- Volume 2, Issue 2, 1948

SUMMARY: Equipment can be readily made in the laboratory for the mechanical preparation of roll-tubes. Roll-tubes so prepared can largely replace Petri dishes both for counting and for single colony isolations.

SUMMARY: The long-chained and short-chained variants of Streptococcus pyogenes, Str. viridans and Str. faecalis possess a structure closely resembling that of rough and smooth bacillary variants. Cocci of the long-chained strains are frequently divided by transverse cell walls into two cells, each containing a single chromatinic body. Short-chained strains fail to form transverse cell walls, the cocci dividing by constriction; they are unicellular and usually contain a pair of chromatinic bodies. Analogous types occur in Staphylococcus aureus and Staph. albus. In one of these types the cells contained a well-defined central granule, dividing with the coccus.

SUMMARY: The growth of Escherichia coli strain B/r on nutrient agar is the same as in broth with regard to the initial lag period and the rate of increase during the exponential phase. On nutrient agar, growth is finally slowed down either when individual colonies contain more than 219 bacteria, or when the total number of bacteria/4 in. plate exceeds 1010.

The rate of mutation to phage resistance was measured by a modification of Demerec’s method of spraying plate cultures with a phage aerosol, and counting resistant survivals. With the phage T1, after twelve bacterial generations on nutrient agar at 37°, the value of 0·74 ± 0·04 mutations/108 bacterial divisions was obtained, and after eighteen bacterial generations, 0·70 ± 0·04.

Resistance to T1 phage is probably brought about by at least five separate mutations, as shown by tests of resistance to other phages. Different strains of E. coli yield different proportions of some of these mutants. Thus strain B/r yielded a lower proportion of the mutant B/1 than did strain B. Furthermore, as different strains of T1 phage may contain varying amounts of the phage mutant T1h, capable of lysing the bacterial mutant B/1, but not B/1, 5, variations in the relative proportions of B/1 and B/1, 5 may also occur when different samples of phage are used. Consequently, constancyof mutation rate can only be guaranteed for given cultures of both bacteria and phage.

SUMMARY: An inhibitor of plant viruses can be isolated from the sap of Phytolacca esculenta by differential precipitation with ethanol followed by adsorption on celite and elution with 10% NaCl. Purified preparations contain 14–15% nitrogen and 8–12% carbohydrate and the inhibitor is probably a glycoprotein. Denaturation leads to loss of inhibiting power. The protein, unless denatured, is unaffected by pepsin and trypsin.

The glycoprotein is isoelectric at about pH 7. It can combine with tobacco mosaic virus, and when salt-free solutions of the two are mixed in certain proportions at pH values between their isoelectric points it precipitates the virus in the form of paracrystalline threads. The glycoprotein also precipitates tomato bushy stunt virus.

When added to several plant viruses, the glycoprotein causes an immediate reduction in infectivity, but has no effect on a bacteriophage. Non-infective mixtures regain infectivity when diluted. No evidence was found for a combining ratio of virus to inhibitor necessary to cause loss of infectivity. The mechanism of virus neutralization is discussed.

Summary: Bacteriophages were isolated from several strains of Pseudomonas pyocyanea and were assayed against an indicator strain. Counts of up to 1010/ml. were obtained in both broth and a denned medium. The phages are stable at 60° but rapidly inactivated at 65°. They are unaffected by 50 % glycerol and are stable at pH 6·5–10·0 but not below 6·5.

About 500 compounds have been tested for their inhibitory action on the development of these bacteriophages. All the sulphonamides, amidines, pyrimidines, organo-metallic compounds, plant extracts, mould cultures and antibiotics tested were inactive at concentrations permitting the growth of the host. Proflavine exerted a viristatic effect while notatin and hydrogen peroxide were lethal in a defined medium but not in broth.

Summary: The influence of certain carbohydrates, nitrogen compounds and accessory substances on the growth and sporulation of Chaetomium globosum and Memnoniella echinata has been studied. It has been confirmed that M. echinata needs an external supply of biotin, which has also a slight effect on the growth of Chaetomium globosum. For C. globosum, a very low level of soluble sugar in the medium was essential for the production of perithecia. With adequate biotin, Memnoniella echinata sporulated in the presence of considerable concentrations of sugar, but at low biotin levels no sporulation occurred until soluble sugar approached exhaustion.

Jute extract stimulated growth and accelerated sporulation of Chaetomium globosum; this was not due to the presence in the extract of any of nine well-known B-group vitamins. Jute extract had no more influence on Memnoniella echinata than would be due to its biotin content.

SUMMARY: When concentrated by precipitation with acid and salts, or by highspeed centrifugation, potato virus X tends to become insoluble though still remaining infective and serologically active. This greatly complicates purification and no method was found that could be relied upon to give good yields of virus with constant properties. Insolubility is correlated with the aggregation of virus particles to form long threads that become entangled, but it is probable that combination of the particles with some cell constituents is also concerned.

Insoluble preparations dissolve slowly when incubated with pH 7·5 borate buffer, and rapidly in the presence of trypsin or chymotrypsin. Both of these enzymes hydrolyse virus X, chymotrypsin being the more effective, but different strains of the virus vary in their susceptibility.

Ribonuclease readily hydrolyses the nucleic acid derived from virus X, but seems to have no enzymic action on the active virus. When mixed with the virus, the enzyme combines with it and reversibly inhibits infectivity. At pH 7 the addition of ribonuclease to soluble virus preparations causes loss of anisotropy of flow, a fall in precipitin titre, and the production of an insoluble complex. Incubation at pH 7·5 with borate buffer slowly dissolves the complex and restores the original properties of the virus; the rate of re-solution is increased by the presence of trypsin. Some preparations of the virus were partially decomposed by incubation with borate buffer, and sometimes the rate of decomposition was increased in the presence of ribonuclease.

Summary: Glutamine is not required for staphylococcal growth, and was found to be synthesized by staphylococci from NH3 and glutamic acid in the presence of glucose. The amount of glutamine found was, however, much less than that of the NH3 reacting. Similar relationships were found in Proteus morganii, Pr. vulgaris, Saccharomyces cerevisiae and S. ellipsoideus. Proteus morganii was, however, outstanding in possessing a powerful glutaminase which could easily be separated from the cells; in this it differed markedly from Pr. vulgaris. Glutaminase activity was also demonstrated in yeasts, where its occurrence has been previously a matter of dispute.

Summary: When using MacConkey broth medium incubated at 44° as a confirmatory test for Bacterium coli Type I, positive reactions were produced by anaerobic lactose-fermenting bacteria such as Clostridium welchii. The use of brilliant green bile broth instead of MacConkey broth was completely successful in suppressing the growth of these organisms at 44°, but did not influence the growth of Bact. coli Type I. Two other coliform organisms, namely, Irregular Type II and Irregular Type VI were found to ferment lactose at 44°. A rapid test for their differentiation from Bact. coli Type I has been based upon the ability of that organism to produce indole at 44°, whereas the two irregular types are indole negative. By subculture of presumptive positive tubes in brilliant green bile broth and peptone water together with incubation at 44°, an estimation of the Bact. coli content of a water sample may be obtained within 48 hr. of primary inoculation.

SUMMARY: Penicillin in concentrations up to 100 units/ml. in broth or synthetic media has no demonstrable effect, after 20 hr. incubation at 37°, on the activities of Staphylococcus K phage, Coli-phage C 36, Coli-dysentery phage S13, a Streptococcal phage and a Bacillus subtilis phage.

The simultaneous action of penicillin and phage on young cultures of Staphylococcus aureus (Oxford) in broth or synthetic medium at 37° produces, under certain conditions, a more rapid lysis than occurs in the presence of penicillin or phage alone.

The phenomenon of accelerated lysis through the joint action of penicillin and phage occurs with other organisms besides Staph. aureus, e.g. B. subtilis and Streptococcus pyogenes, Group C, differing from that with Staph. aureus only in degree.

Penicillin does not affect the adsorption of phage by the organisms. When the amount of antibiotic is sufficient to interfere adversely with the growth of the cell then the multiplication of phage decreases. It is suggested that certain balanced intracellular reactions of metabolism are disturbed by the action of penicillin, and as a result, intermediates essential to growth both of cell and phage cease to be available.

A phage-inhibiting substance was demonstrable in certain instances when Staph. aureus (Oxford) cultures were lysed by penicillin.

Summary: The ultra-violet light and electron microscopes were used to observe the effect of bacteriophage and penicillin on Staphylococcus aureus. Penicillin caused the organisms to swell to almost twice their normal size, and immediately before lysis they became less opaque to electrons and internal structure was evident. The triply segmented cocci seen in the ultra-violet light micrographs to be characteristic of penicillin-treated Staph. aureus appear to be due to the irregular arrest of the process of division, whereby a cell partly divides and then only one of the incipient daughter cells partly divides again.

The adsorption of the Staph. K phage to the surface of Staph. aureus, the latent period during which the coccus enlarges and the phage multiplies, and the lysis of the cell with liberation of phage particles were also recorded optically. Accelerated lysis of Staph. aureus by the combined action of phage and penicillin was not associated with any peculiarity in micrographic appearance, except that with high concentrations of penicillin the number of phage particles released on lysis was diminished.

During air- and freeze-drying the diameter of Staph. aureus contracts by 30–50%.

Electron micrographs of Staph. K bacteriophage, gold-shadowed and unshadowed, show a round head totally opaque to 50 kV. electrons, 50–60 mμ. in diameter, and a slender tail 200–250 mμ. in length.

Summary: By a cup-plate method, the antibiotic activity of fluid cultures of 253 strains of spore-bearing anaerobic bacilli representing over sixteen Clostridium spp. was tested against one strain each of Mycobacterium phlei, Staphylococcus aureus, and Bacterium coli. Two strains of Clostridium sporogenes and two of Cl. bifermentans showed a weak and variable activity against Mycobacterium phlei. The remainder of the tests were all negative.