- Volume 56, Issue 1, 1969

Ultraviolet irradiation of Escherichia coli k 12 bacteria, carrying an autonomous F factor or an F-prime factor, enhances the fertility of the population by increasing the number of cells which can transfer the bacterial chromosome. In contrast, under similar conditions the fertility of irradiated Hfr populations falls in proportion to the survivors. Following irradiation, the effect begins to develop after about 30 min. incubation in broth at 37°, reaches a peak at about 90 min., and thereafter slowly declines. The effect develops with similar kinetics during post-irradiation incubation in minimal medium as in broth, provided the bacteria have been minimal-grown; in the case of broth-grown cells, appearance of the effect in minimal medium is greatly delayed. A comparison of the kinetics of the effect with the growth of the population as a whole shows that the u.v.-induced donor state is not inheritable. Mitomycin C, which resembles u.v. radiation in producing DNA damage repairable by a mechanism involving excision of single-stranded fragments, also induces new donor bacteria. Other agents such as X-rays and methyl methansulphonate (MMS) do not stimulate the production of new donors but may enhance the recombination frequency since cells killed by them may continue to act as chromosome donors. The effect is not shown either by uvr mutants (unable to excise thymine dimers) or by rec mutants (unable to mediate recombination) which carry an F-prime factor. A possible mechanism is suggested whereby the excision of single-stranded fragments of the bacterial chromosome, during the repair of u.v. damage, facilitates pairing with homologous regions of the complementary sex factor strand. A recombination event, mediated by breakage and covalent bonding, then joins a free end of the excised DNA strand to the paired sex factor strand. In this way, recombination connects sex factor and chromosome by only a single strand instead of by the two strands which normally leads to insertion and the formation of an Hfr chromosome. It is postulated that such a structure can be transferred at conjugation, but is incapable of more than one cycle of replication.

Sterilizing doses of γ-radiation for food products (4·5 Mrad.) do not completely inactivate spore toxin or enzymes. The present study concerns spore toxin during and after irradiation of 4 × 106 spores of Clostridium botulinum per g. of canned ground beef irradiated to 4.5 Mrad., at temperatures of – 25°, 0° and + 25°. During subsequent incubation of irradiated samples at 30° for 10 months, periodic analyses showed no viable spores. Toxin was tested on (i) non-homogenized samples and (ii) samples homogenized in a tissue grinder.

In samples irradiated at – 25° and 0° and in un-irradiated un-incubated controls, 4 minimal lethal doses (m.l.d.) of toxin were found in non-homogenized samples immediately after irradiation. The toxin titre of irradiated samples gradually diminished to approximately half of its initial value at the end of 10 months at 30°. In identical sets of homogenized irradiated samples, 8 m.l.d. were found initially and the titre increased to 44 m.l.d. after 2 weeks to 4 months of incubation at 30°. Thereafter, the toxin titre slowly deteriorated to approximately 25 m.l.d. at the end of 10 months.

Samples irradiated at 25° exhibited a definite increase in toxin titre even immediately after irradiation. This seems to suggest the possibility of new toxin synthesis either during or after radiation. The toxin titre in homogenized samples reached 64 m.l.d. after 2 months at 30° and thereafter slowly diminished to 31 m.l.d. at 10 months.

Toxin titres in radiation-damaged spores were small in comparison to un-irradiated C. botulinum growing in beef which reached 1000 to 5000 m.l.d. in 2 days at 30°.

Thin sections of Leptotrichia buccalis were examined with the electron microscope. The organism showed a cell-wall profile characteristic of Gramnegative bacteria. A Gram-positive homofermentative Lactobacillus sp. was included in the study for comparison. The results indicate that L. buccalis should not be included within the Lactobacillaceae.

The growth of Mycobacterium smegmatis was inhibited by lack of zinc in a glycerol + asparagine medium depleted of trace metals but enriched with iron. Addition of zinc to this medium supported full growth of the organism. Both zinc-deficient and zinc-replete cultures had the same exponential doubling time. In zinc-deficient organisms the synthesis of RNA and DNA varied with growth rate in a manner typical for nutrient-induced decreases in a variety of organisms. Zinc-deficiency led to inhibition of nicotinamide synthesis and an increase in phosphate storage as compared with zinc-replete cultures. The pH value of zinc-deficient cultures was preponderantly alkaline. The results are discussed with reference to the metabolic role of zinc ions, and the effects of zinc deficiency is compared to the effects of iron deficiency as previously reported for the same organism.

Crowding of conidia of Glomerella cingulata on potato sucrose agar led to a yeast-like growth of closely opposing and sandwiched streaks whereas the outer edges of the streaks developed fluffy mycelia. In liquid cultures increasing the initial conidial inocula increased the development of conidia and correspondingly decreased the mycelia. Normally, conidia germinate to produce germ tubes which branch and develop into cottony mycelia (M phase). Crowded conditions (3000 or more conidia/mm.2 or about 2 million/ml.) gave rise to secondary conidia with or without short hyphae or pseudomycelia: the yeast-like cultures (Y phase). In addition to crowding, thorough aeration, protection from light radiation below 500 mμ, and a temperature of incubation c. 21° favoured the Y phase development on appropriate media. Bicarbonate or CO2 did not induce Y phase development. Washings and basic CH2Cl2 extracts of conidia (which contained auto-inhibitors) induced preferential development of conidia with concomitant decrease in mycelia. Diffusible substances associated with the conidia, in appropriate concentrations, are considered responsible for inhibition of mycelia and preferential development of Y phase.

A respiratory-competent strain of Saccharomyces cerevisiae (1 D) had mitochondria of diverse morphology. A spontaneous petite strain and petites induced by acriflavine, ultraviolet-radiation or phenethyl alcohol showed similar mitochondrial diversity to the parent strain, but in all the petites except the spontaneous one there were fewer mitochondrial profiles showing cristae. In another respiratory-competent strain (1 1) the mitochondria were of uniform morphology. A phenethyl alcohol-induced petite derived from this strain again showed fewer cristate mitochondrial profiles. There was no evidence of the grossly aberrant and lamellate mitochondrial profiles observed in strain 1 D and its petites. Each of the cultures obtained from an ascospore tetrad from a 1 D × 1 1 cross was induced to form petites with phenethyl alcohol ; all showed a significant reduction in the proportion of mitochondrial profiles with cristae. Two of them showed the mitochondrial diversity characteristic of parent strain 1 D. Other features of mitochondrial morphology appear to be under nuclear control and are strain-dependent. All the petites were cytoplasmic mutants and lacked cytochromes a and b.

Clubroot formation on Sinapis alba plants infected with Plasmodiophora brassicae was not prevented by application of 2,3,5-triiodobenzoic acid or α-naphthylphthalamic acid (inhibitors of polar transport of auxins in plants) to the hypocotyls. However, the fresh weight of club tissue produced on treated plants was lower than on untreated plants. This difference in weight was of the same order as the difference in total fresh weights of treated and untreated plants. Explants of tumour tissue from two Brassica species infected with P. brassicae, unlike explants from healthy roots, produced callus on a tissue culture medium without growth substances. Callus growth from infected tissues depended on the presence of active vegetative plasmodia in the cells. Established clones of Brassica rapa (Golden Ball) callus infected with vegetative plasmodia of P. brassicae, unlike healthy callus clones, grew well on a series of media deficient in one or more growth substances. Growth of infected Golden Ball callus on medium without growth substances was prevented by the tryptamine antagonist xylamidine tosylate and by the monoamine oxidase inhibitor iproniazid. Inhibition was annulled by including abnormal growth of Brassica roots infected with P. brassicae is discussed.

Phenethyl alcohol (PEA) has been found to obstruct genetic transformation in Bacillus subtilis. A concentration of 0·3% (v/v) PEA prevents the development of competence and destroys predeveloped competence without causing a measurable decrease in viable cells. Treatment of competent cells with 0·3 % PEA prior to their exposure to transforming deoxyribonucleic acid (DNA) causes the cells to become incapable of irreversibly incorporating DNA. Treatment of competent cells which have irreversibly incorporated DNA with 0·3 % PEA causes a reduction in the rate of transformation. These observations suggest that PEA affects the ability of competent cells to irreversibly incorporate DNA and also affects the cells’ ability to express the DNA in the form of transformed cells once the DNA is irreversibly incorporated. The possible nature of the effect of PEA on competent cells is discussed.

Three strains of Butyrivibrio fibrisolvens were isolated from the rumen contents of cattle feeding on red (Trifolium pratense L.) or white (T. repens L.) clover. The substrates used in these isolations were plant hemicellulose fractions other than simple insoluble xylan. The strains showed some differences in their ability to grow on various plant polysaccharides and to secrete polysaccharases specific to these polymers. The same type of rumen contents yielded, on polygalacturonic acid media, a strain of Lachnospira multiparus which grew only on pectin and secreted as sole polysaccharase a polygalacturonase. Only one of the three B. fibrisolvens strains grew vigorously on polygalacturonic acid and its polygalacturonase appeared to be different to that of the L. multiparus.

The properties of a temperature-sensitive mutant of Bacillus subtilis in which DNA synthesis is specifically and reversibly inhibited above 40° are described. When incubated at 45° there was no detectable breakdown of DNA in the mutant and the synthesis of deoxynucleoside triphosphates was normal. The temperature-sensitive element affects DNA synthesis in vivo but not in vitro and (unlike an initiator) it appears to be needed for DNA synthesis throughout the replication cycle.

Leakage of 260 mμ-absorbing cell constituents from Pseudomonas aeruginosa was enhanced with increasing concentrations of Polysorbate 80 up to a characteristic maximum at 0·125 %. This relationship applied at 4°, 18° or 37° at bacterial concentrations between 1 × 108 and 9 × 108/ml. and for three strains of this organism, but did not apply with Escherichia coli. Uptake of a fluorescent dye was also greatest at 0·125 % Polysorbate 80. Increasing concentration of NaCl in the suspending liquid progressively decreased leakage. In buffer between pH 5·7 and 9·5, the low degree of leakage varied insignificantly. Polysorbate-treated organisms consistently leaked more readily and also showed greater percentage viability losses on rapidly changing the pH, temperature or NaCl concentration of the suspending solution. There was little correlation between loss of viability and leakage. There was a correlation between loss of viability and the magnitude of the sudden change in any of the colligative properties of the organism’s environment. Uptake of Polysorbate from aqueous solution at 20° was rapid. These results supported the hypothesis that Polysorbate 80 had an immediate effect on the permeability barrier of the cell.

Membranes from Sarcina flava were solubilized by using 1 % (v/v) Lubrol L and a water-soluble detergent-rich carotenoid glycoprotein complex isolated and purified. Homogeneity of the complex was indicated on the ultracentrifuge but molecular weight determinations made on a membrane osmometer indicated that the molecular weight values obtained in the presence of detergent may depend on the presence or absence of salts.

Using lysates and purified DNA, base ratios were determined by the buoyant density ultracentrifugation and the absorbance-temperature transition techniques ; the correspondence was very high between the two methods. The DNAs from a large number of Erwinia species, mostly phytopathogenic and nonphytopathogenic, but including some ‘Erwinia-like’ bacteria from animal sources and from other genera of the family Enterobacteriaceae were studied. The DNA base ratios of these bacteria fell mainly in the range 50–58 % GC, consistent with their taxonomic placement in the family Enterobacteriaceae. Certain assemblages of erwinias could be grouped on the basis of GC content into clearly differentiated clusters, which, accidentally or otherwise, correlated neatly with certain existing nomenclatural groupings. It was not possible, from the GC values alone, to decide whether the present multiplicity of Erwinia species can be justified as separate species, or whether and how they might be combined into fewer species, or whether and how they might be demoted to some infraspecific level. Indeed, it is not clear whether the genus Erwinia, defined as it is solely on the basis of plant habitat, should be maintained or whether its species might not, just as rationally, be distributed throughout other genera of the family Enterobacteriaceae. Classifications of the genus Erwinia on over-all phenotypic similarity result in a small number of specific taxa which, on the basis of the present study, are seen to contain organisms of different GC contents. In the present state of the art, one can only be agnostic about the amount of such differences in GC content which corresponds to separate taxa or to a particular categoreal level (generic, specific, infraspecific) in the present hierarchical taxonomic scheme.