- Volume 36, Issue 1, 1964

Summary: A broad range of typing phages belonging to lytic groups I, II, III, IV and those classified as miscellaneous produced plaques on Staphylococcus aureus strain k1. Two variant forms of this strain were compared, k1n and k1hi. Phages belonging to lytic group II, of both A and B serology, had low efficiencies of plating (EOP) for the k1n variant but had high EOP values for the k1hi variant. In general, of phages from other lytic groups, those of B serology tended to plate with low EOP values on both variants, whereas those of A serology plated with high EOP values. Propagation of several of the phages on both k1 hosts resulted in host-modifications of the phage progeny. Similar host-induced modifications were produced by propagation of the phages on ps 73. The host-modified phages showed striking losses in plating efficiency for their usual propagating hosts. The significance of these findings with respect to genetic classification of the phages and hosts is discussed.

Summary: Propagated on a group II host, phage 3C behaves as a typical group II phage; but after passage on hosts of wide group patterns, Staphylococcus aureus k1 and 73, 3C phage is host-restricted, losing its potential for infecting group II hosts and gaining an increased potential for infecting group III hosts. Such a phage would be considered a group III phage. The change occurs in the first burst of every infected cell. The phage released retains its A serology. Phage 3C can be propagated directly on several group III cocci and in these hosts a similar host-restriction occurs. These observations constitute evidence that host-induced modifications can alter the group lytic spectrum of a specific typing phage.

Summary: The best labelling of Staphylococcus aureus (var. pyogenes) with 2P was achieved when the organisms were grown in broth containing 33 μg. ortho- phosphate/ml. and 150–200 μ c./ml. 2P. Cocci labelled in this way were as virulent to mice as control cocci. The phosphorus content of cocci was proportional to the orthophosphate content of the medium over the range 5 μg.-300 μg. PO4/ml. 2P was released from cocci when they were incubated with phosphate buffer and other media which did not support cell division. 2P was released from labelled dividing cocci grown in nonradioactive media. About 80% of the 2P was released during division; a maximum of about 55% was released from non-dividing cocci over a similar time period. Most of the 2P released from dividing cocci was present in the form of molecules non-diffusible in dialysis (80%); most of the 2P released into phosphate buffer was diffusible. Cocci deficient or rich in phosphorus were fractionated with trichloroacetic acid (TCA), ether, ethanol and perchloric acid. Cocci containing low amounts of P lacked 2P in the fraction soluble in cold TCA, about 80% of which was orthophosphate. Cocci deficient in phosphorus incorporated 2P from 2PO4 solutions more rapidly than enriched cocci. The distributions of 2P in cocci previously rich or deficient in P were similar, but labelling in deficient cocci was increased in the RNA-containing fraction; and that part of the fraction soluble in hot TCA which had a barium salt insoluble at pH 4. The phosphorus level in the cells had no effect on the amount of phosphorus released into phosphate buffer solution.

Summary: In this account of the fine structure of a nitrogen-fixing clover nodule the interactions between host and bacteria have been particularly considered. It is suggested that the bacterial infection is located between the paired membranes of the endoplasmic reticulum, i.e. within the intrareticular space. It is shown that the plant membrane around the infection thread is continuous with the plasmalemma, but that it is fragmentary around the thread tip. Bacteria emerging from the infection thread remain surrounded by a plant membrane, and their subsequent fate (division or rapid swelling) is controlled by the condition of the host cytoplasm. Structural changes occurring in the cytoplasm, nuclear region and wall of emerged bacteria during their transition to the bacteroid form are described, and the development of peripheral carbohydrate platelets in the host cell is illustrated. Supporting observations on two types of ineffective nodules are discussed.

Summary: Nutritional experiments were carried out dealing with: (1) the utilization of sulphur compounds; (2) the utilization of phosphorus compounds;

(3) the action of different mineral salts and their interaction. The results showed that Phytophthora erythroseptica grew well at 28° and pH 6·6 incubated for 20 days; P. parasitica grew well at 28° and pH 6·6 for 17 days; and P. infestans at 20° and pH 4·5 for 16 days. In the adjusted controlled medium pH changes were generally within one pH unit. The best carbon and nitrogen sources are stated. The only satisfactory sulphur sources for P. infestans were sodium sulphate and sodium thiosulphate. These compounds were also among the best S sources for P. erythroseptica and P. parasitica, but a number of other compounds also were utilized equally well, e.g. sodium metabisulphite, sodium sulphite, a-cysteine, sodium sulphide, sodium dithionite and methionine. For all organisms the best phosphorus sources were sodium dihydrogen orthophosphate, sodium metaphosphate and lecithin. Rate of utilization of the phosphorus was an important factor in mycelial yield. Factorial experiments were carried out in which P. erythroseptica and P. parasitica were incubated at 28° for 17 days. Statistical analysis of the results showed that under the given conditions optimal growth measured as mg. dry wt. was obtained in liquid media containing glucose, 25 g./l.; dl-asparagine, 4·0 g./l.; FeSO4.7H2O, 0·001 g./l.; thiamine, 0·8 mg./l.; ZnSO4.7H2O, 1 p.p.m.; H2MoO4, CuSO4. 5H2O, 0·02 p.p.m.; with varying amounts of K2HPO4, MgSO4.7H2O and CaSO4.2H2O for the different species specified. There was a balance between all combinations of K2HPO4, MgSO4.7H2O and CaSO4.2H2O for P. parasitica but not for P. erythroseptica, but there were significant interactions between the salts taken two at a time for both these fungi. There was interaction for all three salts together for both species. Phytophthora infestans and P. parasitica are more exacting in their nutritional requirements than P. erythroseptica.

Summary: Strain d of tobacco necrosis virus (TNV) was transmitted by zoospores of 3 different isolates of Olpidium brassicae (Wor.) Dang, to roots of Mung bean and lettuce grown in modified Hoagland’s solution diluted 1/20. On Mung bean roots necrotic local lesions formed one day after exposure to virus and zoospores. Virus in lettuce was assayed by inoculation to leaves of French bean. Virus transmission was favoured by decreasing salt concentration and increasing the pH value of the nutrient solution and depended also on the concentrations of virus and zoospores. With 105 zoospores/ml. transmission to lettuce was obtained with as little virus as 0·05 μg./l. When the virus concentration was 5 μg./l., 50–100 zoospores/ ml. were effective. Fungus infection as measured by the number of zoosporangia in the root was not strictly correlated with virus infection.

Exposure of roots to virus + zoospore mixture for 1 min. sufficed to infect them with virus. More transmission occurred when virus was added before or together with zoospores, than after. Roots, exposed to zoospores for 10 min., then washed, were more readily infected by TNV when virus was introduced during the first hour or two after zoospore attachment to the root cells than later; there was some transmission even when virus was withheld till 4 hr after washing. Immersing roots, inoculated with fungus and virus, in hot water (60°) killed the fungus but not the virus, and varying the interval between inoculation and heating showed that virus became established after 2–3 hr.

Isolates of Olpidium brassicae naturally contaminated by strain d or a of TNV were freed from contamination by inoculating lettuce roots with dilute zoospore suspensions. Zoospores mixed or naturally contaminated with TNV were partially separated from it by centrifugation. Virus transmission was prevented by adding concentrated homologous antiserum to zoospores that had already been exposed to virus, or by adding very dilute antiserum to virus before mixing it with zoospores. The extent to which transmission was prevented by antisera to other strains of TNV depended on the degree of their serological relationship to strain d. The present evidence does not support the suggestion that TNV is carried inside the fungus.

Qualitative and quantitative analyses of formamide extracts of group F streptococci revealed rhamnose, glucose, galactose, glucosamine and galactosamine as main components. The rhamnose content was high. Streptococci, carrying a type antigen in the cell wall, excreted into the medium also a polysaccharide with the same serological activity. These soluble polysaccharides contained no muramic acid, small amounts of rhamnose, but had a high glucosamine and mannose content. Column chromatography on DEAE-cellulose revealed the formamide extracts to be heterogeneous. Over 90 % of the serological activity could be recovered in one of the fractions. Inhibition reactions of the quantitative precipitation with a number of simple sugars gave indications about the composition of the determinant groups of group and type antigens. Rhamnose was inactive in all cases. The determinant group of the group F antigen is at least a disaccharide with a β-glucosidic moiety. The determinants of the type antigens I and II probably contain n-acetyl-galactosamine. There is evidence for a second determinant in type antigen II. The determinants of type antigens III and V contain a β-glucosidic moiety, that of antigen IV a β-galactosidic moiety.

Summary: Strains of two bacteria, a Pseudomonas and an Achromobacter, which grow with toluene, benzene or certain other related aromatic compounds as sole carbon source were isolated from soil. The use of aromatic compounds by these bacteria was an induced phenomenon. Toluene-grown organisms oxidized without lag toluene, benzene, catechol, 3-methyl- catechol, benzyl alcohol and, more slowly, o- and m-cresol, but not benzaldehyde or benzoic acid. 3-Methylcatechol, acetic acid, pyruvic acid, and a yellow ether-soluble acidic substance which was colourless in acid solution, were detected in toluene-oxidizing cultures. Acetic and pyruvic acids were also formed during the bacterial oxidation of 3-methylcatechol. 3-Methylcatechol is probably an early stage in the bacterial metabolism of toluene; benzaldehyde and benzoic acid seem not to be intermediates in this metabolism.

The inter-relationship between the intracellular concentration of potassium ions and the rate of oxidation of glutamate was investigated in washed Escherichia coli. The time-curve of glutamate oxidation by the potassium-depleted coli showed a marked lag phase and the rate of oxygen uptake increased concurrently with the accumulation of potassium ions. After a constant intracellular potassium concentration was reached, the rate of oxidation remained constant. Carbonyl cyanide m-chlorophenylhydrazone (m-Cl-CCP) and methylene blue inhibited the respiration when added to the reaction mixture during the initial phase of K+ accumulation. The extent of inhibition induced by these compounds was inversely related to the rate of oxidation prevailing at the time of their addition. No inhibition resulted when the substances were added after the K+ accumulation and respiratory rate had reached the steady state values. Pre-incubation with glucose and KCl abolished the initial lag of glutamate oxidation as well as the inhibitory action of m-Cl-CCP. It is concluded that the intracellular concentration rather than the flux of potassium ion governs the control of respiration in E. coli. The possible relation of the mode of respiratory inhibition induced by m-Cl-CCP and methylene blue to the known ability of these compounds to uncouple oxidative phosphorylation, is discussed in terms of the presumed energy requirements of the system mediating the K+ transport.

The biochemical and cultural characters of Micrococcus violagabriellae Castellani were examined, and the cultural conditions necessary for pigment production investigated; iron and manganese were essential for pigmentation. Some pigment-containing material was isolated and its simple properties were examined. Taxonomically, the organism might warrant sub-specific rank as Staphylococcus epidermidis (or S. sapro-phyticus) var. violagabriellae.

Summary: Molar growth yields were measured for Aerobacter aerogenes growing with a number of substrates as sole carbon and energy source in a minimal medium. Under anaerobic conditions the molar growth yield for glucose was 26·1 g. This amount of dry weight is produced at the expense of 2·55 mole of ATP (1·71 mole from glycolysis and 0·84 mole from acetate produced from pyruvate by the thioclastic reaction). The yield per mole ATP is thus 10·2 g., which value is very close to the one found for other micro-organisms. Under aerobic conditions the molar growth yield for glucose is 72·7 g. During growth 1·14 mole of O2 are taken up. The yield per atom oxygen is thus 31·9 g. By dividing the yield per atom oxygen by the yield per mole ATP we find the number of ATP mole formed per atom O. The values found are very close to 3, indicating that the efficiency of oxidative phosphorylation in this organism is the same as that in mitochondria. During the experiments it was observed that growth was maximal before the maximal O2 uptake was reached. The explanation is that during aerobic growth acetate accumulates, which is oxidized after maximal growth. Acetate oxidation after glucose consumption does not contribute to the dry weight.

When serological type strains of Streptococcus faecalis from different workers were compared with those of Sharpe & Shattock (1952) by using precipitin and reciprocal absorption tests, several of these types were found to possess the same type antigens. The location of these type antigens was in the cell wall and they were polysaccharide in nature. Some strains had two of these antigens. The distribution of S. faecalis serotypes is discussed and a typing scheme applicable to all recognized types of group D streptococci is suggested.