Cellular Chemical Composition Research Articles

Failure of Cronobacter sakazakii to acquire direct and cross-protection against lethal stresses after exposure to sublethal ethanol

Abstract Cronobacter sakazakii may encounter subinhibitory concentrations of ethanol stress over its lifecycle. Bacterial tolerance to homologous or heterologous stress agents may be altered as a result of ethanol adaptive responses. Therefore, the tolerance of ethanol-exposed and control cells to subsequent lethal stresses was evaluated in the current work. It was discovered that sublethal ethanol exposure increased the susceptibility to lethal ethanol stress in C. sakazakii as determined by the Weibull model. Furthermore, sublethal ethanol concentration exposure in C. sakazakii did not lead to any cross-tolerance against other stressors such as benzalkonium chloride (120 mg/L), heat (55 °C), cold (4 °C), simulated gastric fluid (pH 3.0), osmotic stress (sorbitol, 0.75 g/mL), and desiccation stress. Analysis of zeta potential, scanning electron microscope, and surface-enhanced Raman spectroscopy spectra revealed that cellular injury and changes in cellular chemical composition may contribute to the reduced resistance of C. sakazakii after ethanol exposure. Furthermore, sublethal ethanol exposure resulted in an elevated proportion of unsaturated fatty acids (USFA), while reducing the proportion of saturated fatty acids (SFA) and the ratio of SFA to USFA. The developed inactivation models can serve as a valuable source of data to support quantitative microbial risk assessment. Moreover, a better understanding of the response of C. sakazakii to sublethal ethanol exposure may provide valuable insights into the prevention and control of C. sakazakii.

The Recent Development of Acoustic Sensors as Effective Chemical Detecting Tools for Biological Cells and Their Bioactivities.

One of the most significant developed technologies is the use of acoustic waves to determine the chemical structures of biological tissues and their bioactivities. In addition, the use of new acoustic techniques for in vivo visualizing and imaging of animal and plant cellular chemical compositions could significantly help pave the way toward advanced analytical technologies. For instance, acoustic wave sensors (AWSs) based on quartz crystal microbalance (QCM) were used to identify the aromas of fermenting tea such as linalool, geraniol, and trans-2-hexenal. Therefore, this review focuses on the use of advanced acoustic technologies for tracking the composition changes in plant and animal tissues. In addition, a few key configurations of the AWS sensors and their different wave pattern applications in biomedical and microfluidic media progress are discussed.

FTIR study on the cellular chemical composition of Allium cepa L. seedlings under salt stress

The current study is to investigate the effects of salinity on cellular chemical composition in four cultivars of onion seedlings (Agrifound rose, Bellary, Prema-178 and Nasik red) at different time intervals through the Fourier-transform infrared (FTIR) spectroscopy. Onion seedlings were treated with 0, 50 and 150 mM NaCl for 3 h and 48 h intervals and dried samples were used for analysing the biochemical changes using FTIR spectroscopy. The results provided insights at 50 and 150 mM NaCl and clear differences were noticed in profiles related to lipids, proteins and carbohydrate functional groups compared with untreated control. In principal component analysis (PCA), the first two principal components (PC1 and PC2) of four onion cultivars and salt treatments explained 95.7 % of total variations whereas 71.8 % of total variations were accounted with FTIR spectra of 4000-1000 cm-1 region. Multiple linear regression (MLR) analysis exhibited significant correlation values of R2 > 0.9. At both the intervals of salt treatment, the Bellary cultivar showed good responses, thereafter Prema-178 cultivar at each concentration when compared with control. In conclusion, FTIR spectroscopy can elucidate early salt stress-induced chemical changes in various compounds as well as the salt adaptive process in onion metabolism.

Parametric mapping of cellular morphology in plant tissue sections by gray level granulometry

BackgroundThe cellular morphology of plant organs is strongly related to other physical properties such as shape, size, growth, mechanical properties or chemical composition. Cell morphology often vary depending on the type of tissue, or on the distance to a specific tissue. A common challenge in quantitative plant histology is to quantify not only the cellular morphology, but also its variations within the image or the organ. Image texture analysis is a fundamental tool in many areas of image analysis, that was proven efficient for plant histology, but at the scale of the whole image.ResultsThis work presents a method that generates a parametric mapping of cellular morphology within images of plant tissues. It is based on gray level granulometry from mathematical morphology for extracting image texture features, and on Centroidal Voronoi Diagram for generating a partition of the image. Resulting granulometric curves can be interpreted either through multivariate data analysis or by using summary features corresponding to the local average cell size. The resulting parametric maps describe the variations of cellular morphology within the organ.ConclusionsWe propose a methodology for the quantification of cellular morphology and of its variations within images of tissue sections. The results should help understanding how the cellular morphology is related to genotypic and / or environmental variations, and clarify the relationships between cellular morphology and chemical composition of cell walls.

How growth conditions of Euglena gracilis cells influence cellular composition as evidenced by Fourier transform infrared spectroscopy and direct infusion high-resolution mass spectrometry

This study aims to assess the influence of growth and culture conditions of the Euglena gracilis protist on the cellular chemical composition. Fourier transform infrared spectroscopy (FT-IR) and direct infusion Orbitrap electrospray ionization mass spectrometry (ESI-MS) were used in tandem for confirmation of functional groups, and to decipher the normalized percentage abundances of compound classes, molecular species, and aromaticity and oxygenation within each growth condition. Abundances of molecular species and compound classes varied with growth and culture conditions. No major differences in the qualitative FT-IR-based screening of functional groups were found between growth conditions. This contrasts with ESI-MS-based results where polyphenols, carbohydrate, and condensed aromatic compounds were the most abundant in dark-grown conditions with glucose supplementation, revealing the differences in physiological states of the cell in different growth conditions. Dark-grown conditions promoted CHO molecular species while light-grown conditions favored CHON, CHOS, and CHONS molecular species. The dark-grown exponential E. gracilis cells grown in glucose-supplemented media showed the greatest normalized abundances of aromatics, carbohydrate, polyphenols, and protein, which are compound classes typically involved in metal binding.

An endophytic Kocuria palustris strain harboring multiple arsenate reductase genes

Aiming at revealing the arsenic (As) resistance of the endophytic Kocuriastrains isolated from roots and stems of Sphaeralcea angustifolia grown at mine tailing, four strains belonging to different clades of Kocuria based upon the phylogeny of 16S rRNA genes were screened for minimum inhibitory concentration (MIC). Only the strain NE1RL3 was defined as an As-resistant bacterium with MICs of 14.4/0.0125mM and 300/20.0mM for As3+ and As5+, respectively, in LB/mineral media. This strain was identified as K. palustris based upon analyses of cellular chemical compositions (cellular fatty acids, isoprenoides, quinones, and sugars), patterns of carbon source, average nucleotide identity of genome and digital DNA-DNA relatedness. Six genes coding to enzymes or proteins for arsenate reduction and arsenite-bumping were detected in the genome, demonstrating that this strain is resistant to As possibly by reducing As5+ to As3+, and then bumping As3+ out of the cell. However, this estimation was not confirmed since no arsenate reduction was detected in a subsequent assay. This study reported for the first time the presence of phylogenetically distinct arsenate reductase genesin a Kocuria strain and evidenced the possible horizontal transfer of these genes among the endophytic bacteria.

Label-free Mass Cytometry for Unveiling Cellular Metabolic Heterogeneity.

Comprehensive analysis of single-cell metabolites is critical since differences in cellular chemical compositions give rise to specialized biological functions. Herein, we propose a label-free mass cytometry by coupling flow cytometry to ESI-MS (named CyESI-MS) for high-coverage and high-throughput detection of cellular metabolites. Cells in suspension were isolated, online extracted by sheath fluid, and lysed during gas-assisted electrospray, followed by real-time MS analysis. Hundreds of metabolites, including nucleotides, amino acids, peptides, carbohydrates, fatty acyls, glycerolipids, glycerophospholipids, and sphingolipids, were detected and identified from one single cell. Discrimination of four types of cancer cell lines and even three subtypes of breast cancer cells was readily achieved using their distinct metabolic profiles. Furthermore, we screened out 102 characteristic ions from 615 detected peak signals for distinguishing breast cancer cell subtypes and identified 40 characteristic molecules which exhibited significant differences among these subtypes and would be potential metabolic markers for clinical diagnosis. CyESI-MS is expected to be a new-generation mass cytometry for studying cell heterogeneity on the metabolic level.

Gold nanoaggregates for probing single-living cell based on surface-enhanced Raman spectroscopy.

Gold nanoparticles are delivered into living cells by transient electroporation method to obtain intracellular surface-enhanced Raman spectroscopy (SERS). The subcellular localization of gold nanoparticles is characterized by transmission electron microscopy, and the forming large gold nanoaggregates are mostly found in the cytoplasm. The SERS detection of cells indicates that this kind of gold nanostructures induces a high signal enhancement of cellular chemical compositions, in addition to less cellular toxicity than that of silver nanoparticles. These results demonstrate that rapid incorporation of gold nanoparticles by electroporation into cells has great potential applications in the studies of cell biology and biomedicine.

Ionizing Radiation Induced Biological Effect on Human Cell HCT116(P53+/+, P53-/-) Observed through Synchrotron-FTIR Microspectroscopy and Imaging

Ionizing radiations exist ubiquitously and can induce cell apoptosis and necrosis. It is important to investigate the involved physico-chemical processes and mechanisms. Many new tools and methods are developed, among which synchrotron-FTIR microspectroscopy also gains concern for its advantage of non-invasive measurement of cellular chemical compositions with spatial resolution. Recently, we employed s-FTIR microspectroscopy to study the biological effect on Rhizopus oryzae induced by energetic-particle-irradiation, and observed the ROS-involved DNA damages, lipid peroxidation and protein denaturation with spatial resolution [Liu et al. J Mol Struct, 2013, 1031:1-8.]. In the present work, we continued to apply s-FTIR microspectroscopy to investigate the ionizing radiation induced biological effect on the human colon cancer cells HCT116 (p53 + / +, p53-/ -). The p53 gene is regarded as an anti-oncogene [Hollstein et al. Science, 1991, 253:49-53]. It was also reported that X-ray radiation could induce the cycle arrest of cells [Michael et al. Cancer Res2003, 63:7176-7184]. In our study, we were able to make use of s-FTIR microspectroscopy to observe and distinguish the cells in G1, S and G2 phases. We employed different forms of ionizing radiations including particle radiation, gamma and X-ray radiations to irradiation the cells. Our preliminary results showed that the radiations indeed induced the DNA damages and protein denaturation but with different extents of effect for different radiations and different (p53 + / +, p53-/-) cells. The FTIR measurements also definitely revealed radiation-induced cell cycle arrest and the p53 dependent radiation sensitivity. This research demonstrated that s-FTIR microspectroscopy is a useful tool for observing and understanding the biological effect on cells under ionizing radiations.

Advances in methods for detection of anaerobic ammonium oxidizing (anammox) bacteria

Anaerobic ammonium oxidation (anammox), the biochemical process oxidizing ammonium into dinitrogen gas using nitrite as an electron acceptor, has only been recognized for its significant role in the global nitrogen cycle not long ago, and its ubiquitous distribution in a wide range of environments has changed our knowledge about the contributors to the global nitrogen cycle. Currently, several groups of methods are used in detection of anammox bacteria based on their physiological and biochemical characteristics, cellular chemical composition, and both 16S rRNA gene and selective functional genes as biomarkers, including hydrazine oxidoreductase and nitrite reductase encoding genes hzo and nirS, respectively. Results from these methods coupling with advances in quantitative PCR, reverse transcription of mRNA genes and stable isotope labeling have improved our understanding on the distribution, diversity, and activity of anammox bacteria in different environments both natural and engineered ones. In this review, we summarize these methods used in detection of anammox bacteria from various environments, highlight the strengths and weakness of these methods, and also discuss the new development potentials on the existing and new techniques in the future.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-011-3230-6) contains supplementary material, which is available to authorized users.

Response of Paenibacillus polymyxa to iron: alternations in cellular chemical composition and the production of fusaricidin type antimicrobial compounds

In this work, growth, cellular chemical composition and production of fusaricidin type antimicrobial compounds by P. polymyxa SQR-21 were compared in tryptone broth supplemented with four concentrations of iron (25, 50, 100 and 200 µM). The data revealed that the growth of P. polymyxa SQR-21 was increased by 3-8% with the increase in concentration of ferric ion (Fe3+). The production of fusaricidin type compounds was increased by 33-49% only up to 50 µM Fe3+ and the highest level of Fe3+ was inhibitory. Increase in the liquid culture Fe3+concentration increased the intracellular protein (2%), intracellular carbohydrate (14%), extracellular protein (7%) and polysaccharide contents (18%) while the intracellular lipid contents were increased (11%) only up to 50 µM Fe3+. In addition, the regulatory effects of Fe3+ were also reflected by the increase in total RNA contents and relative expression of the fusaricidin synthetase gene (FusA) by 3-13 and 35-56%, respectively, up to 50 µM Fe3+, after that a continuous decrease was observed.

Effect of chilling on porcine germinal vesicle stage oocytes at the subcellular level

The potential subcellular consequence of chilling on porcine germinal vesicle (GV) stage oocytes was examined. Prior to in vitro maturation (IVM), Cumulus-oocyte complexes (COCs) freshly collected from antral follicles (3–6 mm in diameter) were evenly divided into four groups and immediately incubated in PVA-TL-HEPES medium at the temperature of 39 °C (control group), 23 °C (room temperature), 15 °C and 10 °C for 10 min, respectively. Following 42 h of IVM at 39 °C, the survival rates were examined. There was no significant difference between the survival rate of 23 °C chilled group and control group (77.92 and 91.89%), but the survival rate of 15 and 10 °C chilled group were significantly decreased (46.34 and 4.81%, P < 0.01). A further experiment on15 °C group showed that most oocytes died from 2 to 4 h of IVM. In order to investigate the effects of chilling on oocytes at the subcellular level, the control and 15 °C chilled group COCs fixed at different time points of the IVM cultures (2, 2.5, 3, 3.5 and 4 h of IVM) were prepared for transmission electron microscope (TEM) observation. As the result, compared with the control group, there were two significant changes in the ultrastructural morphology of 15 °C treatment group: (1) dramatic reduction of heterogeneous lipid, (2) disorganized mitochondria–endoplasmic reticulum–lipid vesicles (M–E–L) combination. These results indicate that 15 °C is a critical chilling temperature for porcine GV stage oocyte and the alteration of cellular chemical composition and the destruction of M–E–L combination maybe responsible for chilling injury of porcine oocyte at this stage.

Isolation and Characterization of a Xanthophyll Aberrant Mutant of the Green Alga Nannochloropsis oculata

Novel mutants (xan1 and xan2) of the unicellular green alga Nannochloropsis oculata are impaired in xanthophyll biosynthesis, thereby producing aberrant levels of xanthophylls. High-performance liquid chromatography (HPLC) analysis revealed that the xan1 and xan2 mutants have double the violaxanthin (V) content, but have significantly decreased lutein content in their cells compared to the wild type. Furthermore, these mutants contain two to three times more zeaxanthin than the wild type under low light (LL) growth conditions. However, this xanthophyll aberration in N. oculata did not affect the normal growth and the major cellular chemical composition of the xan1 strain. The xanthophyll pool size of the LL-grown mutant was 1.8-fold greater than that of the wild type. Under high light (HL) growth conditions, V content was substantially decreased in both the mutant and wild types because of the epoxidation state of the xanthophylls. Under LL growth conditions, the deepoxidation states of the xanthophyll pool sizes were 0.1 and 1.2 in the wild type and the mutant, respectively. However, the deepoxidation states of the xanthophyll pool sizes were 0.78 in the wild type and 0.87 in the mutant under HL growth conditions. We observed that the level of one of the commercially important xanthophylls, zeaxanthin, was higher in the mutant than in the wild type under all culture conditions. This mutant is discussed in terms of its commercial value and potential utilization by the algal biotechnology industry for the production of zeaxanthin.

Tracing the Impact of Adverse Environmental Conditions on the Frost Sensitivity in Trees

The winter hardening of trees is genetically determined; the cellular chemical composition and structure are modified in reaction to cues from the length of daylight and ambient temperature. However, adverse environmental conditions can affect the timing of hardening and dehardening, concentration of cryoprotective compounds and cell membrane stability. The tree nutrient status, air pollutants, infestation of pest and pathogens and extreme weather events can thus increase the probability that the tree will be subjected to freezing temperatures when the hardiness level is not adequate. In this paper, the sensitivity to frost is related to the complex chain of cause and effect among environmental stress factors and to the overall tree vitality. This is discussed in the context of sustainable forestry in a changing climate.

CHANGES IN DOMOIC ACID PRODUCTION AND CELLULAR CHEMICAL COMPOSITION OF THE TOXIGENIC DIATOM PSEUDO‐NITZSCHIA MULTISERIES UNDER PHOSPHATE LIMITATION1

ABSTRACTProduction of domoic acid (DA), a neurotoxin, by the diatom Pseudo‐nitzschia multiseries (previously Nitzschia pungens f. multiseries) Hasle and its cellular chemical composition were studied in phosphate‐limited chemostat continuous cultures and in subsequent batch cultures. Under steady‐state chemostat conditions, DA production increased from 0.01 to 0.26 pg DA · cell−1· d−1 as the growth rate decreased. When the nutrient supply was discontinued (to produce a batch culture), DA production was enhanced by a factor of ca. 3. DA production was temporarily suspended upon addition of phosphate to the batch cultures but resumed 1 d later at a higher rate coincident with the decline of phosphate uptake. In both steady‐state continuous culture and batch culture, more DA was produced when alkaline phosphatase activity (APA) was high. The association of high DA production with high levels of APA and high cellular N:P ratios strongly suggests that phosphate limitation enhances DA production. Also, DA production was high when other primary metabolism (e.g. uptake of carbon, nitrogen, phosphorus and silicon, and cell division) was low, but chlorophyll a and adenosine triphosphate were generally high. This suggests that the synthesis of DA requires a substantial amount of biogenic energy.

Biochemical quality of marine unicellular algae with special emphasis on lipid composition. II. Nannochloropsis sp.

The eustigmatophyte, Nannochloropsis sp., contains relatively large amounts of valuable lipids and is commonly grown as food source for commercial application in aquaculture. In this study we evaluated the effect of environmental conditions such as light intensity, nitrogen availability and temperature on cellular chemical composition with special emphasis on fatty acid distribution in cultures of Nannochloropsis sp. continuously grown under laboratory-controlled conditions. Seasonal variations in cellular chemistry were further assessed using large-scale outdoor shallow ponds, aimed at mass production of Nannochloropsis for application in an aquaculture hatchery. The percentage of the polyunsaturated long-chain fatty acid, eicosapentaenoic acid, in the algal biomass varied between 1.6 and 3.8% (w/w). In spite of significant variations in cellular protein content, the amino acid composition of Nannochloropsis biomass remained fairly constant during the different production seasons. Variations in the fatty acid composition of the algal biomass are attributed to differences in seasonal climatic conditions and diurnal cycles in light intensity and temperature. This study demonstrates the feasibility of culturing algal biomass of a high chemical quality for various applications in aquaculture hatcheries.

Biochemical quality of marine unicellular algae with special emphasis on lipid composition. I. Isochrysis galbana

The prymnesiophyte Isochrysis aff. galbana (T-ISO) contains relatively large amounts of valuable lipids and is commonly grown as a food source for commercial production in aquaculture. The effect of environmental conditions, such as light intensity and nitrogen availability, on cellular chemical composition with special emphasis on lipid content has been studied in continuously grown cultures of I. galbana. High light intensity caused accumulation of carbohydrates, while cellular lipid content slightly decreased. Nitrogen limitation, however, caused an increase of both carbohydrate and lipid content. Variations in cellular lipid content were associated with changes in fatty acid composition. High light conditions or severe nitrogen limitations were associated with an increase in the relative abundance of palmitic (C16:0) and oleic (C18:1 ) acids and a decrease in the percentage of the polyunsaturated fatty acid C18:4. The relative distribution of the highly unsaturated fatty acid docosahexaenoic acid (C22:6) slightly increased under high light conditions and decreased under low nitrogen levels. Thus the polyunsaturated to saturated fatty acid ratio decreased when I. galbana was grown under high irradiance levels or under nitrogen-limiting conditions.

Effect of chemical fixatives on accurate preservation of Escherichia coli and Bacillus subtilis structure in cells prepared by freeze-substitution

Five chemical fixatives were evaluated for their ability to accurately preserve bacterial ultrastructure during freeze-substitution of select Escherichia coli and Bacillus subtilis strains. Radioisotopes were specifically incorporated into the peptidoglycan, lipopolysaccharide, and nucleic acids of E. coli SFK11 and W7 and into the peptidoglycan and RNA of B. subtilis 168 and W23. The ease of extraction of radiolabels, as assessed by liquid scintillation counting during all stages of processing for freeze-substitution, was used as an indicator of cell structural integrity and retention of cellular chemical composition. Subsequent visual examination by electron microscopy was used to confirm ultrastructural conformation. The fixatives used were: 2% (wt/vol) osmium tetroxide and 2% (wt/vol) uranyl acetate; 2% (vol/vol) glutaraldehyde and 2% (wt/vol) uranyl acetate; 2% (vol/vol) acrolein and 2% (wt/vol) uranyl acetate; 2% (wt/vol) gallic acid; and 2% (wt/vol) uranyl acetate. All fixatives were prepared in a substitution solvent of anhydrous acetone. Extraction of cellular constituents depended on the chemical fixative used. A combination of 2% osmium tetroxide-2% uranyl acetate or 2% gallic acid alone resulted in optimum fixation as ascertained by least extraction of radiolabels. In both gram-positive and gram-negative organisms, high levels of radiolabel were detected in the processing fluids in which 2% acrolein-2% uranyl acetate, 2% glutaraldehyde-2% uranyl acetate, or 2% uranyl acetate alone were used as fixatives. Ultrastructural variations were observed in cells freeze-substituted in the presence of different chemical fixatives. We recommend the use of osmium tetroxide and uranyl acetate in acetone for routine freeze-substitution of eubacteria, while gallic acid is recommended for use when microanalytical processing necessitates the omission of osmium.

Effects of 2,4‐dichlorophenoxyacetic acid on Rhizobium sp. in pure culture

AbstractThe effects of 2,4‐Dichlorophenoxyacetic acid on the growth rate, chemical composition, 14C‐2,4‐dichlorophenoxyacetic acid and 45Ca2+ uptake by Rhizobium sp. M 4 able to nodulate Arachis hypogaea were determined. Cellular growth was diminished by the presence of 10−3 M 2,4‐dichlorophenoxyacetic acid in the medium. Alterations in cellular chemical composition, in 14C‐2,4‐dichlorophenoxyacetic acid and in 45Ca2+ uptake were found.

Growth rate and some physiological features of Deleya halophila CCM 3662 at different salt concentrations

The growth rate of the type strain of the new, moderately halophilic species Deleya halophila ( Deleya halophila CCM 3662) at different salt concentrations was determined and its optimal marine salt concentration for growth was established. Oxygen uptake and some cellular chemical composition characteristics of cells grown at their optimal salt concentration (7.5 % w/v) and at a higher concentration (15 % w/v) were compared, showing that both protein and carbohydrate intracellular contents decreased when the external medium salt concentration was increased, with the carbohydrate content reaching 50 % of the value presented by cells cultured at optimal salinity conditions. Oxygen uptake and poly-β-hydroxybutyric acid content were not significantly affected by changes in external salinity.