growth potential, but COX-associated inflammation appears to be necessary for in vivo bone growth. [ABSTRACT FROM AUTHOR]

Growth phase dependent changes of protein composition in the marine bacterium Rhodopirellula baltica were quantitatively monitored by applying the two-dimensional difference gel electrophoresis (2D DIGE) technology. The number of regulated proteins (fold changes in protein abundance > |2|) increased from early (10) to late stationary growth phase (179), with fold changes reaching maximal values of 40. About 110 of these regulated protein spots were analysed by MALDI-TOF-MS and identified by mapping of peptide masses. Results indicate an opposing regulation of tricarboxylic acid cycle and oxidative pentose phosphate cycle, a downregulation of several enzymes involved in amino acid biosynthesis and an upregulation of the alternative sigma factor σ H in stationary phase. Interestingly, 26 proteins of unknown function were up- or downregulated in the stationary phase. Several proteins were specifically regulated during growth on solid surface (agar plates). These proteins could possibly be involved in the development of the different R. baltica morphotypes, i.e. motile swarmer cells and sessile cell aggregates (so-called rosettes). [ABSTRACT FROM AUTHOR]

Reinhardt RA, Sanderfer VJ, Meinberg TA, Nummikoski P, Lee H-M, Marx DB: Local biochemical markers of bone turnover: relationship to subsequent density of healing alveolar bone defects. J Clin Periodontol 2004; 31: –. © Blackwell Munksgaard, 2004. This pilot study was designed to test whether biochemical markers of bone turnover in washes of periosteal or trabecular alveolar bone surfaces could be correlated with increases in bone density of an adjacent healing implant socket. Ten subjects had a canula inserted into the alveolar crest and sterile phosphate-buffered saline was washed over the periosteal and trabecular surfaces and collected. Surgical flaps were reflected, 5 mm diameter bone cores were removed from the bone wash site, and standardized radiographs were taken. The sites were allowed to heal for 12 weeks, and radiographs were repeated. Bone washes of the healing sites were also collected after 2 and 12 weeks. Washes were analysed for bone turnover markers osteocalcin (OC; radioimmunoassay) and C-terminal telopeptide of Type 1 collagen (ICTP; enzyme-linked immunosorbent assay (ELISA)), and blood component albumin (ALB; ELISA). Changes in bone density during healing were determined by radiographic absorptiometry. OC/ALB and ICTP/ALB ratios were higher for trabecular than periosteal washes at baseline ( p⩽0.01). Trabecular OC/ALB and ICTP/ALB were inversely correlated with increasing bone density of the healing bone core socket ( r=−0.72, p=0.03; Pearson's correlation coefficient). Biochemical markers of bone turnover in bone washes of specific alveolar bone sites may prove helpful in predicting how the bone density will increase around healing dental implants. [ABSTRACT FROM AUTHOR]

Background: The study of acclusal therapy in humans with periodontitis is problematic due to potential irreversible bone loss in control subjects. The hypothesis of this pilot investigation was that increased interocclusal contact initiated by short-term occlusal splint disuse would increase tooth mobility and the bone-resorptive cytokine interleukin (IL)-1β in gingival crevicular fluid (GCF), comparable to IL-1-positive genotype and increased periodontitis severity. Methods: Nineteen nonsmoking chronic periodontitis patients using nocturnal occlusal splints and undergoing periodontal maintenance in a private practice were evaluated at five time points: 24 hours after continuous splint use: 1, 2, and 3 days after no occlusal splint use; and 14 days after resumption of customary nighttime splint use. Subjects were evaluated to confirm that the plaque index and gingival index were ≤1.0, and to categorize past periodontitis (moderate or severe) and IL-l genotype (1A+4845 plus IL-1B +3954). Test sites on two anterior teeth vulnerable to occlusal trauma were sampled for mobility. GCF IL-1β, and IL-1 receptor antagonist (ra). Results: Tooth mobility remained low during the 3-day period when patients were not wearing their occlusal appliance. GCF IL-1β decreased after not wearing the appliances (P=0.016), especially at 48 hours. At this time, genotype-positive subjects had higher levels of GCF IL-1β/IL-1ra than genotype-negative subjects (P= 0.045), and patients who had experienced severe periodontitis had higher IL-1β levels than moderate periodontitis subjects (P= 0.004). Conclusions: These findings suggest that short-term discontinuation of occlusal splint therapy in non-smoking periodontitis patients undergoing periodontal maintenance does not result in potential signs of early occlusal trauma (increasing mobility or GCF IL-1β). Longer-term studies may be needed to determine appropriate therapy applications for periodontitis-susceptible patients with definable occlusal discrepancies and/or parafunction. [ABSTRACT FROM AUTHOR]