scholarly journals Hidden Components of Microbiome Diversity Revealed by Cell Wall Lysis with Adaptive Focused Acoustics

2020 ◽  
Author(s):  
Shurjo Kumar Sen ◽  
Giselle F. Wallace ◽  
Sonia Ahluwalia ◽  
Vishal Thovarai ◽  
James Laugharn ◽  
...  
Keyword(s):  
Cell Wall ◽  
Wall Thickness ◽  
Energy Levels ◽  
Functional Characterization ◽  
Cell Wall Thickness ◽  
16S Sequencing ◽  
Alpha And Beta Diversity ◽  
Dna Yield ◽  
Mechanical Lysis

Abstract Within the rapidly evolving field of microbiome sequencing, a primary need exists for experimentally capturing microbiota in a manner as close as possible to their in vivo composition. During microbiome profiling, the first step necessarily involves lysis of the cell wall, releasing nucleic acids for next-generation sequencing. Microbial cell wall thicknesses can vary between 5nm to 80nm; while some species are quite easy to lyse, others are particularly resistant to lysis. Despite this, current chemical/mechanical lysis protocols ignore the possibility that species with different cell wall thicknesses are lysed at differential rates. This creates noise in species compositions and possibly skews current microbiome results in ways that are not currently understood. To develop a cell wall thickness-agnostic lysis protocol, we used Adaptive Focused Acoustics (AFA), a tunable acoustic methodology for processing of biological samples. Using identical aliquots of mouse stool homogenate as the lysis substrate, we compared AFA with chemical/mechanical lysis methodology routinely used in microbiome studies and found that AFA-mediated lysis substantially increases both microbial DNA yield as well as alpha and beta diversity. By starting with lower AFA energy levels, sequentially removing aliquots at each step, and subjecting the remainder to progressively stronger AFA treatment, we developed a sequential lysis method that accounts for differences in cell wall thickness. This method revealed even greater levels of diversity than single-timepoint AFA treatment. 16S sequencing results from the above experiments were verified by shotgun metagenome sequencing of a subset of the AFA samples. We found that lysis-induced noise affects not just species compositions, but also functional characterization of shotgun metagenome data. AFA samples also showed a higher detection of eukaryotic and fungal DNA. We suggest that AFA-mediated lysis produces a truer representation of the native microbiota, and that this method deserves consideration as a potential addition to microbiome lysis protocols.

scholarly journals Hidden components of microbiome diversity revealed by cell wall lysis with Adaptive Focused Acoustics

2020 ◽  
Author(s):  
Giselle F. Wallace ◽  
Sonia Ahluwalia ◽  
Vishal Thovarai ◽  
James Laugharn ◽  
Hamid Khoja ◽  
...  
Keyword(s):  
Cell Wall ◽  
Wall Thickness ◽  
Energy Levels ◽  
Functional Characterization ◽  
Cell Wall Thickness ◽  
16S Sequencing ◽  
Alpha And Beta Diversity ◽  
Dna Yield ◽  
Mechanical Lysis

AbstractWithin the rapidly evolving field of microbiome sequencing, a primary need exists for experimentally capturing microbiota in a manner as close as possible to their in vivo composition. During microbiome profiling, the first step necessarily involves lysis of the cell wall, releasing nucleic acids for next-generation sequencing. Microbial cell wall thicknesses can vary between 5nm to 80nm; while some species are quite easy to lyse, others are particularly resistant to lysis. Despite this, current chemical/mechanical lysis protocols ignore the possibility that species with different cell wall thicknesses are lysed at differential rates. This creates noise in species compositions and possibly skews current microbiome results in ways that are not currently understood. To develop a cell wall thickness-agnostic lysis protocol, we used Adaptive Focused Acoustics (AFA), a tunable acoustic methodology for processing of biological samples. Using identical aliquots of mouse stool homogenate as the lysis substrate, we compared AFA with chemical/mechanical lysis methodology routinely used in microbiome studies and found that AFA-mediated lysis substantially increases both microbial DNA yield as well as alpha and beta diversity. By starting with lower AFA energy levels, sequentially removing aliquots at each step, and subjecting the remainder to progressively stronger AFA treatment, we developed a sequential lysis method that accounts for differences in cell wall thickness. This method revealed even greater levels of diversity than single-timepoint AFA treatment. 16S sequencing results from the above experiments were verified by shotgun metagenome sequencing of a subset of the AFA samples. We found that lysis-induced noise affects not just species compositaions, but also functional characterization of shotgun metagenome data. AFA samples also showed a higher detection of eukaryotic and fungal DNA. We suggest that AFA-mediated lysis produces a truer representation of the native microbiota, and that this method deserves consideration as a potential addition to microbiome lysis protocols.

Properties of Gliricidia Wood (Gliricidia sepium) lntercropped with Cocoa (Theobroma cocoa) in Malaysia

2014 ◽  
Vol 11 (1) ◽  
pp. 51
Author(s):  
Mohd Helmy Ibrahim ◽  
Mohd Nazip Suratman ◽  
Razali Abd Kader
Keyword(s):  
Cell Wall ◽  
Specific Gravity ◽  
Moisture Content ◽  
Wall Thickness ◽  
Age Groups ◽  
Gliricidia Sepium ◽  
Cell Wall Thickness ◽  
Tree Age ◽  
Lumen Diameter ◽  
Significant Difference

Trees planted from agroforestry practices can become valuable resources in meeting the wood requirements of many nations. Gliricidia sepium is an exotic species introduced to the agricultural sector in Malaysia mainly for providing shade for cocoa and coffee plantations. This study investigates its wood physical properties (specific gravity and moisture content) and fibre morphology (length, lumen diameter and cell wall thickness) of G. sepium at three intervals according to age groups ( three, five and seven years of ages). Specific gravity (0.72) was significantly higher at seven years ofage as compared to five (0.41) and three (0.35) years age group with a mean of 0.43 (p<0.05). Mean moisture content was 58.3% with no significant difference existing between the tree age groups. Fibre diameter (22.4 mm) was significantly lower (p<0.05) for the trees which were three years of age when compared to five and seven years age groups (26.6 mm and 24. 7 mm), respectively. Means of fibre length, lumen diameter and cell wall thickness were 0.83 mm, 18.3 mm, and 6.2 mm, respectively, with no significant differences detected between trees in all age groups. Further calculation on the coefficient of suppleness and runkel ratio suggest that wood from G.sepium may have the potential for insulation board manufacturing and paper making. However, future studies should experiment the utilisation of this species for these products to determine its full potential.

Characterization of microstructures of SAN foam core using micro-computed tomography

2021 ◽  
pp. 026248932110068
Author(s):  
Youming Chen ◽  
Raj Das ◽  
Hui Wang ◽  
Mark Battley
Keyword(s):  
Cell Wall ◽  
Cell Size ◽  
Wall Thickness ◽  
Strain Localization ◽  
Cell Wall Thickness ◽  
Micro Computed Tomography ◽  
Average Cell Size ◽  
Average Cell ◽  
3D Images ◽  
Compressive Tests

In this study, the microstructure of a SAN foam was imaged using a micro-CT scanner. Through image processing and analysis, variations in density, cell wall thickness and cell size in the foam were quantitatively explored. It is found that cells in the foam are not elongated in the thickness (or rise) direction of foam sheets, but rather equiaxed. Cell walls in the foam are significantly straight. Density, cell size and cell wall thickness all vary along the thickness direction of foam sheets. The low density in the vicinity of one face of foam sheets leads to low compressive stiffness and strength, resulting in the strain localization observed in our previous compressive tests. For M80, large open cells on the top face of foam sheets are likely to buckle in compressive tests, therefore being another potential contributor to the strain localization as well. The average cell wall thickness measured from 2D slice images is around 1.4 times that measured from 3D images, and the average cell size measured from 2D slice images is about 13.8% smaller than that measured from 3D images. The dispersions of cell wall thickness measured from 2D slice images are 1.16–1.20 times those measured from 3D images. The dispersions of cell size measured from 2D slice images are 1.12–1.36 times those measured from 3D images.

Summer temperature in northeastern Siberia since 1642 reconstructed from tracheid dimensions and cell numbers of Larix cajanderi

2003 ◽  
Vol 33 (10) ◽  
pp. 1905-1914 ◽  
Author(s):  
Irina P Panyushkina ◽  
Malcolm K Hughes ◽  
Eugene A Vaganov ◽  
Martin AR Munro
Keyword(s):  
Cell Wall ◽  
Wall Thickness ◽  
Cell Number ◽  
Cell Wall Thickness ◽  
Cell Dimension ◽  
Cell Numbers ◽  
Age Related ◽  
Cambial Age ◽  
Northeastern Siberia ◽  
Larix Cajanderi

We reconstructed air temperature for two periods in the growth season from cell dimension and cell number variability in cross-dated tree rings of Larix cajanderi Mayr. from northeastern Siberia. Thirteen tree-ring chronologies based on cell size, cell wall thickness, and cell number were developed for AD 1642–1993. No clear evidence was found of an age-related trend in cell dimensions in the sampled materials, but cell numbers were correlated with cambial age. The chronologies contain strong temperature signals associated with the timing of xylem growth. We obtained reliable reconstructions of mean June temperature from the total cell number and July–September temperature from the cell wall thickness of latewood. June temperature and July–September temperature covaried for most of the period from AD 1642 to AD 1978. After that time, June temperature became cooler relative to July–September temperature. This difference caused disproportional changes in earlywood tracheids because of the late start of growth and cool conditions in June followed by warming during the rest of the season. The identification of this unusual recent change has shown that intraseasonal resolution may be achieved by cell dimension and cell number chronologies.

Variation in xylem anatomy of selected populations of lodgepole pine

2001 ◽  
Vol 31 (11) ◽  
pp. 2049-2057 ◽  
Author(s):  
Tongli Wang ◽  
Sally N Aitken
Keyword(s):  
Cell Wall ◽  
Wall Thickness ◽  
Slow Growth ◽  
Lodgepole Pine ◽  
High Density ◽  
Cell Wall Thickness ◽  
Fast Growth ◽  
Low Density ◽  
Xylem Anatomy ◽  
Latewood Density

Variation in xylem anatomy among selected populations of lodgepole pine (Pinus contorta Dougl. ex Loud. var. latifolia Engelm.) was examined using digital image analysis based on an annual growth ring (age 10) per tree. Four subpopulations were selected using the following criteria for height growth and wood density: (i) fast growth and high density; (ii) slow growth and high density; (iii) fast growth and low density; and (iv) slow growth and low density. Significant differences were found among subpopulations for several anatomical parameters including tracheid density, lumen size, and cell wall thickness that may affect the economic value and utilization of wood. Principal component analysis indicate that the first four principal components (PCs) were associated with (i) ring area (PC1), (ii) earlywood density (PC2), (iii) latewood density (PC3), and (iv) lumen shape in earlywood (PC4), suggesting that these aspects of wood properties and growth are controlled by different sets of genes. Relative contributions of total number of tracheids, tracheid lumen size, and cell wall thickness to ring area and correlations between cell wall area proportion and X-ray density are discussed.

scholarly journals Host Barriers and Responses to Uncinula necator in Developing Grape Berries

2004 ◽  
Vol 94 (5) ◽  
pp. 438-445 ◽  
Author(s):  
Andrea Ficke ◽  
David M. Gadoury ◽  
Robert C. Seem ◽  
Dale Godfrey ◽  
Ian B. Dry
Keyword(s):  
Cell Wall ◽  
Powdery Mildew ◽  
Wall Thickness ◽  
Infection Process ◽  
Cell Wall Thickness ◽  
Antifungal Compound ◽  
Uncinula Necator ◽  
Grape Berries ◽  
Pathogenesis Related ◽  
Cuticle Thickness

Grape berries are highly susceptible to powdery mildew 1 week after bloom but acquire ontogenic resistance 2 to 3 weeks later. We recently demonstrated that germinating conidia of the grape powdery mildew pathogen (Uncinula necator) cease development before penetration of the cuticle on older resistant berries. The mechanism that halts U. necator at that particular stage was not known. Several previous studies investigated potential host barriers or cell responses to powdery mildew in berries and leaves, but none included observation of the direct effect of these factors on pathogen development. We found that cuticle thickness increased with berry age, but that ingress by the pathogen halted before formation of a visible penetration pore. Cell wall thickness remained unchanged over the first 4 weeks after bloom, the time during which berries progressed from highly susceptible to nearly immune. Autofluorescent polyphenolic compounds accumulated at a higher frequency beneath appressoria on highly susceptible berries than on highly resistant berries; and oxidation of the above phenolics, indicated by cell discoloration, developed at a significantly higher frequency on susceptible berries. Beneath the first-formed appressoria of all germinated conidia, papillae occurred at a significantly higher frequency on 2- to 5-day-old berries than on 30- to 31-day-old fruit. The relatively few papillae observed on older berries were, in most cases (82.8 to 97.3%), found beneath appressoria of conidia that had failed to produce secondary hyphae. This contrasted with the more abundantly produced papillae on younger berries, where only 35.4 to 41.0% were located beneath appressoria of conidia that had failed to produce secondary hyphae. A pathogenesis-related gene (VvPR-1) was much more highly induced in susceptible berries than in resistant berries after inoculation with U. necator. In contrast, a germin-like protein (VvGLP3) was expressed within 16 h of inoculation in resistant, but not in susceptible berries. Our results suggest that several putative barriers to infection, i.e., cuticle and cell wall thickness, antimicrobial phenolics, and two previously described pathogenesis-related proteins, are not principal causes in halting pathogen ingress on ontogenically resistant berries, but rather that infection is halted by one or more of the following: (i) a preformed physical or biochemical barrier near the cuticle surface, or (ii) the rapid synthesis of an antifungal compound in older berries during the first few hours of the infection process.

scholarly journals Differences in cell wall of thin and thick filaments of cyanobacterium Aphanizomenon gracile SAG 31.79 and their implications for different resistance to Daphnia grazing

2016 ◽  
Author(s):  
Lukasz Wejnerowski ◽  
Slawek Cerbin ◽  
Maria K. Wojciechowicz ◽  
Marcin K. Dziuba
Keyword(s):  
Cell Wall ◽  
Wall Thickness ◽  
Cell Walls ◽  
Cell Wall Thickness ◽  
Filamentous Cyanobacterium ◽  
Thin Sections ◽  
Thick Filaments ◽  
Transmission Electron ◽  
The Difference ◽  
Aphanizomenon Gracile

<p>Recent studies have shown that the filamentous cyanobacterium <em>Aphanizomenon gracile</em> Lemmermann, strain SAG 31.79, consists of two types of filaments that differ in thickness. These two types are known to vary in resistance to <em>Daphnia</em> <em>magna</em> grazing: thin filaments (&lt;2.5 µm) are more vulnerable to grazing than the thick ones (&gt;2.5 µm). In this study, we investigated whether the difference in the vulnerability to grazing of thin and thick filaments is a result of different thickness of their cell walls, a filament stiffness determinant. We expected thick filaments to have thicker cell walls than the thin ones. Additionally, we analysed whether cell wall thickness correlates with filament thickness regardless of the filament type. A morphometric analysis of cell walls was performed using transmission electron micrographs of ultra-thin sections of the batch-cultured cyanobacterial material.  Our study revealed that the thin type of filaments had thinner cell walls than the thick filaments. Moreover, cell wall thickness was positively correlated with filament thickness. TEM (transmission electron microscopy) observations also revealed that the thin type of filaments was often at different stages of autocatalytic cell destruction, which was mainly manifested in the increase in cell vacuolization and degradation of the cytoplasm content. Based on our findings, we assume that previously reported higher resistance of thick filaments to <em>Daphnia</em> grazing results from greater stiffness and excellent physiological conditions of thick filaments. </p>

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