- Volume 10, Issue 2, 1954

SUMMARY: The speed of development of narrow zones of opacity due to massed growth of leptospira a few millimetres below the surface in cultures in soft agar, a phenomenon first fully described by Dinger (1932), has been used as a criterion of the nutritional value of media for certain pathogenic leptospira. In applying this test to assess the value of the addition of peptone to a basal medium of 10% (v/v) rabbit serum in diluted meat extract, the impression was formed that peptones contained substances which promoted growth and others which were inhibitory. Witte’s peptone was the most constantly favourable of those which were tested. Tryptic digest broth (Hartley’s) gave results at least as good. Inhibitory effects with peptone may develop on autoclaving, especially at high pH values, or on leaving sterilized solutions to stand for considerable periods at room temperature. The value of haemoglobin (as laked blood) may depend on its catalase content since Czeka-lowski, McLeod & Rodican (1953) showed that leptospira did not produce this enzyme. The range of pH suitable for the development of cultures is narrow with pH 7·6 as optimum; but the leptospira are more tolerant to divergences on the alkaline side in the media at the time of inoculation.

SUMMARY: Hyaluronidase is formed as a constitutive enzyme by Staphylococcus aureus growing in either broth or casein hydrolysate media. The lag in the appearance of hyaluronidase activity after inoculation of a culture is longer than the lag in growth. A change in the rate of formation of hyaluronidase therefore occurs after exponential growth has been established. Activity per unit volume of culture supernatant increases exponentially faster after this initial lag than the mass of bacteria; it stops increasing abruptly sooner than growth. Coagulase appears, after inoculation, without lag additional to that in growth but increases at a slower rate than growth until it too ceases to increase before growth stops. Evidence is presented to show that changes in enzymic activity of the culture represent enzyme formation and not secondary changes in formed protein or secretion phenomena. The changes in rate of enzyme formation are connected with processes controlling enzymic specificity of the proteins rather than with general processes involved in manufacturing extracellular protein.

SUMMARY: Certain single-gene acetate-requiring mutants of Neurospora crassa (ac) are deficient in their ability to oxidize pyruvate directly, but can decarboxylate pyruvate to acetaldehyde and can form ethanol. The growth characteristics of the ac mutants indicate that they can form acetate from glucose or ethanol. ac Mutants are inhibited by glucose or sucrose. This glucose inhibition is relieved by the single-gene suppressor mutations sp and car which lower the activity of pyruvic carboxylase, decrease the amount of ethanol formed and permit some growth of ac sp and ac car strains in absence of added acetate. The sp mutation also lowers ethanol dehydrogenase activity, but this activity can be partially restored by growth of sp strains in the presence of ethanol. As a result of the nearly complete block in their pyruvate metabolism, ac sp strains are forced to metabolize glucose by pathways which produce more of the initial respiratory CO2 from the C-1 carbon of glucose.

SUMMARY: Radiopenicillin is strongly bound by ultra-microscopic lipid-containing particles liberated on mechanical rupture of Staphylococcus aureus cells. The binding resembles that of intact cells in that it is irreversible and only occurs to a limited extent, but differs in that 7–12 times as much penicillin is bound per unit dry weight of material. The supernatant after centrifuging down the lipid particles decreases the titre of added penicillin as indicated by diffusion assay, possibly by a small irreversible inactivation superimposed upon a ‘reversible’ type of binding.

There is a correlation between the distribution of (a) the penicillin-binding component (p.b.c.), (b) 35S from radiopenicillin pretreated cells, and (c) lipid phosphorus, in the three fractions produced on rupture either in distilled water or in a formaldehyde solution. Rupture in formalin appears to allow the cells walls to retain most of the lipid particles and p.b.c. but only a little extra of the dry weight of the cells. Thus penicillin reacts with a lipid-containing fraction close to the cell wall in intact organisms. At least as much more p.b.c. is liberated on rupture of the cells as was available to the penicillin in the intact cell, but p.b.c. is somewhat unstable after rupture. These data are discussed in the light of evidence in the literature that penicillin may react initially with the osmotic barrier of bacteria.

SUMMARY: Staphylococcal bacteriophage K was concentrated and purified from broth lysates without the use of high-speed centrifugation, which inactivates this phage. After precipitation with 30 % (v/v) acetone in water in the cold, the phage was further purified by treatment with isobutanol and chloroform, and concentrated by pressure filtration in reinforced collodion sacs. Electron microscopical examination revealed the presence of empty phage heads and tails, in addition to intact phage in the purified concentrate. Experiments indicate that while these entities could be made by disintegration of the phage, they were already present in this form in the original lysate. Study of the structure of the phage indicates the presence of a removable solid rod or plug within the tail which terminates in the terminal bob. These findings are discussed in relation to the structure of other phages, and their physiology.

SUMMARY: The isolation, morphology, cultural characteristics and growth requirements of an autotroph which oxidizes thiocyanate are described. It is suggested that the organism be assigned to the genus Thiobacillus.

SUMMARY: Leuconostoc mesenteroides P60 (Streptococcus equinus P60) requires serine for growth under normal conditions. On a medium containing relatively high concentrations of glycine it can dispense with serine provided that (a) pyridoxal is present, and (b) either Leucovorin (folinic acid) is added or the atmosphere is enriched with CO2. Increasing concentrations of CO2 or Leucovorin decrease the concentration of glycine required to support growth without added serine. Pyridoxal is not necessary for optimal growth with added serine, but the required concentration of the latter is twice as great.

Higher concentrations of p-aminobenzoic acid or Leucovorin are required for growth on glycine than on serine. Leucovorin replaces p-aminobenzoic acid for growth on serine, and both this factor and CO2 for growth on glycine. With the basal medium used (containing purines and all other amino-acids) thymidine supports growth in the absence of p-aminobenzoic acid only when serine is added.

Growth is not inhibited by sulphonamide when Leucovorin is present; with p-aminobenzoic acid there is the usual competition. Pteroylglutamic acid is inactive with this organism.

A valid assay for serine is possible, even with high concentrations of glycine present, when precautions are taken to destroy either Leucovorin or pyridoxal in the samples, and to prevent the atmosphere becoming enriched with CO2.

SUMMARY: When 0·01% chymotrypsin is added to mixtures of rhizobium bacteriophage and bacteria in liquid cultures, the multiplication of phage is prevented and the phage gradually becomes inactive. The rate and extent to which phage and bacteria combine are unaffected by the chymotrypsin whose effect seems directed against an early stage in the interaction between the two. This stage persists, on average, for less than 1 min. from the moment of combination. A phage particle combined with a bacterium becomes inactive and the bacterium is thus protected against lysis and remains able to multiply. Chymotrypsin does not interfere with combination between phage and bacteria killed by ultraviolet radiation: the combination leads to loss of phage activity irrespective of the presence of chymotrypsin. The multiplication of the phage is unaffected by previous incubation of either the phage or of the host bacteria separately with 0·01% chymotrypsin.

Chymotrypsin has no effect on phage-host interaction in an agar medium.

SUMMARY: The tovarious stages of the reproductive cycle in freshly transplanted cultures of avian tubercle bacilli were recorded by a series of successive electron micrographs. An early stage of cytoplasmic differentiation is described in which discrete cellular units form inside the elongating filaments. Their significance and relation to the vacuolization seen at later stages is discussed. Evidence is produced to show that the dense bodies of mycobacteria are not nuclei. Free granules do not appear to play any part in the reproductive cycle in the normal type of development. In mycelial variants lysis is often seen to initiate new growth which starts from submicroscopical elements and micro-rods.

SUMMARY: Mechanically disrupted cells of polymyxin E sensitive strains of Pseudomonas denitrificans and Escherichia coli and a resistant strain of Staphylococcus aureus were studied in the electron microscope. The occurrence of a secondary layer or membrane was observed within the outer cell walls derived from Ps. denitrificans and E. coli; a similar structure was not observed in cell walls prepared from Staph. aureus. Treatment of the disrupted cells of Ps. denitrificans and E. coli with polymyxin E resulted in marked morphological changes, whilst similar exposure of Staph. aureus cell walls to the antibiotic produced no significant changes in their morphology.

Penicillin-sensitive and penicillin-resistant colonies of Staphylococcus aureus, belonging to the same phage pattern, were found in six out of 200 apparently resistant cultures isolated from infective processes. The in vitro instability of peni-cillinase-producing strains of Staph. aureus was confirmed. It was found that growth with Streptococcus haemolyticus or in the presence of other antibiotics did not greatly influence the natural development of sensitive variants. This process, however, tended to be accelerated by growth at 44°, although some strains did not yield any sensitive variants in spite of prolonged incubation. Some inherited factor is considered to be primarily responsible for the production of penicillinase.

The cell wall of a bacillus, related to Bacillus anthracis, reacts specifically with the homologous antibody. This reaction can be made clearly visible by phase contrast when cell wall and cytoplasmic membrane are separated by mild lysozyme digestion, or when the cytoplasm is partially autolysed or digested with trypsin. The same antibody reacts with the transverse septa and polar bodies of the capsule, rendering them visible by phase contrast. These septa are in continuation of the cross walls of the cell, and sometimes give the appearance of division.

By these means the morphological locations of the chemically isolable fractions of serologically active bacterial extracts can be defined. These substances in the cell wall and in the septa and polar bodies of the capsule appear to be polysaccharide. The d-glutamic acid polypeptide element of the capsule does not occur in the cell wall.

A non-specific cell-wall reaction can be obtained with proteins at the appropriate values of pH, analogous to the non-specific capsular reaction previously reported.

The flagella of certain large spirilla appear to be compound structures composed of a large number of individual fibres. These arise in bundles from a single blepharoplast, whereas in other bacteria each flagellum arises separately. The flagella of spirilla are thus, in some respects, intermediate between those of other bacteria and of flagellates.

Saccharomyces rouxii is shown to be tolerant of a wide range of acidities and of sugar and salt concentrations, but sensitive to temperatures above 35° unless protected by a small quantity of sugar in the medium.

A limited investigation into the vitamin nutrition and nitrogen requirements of the yeast is reported.

SUMMARY: Seven hundred and thirty-six strains of Streptococcus pyogenes were examined for production of glucuronidase. The types most commonly found producing the enzyme were 9 and 28. Among other types there appeared to be some inverse relation between production of glucuronidase and of the M antigen.

SUMMARY: Cytological studies of Oscillospira guilliermondi from the caecal contents of guinea-pigs show that this organism consists of trichomes with numerous disklike cells containing nuclei in the form of paired rods. They offer a peculiarly clear illustration of this typically eubacterial form of nucleus.