- Volume 135, Issue 4, 1989

The effects of the antibiotic validamycin A on the inositol content and hyphal branching of Rhizoctonia cerealis were investigated. Inositol was found associated with the cell wall and as a lipid extractable fraction composed of phospholipid, phosphosphingolipid and glycolipid. Validamycin A decreased the inositol content of the phospholipid fraction by 57% and the phosphosphingolipid fraction by 40%, but had no effect on the glycolipid fraction. The phosphatidylinositol (the only inositol-containing phospholipid detected) content of the biomass was reduced by 53 % in the presence of validamycin A and this decrease was compensated for by rises in the levels of phosphatidylserine and an unknown, minor phospholipid. Over the concentration range 1 to 5000 µm, validamycin A caused decreases in the colony radial growth rate, internode length (the distance between adjacent branches on a leading hypha) and inositol content of R. cerealis colonies, and a positive correlation was found between inositol content and internode length. Addition of inositol to the medium only slightly antagonized the effect of the antibiotic on mycelial morphology and did not restore phosphatidylinositol levels of the biomass to control values.

The regulation of the enzymes associated with one-carbon metabolism and the assimilation of nitrogen, together with the cellular composition of Hyphomicrobium X, were investigated. The effect of changing the methanol-carbon concentration with the NH+ 4-nitrogen concentration remaining constant (C: N ratio) in the medium during chemostat growth at a constant dilution rate was studied. As the medium changed from a C-limitation to a dual C- and N- and finally a N-limitation, the culture gradually passed through three definite growth phases. In response to these environmental conditions the cellular composition and the specific enzyme activity patterns changed. The C-content of the cells changed very little. The N- and protein-content was constant over C-limiting conditions, but under dual C- and N-limiting and N-limiting conditions an accumulation of poly-β-hydroxybutyrate (PHB) occurred and as a consequence the N-content and protein-content of the cells decreased. The enzyme associated with N-assimilation during C-limitation was an NADP+-dependent glutamate dehydrogenase which was replaced by the high affinity glutamine synthetase and glutamate synthase pathway immediately the NH+ 4-N concentration in the medium became limiting. Similarly the specific activity of methanol dehydrogenase, which was high during C-limiting conditions, dropped to a low level as the NH+ 4-N concentration decreased. Finally carbon balances were constructed throughout the experiment which showed that irrespective of the C:N ratio in the medium during C-limitation, the methanol-carbon was fluxed into biomass and CO2 only; during dual limitation the carbon was channelled into biomass, CO2 and PHB; and finally when the growth was in the presence of excess carbon no methanol-carbon was directed into over-metabolite production but, instead, the excess carbon was oxidized through the dissimilatory pathway.

The effect of growth conditions on the production of alginate by highly mucoid strains of Pseudomonas mendocina was investigated, using pH-controlled batch and continuous cultures. In batch culture, alginate was produced both during the exponential and stationary phases of growth. A polysaccharide concentration of 20 g 1−1 was obtained from medium containing glucose (50 g l−1). In steady-state continuous culture under nitrogen limitation (dilution rate, D = 0·05 h−1), a similar alginate concentration was attained; more than 60% of the utilized glucose was converted to alginate and the polysaccharide concentration was ten-fold the biomass concentration. Increasing the dilution rate above 0·1 h−1 decreased the proportion of glucose converted to alginate although the biomass was unchanged. The optimum dissolved oxygen tension (d.o.t.) for alginate production in nitrogen-limited continuous culture was approximately 40 mm Hg (1 mm Hg is approximately 133 Pa). The respiration rate and bacterial density were independent of d.o.t. above 10 mm Hg. Under oxygen limitation, no alginate was produced but acetate was excreted into the culture fluid. Variants producing little alginate arose during growth in continuous culture. Under carbon limitation, alginate production was substantially decreased but not eliminated.

The synthesis of the pyrimidine biosynthetic enzymes is repressed by the pyrimidine nucleotide end-products of the pathway. However, purine nucleotides also play a role. In this study, I have measured expression of the pyr genes (pyrA-E) in Salmonella typhimurium strains harbouring mutations that permit manipulation of the intracellular pools of both pyrimidine and purine nucleotides. The results identify the effectory purine compound as being a guanine nucleotide; it is probably GTP, but it may be GDP or GMP. The synthesis of carbamoylphosphate synthase, encoded by pyrA, and particularly dihydroorotase, encoded by pyrC, and dihydroorotate dehydrogenase, encoded by pyrD, is stimulated by the guanine nucleotide, while the synthesis of aspartate transcarbamoylase, encoded by pyrBI, and orotate phosphoribosyltransferase, encoded by pyrE, is inhibited by guanine nucleotides. The regulatory pattern of each pyr gene is discussed in relation to present knowledge on gene structure and regulatory mechanism.

The effect of light and dark incubation on the ultrastructure, pigments and lipids of five marine cyanobacteria was studied. All light-grown cyanobacteria contained in their lipid extracts the four major lipid classes characteristic of chloroplasts, namely monogalactosyldiacylglycerols, digalactosyldiacylglycerols, sulphoquinovosyldiacylglycerols, and phosphatidyl glycerols. However, the fatty acid patterns of the cyanobacterial total lipids and individual lipid classes were different from those of chloroplasts, and responded differently to dark incubation. In two cyanobacteria the fatty acid patterns of total lipids from light- and dark-incubated cultures were similar, but in the other three, dark incubation was associated with increased levels of oleic acid in the total lipids and individual lipid classes. The decrease of linolenic acid known to occur in lipids from photosynthetic eukaryotes in response to dark incubation was recorded only in Spirulina subsalsa. However, linolenic acid was not esterified in monogalactosyldiacylglycerols, as it is in chloroplasts. We also found the class of alcohol glycosides in three non-heterocystous cyanobacteria.

Plasma membranes were isolated from the acellular slime mould Physarum polycephalum by differential centrifugation and an aqueous two-phase polymer method. ATPase activity in the membrane fraction was optimal at pH 6·5 and was severely inhibited by vanadate but resistant to oligomycin. The protein components of the plasma membrane were analysed by polyacrylamide gel electrophoresis. Glycoproteins were located on Western blots by incubation of the sheets with lectin-peroxidase reagents. The results indicated that most of the membrane proteins were glycosylated. Pulse-chase experiments with [3H]glucosamine showed that when the plasmodia were induced to differentiate to macrocysts, turnover of the membrane glycoproteins occurred more rapidly than in growing plasmodia.

Five strains of Gram-negative denitrifying bacteria that used various ketones as sole carbon and energy sources were isolated from activated sludge from a municipal sewage plant. Three strains are related to the genus Pseudomonas; two non-motile species have not yet been affiliated. All strains grew well with ketones and fatty acids (C2 to C7), but sugars were seldom utilized. The physiology of anaerobic acetone degradation was studied with strain BunN, which was originally enriched with butanone. Bicarbonate was essential for growth with acetone under anaerobic and aerobic conditions, but not if acetate or 3-hydroxybutyrate were used as substrates. An apparent K s value of 5·6 mm-bicarbonate was determined for growth with acetone in batch culture. The molar growth yield was 24·8–29·8 g dry cell matter (mol acetone consumed)−1, with nitrate as the electron acceptor in batch culture; it varied slightly with the extent of poly-β-hydroxybutyric acid (PHB) formation. During growth with acetone, 14CO2 was incorporated mainly into the C-1 atom of the monomers of the storage polymer PHB. With 3-hydroxybutyrate as substrate, 14CO2 incorporation into PHB was negligible. The results provide evidence that acetone is channelled into the intermediary metabolism of this strain via carboxylation to acetoacetate.

The rRNA gene restriction patterns of 110 strains belonging to 12 staphylococcal species have been determined. The strains, isolated from various sources, were epidemiologically unrelated. Total DNA was cleaved with restriction enzymes HindIII and EcoRI, electrophoretically separated and probed with radiolabelled 16S rDNA from Bacillus subtilis inserted in a plasmid vector, pBR322. Fourty-four distinct Hindlll patterns and 44 distinct EcoRl patterns were observed. Strains belonging to different species had different patterns. Although distinct patterns were also observed within some species, a core of common bands could be discerned within each species or subspecies. Analysis of the patterns revealed two taxa in Staphylococcusxylosus which were not evident using phenotypic characteristics. Of 18 strains which were difficult to identify using phenotypic schemes, 15 showed patterns typical of known species. The three remaining atypical strains showed unusual patterns and may belong either to a known species, not included in the study, or to a new species. Since various patterns were observed within some species (e.g. S. aureus and S. epidermidis), rRNA gene restriction patterns may have epidemiological, as well as taxonomic interest.