It is crucial to the reproducibility of results and their proper interpretation that the conditions under which experiments are carried out be defined with rigour and consistency. In this review we attempt to clarify the differences and interrelationships among steady, balanced and exponential states of culture growth. Basic thermodynamic concepts are used to introduce the idea of steady-state growth in open, biological systems. The classical, sometimes conflicting, definitions of steady-state and balanced growth are presented, and a consistent terminology is proposed. The conditions under which a culture in balanced growth is also in exponential growth and in steady-state growth are indicated. It is pointed out that steady-state growth always implies both balanced and exponential growth, and examples in which the converse does not hold are described. More complex situations are then characterized and the terminology extended accordingly. This leads to the notion of normal growth and growth that can be synchronous or otherwise unbalanced but still reproducible, and to the condition of approximate steady state manifested by growth in batch culture and by asymmetrically dividing cells, which is analysed in some detail.

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Experimental verification is provided for the theoretical expressions (see preceding article, I. Theory) describing the electrical processes that take place during the passage of an aqueous suspension of rigid, nonconducting spheres (ragweed pollen) through an orifice across which there exists an electrical field, for a large range of orifice dimensions; the instrumentation developed is considered in some detail. The effective length of an orifice as deduced from conductivity measurements is shown to be essentially the same as that predicted theoretically. Absolute volume distributions are presented of a suspension of polystyrene latex spheres as determined electrically (mean 11.17 mu(3), c. v. 4.2%) and with an electron microscope (mean 11.01 mu(3), c. v. 4.1%). Conflicting experimental results reported in the literature are discussed.

Escherichia coli nucleoids were visualized after the DNA of OsO4-fixed but hydrated cells was stained with the fluorochrome DAPI (4',6-diamidino-2-phenylindole dihydrochloride hydrate). In slowly growing cells, the nucleoids are rod shaped and seem to move along the major cell axis, whereas in rapidly growing, wider cells they consist of two-to four-lobed structures that often appear to advance along axes lying perpendicular or oblique to the major axis of the cell. To test the idea that the increase in cell diameter following nutritional shift-up is caused by the increased amount of DNA in the nucleoid, the cells were subjected to DNA synthesis inhibition. In the absence of DNA replication, the nucleoids continued to move in the growing filaments and were pulled apart into small domains along the length of the cell. When these cells were then transferred to a richer medium, their diameters increased, especially in the region enclosing the nucleoid. It thus appears that the nucleoid motive force does not depend on DNA synthesis and that cell diameter is determined not by the amount of DNA per chromosome but rather by the synthetic activity surrounding the nucleoid. Under the non-steady-state but balanced growth conditions induced by thymine limitation, nucleoids become separated into small lobules, often lying in asymmetric configurations along the cell periphery, and oblique and asymmetric division planes occur in more than half of the constricting cells. We suggest that such irregular DNA movement affects both the angle of the division plane and its position.A bacterial cell maintains its DNA in a central space called the nucleoid. In living cells, this domain can be visualized either by phase contrast in the presence of gelatin (25) or by fluorescence microscopy with a fluorochrome (13). Fixing the cells with osmium tetroxide results in nucleoids that are more compact (5, 39) but that still retain characteristic shapes which seem to reflect the replication and segregation states of the chromosome (see also references 7, 16, and 32). These distinct nucleoid shapes indicate that the DNA is not dispersed randomly throughout the cytoplasm but that most of it is confined to a region separated from the cytoplasmic phase by some unknown chemical property (38). This phenomenon does not preclude the presence throughout the cytoplasm of short loops of DNA that extend towards the plasma membrane and are actively involved in transcription and translation (14,33). That such connections are indeed effective in shaping the nucleoid is suggested by the old observation that treatment of cells with chloramphenicol causes the nucleoid to round off (21).Several models for the various mechanisms and forces involved in the movement and positioning of the chromosomes within the cell have been advanced. The classical replicon hypothesis (20) considered envelope growth between postulated sites of chromosomal attachment to the plasma membrane. However, the rigid peptidoglycan layer has been shown to be synthesized in a dispersed mode ...

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