- Volume 48, Issue 2, 1967

SUMMARY: Organisms belonging to several genera of the order Actinomycetales were examined using the ‘Stereoscan’ electron microscope of the Cambridge Instrument Company. Unlike the transmission electron microscope, this instrument provides surface views of fine structures without the need for preparation of replicas. Using a simple preparation procedure, surface views of intact sporing structures of several actinomycetes were observed over a range of magnifications. The potential value of using scanning electron microscopes for the examination of actinomycetes was assessed.

SUMMARY: Serum-sensitive mutants have been derived from serum-resistant smooth virulent Salmonella typhimurium and S. enteritidis strains by selection for resistance to cephalosporin or penicillin. Chemical analyses of the lipopolysaccharides of these mutants reveal that they belong to at least three different rough or semi-rough classes. Partial or total loss from the lipopolysaccharide of the sugars responsible for O antigenicity resulted in loss of virulence, as well as increased sensitivity to the bactericidal effect of antibody plus complement. However, such loss is not necessary for serum sensitivity because two serum-sensitive mutants possessed lipopolysaccharides indistinguishable from the smooth serum-resistant parents and were nearly as virulent.

SUMMARY: Cultures of Escherichia coli b which had received pretreatment with dihydrostreptomycin were considered to consist of two subpopulations corresponding to the two fractions of culture samples that did and did not survive through plating and incubation to produce visible colonies. No initial assumptions were made about the viability of the subpopulations at the time that the samples were taken. A general-purpose analogue computer was then used both in mathematical analysis and mathematical synthesis to investigate the growth characteristics of the survivors and non-survivors as functions both of time and of pretreatment.

SUMMARY: Dwarf colony variants of Staphylococcus aureus have been described as a causative agent of bovine mastitis in Israel. These strains were reported as auxotrophic for thiamine, being unable to synthesize the vitamin from its pyrimidine and thiazole moieties. In the present paper it is shown that strains with two different deficiencies are involved. One type is unable to concentrate the thiazole moiety and the other is unable to phosphorylate the pyrimidine portion of the vitamin. The results indicate the existence of independent permeases in S. aureus for these thiamine moieties.

SUMMARY: The influence of casein hydrolysate, tryptophan, glucose, magnesium sulphate and different buffer solutions on the rate of multiplication of Bacillus subtilis strain 168 was determined. Inoculum size was shown to influence the generation time and the increase in the mass of bacteria was found to be inversely proportional to the size of the inoculum. Bacillus subtilis 168 forms chains especially in minimal media. In complete media chains are formed by only a small proportion of the bacteria and sometimes not at all.A simple transformation method was used to follow the development of competence in cultures of Bacillus subtilis 168 in media inoculated with different numbers of bacteria. The estimation of the number of transformants was subject to greater error than estimation of the viable count. This error in the estimation of the number of transformants is due mainly to the thread-like character of DNA, which is difficult to dilute accurately. The smaller the inoculum of bacteria, the shorter the time for the rise phase of competence. Optimal competence is reached when about 106 bacteria/ml. are inoculated into the medium. The rise phase of competence takes a shorter time than the decline phase.An equation was developed from which it is easy to calculate approximately the onset of the peak of competence. Precise determination of this peak can be achieved by controlling the density of the bacterial suspension.

SUMMARY: Isolated lipopolysaccharide from a rough mutant of Salmonella minnesota was shown to act as a receptor for Felix 0-1 bacteriophage. The isolated receptor structure could be dissociated by sodium deoxycholate with subsequent loss of receptor activity. When the deoxycholate was removed by dialysis the receptor activity was restored. Ultrasonic treatment destroyed the receptor activity and electron micrographs indicated that the ultrasonic treatment had altered the structure of the lipopolysaccharide. The experiments suggest that the inactivation of phages can be used as a test of the biological activity of a lipopolysaccharide. The interaction between the 0-1 phage and the receptor structure was also studied with the aid of electron microscopy.

SUMMARY: As conidia of Neurospora crassa aged, the activity of NADP-specific glutamic acid dehydrogenase (NADP-GDH) decreased to negligible values. Subsequent to this decrease, a significant increase occurred in the activity of the NAD-specific glutamic acid dehydrogenase (NAD-GDH). Incubation of aged conidia in basal medium resulted in over a 100-fold increase in NADP-GDH activity within 3 hr. Although no net protein synthesis occurred during these early stages of germination, a turnover in protein was observed. The data presented are consistent with de novo synthesis of NADP-GDH. Development of NADP-GDH activity was dependent upon an appropriate carbon source and pH value. An exogenous nitrogen source was not required. The data do not directly support reciprocal regulation of the synthesis of NAD-GDH and NADP-GDH in Neurospora as postulated by Sanwal & Lata (1962a, b).

SUMMARY: The cell-bound penicillinase of Bacillus licheniformis strain 749/c was examined since this material appears to be an intermediate in the formation of the exopenicillinase. The enzyme remained attached to fragments of the cell membrane during lysis of the cell. It was not released from these fragments by a variety of physical or chemical treatments (urea, hydroxylamine, deoxycholate, ultrasound, etc.). Free penicillinase, very similar if not identical, to the exoenzyme, was produced by treatment with trypsin, Streptomyces griseus proteinase or papain, but not by chymotrypsin, lipases or nucleases. The cell-bound enzyme appeared to be covalently linked to the cell membrane through a peptide chain. Trypsin probably cleaved this chain at a linkage involving the carboxyl group of lysine or arginine. The enzymic characteristics of the bound penicillinase are essentially equivalent to those of the exoenzyme. Partial purification of the membrane-bound enzyme is described.

SUMMARY: Organisms (logarithmic growth phase) of Bacillus licheniformis strain 749/C when washed and resuspended in buffer ceased to produce penicillinase but continued to release their cell-bound enzyme as typical exopenicillinase. In the first 1-2 hr, 15-30% of the bound enzyme was liberated; the process then became very slow. Liberation was dependent upon temperature and pH value; it was not inhibited by chloramphenicol. Release was apparently not a result of membrane damage, since there was no leakage of intracellular α-glucosidase. Lysis of the bacilli decreased sharply the rate of release of coenzyme. Membrane preparations active in releasing penicillinase were obtained by lysing the bacilli with lysozyme in the absence of added Mg2+ and subsequently adding 0.05 m-MgSO4 to prevent dispersal of the membrane segments. The liberation of penicillinase by this fraction had the same general enzymic properties as had the process with intact bacilli. Disruption of the membranes also eliminated most of the penicillinase-releasing activity. The importance of membrane structure in the liberation process is discussed.

SUMMARY: A series of fungal melanoproteins (mol. wt 10,000-70,000) was isolated from culture filtrates of Venturia inaequalis, and partially characterized by gel filtration and acid hydrolysis. Petiole injection of aqueous solutions of the melanoproteins with non-phytotoxic marker compounds, into apple shoots, produced specific effects on the transport of solutes within the leaves. The effects were not reproduced by substitution of melanoprotein by egg-white lysozyme, rabbit haemoglobin, bovine serum albumen or deoxyribonuclease (EC 3.1.4.5). Inhibition of leaf expansion by injected melanoproteins was observed.

The application of a melanoprotein with spore inoculum of Venturia inaequalis on to leaves of a susceptible apple variety caused a great increase in lesion development. This effect was not reproduced by the use of other proteins. The decreased lesion-stimulating activity of melanoprotein produced by V. inaequalis after storage of cultures at 0° was correlated with decreased effects on solute transport in the host. Radioactive material arising from spore inoculum labelled with [14C] dl-alanine was detected in the vascular system of test plants. The distribution of this material in the infected leaf was similar to that given by petiole injection of an indicator compound with melanoprotein in healthy plants.

SUMMARY: N-Methyl-N'-nitro-N-nitrosoguanidine (NTG), which is known to be a very effective mutagen in many systems, induces petite mutations when cells of Saccharomyces cerevisiae are treated in acetate buffer. Fifty per cent or more of the survivors may be petite mutants. NTG is a real mutagen in the process since the number of mutants increases at short times of exposure to NTG although the total number of cells decreases. Growing cells are more susceptible to killing and mutation than are cells in buffer. The production of petites may cause difficulties when screening for rare auxotrophs.

SUMMARY: Chlorophyll formation in non-growing etiolated cells of Euglena gracilis var. bacillaris is inhibited by certain antimetabolite analogues of purines, pyrimidines and amino acids. The inhibitory effects of bromo-and nitro-uracils were annulled by uracil and thymine; those of fluoro-and thio-urneils were not. Ethionine inhibition was completely annulled by methionine. All the base analogues were more inhibitory for the dark growth of the alga than in the light and an adaptation to these antimetabolites was apparent during growth. The growth inhibitory effects of some of these base analogues was annulled by several Krebs's cycle intermediates; the latter in themselves stimulated growth of photosynthesizing euglenas although they were not utilized in the dark with any degree of efficiency. Long-term exposure of non-proliferating euglenas, green or etiolated, to 5-fluoro-uracil had a profound effect on the chloroplast integrity of the organisms. Such euglenas on further subculture in an adequate growth medium were permanently bleached. The effect of 5-bromo-uracil, though similar, was less marked.

SUMMARY: The acetone-extractable antibacterial components of Isochrysis galbana have been resolved by paper-chromatography and bioautography. Two pigmented products with similar antibacterial properties were obtained which were tentatively identified from their absorption spectra as chlorophyll a derivatives, probably pheophytin a and an atypical chlorophyllide a.

SUMMARY: In several bacterial species the occurrence of competence in transformation is strictly related to the production of a competence factor (c.f.). In the present work this factor was isolated from the cell surface of Streptococcus group H strain challis by the centrifugation of competent bacteria at pH 2.0, followed by thermal shock. Streptococci of strain wicky adsorb c.f. isolated from the challis organisms because supernatant fluids containing c.f. lose their activity after having been in contact with organisms. It was possible to ‘tear off’ again from such organisms of strain wicky the c.f. they had adsorbed. The re-isolated c.f. was still active in the transformation systems used. Some biochemical properties of the c.f. from strain challis are described. The factor was relatively heat resistant and sensitive to proteolytic enzymes. Streptococcal c.f. or a fragment of it seems to be a protein or polypeptide of fairly low molecular weight. This streptococcal c.f. is probably different from the pneumococcal c.f.

SUMMARY: Pityrosporum ovale did not show the normal swelling and shrinking such as is undergone by Saccharomyces cerevisiae in media of different osmotic pressures. Pityrosporum ovale shrank slightly in media below 0.33 osmolar and remained at constant volume between 0.33 and 3.4 osmolar, while S. cerevisiae swelled below 0.10 osmolar and shrank markedly and progressively from 0.33 to 3.4 osmolar. The resistance of P. ovale to this effect is believed to be associated with its survival on the scalp in an environment low in water and high in salt from sweat, and to be a property of the remarkable construction of its cell wall as revealed by electron microscopy.

SUMMARY: The properties of a bacteriocin derived from Lactobacillus fermenti strain 466 were investigated. The bacteriocin was present in low titre in supernatant fluids from overnight broth cultures and was not inducible by ultraviolet radiation. It was purified and concentrated to a titre of 1/1000 by dialysis, chromatography on Sephadex G100 and calcium phosphate gel columns. The bacteriocin is heat stable, and sensitive to trypsin and pepsin but not to lysozyme. No migration was demonstrated in electrophoretic fields in agar gel. Electron microscopy of the bacteriocin did not show any phage components. The bacteriocin is a macromolecular lipocarbohydrate protein which consists of 16 amino acids, four sugars, hexosamine and phosphorus. The biological activity of this complex is dependent on its structural integrity.