- Volume 42, Issue 3, 1966

SUMMARY: When concentrated purified influenza virus PR 8 was incubated at 37° with caseinase C in phosphate buffer at pH 7 and ionic strength 0.01 or 0.1, the neuraminidase activity was released from the virus particle and progressively destroyed. Under these conditions a material was released from virus which neutralized the haemagglutination inhibiting antibodies of anti-PR 8 rabbit serum. This material and neuraminidase, released by caseinase C, were both sedimented by centrifugation at 125,000 g. Neither was adsorbed to fowl red cells. After 4-hr incubation with caseinase C in phosphate buffer at pH 7 and ionic strength 0.1, the infectivity of influenza virus PR 8 suspension was considerably decreased, but its haemagglutinating activity was unchanged and remained bound to the virus particles. Under these conditions, most of the surface projections of the virus particles were removed.

SUMMARY: The intracellular concentrations of 14C-streptomycin accumulated by Escherichia coli strain B during growth in the presence of different extracellular concentrations of the compound have been measured. They increased logarithmically with time at rates proportional to the logarithm of the extracellular concentration; the growth rates declined linearly with time at rates which were also proportional to the extracellular concentrations. Thus the same decrease in growth rate resulted from different intracellular concentrations of streptomycin, according to the conditions of uptake. After removal of extracellular streptomycin, the growth rates remained constant for several hours, during which time 40-60% of the total cell-bound radioactivity was lost. Of the radioactivity lost from the organisms during growth after removal of extracellular streptomycin, 50-80% was recovered from culture filtrates but did not behave as streptomycin with respect to adsorption to charcoal.

SUMMARY: The intracellular concentrations of 14C-streptomycin accumulated by Escherichia coli strain B during growth in the presence of different extracellular concentrations of the compound have been measured. They increased logarithmically with time at rates proportional to the logarithm of the extracellular concentration; the growth rates declined linearly with time at rates which were also proportional to the extracellular concentrations. Thus the same decrease in growth rate resulted from different intracellular concentrations of streptomycin, according to the conditions of uptake. After removal of extracellular streptomycin, the growth rates remained constant for several hours, during which time 40-60% of the total cell-bound radioactivity was lost. Of the radioactivity lost from the organisms during growth after removal of extracellular streptomycin, 50-80% was recovered from culture filtrates but did not behave as streptomycin with respect to adsorption to charcoal.

SUMMARY: The resistance of 148 clinical isolates of Gram-negative bacteria to ampicillin and benzylpenicillin was determined in serial-dilution sensitivity tests, together with the extent of inactivation of the penicillins, and the production of 6-aminopenicillanic acid (6-APA). Many of the cultures were resistant to the penicillins and inactivated the compounds, but only one culture, a strain of Escherichia coli, showed penicillin acylase activity, as indicated by the production of 6-APA. However, the cultural conditions prevailing in serial-dilution tests, namely, stationary culture at 37°, although favourable for the functioning of penicillin acylases, were shown to be highly unfavourable for production of the enzyme by coliform bacteria.

SUMMARY: The resistance of 148 clinical isolates of Gram-negative bacteria to ampicillin and benzylpenicillin was determined in serial-dilution sensitivity tests, together with the extent of inactivation of the penicillins, and the production of 6-aminopenicillanic acid (6-APA). Many of the cultures were resistant to the penicillins and inactivated the compounds, but only one culture, a strain of Escherichia coli, showed penicillin acylase activity, as indicated by the production of 6-APA. However, the cultural conditions prevailing in serial-dilution tests, namely, stationary culture at 37°, although favourable for the functioning of penicillin acylases, were shown to be highly unfavourable for production of the enzyme by coliform bacteria.

SUMMARY: Suspensions of Chromatium D oxidized thiosulphate completely to sulphate (in the light) under anaerobic conditions in the presence of carbon dioxide. During the oxidation, intracellular sulphur accumulated transiently and sulphate production followed a biphasic pattern. Using inner-35 S-thiosulphate, the initial burst of sulphate production, which accounted for about half of the total yield, occurred principally at the expense of the inner (SO3-) atom of thiosulphate; there was no intracellular accumulation of labelled sulphur. The radioactivity from outer-35S-thiosulphate accumulated transiently within the organisms and was transferred to sulphate at a rate which was similar to the second phase of sulphate production; most of the outer (S-) atom therefore passed through the stage of endogenous sulphur. Extracts of the organism catalysed the cyanolysis of thiosulphate to give sulphite and thiocyanate. An assay for this enzyme, rhodanese, based on the spontaneous reduction of 2,6-dichlorophenol-indophenol by sulphite was developed. The enzyme was also detected in extracts of Athiorhodaceae. The enzyme was partially purified from extracts of Chromatium D and resolved into two active fractions. It is concluded that endogenous sulphur is an intermediate in the oxidation of thiosulphate by Chromatium D and that the cleavage of the S—S bond in the molecule is a key step in the oxidation process. Rhodanese, which catalyses this type of reaction, may be concerned in this cleavage.

SUMMARY: Suspensions of Chromatium D oxidized thiosulphate completely to sulphate (in the light) under anaerobic conditions in the presence of carbon dioxide. During the oxidation, intracellular sulphur accumulated transiently and sulphate production followed a biphasic pattern. Using inner-35 S-thiosulphate, the initial burst of sulphate production, which accounted for about half of the total yield, occurred principally at the expense of the inner (SO3-) atom of thiosulphate; there was no intracellular accumulation of labelled sulphur. The radioactivity from outer-35S-thiosulphate accumulated transiently within the organisms and was transferred to sulphate at a rate which was similar to the second phase of sulphate production; most of the outer (S-) atom therefore passed through the stage of endogenous sulphur. Extracts of the organism catalysed the cyanolysis of thiosulphate to give sulphite and thiocyanate. An assay for this enzyme, rhodanese, based on the spontaneous reduction of 2,6-dichlorophenol-indophenol by sulphite was developed. The enzyme was also detected in extracts of Athiorhodaceae. The enzyme was partially purified from extracts of Chromatium D and resolved into two active fractions. It is concluded that endogenous sulphur is an intermediate in the oxidation of thiosulphate by Chromatium D and that the cleavage of the S—S bond in the molecule is a key step in the oxidation process. Rhodanese, which catalyses this type of reaction, may be concerned in this cleavage.

SUMMARY: Sulphate was progressively replaced by tetrathionate as the end product of thiosulphate oxidation by suspensions of Chromatium D when the pH value was decreased from 7.3 to 6.25; tetrathionate was not itself oxidized to sulphate within this range. The effect of pH value on the oxidation of endogenous sulphur was less marked than on the production of sulphate from thiosulphate. Extracts of Chromatium D catalysed the oxidation of thiosulphate to tetrathionate in the presence of ferricyanide; pH 5.0 was the optimum of the purified enzyme. Tetrathionate inhibited growth and the complete oxidation of thiosulphate by suspensions. At pH 6.75 where sulphur accumulated during thiosulphate utilization and sulphate and tetrathionate were formed, added tetrathionate inhibited the accumulation of endogenous sulphur but not the production of tetrathionate from thiosulphate. Tetrathionate had no effect on the oxidation of endogenous sulphur by suspensions. It is concluded that in Chromatium D thiosulphate is metabolized by two alternative pathways depending on the conditions: (1) the cleavage of one molecule of thiosulphate in a reaction similar to that catalysed by thiosulphate sulphur-transferase E.C.2.8.1.1. (rhodanese); (2) the oxidative coupling of two molecules of thiosulphate to give tetrathionate, catalysed by the thiosulphate-oxidizing enzyme which only operates at low pH values.

SUMMARY: Sulphate was progressively replaced by tetrathionate as the end product of thiosulphate oxidation by suspensions of Chromatium D when the pH value was decreased from 7.3 to 6.25; tetrathionate was not itself oxidized to sulphate within this range. The effect of pH value on the oxidation of endogenous sulphur was less marked than on the production of sulphate from thiosulphate. Extracts of Chromatium D catalysed the oxidation of thiosulphate to tetrathionate in the presence of ferricyanide; pH 5.0 was the optimum of the purified enzyme. Tetrathionate inhibited growth and the complete oxidation of thiosulphate by suspensions. At pH 6.75 where sulphur accumulated during thiosulphate utilization and sulphate and tetrathionate were formed, added tetrathionate inhibited the accumulation of endogenous sulphur but not the production of tetrathionate from thiosulphate. Tetrathionate had no effect on the oxidation of endogenous sulphur by suspensions. It is concluded that in Chromatium D thiosulphate is metabolized by two alternative pathways depending on the conditions: (1) the cleavage of one molecule of thiosulphate in a reaction similar to that catalysed by thiosulphate sulphur-transferase E.C.2.8.1.1. (rhodanese); (2) the oxidative coupling of two molecules of thiosulphate to give tetrathionate, catalysed by the thiosulphate-oxidizing enzyme which only operates at low pH values.

SUMMARY: Strains classified as Lipomyces lipoferus and L. starkeyi produce specific extracellular acidic heteropolysaccharides. The polymer from L. lipoferus contains mannose and glucuronic acid; the polymer from L. starkeyi contains, in addition, galactose and a trisaccharide. This trisaccharide is composed of mannose and glucuronic acid in the molar ratio 1:2. The difference in polysaccharide composition may be the most significant physiological basis yet discovered for separation of species in the genus Lipomyces.

SUMMARY: Strains classified as Lipomyces lipoferus and L. starkeyi produce specific extracellular acidic heteropolysaccharides. The polymer from L. lipoferus contains mannose and glucuronic acid; the polymer from L. starkeyi contains, in addition, galactose and a trisaccharide. This trisaccharide is composed of mannose and glucuronic acid in the molar ratio 1:2. The difference in polysaccharide composition may be the most significant physiological basis yet discovered for separation of species in the genus Lipomyces.

SUMMARY: The following effects of different partial pressures of oxygen p(O2) on the growth and on the oxidation of ammonium to nitrite by Nitrosocystis oceanus in liquid and on solid sea-water media were observed. Nearly pure O2 was toxic to the organism when it was exposed to the oxygen on agar medium. In normal air the growth on agar medium was variable; usually a large proportion of the inoculum organisms never started to divide. At low p(O2) values the formation of microcolonies took place. In liquid medium the organism was less affected by high p(O2) values than on agar. The lower limit of oxygen concentration which permitted oxidation of ammonium in liquid medium corresponded to about 0.05 ml. O2/l. In respiration experiments a high p(O2) in the gas phase resulted in twice as much oxygen uptake at the beginning as from air. Respiration at low p(O2) was comparatively high. These results seem to indicate that the inorganic respiration is accelerated by an increased p(O2) whereas growth is inhibited. Possible explanations are discussed.

SUMMARY: The following effects of different partial pressures of oxygen p(O2) on the growth and on the oxidation of ammonium to nitrite by Nitrosocystis oceanus in liquid and on solid sea-water media were observed. Nearly pure O2 was toxic to the organism when it was exposed to the oxygen on agar medium. In normal air the growth on agar medium was variable; usually a large proportion of the inoculum organisms never started to divide. At low p(O2) values the formation of microcolonies took place. In liquid medium the organism was less affected by high p(O2) values than on agar. The lower limit of oxygen concentration which permitted oxidation of ammonium in liquid medium corresponded to about 0.05 ml. O2/l. In respiration experiments a high p(O2) in the gas phase resulted in twice as much oxygen uptake at the beginning as from air. Respiration at low p(O2) was comparatively high. These results seem to indicate that the inorganic respiration is accelerated by an increased p(O2) whereas growth is inhibited. Possible explanations are discussed.

SUMMARY: The passive haemagglutination test and the sensitivity of the haemagglutination inhibition test were investigated by using two plant viruses (tobacco mosaic virus and latent carnation virus) and an insect virus, tipula iridescent virus. The minimum quantities detectable by these methods were of the order of 0.26 to 0.015 μg. There was no direct agglutination by tobacco mosaic virus or tipula iridescent virus of erythrocytes from a variety of animals.

SUMMARY: The passive haemagglutination test and the sensitivity of the haemagglutination inhibition test were investigated by using two plant viruses (tobacco mosaic virus and latent carnation virus) and an insect virus, tipula iridescent virus. The minimum quantities detectable by these methods were of the order of 0.26 to 0.015 μg. There was no direct agglutination by tobacco mosaic virus or tipula iridescent virus of erythrocytes from a variety of animals.

SUMMARY: Fifty strains of Staphylococcus aureus resistant to penicillin or tetracycline or both were examined for loss of these two resistances as the result of growth at 43-44°. Twelve strains showed a loss of penicillin resistance under the experimental conditions, and of these twelve, three showed a loss of tetracycline resistance. The two resistances were lost independently; in strains in which both resistances were lost most of the sensitive variants had lost one or other resistance but not both. However, penicillin resistance was lost only in strains that were also tetracycline-resistant and vice versa. All strains in which a loss of resistance occurred had similar phage-typing patterns and all belonged to the ‘52, 52A, 80, 81 complex’ of strains. Not all strains in the complex, however, showed a loss of resistance. Both penicillin and tetracycline resistance were transduced into suitable sensitive recipients. The results showed that the heat-sensitivity of the transduced resistance was the same as in the donor in which the transducing phage was propagated. The results are consistent with the hypothesis that these two resistances are carried by two separate plasmids, at least in certain strains of S. aureus.

Nine multiply-resistant strains of Staphylococcus aureus were examined for loss of resistance to antibiotics other than penicillin and tetracycline as the result of growth at elevated temperatures. These other antibiotics included streptomycin, erythromycin, novobiocin, oleandomycin, neomycin and bacitracin. In no case was any loss of resistance observed.

SUMMARY: Fifty strains of Staphylococcus aureus resistant to penicillin or tetracycline or both were examined for loss of these two resistances as the result of growth at 43-44°. Twelve strains showed a loss of penicillin resistance under the experimental conditions, and of these twelve, three showed a loss of tetracycline resistance. The two resistances were lost independently; in strains in which both resistances were lost most of the sensitive variants had lost one or other resistance but not both. However, penicillin resistance was lost only in strains that were also tetracycline-resistant and vice versa. All strains in which a loss of resistance occurred had similar phage-typing patterns and all belonged to the ‘52, 52A, 80, 81 complex’ of strains. Not all strains in the complex, however, showed a loss of resistance. Both penicillin and tetracycline resistance were transduced into suitable sensitive recipients. The results showed that the heat-sensitivity of the transduced resistance was the same as in the donor in which the transducing phage was propagated. The results are consistent with the hypothesis that these two resistances are carried by two separate plasmids, at least in certain strains of S. aureus.

Nine multiply-resistant strains of Staphylococcus aureus were examined for loss of resistance to antibiotics other than penicillin and tetracycline as the result of growth at elevated temperatures. These other antibiotics included streptomycin, erythromycin, novobiocin, oleandomycin, neomycin and bacitracin. In no case was any loss of resistance observed.

SUMMARY: Spherical protoplast-like bodies were released from the mycelia of Phytophthora cinnamomi and P. parasitica on digestion of their hyphal walls with an extracellular enzyme preparation from Streptomyces sp. in media of appropriate osmotic pressure. Protoplast liberation was observed when using as osmotic stabilizer either sucrose or mannitol (0.2 M-0.8 M); maximum response at about 0.4 M-0.6 M. Although hyphal wall digestion was always extensive, the amount of protoplast release varied widely under seemingly similar conditions. Protoplasts were liberated from the phytophthora mycelium by two mechanisms: bud-like emergence in which cytoplasm was squeezed through a small pore in the wall usually located at or near a hyphal apex; or intercalary hyphal swelling, which consisted of the swelling and rounding off of short hyphal segments whose walls became progressively thinner until they were no longer visible. Both types of protoplast were osmotically sensitive and disintegrated upon dilution of the suspending media.

SUMMARY: Spherical protoplast-like bodies were released from the mycelia of Phytophthora cinnamomi and P. parasitica on digestion of their hyphal walls with an extracellular enzyme preparation from Streptomyces sp. in media of appropriate osmotic pressure. Protoplast liberation was observed when using as osmotic stabilizer either sucrose or mannitol (0.2 M-0.8 M); maximum response at about 0.4 M-0.6 M. Although hyphal wall digestion was always extensive, the amount of protoplast release varied widely under seemingly similar conditions. Protoplasts were liberated from the phytophthora mycelium by two mechanisms: bud-like emergence in which cytoplasm was squeezed through a small pore in the wall usually located at or near a hyphal apex; or intercalary hyphal swelling, which consisted of the swelling and rounding off of short hyphal segments whose walls became progressively thinner until they were no longer visible. Both types of protoplast were osmotically sensitive and disintegrated upon dilution of the suspending media.