- Volume 82, Issue 2, 1974

Summary: A culture of Staphylococcus aureus harbouring a temperature-sensitive plasmid carrying genes for resistance to penicillin and cadmium ions, loses this plasmid and the associated phenotype when grown at the restrictive temperature. However, incubation at the restrictive temperature in the presence of cadmium ions results in a small number of cadmium-resistant cells. The properties of these surviving cells have been investigated and evidence is presented in support of the hypothesis that at least some of these cadmium-resistant cells are the result of integration of the plasmid into some other replicon, possibly the chromosome.

SUMMARY: Mutants of Salmonella typhimurium lt2 requiring acetate or succinate for aerobic growth on glucose were isolated. One acetate-requiring mutant, s8, lacked the activity of the overall pyruvate dehydrogenase complex due to a deficiency in the pyruvate dehydrogenase component (E1p) and was therefore designated an aceE mutant. Another mutant, s6, which required succinate (or lysine plus methionine), lacked activities of the overall α-ketoglutarate dehydrogenase complex and the α-ketoglutarate dehydrogenase component (E1 kg) and was designated a sucA mutant. Genetic studies with these mutants established that like Escherichia coli k12, Salmonella typhimurium lt2 has an aceE gene linked to aziA and aroP thus: leu-aziA-aroP-aceE, and a sue A gene linked to nadA thus: sucA-nadA-gal.

Two deletion strains, sm16 and sm51, which required acetate, or better, acetate plus lysine plus methionine, were found to lack the overall activities of both α-keto acid dehydrogenase complexes. Biochemical and immunological studies showed this to be due to deficiences in pyruvate dehydrogenase and dihydrolipoyl- transacetylase (the E1p and E2p components of the pyruvate complex) and failure to synthesize lipoamide dehydrogenase (the E3 components of both complexes). These deletion strains also behaved as if they lacked the general aromatic permease (aroP), and sm51, which requires nicotinate, was probably deleted for the nadC gene. Genetic studies confirmed that these strains were deleted in the leu-aziA-nadC- aroP-aceE,aceF,lpd-pan region. The results also indicated that the gene-protein- relationships of the α-keto acid dehydrogenase complexes are similar in S. typhimurium and E. coli.

Summary: Coprinus lagopus is inhibited by the analogue of phenylalanine, p-fluorophenyl-alanine (pFPA). Mutant strain f 7, selected for growth on pFPA, is a slow grower which is not inhibited by the analogue. Over a prolonged period of growth, the amount of [14C] (pFPA incorporated into the trichloroacetic acid-insoluble fraction of resistant f 7 mycelium is significant, but is less than 20 % of the amount incorporated into similar fractions of the mycelium of the parental sensitive strain tc1. Resistant f 7 grown on minimal medium, excretes into the medium p-hydroxy-phenylpyruvic acid, phenylpyruvic acid and a number of unidentified phenolic compounds. The sensitive tci strain grown on minimal medium does not excrete, but when grown on media supplemented with phenylalanine or shikimate excretes traces of these compounds. The total biosynthesis of phenylalanine over 24 h by sensitive strain tc1 is greatly reduced by the presence of exogenously supplied phenylalanine. No such reduction in synthesis is observed in the resistant mutant f 7. It is inferred that the mutation impairs the regulation of phenylalanine biosynthesis, probably by the loss of feedback control at either the 3-deoxy-d-amftmo-heptulo-sonate 7-phosphate (DAHP) synthetase or chorismate mutase.

Summary: Seven mutants which have low activities of vegetative (phosphate-repressible) alkaline phosphatase have been mapped by transduction crosses. These mutations do not affect the production of the alkaline phosphatase associated with sporulation. All the evidence available suggests that the activities produced in phosphate- starvation and during sporulation are attributable to the same protein and derive, presumably, from a common structural gene. It thus appears that alkaline phosphatase has distinct sporulation and vegetative controls and that all the mutations so far examined have been of the regulator type. The position on the chromosome of the structural gene is still in doubt.

Summary: We have developed a rapid and simple technique for examining chromosomes of the cellular slime mould Dictyostelium discoideum, based on standard cytological techniques involving fixation in methanol/acetic acid and staining with Giemsa stain. The haploid chromosome set is analysed. Four diploid strains are examined cytologically in order to determine their ploidal stability and to determine the pathway of haploidization in the parasexual cycle. In the absence of cloning, diploid strains eventually revert to haploid. Haploidization proceeds via transient aneuploidy. Aneuploids of all classes, n +1 through to 2n + 2, have been demonstrated cytologically. It is concluded that aneuploids can divide but that they are unstable and readily lose chromosomes to form aneuploids of lower chromosome numbers and eventually haploids. Hence, the process of haploidization is similar to that found in the parasexual cycle of Aspergillus nidulans.

Summary: Antigens of 32 ineffective, antibiotic-resistant or antimetabolite-resistant mutants from two effective parent strains of Rhizobium trifolii (t1 and t42) were cross-reacted with antiserum prepared against the parent and six other strains of R. trifolii. Fifteen of the mutants were compared with the parent strains by gel- electrophoresis of soluble and cell envelope (bacterial wall plus plasma membrane) proteins and structural examination of nodules. Six mutants of t1 and three mutants of t42 appeared to have an altered cell envelope and released internal antigens more readily from intact rhizobia than did the parent strains. The same six mutants of t1 formed tumour-like growths on Trifolium pratense roots which contained no infection-threads or rhizobia. The three mutants of t42 formed ineffective nodules which differed in structure. Ineffective nodules produced by the other mutants were defective at various stages of development. No qualitative differences in soluble or cell envelope proteins were found between mutants and parent strains, but the cell envelope fraction of some mutants differed quantitatively.

Summary: Factors have been concentrated from human serum and seminal plasma which, in the presence of glucose and at 37 °C, will induce germination and support mycelial development of blastospores of Candida albicans. The most active fraction from serum contained 98 % albumin but the inactivity of a number of commercial samples of albumin and comparison with the factor from seminal plasma indicated that the active component was probably not albumin. The active fraction from seminal plasma was of lower molecular weight than that from serum and had considerably greater germ-tube producing activity/mg protein. Mycelial development in serum declined after 24 h incubation but removal of cells at this time and reinoculation again gave rise to mycelial growth. An inhibitor of germination (but not of growth) accumulated after 3 to 6 days. Many non-peptide components of serum have been tested for germ-tube-producing activity with negative results.

SUMMARY: When Arthrobacter globiformis, A. simplex, A. crystallopoietes and A. atrocyaneus were grown in a chemostat, their morphology was related to growth rate. A transition from rod to coccus occurred at growth rates characteristic for each species; the higher the rate at which this transition occurred, the higher was the maximum specific growth rate. Organism size, concentration and viability were determined for A. globiformis at a range of dilution rates between 0·01 h−1 and washout. Yield remained constant except at low dilution rates, although the number of organisms varied because of the changes in their size and shape. The percentage of viable organisms in the chemostat remained almost constant at dilution rates above 0·1 h−1 (92 %) and was only slightly lowered even at dilution rates as low as 0·01 h−1.

Rods grown in the chemostat and then placed in nutrient-free solutions divided and produced cocci which survived for about 56 days. Cocci survived for about 70 days. The respiration rate of freshly harvested rods was higher than that of cocci but within two days both fell to low levels.

Summary: When exposed to light, cultures of Penicillium isariiforme showed a sharp decrease in citric acid accumulation in the medium compared with parallel cultures in continuous darkness, and a rise in the production of other low molecular weight intracellular intermediates, lipids, nucleic acids and protein. The significance of these phenomena is discussed.

SUMMARY: Streptococcus sanguis, S. bovis, S. mutans and S. salivarius, and also Escherichia coli for comparison, were grown separately in a chemostat under ammonia- and glucose-limitation. Bacterial extracts were assayed for ammonia-assimilating enzymes. For E. coli the level of glutamate dehydrogenase decreased and the levels of glutamine synthetase and glutamine (amide) : 2-oxoglutarate aminotransferase (glutamate synthase) increased following a change from glucose- to ammonia- limitation. In contrast, ammonia-limited streptococci contained much higher levels of glutamate dehydrogenase than glucose-limited streptococci, and although a glutamine synthetase was detected in S. bovis, glutamate synthase could not be detected in any of the streptococci. Low activity of a NAD-linked alanine dehydrogenase was found in S. bovis and S. sanguis. These results suggest that the only pathway of ammonia assimilation in streptococci involves glutamate dehydrogenase, irrespective of the nature of the growth limitation. Glutamate dehydrogenase extracted from the different species of Streptococcus had similar Michaelis constants and optimum pH values.

SUMMARY: The synthesis of trisporic acids, the sexual hormones, proceeds in a similar way in mated cultures of the heterothallic fungi Mucor mucedo and Blakeslea trispora. In both species precursors are secreted by both mating types and these can be converted to trisporic acids by the sexual partner. Precursor production is strongly stimulated by trisporic acids. In homothallic species the presence of biologically active substances can be demonstrated, which have characteristics similar to those found in heterothallic strains. Moreover, precursors of trisporic acids isolated from Mucor mucedo are readily converted to trisporic acids by homothallic species. These results suggest a close relationship between the hormonal systems of homothallic and heterothallic species.

SUMMARY: Of 53 steroids tested for their ability to induce the formation of antheridial branches in the water mould Achlya ambisexualis Raper, strain e87,34 were inactive, including 18 steroids having hormonal activity in mammals and six related to antheridiol, a C-29 steroid controlling the formation of antheridia in Achlya. The C-22, C-23 stereoisomers of antheridiol and two intermediates in its synthesis had about 10−3 to 10−6 the activity of antheridiol. Certain modifications in the side chain of antheridiol resulted in a loss of activity.

SUMMARY: During the transition from the mycelial to the yeast form of the dimorphic fungus Histoplasma capsulatum, RNA synthesis decreased to 26 ·7 % of the normal level for at least 4 h. There was a decrease in total RNA on the first day of mycelial → yeast transition. Starting on the second day, RNA synthesis in the mycelial → yeast transition phase resumed and this event was paralleled by the emergence of bud-like constrictions on hyphal tips.

SUMMARY: A strain of Claviceps fusiformis produced clavine alkaloid (principally agroclavine) in yields of 4 mg/ml within 6 days when grown in a sucrose-ammonium sulphate-inorganic salts medium in 400 l stirred fermenters. Alkaloid accumulated rapidly during the growth phase, most of the synthesis coinciding with a mycelial growth form consisting of plectenchymatic hyphae, the cells of which resembled the constituent cells of naturally occurring C. fusiformis sclerotia. The broth was rendered low in glucan by the activity of a α-glucanase and this, together with increased fermenter shaft speed, ensured that oxygen supply was not limiting throughout the fermentation. Alkaloid was extracted efficiently from the glucan- free culture filtrate by a solvent extraction procedure involving sequential transfer of the product into n-butanol, aqueous tartaric acid and chloroform, followed by crystallization from acetone. Improved alkaloid yield (6 mg/ml) was obtained by using a modified medium containing increased concentrations of magnesium sulphate, zinc sulphate and potassium dihydrogen orthophosphate, in a 400 l multi-stage fermentation.

SUMMARY: Mycobacterium lepraemurium multiplies in a cell-free liquid medium, referred to as NC-5, which is enriched Kirchner medium plus goat serum, α-ketoglutaric acid, cytochrome c, haemin, and l-cysteine. At 30 °C, the bacilli gradually elongated before multiplying 100- to 1000-fold. The maximum number of bacilli is reached after 8 weeks’ incubation. The generation time of Mycobacterium lepraemurium is between 8 and 14 days, depending upon the size of inoculum. Bacilli grown in NC-5 medium maintain their capacity to produce leprosy in mice. Optimum growth was with the basal medium at pH 7·3 using a small number of bacilli in the inoculum.