The growth of bacteriophage and lysis of the host

1. A new strain of B. coli and of phage active against it is described, and the relation between phage growth and lysis has been studied. It has been found that the phage can lyse these bacteria in two distinct ways, which have been designated lysis from within and lysis from without. 2. Lysis from within is caused by infection of a bacterium by a single phage particle and multiplication of this particle up to a threshold value. The cell contents are then liberated into solution without deformation of the cell wall. 3. Lysis from without is caused by adsorption of phage above a threshold value. The cell contents are liberated by a distension and destruction of the cell wall. The adsorbed phage is not retrieved upon lysis. No new phage is formed. 4. The maximum yield of phage in a lysis from within is equal to the adsorption capacity. 5. Liberation of phage from a culture in which the bacteria have been singly infected proceeds at a constant rate, after the lapse of a minimum latent period, until all the infected bacteria are lysed. 6. If the bacteria are originally not highly in excess, this liberation is soon counterbalanced by multiple adsorption of the liberated phage to bacteria that are already infected. This leads to a reduction of the final yield

Adsorption of bacteriophage under various physiological conditions of the host

The first step in the growth of bacteriophage is the combination of phage with the susceptible bacterial host. The rate of this combination is, under simple conditions, proportional to both the bacterial concentration and to the phage concentration. Various aspects of this process have been studied quantitatively by previous workers (1, 2). Their results will be analyzed and discussed in the sections entitled "Residual free phage," and "Theory of adsorption rates." The main purpose of this paper was the study of a detail of the adsorption process that had not previously received attention, namely the dependence of the rate constant of adsorption on the physiological state of the bacterial host. Such a dependence must be anticipated for two reasons. First, it is known that the size of a bacterium changes very considerably depending on its phase of growth in a given culture medium, and an increased cell surface should lead to an increase of the adsorption rate on to a given number of bacteria. Second, for motile bacteria, like E. coli, the adsorption will be faster when the bacteria move about rapidly than when their motility is reduced by adverse physiological conditions. Our experiments show that the rate constant under optimal conditions is more than sixty times greater than under poor conditions

Photoreactions in Phycomyces. Responses to the stimulation of narrow test areas with ultraviolet light

Equipment has been developed for ultraviolet illumination of sharply bounded test areas of the growing zone of sporangiophores of Phycomyces. The growing zone is opaque for this light and the tropic responses are negative. Periodic short narrow stimuli on alternating sides produce periodic tropic responses when applied at x > 0.5 mm, but none for x 0.1 mm. The lag between stimulus and response is 3.3 min. for any x > 0.5 mm. For smaller x the lag increases progressively. In all cases the tropic bend occurs at values of x > 0.5 mm. Sustained tropic stimuli, applied at constant height relative to ground, produce relatively sharp tropic bends. The center of the bend is at all times close to the simultaneous position of the stimulated area. The boundaries of a light-adapted zone move less than 0.1 mm in 10 min. relative to the sporangium. It is concluded that the receiving and adapting structures do not move relative to the sporangium, and that the responding system does not move relative to ground. The two systems move relative to each other with the speed of growth. The responding system does not extend above x = 0.5 mm

Effects of Cold Periods on the Stimulus-Response System of Phycomyces

The sporangiophores of Phycomyces do not exhibit phototropic responses when growth is arrested reversibly by cooling to 1°C. Unilateral UV stimuli (254 nm) applied during cold periods are stored for at least 2 hr and produce tropic responses away from the light after warm-up. During the cold period dark adaptation proceeds at a rate which decreases with the temperature

Photoreactions in Phycomyes: growth and tropic responses to the stimulation of narrow test areas

Sporangiophores of Phycomyces in stage IV b have been stimulated by parallel light in test areas 0.2 mm. wide. The growth responses to large stimuli are very large, owing probably to light scattered within the specimen. For medium stimuli the sensitive zone coincides with the growth response zone obtained previously and excludes the region of maximum stretch. Sustained stimulations were used to elicit tropic responses. The bends formed travel away from the sporangium at a speed equal to the growth speed. Thus they remain very close to the stimulus when this is held at a constant level relative to ground but separate from it for stimuli programmed differently. The existence of a protoplasmic structure, the "inner wall," with the following properties is postulated: it is attached to the lower, non-growing part of the sporangiophore and grows by addition above the sensitive zone. It neither stretches nor twists in the sensitive zone. It is the seat of the light receptors and gives growth and tropic responses. The cell wall follows its bends by elastic stretch

The growth of bacteriophage

1. An anti-Escherichia coli phage has been isolated and its behavior studied. 2. A plaque counting method for this phage is described, and shown to give a number of plaques which is proportional to the phage concentration. The number of plaques is shown to be independent of agar concentration, temperature of plate incubation, and concentration of the suspension of plating bacteria. 3. The efficiency of plating, i.e. the probability of plaque formation by a phage particle, depends somewhat on the culture of bacteria used for plating, and averages around 0.4. 4. Methods are described to avoid the inactivation of phage by substances in the fresh lysates. 5. The growth of phage can be divided into three periods: adsorption of the phage on the bacterium, growth upon or within the bacterium (latent period), and the release of the phage (burst). 6. The rate of adsorption of phage was found to be proportional to the concentration of phage and to the concentration of bacteria. The rate constant ka is 1.2 x 10–9 cm.8/min. at 15°C. and 1.9 x 10–9 cm.8/min. at 25°. 7. The average latent period varies with the temperature in the same way as the division period of the bacteria. 8. The latent period before a burst of individual infected bacteria varies under constant conditions between a minimal value and about twice this value. 9. The average latent period and the average burst size are neither increased nor decreased by a fourfold infection of the bacteria with phage. 10. The average burst size is independent of the temperature, and is about 60 phage particles per bacterium. 11. The individual bursts vary in size from a few particles to about 200. The same variability is found when the early bursts are measured separately, and when all the bursts are measured at a late time

Responses of Phycomyces indicating optical excitation of the lowest triplet state of riboflavin

Phototropic and light growth responses of the sporangiophore of Phycomyces have been elicited using tunable laser stimulation from 575 to 630 nm. The growth response shows additional components of the action spectrum with a sharp peak at 595 nm, a sharp cut-off at 585 nm, and a tail extending beyond 630 nm. The integral over the electronic transition (f-value) is 1.5 x 10^-9 times that at 455 nm. These parameters indicate a direct transition from the ground state to the lowest triplet state of riboflavin

Interplay between the Reactions to Light and to Gravity in Phycomyces

Sporangiophores of Phycomyces do not grow directly towards a horizontal beam of light, but equilibrate at an angle of about 30° above the horizontal. After describing several related observations, this paper suggests that the dioptric properties of an obliquely illuminated cylindrical lens, illustrated by a dummy cell, as well as a negative geotropic response, play major roles in determining the direction of growth. The shift of the equilibrium direction of growth towards the vertical, or a purely geotropic response, over a tenfold range of very low intensities (around 10^6 quanta/cm^2 sec., or 10^-13 watt/cm^2) has been studied, and an action spectrum made, measuring the quantum fluxes producing a standard intermediate equilibrium direction of growth at different wavelengths. This may differ from the action spectra at higher intensities in lacking conspicuous maxima from 370 to 490 mµ. However, in the ultraviolet it parallels the other spectra, although without showing the much higher quantum efficiency of ultraviolet relative to visible light previously noted. Possible interpretations are discussed

Absorption and Screening in Phycomyces

In vivo absorption measurements were made through the photosensitive zones of Phycomyces sporangiophores and absorption spectra are presented for various growth media and for wavelengths between 400 and 580 mµ. As in mycelia, ß-carotene was the major pigment ordinarily found. The addition of diphenylamine to the growth media caused a decrease in ß-carotene and an increase in certain other carotenoids. Growth in the dark substantially reduced the amount of ß-carotene in the photosensitive zone; however, growth on a lactate medium failed to suppress ß-carotene in the growing zone although the mycelia appeared almost colorless. Also when diphenylamine was added to the medium the absorption in the growing zone at 460 mµ was not diminished although the colored carotenoids in the bulk of the sporangiophore were drastically reduced. Absorption which is characteristic of the action spectra was not found. Sporangiophores immersed in fluids with a critical refractive index show neither positive nor negative tropism. Measurements were made of the critical refractive indices for light at 495 and 510 mµ. The critical indices differed only slightly. Assuming primary photoreceptors at the cell wall, the change in screening due to absorption appears too large to be counterbalanced solely by a simple effect of the focusing change. The possibility is therefore advanced that the receptors are internal to most of the cytoplasm; i.e., near the vacuole

Meiosis in Phycomyces

A four-factor cross between two strains of Phycomyces involving two auxotrophic, one color, and the mating type marker is described. Samples of 40 germspores from 84 individual fertile germsporangia were characterized. The results show: (i) The germspores of a germsporangium are derived from one meiosis in approximately 78% of the cases. (ii) The four markers are on separate chromosomes. They are nonselective. (iii) Analysis of a large sample of germspores from 106 pooled germsporangia confirms that the four markers are unlinked. (iv) From the ditype/tetratype ratios it is inferred that each marker is located about 15 map units from its centromere