- Volume 27, Issue 2, 1962

SUMMARY: Bacteriophage øR was examined in the electron microscope by the metal shadowing and negative-contrast techniques. The particle was a very small icosahedron and appeared to possess an extremely small tail. Some information was also obtained about the structure of the protein coat.

SUMMARY: Bacteriophage øR was prepared and purified in sufficient quantity to determine the composition and base ratios of its nucleic acid. The ratios are such that the complementary double-stranded structure usually found in deoxyribonucleic acid (DNA) seems unlikely to be present. This observation and the accessibility of the DNA of phage øR to the action of formaldehyde support the view that phage øR, like phage øX-174 which it resembles, carries its nucleic acid in a single strand.

SUMMARY: The sensitivity to antibiotics and the reactions to antibiotic-producing bacteria, streptomyces and fungi of a range of bacteria isolated from soil, leaves and forest litter showed broad differences between the Gram-positive and Gram-negative organisms, the fermentative and non-fermentative ones, and, to a lesser extent, between those of leaf origin and those of soil origin. These differences are not alone sufficient to explain the differences between the leaf flora and the soil flora. Other factors, including the capacity to produce antibiotics, are also important.

SUMMARY: Mutant strains of Aspergillus nidulans requiring methionine for growth were isolated and their growth responses to inorganic and organic sulphur sources studied. It is suggested that methionine is synthesized in this mould from inorganic sulphate through sulphite, thiosulphate and cysteine. The mutant strains genetically blocked in the reaction from thiosulphate to cysteine could be divided into two genetically different groups by means of the heterokaryosis test; heterokaryons formed between these two groups were able to grow on sulphate as sole sulphur source because of the syntrophic action of two different types of nuclei. It was further shown that cysteine-S-sulphonate supported good growth of one of the two groups, whereas the other group showed no growth on this compound. It was concluded that the metabolic conversion of thiosulphate to cysteine involves cysteine-S-sulphonate as an intermediate. Mutant strains responded to sulphide in the same way as to thiosulphate. From this and other evidence, it is suggested that sulphide is utilized by way of thiosulphate.

SUMMARY: A rapid centrifugation method for purifying foot-and-mouth disease virus (FMDV) combined an isodensity separation below a moving zone separation in the same tube. Two ml. of crude infectious fluid were introduced over a 3 ml. nonlinear CsCl gradient in a swinging bucket rotor. This preformed gradient provided a gradual increase from density 1.0 to 1.3 g./ml. and a step from 1.3 to 1.6 g./ml. After centrifugation for 4 hr. at 37,000 rev./min. (120,000 g) and 4°, a 1 mm. wide light scattering zone was observed with type A virus near the bottom, clearly separated below debris extending from density 1.3 to the meniscus. The narrow light scattering zone contained (40 ± 12%) of the FMDV infectivity and its CsCl isodensity was 1.43 ± 0.01 g./ml. Southern bean mosaic virus and bacteriophage øX174 behaved similarly and were useful as density markers. Virus suspensions concentrated an average of 8 fold retained (47 ± 16) % of their infectious units and were studied in the analytical ultracentrifuge directly. Dialysed concentrated virus revealed characteristic particles in an electron microscope. Exposure to concentrated impure CsCl decreases the stability of the infectivity to such an extent that in 40% CsCl its half-life is about 4 days.

SUMMARY: Samples of a liquid culture of Proteus L9 were transferred to agar blocks containing the same medium as the liquid culture. Photomicrographs were made of the slide cultures thus obtained, and the diameters of a large number of individual L elements determined. The measurements indicated that the elements of the L cultures studied could be assigned to two classes (I and II) with regard to their size. The elements of class I had an average diameter of 0.33 μ (standard deviation 0.071; standard error of the mean 0.006 μ); the corresponding figures for class II were 0.94, 0.21 and 0.02 μ, respectively. The growth of individual L elements in the slide cultures was followed during incubation at 30°. Only elements having a diameter > 0.6 to 0.7 μ, i.e. elements belonging to class II, enlarged measurably or formed microcolonies. Calculations based on measurements made on L elements growing on streak plates confirmed these observations.

SUMMARY: The antibiotic powers of some common soil and leaf bacteria, streptomyces and moulds were tested against soil and leaf yeasts. Twelve of the seventeen bacteria used, ten of the eleven streptomyces strains and seven of the twenty-two moulds were inhibitory. Pseudomonas chlororaphis, Aeromonas sp., five streptomyces and Trichoderma viride inhibited 70% or more of the twenty-five yeast species used. The most frequently isolated soil yeasts were amongst those most sensitive to antibiotics. Only one species of leaf yeast but several soil yeasts were inhibited by leaf bacteria and moulds. As streptomyces do not occur on leaves, it seems that the wide seasonal fluctuations in kinds and numbers of yeasts on leaves are due rather to nutritional and physical factors than to antibiotic ones.

SUMMARY: Hitherto only limited multiplication of Mycobacterium lepraemurium has been obtained in cell cultures; in cultures of rat fibroblasts (strain 14 pf) used by Garbutt, Rees & Barr (1958) growth of bacilli was limited to one or two generations. The present work shows that more continuous intracellular growth of M. lepraemurium can be achieved in cultures of rat fibroblasts by repeatedly subculturing the infected cells. The results suggest that multiplication of the bacteria is maintained only when a high proportion (50-75%) of the infected cells are transferred at each subculture. In one experiment, continued for 156 days, the increase in the number of bacteria was equivalent to 8 generations and the bacteria recovered from the cells were still infectious for mice. Quantitative electron microscopy also was used to follow the viability of bacteria from the cell cultures.

SUMMARY: Salmonella typhimurium will grow in simple defined media containing proline or glutamate as sole carbon source; in these media the organisms grew at 0.7 and 0.9 doublings/hr., respectively. Since most of the proline fed to the organisms is converted endogenously to cell material via glutamate, an attempt was made to see what reaction limited growth when the organisms were growing in proline as sole carbon source. The results suggest that the step which controls the conversion of proline to glutamate endogenously is the rate-limiting step. The uptake of proline from the medium to the free amino-acid pool did not appear to be rate limiting under these conditions. There was no evidence that the need to synthesize additional enzyme proteins (the proline degradative pathway) was responsible for the decreased growth rate of organisms growing on proline alone.

SUMMARY: Salmonella typhimurium was grown in a number of media containing compounds related either to the tricarboxylic acid (Krebs) cycle or to glucose metabolism. Organisms grown on compounds related to the Krebs cycle are pink when looked at by reflected light, whereas those grown on compounds related to glucose are white. Organisms growing in media containing compounds from one group continue to grow when transferred to media containing another compound of the same group, but not when transferred to media containing a compound from the other group. Growth of organisms in a mixture of one compound from each group leads to the formation of white organisms, i.e. the glucose-like state appears to predominate.

SUMMARY: When incubated in tissue-culture medium, inclusion blennorrhoea virus lost infectivity for HeLa cells at 37° but not at 30°. The rate of adsorption of virus to HeLa cell monolayers was dependent on temperature and on the volume of the inoculum. When the volume of medium was minimal, the virus was adsorbed before a detectable proportion was inactivated. Adsorption was complete after 7-8 hr. at 30° and 5-6 hr. at 37°.

At 30° the intracellular virus went into eclipse and maturation was retarded. When the temperature was raised to 37° maturation was rapid so that, for practical purposes, replication was synchronized during this period. In singly infected HeLa cells, at 37° infective virus was not detected for 22-23 hr., after which the progeny increased exponentially until at 34-38 hr. 35-60 infectious particles/infected cell were formed. The total number of particles/inclusion seen in Giemsastained preparations exceeded the number of infectious units/inclusion.

After 42-48 hr. infective virus was found in the supernatant and the number of intracellular infectious units began to decrease. Unlike vaccinia, another large virus, the virus progeny did not directly infect adjacent cells.

SUMMARY: A strain of Pseudomonas aeruginosa was obtained which was able to grow on acetamide or propionamide as sole source of carbon and nitrogen. When grown on these amides, whole bacteria and cell-free extracts rapidly hydrolysed acetamide, glycollamide, acrylamide and propionamide and slowly hydrolysed formamide and butyramide. N-Methylformamide, N-methylacetamide, N-ethylacetamide, N-acetylacetamide, N-methylpropionamide, N-ethylpropionamide, lactamide and methyl carbamate were found to be non-substrate inducers of the amidase when the organism was grown in succinate + ammonium chloride medium. N-Methylformamide, N-methylacetamide, lactamide and methyl carbamate did not inhibit propionamide hydrolysis by whole bacteria, but under the same conditions glycine amide, iodoacetamide and urea were effective inhibitors of amidase activity. N-Phenylacetamide, cyanoacetamide, glycine amide, sarcosine amide, β-hydroxy-propionamide and thioacetamide were neither substrates nor inducers of the amidase in this strain, but inhibited amidase induction by N-methylacetamide in succinate + ammonium chloride medium. Formamide also inhibited amidase induction under the same conditions.

SUMMARY: Fifteen strains of Haemophilus influenzae were found to require for growth, in addition to coenzyme 1 (diphosphopyridine nucleotide; DPN) and haematin, the following substances: pantothenic acid, thiamine, uracil. Some of the strains also required a purine, accepting xanthine, hypoxanthine or guanine, but not adenine. Cysteine (or glutathione) was also needed for luxuriant growth. A medium is described which yields crops of about 1010 organisms/ml. after incubation for 18 hr. at 34°. Sheep red cells, but not horse red cells, contain a DPN-ase, located in the stroma, which rapidly destroys any DPN added to them and also destroys the contained DPN when the cells are lysed.

SUMMARY: Bordetella pertussis grew profusely and retained its normal minute coccobacillary form when grown under 0.4 mm. of fluid lying on a Cohen & Wheeler agar medium + charcoal. Microbes grown in this way were fully antigenic in terms of the mouse protection test. It was easy to obtain harvests of 1.2 x 1011 organisms/ml.

SUMMARY: In the fluid culture of Streptococcus pneumoniue in glucose + serum broth the pa.rt played by the serum appeared to be solely the regulation of pH value, and may be substituted by a suitable amount of sodium carbonate. The reason for the autolysis experienced in the fluid culture of pneumococci would appear to be due to hydrogen peroxide production. The addition of cysteine to a glucose + carbonate broth precluded the autolytic effect either by the cysteine giving an anaerobic condition of growth or by destroying H2O2.

SUMMARY: An inositol-dependent yeast Kloeckera brevis was grown in a medium limited in inositol and the fate of the inositol determined. All of the inositol was taken in by the yeast and combined in an organic form. Fractionation of the yeast after disruption in a Hughes press showed that 31-38% of the combined inositol was soluble in trichloroacetic acid solution and 43-65% was soluble in neutral or acidified lipid solvents. The trichloroacetic acid extract contained three inositol components; one minor one was identified as inositol monophosphate. The major component was a neutral derivative of mesoinositol which was not identified as any known naturally-occurring form of combined inositol. The lipid extracted with neutral solvents contained an inositide whose properties were those of phosphatidyl inositol. The lipid inositol extracted with acidified solvents appeared to be present in a lipoprotein.

SUMMARY: Four strains of Leptospira were preserved by drying, in vacuo, small volumes (0.04 ml.) of suspension of the organism from the liquid state. The leptospires were suspended and dried in a mixture of equal parts of Korthof’s medium and 20% (w/v) glucose. Recoveries of the organisms after storage for one year were appreciably higher from desiccates stored at 4° than from those stored at about 20°.

SUMMARY: Alteration of the tonicity of a modified Edward medium by means of NaCl or other solutes resulted in very marked effects on the growth of several Mycoplasma strains. With both fluid and solid media, all grew best at about 10 atmospheres osmotic pressure (water activity, aw , 0.9925). The most exacting species, Mycoplasma gallisepticum, failed to grow outside the range 6.8-14 atmospheres (aw 0.995-0.990), while the least exacting, M. laidlawii, multiplied at up to 27 atmospheres (aw 0.980. Osmotic requirements were not appreciably altered by serial growth in hypertonic or hypotonic media but were, to some extent, conditioned by the serum content of the medium.