- Volume 87, Issue 1, 1975

SUMMARY: The intracellular esterases of 80 strains of Proteus and Providencia were analysed by the acrylamide-agarose zymogram technique using several synthetic substrates.

The esterase bands were classified in five main groups. The αA-esterase bands hydrolysed α-naphthyl acetate and were resistant or relatively insensitive to di-iso-fluoropropyl phosphate (DFP). The αB-esterase band hydrolysed both α-naphthyl acetate and α-naphthyl butyrate and were very sensitive to DFP. Both groups of esterase bands were inactivated by heat. The βA- and βB-esterase bands hydrolysed β-naphthyl acetate and were sensitive to DFP; these were distinguishable by the difference in their relative activity towards β-naphthyl butyrate and in their relative stability to heat. The αβ-esterase bands hydrolysed α- and β-naphthyl acetates and α- and β-naphthyl butyrates; they were inactivated by heat and were sensitive to DFP.

The distribution of these esterase bands among the strains of Proteus and Providencia and their electrophoretic patterns established esterase profile types which correlate with the classification based on traditional bacteriological tests. The degree of inter-strain similarity in esterase pattern varied highly among species. The homogeneity of Proteus mirabilis and especially of Providencia stuartii contrasted with the heterogeneity of other species. This disparity suggests that the bacteria of the tribe Proteae have not the same degree of intra-specific differentiation in physico-chemical properties of esterases.

SUMMARY: Keto acids and free amino acids were assayed in the cells and the medium of Escherichia coli B growing in the presence of chloramphenicol, cycloserines, aminooxyacetate, and limiting nitrogen source.

Under these growth-limiting conditions the cells accumulated ketoglutarate and ‘ketovaline’ but no other keto acids. In all experiments only ketoglutarate, pyruvate, and ‘ketovaline’ were found in the medium. Amino acids are released into the medium in the early phases of growth and the composition of the extracellular amino acids is similar to that of the amino acid pool. The concentrations of free amino acids were 103–104 times higher in the cell than in the medium. The internal pool composition is fixed under all growth-limiting conditions. In the presence of the drugs the cells release amino acids into the medium.

SUMMARY: Several mutants obtained from smooth Salmonella typhimurium strains by selection for resistance to Felix O (FO) phage [whose receptor site includes the N-acetylglucosamine branch of the lipopolysaccharide (LPS) core] were smooth in cultural properties, antigenic character and phage sensitivity pattern (except for their FO resistance). However, the affected genes of several such ‘FOR’ (FO-resistant) mutants were shown by transduction to map in the short cysE-pyrE segment, which includes nearly all known rfa genes responsible for synthesis of LPS core. All of seven FOR mutants differed from their parents, and resembled rfa mutants with defects in the deeper part of the LPS core, by increased sensitivity to various antibiotics. One FOR mutant was non-virulent (LD50 > 107, compared with < 100 for its parent); lt7 derivatives given this FOR gene by co-transduction with cysE + were likewise non-virulent. It is inferred that FOR mutations affect the assembly of the inner part of the LPS core, perhaps causing incomplete blocks in glycosyl transferase reactions.

SUMMARY

The sensitivity of Candida albicans, grown in batch culture at 37°C, to ampho-tericin methyl ester (AME), judged by the concentration of AME required to induce a standard rate of leakage of K+ from suspensions of organisms, decreased with the time of growth. Organisms in exponential growth were sensitive to 0·1 to 0·2 μg AME/ml while organisms in the stationary phase were resistant to 4 to 60 μg AME/ml, depending on the initial concentration of glucose in the medium and the length of time for which incubation had been continued. When the initial concentration of glucose was low (0·1%, w/v), the AME resistance rose during the early stationary phase and then, after 40 h incubation at 37°C, decreased again. Sphaeroplasts were prepared from organisms at different phases of growth and did not show these changes in AME sensitivity, but remained highly sensitive for growth up to 40 h.

Sphaeroplasts were prepared by treating suspensions of organisms with mercap-toethanol and then digesting with Streptomyces enzyme preparation. Addition of the material extracted by the digestion to suspensions of exponential-phase organisms or sphaeroplasts increased their AME resistance. Fractionation of the digest showed that the antagonistic material was contained in the neutral lipid fraction. Pure lipids fell into the following order of decreasing antagonism to AME when added together with the antibiotic to suspensions of exponential-phase organisms: sterol esters (ergosterol esters > cholesterol esters; unsaturated fatty acid esters > saturated fatty acid esters), sterols, triglycerides, unsaturated fatty acids, saturated fatty acids. The amount of antagonistic material released from stationary organisms was not markedly greater than that from exponential-phase organisms and analysis of the lipid content of wall preparations showed that the content of total lipid, neutral lipid and triglyceride of 40 h organisms was not more than 75,25 and 30%, respectively, greater than that of exponential-phase organisms.

The AME resistance of stationary-phase organisms decreased rapidly if suspensions were incubated with glucose or mercaptoethanol. The decrease in the presence of glucose was prevented by metabolic inhibitors, especially SH binding agents. Treatment of organisms with either iodoacetamide or N-ethylmaleimide gave a rapid increase in AME resistance, amounting in some cases to 5- to 15-fold. The effect of iodoacetamide decreased as the organisms passed into the stationary phase and their intrinsic resistance increased. Evidence is presented which suggests that the degree of reduction of SH groups in the cell surface is an important factor in determining AME resistance.

SUMMARY: Two hundred and thirty-three strains of coryneform bacteria, including representatives of the genera Arthrobacter, Brevibacterium, Cellulomonas, Corynebac-terium, Erysipelothrix, Jensenia, Kurthia, Listeria, Microbacterium, Mycobacterium, Nocardia and Propionibacterium and other related bacteria, were studied using 173 morphological, physiological and biochemical tests. The bacteria were grown on a soil extract medium which allowed growth of all the strains, and all were incubated at 30 °C. The results were subjected to computer analysis. The majority of the strains grouped into eight main clusters representing: (A) Lactobacillus, Listeria, Microbacterium thermosphactum and Streptococcus faecalis; (B) Erysipelothrix and Streptococcus pyogenes; (C) animal corynebacteria and Microbacterium flavum; (D) Cellulomonas and related bacteria; (E) Propionibacterium; (F) Arthrobacter, Brevibacterium, Kurthia and Mycobacterium rhodochrous; (G) plant pathogenic corynebacteria; (H) Nocardia. Based on these clusters, several recommendations are made regarding the classification of the coryneform area. (i) The members of clusters A and B at present placed in the Corynebacteriaceae would seem better moved to the Lactobacillaceae. (ii) The genus Corynebacterium would best be retained for the type species C. diphtheriae, closely related animal corynebacteria and Micro. flavum. (iii) Cellulomonas and Propionibacterium are distinct taxa more closely related to Corynebacterium than to either Arthrobacter or Lactobacillus. (iv) Clusters F and G are evidently heterogeneous. In particular the positions of Kurthia and the plant pathogenic corynebacteria are unclear. Arthrobacter is a large loose taxon and it is premature to decide on its taxonomic rank. The genus Brevibacterium should be retained for B. linens and closely related strains. (v) The cellulolytic forms of Nocardia should be removed from the genus; they are however quite distinct from Cellulomonas.