- Volume 28, Issue 2, 1962

SUMMARY: During the resumption of development in cultures of Nostoc muscorum A, grown in the dark, and then exposed to light, cellular attachment occurs. This process is believed to be prerequisite for the formation of heterocystous filaments. The significance of anastomosis in the developmental cycle of Nostoc is discussed and a tentative scheme for an alternation of sporogenous and heterocystous generations is presented.

SUMMARY: Electron microscopy of apothecia of a species of a discomycete, Dasyscyphus sp., provided the opportunity to observe various aspects of ascospore formation. This report presents a series of micrographs arranged to represent what are interpreted to be successive stages in the process.

SUMMARY: The enzymic conversion of citrulline argininosuccinate arginine in wild-type Neurospora crassa has been shown to be essentially like that in mammalian tissues in all respects tested. The enzymes responsible for the two reactions (condensing enzyme and argininosuccinase) have been partially separated. Five mutants at the arg-1 locus have normal arginino-succinase and little or no condensing activity. The lack of condensing activity appears to be due to a simple absence of enzyme, alternatives such as inhibitor production or increased ATPase competition having been ruled out. Small amounts of apparent condensing activity, detected in the substrate-disappearance assay, were shown to be due to side reactions. When grown at the usual high arginine concentrations, three mutants at the arg-10 locus, which are known to lack argininosuccinase, have normal condensing activity. However, when arg-10 strains are grown at low arginine concentrations, the resulting extracts have very little condensing activity. This also appears to be due to a simple loss of condensing enzyme. It has not been determined whether the low activity is a specific secondary effect of arg-10 on the condensing enzyme, or whether it is merely a non-specific effect of the inadequate supplementation, which might cause reductions in many enzyme activities. Despite its very low condensing activity in vitro, arg-10 grown at low arginine concentrations must be active at some time in vivo, since its mycelium accumulates argininosuccinate but not citrulline. In contrast, an arg-1 arg-10 double mutant, grown at low arginine concentrations, must be inactive in vivo, since it accumulates citrulline but not argininosuccinate. It is concluded that arg-1 is probably the primary locus controlling the synthesis of condensing enzyme.

SUMMARY: A cytological investigation of cell walls, lipid granules, metachromatic granules and chromatinic bodies of Nocardia rubra revealed that each of these structures existed as separable entities. Multicellularity was not observed, as indicated by the lack of septa in non-fragmenting organisms, but lipid inclusions were frequently found in close association with cell walls. Metachromatic granules were separable from lipid inclusions and were found to be homologous with electron opaque granules. No evidence for the origin of metachromatic granules as cytoplasmic condensations upon cell septa was found. Acid hydrolysis or ribonuclease treatment revealed Giemsa-staining chromatinic structures which were removed by deoxyribonuclease digestion. Metachromatic granules were not removed by ribonuclease, deoxyribonuclease hydrolysis or a combination of both, but these granules were removed by acid hydrolysis.

SUMMARY: The nucleic acid precursor requirements of Mycoplasma laidlawii strain A were determined by using a partially defined medium. Adenine, guanosine and cytidine were found to be the minimal growth requirements. However, best growth was obtained with undegraded RNA or oligoribo-nucleotides. Thymidine was not essential for growth when folinic acid was present in the medium; folic acid was completely inactive. M. laidlawii utilized the 2#and 3#-ribonucleotides most poorly. The dephosphorylation of these mononucleotides to nucleosides rendered them growth promoting.

SUMMARY: Acetoacetic acid is a product of degradation of homogentisic acid by cell-free extracts of a vibrio grown with phenylacetic acid as sole carbon source. 4-Maleylacetoacetate was identified as a reaction intermediate; reduced glutathione was a cofactor for this enzymic degradation. These reactions are similar to those catalysed by enzymes of rat liver.

SUMMARY: Bacteria in cultures of Salmonella typhimurium LT2 were peritrichously flagellated when grown in nutrient broth at 370; but most were non-flagellated when grown for 6 mean-generation times or more at 440. When a culture growing exponentially at 370 was transferred to 440, growth continued at about the same rate; but the synthesis of new flagella was largely curtailed. The fate of the parental flagella was studied by staining and counting flagella on bacteria from samples taken during growth at 440 of cultures first grown at 370. After 3 mean-generation-times the average number of flagella/flagellated bacterium had fallen from about 8 to about 2 and the proportion of flagellated bacteria from about 100 % to about 60 %. The distribution of numbers of flagella/bacterium was at all times unimodal, with the mode decreasing from about 8 to 0. In non-growing cultures at 440 there was little or no change in the average number of flagella/bacterium, in the proportion of flagellated bacteria, or in the distribution of numbers of flagella/bacterium. It is inferred that parental flagella are neither rapidly shed at 440 nor retained entirely by one daughter cell at each division but are distributed about equally between the two daughter cells.

SUMMARY: Suspensions of Entodinium caudatum containing less than one viable bacterium per ten protozoa were prepared by aerobic incubation of the protozoa with penicillin, streptomycin, dihydrostreptomycin and neomycin in the presence of autoclaved rumen fluid and rice starch grains. Unfortunately these almost axenic Entodinia could be maintained alive for only 3-4 days. Replacement of the rice starch grains in the medium used for bacterial removal by soluble starch produced almost bacteria-free protozoa which were devoid of food storage materials and died more rapidly than usual. The life of these protozoa was prolonged by adding any of the following: rice starch grains, soluble starch, maltose, glucose and some other sugars.

SUMMARY: An approach has been made to the problem of how the synthesis of coenzymes is regulated. Two aspects of the problem have been studied, especially as they concern the synthesis of flavins by bacteria:

(1) How are coenzymes prevented from being synthesized as fast as amino acids or nucleic acid bases?

(2) How is coenzyme synthesis adjusted to the often changing physiological needs of bacteria?

Evidence is presented that flavins cannot inhibit the activity of enzymes in the flavin biosynthetic pathway of Escherichia coli, but the amount of these enzymes can be made to vary by a factor of at least two. Repression might, therefore, account for the low rate of flavin synthesis. The possibility that repression rather than feedback inhibition also accounts for the low rate of synthesis of other coenzymes is discussed.

Flavin synthesis is not as precisely adjusted to the physiological needs of bacteria as are syntheses of major metabolites for the following reasons:

(1) Flavins are greatly overproduced by bacteria during exponential growth; the ratio of flavins excreted to flavins retained in the cells is between 0.8 and 8 for all strains and cultural conditions tested.

(2) Flavin synthesis is not tightly geared to growth; thus, flavin synthesis goes on uninterrupted for more than an hour when the growth rate of E. coli or Pseudormonas fluorescens is abruptly reduced from a rapid rate to zero; also growth goes on uninterrupted for over an hour when the flavin supply is abruptly cut off from rapidly growing lactic acid bacteria. Evidently the control mechanism in the flavin pathway is not very sensitive to physiological needs. This conclusion probably applies to other coenzymes as well.

Some incidental findings of interest from other points of view were:

(1) Although internal flavins can get out of E. coli, external flavins apparently cannot enter. This could account for the absence of flavinless mutants.

(2) After brief treatment with penicillin, E. coli becomes permeable to external flavins while remaining both impermeable to inulin and capable of synthesizing flavins.

(3) Less than 4 % of the intracellular flavins of E. coli are free, in a form that can be extracted with n-butanol (5 %, v/v), toluene (0.05 %, v/v), cetyl trimethylammonium bromide (0.001 %, w/v) or distilled water. The remaining flavins are bound, in a form that can be extracted with trichloracetic acid (5 %, w/v). The intracellular concentration of free flavins in E. coli is estimated to be less than 4 x 10−6M.

SUMMARY: An anaerobic actinomycete isolated from the lachrymal duct of a case of human lachrymal canaliculitis is described. Although the organism was described previously as Actinomyces israelii, more recent observations show it differs significantly from members of this genus in metabolism, physiology and cell-wall composition. The organism is similar to the Actinomyces in its production of true branching mycelial elements in vivo and in vitro, catalase negativity, pathogenicity for experimental animals, and amino acid cell-wall composition. It is similar to the propionic acid bacteria in its fermentation of glucose to propionate, acetate, and CO2, its morphological variation, formation of a dull orange colour, and the presence of diaminopimelic acid in its cell wall. Other characteristics such as the formation of DL-lactic acid, absence of a CO2 requirement either anaerobically or aerobically, inability to ferment glycerol or lactate, and sugar and amino sugar composition of the cell wall also serve to differentiate this organism from other strains of Actinomyces and from Propionibacterium. In view of these observations the organism is reclassified and named A. propionicus, sp.nov. The possible phylogenetic relation of A. propionicus to certain other branching filamentous organisms is discussed.

SUMMARY: The growth of Saccharomyces pastorianus was 50% inhibited by 0.013 p.p.m. cycloheximide, and completely inhibited by 0.100 p.p.m. in liquid medium after 24 hr. at 280. At subinhibitory concentrations of cycloheximide abnormally large cells and cell aggregates were formed, probably as a result of interference by cycloheximide with the synthesis of structural cell-wall materials. When S. pastorianus was grown in media with successively higher concentrations of cycloheximide partially resistant organisms were developed. These did not form aggregates and giant cells in the presence of cycloheximide, but on back inoculation to cycloheximide-free media they were ‘slow growers’ and formed small colonies on nutrient agar. After a few transfers on cyclo heximide-free media the resistant organisms reverted to the normal ‘fast growing’ type. The resistance to cycloheximide was not the result of any acquired ability to decompose or inactivate the compound.

SUMMARY: The physiological properties of 229 strains of mycobacteria (photochromogens, scotochromogens, non-photochromogens and rapid growers) and others already classified were analysed. According to their metabolic capacities and using the method proposed by Sneath, three branches were established.

Branch I is formed by micro-organisms with a high metabolic capacity and rapid rate of growth. It includes Mycobacterium smegmatis, M. phlei, M. peregrinum sp.nov. and many unnamed strains with great disparity in their characteristics, that were placed in a separate branch labelled ‘irregular’.

Branch II includes micro-organisms that utilize only a limited number of carbohydrates. Belonging to this branch are Mycobacterium fortuitum, M. marinm, M. piscium and M. thamnopheos. Three new species are described, two of them rapid growers, M. acapulcensis sp.nov. and M. runyonii sp.nov. and a scotochromogen, M. flavescens sp.nov.

Branch III, formed by Mycobacterium kansasii (photochromogen), M. avium (non-photochromogen), M. marianum (scotochromogen) and M. gordonae sp.nov. (scotochromogen). All are slow growing micro-organisms.

Neotype strains of old species are proposed and holotypes of the new species are designated. A key for the identification of these species and a taxonomic tree of mycobacteria are described.

SUMMARY: The exponential growth rate of a streptomycin-dependent strain of Escherichia coli was proportional to streptomycin concentration until a critical concentration was reached, above which it was independent of streptomycin concentration. The value of the critical concentration changed with a change either in the carbon source, or in the temperature of cultivation. Below the critical concentration, the macromolecular composition of the cells was affected by the external streptomycin concentration: as this decreased, the ribonucleic acid (RNA) content of the organisms increased, and the protein content decreased. When external streptomycin was removed, streptomycin-dependent organisms continued to grow for many hours. Growth was at first exponential, the extent and duration of this phase being functions of the concentration of streptomycin to which the organisms had previously been exposed. This phase was followed by a much longer period of arithmetic growth, unaccompanied by cell division, during which the organisms elongated progressively. Growth in the absence of streptomycin caused changes in the macromolecular composition of the organisms which were similar in nature to those produced by growth with a subcritical concentration of streptomycin, but much more pronounced. The greatly increased total RNA content of these organisms was not accompanied by grossly detectable qualitative changes in the RNA content of the organisms. In the absence of streptomycin, the synthesis of some enzymes was either arrested or decreased in rate; the synthesis of others was unaffected. This leads to an imbalance in the enzymic constitution. These differential effects on enzyme synthesis appeared to be random. Growth in absence of streptomycin did not seem to affect deoxyribonucleic acid (DNA) synthesis or function, as shown by the ability of a lysogenic streptomycin-dependent strain to produce infective phage under such conditions. The re-introduction of streptomycin to a culture growing arithmetically as a consequence of streptomycin depletion caused a resumption of DNA synthesis at the normal exponential rate. The rate of protein synthesis also soon increased, but attained its normal exponential rate more slowly. RNA synthesis was wholly arrested until the RNA content of the organisms had fallen to a normal value, and then resumed at the normal exponential rate. Grown in the presence of a greater than critical concentration of streptomycin, the streptomycin-dependent organism bound irreversibly about 250,000 molecules of streptomycin, half of which could be extracted with hot water, and the remainder with hot perchloric acid. A new hypothesis concerning the location and nature of the genetically determined intracellular lesion which results in streptomycin dependence is developed on the basis of these facts.

SUMMARY: Twenty-two staphylococcal bacteriophages representing serological groups A, B, F and L were examined for activity after incubation of free phage in 10 % (w/v) NaCl Trypticase soy broth for 24 hr. at 300. The mean decrease in plaque-forming units for ten phages of serological group A was 13 %; in no instance was inactivation of any group A phage greater than 33 %. All but one of nine group B phages were 88 % or more inactive after incubation in the 10 % NaCl broth; the single exception was phage 83 with slightly more than 40 % inactivation in both 0.5 % or 10 % NaCl broth. Serological group F phage 42D resembled phages of group A, whereas phage 77 of group F and group L phage 187 were intermediate between group A and B phages with respect to NaCl stability.

SUMMARY: A new Saccharomyces species has been isolated from ‘alpechin’. It is distinguished from other species of the genus by its ability to ferment and assimilate glucose, maltose and melibiose. Galactose, sucrose, lactose and raffinose are not assimilated. The melibiase found in S. hienipiensis can hydrolyse raffinose.

SUMMARY: The rate of O2 utilization by both pigmented and non-pigmented strains of Pseudomonas aeruginosa was greater with 2 % (w/v) Casitone as substrate than with 0.1 M-glucose as substrate. P. aeruginosa readily catabolized Casitone, amino acids and other nitrogenous compounds with the production of an alkaline reaction even in the presence of 0.5% (w/v) glucose. Other species of Pseudomonas studied varied in their utilization of these compounds, suggesting a different and possibly satisfactory approach to the identification and classification of species within the genus Pseudomonas.