Flow-microfluorometric analysis of Escherichia coli, Rhizobium meliloti, and Rhizobium japonicum at different stages of the growth cycle

1977 ◽  
Vol 23 (9) ◽  
pp. 1165-1169 ◽  
Author(s):  
Alan S. Paau ◽  
Joe R. Cowles ◽  
James Oro
Keyword(s):  
Escherichia Coli ◽  
Nucleic Acid ◽  
Stationary Phase ◽  
Cell Size ◽  
Acid Content ◽  
Rhizobium Meliloti ◽  
Growth Cycle ◽  
Rhizobium Japonicum ◽  
Nucleic Acid Content ◽  
E Coli

The applicability of flow-microfluorometry (FMF) to the study of bacterial samples was investigated on cultures of Rhizobium meliloti, Rhizobium japonicum, and Escherichia coli using fluorescent and light-scattering signals. This technique which analyzes individual bacterial cells in a population was used to monitor the relative change in nucleic acid content and cell size during the growth cycle of the three microorganisms which were known to have different growth rates.Early log-phase E. coli cells contained at least eightfold more nucleic acid and were significantly larger than the stationary-phase cells. Cultures of early log-phase R. meliloti cells contained three to four-fold more nucleic acid and were slightly larger than cells in the stationary phase. Rhizobium japonicum had very little change in either parameter. In general, the amount of change in both cell size and nucleic acid content upon initiation of log-phase growth was related to the overall growth rate of the organisms, with E. coli experiencing the greatest change and R. japonicum the least. Results obtained by FMF analyses, therefore, were consistent with observations reported by earlier workers. Cultures of R. meliloti also were used to demonstrate that the intensity of the fluorescent signals was sensitive to digestion by DNase and RNase and to prolonged storage and fixation. The potential use of FMF in the study of microorganisms is discussed.

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

2002 ◽  
Vol 28 ◽  
pp. 131-140 ◽  
Author(s):  
P Lebaron ◽  
P Servais ◽  
AC Baudoux ◽  
M Bourrain ◽  
C Courties ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Nucleic Acid ◽  
Cell Size ◽  
Acid Content ◽  
Specific Activity ◽  
Nucleic Acid Content

Studies on bacteroid size and nucleic acid content of alfalfa bacteroids fractionated by velocity sedimentation

1978 ◽  
Vol 24 (10) ◽  
pp. 1283-1287 ◽  
Author(s):  
Alan S. Paau ◽  
Joe R. Cowles
Keyword(s):  
Nucleic Acid ◽  
Acid Content ◽  
Rhizobium Meliloti ◽  
Velocity Sedimentation ◽  
Nucleic Acid Content ◽  
Free Living ◽  
Positive Correlation ◽  
Fraction I

A velocity sedimentation procedure was described to fractionate bacteroids of alfalfa nodules into four subpopulations. Bacteroids in these subpopulations were different in size and nucleic acid content as determined by microscopy and flow-microfluorbmetry (FMF). The slowest-sedimenting bacteroids (fraction I) were small and resembled free-living Rhizobium meliloti both in size and nucleic acid content. The fastest-sedimenting bacteroids (fraction IV) were 2 to 3 times longer and contained 3 to 4 times more nucleic acid than the small bacteroids in fraction I and free-living R. meliloti. A positive correlation was established between bacteroid size and relative nucleic acid content of bacteroids in alfalfa nodules.

scholarly journals Characterization of the Population of the Sulfur-Oxidizing Symbiont of Codakia orbicularis (Bivalvia, Lucinidae) by Single-Cell Analyses

2007 ◽  
Vol 73 (7) ◽  
pp. 2101-2109 ◽  
Author(s):  
Audrey Caro ◽  
Olivier Gros ◽  
Patrice Got ◽  
Rutger De Wit ◽  
Marc Troussellier
Keyword(s):  
Nucleic Acid ◽  
Single Cell ◽  
Cell Size ◽  
Acid Content ◽  
Size Analysis ◽  
Nucleic Acid Content ◽  
Tropical Environments ◽  
Multiple Copies ◽  
Sulfur Oxidizing ◽  
Light Side

ABSTRACT We investigated the characteristics of the sulfur-oxidizing symbiont hosted in the gills of Codakia orbicularis, a bivalve living in shallow marine tropical environments. Special attention was paid to describing the heterogeneity of the population by using single-cell approaches including flow cytometry (FCM) and different microscopic techniques and by analyzing a cell size fractionation experiment. Up to seven different subpopulations were distinguished by FCM based on nucleic acid content and light side scattering of the cells. The cell size analysis of symbionts showed that the symbiotic population was very heterogeneous in size, i.e., ranging from 0.5 to 5 μm in length, with variable amounts of intracellular sulfur. The side-scatter signal analyzed by FCM, which is often taken as a proxy of cell size, was greatly influenced by the sulfur content of the symbionts. FCM revealed an important heterogeneity in the relative nucleic acid content among the subclasses. The larger cells contained exceptionally high levels of nucleic acids, suggesting that these cells contained multiple copies of their genome, i.e., ranging from one copy for the smaller cells to more than four copies for the larger cells. The proportion of respiring symbionts (5-cyano-2,3-ditolyl-terazolium chloride positive) in the bacteriocytes of Codakia revealed that around 80% of the symbionts hosted by Codakia maintain respiratory activity throughout the year. These data allowed us to gain insight into the functioning of the symbionts within the host and to propose some hypotheses on how the growth of the symbionts is controlled by the host.

scholarly journals THE RIBONUCLEIC ACID AND DEOXYRIBONUCLEIC ACID CONTENT OF MONONUCLEAR LEUKOCYTES OF DIFFERENT SIZES STUDIED BY ULTRAVIOLET CYTOPHOTOMETRY

1973 ◽  
Vol 21 (7) ◽  
pp. 628-633 ◽  
Author(s):  
ALFREDO MARIANO GARCIA ◽  
PATRICIA A. N. SULLIVAN
Keyword(s):  
Nucleic Acid ◽  
Cell Size ◽  
Dna Content ◽  
Random Error ◽  
Acid Content ◽  
Deoxyribonucleic Acid ◽  
Nucleic Acid Content ◽  
Mononuclear Leukocytes ◽  
Total Nucleic Acid ◽  
Dna And Rna

Rat mononuclears (lymphocytes and monocytes) were studied for total nucleic acid content by means of ultraviolet cytophotometry. Another set was treated with ribonuclease, and deoxyribonucleic acid (DNA) was measured using the same technique. It was found that total nucleic acid content (DNA and RNA) increases linearly with cell size from about 20 units in lymphocytes having 5 µ in diameter up to around 30 units in cells having 12-14 µ in diameter; this is to say, an almost 50% increase for a 6-7-fold enlargement. After ribonuclease treatment, however, the value of the integrated extinction (DNA) tends to remain constant for different cell sizes. A 650% variation in area is accompanied by a DNA change of less than 6%. The differences between treated and nontreated cells are nonsignificant for populations having up to 7.0-7.5 µ in diameter, which implies that small lymphocytes either have a negligible amount of RNA or that the instrument is not sensitive enough to detect it (less than 7% of the DNA content, this figure being the random error of our technique). These differences become highly significant for mononuclears having 8 µ or more in diameter. Therefore, while DNA tends to be constant and independent from cell size, RNA content tends to be harmoniously inconstant, since it is correlated with cell (and nuclear) size and degree of chromatin diffusion.

scholarly journals Characteristics and Dynamics of Bacterial Populations during Postantibiotic Effect Determined by Flow Cytometry

1998 ◽  
Vol 42 (5) ◽  
pp. 1005-1011 ◽  
Author(s):  
Magnús Gottfredsson ◽  
Helga Erlendsdóttir ◽  
Ásbjörn Sigfússon ◽  
Sigurdur Gudmundsson
Keyword(s):  
Flow Cytometry ◽  
Nucleic Acid ◽  
Fluorescence Microscopy ◽  
Acid Content ◽  
Quantitative Information ◽  
Nucleic Acid Content ◽  
Dependent Manner ◽  
Postantibiotic Effect ◽  
E Coli ◽  
Drug Removal

ABSTRACT Changes in bacterial ultrastructure after antibiotic exposure and during the postantibiotic effect (PAE) have been demonstrated by electron microscopy (EM). However, EM is qualitative and subject to individual interpretation. In contrast, flow cytometry gives qualitative and quantitative information. The sizes and nucleic acid contents of Escherichia coli and Pseudomonas aeruginosa were studied during antimicrobial exposure as well as during the PAE period by staining the organisms with propidium iodide and analyzing them with flow cytometry and fluorescence microscopy. The effects of ampicillin, ceftriaxone, ciprofloxacin, gentamicin, and rifampin were studied for E. coli, whereas for P. aeruginosa imipenem and ciprofloxacin were investigated. After exposure of E. coli to ampicillin, ceftriaxone, and ciprofloxacin, filamentous organisms were observed by fluorescence microscopy. These changes in morphology were reflected by increased forward light scatter (FSC) and nucleic acid content as measured by flow cytometry. For the β-lactams the extent of filamentation increased in a dose-dependent manner after drug removal, resulting in formation of distinct subpopulations of bacteria. These changes peaked at 20 to 35 min, and bacteria returned to normal after 90 min after drug removal. In contrast, the subpopulations induced by ciprofloxacin did not return to normal until >180 min after the end of the classically defined PAE. Rifampin resulted in formation of small organisms with low FSC, whereas no distinctive characteristics were noted after gentamicin exposure. For P. aeruginosa an identifiable subpopulation of large globoid cells and increased nucleic acid content was detected after exposure to imipenem. These changes persisted past the PAE, as defined by viability counting. Swollen organisms with increased FSC were detected after ciprofloxacin exposure, even persisting during bacterial growth. In summary, for β-lactam antibiotics and ciprofloxacin, the PAE is characterized by dynamic formation of enlarged cell populations of increased nucleic acid content, whereas rifampin induces a decrease in size and nucleic acid content in the organisms. Flow cytometry is an ideal method for future studies of bacterial phenotypic characteristics during the PAE.

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