- Volume 104, Issue 2, 1978

A lipopolysaccharide was isolated from Neisseria meningitidis group B by phenol/water extraction and purified by differential ultracentrifugation. This preparation exhibited endotoxic properties as shown by the limulus-lysate assay. Mild acid hydrolysis of the lipopolysaccharide yielded a lipid A fraction and a polysaccharide fraction. The lipid A fraction contained fatty acids, phosphorus and glucosamine. Analysis of the polysaccharide fraction revealed the presence of glucose, galactose, glucosamine, 2-keto-3-deoxyoctonic acid and phosphorus. There was no heptose.

4,9-Dihydroxyphenazine-1,6-dicarboxylic acid dimethylester, the ester form of a proposed ‘missing link’ in the biosynthesis of phenazines, has been isolated from a strain of Pseudomonas cepacia.

Two inducible polygalacturonases (PG-1 and PG-2) from culture filtrates of Trichoderma koningii were purified to homogeneity by CM-cellulose chromatography and isoelectric focusing in a narrow pH range (pH 6 to 8). They were both hydrolytic enzymes classifiable as endopolygalacturonases [poly(1,4-α-d-galacturonide) glycanohydrolase; EC 3.2.1.15]. PG-1 and PG-2, focusing at pH 6·41 and 6·57 respectively, each consisted of a single polypeptide chain having an apparent molecular weight of 32000 as determined by gel filtration on Sephadex G-100; they were both glycoproteins and had carbohydrate contents of 0·033 and 0·062 mg sugar (mg protein)−1 respectively. When the isoenzymes were incubated with different plant tissues, they were not absorbed by any of them.

Saccharomyces cerevisiae can accommodate to the presence of toxic levels of Cd2+. This adaptation can also be induced, though to a lesser degree, by pre-growth of the yeast in 50 μ m-Zn2+. Growth of Cd2+-adapted yeast through several passages in Cd2+-free medium leads to a progressive decrease in Cd2+-tolerance by the yeast, suggesting that the adaptation did not involve selection of a Cd2+-resistant mutant. Chromatography on Sephadex G-75 of the soluble fraction from Cd2+-adapted yeast indicated that no metallothionein-like protein was present. This suggests that the mechanism of adaptation is unlike that of the higher eukaryotes.

Eight strains of thermophilic bacteria were examined for the presence of covalently closed circular deoxyribonucleic acid molecules by caesium chloride-ethidium bromide density gradient centrifugation. Four of the eight strains tested, Thermus flavus Bs1, AT61, AT62 and Thermus thermophilus HB8 carried covalently closed circular DNA molecules. Thermus flavus BS1 harboured two species of plasmids with molecular weights of 6·1 × 106 and 17·0 × 106 as determined by electron microscopy. Thermus thermophilus HB8, T. flavus AT61 and T. flavus AT62 carried plasmids with molecular weights of 6·2 × 106, 6·6 × 106 and 6·6 × 106, respectively. Plasmids from T. flavus at61 and at62 were indistinguishable in their electrophoretic patterns in agarose or acrylamide gel after digestion with various restriction endonucleases. This is the first evidence for the presence of plasmids in extremely thermophilic bacteria, though their functions are unknown.

Wild-type R plasmids of the P1 incompatibility group mediated the transfer of chromosomal genes in Rhizobium leguminosarum, but only at very low frequencies. Two P1 R plasmids, which had originally been selected in Pseudomonas aeruginosa for enhanced donor properties, promoted much higher levels of gene transfer in R. leguminosarum. One of these, R68.45, was used for linkage mapping. All markers tested mapped on a single circular linkage group. Segments of donor chromosome up to at least one-seventh of its total length were transferred and integrated into the recipient chromosome.

SUMMARY: Using R68.45 as a conjugative plasmid in crosses between strains of Rhizobium leguminosarum and R. meliloti, transfer of chromosomal genes occurred. Haploid recombinants were formed in which rif and str alleles were transferred from R. leguminosarum to R. meliloti. In crosses in the reverse direction haploid recombinants arose at barely detectable frequencies and progeny were isolated which were high frequency donors of the selected allele. Genetic evidence showed that such progeny harboured R-primes in which sections of the R. meliloti chromosome were inserted into R68.45. Four different classes of R68.45-prime, each carrying a different prototrophic marker, were obtained.

In experimental models of cholera, some strains of Vibrio cholerae harbouring P or V or both plasmids were markedly less pathogenic than isogenic strains devoid of these plasmids. This effect was due to decreased production of toxin by P+, V+ or P+V+ bacteria compared with their parents.

Phylogenetic and epidemiological relatedness among transferable plasmids belonging to the IncC, IncM and IncH incompatibility groups has been studied by DNA-DNA filter hybridization. Hybridization was carried out on nitrocellulose microfilters, at low temperature, in formamide and under paraffin oil. The degree of hybridization among plasmids belonging to the IncC and IncM groups supported the conclusions drawn from genetic classification. Studies on relatedness among plasmids belonging to the IncH group allowed their classification into three phylogenetic sub-groups. Comparison of DNA sequences of three plasmids sharing the same genetic properties and isolated from different bacterial species suggested an epidemiological spread of the same plasmid.

In a culture of Escherichia coli K12 gal (γdg), cells which form large colonies on agar plates containing galactose and thiomethyl β-d-galactoside (TMG) appear at high frequency. These clones are resistant to growth inhibition by TMG on galactose minimal medium. Biochemical studies of the steady-state levels of galactokinase and UDPgalactose 4-epimerase suggest that the resistant clones have extra copies of the genes for the galactose-metabolizing enzymes. The mutation for TMG resistance is not located in either the bacterial or the bacteriophage genome, but is probably due to an aberrant association between cell and prophage DNA.

Mapping the TMG-resistant characteristic by phage P1 indicates that TMG-resistant bacteria possess at least two gal + operons, one of which is cotransducible with bio +. In addition, TMG-resistant bacteria behave like λdg polylysogens when challenged with the phage λcI90c17. From these genetic experiments we conclude that TMG-resistant bacteria arise by duplication of the λ dg prophage. Finally, gal + bacteria which carry a single, additional, λ dg prophage are TMG-resistant. TMG resistance is probably a gal + gene dosage effect.

SUMMARY: Chlamydomonas reinhardii populations in nitrite-limited chemostat culture exhibited damped oscillations in cell number following sharp changes in dilution rate. These oscillations had a period of about 70 h and damped out after not more than two clearly observable cycles. Residual nitrogen in the culture medium during these cycles remained very low (less than 1 μmol N 1−1). The oscillations were apparently caused by the presence of a complex delay of two distinct physiological components in the relationship between cell division rate and environmental limiting nutrient concentration.

SUMMARY: Arginine and glutamine, which prevents the catabolism of arginine, are accumulated when conidia of auxotrophic mutants of Neurospora crassa are deprived of an amino acid. This accumulation requires sources of carbon and nitrogen, an intact arginine biosynthetic pathway, de novo synthesis of pyrimidines and protein synthesis. There is coordination between the synthesis of glutamine and the synthesis and catabolism of arginine. Conditions which allow these amino acids to accumulate prevent exogenous arginine from being catabolized.

Since more than half of the accumulated amino nitrogen is stored as glutamine and arginine, it is proposed that these amino acids are nitrogen and/or carbon reservoirs under non-growth conditions.

SUMMARY: Mycelium of Neurospora crassa accumulates arginine and glutamine when deprived of an amino acid or pyrimidines, or in the presence of cycloheximide or at the end of exponential growth. A large proportion of the accumulated arginine is sequestered in an osmotically sensitive compartment. Electron microscopy of this fraction shows it consists of mitochondria and electron-dense bodies. Only a small part of the accumulated glutamine was sequestered in this fraction.

Mycelium of this fungus, when deprived of an amino acid, did not catabolize arginine due to a lack of arginase induction. A relationship is proposed between the sequestered arginine, its accumulation, and the lack of induction of arginase. The accumulated glutamine in the cytosol also prevents the catabolism of soluble arginine. Guanosine tetra- and pentaphosphate were not found in the mycelium of N. crassa during the deprivation of an amino acid or after cycloheximide treatment.

Rates of nitrogenase synthesis by Klebsiella pneumoniae were measured by pulse-labelling organisms with a mixture of 14C-labelled amino acids followed by sodium dodecyl sulphate gel electrophoresis and autoradiography. Populations from an NH4+-repressed, SO4 2−-limited chemostat (0·46 mg dry wt m1−1), when released from NH4+ repression, simultaneously synthesized detectable quantities of the three nitrogenase polypeptides 45 min before acetylene-reducing activity was observed. Exposure of populations synthesizing nitrogenase to air or NH4+ (200 μg N m1−1) repressed synthesis of both component proteins simultaneously, the rate initially decreasing by half in 11 to 12 min; in the presence of NH4+ a second slower phase with an approximate half-life of 30 min was observed. With 5% O2 in N2 the half-lives for the decreases in the rates of synthesis were 30 min for the Fe protein and 33 min for the Mo-Fe protein. Oxygen also repressed nitrogenase in a glutamine synthetase constitutive derivative of K. pneumoniae (strain SK24) which escapes NH4+ repression. Regulation of nitrogenase by O2 may therefore be independent of glutamine synthetase.

Factors influencing the interaction between Candida albicans and the polyenoic antibiotics nystatin and amphotericin B have been investigated using a K+-specific electrode to measure polyene-mediated efflux of cellular K+. In batch cultures, sensitivity was a function of culture age. Using continuous (chemostat) cultures, the influence of growth-limiting substrate, specific growth rate, growth temperature and growth pH were examined. Carbon-limited cultures showed the highest sensitivity of those substrates tested, and susceptibility increased with growth rate. Within the range 22 to 42 °C, growth at lower temperatures resulted in increased sensitivity, whilst a similar trend was observed when the growth pH of cultures was reduced. Further, under the conditions tested, there were considerable variations in free intracellular K+ concentrations.