High Throughput Sequencing Detect Gingivitis and Periodontal Oral Bacteria in Alzheimer’s Disease Autopsy Brains

The sporadic form of Alzheimer’s disease (AD) is the most common form of dementia characterized microscopically by the presence of amyloid-beta (Aβ) plaques and tau-neurofibrillary tangles with clinical presentation of cognitive deficit. Its etiology remains obscure. The cited literature from epidemiological studies suggests the presence of gingivitis and periodontitis almost double the risk for AD over a 10-year period. This feasibility study used 16S rDNA high throughput sequencing to evaluate the bacterial components of the oral microbiome in snap frozen human AD and non-AD control brains. Specimens (n = 20), 10 from AD brains and 10 from non-AD age-matched brains were subjected to molecular barcoding “blindly” with high throughput sequencing. Initial PCR treatment, using 14 different primer sets separately and in combination, identified 4 (40%) positive samples in the AD-group and 6 (60%) positive samples in the non-AD-group with bacterial species associated with the oral and the gastrointestinal tract. Brain cell lysates were validated for the presence of bacterial peptidoglycan and showed 8 out of 10 brains to be positive for this universal bacterial protein in both the AD and non-AD groups. Actinomycetales and Prevotella (bacterial marker for gingivitis) and Treponema and Veillonella, (periodontitis) were present exclusively in the AD group. This study confirmed Actinomycetales and Bacteroidales (Treponema and Veillonella species) were exclusively isolated from AD brain tissue, and supports other epidemiological which demonstrate gingivitis and periodontal disease to be associated with AD

Can oral infection be a risk factor for Alzheimer’s disease?

Alzheimer’s disease (AD) is a scourge of longevity that will drain enormous resources from public health budgets in the future. Currently, there is no diagnostic biomarker and/or treatment for this most common form of dementia in humans. AD can be of early familial-onset or sporadic with a late-onset. Apart from the two main hallmarks, amyloid-beta and neurofibrillary tangles, inflammation is a characteristic feature of AD neuropathology. Inflammation may be caused by a local central nervous system insult and/or by peripheral infections. Numerous microorganisms are suspected in AD brains ranging from bacteria (mainly oral and non-oral Treponema species), viruses (Herpes simplex type I) and yeasts (Candida species). A causal relationship between periodontal pathogens/non-oral Treponema species of bacteria has been proposed via the amyloid-beta and inflammatory links. Periodontitis constitutes a peripheral oral infection that can provide the brain with intact bacteria and virulence factors and inflammatory mediators due to daily, transient bacteraemias. If and when genetic risk factors meet environmental risk factors in the brain, disease is expressed, in which neurocognition may be impacted, leading to the development of dementia. To achieve the goal of finding a diagnostic biomarker and possible prophylactic treatment for AD, there is an initial need to solve the etiological puzzle contributing to its pathogenesis. This review therefore addresses oral infection as the plausible aetiology of late onset AD (LOAD)

Assessing the role of Porphyromonas gingivalis in periodontitis to determine a causative relationship with Alzheimer’s disease

Chronic periodontitis of ten years duration is reported to become a twofold risk factor for the development of Alzheimer’s disease (AD). Periodontitis is modifiable, and this fits with the current action plan for preventing AD. However, until periodontitis, becomes acknowledged as a firm risk factor for AD this risk will continue. Here we put forward our own argument based on the current literature for in vivo infection-mediated periodontal disease models supporting the antimicrobial protection hypothesis of AD and interventional studies supporting the causal links. Oral infections with Porphyromonas gingivalis, or introduction of its lipopolysaccharide (LPS), in various mouse models has demonstrated the development of key neuropathological hallmark lesions defining AD. These are extracellular amyloid-beta plaques, phosphorylated tau, neurofibrillary tangles, widespread acute and chronic inflammation, blood-brain barrier defects together with the clinical phenotype showing impaired learning and spatial memory. Live P. gingivalis and its LPS (commercial or from “microbullets”) are powerful peripheral and intracerebral inflammatory signaling initiators, and this has direct implications on memory and lesion development. Maintaining a healthy oral microbiome and managing periodontal disease with regular surveillance and good oral hygiene throughout life is likely to reduce the unnecessary burden of AD in some individuals

Chronic Porphyromonas gingivalis infection accelerates the occurrence of age-related granules in ApoE-/- mice brains

This study explored the origin of age-related granules in the apolipoprotein E gene knockout (ApoE−/−) B6 background mice brains following chronic gingival infection with Porphyromonas gingivalis for 24 weeks. Intracerebral localization of P. gingivalis was detected by fluorescence in situ hybridization (FISH) and its protease by immunohistochemistry. The age-related granules were observed by periodic acid–Schiff (PAS), silver impregnation, and immunostaining. FISH showed intracerebral dissemination of P. gingivalis cells (p = 0.001). PAS and silver impregnation demonstrated the presence of larger inclusions restricted to the CA1, CA2, and dentate gyrus sectors of the hippocampus. A specific monoclonal antibody to bacterial peptidoglycan detected clusters of granules with variable sizes in mice brains infected with P. gingivalis (p = 0.004), and also highlighted areas of diffuse punctate staining equating to physical tissue damage. Mouse immunoglobulin G was observed in the capillaries of the cerebral parenchyma of all P. gingivalis–infected brains (p = 0.001), and on pyramidal neurons in some severely affected mice, compared with the sham-infected mice. Gingipains was also observed in microvessels of the hippocampus in the infected mice. This study supports the possibility of early appearance of age-related granules in ApoE−/− mice following inflammation-mediated tissue injury, accompanied by loss of cerebral blood-brain barrier integrity

Risk assessment for the spread of Serratia marcescens within dental-unit waterline systems using Vermamoeba vermiformis

Vermamoeba vermiformis is associated with the biofilm ecology of dental-unit waterlines (DUWLs). This study investigated whether V. vermiformis is able to act as a vector for potentially pathogenic bacteria and so aid their dispersal within DUWL systems. Clinical dental water was initially examined for Legionella species by inoculating it onto Legionella selective-medium plates. The molecular identity/profile of the glassy colonies obtained indicated none of these isolates were Legionella species. During this work bacterial colonies were identified as a non-pigmented Serratia marcescens. As the water was from a clinical DUWL which had been treated with Alpron™ this prompted the question as to whether S. marcescens had developed resistance to the biocide. Exposure to Alpron™ indicated that this dental biocide was effective, under laboratory conditions, against S. marcescens at up to 1x108 colony forming units/millilitre (cfu/ml). V. vermiformis was cultured for eight weeks on cells of S. marcescens and Escherichia coli. Subsequent electron microscopy showed that V. vermiformis grew equally well on S. marcescens and E. coli (p = 0.0001). Failure to detect the presence of S. marcescens within the encysted amoebae suggests that V. vermiformis is unlikely to act as a vector supporting the growth of this newly isolated, nosocomial bacterium

In-situ characterization of the Hamamatsu R5912-HQE photomultiplier tubes used in the DEAP-3600 experiment

The Hamamatsu R5912-HQE photomultiplier-tube (PMT) is a novel high-quantum efficiency PMT. It is currently used in the DEAP-3600 dark matter detector and is of significant interest for future dark matter and neutrino experiments where high signal yields are needed. We report on the methods developed for in-situ characterization and monitoring of DEAP's 255 R5912-HQE PMTs. This includes a detailed discussion of typical measured single-photoelectron charge distributions, correlated noise (afterpulsing), dark noise, double, and late pulsing characteristics. The characterization is performed during the detector commissioning phase using laser light injected through a light diffusing sphere and during normal detector operation using LED light injected through optical fibres

The Porphyromonas gingivalis/host interactome shows enrichment in GWASdb genes related to Alzheimer’s disease, diabetes and cardiovascular diseases

Periodontal disease is of established aetiology in which polymicrobial synergistic ecology has become dysbiotic under the influence of Porphyromonas gingivalis. Following breakdown of the host’s protective oral tissue barriers, P. gingivalis migrates to developing inflammatory pathologies that associate with Alzheimer’s disease (AD). Periodontal disease is a risk factor for cardiovascular disorders (CVD), type II diabetes mellitus (T2DM), AD and other chronic diseases, whilst T2DM exacerbates periodontitis. This study analysed the relationship between the P. gingivalis/host interactome and the genes identified in genome-wide association studies (GWAS) for the aforementioned conditions using data from GWASdb (P<1E-03) and, in some cases, from the NCBI/EBI GWAS database (P< 1E-05). Gene expression data from periodontitis or P. gingivalis microarray was compared to microarray datasets from the AD hippocampus and/or from carotid artery plaques. The results demonstrated that the host genes of the P. gingivalis interactome were significantly enriched in genes deposited in GWASdb genes related to cognitive disorders, AD and dementia, and its co-morbid conditions T2DM, obesity, and CVD. The P. gingivalis/host interactome was also enriched in GWAS genes from the more stringent NCBI-EBI database for AD, atherosclerosis and T2DM. The misregulated genes in periodontitis tissue or P. gingivalis infected macrophages also matched those in the AD hippocampus or atherosclerotic plaques. Together, these data suggest important gene/environment interactions between P. gingivalis and susceptibility genes or gene expression changes in conditions where periodontal disease is a contributory factor

Complement in the pathogenesis of Alzheimer's disease

The emergence of complement as an important player in normal brain development and pathological remodelling has come as a major surprise to most scientists working in neuroscience and almost all those working in complement. That a system, evolved to protect the host against infection, should have these unanticipated roles has forced a rethink about what complement might be doing in the brain in health and disease, where it is coming from, and whether we can, or indeed should, manipulate complement in the brain to improve function or restore homeostasis. Complement has been implicated in diverse neurological and neuropsychiatric diseases well reviewed elsewhere, from depression through epilepsy to demyelination and dementia, in most complement drives inflammation to exacerbate the disease. Here, I will focus on just one disease, the most common cause of dementia, Alzheimer’s disease. I will briefly review the current understanding of what complement does in the normal brain, noting, in particular, the many gaps in understanding, then describe how complement may influence the genesis and progression of pathology in Alzheimer’s disease. Finally, I will discuss the problems and pitfalls of therapeutic inhibition of complement in the Alzheimer brain

Identification and Clonal Characterisation of a Progenitor Cell Sub-Population in Normal Human Articular Cartilage

Background: Articular cartilage displays a poor repair capacity. The aim of cell-based therapies for cartilage defects is to repair damaged joint surfaces with a functional replacement tissue. Currently, chondrocytes removed from a healthy region of the cartilage are used but they are unable to retain their phenotype in expanded culture. The resulting repair tissue is fibrocartilaginous rather than hyaline, potentially compromising long-term repair. Mesenchymal stem cells, particularly bone marrow stromal cells (BMSC), are of interest for cartilage repair due to their inherent replicative potential. However, chondrocyte differentiated BMSCs display an endochondral phenotype, that is, can terminally differentiate and form a calcified matrix, leading to failure in long-term defect repair. Here, we investigate the isolation and characterisation of a human cartilage progenitor population that is resident within permanent adult articular cartilage. Methods and Findings: Human articular cartilage samples were digested and clonal populations isolated using a differential adhesion assay to fibronectin. Clonal cell lines were expanded in growth media to high population doublings and karyotype analysis performed. We present data to show that this cell population demonstrates a restricted differential potential during chondrogenic induction in a 3D pellet culture system. Furthermore, evidence of high telomerase activity and maintenance of telomere length, characteristic of a mesenchymal stem cell population, were observed in this clonal cell population. Lastly, as proof of principle, we carried out a pilot repair study in a goat in vivo model demonstrating the ability of goat cartilage progenitors to form a cartilage-like repair tissue in a chondral defect. Conclusions: In conclusion, we propose that we have identified and characterised a novel cartilage progenitor population resident in human articular cartilage which will greatly benefit future cell-based cartilage repair therapies due to its ability to maintain chondrogenicity upon extensive expansion unlike full-depth chondrocytes that lose this ability at only seven population doublings