New Method of Isolating Salmonellae from Milk

The use of a cotton gauze swab and subsequent culture of the swab was found to be a more sensitive method for isolating Salmonella from liquid milk than the revised procedure of North. The swab method was found to be as sensitive as the North procedure for recovering Salmonella when incubated at 37 C but more sensitive when incubated at 43 C. Incubation of the swab cultures at the elevated temperature of 43 C gave good results when Salmonella was present at levels as low as one per liter. Swabs exposed to milk contaminated with 100 Salmonella per liter remained positive even when subsequently washed for 2 hr in noncontaminated milk. Bismuth sulfite agar and Brilliant Green sulfadiazine agar were equally effective for isolating Salmonella from broth cultures; use of both media resulted in maximal isolations.     

The number of Na:K pump sites on red blood cells from HK and LK lambs

Lambs of known genotype with respect to the locus determining cation composition of red cells were obtained by selective matings. Numbers of K⁺ pump sites per cell were determined on HK and LK lambs 10–20 days postnatal by simultaneously determining [³H]ouabain binding and inhibition of active K⁺ transport. Red cells from HK lambs were indistinguishable from adult HK cells with regard to the K⁺ pump flux and number of pump sites. Cells from genetically LK lambs had pump fluxes and numbers of pump sites intermediate between those from adult HK and LK sheep. The results suggest that the change in cation composition and in the K⁺ pump during the first 60 days in genetically LK lambs can be correlated with a reduced number of K⁺ pump sites.     

Immunological and Biophysical Separation of Dengue-2 Antigens

Antigenic compositions of slowly sedimenting dengue-2 hemagglutinin (SHA) and soluble complement-fixing antigen (SCF) were compared with the virion (rapidly sedimenting hemagglutinin, RHA) by radioimmune precipitation (RIP), RIP inhibition, kinetic neutralization, and neutralization blocking tests with the use of hyperimmune mouse ascitic fluids. RHA and SHA were unable to inhibit completely the RIP of each other by anti-RHA, and neutralization by anti-RHA was not blocked by SHA. This indicated that SHA is serologically related, but not identical, to RHA. SHA differed from RHA in that SHA lacked the “core” polypeptide but contained the two envelope polypeptides. In addition, SHA contained a polypeptide with a molecular weight of 16,500 daltons and a suggestion of several other proteins. These data, when considered with other evidence, suggest that SHA is a special form of “incomplete virus.” SCF was unable to inhibit the RIP of SHA or RHA or to block neutralizing antibodies. Further, anti-SCF did not neutralize RHA or precipitate significant levels of SHA or RHA. Polyacrylamide gel electrophoresis separated SCF from structural polypeptides by molecular size. This evidence suggests that SCF is a nonstructural antigen.     

Histidyl-Transfer Ribonucleic Acid Synthetase in Positive Control of the Histidine Operon in Salmonella typhimurium

Autolysin-like enzymes appear to be responsible for cell separation in Agmenellum quadruplicatum. Mutants that are impaired in cell separation and grow as chains exhibit reduced cell lytic activity. Lysozyme, extracted autolysin, and antibiotics that affect peptidoglycan synthesis phenotypically suppress chain formation. Various aspects of the regulation of the cell separation process were also examined. Studies involving antibiotic inhibitors of macromolecular synthesis and general growth inhibitors provided no evidence for the active regulation of the cell separation process during the latter portion of the division cycle. Evidence was obtained, however, for the partial restriction of peptidogly-can hydrolysis by unknown secondary modifications. The thin electron-dense layer of peptidoglycan along the sides of cells was much more resistant to hydrolysis by egg-white lysozyme than was the septum between daughter cells. The middle portion of the septum was more sensitive than was the layer immediately adjacent to the cytoplasmic membrane. Under conditions that would not osmotically stabilize spheroplasts, lysozyme facilitates rapid cell separation in chain-forming mutants with little leakage of cellular protein or loss of viability.     

Culture Medium for Enterobacteria

A new minimal medium for enterobacteria has been developed. It supports growth of Escherichia coli and Salmonella typhimurium at rates comparable to those of any of the traditional media that have high phosphate concentrations, but each of the macronutrients (phosphate, sulfate, and nitrogen) is present at a sufficiently low level to permit isotopic labeling. Buffering capacity is provided by an organic dipolar ion, morpholinopropane sulfonate, which has a desirable pK (7.2) and no apparent inhibitory effect on growth. The medium has been developed with the objectives of (i) providing reproducibility of chemical composition, (ii) meeting the experimentally determined nutritional needs of the cell, (iii) avoiding an unnecessary excess of the major ionic species, (iv) facilitating the adjustment of the levels of individual ionic species, both for isotopic labeling and for nutritional studies, (v) supplying a complete array of micronutrients, (vi) setting a particular ion as the crop-limiting factor when the carbon and energy source is in excess, and (vii) providing maximal convenience in the manufacture and storage of the medium.     

Reactions of polycyclic hydrocarbon epoxides with RNA and polyribonucleotides

RNA, poly(G) and poly(A) were reacted with benz[a]anthracene 5,6-oxide or with 7,12-dimethylbenz[a]anthracene 5,6-oxide and hydrolysates of the alkylated polymers examined using a combination of Sephadex LH20 column chromatography and thin-layer chromatography on silica gel. The results show that two RNA products are formed in reactions with benz[a]anthracene 5,6-oxide, one resulting from reaction with guanine and the other from reaction with adenine. With 7,12-dimethylbenz[a]anthracene 5,6-oxide, six RNA products appeared to be formed, two resulting from reactions with guanine and three from alkylation of adenine; the other product has not been identified.     

Restricted pH ranges and reduced yields for bacterial growth under pressure

Hydrostatic pressure was found to cause a marked narrowing of pH ranges for growth and reductions in growth yields for a variety of bacteria. In many cases, reduced yields under pressure could be directly related to increased sensitivities to metabolic acids that accumulated in the enclosed culture vessels used. Magnesium and calcium ions partially reversed increases in sensitivities of representative gram-positive and gram-negative bacteria to low, but not high, pH. Growth inhibition of these organisms at both extremes of pH was associated with enhanced loss of K⁺ from pressurized cells. Inhibited cells in alkaline media also lysed under pressure, but microscopically observable lysis was clearly a secondary phenomenon because it occurred slowly. Apparent volumes for growth-inhibitory protonation-deprotonation reactions were calculated on the basis of measured shifts in inhibitory pH with pressure. The values ranged from 99 to 431 ml/mole, and their magnitudes indicated that growth inhibition by acids or bases involves cooperative changes in polymeric interactions such as those which accompany protein denaturation.     

Recombination of human erythrocyte apoprotein and lipid. II. Visualization of apoprotein-lipid bilayer complex by freeze-etching

Bilayers of human erythrocyte apoprotein-lipid complexes were made by dipping a mica plate through monolayers of the complex formed at the air-water interface. Stearic acid and erythrocyte lipid alone served as controls. Freeze-fracture images of the complex at high lipid surface pressures (30 dynes/cm) showed particles (average diameter, 109 Å ± 18 Å) similar to those of erythrocyte ghosts (average diameter, 102 Å ± 19 Å). Control surfaces were smooth. We conclude that part or all of the protein molecule penetrated into the lipid bilayer and that erythrocyte apoprotein-lipid complexes yield fracture faces similar to the native erythrocyte membrane.     

Effects of Estrogen on Glucose Uptake by Rat Muscle

The isolated rat diaphragm was used to study the effects of 17β-estradiol on basal and insulin-mediated glucose uptake. Rats were injected with estradiol for 2 wk in daily doses of 10 μg/100 g of body weight and were compared to untreated control animals. Estrogen treatment resulted in a 16% decrease in basal glucose uptake by diaphragm muscle as compared to controls. In contrast, in the presence of insulin, glucose uptake by muscle increased 103% above basal in estradiol-treated animals as compared to a 38% rise in the control group. The absolute rate of glucose uptake induced by insulin in the estradiol treated animals (5.8 mg/g/hr) was 22% higher than in controls. These findings were not accompanied by changes in weight gain, plasma glucose and plasma immunoreactive insulin concentrations in the treated animals. In vitro incubation of diaphragm muscle with estradiol did not have an effect on basal or insulin-mediated glucose uptake.