- Volume 45, Issue 3, 1966

SUMMARY: Phage-like objects obtained from bacteriocinogenic strains of Escherichia coli, Pseudomonas aeruginosa and Listeria monocytogenes by the differential centrifugation of culture fluids were examined in the electron microscope. Particles with small heads and contractile tails were found associated with strains of E. coli. The three P. aeruginosa strains all produced numerous headless contractile tails in broth cultures together with a very few phage-like objects with flexible non-contractile tails. The headless tails possessed cores which appeared both solid and hollow in the electron microscope, indicating the presence of some unidentified substance, perhaps nucleic acid. The only type of particle found associated with L. monocytogenes was a large bacteriophage with a complex contractile tail. The structure of these particles is described and discussed, and it is concluded that some of them are the bactericidal principles of spontaneously released bacteriocins.

SUMMARY: Theoretical cell-size distributions for populations of growing cells are calculated for different models of cell growth and for different degrees of variability in size of cells at division. From these computations, it is concluded that the coefficient of variation (c.v.) is almost independent of the relationship of growth rate to cell size. It is 20% if there is no variability in the cell size at division. For a case typical for enteric rod-shaped bacteria, the variability in cell size at division is about 10% and the calculated c.v. in cell size of the population in this case increases to 22-23%. Calculations based on the microscopic observations of others are in the range of 20-25%. It is proposed that the c.v. of the size distribution serve as a standard in assessing the accuracy of the electronic instruments that size bacteria.

Evidently, only the higher moments of the population cell size distribution contain information bearing on the growth dependence of the organisms on their size. It is pointed out that this means that the Collins-Richmond principle must be applied only to precise and accurate data.

SUMMARY: ‘Lucerne’ auxotrophs of Verticillium albo-atrum produced by ultraviolet irradiation and derived from the same parental strain formed heterokaryons on minimal agar 4-5 weeks after inoculation. A degree of heterokaryon incompatibility was shown between auxotrophs derived from different isolates of V. albo-atrum and one of V. dahliae. Conidial methods of heterokaryon synthesis failed but transference of agar inocula from complete medium agar cultures to minimal medium agar was successful. Conidia were uni-nucleate and nuclear migration subsequent to germ-tube fusion was observed. Heterokaryons grew slowly and were unstable, but by using di-auxotrophs a greater stability was achieved and a number were subcultured successfully for more than a year. Spore analysis yielded colonies of both parental types with various ratios. A heterokaryon test between a hyaline and normal dark auxotroph of V. albo-atrum indicated that an important system controlling morphogenesis might be linked with a cytoplasmic factor. A hypothesis is proposed to account for the production and degree of stability of the hyaéine variants.

SUMMARY: In the leaves of their natural (homologous) hosts, the bean and cherry respectively, Pseudomonas phaseolicola and P. morsprunorum increased logarithmically for at least 4 days after inoculation. Transition into the stationary phase was gradual and accompanied by water congestion of the infected tissues, followed by typical field symptoms. Increases in the inoculum dose had relatively little or no effect on the generation time but growth ceased earlier at the higher doses. In the reciprocal (heterologous) combinations, logarithmic growth was abruptly terminated after 2-3 days, due apparently to a specific defensive reaction in the host. This coincided at the higher inoculum doses with the appearance of dry necrotic symptoms in the leaves. No macroscopic symptoms were observed with the lower doses, and with the lowest dose in bean there was an acceleration of leaf maturation in the presence of heterologous organisms. Generation times were lower in heterologous combinations but increased markedly with the inoculum dose. The growth of the pear strain of P. syringae in bean and cherry leaves showed typical heterologous characteristics. The final yields of bacteria per unit inoculum were invariably higher in homologous combinations, but they decreased with increasing dose, whereas heterologous yields increased. The differences in net growth response were therefore greatest at the lowest doses. This suggested that host specificity in the field was associated with factors controlling growth of the organisms in vivo from small initial inocula.

SUMMARY: Cholesterol enrichment cultures from soil samples yielded various Nocardia and Streptomyces spp. capable of metabolizing cholesterol aerobically as a sole carbon source. One of these isolates, designated Streptomyces 14PH8, formed haloes (cholesterol-free zones surrounding colonies) on cholesterol mineral salts agar. This organism was selected for further study as it was able to utilize 80-100% of 0·1% (w/v) cholesterol in a mineral salts medium in 6 days. Oxidation of the sterol was initiated by a cholesterol dehydrogenase, giving 4-cholestene-3-one. The latter compound was then hydroxylated to form 4-cholestene-4-ol-3-one. Isotopic tracer studies revealed all of carbon-4 and most of carbon-26 of cholesterol were converted to 14C02. However, some of carbon-26 was converted to cell material.

Streptomyces 14ph8 gave several variants, one of which (Streptomyces 14ph8 no. 2 var. A) could be reclassified as a Nocardia, using the same method of Gordon & Smith (1955).

SUMMARY: Red blood cells from a group of fowls were sensitive to the haemag-glutinins of vaccinia and Coxsackie A7 viruses but the red cells from certain fowls exhibited differences in their relative sensitivities. Differences between the reaction of these haemagglutinins with susceptible red cells of the same sensitivity were shown by treatment with chemical and physical agents. There were differences in the effect of pH value on the haemagglutination titres and, unlike Coxsackie A7 hemagglutinin, the adsorption of vaccinia haemagglutinin to susceptible red cells was inhibited by divalent cations. Red cell receptors for both haemagglutinins were insensitive to RDE (receptor destroying enzyme) but were inactivated by treatment with potassium periodate, papain or α-chymotrypsin. There were quantitative differences in the degrees or rates of receptor destruction by these reagents. Haemagglutination by vaccinia and Coxsackie A7 haemagglutinins was inhibited only by homologous antiserum. These qualitative and quantitative differences indicate separate red cell receptors for the two haemagglutinins.

SUMMARY: The colicinogenic factor colI derived from Shigella sonnei colicine type 2 (Abbott & Shannon, 1958) was transmitted between different strains of Shigella flexneri in serotypes 1a, 1b, 2a, 2b, 3a, 3c, 4a, 4b, 4c, 5a, X and Y when donor and acceptor strains were grown together in mixed culture in aerobic static broth for a period of 20 hr or 48 hr at 37·. The rate of transfer was influenced by the state of fimbriation of the acceptor bacteria, but not by that of the donor bacteria. The proportion of acceptor organisms acquiring colI was very high, e.g. 15-40 % in 20 hr and 50-95 % in 48 hr, when the acceptor was genotypically fimbriate and in the fimbriate phase (Fim+F); but it was much lower, e.g. 0-5 % in 20 hr and 1-15 % in 48 hr, when the acceptor was either genotypically non-fimbriate (Fim−) or genotypically fimbriate but in the non-fimbriate phase (Fim+N). Although it facilitated the transmission of colI, the presence of fimbriae was not essential for transmission, since transmission at a low rate was obtained in most crosses of Fim− donors with Fim− acceptors. Only 1 out of 17 strains of S. flexneri tested as acceptors failed to acquire colI in any test. Transmission did not take place within the first 2 hr of mixed culture; it occurred mainly during the period of slow growth after the end of the exponential phase (i.e. between 8 and 48 hr) when, with fimbriate organisms, a pellicle of bacteria had developed on the surface of the broth. The rate of transmission was very much decreased when pellicle formation was prevented by intermittent or continuous agitation of the culture. Experiments in which the donor bacteria were killed by adding streptomycin at 8 hr showed that an extensive ‘epidemic’ spread of colI took place within the acceptor population during the later stages of culture. When stationary-phase (1-10 days) broth cultures of donor and acceptor bacteria were mixed together and incubated without the addition of fresh broth, the rates of transfer at 20 and 48 hr were as high as in the mixed cultures grown from small inocula. Transmission also occurred with high frequency in a defined medium containing glucose, NH4 + and nicotinic acid.

SUMMARY: The reduction of ferricyanide by resting and actively growing Escherichia coli was studied. Under anaerobic conditions ferricyanide acted as a hydrogen acceptor for the complete oxidation of glucose and TCA cycle intermediates. However, reduction of ferricyanide was not coupled to ATP formation. The anaerobic molar growth yield from glucose in the presence of ferricyanide was even less than in its absence. Ferricyanide repressed the synthesis of formate hydrogenylase.

SUMMARY: Two thy − Hfr strains of Escherichia coli K12 were mated with stationary phase W677 F− at intervals during thymine starvation. With both strains, Hfr bacteria retained their capability of chromosome transfer for a period after loss of colony-forming ability. Eventual loss of transfer capability in strain HfrC thy − was associated with inactivation of some early stage of the transfer process. Thymine starved HfrC thy − transferred the genetic markers pro, thr, leu and thi with a normal gradient but HfrB1 thy − showed marker inactivation. The extent of marker inactivation in this strain was consistent with the presence of 10-20 breaks/chromosome. It is suggested that the strains may have behaved differently in these experiments as a result of differences in their ability to modify primary genetic lesions resulting from thymine starvation.

SUMMARY: Virions of the papillomaviruses of man, cattle, dog and rabbit were compared by immunodiffusion in agar. Antigen reactants were prepared from saline extracts of warts, and consisted either of crude virus-particle concentrates, or of ‘full’ or ‘empty’ fractions from equilibrium density gradients. There were no cross-reactions between the wart viruses of different species. Within any one species, ‘full’ and ‘empty’ particles gave antigenically identical single bands of precipitate, whether derived from the same or from different warts; but some old preparations of ‘empty’ particles gave a second band.

SUMMARY: Streptococcus lactis was grown in batch culture in a complex organic medium. Growth of the culture was followed by estimating the extinction, the dry weight, protein, DNA and RNA of the organism; basic peptides were estimated by electrophoresis and nisin by bio-assay. By the end of the lag phase of growth when active DNA and RNA synthesis were already proceeding, the nisin which had been introduced with the inoculum could not be recovered, although the cocci contained other basic peptides. Rapid nisin synthesis started after an increase of about 50 % in the dry weight of the cocci had taken place. Initially, high molecular weight nisin was made, the concentration of which decreased as the incubation proceeded. This was followed by the production of lower molecular weight nisin which could be recovered 24 hr after the end of the incubation. Inocula of organisms derived at intervals from the parent culture were tested in secondary cultures for length of lag of growth by re-inoculation into fresh medium of the same composition. During the lag phase of growth of the parent culture, the length of lag of growth of the secondary cultures fell to about half the original time. The shortest lag of the secondary cultures coincided with commencement of logarithmic growth of the parent culture and with the disappearance of cellular nisin from the parent culture. The length of the lag of the secondary cultures returned to their original value as the parent culture reached stationary phase and coincided with maximum nisin concentration in the parent culture. It is suggested that the length of lag of growth is related to the presence of basic peptides.

SUMMARY: The process of regeneration of Fusarium culmorum protoplasts, obtained by means of strepzyme RA, has been followed. Protoplast regeneration started with the formation of cellular aggregates originating a mass of globular forms which later gave rise to the formation of a pseudomycelial form. Regeneration of protoplasts was formed in three different ways: (a) by means of a chain of yeast-like forms and later originating a germ tube; (b) by direct formation of the germ tube from the protoplast; (c) through a complicated process with the formation of various globular forms and giving rise to the formation of the germ tube at the end. Maximum regeneration, about 80 % of the protoplasts, was found in the presence of 2% (w/v) sucrose (or 1% (w/v) glucose + 1% (w/v) fructose) + 10% (w/v) sorbose + 0·2% agar + the mineral salts of the Czapek medium. No other sugars were able to substitute for sucrose and sorbose. Agar was the best substance for regeneration; gelatin produced an inhibition. Regeneration was greater under acid conditions, alkaline pH values interfering with the phenomenon. The number of nuclei/protoplast varied from 1 to 4, the lack of them in some spherical forms perhaps being the cause of failure to regenerate. No differences were found in the regenerative process as between protoplasts obtained by the use of snail or by microbial lytic enzymes.

SUMMARY: The growth cycle of Nocardia corallina involves a period of coenocytic hyphal development terminated by fragmentation. Hyphal elongation occurred by insertion of new material at the hyphal tips, not by intercalary growth. Exceptions occurred at points of branch initiation. Development of cross-walls preceded fragmentation and the new tips so produced were also capable of elongation. The fragmentation process appeared to be initiated by agents which accumulated in the medium, since used broth from actively fragmenting cultures stimulated earlier fragmentation in assay inocula. This effect was not shown by media from cultures in the coenocytic hyphal phase, nor by old resting cultures. The population density also affected fragmentation in a manner which suggested that accumulation of threshold concentrations of a diffusible metabolic product was required. A Millipore filter technique for assaying the effects of agents involved in fragmentation is described.

SUMMARY: Verticillium albo-atrum mutant m5·6 which produces phytoene, β- and γ-carotene, neolycopene A, lycopene and neurosporaxanthin was subjected to the effect of the following inhibitors of carotenoid synthesis: diphenylamine, 2-hydroxydiphenyl, β-ionone, methylheptenone. Certain intermediates in the Porter-Anderson (1962) pathway for carotenoid biosynthesis not previously present were then observed, namely: phytofluene, β-zeacarotene, ζ-carotene, neurosporene. When the mutant was grown in the presence of diphenylamine, washed free from it and then grown again in 1 % glucose in m/15 KH2PO4, the more unsaturated polyenes seem to be formed at the expense of the more saturated ones. There was also evidence that γ-carotene was formed via β-zeacarotene and the sequence of carotenoid formation seems to agree very closely to that suggested by Porter & Anderson (1962).

SUMMARY: The sulphur-containing amino acids L-cysteine, L-cystine, L-methionine, L-homocysteine thiolactone, L-homocystine and glutathione at 10−4 to 10−5 M inhibited sclerotial formation without inhibition of mycelial growth and induced basidia formation in the fungus Sclerotium rolfsii Sacc. Higher concentrations inhibited linear growth and production of sclerotia. The effect of L-cysteine was competitively antagonized by iodoacetic acid at a molar ratio of 30:1. Iodoacetic acid induced sclerotial formation at 3 x 10−5 to 10−4 M, sclerotia being produced in circles around inoculation point. The possible role of -SH groups in the morphogenesis of S. rolfsii is discussed.

SUMMARY: The variations in potassium, magnesium and phosphorus contents of Aerobacter aerogenes grown in a chemostat, were measured as functions of the RNA content of the organisms. The RNA content was varied by altering temperature at a fixed dilution rate under potassium-limiting conditions, and by varying the growth rate at a fixed temperature under conditions of both magnesium- and phosphate-limitation. Changes in RNA were accompanied by corresponding changes in these other cellular components such that a molar stoichiometry close to 1:4:5:8 for magnesium, potassium, RNA (nucleotide) and phosphorus, respectively, was maintained. The only significant deviation from this ratio was observed in phosphate-limited organisms at low growth rates; these organisms possessed considerable amounts of carbohydrate. It is suggested that potassium, magnesium and phosphate are implicated in polysaccharide synthesis, thus changing the quantitative relationship between these three components and RNA previously observed. The results support the suggestion that most of the potassium in growing A. aerogenes is required to maintain the functional state of ribosomal particles.