Nucleic Acid Content Of Cells Research Articles

Coexistence of dominant groups in marine bacterioplankton community—a combination of experimental and modelling approaches

The population dynamics in the coastal bacterioplankton community off Plymouth (UK) was studied in samples proportionally diluted (99%, 90%, 66% and 0%) with sterile seawater, incubated in the dark for 2–4 days and monitored by flow cytometry. Nucleic acid content of cells, stained with SYBR Green I DNA specific dye, was used as an index of a genome size. Using flow sorting and fluorescence in situ hybridization (FISH) with a set of ribosomal RNA targeted oligonucleotide probes, the phylogenetic composition of dominant cytometric groups of bacterioplankton was determined to be similar during growth in the dilution series. The proportion of the low nucleic acid (LNA) group decreased and correspondingly the high nucleic acid (HNA) groups increased with dilution. The assimilation rates of free amino acids, a highly labile nutrient pool, were determined by flow sorting the dominant groups after short incubations with 35S-methionine tracer. The relative cellular amino acid assimilation by the LNA cells increased with dilution, while the activity of the HNA cells either decreased or remained unchanged. However, highly metabolically active LNA bacteria were overgrown by the HNA bacteria, presumably because the small genome size—an adaptation to living in an oligotrophic environment—did not allow the LNA group to grow sufficiently fast to compete with the HNA group under experimentally reduced grazing pressure. To examine the experimental results a numerical model of bacterioplankton population dynamics was formulated based on the hypothesis that the LNA cells consume only a labile fraction of organic nutrients (amino acids etc.), while the HNA cells feed on both the labile and more refractory sources of nutrients, and that in the absence of phytoplankton the labile source of nutrients is produced entirely by the bacterivorous flagellates. The model simulations gave credence to the hypothesized primary dependence of the LNA group on labile organic nutrients recycled within the microbial loop.

Variations of bacterial-specific activity with cell size and nucleic acid content assessed by flow cytometry

AME Aquatic Microbial Ecology Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials AME 28:131-140 (2002) - doi:10.3354/ame028131 Variations of bacterial-specific activity with cell size and nucleic acid content assessed by flow cytometry Philippe Lebaron*, Pierre Servais**, Anne-Claire Baudoux, Muriel Bourrain, Claude Courties, Nathalie Parthuisot Observatoire Océanologique, Université Pierre et Marie Curie, UMR 7621-7628 CNRS-INSU, BP 44, 66651 Banyuls-sur-Mer cedex, France *E-mail: lebaron@obs-banyuls.fr **Present address: Ecologie des Systèmes Aquatiques, Université Libre de Bruxelles, Campus Plaine, CP 221, 1050 Bruxelles, Belgium ABSTRACT: In most aquatic bacterial communities, it is possible to discriminate bacterial cells with a high nucleic acid content (HNA) from those with a low nucleic acid content (LNA) by flow cytometry. The distribution of leucine incorporation rate per cell (specific activity) within the fraction of bacteria with a high apparent nucleic acid content was investigated in coastal seawater using flow cytometry and cell sorting techniques. Bacterial cells from a natural seawater sample were first labeled with tritiated leucine, stained with a fluorescent nucleic acid stain and then sorted based on their fluorescence and scatter signals, assuming that fluorescence was proportional to the cellular nucleic acid content and scatter signals to biovolume. Our results clearly demonstrated that specific activity was heterogeneously distributed within the group of HNA cells and increased with both scatter and fluorescence signals. This shows that important differences in terms of cell-specific rates of activity exist within the HNA group and that specific activity is positively correlated with both the nucleic acid content of cells and their biovolume. We hypothesized that this heterogeneity may be partly due to the structure of bacterial communities. Therefore, further investigations were made by analyzing the nucleic acid content and scatter properties of a set of 10 species isolated from the same coastal area. Our results confirm that both fluorescence and scatter values vary greatly between species at a given growth stage and between growth stages for a given species. Variations reported at different growth stages suggest that both parameters are sensitive to the metabolic activity of individual cells and confirm the positive relationship reported in this study between scatter and fluorescence parameters and specific activity within the HNA cellular fraction of natural communities. Interestingly, it was shown that HNA and LNA cells were present in late stationary phase cultures. This suggests that LNA cells found in natural communities may be dead or dying cells. KEY WORDS: Nucleic acid content · Scatter · Cellular activity · Flow cytometry · Cell sorting Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in AME Vol. 28, No. 2. Online publication date: June 26, 2002 Print ISSN: 0948-3055; Online ISSN: 1616-1564 Copyright © 2002 Inter-Research.

Advances and New Concepts in Oral and Maxillofacial Pathology

Improvement in the treatment of patients has come through new surgical techniques, improved modalities of therapy, and better understanding of disease processes. The role of the pathologist is to elucidate and clarify the pathogenesis of disease. For many years, advances in the understanding of neoplastic disease have been achieved by pathologists through the recognition and classification of tumors via standard light microscopy using routine hematoxylin and eosin staining, occasionally complemented by specific histochemical stains. The advent of electron microscopy yielded further morphologic information but was limited in application by its expense, the size of the operating unit, and the requirement of fresh tissue. Over the past 2 decades, further characterization of tumors became feasible using newly developed diagnostic techniques. In addition, the ease of obtaining tissue samples has been enhanced by new approaches such as fine-needle aspiration biopsy. Analysis of the molecular characteristics of certain tumors has become the standard of care, contributing to more specific classification and treatment recommendations. Recent molecular biologic advances have made such analyses not only possible but relatively convenient. Labeled probes have been developed that help to analyze nucleic acid sequences. Techniques used to identify and quantify the nucleic acid content of cells include filter hybridization (dot blot, Southern blot, Northern blot, etc.), DNA or RNA in-situ hybridization, quantification of argyrophilic nucleolar organizer regions (AgNOR), and flow cytometric analysis. Cellular proteins other than nucleic acids can be recognized by immunohistochemistry and Western blot techniques. The use of polymerase chain reaction (PCR) has added to the efficacy of many of these techniques by providing a means of nucleic-acid amplification during testing. This technique allows the amplification of small quantities of nucleic acid (body fluid, peripheral white cells, aspirates, cytology, etc.). It is critical to distinguish among various types of tumors, because treatment is often determined by tumor type. For example, it is no longer acceptable to rely on clinical and microscopic evaluation alone to determine if a malignant round-cell tumor represents a leukemic infiltrate, lymphoma, neuroblastoma, rhabdomyosarcoma, or poorly differentiated carcinoma. Instead, immunohistochemical markers are used to help distinguish whether the tissue of origin is epithelial, neurogenic, myogenic, lymphoid, or metastatic. DNA and RNA in-situ hybridization are available to identify intracellular pathogens. AgNOR and flow cytometric analysis are used to evaluate cell kinetics in an attempt to predict biologic behavior of tumors and patient response to treatment. The focus of this discussion is not meant to be inclusive of all of the most recent diagnostic techniques currently available, but it presents an overview of the most commonly applied new approaches to diagnosis.

The effect of CO, KCN and NaN 3 on the nucleic acid content of Ehrlich ascites tumor cells

Ehrlich ascites cells were treated in vivo with a 2% aqueous solution of carbon monoxide or KCN or NaN 3. On the ninth day after transplantation the quantity of ascitic fluid, the number and average size of cells and the total packed cell volume (TPCV) were determined. Nucleic acid fractions of the cells were isolated and measured on the basis of their phosphorous content. Smears of the cells were stained with methyl-green pyronin and by the Feulgen method. It was found that after treatment with carbon monoxide the nucleic acid content of the cells increased similarly to that already shown for yeast cells. After treatment with KCN or NaN 3 the nucleic acid content of the cells decreased.

The nucleic acid content of cells in the meristematic elongating and fully elongated segments of roots of Vicia faba

By combined cell counting and biochemical assay, the mean DNA and RNA content of the cells in the root tip of Vicia faba seedlings has been measured. The values for segments 0–2, 2–4, 4–8 and 12–20 mm behind the root cap were found to be, for DNAP, 3.5 7 ± 0.2 0, 4.00 ± 0.20, 4.5 5 ± 0.3 7 and 4.7 0 ± 0.3 2, 10 −12 grams, and for RNAP, 14.2 ± 0.54, 16.1 ± 0.63, 18.0 ± 0.99, and 16.5 ± 0.86, 10 −12 grams. The approximate amounts of phosphorus per cell in other forms were also estimated. The nucleic acid determinations show that the mean DNA content per cell steadily increases with distance from the root apex, being 30 per cent greater in the proximal segment, which is composed mainly of fully differentiated cells, than in the terminal segment, which is entirely meristematic. Thus, if the average meristematic cell at the commencement of interphase is assumed to have a diploid content of DNA, then the mean DNA content of the differentiated cells is approximately tetraploid.

Effects of cortisone on regenerating rat liver.

The effects of continuous administration of cortisone on the metabolism of regenerating rat liver have been studied. Whereas the restoration of the weight of the liver after partial hepatectomy was not markedly affected by cortisone, the multiplication of cells was reduced to a significant degree after the first 2 days of regeneration. Liver restoration in terms of nucleic acids was similarly inhibited by cortisone. The results are consistent with the interpretation that the inhibition of cell multiplication in this system is dependent on and keeps pace with the inhibition of nucleic acid synthesis by this drug. At almost any time after hepatectomy, the nucleic acid content of the liver cells was the same in treated and in untreated animals. In ancillary studies, it was shown that cortisone caused the cells of regenerating liver to be increased in size and weight through the increased infiltration of lipids. Changes in water, protein, and carbohydrate content of the liver cells did not contribute to this increase in the weight of the cells. Since all animals were treated with cortisone for 5 days before hepatectomy, data were also obtained on the effect of this agent on the resting liver. This course of treatment brought about a significant decrease in the number of cells per unit wet weight and in the water content of the livers. The nucleic acid content of the cells at hepatectomy, on the other hand, was unchanged.

Changes in nucleic acids of cells of mouse mammary carcinoma following exposure to X-ray of low dosage

1. 1. X-radiation of 150 roentgens caused a continuous decrease in the nucleic acid content of cells of a mouse mammary carcinoma uniformly suspended in Ringer's solution. A dosage of 100 r. produced a variable increase in cellular nucleic acids, which appeared to be correlated with high RNAP DNAP ratios. We believe that this dosage of X-ray caused some increased susceptibility of DNA to alkaline hydrolysis, as shown by the Schmidt-Thannhauser technic (6), resulting in an abnormal increase in RNAP from 30 to 90 minutes after exposure. A dosage of 75 r. produced changes in cellular nucleic acid difficult to interpret. 2. 2. The increase in nucleic acids following X-radiation of 100 r. was interpreted as a synthesis from precursors existing in the cell rather than a recovery from a temporary inhibition in their development. 3. 3. Increase and decrease in RNAP following X-radiation are shown graphically in the accompanying figures in two ways; one, associated with DNAP and taking place up and down along the lines of regression, and the other, not associated with DNAP and occurring in a perpendicular line up and down between lines of regression at points with fixed values of DNAP. In our experiments recorded here, these values of cellular DNAP are 1.00, 1.20 and 1.50 × 10 9− mg.