Natural medicine HLXL targets multiple pathways of amyloid-mediated neuroinflammation and immune response in treating alzheimer's disease.

Background: Based on the complex pathology of AD, a single chemical approach may not be sufficient to deal simultaneously with multiple pathways of amyloid-tau neuroinflammation. A polydrug approach which contains multiple bioactive components targeting multiple pathways in AD would be more appropriate. Here we focused on a Chinese medicine (HLXL), which contains 56 bioactive natural products identified in 11 medicinal plants and displays potent anti-inflammatory and immuno-modulatory activity.

Age-dependent changes in Porphyromonas gingivalis and Prevotella species/phylotypes in healthy gingiva and inflamed/diseased sub-gingival sites.

Objectives: Early colonisation of oral surfaces by periodontal pathogens presents a significant risk factor for subsequent development of destructive disease affecting tissues that support the dentition. The aims of the present study were to establish the age-dependent relationship between sub-gingival profiles of 22 Prevotella species/phylotypes in children, adolescents and adults from an isolated Aboriginal community and, further, to use this information to identify Prevotella species that could serve as microbial risk indicators.

Materials And Methods: DNA isolated from sub-gingival plaque samples (three healthy sites and three inflamed/diseased sites) from adults, adolescents and children was screened for Porphyromonas gingivalis load and 22 Prevotella species/phylotypes by species-specific PCR.

Lactobacilli are prominent in the initial stages of polymicrobial infection of dental pulp.

In earlier studies we used molecular methods to identify the major bacterial consortia associated with advanced dentin caries. These consortia are dominated by bacteria from the families Lactobacillaceae, Streptococcaceae, Veillonellaceae (formerly Acidaminococcaceae), Eubacteriaceae, and Lachnospiraceae from the phylum Firmicutes; Coriobacteriaceae, Bifidobacteriaceae, and Propionibacteriaceae from the phylum Actinobacteria; and Prevotellaceae from the phylum Bacteroidetes, as well as fusobacteria. The phases of infection of vital pulp tissue by dentin microorganisms remain obscure.

An observational study of the microbiome of the maternal pouch and saliva of the tammar wallaby, , and of the gastrointestinal tract of the pouch young.

Marsupial mammals, born in an extremely atricial state with no functional immune system, offer a unique opportunity to investigate both the developing microbiome and its relationship to that of the mother and the potential influence of this microbiome upon the development of the immune system. In this study we used a well-established marsupial model animal, Macropus eugenii, the tammar wallaby, to document the microbiome of three related sites: the maternal pouch and saliva, and the gastrointestinal tract (GIT) of the young animal. We used molecular-based methods, targeting the 16S rDNA gene to determine the bacterial diversity at these study sites.

Lysine gingipain (kgp) biovars of Porphyromonas gingivalis exhibit differential distribution on oral mucosal sites.

A predominant kgp biovar colonized subgingival sites and buccal and tongue mucosa in 45 of 56 adults in an isolated community. The presence of biovars 381, W83, and W83v, but not HG66, correlated with the Porphyromonas gingivalis load at diseased sites. Biovars W83 and W83v poorly colonized tongue and buccal mucosa.

Methods for optimizing DNA extraction before quantifying oral bacterial numbers by real-time PCR.

Methods for the optimal extraction of genomic DNA for real-time PCR enumeration of oral bacteria using the muramidase, mutanolysin, were developed using a simple in vitro oral flora model comprised of the facultative anaerobic gram-positive bacteria, Lactobacillus acidophilus and Streptococcus mutans, the gram-positive anaerobe, Parvimonas micra, and the gram-negative anaerobes, Porphyromonas gingivalis, Prevotella melaninogenica and Fusobacterium nucleatum. Traditional, as well as more elaborate, methods of quantifying bacterial numbers, including colony counting and estimation of DNA content using 4',6-diamino-2-phenylindole were compared in order to validate the real-time PCR approach. Evidence was obtained that P.

Identification and expression of a novel marsupial cathelicidin from the tammar wallaby (Macropus eugenii).

Cathelicidins are important components of the innate immune system and have been identified in skin and epithelia of a range of mammals. In this study molecular techniques, including RACE-PCR, were used to identify the full cDNA sequence of a cathelicidin gene, MaeuCath8, from the Australian marsupial, the tammar wallaby, Macropus eugenii. This cathelicidin was not homologous to other such genes previously isolated from a tammar wallaby mammary gland EST library, however, it did contain 4 conserved cysteine residues which characterise the pre-propeptide and had 80% identity with a previously isolated bandicoot cathelicidin.

Immunohistochemistry using antibodies to the cathelicidin LL37/hCAP18 in the tammar wallaby, Macropus eugenii.

Antibodies to the human cathelicidin, hCAP18 have been used to examine epithelial tissues of adult and pouch young marsupials. Immunoreactivity was observed in skin, gastrointestinal tract, lung and mammary node of adults as well as skin, gastrointestinal tract, lung and bone marrow of pouch young. The locations of expression were similar to that reported in human tissues.

The microbiome of the cloacal openings of the urogenital and anal tracts of the tammar wallaby, .

The bacterial diversity of the openings of the urogenital and anal tracts of the adult female tammar wallaby, Macropus eugenii, was determined in order to ascertain whether the physical proximity of the openings of these tracts within the cloaca affected the two populations of bacteria. Terminal restriction fragment length polymorphism (T-RFLP) analyses of 42 wallabies identified 81 different terminal fragments, indicative of diverse and complex microbiomes at these anatomical locations. Subsequent amplified rDNA restriction analysis (ARDRA) identified 72 phylotypes from the urogenital tract and 50 from the anal tract.

Structure of N-acetyl-beta-D-glucosaminidase (GcnA) from the endocarditis pathogen Streptococcus gordonii and its complex with the mechanism-based inhibitor NAG-thiazoline.

The crystal structure of GcnA, an N-acetyl-beta-D-glucosaminidase from Streptococcus gordonii, was solved by multiple wavelength anomalous dispersion phasing using crystals of selenomethionine-substituted protein. GcnA is a homodimer with subunits each comprised of three domains. The structure of the C-terminal alpha-helical domain has not been observed previously and forms a large dimerisation interface.

Veillonella denticariosi sp. nov., isolated from human carious dentine.

Selective culture of human carious dentine for Veillonella strains resulted in the isolation of two strains of a Gram-negative, coccus-shaped bacterium that has not been described previously. Comparative 16S rRNA and dnaK gene sequence analysis indicated that the two strains were homogeneous and comprised a distinct lineage within the genus Veillonella, phylogenetically most closely related to Veillonella rodentium. This was supported by DNA-DNA hybridization, which showed clearly that the two strains were similar and distinct from other Veillonella species, and the production of major cellular fatty acids (C(13 : 0) and C(17 : 1)omega8), which is consistent with other members of the genus Veillonella.

Interventions for prevention of otitis media may be most effective if implemented in the first weeks of life.

Objective: For Indigenous Australian children living in remote communities, onset of otitis media commences within weeks of birth and is associated with early nasopharyngeal colonisation with multiple respiratory bacterial pathogens: Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. The high prevalence of eardrum perforation and the failure of standard therapies to cure or prevent OM in this population require urgent attention. The objective of this study was to measure the changes in nasopharyngeal bacterial flora between birth and first episode of otitis media.

Measuring nasal bacterial load and its association with otitis media.

Background: Nasal colonisation with otitis media (OM) pathogens, particularly Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis, is a precursor to the onset of OM. Many children experience asymptomatic nasal carriage of these pathogens whereas others will progress to otitis media with effusion (OME) or suppurative OM. We observed a disparity in the prevalence of suppurative OM between Aboriginal children living in remote communities and non-Aboriginal children attending child-care centres; up to 60% and <1%, respectively.

Two-dimensional fluorescence difference gel electrophoretic analysis of Streptococcus mutans biofilms.

Compared with traditional two-dimensional (2D) proteome analysis of Streptococcus mutans grown as a biofilm from a planktonic culture at steady state (Rathsam et al., Microbiol. 2005, 151, 1823-1837), the use of 2D fluorescence difference gel electrophoresis (DIGE) led to a 3-fold increase in the number of identified protein spots that were significantly altered in their level of expression (P < 0.

Molecular analysis of microbial diversity in advanced caries.

Real-time PCR analysis of the total bacterial load in advanced carious lesions has shown that the total load exceeds the number of cultivable bacteria. This suggests that an unresolved complexity exists in bacteria associated with advanced caries. In this report, the profile of the microflora of carious dentine was explored by using DNA extracted from 10 lesions selected on the basis of comparable total microbial load and on the relative abundance of Prevotella spp.

Carious dentine provides a habitat for a complex array of novel Prevotella-like bacteria.

Previous analysis of the microbiology of advanced caries by culture and real-time PCR emphasized the high incidence and abundance of gram-negative anaerobic species, particularly Prevotella-like bacteria. The diversity of Prevotella-like bacteria was further explored by analyzing pooled bacterial DNA from lesions of carious dentine. This was achieved by amplification of a region of the 16S ribosomal DNA with a Prevotella genus-specific forward primer and a universal bacterial reverse primer, followed by cloning and sequencing.

Distribution of Porphyromonas gingivalis biotypes defined by alleles of the kgp (Lys-gingipain) gene.

Paired subgingival plaque samples representing the most-diseased and least-diseased sites were collected from 34 adult patients with diagnosed chronic periodontitis. The percentage of Porphyromonas gingivalis relative to the total anaerobic and gram-negative bacterial load at each site was determined by real-time PCR. Based on variations in the noncatalytic C terminus of the Lys-gingipain (Kgp), it was reasoned that DNA sequence variation in the 3'-coding region of the kgp gene might determine functional biotypes.

Quantitative analysis of diverse Lactobacillus species present in advanced dental caries.

Our previous analysis of 65 advanced dental caries lesions by traditional culture techniques indicated that lactobacilli were numerous in the advancing front of the progressive lesion. Production of organic acids by lactobacilli is considered to be important in causing decalcification of the dentinal matrix. The present study was undertaken to define more precisely the diversity of lactobacilli found in this environment and to quantify the major species and phylotypes relative to total load of lactobacilli by real-time PCR.

Characterisation of a novel homodimeric N-acetyl-beta-D-glucosaminidase from Streptococcus gordonii.

An N-acetyl-beta-D-glucosaminidase (GcnA) from Streptococcus gordonii FSS2 was cloned and sequenced. GcnA had a deduced molecular mass of 72,120 Da. The molecular weight after gel-filtration chromatography was 140,000 Da and by SDS-PAGE was 70,000 Da, indicating that the native protein was a homodimer.

Proteome analysis of Streptococcus mutans metabolic phenotype during acid tolerance.

Two-dimensional gel electrophoretic analysis of the proteome of Streptococcus mutans grown at a steady state in a glucose-limited anaerobic continuous culture revealed a number of proteins that were differentially expressed when the growth pH was lowered from pH 7.0 to pH 5.0.

Stress-responsive proteins are upregulated in Streptococcus mutans during acid tolerance.

Streptococcus mutans is an important pathogen in the initiation of dental caries as the bacterium remains metabolically active when the environment becomes acidic. The mechanisms underlying this ability to survive and proliferate at low pH remain an area of intense investigation. Differential two-dimensional electrophoretic proteome analysis of S.

Cellular and extracellular proteome analysis of Streptococcus mutans grown in a chemostat.

The oral pathogen, Streptococcus mutans, was grown under glucose limitation in a chemostat at pH 7.0 and a dilution rate of 0.1 h(-1) to mimic the conditions prevailing in a healthy human oral cavity in between meal times.

Fructan formation in transgenic white clover expressing a fructosyltransferase from Streptococcus salivarius.

White clover (Trifolium repens L.) is an important pasture legume that does not normally accumulate fructan as a storage carbohydrate. We have generated transgenic white clover plants that accumulate fructan, by expressing the fructosyltransferase (Ftf) enzyme from the bacterium Streptococcus salivarius under the control of the cauliflower mosaic virus (CaMV) 35S promoter.

Expression of a streptococcal glucosyltransferase as a fusion to a solute-binding protein in Lactobacillus fermentum BR11.

BspA is a non-covalently anchored cystine-binding protein from Lactobacillus fermentum BR11. It has previously been used to present antigens derived from infectious organisms on the L. fermentum BR11 cell surface.

Quantitative microbiological study of human carious dentine by culture and real-time PCR: association of anaerobes with histopathological changes in chronic pulpitis.

The bacteria found in carious dentine were correlated with the tissue response of the dental pulps of 65 teeth extracted from patients with advanced caries and pulpitis. Standardized homogenates of carious dentine were plated onto selective and nonselective media under anaerobic and microaerophilic conditions. In addition, real-time PCR was used to quantify the recovery of anaerobic bacteria.