Comparison of image analysis software packages in the assessment of adhesion of microorganisms to mucosal epithelium using confocal laser scanning microscopy

We have compared current image analysis software packages in order to find the most useful one for assessing microbial adhesion and inhibition of adhesion to tissue sections. We have used organisms of different sizes, the bacterium Helicobacter pylori and the yeast Candida albicans. Adhesion of FITC-labelled H. pylori and C. albicans was assessed by confocal microscopy. Four different Image analysis software packages, NIH-Image, IP Lab, Image Pro+, and Metamorph, were compared for their ability to quantify adhesion of the two organisms and several quantification methods were devised for each package. For both organisms, the dynamic range that could be detected by the software packages was 1×106?1×109 cells/ml. Of the four software packages tested, our results showed that Metamorph software, using our ?Region of Interest? method, with the software's ?Standard Area Method? of counting, was the most suitable for quantifying adhesion of both organisms because of its unique ability to separate clumps of microbial cells. Moreover, fewer steps were required. By pre-incubating H. pylori with the glycoconjugate Lewis b-HSA, an inhibition of binding of 48.8% was achieved using 250 ?g/ml Lewis b-HSA. The method we have devised using Metamorph software, provides a simple, quick and accurate way of quantifying adhesion and inhibition of adhesion of microbial cells to the epithelial surface of tissue sections. The method can be applied to organisms ranging in size from small bacteria to larger yeast cells

Strong terahertz absorption using SiO2/Al based metamaterial structures

The article of record as published may be found at http://dx.doi.org/10.1063/1.3693407Metamaterial absorbers with nearly 100% absorption in the terahertz (THz) spectral band have been designed and fabricated using a periodic array of aluminum (Al) squares and an Al ground plane separated by a thin silicon dioxide (SiO2) dielectric film. The entire structure is less than 1.6 mm thick making it suitable for the fabrication of microbolometers or bi-material sensors for THz imaging. Films with different dielectric layer thicknesses exhibited resonant absorption at 4.1, 4.2, and 4.5 THz with strengths of 98%, 95%, and 88%, respectively. The measured absorption spectra are in good agreement with simulations using finite element modeling

POLARIZATION IN PION-PROTON SCATTERING FROM 670-3750 MeV/c

Using a polarized proton target, we have measured the polarization parameter P({theta}) in pion-proton scattering for both positive and negative pions. Because there seems to be a great deal of current interest in the analysis of pion-proton scattering we wish to present these experimental results at this time even though we have not yet completed their analysis. The measurement consisted of scattering pions from polarized target protons and observing the asymmetry in scattered intensity, I({theta}), as the target protons spin directions were reversed. The intensity for scattering from a target of polarization P{sub T} is I({theta}){sub pol.} = I({theta}){sub unpol.} (1 + P({theta})P{sub T}), where the parameter P({theta}) is the same as the recoil proton polarization in scattering pions from unpolarized protons under the assumption that parity is conserved in the process

ASYMMETRY IN n+ PHOTOPRODUCTION FROM A POLARIZED TARGET AT 5 AND16 GeV

The authors have measured the asymmetry in the cross section for the reaction {gamma}p {yields} {pi}{sup +}n between the two stages of polarization of the initial proton normal to the plane of scattering. The initial laboratory photon energies, k, were 5 GeV and 16 GeV, and the regions of momentum transfer, t, covered were 0.14 {le} {radical}-t {le} 1.01 GeV/c and 0.14 {le} {radical}-t {le} 0.78 GeV/c respectively. A butanol polarized target was used with the SLAC 20 GeV/c magnetic spectrometer. The data show a sizeable asymmetry at both 5 GeV and 16 GeV. The 16 GeV data peak at {radical}-t {approx} 0.30 GeV/c with an asymmetry of about -0.70, and the 5 GeV data pak at {radical}-t {approx} 0.80 GeV/c with an asymmetry of about -0.70. The direction of our normal to the scattering plane is along (photon in) x (pion out)

Genome Sequence of Fusobacterium nucleatum Subspecies Polymorphum — a Genetically Tractable Fusobacterium

Fusobacterium nucleatum is a prominent member of the oral microbiota and is a common cause of human infection. F. nucleatum includes five subspecies: polymorphum, nucleatum, vincentii, fusiforme, and animalis. F. nucleatum subsp. polymorphum ATCC 10953 has been well characterized phenotypically and, in contrast to previously sequenced strains, is amenable to gene transfer. We sequenced and annotated the 2,429,698 bp genome of F. nucleatum subsp. polymorphum ATCC 10953. Plasmid pFN3 from the strain was also sequenced and analyzed. When compared to the other two available fusobacterial genomes (F. nucleatum subsp. nucleatum, and F. nucleatum subsp. vincentii) 627 open reading frames unique to F. nucleatum subsp. polymorphum ATCC 10953 were identified. A large percentage of these mapped within one of 28 regions or islands containing five or more genes. Seventeen percent of the clustered proteins that demonstrated similarity were most similar to proteins from the clostridia, with others being most similar to proteins from other gram-positive organisms such as Bacillus and Streptococcus. A ten kilobase region homologous to the Salmonella typhimurium propanediol utilization locus was identified, as was a prophage and integrated conjugal plasmid. The genome contains five composite ribozyme/transposons, similar to the CdISt IStrons described in Clostridium difficile. IStrons are not present in the other fusobacterial genomes. These findings indicate that F. nucleatum subsp. polymorphum is proficient at horizontal gene transfer and that exchange with the Firmicutes, particularly the Clostridia, is common

Hydrolysis of γ -glutamyl linkages by Fusobacterium nucleatum

The cell extracts of two human oral strains (FN2 and FN3) of Fusobacterium nucleatum displayed exceptionally high γ -glutamylpeptidase activity as determined with N-γ - l -glutamyl-2-naphthylamine as substrate. This activity was so dominant that the hydrolysis of other N -aminoacyl-2-naphthylamines progressed at a rate <10% of the former. Two major enzymes (I and II) were partially purified from FN2. I had a molecular weight of 115,000 and did not hydrolyze γ -glutamylcysteinylglycine (glutathione). II had a molecular weight of 70,000 and rapidly liberated only glutamic acid from glutathione. Strain FN3 contained several enzymes hydrolyzing γ -glu-2NA. Direct anion exchange chromatography of FN3 cell extracts separated one enzyme that liberated both glutamic acid and glycine from glutathione, one that was inactive against glutathione (but hydrolyzed γ -glu-2NA), and one that liberated only glutamic acid. Although γ -glu-2NA was a good synthetic substrate, glutathione was hydrolyzed at least 500 times faster by an enzyme present in both strains. These results indicate that the presence of γ -glutamylpeptidase activity is very characteristic of these F. nucleatum strains.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/41334/1/284_2005_Article_BF02094016.pd

Amplified and selective assay of collagens by enzymatic and fluorescent reactions

Sensitive and selective assay of collagen is of substantial importance to the diagnostic study of health- and aging-related failures. In this paper, we describe a highly specific and sensitive method for the assay of whole collagens in biological samples using a novel fluorogenic reagent, 3,4- dihydroxyphenylacetic acid (3,4-DHPAA). The 3,4-DHPAA reagent can selectively detectN-terminal Gly-containing peptides (NGPs) in the presence of sodium borate and NaIO4. Under conditions optimized, this assay format for collagen, termed 3,4-DHPAA assay method showed a good linear relationship between the amplified FL signals and the collagen concentrations from 0.18 to 12 μg/ml. Therefore the sensitive determination of intracellular collagens in cheek tissue and HeLa cells was individually possible without any separation protocol. The dual recognitions of the collagens in the samples could be performed by the enzymatic digestion and the FL reaction. The proposed assay method enables the determination facile, specific, sensitive and quantitative for biogenic collagens