- Volume 132, Issue 8, 1986

Summary: Anacystis nidulans (Synechococcus PCC 6301) is an obligate phototrophic cyanobacterium. When light-grown cultures of Anacystis are transferred to the dark, the on going cell cycles are aborted. To characterize the fates of cell cycle events in the dark, synchronized cultures of A. nidulans,taken at various phases of growth, were placed in the dark and the macromolecular contents and cell numbers were determined. Cell number did not increase in any culture in the dark. Protein and RNA contents remained the same. However, cultures in the last hour of their respective synthesis periods showed detectable increases in protein and RNA contents. In cultures in the early stages of DNA synthesis, no sustained increase in DNA was observed, indicating that DNA replication was not completed in the dark by these cultures. However, incorporation of 32P in the DNA fraction in the dark suggested that DNA replication was completed for cultures in the last stages of DNA synthesis. These results suggest that macromolecular synthesis and cell septum formation were curtailed (with the exceptions indicated above) and further progress in the cell cycle stopped in the dark.

Summary: The cyanobacterium Oscillatoria agardhii was grown in continuous cultures with light periods of various duration at a constant irradiance. Growth at shorter light periods led to light-limited cultures. P max was the only photosynthetic parameter that reflected increastng pigment contents at shorter light periods; α and q O 2, were maximal with light periods of8 h and less. The dynamics of the carbohydrate pool with light/dark cycles are described in a concept ofthree maxima: a maximum accumulation rate, a maximum content and a maximum consumption rate. With shorter light periods the growth yield on carbon increased as did yield values for dark growth on carbohydrate. Rates ofprotein synthesis were equal in the light and dark for light periods >8 h; with shorter light periods the rates of protein synthesis in the dark showed a severe drop. From the response of O. agardhii to changes in light/dark cycle we distinguished four ranges in which regulation ofgrowth differed. The central role ofthe photosynthetic apparatus in the different responses to changes in either irradiance or light period is stressed.

Summary: L-Lysine transport in Corynebacterium glutamicum was investigated. The bacterium was shown to possess a highly specific, energy-dependent system of active lysine transport. The system transferred lysine into the cells and exchanged intra-and extracellular lysine. Mutations in the transport system did not lead to overproduction ofthe amino acid. Resting cells ofthe parent strain, or of its lysine-producer derivatives with a defective transport system, failed to excrete lysine into the medium. An efflux ofintracellular lysine could be induced by a hyperosmotic shock, and by different membrane-active substances. It has been suggested that C. glutamicum cells are equipped with channels (pores) for excreting lysine from the cytoplasm. These channels appeared to open in response to an increase in the intracellular lysine concentration. The channel permeability also depends on the membrane structure.

summary: The transport and hydrolysis of several radioactive di- and tripeptides in Saccharomyces cerevisiae was studied. A peptide-transport-deficient mutant isolated on the basis of its resistance to nikkomycin Z lost most of its capacity to take up di- and tripeptides. The transport kinetics of [14C]methionylglycine, [14C]methionylsarcosine and [3H]nikkomycin Z indicated that peptide transport is not dependent on intracellular hydrolysis. Intact cells had some peptidase activity towards methionylsarcosine but not towards nikkomycin Z. The relationship between this activity and peptide transport is discussed.

Summary: Uptake of 54Mn from 10 nM-Mn2+ was shown to be both energy-and pH-dependent. Analysis of the uptake kinetics revealed an apparent half-saturation constant, K t of 16·4 nM-Mn2+ and a maximal rate of transport, V max,of l·01 nmol Mn2+ (g dry wt)−1 min−1 Mn2+ uptake was highly specific, being unaffected by 100-fold molar excess of Mg2+, zn2+, Ca2+, Co2+, Ni2+ and Cu2+; however, uptake was inhibited 30 to 40% by 1000-fold molar excess of Mg2+, Zn2+, Ca2+, Co2+ and Ni2+. Zn2+ competitively inhibited Mn2+ uptake, theK i value being approximately 500-fold greater than the Kt for Mn2+. Efflux studies indicated that metabolic exchange of 54Mn occurred to a small extent. Cellular Mn2+ contents remained relatively constant during growth in batch culture. The Mn2+ transport system observed appears to be analogous to the specific metal transport systems reported in bacteria.

Pseudomonas putida PP3 carrying dehalogenases I and II and Pseudomonas aeruginosa PAU3 carrying dehalogenase I coded for by plasmid pUU2 were able to grow on 2-monochloropropionic acid (2MCPA). Neither strain utilized 2-chloropropionamide (2CPA) as a carbon or nitrogen source for growth. Mutations in both strains to 2Cpa+ phenotypes (designated P. putida PPW3 and P. aeruginosa PAU5, respectively) involved the expression of an acquired 2CPA-amidase activity. The amidase followed by dehalogenase reactions in these strains constituted a novel metabolic pathway for growth on 2CPA. P. putida PPW3 synthesized a constitutive amidase of molecular mass 59 kDa consisting of two identical subunits of 29 kDa. For those amides tested this acquired enzyme was most active against chlorinated aliphatic amides, although substrate affinities (K m) and maximum rates of activity (V max) were poor. P. aeruginosa PAU5 acquired a 2Cpa+ phenotype by overproducing the A-amidase normally used by this species to hydrolyse aliphatic amides. The A-amidase had only slight activity towards 2CPA. However, with constitutive synthesis the mutant grew on the chlorinated substrates. Chloroacetamide (CAA) was a toxic substrate analogue for these Pseudomonas strains. A strain resistant to CAA was isolated from P. aeruginosa PAU5 when exposed to 1–10 mM-CAA. This mutant, P. aeruginosa PAU6, synthesized an inducible A-amidase. CAA-resistance depended upon the simultaneous expression of CAA-inducible amidase and dehalogenase activities.

Summary: Pseudomonas putida mt-2 (ATCC 33015) carrying the TOL plasmid pWW0 could adapt to growth on the aromatic amines aniline and m-and p-toluidine. In strain UCC2, a derivative adapted to rapid growth on these compounds, they were oxidatively deaminated to catechol or 4-methylcatechol, which in turn were dissimilated by a meta-cleavage pathway. The aniline/toluidine oxygenase and the meta-cleavage pathway enzymes were inducible by aromatic amines. Evidence is presented that in strain UCC2, plasmid pWWO has undergone deletion of its catabolic genes, and that it is a novel plasmid, pTDNl, which is involved in the catabolism of aniline and m-and p-toluidine. The meta-cleavage pathway genes which are carried by pTDN 1 were shown not to have originated in pWW0.

Summary: Azospirillum Lipoferum strain D-2 produces the phenolate siderophores 2,3-dihydroxybenzoic acid, 3,5-dihydroxy benzoic acid and salicylic acid under iron-starved conditions. Lysine and leucine were identified as the amino acid conjugates of the siderophores. The concentration of siderophores in the culture supernatant was maximal after 20 h growth. Iron-binding proteins were present in membranes of cultures grown under iron starvation. Iron uptake was enhanced in the presence of each of the siderophores.

Summary: 1,3-β-D-Glucanase and chitinase activities were induced in Trichoderma harzianum when glucose or cell walls of Rhizoctonia solani (anastomosis group AG2) were used as sole carbon source. Electrophoresis showed that more proteins were induced by cell walls than by glucose. The composition, as revealed by electrophoresis, of the T. harzianum extracellular proteins was similar when grown on R. solani AG2 and AG4 cell walls but different for R. solani AG1 cell walls. Several major proteins were induced in all strains of T. harzianum but there were strain differences in the number and intensity of other proteins. The number of induced proteins was less for four strains of T. viride and their composition was different from those of five strains of T. harzianum. The results indicate that chitinase, 1,3-β-D-glucanase and a large number of other extracellular proteins may be involved in the degradation of R. solani cell walls.

summury: Of 33 filamentous fungi, representing five taxonomicsubdivisions, 31 were able to grow on heat-killed Bacillus subtilis cells as sole C,N and P source. Two types of decomposition were observed: cytolysis, in which the bacterial cytoplasm was rapidly degraded, leaving apparently empty cell walls, and bacteriolysis, in which the entire bacterial cell gradually disintegrated. Supernatants of cultures in which the latter type of attack occurred contained enzymes capable of dissolving bacterial cell walls. Most of these enzymes were glycosidases with pH optima of 2·0-3·9.Two were peptidases and/or amidases with pH optima of7·8–8·3.

Summary: Some of the phenotypic characteristics of a slow-growing, nongerminating variant of a commonly studied strain of Candida albicans are described. The variant arose as a chance isolate. The rate of occurrence was about 0·1% and the reversion rate was about 1 per 106 cells. The colony size was typically smaller than that of the parent and the yeast cells tended not to separate from one another so that catenulate strands of cells (pseudohyphae) were formed. Under standard conditions the generation time of the small-colony variant in liquid shake cultures was about twice that of the parental strain. Growth of the variant was suppressed by antimycin A, indicating that the small colony form was not the consequence of a defect in the cytochrome system. The colony size of the variant was not influenced by chlorobenzotriazole, which suggested that adenine metabolism was not involved in the small-colony phenotype. The pseudohyphal growth pattern was not relieved by high concentrations of utilizable carbohydrates, which means the catenulate microscopic appearance of the yeast cells was not simply an exaggeration of the normal growth pattern of isolates of C. albicans but more probably represented the growth of a cell-cycle mutant defective at the cell separation step. The cytoplasmic proteins of the variant and the parent were very similar though some unique peptides were displayed by each.

Summary: Yeast and mycelial forms of Candida albicans ATCC 10231, growing together in 12 h and in 96 h cultures, were separated and their lipids were extracted and characterized. The total lipid content of the yeast forms was always lower than that of the mycelial forms. In 12 h cultures the lipids from the two morphological forms consisted mainly of polar compounds, viz. phospholipids and glycolipids. In 96 h cultures both the yeast and mycelial forms accumulated substantial amounts of apolar compounds, mainly steryl esters and triacylglycerols. The mycelial forms were more active than the yeast forms in this respect. Major differences in the lipid composition between the two morphological forms involved the contents of sterols and complex lipids that contain sterols. As a rule, the yeast lipids contained much larger proportions of free sterols than the mycelial lipids. However, the mycelial lipids contained several times more sterols than the yeast forms but bound as steryl glycosides, esterified steryl glycosides and steryl esters. Steryl glycosides and esterified steryl glycosides occurred in yeast lipids only in traces, if at all. The major steryl glycoside in the mycelial forms was unequivocally identified as cholesteryl mannoside. At both phases of growth the apolar and polar lipid fractions from the mycelial forms contained higher levels of polyunsaturated fatty acids (18:2 and 18:3) but lower levels of oleic acid (18:1) than the corresponding fractions from the yeast forms. The lipid content and composition of 12 h and 96 h yeast and mycelial forms of C. albicans KCCC 14172, a clinical isolate, were almost identical with those of C. albicans ATCC 10231.

Summary: DNA was isolated from Mycobacterium gadium with high purity. Its G + C content was between 64 and 67 mol%. The homology of M. gadium DNA with DNA from three other rapidly growing mycobacteria was less than 22%, which indicates that M. gadium is a discrete genomic species. Analysis of the DNAs with restriction endonucleases supported this finding.

Summary: Characteristics of bacteriophage KM1, which infects Clostridium butyricum, are described, and several important properties are compared with those of phage 5, which infects the same species. Phage KM1 has a hexagonal head and a tail with a contractile sheath. Agarose gel electrophoresis patterns of BglII, EcoRI and HindIII fragments of the two phage DNAs differed greatly. The G+C contents were 30·5 mol% for phage KM1 and 35·4 mol% for phage 5. Among the 32 strains of clostridia tested, phage KM1 infected only C. butyricum MII588. Other properties of phage KM1 are also discussed.

Summary: A soil isolate, 4B, which had been previously assigned to the genus Pseudomonas and shown to be capable of reducing C2H2 with simple phenolic compounds as sole carbon source, was further characterized in comparison with two other diazotrophs which were identified as pseudomonads. The DNA base composition of 4B was 60·2 mol% G + C. Plasmid DNA was not detected in alkaline SDS lysates of 4B by agarose gel electrophoresis. Comparable maximum C2H2 reduction activities in 4B were observed under microaerobic conditions (pO2 about 0·003 atm) with either 28 mM-glucose or 5 mM-protocatechuate as carbon source. N2 fixation was confirmed by the cellular incorporation of 15N2 in cultures of 4B grown in N-free medium. Extensive biochemical tests, including the carbon utilization pattern, demonstrated that 4B was closely related to Pseudomonas delafieldii (ATCC 17505) although the latter did not fix N2. 4B had metabolic patterns different from the two other strains reported to be diazotrophic pseudomonads; all three contained DNA homologous to the nifHDK genes of Klebsiella pneumoniae M5A1.

Summary: Gas chromatography was used to determine the long-chain fatty acid composition of 33 strains representing 17 genera of the Saccharomycetaceae, Endomycetaceae, Metschnikowiaceae and Saccharomycodaceae. The similarities between the strains were calculated on the basis of fatty acid composition, resulting in three groups. Group I strains contained oleic acid (C18 : 1) as the major fatty acid, while group II strains had substantial proportions of both oleic acid and linoleic acid (C18 : 2). Strains of group III were characterized by a high percentage of palmitoleic acid (C16 : 1). The one strain of Metschnikowia reukaufii studied fell within the range of variation of group II, but it contained a slightly higher proportion of palmitoleic acid than the other strains in this group.

Summary: A mutant of Candida albicans strain MEN, which was unable to produce mycelia in SSV medium and in horse serum at 37 °C, was isolated by a physical separation procedure. The mutant was shown to be derived from the parental strain by growth and morphology studies, sugar uptake and fermentation patterns, and the presence of genetic markers.