- Volume 85, Issue 2, 1974

Summary: The major pigments induced by parahydroxymercuribenzoate and parachloromercuribenzoate (both SH group inhibitors) in the fungus Cephalosporium diospyri have been characterized as carotenoids. Those present in both chemically-induced and light-induced cells include phytoene, β-carotene, γ-carotene, lycopene, torulene and neurosporaxanthin. Two other organomercurial compounds, phenyl mercuric acetate and phenyl mercuric chloride, were also found to induce pigment formation. These pigments have the absorption properties of carotenoids, but have not been fully characterized. A range of other SH group inhibitors tested failed to induce any pigment formation.

Summary: Isolates of 26 fungal species from broiler house litter were screened for their ability to metabolize and methylate 2,3,4,6-tetrachlorophenol over a five-day period, by adding the chlorophenol to cultures after mycelial pellets were established on a complete growth medium. Under these conditions, 99 of the 116 isolates tested metabolized the chlorophenol and 68 of these produced 2,3,4,6-tetrachloro-anisole. The proportion of the chlorophenol methylated to the chloroanisole differed widely with the isolate, even within species. The highest methylation was observed with Penicillium corylophilum; certain other isolates, notably of P. brevicompactum, metabolized almost all the chlorophenol without forming the chloroanisole. Progress studies with these species suggested that there is more than one route for the metabolism of 2,3,4,6-tetrachlorophenol. In tests with suspensions of mixed bacterial populations from broiler house litter, the chlorophenol was metabolized but no methylation was detected.

Summary: Washed suspensions of Epidinium spp. grown in vitro and incubated anaero-bically in salts medium engulfed all the bacteria tested at rates ranging from 60 to 7000/protozoon/h at a density of 109/ml. The rate of uptake of bacteria was also compared by the volume of medium cleared of bacteria/h. On both bases Proteus mirabilis was taken up most rapidly. Some of the bacteria were killed and digested inside the protozoa. The Epidinium spp. took up a wide range of amino acids at rates varying from 6 to 250 nmol/106 protozoa/h and incorporated them into protein, mostly without conversion to other amino acids. The protozoa also incorporated purines and pyrimidines into nucleic acid and interconverted adenine and guanine and also uracil and cytosine.

Epidinium ecaudatum caudatum took up [14C]gIucose more rapidly than Entodinium caudatum and used the glucose carbon for the synthesis of an intracellular glucose polysaccharide and at least eight amino acids, 15 % of the 14C in the protozoa appearing in protein. The protozoa also synthesized protein from starch grains. Evidence has been obtained that Epi. ecaudatum caudatum cannot obtain all the protein it requires for growth from bacteria, free amino acids, glucose or starch and must incorporate protein from the wholemeal flour on which it is grown.

Summary: Washed suspensions of three Epidinium spp., grown in vitro and incubated anaerobically in salts solution, released into the medium protein containing hexokinase and an enzyme which lysed Bacillus megaterium, B. subtilis and Micrococcus lysodeikticus. Material from the lysate, containing bound amino acids, was incorporated by Epidinium spp. at a rate that would allow only very slow growth if it were the sole source of nitrogen. Although there was no massive cell lysis the enzymes were liberated as the result of a release of whole cell contents by an unknown mechanism. In the presence of other rumen protozoa, protein, but no lytic enzyme, appeared in the medium. No good evidence was obtained for the existence in the rumen of a lytic enzyme associated with the presence of Epidinium spp.

Summary: The technique of autoradiography in the electron microscope has been used to determine the site of incorporation of tracer from tritium-labelled glucose, amino acids and bacteria by Entodinium caudatum and the ovine form of Epidinium ecaudatum caudatum. Both protozoa incorporated tracer from glucose principally into protozoal polysaccharide and intracellular bacteria. The results with the bacteria confirmed previous findings that Entodinium caudatum digested Bacillus megaterium rapidly in vesicles in the endoplasm, whereas Epidinium spp. lysed the bacteria in the medium before incorporation. These products were then taken up both by the protozoa and the bacteria attached to their surface. In the presence of Entodinium caudatum label from tritiated Escherichia coli was found principally in the endoplasm, whereas that from glycine was present in the ectoplasm also.

Summary: Two Pseudomonas sp. able to grow with folic acid as sole source of carbon and nitrogen were isolated from soil. Strain FO1 converts folic acid to pteroic acid which accumulates in the culture medium. Strain FO8 grows with various pteridine compounds. From the culture medium of this strain the following compounds were isolated: deaminated folic acid, pteroic acid, deaminated pteroic acid, and lumazine-6-carboxylic acid. Extracts deaminated various pteridines with high activity. A pathway of folic acid degradation is proposed.

Summary: A mutant of Bacillus subtilis which required riboflavin for growth was grown with limiting or excess riboflavin. Riboflavin deficiency decreased the content of flavin, particularly of FMN in the particulate fraction. The activities of particulate succinate and NADH dehydrogenases also decreased, whereas α-glycerophosphate dehydrogenase activity did not alter. When riboflavin was added back to a starved culture, the bacterial flavin content recovered quickly, even when chloramphenicol was present. Succinate dehydrogenase activity also recovered, but this increase was inhibited by chloramphenicol.

Summary: A survey of phenol-water extracts of 17 different Fusobacterium and Bacteroides species, including subspecies, and of Leptotrichia buccalis has been carried out using colorimetric and chromatographic techniques to demonstrate the presence of heptose and 2-keto-3-deoxy-octonate. Both sugars were found in the water phase of the extracts from Fusobacterium strains and Leptotrichia buccalis. Neither heptose nor 2-keto-3-deoxy-octonate were detected in the water or the phenol phase of extracts from strains classified as Bacteroides.

Summary: Mucopeptide synthesis in rodA and rodB mutants of Bacillus subtilis has been examined by studying the incorporation of N-acetyl[1-14C]glucosamine. It has also been examined in a rodA ald mutant by the incorporation of D-[1-14C]alanine. In the latter the changes in the ester-linked d-alanine of the teichoic acids was also studied. When the growth temperature for the rodA mutant is changed from 30 to 45 °C and the organisms become round, mucopeptide synthesis is increased so that the bacteria finally contain 4 to 5 times as much mucopeptide per unit of dry weight. The change in rate of synthesis is immediate. The change in proportion of ester-linked alanine in the walls is entirely due to dilution of the existing material by additional mucopeptide. This suggests that the large reduction in the proportion of teichoic acid in the walls may similarly be due to dilution by increased mucopeptide. On changing the growth temperature from 45 to 30 °C the time course for the formation of mucopeptide is complicated, having an initial phase of rapid deceleration and a secondary phase of acceleration to a rate characteristic for the lower temperature. In both the shifts, up and down, the morphological changes accurately reflect the periods of maximum alteration in mucopeptide content. There are no similar changes in rodB mutants, but a small reduction occurs in the specific activity of the mucopeptide-N-acetyl hexosamines, when N-acetyl[1-14C]-glucosamine is present in the medium and l-glutamic acid is added to convert the round forms to rods. None of the changes occur in the parent organisms not carrying the rod mutations.

Summary: We have analysed the pattern of concentric waves visible at the onset of aggregation of amoebae of Dictyostelium discoideum and have shown that each wave consists of a light band of elongated, moving cells, and a darker interband zone of rounded cells. Our analysis supports earlier suggestions that aggregation occurs in response to chemotactic signals emanating from a centre and which are propagated outwards through the field by a relay mechanism. The width of each band of moving cells corresponds to the distance the signal is propagated during the time that the cells remain elongated after stimulation, and does not vary with signal frequency. The width of the interbands of non-moving cells depends upon the distance the signal is propagated between signalling events, and varies with the signal frequency which increases during aggregation. The velocity of signal propagation decreases slightly with increase in the density of the monolayer of aggregating cells. We have shown by time-lapse films taken at high magnification that the signal is relayed radially outwards in steps of approximately 57 μm (relay zones) and that the response of each successive zone occurs approximately 12 s after the previous one (relay time). We have attempted to demonstrate the existence of a refractory period for chemotactic responsiveness. Our results indicate that such a refractory period, if it exists, cannot be more than 12 s.

Summary: The spore of Nosema algerae from the tissues of Anopheles stephensi has a wall consisting principally of electron-dense exospore and electron-lucent endospore. The polar filament, composed of two membranes around an amorphous or tubular core, described eleven coils in the posterior half of the spore and joined anteriorly with the umbrella-shaped polar sac. The polaroplast consisted of close-packed single membranes and sacs disposed around the anterior straight part of the filament. There was no posterior vacuole. There were two nuclei in a central area of cytoplasm containing abundant ribosomes and cysternae of endoplasmic reticulum. The cytoplasm beneath the plasmalemma and around the coils of the polar filament lacked ribosomes. Various hypotheses for filament extrusion are discussed in the light of the present observations. It is proposed that the positions of the outer membranes of the polar filament are inverted during extrusion and that the core becomes a coat to the filament. Some of the cytoplasm, together with the nuclei, passes through the filament and probably acquires a membrane from the sealed tip of the filament to become the sporoplasm. The polaroplast does not participate structurally in extension of the filament.

Summary: An attempt was made to define the basis of temperature-induced aberrant wall biosynthesis in Cryptococcus diffluens. A quantitative determination of the sulphydryl content of the cell surface of C. diffluens as an index of a protein disulphide reductase activity failed to reveal differences between cells cultured at permissive and non-permissive growth temperatures. Assays of β-1,3-exo-glucanase showed no abnormal activities which could be responsible for the temperature-induced aberrations. Cryptococcus diffluens excreted low molecular weight compounds non-specifically: UMP and AMP were the predominant nucleotides excreted. Uridine diphosphoglucose, a precursor of glucan biosynthesis, was accumulated and excreted by C. diffluens at non-permissive but not at permissive growth temperatures. A correlation exists between the rate of yeast multiplication at permissive temperatures and the extent of residual growth upon subsequent exposure to a non-permissive temperature.

Summary: Mutuants of Escheriachia coli were induced by u.v. light and selected by the criterion that they formed small colonies on a nutrient agar at normal temperature. Nine slow-growing mutant strains were isolated. These mutants were characterized and compared during exponential growth in a nutrient medium with respect to doubling time, average cell mass, and DNA and RNA contents. The mutants showed only two groupings of doubling times, approximately 60 min and 35 min, as compared with 20 min for the parent. The Mutants were heterogeneous with regard cell mass, ranging from considerably larger than the parent to smaller. There was no constancy between average cell mass and the population doubling time. RNA:mass and DNA:mass ratios were similar among mutant strains with the same doubling time despite large variations in average cell mass.

Summary: d-Glutamate inhibited hyphal extension, the degree of inhibition depending on the identity of the glutamate dehydrogenase (GDH) enzyme present in the mycelium. Mycelia were most sensitive to inhibition on media which promoted formation of the NAD-linked GDH. The activity of this enzyme was inhibited by d-glutamate in experiments with cell-free extracts, the inhibitions being noncompetitive or mixed. Similarly the activity of the NADP-linked enzyme (GDHNADP) was inhibited in its utilization of ammonium ions (in the amination reaction) and l-glutamate (in the deamination reaction). In contrast, 2-oxoglutarate was a co-operative substrate for GDHNADP in the amination reaction and d-glutamate acted as an allosteric activator. d-Glutamate was also able to derepress the synthesis of GDHNADP; the NAD-linked enzyme was hardly affected.