Use of Fluorescent Microspheres as a Tool To Investigate Bacterial Interactions with Growing Plants

2005 ◽  
Vol 68 (4) ◽  
pp. 870-873 ◽  
Author(s):  
ETHAN B. SOLOMON ◽  
KARL R. MATTHEWS
Keyword(s):  
Foodborne Pathogens ◽  
Laser Scanning ◽  
Leaf Tissue ◽  
Laser Scanning Confocal Microscopy ◽  
Escherichia Coli O157 ◽  
Fluorescent Microspheres ◽  
E Coli ◽  
Bacterial Interactions ◽  
Scanning Confocal Microscopy ◽  
Tissue Sections

Foodborne pathogens may exist as endophytes of growing plants. The internalization of Escherichia coli O157:H7 or other foodborne pathogens in growing lettuce plants may be independent of microbial factors. Mature lettuce plants were surface irrigated with E. coli O157:H7 or with FluoSpheres (fluorescent microspheres) and harvested 1, 3, and 5 days post-exposure. FluoSpheres were utilized as a bacterial surrogate. Microscopic examination of root, stem, and leaf tissue sections revealed that FluoSpheres were internalized into growing plants. Laser scanning confocal microscopy revealed that FluoSpheres were present within the root tissue and leaf stem tissue. The presence of FluoSpheres in internal portions of stem and leaf tissue suggests transport of the spheres from the root upward into the edible tissue. The level of uptake of FluoSpheres and E. coli O157:H7 was quantified using filtration. Numbers of FluoSpheres and E. coli O157:H7 cells in plant tissue were similar. The entry of E. coli O157:H7 into lettuce plants may be a passive event because the concentration of FluoSpheres was similar to that of the pathogen.

Automated 3-D cell population analysis in thick tissue sections using laser-scanning confocal-microscopy data

1993 ◽  
Vol 51 ◽  
pp. 562-563
Author(s):  
Hakan Ancin
Keyword(s):  
Laser Scanning ◽  
Population Analysis ◽  
Three Dimensional ◽  
Large Data ◽  
Laser Scanning Confocal Microscopy ◽  
Tissue Slices ◽  
Scanning Confocal Microscopy ◽  
Tissue Sections ◽  
Spatially Adaptive ◽  
Microscopy Data

This paper presents methods for performing detailed quantitative automated three dimensional (3-D) analysis of cell populations in thick tissue sections while preserving the relative 3-D locations of cells. Specifically, the method disambiguates overlapping clusters of cells, and accurately measures the volume, 3-D location, and shape parameters for each cell. Finally, the entire population of cells is analyzed to detect patterns and groupings with respect to various combinations of cell properties. All of the above is accomplished with zero subjective bias.In this method, a laser-scanning confocal light microscope (LSCM) is used to collect optical sections through the entire thickness (100 - 500μm) of fluorescently-labelled tissue slices. The acquired stack of optical slices is first subjected to axial deblurring using the expectation maximization (EM) algorithm. The resulting isotropic 3-D image is segmented using a spatially-adaptive Poisson based image segmentation algorithm with region-dependent smoothing parameters. Extracting the voxels that were labelled as "foreground" into an active voxel data structure results in a large data reduction.

scholarly journals Activities associated with the putative replication initiation protein of Coconut foliar decay virus, a tentative member of the genus Nanovirus

2000 ◽  
Vol 81 (12) ◽  
pp. 3099-3106 ◽  
Author(s):  
Andres Merits ◽  
Oleg N. Fedorkin ◽  
Deyin Guo ◽  
Natalia O. Kalinina ◽  
Sergey Yu. Morozov
Keyword(s):  
Laser Scanning ◽  
Recombinant Baculovirus ◽  
Laser Scanning Confocal Microscopy ◽  
Replication Initiation ◽  
Rep Protein ◽  
Ssdna Binding ◽  
E Coli ◽  
Replication Initiation Protein ◽  
Scanning Confocal Microscopy ◽  
Tentative Member

The putative replication initiation protein (Rep) of Coconut foliar decay virus (CFDV) was expressed as a 6× His recombinant protein in E. coli and in recombinant baculovirus. Purified 6× His–Rep protein was demonstrated to possess sequence non-specific RNA- and ssDNA-binding activities as well as magnesium-dependent ATPase/GTPase activity. The yeast two-hybrid system revealed that CFDV Rep could interact with itself. Subcellular distribution of the CFDV Rep was studied by fractionation of insect cells infected with recombinant baculovirus expressing the 6× His–Rep protein and by laser scanning confocal microscopy of Nicotiana benthamiana epidermal cells bombarded with a construct encoding CFDV Rep fused to GFP. It was shown that CFDV Rep associated predominantly with nuclei and membranes of infected/transfected cells. These activities of CFDV-encoded Rep are very similar to those reported for Reps of geminiviruses.

Penetration of Surface-Inoculated Bacteria as a Result of Hydrodynamic Shock Wave Treatment of Beef Steaks

2002 ◽  
Vol 65 (4) ◽  
pp. 616-620 ◽  
Author(s):  
T. A. LORCA ◽  
M. D. PIERSON ◽  
J. R. CLAUS ◽  
J. D. EIFERT ◽  
J. E. MARCY ◽  
...  
Keyword(s):  
Shock Wave ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Laser Scanning Confocal Microscopy ◽  
Exterior Surface ◽  
Safety Hazard ◽  
E Coli ◽  
Scanning Confocal Microscopy ◽  
Beef Steaks ◽  
Green Fluorescent

The top surface of the raw eye of round steaks was inoculated with either green fluorescent protein (GFP)-labeled Escherichia coli (E. coli-GFP) or rifampin-resistant E. coli (E. coli-rif). Cryostat sampling in concert with laser scanning confocal microscopy (LSCM) or plating onto antibiotic selective agar was used to determine if hydrodynamic shock wave (HSW) treatment resulted in the movement of the inoculated bacteria from the outer inoculated surface to the interior of intact beef steaks. HSW treatment induced the movement of both marker bacteria into the steaks to a maximum depth of 300 μm (0.3 mm). Because popular steak-cooking techniques involve the application of heat from the exterior surface of the steak to achieve internal temperatures ranging from 55 to 82°C, the extent of bacterial penetration observed in HSW-treated steaks does not appear to pose a safety hazard to consumers.

scholarly journals Morphophysiological Changes in Staphylococcus aureus Biofilms Treated with Plasma-Activated Hydrogen Peroxide Solution

2021 ◽  
Vol 11 (24) ◽  
pp. 11597
Author(s):  
Jianying Zhao ◽  
Jing Qian ◽  
Ji Luo ◽  
Mingming Huang ◽  
Wenjing Yan ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Staphylococcus Aureus ◽  
Foodborne Pathogens ◽  
Laser Scanning ◽  
Laser Scanning Confocal Microscopy ◽  
Hydrogen Peroxide Solution ◽  
Metabolic Capacity ◽  
Electron Microscopic ◽  
Scanning Confocal Microscopy ◽  
Barrier Discharge Plasma

Plasma-activated solution has attracted more attention in the food industry due to no chemical residue and good bacteriostatic properties. This study aimed to evaluate the effects of plasma-activated hydrogen peroxide solution (PAH) on the morphophysiology of Staphylococcus aureus biofilms. PAH was prepared using dielectric-barrier-discharge plasma and incubated with S. aureus biofilms for 0–40 min. Changes in biofilm morphophysiology were evaluated with laser scanning confocal microscopy, electron microscopic images, reactive oxygen species (ROS) content, metabolic capacity, and 1% agarose gel. Results indicated that the population of S. aureus in the biofilms was reduced by 4.04-log after incubation with PAH for 30 min. The thickness and metabolic capacity of biofilms were decreased, the ROS content and DNA fragments of bacteria increased after PAH treatments. Data suggested that PAH treatments significantly destroyed the morphophysiology of S. aureus (ATCC 6538) biofilms and could be considered as a valuable anti-biofilm technology to reduce foodborne pathogens on food and/or in food facilities.

scholarly journals Transmission of Escherichia coli O157:H7 from Contaminated Manure and Irrigation Water to Lettuce Plant Tissue and Its Subsequent Internalization

2002 ◽  
Vol 68 (1) ◽  
pp. 397-400 ◽  
Author(s):  
Ethan B. Solomon ◽  
Sima Yaron ◽  
Karl R. Matthews
Keyword(s):  
Escherichia Coli ◽  
Irrigation Water ◽  
Plant Tissue ◽  
Laser Scanning ◽  
Laser Scanning Confocal Microscopy ◽  
Epifluorescence Microscopy ◽  
Lettuce Plant ◽  
E Coli ◽  
Scanning Confocal Microscopy ◽  
Viable Cells

ABSTRACT The transmission of Escherichia coli O157:H7 from manure-contaminated soil and irrigation water to lettuce plants was demonstrated using laser scanning confocal microscopy, epifluorescence microscopy, and recovery of viable cells from the inner tissues of plants. E. coli O157:H7 migrated to internal locations in plant tissue and was thus protected from the action of sanitizing agents by virtue of its inaccessibility. Experiments demonstrate that E. coli O157:H7 can enter the lettuce plant through the root system and migrate throughout the edible portion of the plant.

A Microscopic Method to Visualize Escherichia coli Interaction with Beef Muscle

2000 ◽  
Vol 63 (4) ◽  
pp. 427-433 ◽  
Author(s):  
P. PRACHAIYO ◽  
L. A. McLANDSBOROUGH
Keyword(s):  
Escherichia Coli ◽  
Laser Scanning ◽  
Fluorescent Dyes ◽  
Fluorescent Protein ◽  
Nile Red ◽  
Laser Scanning Confocal Microscopy ◽  
E Coli ◽  
Scanning Confocal Microscopy ◽  
Meat Samples ◽  
Beef Muscle

The genetic determinant for enhanced green fluorescent protein (EGFP) was introduced into Escherichia coli JM109 (ATCC 53323) and E. coli O157:H7 (ATCC 43895) on plasmid EGFP. The expression of EGFP did not change the growth kinetics or surface properties tested (hydrophobicity and electrophoretic mobility). Microscope slides were modified to allow for optimal viewing of thick meat samples with an inverted microscope. Two fluorescent dyes, nile red and Cy3 were used to stain for lipid and protein portions of beef muscle, respectively. Laser scanning confocal microscopy was used to observe interaction of the EGFP-expressing E. coli strains and the fluorescently stained muscle components without changing the spatial and temporal environment of the organisms.

Laser Scanning Confocal Microscopy and Three-Dimesnsional Reconstruction of the Mitotic Spindles of Unequally Dividing Embryonic Cells

1990 ◽  
Vol 48 (3) ◽  
pp. 166-167
Author(s):  
J. Holy ◽  
G. Schatten
Keyword(s):  
Confocal Microscopy ◽  
Mitotic Spindle ◽  
Laser Scanning ◽  
Three Dimensional ◽  
Laser Scanning Confocal Microscopy ◽  
Two Dimensions ◽  
Embryonic Cells ◽  
Mitotic Spindles ◽  
Cleavage Division ◽  
Scanning Confocal Microscopy

One of the classic limitations of light microscopy has been the fact that three dimensional biological events could only be visualized in two dimensions. Recently, this shortcoming has been overcome by combining the technologies of laser scanning confocal microscopy (LSCM) and computer processing of microscopical data by volume rendering methods. We have employed these techniques to examine morphogenetic events characterizing early development of sea urchin embryos. Specifically, the fourth cleavage division was examined because it is at this point that the first morphological signs of cell differentiation appear, manifested in the production of macromeres and micromeres by unequally dividing vegetal blastomeres.The mitotic spindle within vegetal blastomeres undergoing unequal cleavage are highly polarized and develop specialized, flattened asters toward the micromere pole. In order to reconstruct the three-dimensional features of these spindles, both isolated spindles and intact, extracted embryos were fluorescently labeled with antibodies directed against either centrosomes or tubulin.

scholarly journals Advanced Static and Dynamic Fluorescence Microscopy Techniques to Investigate Drug Delivery Systems

Pharmaceutics ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 861
Author(s):  
Jacopo Cardellini ◽  
Arianna Balestri ◽  
Costanza Montis ◽  
Debora Berti
Keyword(s):  
Drug Delivery ◽  
Fluorescence Microscopy ◽  
Drug Delivery Systems ◽  
Laser Scanning ◽  
Super Resolution ◽  
Laser Scanning Confocal Microscopy ◽  
Delivery Systems ◽  
Scanning Confocal Microscopy ◽  
Biological Phenomena

In the past decade(s), fluorescence microscopy and laser scanning confocal microscopy (LSCM) have been widely employed to investigate biological and biomimetic systems for pharmaceutical applications, to determine the localization of drugs in tissues or entire organisms or the extent of their cellular uptake (in vitro). However, the diffraction limit of light, which limits the resolution to hundreds of nanometers, has for long time restricted the extent and quality of information and insight achievable through these techniques. The advent of super-resolution microscopic techniques, recognized with the 2014 Nobel prize in Chemistry, revolutionized the field thanks to the possibility to achieve nanometric resolution, i.e., the typical scale length of chemical and biological phenomena. Since then, fluorescence microscopy-related techniques have acquired renewed interest for the scientific community, both from the perspective of instrument/techniques development and from the perspective of the advanced scientific applications. In this contribution we will review the application of these techniques to the field of drug delivery, discussing how the latest advancements of static and dynamic methodologies have tremendously expanded the experimental opportunities for the characterization of drug delivery systems and for the understanding of their behaviour in biologically relevant environments.

Observation of Fine Structure in Bicontinuous Phase-Separated Domains of a Polymer Blend by Laser Scanning Confocal Microscopy

2001 ◽  
Vol 34 (15) ◽  
pp. 5186-5191 ◽  
Author(s):  
Hiroshi Jinnai ◽  
Hiroshi Yoshida ◽  
Kohtaro Kimishima ◽  
Yoshinori Funaki ◽  
Yoshitsugu Hirokawa ◽  
...  
Keyword(s):  
Fine Structure ◽  
Confocal Microscopy ◽  
Polymer Blend ◽  
Laser Scanning ◽  
Laser Scanning Confocal Microscopy ◽  
Scanning Confocal Microscopy

scholarly journals The DNA content of trophozoites and cysts of Giardia lamblia by microdensitometric quantitation of Feulgen staining and examination by laser scanning confocal microscopy.

1994 ◽  
Vol 42 (11) ◽  
pp. 1413-1416 ◽  
Author(s):  
S L Erlandsen ◽  
E M Rasch
Keyword(s):  
Confocal Microscopy ◽  
Genome Size ◽  
Dna Content ◽  
Giardia Lamblia ◽  
Laser Scanning ◽  
Laser Scanning Confocal Microscopy ◽  
Evolutionary Development ◽  
Scanning Confocal Microscopy ◽  
Dividing Cells ◽  
C Value

We investigated direct measurement of the DNA content of the parasitic intestinal flagellate Giardia lamblia through quantitation by Feulgen microspectrophotometry and also by visualization of Feulgen-stained DNA chromosomes within dividing cells by laser scanning confocal microscopy. Individual trophozoites of Giardia (binucleate) contained 0.144 +/- 0.018 pg of DNA/cell or 0.072 pg DNA/nucleus. Giardia lamblia cysts (quadranucleate) contained 0.313 +/- 0.003 pg DNA or 0.078 pg DNA/nucleus. The genome size (C) value per nucleus ranged between 6.5-7.1 x 10(7) BP for trophozoites and cysts, respectively. Confocal microscopic examination of Giardia trophozoites undergoing binary fission revealed five chromosome-like bodies within each nucleus. Further information about genome size and DNA content within different Giardia species may help to clarify the pivotal role of these primitive eukaryotic cells in evolutionary development.

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