- Volume 2, Issue 3, 1948

SUMMARY: Forty-nine strains of vibrio were isolated from 32 of 43 samples of fresh water collected in various parts of England and Wales during the period October 1945 to July 1946. More than one kind of vibrio were found in 14 of the samples.

The types of vibrio isolated were heterogeneous in their fermentation reactions. Only three gave a cholera-red reaction. Twenty-five produced haemolysin for goat erythrocytes and all the haemolysins tested were filterable through gradacol membranes of 75 mμ pore diameter; an antiserum prepared against the haemolysin of one strain neutralized those of the other haemolytic water strains and of an El Tor vibrio.

Only one serological group of five strains related in ‘O’ and ‘H’ antigens was found; a few other strains showed H relationship and one (or two) possessed the H antigen of Vibrio cholerae; otherwise the organisms isolated were serologically heterogeneous with little antigenic similarity to one another or to known Asiatic vibrio types.

Summary: There is an apparent cycle of nuclear reorganization in Lactobacillus sp. Nuclear fusion occurs between the units of a multicellular bacillus. The organism then increases in length, the nucleoplasm being redistributed throughout the resulting filament, which by fragmentation returns to the bacillary condition. A very similar cycle is seen in Streptococcus faecalis.

Summary: Variation in the Actinomyces coelicolor species-group comprises loss of pigment and aerial mycelium, and occasionally of agar liquefaction. Stable variants may arise from degenerate, aged, vegetative mycelium, but do not normally do so when the vegetative mycelium is kept in vigorous condition by frequent subcultivation in suitable media. Single spore isolations from the aerial mycelium of typical and of variant colonies show that there may be inherent differences in the sister-spores of the same chain. Thus, in an agar-liquefying strain 3 of 15 spores had lost the power to produce pigment and to liquefy agar; and an atypical colony of the same strain yielded three viable isolates each with a strong tendency towards sectoring, with the ultimate production of a colourless, non-agar-liquefying variant as well as the typical growth. A non-agar-liquefying strain, which by prolonged cultivation in the vegetative phase had lost its power of producing the red-blue indicator pigment, yielded a variant giving rise to sectored colonies with occasional restoration of the blue pigment. Spontaneous occurrence of variants may be detected in certain spores of the aerial mycelium of a well-grown typical colony, although it is more readily seen in the spores of degenerate colonies which have been rendered atypical by artificial methods of cultivation.

Summary: The change in the Gram staining reaction which occurs when heat-killed Gram-positive Clostridium welchii and Staphylococcus albus are incubated with lysozyme is due to the removal of the ribonucleic acid component of the Gram complex, and is brought about by the hydrolysis of certain sugar linkages in polysaccharides located at the cell surface.

Summary: The production of filamentous forms of Clostridium welchii which occurs in complex media containing certain commercial and chemically treated peptones is due to a deficiency of ionized magnesium. Such filaments revert to cells of normal morphological appearance on subculture in a medium containing free magnesium ions, but the change cannot be brought about by the presence of metallic ions other than magnesium. It is, therefore, suggested that the latter is essential for the activity of the cell-dividing mechanism.

The presence of a growth-inhibitory factor in certain peptones has been established. The active agent appears to be a fatty acid and may be extracted from acidified peptone solutions with ether or chloroform. The presence of the inhibitory substance in peptone markedly decreases the crop yield of Cl. welchii but has no direct influence on the production of filaments.

Summary: A commercially available powdered alumina is very suitable for breaking bacterial cells according to the method of Wiggert, Silverman, Utter & Werkman (1940). Extracts have been prepared from streptococci in this way, which are capable of glycolysis, the deamination of adenosine diphosphate, and the production of NH3 from arginine.

Summary: The cultural characters of a strain of Lactobacillus arabinosus and L. helveticus used in the microbiological assay of riboflavin and nicotinic acid proved to be typical, excepting that neither strain fermented xylose and the strain of L. arabinosus fermented rhamnose but not raffinose. Neither produced catalase, so that a strongly positive catalase test in these cultures indicates probable contamination by common air-borne micro-organisms.

Modifications of the medium to achieve maximum acid production were investigated by altering, omitting or adding various constituents. A modified medium was adopted allowing a slightly higher acid production. In this medium the maximum acid production was obtained in the presence of natural substances like peptone. The maximal acid production obtainable with nicotinic acid alone was slightly lower, suggesting an additional specific factor in peptone. No confirmatory evidence, however, could be obtained of its existence. Linoleic acid in a concentration of 640 μg./10 ml. medium depressed the acid production, and its action was antagonized by cholesterol. Other fatty acids and lipids had no effect.

A good reproducibility and a small coefficient of variation was found between tubes at various levels of nicotinic acid within any one assay, but between separate assays the variation was high (coefficient of variation = 12–19%).

Summary: A modified basal medium was employed for the assay of nicotinic acid contained in various biological materials. Of the various possible comparisons of test material with standard nicotinic acid in the assay, the ‘internal standard’ method is preferable; by assaying the standard nicotinic acid in the presence of the test material, any discrepancies due to substances stimulating or inhibiting the nicotinic acid effect are minimized.

Of the different extraction procedures tested, treatment of samples with n-NaOH and n-H2SO4 gave the most satisfactory results; both n-NaOH and n-H2SO4 appeared to convert a ‘precursor’ in potato and bran, into nicotinic acid. 2n-HCl extracts yielded lower values, but the conversion of the precursor was less effective with potato than with bran, suggesting different precursors in the two vegetable substances.

The results of microbiological and chemical assay were in good agreement. The coefficient of variation of the microbiological results was of the order of 9%.

Summary: A number of different organisms were subjected to violent shaking with minute round glass particles. Vegetative bacteria, spores, and acid-fast species were killed by this treatment, though at varying rates.

SUMMARY: The nuclear changes leading to spore formation in Bacillus anthracis were investigated in 35 strains. Fusion of four chromatinic bodies into one mass and its subsequent break up into four bodies was observed. One of these became incorporated in the spore; the other three disintegrated. Some cells showed no fusion before spore formation; in these one chromatinic body was incorporated in the spore and the other three disintegrated. The percentage of cells in which fusion occurred varied with the strain. Rough colony strains showing a high incidence of fusion tended readily to throw smooth colony variants. Strains showing a low incidence of fusion were stable in this respect. The incidence of variants of other types bore no relation to the incidence of nuclear fusion in the strain.

SUMMARY: Among a large number of strains of aerobic sporing bacilli only Bacillus cereus, B. mycoides and, to a lesser extent, B. anthracis were able to produce turbidity and formation of a curd in saline extract of egg-yolk. None of the other species tested, namely, B. alvei, B. alcalophilus, B. brevis, B. carotarum, B. circulans, B. coagulans, B. fusiformis, B. licheniformis, B. macerans, B. megatherium, B. pumilus, B. polymyxa, B. orpheus, B. repens and B. subtilis caused any opalescence in the yolk medium. The yolk reaction was due to the action of phospholipinase produced by the organisms. As in the case of Cl. welchii α-toxin, the yolk curd-forming activity of B. cereus and B. mycoides also was associated with haemolytic activity. The substances responsible for these activities appear to be similar, and may be identical. The yolk reaction has proved useful for the rapid identification of B. cereus, as it is more specific than any of the other distinguishing tests hitherto employed. The three positive organisms, B. cereus, B. mycoides and B. anthracis, have been considered by previous workers on the grounds of morphology and antigenic structure to be closely related.

SUMMARY: Gladiolic acid is produced when Penicillium gladioli is grown on a wide range of culture media. The main factor influencing its production and accumulation is the pH drift of the medium, high yields being associated with a characteristic pH drift, consisting of an initial fall to about pH 4·0 followed by a steady but not too rapid rise. Continued low pH is unfavourable; too rapid a rise of pH is also unfavourable since gladiolic acid tends to disappear from the medium when pH 6·0 is reached, the disappearance being very rapid at pH 7·0 or above. The effects of variation of the initial pH of the medium, of glucose concentration, of variation of nitrogen source and of additions of certain organic acids are all explicable in terms of their effect on pH drift.

The antibiotic is best extracted from culture filtrates by treatment with activated charcoal after adjustment to pH 4·0, elution of the charcoal with ether, and re-crystallization from water after evaporation of the ether. Yields of the order of 300 mg./l. are obtained.

Gladiolic acid is highly fungistatic if tested at low pH; the toxic effect is due to the undissociated molecules only and at pH 7·0, when dissociation is virtually complete, gladiolic acid is almost inactive. At pH 3·5, the least concentration inhibiting germination of fungus spores varies from 0·9 µg./ml. for Fusarium graminearum to 250 µg./ml. for Trichoderma viride. It is not highly antibacterial in broth, many organisms growing freely in the presence of 500 µg./ml. This low activity is thought to be due in part to the dissociation of gladiolic acid at pH 7·0 and in part to inactivation by certain broth constituents. This view is supported by the observation that bacterial cells suspended in gladiolic acid solutions (100 µg./ml.) in buffer at pH 4·0 are rapidly killed. This bactericidal effect occurs with both Gram-positive and Gram-negative organisms.

Gladiolic acid in solution is relatively stable in the range pH 3·0–8·0. In the presence of ammonium salts or certain amino-acids, notably p-aminobenzoic acid, it is rapidly inactivated, coloured complexes being formed. The reaction with ammonium salts is dependent on pH, not proceeding at pH 3·5 but proceeding rapidly at pH 7·0. The rapid disappearance of gladiolic acid in culture when the pH rises above 6·0 is possibly associated with this type of reaction.