Isolation and characterization of phosphothioredoxin from Excherichia coli. Thioredoxin was isolated as a phosphoprotein from actively growing cultures of Escherichia coli. Labeling was performed in vivo by growing cells in the presence of 32P-labeled inorganic phosphate, and phosphothioredoxin was purified in one step by immunoabsorption to a thioredoxin antibody column. The stoichiometry of phosphate bound was 0.7 to 0.8 mol of phosphate/mol of thioredoxin. The phospho-amino acid linkage was identified as a thiol phosphate by several criteria: (a) the maximum lability of the phosphate bond was between pH 2.5 and 3.5 (t1/2 (37 degrees) = 200 h (pH 7 to 8); 0.4 h (pH 3.0); 200 h (pH 1.0)); (b) the phosphate linkage was very labile in the presence of iodine at neutral pH (t1/2 less than 1 min); and the phosphopeptide was identified as Cys32-Gly-Pro-Cys35-Lys, the same sequence previously implicated as the active site for disulfide-linked oxidation-reduction reactions. Phosphate was distributed on either cysteine, with 60% of the phosphate bound to cysteine32. Results are discussed in terms of the possible role of phosphothioredoxin as an intermediate in phosphotransferase reactions.


Isotope effects in 3-dehydroquinate synthase and dehydratase. Mechanistic implications. The mechanism of 3-dehydroquinate synthase was explored by incubating partially purified enzyme with mixtures of [1-14C]3-deoxy-D-arabino-heptulosonic acid 7-phosphate (DAHP) and one of the specifically tritiated substrates [4-3H]DAHP, [5-3H]DAHP, [6-3H]DAHP, (7RS)-[7-3H]DAHP, (7R)-[7-3H]DAHP, or (7S)-[7-3H]DAHP. Kinetic and secondary 3H isotope effects were calculated from 3H:14C ratios obtained in unreacted DAHP, 3-dehydroquinate, and 3-dehydroshikimate. 3H was not incorporated from the medium into 3-dehydroquinate, indicating that a carbanion (or methyl group) at C-7 is not formed. A kinetic isotope effect kH/k3H of 1.7 was observed at C-5, and afforded support for a mechanism involving oxidation of C-5 with NAD. A similar kinetic isotope effect was found at C-6 owing to removal of a proton in elimination of phosphate, which is reasonably assumed to be the next step in 3-dehydroquinate synthase. Hydrogen at C-7 of DAHP was not lost in the cyclization step of the reaction, indicating that the enol formed in phosphate elimination participated directly in an aldolase-type reaction with the carbonyl at C-2. In the dehydration of 3-dehydroquinate to 3-dehydroshikimate the (7R) proton from (7RS)- or (7R)-[7-3H]DAHP is lost, indicating that the 7R proton occupies the 2R position in dehydroquinate. Hence the cyclization step occurs with inversion of configuration at C-7. A kinetic isotope effect kH/k3H = 2.3 was observed in the conversion of (2R)-[2-3H]dehydroquinate to dehydroshikimate. Hence loss of a proton from the enzyme-dehydroquinate imine contributed to rate limitation in the reaction.


Purification and properties of a major cytochrome b peptide from baker's yeast. A major cytochrome b peptide was purified from yeast mitochondria by a procedure involving solubilization in deoxycholic and cholic acids, ammonium sulfate fractionation, proteolytic digestion, and sucrose gradient centrifugation in the presence of Tween 80. The homogeneity of the purified protein was established by the criteria that the product was spectrally pure and yielded a single band on both sodium dodecyl sulfate polyacrylamide gel electrophoresis, and by gel isoelectric focusing. The purified cytochrome b polypeptide had absorption maxima at 562, 532, and 430 nm in the reduced form and at 525 to 570 nm and 419 nm in the oxidized form. The reduced minus oxidized difference spectra revealed absorption bands at 562, 532, and 430 nm at room temperature and 559, 529, and 429 nm at 77 K, respectively. The heme group was identified as protoheme by formation of the reduced pyridine hemochromogen. Treatment of the reduced form with carbon monoxide affected the absorption spectrum, indicating that the isolated hemoprotein was modified compared to native cytochrome b. The apparent molecular weight of the preparation was 28,000 based on sodium dodecyl sulfate polyacrylamide-gel electrophoresis and 28,800 based on sucrose gradient centrifugation. The isolated cytochrome b polypeptide showed a strong tendency to aggregate.


Inactivation of uridine nucleosidase in yeast. Purification and properties of an inactivating protein. It has been previously demonstrated in our laboratory that uridine nucleosidase (EC 3.2.2.3) is subjected in yeast to inactivation. An inactivating fraction has been isolated and purified to homogeneity with a procedure which includes gel filtration, adsorption chromatography, and electrofocusing techniques. The molecular weight of the enzyme, estimated either by sodium dodecyl sulfate disc gel electrophoresis or by gel filtration is approximately 44,000. No quaternary structure was evidenced. The inactivating activity possesses proteolytic activity against casein and hemoglobin with pH optima of 2.5 and 3.2, respectively. The optimal pH for uridine nucleosidase inactivation is around 4.7. The inactivating activity as well as the proteolytic activity of the preparation can be inhibited by IA but not by IB2 and IC, yeast macromolecular inhibitors for proteinase A (EC 3.4.23.8), B (EC 3.4.22.9), and C (EC 3.4.12.8), respectively. The apparent isoelectric point is pH 4.03. The carbohydrate content is 8.5%. A comparison of the properties of the inactivating protein with those of known yeast proteinases leads to the conclusion that it is identical with the enzyme previously designated as proteinase A, which for the first time has been obtained homogeneous and characterized. It has been shown that proteinase A could play a physiological role in the uridine nucleosidase inactivation process when it is associated, as a complex, with proteinase B.


Effects of sulfhydryl reagents on the binding and release of penicillin G by D-alanine carboxypeptidase IA of Escherichia coli. Purified D-alanine carboxypeptidase IA of Escherichia coli is inhibited by penicillin G and binds penicillin G reversibly. The binding of penicillin to the enzyme is relatively insensitive to sulfhydryl reagents, while release of penicillin from the enzyme is severely inhibited by these reagents. The inhibition of release parallels the inhibition of carboxypeptidase activity by the sulfhydryl reagents. In the presence of the sulfhydryl reagent p-chloromercuribenzoate, an acyl-enzyme intermediate, produced by the reaction of carboxypeptidase IA with diacetyl-L-lysyl-D-alanyl-D-alanine, accumulates and can be isolated. These results indicate that binding of penicillin to carboxypeptidase IA occurs by an acylation step of the carboxypeptidase reaction, while penicillin release occurs by a deacylation step of the reaction. Only the latter is inhibited by sulfhydryl reagents.


The threonine-sensitive homoserine dehydrogenase and aspartokinase activities of Escherichia coli K12. Carboxymethylation of the enzyme: threonine binding and inhibition are functionally dissociable. The inactivation of the aspartokinase I-homoserine dehydrogenase I by iodoacetic acid and the effect on the sensitivity to its inhibitor, L-threonine, were examined. Both aspartokinase and homoserine dehydrogenase inactivation, as well as the dehydrogenase desensitization toward L-threonine occur as a pseudo-first order process. During its inactivation, the aspartokinase remains sensitive to L-threonine. At 50% inactivation, the inhibition curve of the aspartokinase by L-threonine displays homotropic cooperative effects. This alkylated protein retains eight binding sites for L-threonine. During the carboxymethylation, the protein remains in the tetrameric form until half of the kinase activity is lost. At the end of the inactivation aggregate forms and dimers appear.


Binding of antibodies to acetylcholine receptors in Electrophorus and Torpedo electroplax membranes. Antisera against purified acetylcholine receptors from the electric tissues of Torpedo californica and of Electrophorus electricus were raised in rabbits. The antisera contain antibodies which bind to both autologous and heterologous receptors in solution as shown by an immunoprecipitation assay. Antibodies in both types of antisera bind specifically to the postjunctional membrane on the innervated surface of the intact electroplax from Electrophorus electric tissue as demonstrated by an indirect immunohistochemical procedure using horseradish peroxidase conjugated to anti-rabbit IgG. Only anti-Electrophorus receptor antisera, however, cause inhibition of the receptor-mediated depolarization of the intact Electrophorus electroplax. The lack of inhibition by anti-Torpedo receptor antibodies, which do bind, suggests that the receptor does not undergo extensive movement during activity. The binding of anti-Torpedo antibodies to receptor-rich vesicles prepared by subcellular fractionation of Torpedo electric tissue was demonstrated by both direct and indirect immunohistochemical methods using ferritin conjugates. These vesicles can be conveniently collected and prepared for electron microscopy on Millipore filters, a procedure requiring only 25 micrograms of membrane protein per filter. In addition, it was possible to visualize the binding of anti-Torpedo receptor antibodies directly, without ferritin. These anti-Torpedo receptor antibodies, however, do not inhibit the binding of acetylcholine or of alpha-neurotoxin to receptor in Torpedo microsacs but do inhibit binding of alpha-neurotoxin to Torpedo receptor in Triton X-100 solution. It is likely that the principal antigenic determinants on receptor are at sites other than the acetylcholine-binding sites and that inhibition of receptor function, when it occurs, may be due to a stabilization by antibody binding of an inactive conformational state.


Chlorotetracycline as a fluorescent Ca2+ probe in pancreatic islet cells. Pancreatic islets, or suspensions of islet cells, from noninbred ob/ob-mice were incubated with chlorotetracycline and analyzed for Ca2+-dependent fluorescence in a microscope. Unless logarithmically transformed, signals from islets were asymmetrically distributed with unstable variance. Signals from cells pelleted in glass capillaries were more homogeneous and depended linearly on the thickness of the sample. The effect of sample thickness and a significant enhancement of fluorescence by alloxan suggest that beta-cells were involved in producing the signal from whole islets. The signal from dispersed cells was probably diagnostic of Ca2+ in beta-cell plasma membranes because it was suppressed by La3+ and had a spectrum indicative of an apolar micromilieu; fluorescent staining of cell surfaces was directly seen at high magnification. Fluorescence from cells was enhanced by 0.5-10 mM Ca2+ in a dose-dependent manner, whereas less than 0.5 mM Ca2+ saturated the probe alone in methanol. The signal from islets or dispersed cells was suppressed by 5 mM theophylline; that from cells was also suppressed by 0.5 mM 3-isobutyl-1-methylxanthine, 1.2 or 15 mM Mg2+, 3-20 mM D-glucose, and, to a lesser extent, 20 mM 3-O-methyl-D-glucose. D-glucose was more inhibitory in the absence than in the presence of Mg2+, as if Mg2+ and D-glucose influenced the same Ca2+ pool. L-glucose, D-mannopheptulose, or diazoxide had no noticeable effect and 20 mM bicarbonate was stimulatory. The results suggest that microscopy of chlorotetracycline-stained cells can aid in characterizing calcium pools of importance for secretion. Initiation of insulin release may be associated with an increas


Freeze-fracturing in ultrahigh vacuum at -196 degrees C. Conventional freeze-etching is carried out in a vacuum of approximately 10(-6) torr and at a specimen temperature of -100 degrees C. The relatively poor topographic resolution of most freeze-etch replicas, and the lack of complementarity of morphological details in double replicas have been thought to be caused by structural distortions during fracturing, and radiation damage during replication. Both phenomena can be reduced by lowering the specimen temperature. To prevent condensation of residual gases (especially H2O) on the fracture faces at lower specimen temperature, an improved vacuum is required. Therefore, an ultrahigh vacuum freeze-fracture apparatus has been developed which allows fracturing and Pt/C-shadowing of specimens at -196 degrees C while maintaining a vacuum of 10(-9) torr. It consists of a modified Balzers BA 350 ultrahigh vacuum (UHV) unit, equipped with an airlock which enables the input of nonhoar-frosted specimens directly into the evacuated bell jar. A comparison of the paracrystalline plasmalemma structure in yeast cells portrayed by the conventional technique and by UHV-freeze-fracturing at -196 degrees C shows the improved topographic resolution which has been achieved with the new technique. The improvement is explained by less structural distortions during fracturing at lower temperatures. The particles of the paracrystalline regions on the P face are more regularly arranged and exhibit a craterlike substructure which corresponds with a ringlike depression in the E face. The optical diffraction patterns of these paracrystalline regions demonstrate the improvement of the structural record by showing well-defined third- and fourth-order spots.


External fibronectin of cultured human fibroblasts is predominantly a matrix protein. The distribution of a major glycoprotein (fibronectin) of human fibroblast cultures was studied in immunoelectron microscopy with peroxidase- or ferritin-labeled antibodies. External fibronectin was visualized in pericellular structures, in some areas on the growth substratum, and to a lesser degree in close association with the upper and lower surface membranes of the cell. The pericellular fibronectin-containing structures consisted of amorphous or vaguely fibrillar material forming strands or patches, 50-500 nm in diameter; the structures appeared to mediate distant cell-to-cell and cell-to-substrate contacts. When in close association with the plasma membrane, fibronectin markers were seen as discrete patches. The exact relationship between this form of fibronectin and the plasma membrane, however, remained open. Filamentous material was commonly seen in the cortical cytoplasm under patches of membrane-associated fibronectin. The distribution that we observed is consistent with the proposed roles of fibronectin in cell interactions with neighboring structures and with its presence in vivo as an extracellular glycoprotein in connective tissue matrix and basal laminae.


Intracellular localization of the high molecular weight microtubule accessory protein by indirect immunofluorescence. Microtubule accessory proteins were isolated from porcine brain microtubules by phosphocellulose chromatography, and the high molecular weight protein (HMW protein), purified from this microtubule-associated fraction by electrophoretic elution from SDS gels, was used to raise antisera in rabbits. In agarose double diffusion tests, the antiserum obtained forms precipitin lines with purified HMW protein but not with tau protein or tubulin. When rat glial cells (strain C6) are examined by indirect immunofluorescence, this serum specifically stains a colchicine-sensitive filamentous cytoplasmic network in interphase cells, a network indistinguishable from that seen when cells are treated with antitubulin serum. In dividing cells, specific staining of the mitotic spindle and the stem body is observed with the antiserum to HMW protein. These studies indicate that HMW protein, like tau protein, is associated with microtubules in intact cells.


Intracellular calcium as a modulator of transepithelial permeability to water in frog urinary bladder. The divalent cation ionophore A 23187 was used to evaluate the action of intracellular calcium on net transepithelial water movement across the isolated frog urinary bladder. Incubation with the ionophore increases the net basal water flux in a dose-dependent fashion but independent of the extracellular calcium concentration. Bladders pretreated with A 23187 and exposed thereafter to an increase in calcium concentration exhibit a water permeability that under certain conditions can be comparable to that achieved with antidiuretic hormone (ADH). Lowering the serosal calcium at the peak of the hydrosmotic responses to both ADH and A 23187 inhibited the maintenance of the net water flux. The action of a supramaximal dose of ADH is blunted in bladders pretreated with A 23187, while the hydrosmotic effects of a submaximal dose are enhanced when the ionophore is added together with the hormone. The results show that an increase in transepithelial water movement can be triggered by calcium and that serosal calcium is needed to sustain the response. This hydrosmotic response may be dependent upon the rate at which intracellular calcium concentrations change and on the absolute concentration attained. It is suggested that calcium is involved in the action of ADH on water permeability and may act as a modulator of the hydrosmotic response.


High-pressure liquid chromatographic analysis of diazepam, oxazepam and N-desmethyldiazepam in human blood. We describe a rapid method for precisely measuring concentrations of diazepam, oxazepam and N-desmethyldiazepam in blood by high-pressure liquid chromatography. The drugs, together with an internal standard, prazepam, are extracted from 2 ml of blood and analyzed isocratically on a reversed-phase column with a mobile phase consisting of acetonitrile-0.01 M sodium acetate buffer (35:65 v/v). The eluted drugs are detected by their absorption at 240 nm. The sensitivity of this method is 30 microgram/l for oxazepam and N-desmethyldiazepam, 40 microgram/l for diazapam, for 2-ml blood samples. Relative recovery of added drugs to blood varies from 91 to 110%. The day-to-day precision (coefficient of variation) established by 10 replicate analyses was 2.8 to 9.6%.


High-performance liquid chromatographic determination of benzodiazepines in human plasma. A simple and sensitive method for determination of benzodiazepines in plasma has been developed using high-performance liquid chromatography in a reverse-phase mode. The method is illustrated by application to plasma samples containing diazepam and N-desmethyldiazepam at concentrations which would be encountered during therapy, with limits of detection of 10 ng/ml and 2 ng/ml for diazepam and N-desmethyldiazepam, respectively.


Use of Autobac 1 for rapid assimilation testing of Candida and Torulopsis species. We devised a system of presumptive identification of some yeasts that uses the Autobac 1 (Pfizer Inc.) instrument to detect carbon assimilation by 218 strains of Candida and Torulopsis. This system compared favorably with a conventional system of yeast identification and also with the Uni-Yeast-Tek (Corning Medical) and API (Analytab Products Inc.) methods.


Evaluation of three test procedures for identification of Staphylococcus aureus from clinical sources. A total of 520 clinical and environmental isolates of the family Micrococcaceae that fermented glucose anaerobically were tested for their ability to produce coagulase, thermostable nuclease, and deoxyribonuclease. Of these, 450 isolates coagulated rabbit plasma, produced thermostable nuclease, and were identified as Staphylococcus aureus, 447 of which produced a 3+ to 4+ clot. The remaining three isolates produced a 2+ clot, deoxyribonuclease, and thermostable nuclease. It was found that three of the S. aureus isolates failed to produce deoxyribonuclease. A total of 70 isolates which did not coagulate rabbit plasma and which were thermostable nuclease negative were identified as S. epidermidis. Three of them produced deoxyribonuclease. It is suggested that the thermostable nuclease test be performed on all isolates producing a 2+ (or 1+) clot in the coagulase test before identifying them as S. aureus.


Pigment production by Cryptococcus neoformans and other Cryptococcus species from aminophenols and diaminobenzenes. Cryptococcus neoformans and other Cryptococcus species can produce pigment(s) from many aminophenol and diaminobenzene compounds. Pigment production from these compounds is similar to the conversion of diphenols to melanin by C. neoformans. Several pigmentation patterns (resulting in the identification or grouping of Cryptococcus species) have been observed by using diaminobenzene and aminophenol compounds as substrates. The most common pigmentation pattern observed was pigment production by both C. neoformans and C. terreus. In contrast to the diphenols, only two aminophenols (4-hydroxymetanilamide and 3-aminotyrosine) were found to be highly specific as substrates. They allowed only C. neoformans to produce pigment. When 4-aminosalicylic acid was the substrate, a unique pattern was observed because only C. terreus, C. diffluens, and C. albidus produced pigment. Finally, a pattern was observed in which C. neoformans produced large amounts of pigment from aminophenol and diaminobenzene compounds, whereas the other Cryptococcus species produced smaller amounts. A simplified scheme with three substrates resulted in the identification of C. terreus and C. neoformans as well as two groups of other Cryptococcus species, group I (C. albidus and C. diffluens) and group II (C. laurentii and C. luteolus).


Hemolytic mutants of group A Streptococcus pyogenes. Hemolytic mutants of Lancefield strain SS-95 and ATCC 19615 Streptococcus pyogenes were produced by treatment with N-methyl-N'-nitro-N-nitrosoguanidine. These mutants contained the same levels of streptolysin O, nicotinamide adenine dinucleotidase, deoxyribonuclease, and hyaluronidase. The mutants were deficient in streptolysin S, as was the naturally occurring nonhemolytic Lowry strain. The mutants retained their pathogenicity for mice and, when reisolated from the dead animals, produced the mutant hemolytic pattern.


Analysis of group B streptococcal types associated with disease in human infants and adults. It is important to resolve existing differences of opinion regarding group B streptococcal type distribution in human disease because of the relevance of type prevalence to future programs of prevention. This report compares data obtained from typing 392 group B streptococci isolated from systemic infections in both infants and adults in the United States from 1972 through 1975. The data showed a substantial predominance of type III among strains isolated from cases of infant meningitis and from "late-onset" septicemia but did not confirm a prior report that type Ia causes most cases of "early-onset" infant septicemia. Type II was the predominant serotype among 11 cerebrospinal fluid isolates from adults. The fact that over one-fourth of the isolates were types other than Ia or III means that future epidemiological studies, including definition of immunological factors, must include all five group B types.


Fermentation of raffinose by lactose-fermenting strains of Yersinia enterocolitica and by sucrose-fermenting strains of Escherichia coli. Introduction of plasmids carrying the lacY gene (lactose permease gene) into Yersinia enterocolitica results in cells being able to ferment both lactose and raffinose. Transfer of such plasmids into Escherichia coli C600 (lacY) confers ability to ferment lactose but not raffinose. Derivatives of C600 that ferment both lactose and sucrose (Lac+ Scr+ strains) are able to ferment raffinose, but do not grow well on raffinose minimal medium. Fermentation of raffinose by Lac+ strains of Y. enterocolitica, and by Lac+ Scr+ strains of E. coli, is explained in terms of transport of raffinose via the lac permease and subsequent breakdown catalyzed by invertase.


Detection of group B streptococcal antibodies in human sera by radioimmunoassay: concentrations of type-specific antibodies in sera of adults and infants infected with group B streptococci. Current interest in determining the possible protective role of antibodies against group B streptococcal disease prompted this study of the feasibility of using a radioimmunoassay to measure type-specific immunity in humans. The radioimmunoassay was standardized as a quantitative test for antibodies against the carbohydrate (CHO) antigens of all five group B types. The data showed that the CHO antigens extracted by a cold trichloroacetic acid-sonification method measure more antibodies than do the corresponding CHO antigens extracted by hot hydrochloric acid; that the Ia CHOs extracted from two different types, Ia and Ic, measure the same quantity of Ia antibodies; and that human sera contain antibodies reactive with all five type-specific CHOs. No evidence of "protective" antibody was found in the serum samples studied, although there was evidence of and antibody response in adults to prolonged colonization by group B streptococci. The wide ranges of antibody concentration in a serum bank collection, the broad reactivity of all human sera tested, and the mixed populations of antibodies in human sera that react with different determinants on the same type-specific CHO antigen (type III) indicate that further studies must be done to better define normal and susceptible populations and to determine antigenic components important in protection.


Quantitative fluorescent immunoassay of antibodies to, and surface antigens of, Actinomyces viscosus. Optimal conditions for a fluorescence immunoassay of antibodies to, and surface antigens of, Actinomyces viscosus ATCC 19246 are described. In the standard fluorescence immunoassay, 10(8) colony-forming units of A. viscosus reacted with an antibody preparation, were washed, and then were treated with an excess of fluorescein-conjugated goat anti-rabbit immunoglobulin G. After another set of washes, fluorescence was determined in a spectofluorometer; in most cases excitation was at 485 nm, with emission measured at 525 nm. These conditions minimized interference from light scatter and stray light. Under appropriate conditions, antibodies to A. viscosus could be readily determined, with the fluorescence of the specific antibody-treated cells more than five times the fluorescence of controls treated with normal rabbit serum. Organisms coated with specific antibody could be detected at levels approaching 10(5) colony-forming units per ml. The standard fluorescence immunoassay procedure was readily adapted to the measurement of either particulate or soluble surface antigens of A. viscosus by competition of the antigen with a fixed amount of antibody in the standard assay system; the competition resulted in an antigen dose-dependent inhibition of fluorescence. The fluorescent immunoassay system thus appears to be a general one that could be applied to other microbial systems as well.


Evaluation of a semiautomated prestandardized immunofluorescence test system for detection of anti-native deoxyribonucleic acid antibodies. An immunofluorescence-native deoxyribonucleic acid (nDNA) antibody test system (Zeus Scientific, Inc.) was compared with a radioimmunoassay procedure (FARR assay) for detecting anti-nDNA antibodies in human serum. Double-blind studies of split samples obtained from 236 patients showed an 80% correlation between the immunofluorescence-nDNA antibody test system and the radioimmunoassay procedure. Studies of sera from patients with known diagnoses showed positive nDNA antibody findings in biopsy-prove n systemic lupus erythematosus only. The immunofluorescence-nDNA antibody test system provides a reliable, simple, and economically feasible alternative method for detecting nDNA antibodies that can be employed in any laboratory equipped with a fluorescence microscope.


Multiple-inocula (replicator) CAMP test for presumptive identification of group B streptococci. A replicator method for performing the CAMP test is described. Of 304 group B streptococci tested, 303 (99.7%) were CAMP positive. None of 1,093 non-group B streptococci was CAMP positive.


Titration of human serum antibodies to Toxoplasma gondii with a simple fluorometric assay. A new technique, FIAX, has been evaluated for the serodiagnosis of toxoplasmosis. It is based on a "dipstick" principle, and a special fluorometer is used to perform the indirect immunofluorescence test. The procedure appears to be simple and rapid and merits consideration as a useful serological test for toxoplasmosis.


Rapid sodium chloride tolerance test for presumptive identification of enterococci. A rapid test is described which distinguishes enterococci from other group D streptococci in 8 to 24 h by use of a medium containing brain heart infusion agar, NaCl, dextrose, and bromocresol purple.


Sterilisation in the laboratory autoclave using direct air displacement by steam. A device using a steam injection funnel is described by means of which air can be driven quickly and surely from an autoclave load. It is simple and inexpensive, necessitates no changes in the working routine of a microbiology laboratory, and does not interfere with the operation of the autoclave in its normal mode.


Synergy between sulphonamide and trimethoprim in the presence of pus. Synergy between sulphadiazine and trimethoprim against Escherichia coli has been demonstrated in culture media not containing lysed horse blood despite the presence of pus or a pus extract which inhibited the action of each drug separately. Synergy may therefore be important where pus is present in vivo.


Effect of thymidine on activity of trimethoprim and sulphamethoxazole. Thymidine at levels as low as 0.05 mg/1 reduces the activities of sulphamethoxazole and trimethoprim and their combination in vitro. Using a biological assay procedure, levels of thymidine greater than this were interpreted as being present in urine. The addition of sulphamethoxazole and trimethoprim, singly or in combination, to urine obtained from patients with urinary tract infections showed that all the antibacterial effect towards sensitive organisms was due to the trimethoprim component. It is suggested that trimethoprim should replace the combination co-trimoxazole for the treatment of some lower urinary tract infections, and that laboratory media, if they are to resemble the clinical environment, should contain thymidine.


Isolation of chlamydiae in untreated and Cytochalasin B treated McCoy cells. A comparison was made between untreated McCoy cells and McCoy cells treated with Cytochalasin B for the isolation of chlamydiae of subgroup A. Chlamydiae were isolated in both cell systems from 125 specimens, whereas six agents were isolated only in untreated cultures and seven agents were isolated only in Cytochalasin B treated cultures.


The distribution of substance P immunoreactive fibers in the rat central nervous system. A detailed account of the distribution of immunoreactive substance P-containing structures in the rat central nervous system is presented, from results obtained by applying an indirect immunofluorescent technique. High densities of substance P-containing nerve terminals were present in sensory nuclei and other non-sensory structures such as thalamus, hypothalamus and extrapyramidal system. Substance P-reactive neuron cell bodies were present in spinal root ganglia, nucleus habenulae medialis, nucleus interpeduncularis, caudoputamen and globus pallidus. Most of the neocortex and the cerebellar cortices had no substance P-positive elements. The results support the hypothesis that substance P may be a widespread neurotransmitter in the central nervous system.


A contribution to the electron microscopic morphometric analysis of peripheral nerve. Several aspects of data collection and analyses of peripheral nerve experiments employing light and electron microscopic morphometric techniques have not been adequately discussed in the literature. From statistical tests performed on nerve data, it was found that light compared with electron microscopic morphometry underestimates the number of small fibers. An optimum sampling strategy must take into account a potential bias toward small fibers introduced by measuring fibers from electronmicrographs. It must also take into account a potential bias introduced by the non-random distribution of nerve fibers of different sizes in nerves. These biases are offset by sampling a large enough number of fibers from large enough area electron micrographs. A method is presented for analysing periopheral nerve data using the nested analysis of variance. This requires first dividing the usual bimodal nerve fiber distribution into component normally distributed parts. The number of fibers in the two portions of a bimodal distribution must be considered in data analysis. Knowledge of the variances of parameters to be studied in any particular nerve is necessary for optimum sampling strategies.


A psychological rationale for the use of nitrous oxide psychosedation for children. N2O is, perhaps, the best adjunct, available today, in the management of emotional factors which influence problem behavior. It is not panacea, however, and should be used only in conjunction with sound psychological methods. The use of N2O must not be thought of as a method by which dentists can elude their responsibility to deal logically with the fears and anxieties of the patient. The fears and anxieties are frequently the result of rational mental functions flowing logically from the individual's previous dental history and learning experience. Viewed thusly, patients can be assisted to overcome the problems at a logical level, and N2O or other sedatives may be mere tools to assist the patient in overcoming the emotional elements of the situation. The real worth of N2O can only be evaluated intelligently by those who learn to "demand and to expect rational explanation in addition to empirical discovery".


The treatment of chronic renal failure by transplantation. The entire transplantation experience at the Peter Bent Brigham Hospital has been reviewed. This extends between 1951 and 1976, a 25 year period. Information on all patients has been undated through September 1976. Four different periods in the total experience have been compared, as determined by variations in immunosuppressive protocols. Although survival of renal allografts have plateaued over the past several years, the progressive decline in recipient mortality makes renal transplantation an increasingly attractive alternative to dialysis. Transplantation should be used in close conjunction with dialysis in the long-term treatment of patients with chronic renal failure.


Incidence and prevalence of home dialysis. The proportion of patients on home hemodialysis in the United States has declined continuously since 1972 despite an increasing total number of patients treated by dialysis. Regional variations and the experience in Europe are described. The reasons for the changing pattern of dialysis in the United States are discussed, together with possible measures which could help to reverse the present trend of increasing use of more expensive center dialysis which has features making rehabilitation more difficult for many patients.


Home dialysis in New York State.

Home dialysis in New York State. A description of New York State's program for providing assistance to patients on home dialysis. This program, which antedates the Medicare benefits provided under P.L. 92-603, Section 299I, was designated a Medicare provider in September 1975. Through its program, the Institute is able to provide home dialysis patients in the State with equipment, supplies and medications at a savings to the payors of these services, and presents a viable alternative to in-center dialysis.


The erupting first permanent molar--its place in stratified clinical field studies. Erupting first permanent molars of 5 1/2 to 6 1/2 year-olds are eminently suited for the clinical testing of potential caries-inhibiting techniques. Due to the rapid onset of caries in these teeth, accurate monitoring of new materials/regimes can be instituted and meaningful data obtained within one year of trial commencement. In addition, by concentrating on this age-group and by also stratifying for socio-economic status, deciduous caries experience and three-month wide age-bands, different trials' data can be readily compared. A further benefit is that these most caries-prone teeth can be protected thus saving the need for block injections in young subjects and alleviating later orthodontic problems.


Improving interrater agreement in evaluation in dentistry by the use of comparison stimuli. Research shows that use of rating scales with precisely defined scale points improves interrater agreement. Instead of words, scale points may be defined by products for which ratings have been established by consensual agreement. These products serve as standards for raters. In two studies, raters assessed student products using conventional rating scales and identical scales together with comparison stimuli (photographic exemplars) of consensually established values of 3 (adequate) for each criterion. Results of the first study (but not the second) clearly confirmed the contribution of exemplars to interrater agreement. Findings suggested that single exemplars significantly enhance agreement for criteria on which the ideal is an end point on a continuum. Multiple exemplars may be necessary for other criteria. Comparison stimuli offered promise for improving interrater agreement, but further work is necessary to adapt their use for complex criteria that frequently define the quality of dental preparations.


Egocentrism and classroom adjustment.

Egocentrism and classroom adjustment. This study examined the relationship between persistent egocentrism and adjustment status in normal school children. Fifty-three fourth-graders (33 males, 20 females) were administered measures of egocentrism and peer sociometric status, and indices of classroom adjustment were obtained from teachers. Degree of egocentrism was shown to correlate with classroom adjustment for both boys and girls and with peer group status for boys. The pattern of correlations suggested that the negative consequences of persistent egocentrism were different for the two sexes, with high egocentrism associated with shy, anxious behavior for girls and acting-out and learning difficulties for boys.


Protein in the U.S. diet.

Protein in the U.S. diet. Nationwide studies carried out between 1965 and 1975 have been reviewed to determine: (a) The adequacy of dietary protein nutrition in the United States, (b) the level of protein available from the national food supply, and (c) the effects of excessive amounts of protein in the diet. On the basis of the studies reviewed, it can be concluded that the diets of most Americans are adequate in quantity and quality of protein.


Albert Mueller-Deham: distinguished geriatric clinician. Modern geriatrics is the product of notable clinicians, past and present. However, recognition has been unequal. Pioneers whose publications were pre-eminent in their time have not received adequate identification. In consequence, the field is poorer for such neglect. Albert Mueller-Deham (1881-1971) of Vienna did excellent work in geriatrics. His clinical abilities were based on long staff attendance at a major hospital and by his insistence on attendance, with his staff, at autopsies. His key textbook was published in 1937 in German. His career in Austria was disrupted by political forces but, with energy and ability, he continued it with distinction in the United States. After retirement at age 70, he turned to social writings. A bio-bibliography of this noteworthy man is based on several letters, a personal narrative that he wrote in response to questions about his years in geriatrics, notes from one of his staff, and recollections by his co-author in 1942 of the revised American edition of the book. Students of aging can benefit by recourse to his classical text with its group of clinical principles.


Microvascular second toe transfer for digital reconstruction. Reconstruction of a severely damaged hand, with multiple amputations of digits, presents a difficult reconstructive problem. The development of a safe method of transfer of the great toe for thumb reconstruction, using the dorsalis pedis artery and the saphenous vein, suggested the possibility of one-stage microvascular transfer of the second toe for reconstruction of fingers. In two patients a one-stage transfer of the second toe was done to replace an index finger and in another for restoration of part of the left ring finger. The transplants survived without anticoagulants and vessel complications were not encountered. Sensory return was adequate and a significant improvement in function was achieved.


Multilating multidigital injuries: use of a free microvascular flap from a nonreplantable part. In severe hand injuries, all usable parts should be salvaged from amputated nonreplantable areas. A case is presented in which a free flap taken from an amputated finger was used to reconstruct a severely injured but viable remaining digit. This was done as part of the primary reconstructive effort and provided a well vascularized protective cover for the surviving injured digit.


The effect of antimicrobial feed additives on the colonization of the alimentary tract of chickens by Salmonella typhimurium. Groups of 33 chickens were fed continuously on diets containing feed additives that are employed commercially for a variety of purposes, and were infected orally when 4 days old with a nalidixic acid-resistant mutant of Salmonella typhimurium. The amount of S. typhimurium organisms excreted in their faeces was estimated by culturing them at intervals and in a standard manner on brilliant green agar containing sodium nalidixate; when the chickens were killed their caecal contents were examined by the same technique.Avoparcin and lincomycin, like nitrovin and tylosin (Smith & Tucker, 1975b), favoured colonization of the alimentary tract by the S. typhimurium organisms as shown by the fact that the chickens to which they were fed excreted these organisms in their faeces in higher concentration and for longer periods of time than did chickens fed on non-medicated diets. Amprolium, monensin, dimetridazole, arsenilic acid and nitro-hydroxyphenylarsonate had no obvious effect on the salmonella excretion pattern.When only five chickens in each group were experimentally infected so that the effect of the feed additives on infections acquired by contact could be monitored, avoparcin, lincomycin, nitrovin and tylosin again favoured colonization of the alimentary tract with the S. typhimurium organisms and so did dimetridazole. Arsenilic acid, in contrast, hindered the development of infection. Amprolium, monensin and nitro-hydroxyphenylarsonate were without obvious effect.Many of the chickens that were fed on diets that favoured S. typhimurium colonization, but not those fed on non-medicated diets, were still excreting S. typhimurium organisms in their faeces when they were killed at 56 days of age, the age at which broiler chickens kept under commercial conditions are usually slaughtered.


Prophylaxis against tetanus in non-immune patients with wounds: the role of antibiotics and of human antitetanus globulin. The potential value of oral erythromycin for antitetanus prophylaxis in non-immune patients with open wounds was assessed. Serum obtained by venepuncture from health persons 2 h after an oral dose of an erythromycin preparation was used as a culture medium rendered anaerobic by addition of cooked meat. Strains of Clostridium tetani inoculated into these sera failed to multiply when the donor had taken 500 mg of erythromycin estolate before a meal; other erythromycin preparations and the estolate at a dosage of 250 mg were ineffective or inconsistent in their inhibition of the growth of Cl. tetani. Human antitetanus globulin (ATG) was given to 12 patients, 9 with severe injuries and 3 with extensive burns, all of whom were judged, from their history, to be non-immune (or with expired immunity); all except one had received large intravenous infusions of blood and/or other fluids. Serum antitoxin assays by a mouse protection technique on days 0, 1--2, 3--5, 6--10 and 14+ showed no detectable antitoxin (less than 0.01) unit/ml) in the initial (pre-ATG) sample from three patients with severe injuries and in one with extensive burns. All the patients in the severely injured group showed an early appearance or increase in tetanus antitoxin to protective titres. Two of the three severely burned patients showed, respectively, a delayed appearance or an increase in antitoxin; the other burned patients showed a reduction from the initial pre-ATG titre, followed by a return to that titre after day 5.


Genetic differences in BCG-induced resistance to Schistosoma mansoni are not controlled by genes within the major histocompatibility complex of the mouse. Induction of nonspecific resistance to Schistosoma mansoni infection after the i.v. injection of viable BCG was investigated in outbred mice and a panel of inbred and H-2 congenic strains. Significant protection was induced in CF1, A/J, C57BL/6, C57BL/10, DBA/2, C57BR, and SJL mice. BALB/c mice were not protected whereas CBA and C3H mice expressed intermediate degrees of protection. Expression of the protective phenomenon is not controlled by genes within the MHC as shown by the marked differences in response between BALB/c and DBA/2 (H-2d) as well as between C57BR and C3H (H-2k) mice. H-2 congenic strains with C57BL/10 background (B10.A and B10.D2) were high responders. BALB.B10 mice carrying the high responder (B10) MHC on the nonresponder (BALB/c) background were not protected. The degree of splenic hypertrophy did not correlate with the expression of nonspecific resistance. These results demonstrate that, in addition to controlling specific immune responses, genetic differences influence the nonspecific protective phenomena related to BCG administration as well.


Immunologic characterization of hairy cell leukemias in continuous culture. The immunobiologic characteristics of three continuous cell lines established from hairy cell leukemia cells were investigated. All three cell lines continued to produce tartrate-resistant acid phosphatase, the enzymatic marker of hairy cells. Two of these cell lines were B lymphoid in nature. They carried Fc and C receptors, had surface and internal immunoglobulin, and did not form spontaneous sheep red blood cell rosettes. Experiments employing biosynthetic radiolabeling of immunoglobulin demonstrated distinctive immunoglobulin kinetics for each of these two hairy cell lines. One cell line remained quite similar to the original hairy cells from which it was derived whereas the other B lymphoid hairy cell line had undergone a switch in the immunoglobulin isotype produced. The third hairy cell leukemia line was shown to be of thymic derivation. These cells formed spontaneous sheep red blood cell rosettes and did not carry Fc or C receptors. The spontaneous sheep red blood cell rosette-forming cells contained tartrate-resistant acid phosphatase. They did not possess surface on internal immunoglobulin and did not synthesize immunoglobulin in vitro. Hairy cell leukemia cells maintained in permanent cell culture retain their immunobiologic properties and offer the opportunity for indepth study of these unusual cells.


A newly described activity of guinea pig IgG1 antibodies: transfer of cutaneous basophil reactions. Hapten-specific delayed time course skin reactions containing predominant accumulations of basophils and eosinophils were elicited in newborn guinea pigs after i.v. transfer of small amounts of oxazolone immune serum. The immune serum was fractionated by column chromatography procedures, and the fractions were examined for their ability in transferring this form of cutaneous basophil hypersensitivity (CBH). Only the 7S IgG-containing peak from Sephadex G-200 columns, and only the IgG1-containing fractions from DEAE columns, transferred CBH. An affinity column of bound oxazolone removed the activity from immune serum, and it could be recovered from the column by eluting with soluble oxazolone. About 35 microgram of purified IgG1 anti-oxazolone antibody could systemically transfer CBH reactivity. An immunoadsorbant column of anti-IgG1 removed this activity, but a column of anti-IgG2 did not. None of the procedures were able to separate activity in transferring CBH from passive cutaneous anaphylactic (PCA) activity classically associated with guinea pig IgG1 antibody. IgG1 from 8-day immune and 31-day hyperimmune donors were both effective. The average association constant of 8-day antibody was 8 X 10(-4) M-1. Transfer of cutaneous basophil reactions can be mediated by low affinity serum 7S IgG1 antibody.


Enhancement of carrier-specific helper T cell function by the synthetic adjuvant, N-acetyl muramyl-L-alanyl-D-isoglutamine (MDP). The adjuvant effect of a synthetic peptidoglycan, muramyl dipeptide (N-acetyl muramyl-L-alanyl-D-isoglutamine, MDP), was studied by using the anti-Tnp PFC and hemagglutinin responses of BALB/c mice to hapten-carrier conjugates. Administration of Tnp-OVA and MDP in saline to mice, followed 2 weeks later by a boost of Tnp-OVA in saline, led to significantly higher IgM and IgG anti-Tnp PFC and total anti-Tnp-hemagglutinin responses than those obtained in mice not treated with MDP in the initial immunization. A similar adjuvant effect by MDP on anti-hapten PFC responses was seen if mice were primed with KLH together with MDP and challenged with Tnp-KLH 2 weeks later. This apparent effect on carrier priming for helper function was confirmed and quantitated by double adoptive transfer experiments with graded numbers of spleen cells from KLH +/- MDP-primed mice and a fixed number of hapten-primed spleen cells from syngeneic Tnp-OVA immunized animals. These data suggest that at least one mode of action of the synthetic adjuvant MDP is via the enhanced stimulation of the helper T cell function.


A solid-phase radioimmunoassay for detection of human antibodies. I. Measurement of IgG antibody to bee venom antigens. A solid-phase radioimmunoassay (SPRA) has been developed to measure IgG antibodies to bee venom (BV) and phospholipiase A2 (PLA) in human sera. The principle of the test is similar to that of the radioallergosorbent test (RAST) measuring IgE antibody. Cyanogen-bromide-activated paper discs coupled with BV or PLA followed by supplementary coupling with human serum albumin were incubated with standard or test sera, washed, and incubated with 125I-labeled anti IgG. The serum levels of the IgG antibody have been temporarily expressed in arbitrary units. the reaction between the antigen and antibody was specific and the results were reproducible. Sera from 19 beekeepers, 42 beesting-sensitive patients and 20 blood donors (controls) were assayed by the SPRA. IgG antibodies to BV and PLA could not be detected (less than 4 U/ml) in all control sera, in 25 of the 42 patients and in one beekeeper. The IgG antibodies in 17 patients ranged between 5 to 58 U/ml (mean 7.6 U/ml), and in the 18 beekeepers ranged between 8 to 160 U/ml (mean 59 U/ml).


The use of detergents and immunoperoxidase reagents for the ultrastructural demonstration of internal immunoglobulin in lymph cells. Lymph-borne immunoblasts were fixed in dilute glutaraldehyde and then treated with saponin. This treatment made most parts of the cells permeable to ferritin, so that anti-immunoglobulin (Ig) antibodies which had been conjugated to horse radish peroxidase (HRP) had no difficulty in gaining access to Ig which thus could be demonstrated at an ultrastructural level. Best results were obtained by fixing the cells in 0.1% glutaraldehyde for 7 min and then treating them with a 1% solution of saponin for 100 min at 55 degrees C before exposing them to the Ig-HRP conjugate. The method yielded reproducible results and although it causes a small amount of ultrastructural damage, it may be of value in detecting a variety of intracellular antigens.


An indirect, quantifiable assay for cytophilic antibody. An indirect, quantifiable method for in vitro measurement of anti-SRBC antibody cytophilic for macrophages is described. The procedure consists of adding dilutions of normal or immune serum to 4 X 10(6) macrophages in 35 mm plastic petri dishes for 1 h at 4 degrees C. The plates are throughly washed to remove unbound antibody and 2 X 10(7) 51Cr-SRBC are added for 2 h at 4 degrees C. Total cpm in the plates is determined, non-bound 51Cr-SRBC are removed by washing and the plates are re-counted. Percent binding is calculated from these two values. The assay provides data consistent with visual determinations of number of SRBC/100 macrophages but has less variability and is performed more rapidly and easily than the visual method.


A prospective study of enteropathogenic Escherichia coli in endemic diarrheal disease. The rate of isolation of Escherichia coli belonging to the traditional serotypes enteropathogenic for infants was studied prospectively in two groups. Group 1 consisted of children with diarrhea and of controls without gastrointestinal disease who were matched for age and inpatient or outpatient status. Group 2 consisted of families entered in a prospective study of rotavirus infections. In group 1 enteropathogenic Escherichia coli were found in 13 (6%) of 220 children younger than 12 months of age and in nine (6%) of 143 children 12--35 months of age, all of whom had diarrhea. Enteropathogenic E. coli were found in only one of an equal number of matched controls (P = 0.002 and 0.004, respectively). In group 2 enteropathogenic E. coli were present in seven (18%) of 38 specimens obtained during diarrheal episodes, as compared with five (1%) of 492 specimens obtained when there was no diarrhea (P less than 0.001). The enteropathogenic E. coli isolated were not enterotoxigenic. The most common serogroup was O111, but many different O:H serotypes were detected. Thus, the association of enteropathogenic E. coli with endemic diarrhea was significant, even though no enteropathogenic mechanism was apparent.


Hydrocarbon and multibranched ester waxes from the uropygial gland secretion of grebes (Podicipediformes). The uropygial gland secretion of some grebes (Podicipediformes) had been shown to contain saturated and unsaturated aliphatic hydrocarbons and monoester waxes. While ester waxes are common constituents of preen gland secretions, nonisoprenoid hydrocarbons have not been detected hitherto. The wax constituents are very complex, belonging to several multibranched homologous series, including unusual acids with ethyl branches. The waxes were identified by gas-liquid chromatography-mass spectrometry and equivalent chain length comparisons. A method for the prediction of equivalent chain length values of unknown methyl esters is offered. The results are discussed from a chemotaxonomic viewpoint.


The steroids of 2000-year-old human coprolites. Six samples of human coprolites, some more than 2,000 years old, were analyzed for fecal steroid composition. Despite this very lengthy period of storage, the fecal steroids of coprolites were remarkably similar to those of stool samples collected today. The sterol nucleus was clearly rather stable under the dry environmental conditions of the Nevada Caves. The steroid content (microgram/g dried weight) of coprolite was low in comparison to that of modern man. The bile acid/cholesterol and plant sterol/cholesterol ratios of the coprolite, however, were similar to these ratios of the stools of modern man. In the six coprolites, an average 73% of the neutral steroids was digitonin-precipitable. This precipitate was composed of cholesterol and three plant sterols (campesterol, stigmasterol, and beta-sitosterol) and their bacteria-modified products. A portion of the neutral steroids had been converted to products tentatively identified as epimers of these steroids. Individual bile acids were identified in the coprolite. The bile acid composition of the coprolite was similar to that of the stool of modern man.


Inhibition by melatonin of the pituitary response to luteinizing hormone releasing hormone in vivo. The rate of secretion of 17-oxosteroids by the testes of anaesthetized dogs in vivo was used as an index of LH secretion. Intracarotid injection of luteinizing hormone releasing hormone (LH-RH, 1, 5 or 10 microgram/kg body wt) resulted in an increase in the testicular 17-oxosteroid secretion which was roughly proportional to the dose administered and which reached a maximum 60 min after the injection. Testicular output of 17-oxosteroids was unaffected by administration of melatonin (10 or 100 microgram/kg body wt) into the carotid artery. When LH-RH (5 microgram/kg) was injected into the carotid artery 3 h after intracarotid injection of melatonin (10 or 100 microgram/kg), the testicular response to LH-RH was considerably diminished. Pretreatment with melatonin (100 microgram/kg) did not alter the testicular response to human chorionic gonadotrophin (20 i.u./kg body wt) given i.v. Is is concluded that melatonin may act directly on the anterior pituitary gland in dogs to inhibit the LH-RH-induced release of LH.


Localization of trophoblast-defined surface antigens during early mouse embryogenesis. The binding pattern of a rabbit antiserum raised against mouse-ectoplacental-cone trophoblast on component cell populations in the pre-implantation and early post-implantation mouse embryo has been examined at the electron-microscope level using an immunoperoxidase-labelling technique. Binding was not detectable on the 1-cell stage, appeared at low levels at the 8-cell stage and was heavy on the trophectoderm and its trophoblast giant cell and extra-embryonic ectoderm descendants in the post-implantation embryo. In contrast, immunosurgically isolated 3 1/2-day inner cell masses (ICM) showed only slight labelling, whilst ICM derivatives in the 7 1/2-day embryo were unlabelled. The results indicate that the antiserum may be identifying a trophoblast-specific surface determinant(s), which appears with the differentiation of the trohectoderm and is maintained on some of the cell populations derived from this tissue at least until the early post-implantation stages.


The biochemical and genetic basis for high frequency thiomethyl galactoside resistance in lambda,lambdadg lysogens of Escherichia coli. In a culture of Escherichia coli K12 gal (lambdadg), cells which form large colonies on agar plates containing galactose and thiomethyl beta-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 posses at least two GAL+ OPERONS, ONE OF WHICH IS COTRANSDUCIBLe with bio+. In addition, TMG-resistant bacteria behave like lambdadg polylysogens when challenged with the phage lambdaI90c17. From these genetic experiments we conclude that TMG-resistant bacteria arise by duplication of the lambdadg prophage. Finally, gal+ bacteria which carry a single, additional, lambdadg prophage are TMG-resistant. TMG resistance is probably a gal+ gene dosage effect.


Antibodies against outer membrane proteins in rabbit antisera prepared against Escherichia coli O26 K60. Antibodies directed against the protein constituents of the outer envelope membrane of Escherichia coli O26 K60 were demonstrated in antisera elicited in rabbits against three different preparations of the bacterium. Outer membraned solubilized by sodium dodecyl sulphate were applied to the antisera in an interfacial precipitin test, followed by polyacrylamide gel electrophoretic analysis of the resulting immunecomplexes. Protein profiles showed a complete outer membrane protein pattern, indicating the antigenic character of these proteins. Antisera containing antibodies against outer membrane proteins and free of reactive antibodies against lipopolysaccharide showed relatively low agglutinating activities against the bacteria. The antibodies against the protein constituents of the outer membrane belong mainly to the 7S class immunoglobulins, as indicated by 2-mercaptoethanol treatment of the antisera.


A persistent infection of baby hamster kidney-21 cells with mumps virus and the role of temperature-sensitive variants. A persistent infection of baby hamster kidney-21 (BHK-21) cells with mumps virus (BHKpi) was maintained for over 60 cell passages in the absence of antiserum. Viral persistence was demonstrated in the cultures by hemadsorption, immunofluorescence, multinucleate syncytia, and released mumps virus at the level of 10(2)--10(3) fluorescent focus-forming units/ml. No detectable levels of interferon were found in cultures persistently infected with mumps virus. Approximately 85--95% of the cells contained viral antigens. Nuclear fluorescence was observed in the persistently infected cells. Mumps virus from persistently infected clutures (MuVpi) was more heat-labile than wild-type mumps (MuVo) when subjected to 40 degrees C. BHKpi cells had a more rapid doubling time and a higher cloning efficiency in soft agar in comparison to BHK-21 cells. MuVpi was also found to be temperature-sensitive. The temperature-sensitivity of MuVpi was determined by the efficiency of plating at 33 degrees and 39 degrees C. MuVpi readily established a persistent infection in BHK-21 cells with less cytopathology than MuVo, and released temperature-sensitive virus.


Clinical and serologic effects of live attenuated serum inhibitor-resistant influenza B vaccine in seronegative adults. The clinical effects, nasal and serum antibody responses, and virus excretion of a live attenuated serum inhibitor-resistant influenza B virus vaccine, R75, was evaluated in 43 seronegative healthy adults by a random double-blind study. Symptom responses were minimal and were not significantly different between vaccine and placebo groups. No fevers, abnormalities in physical examination or laboratory testing developed during 4 weeks of observation. Among vaccinees, 10 (48%) developed serum hemagglutination-inhibition (HI) antibodies, 16 (76%) developed serum neutralization (N) antibodies and 4 (19%) developed nasal N antibodies. The GMT responses from study day 0 to day 28 were 4.0 to 10.4 for serum HI, 1.8 to 9.8 for serum N, and 1.0 to 1.4 for nasal N. There were no significant titer changes in the placebo group. No virus excretion was detected. Although there are some questions concerning the relationship of antibody levels to protection, the low antibody responses in this study are an indication that R75 is not sufficiently immunogenic.


Studies on the fine structural localization of zinc iodide-osmium reaction in the brain. III. Some characteristics of localization in the synaptosomes. Synaptosomes from rat cerebral cortex were impregnated in the zinc iodide--osmium (ZIO) solution. The fine structural localization of the ZIO impregnation product was studied and, in addition, the function-dependent features of the reaction were examined after electrical stimulation or potassium chloride treatment. It was revealed that: (i) Aldehyde prefixation resulted in an increase in the number of reactive synaptic vesicles in all types of synaptosomes; (ii) Electrical stimulation decreased the number of reactive vesicles in a voltage dependent manner; (iii) Potassium chloride treatment also reduced the reactivity of vesicles; the reduction was dependent on the concentration of potassium and duration of treatment; (iv) Experimental interventions leading to the release of neurotransmitters from the synaptic vesicles and to fatigue of the nerve terminals also resulted in a decrease of the ZIO-reaction product of synaptic vesicles in a manner proportional to the strength of stimuli.


Wire closure of intracranial arteriotomy. Technical note. A technique is desdribed for closure of intracranial arteriotomies in areas of difficult exposure, using wire sutures that can be twisted rather than tied.


Eikenella osteomyelitis of the mandible associated with anemia of chronic disease. An unusual case of mixed bacterial osteomyelitis has been reported. The prodominant organism, E corrodens, normally is an inhabitant of the aerodigestive tract; its pathogenicity should not be overlooked in the diagnosis and treatment of oral infections. In addition, descriptions of anemia of chronic disease and of the spontaneous healing of a large defect of the bone have been presented.


The effectiveness of the short- and long-term use of crystallized theophylline in asthmatic children. The bioavailability, duration of action, and efficacy of a crystallized tablet form of theophylline were studied in 16 nonsteroid-dependent asthmatic children. All required bronchodilator drugs daily for control of symptoms. Theophylline 125 mg, ephedrine SO4 30 mg, T + E, or placebo were given in a randomized, double-blind, crossover design on four separate days. Pulmonary function tests (FVC, FEV1, FEF25-75) and serum T levels were determined at 0, 4, 1, 2, 4, and 6 hours on both day one and after day 7 of a every-six-hour drug dosage schedule. Mean maximum T levels were achieved at two hours with a peak mean of 2.94 microgram/ml +/- 0.24 SEM on day one. On day 8, the maximum T levels were higher, with a peak mean at two hours of 4.69 microgram/ml +/- 0.49 SEM. Computer analyses for pharmokinetics are compatible with 100% absorption of this preparation. Pulmonary function tests were significantly improved (FEV1 20% and FEF25-75 15%) at T levels of 2 to 5 microgram/ml. Addition of E to the T regimen further improved pulmonary function only on day one and had no effect on the last study day.


Differences in Hand Test responses of healthy females across the life-span. The Hand Test was administered to 175 females ranging from 17 to 72 years. Subjects were all "community-living," within normal range of hearing and vision, in good health, and had a modal education level of high school graduate. Results were consistent with previous findings using projective techniques with the elderly, in that a depletion and constriction of personality appears to be concomitant with the normal aging process.


A new clapsing system for unilateral distal-extension removable partial dentures. The background, rationale, and technique for fabricating the remodified Hart-Dunn attachment system have been presented. This subpontic attachment is a product of clinical experience in managing the unilateral distal extension removable partial denture patient. Minimal retention, reduced torque throughout the arch during function, and distribution of force according to sound mechanical principles make this remodified subpontic attachment the preferred method of treating most unilateral distal extension removable partial denture patients.


Stresses from loading distal-extension removable partial dentures. A concentrated load of 222 N in the region of the central fossa of the second molar resulted in stresses in the supporting bone which increased gradually from zero in the region of the retromolar pad to about 25 MN/m2 distal to the second molar. Stresses in the alveolar bone supporting the second premolar were about 30 MN/m2 at the cemento-enamel junction, decreased to about 20 MN/m2 distal to the apex of the root, increased to 100 MN/m2 at the apex, and decreased to 3 MN/m2 near the mesial upper third of the root surface. When a distributed load of 222 N was applied to the first and second molar region, the stress distribution was similar to that observed for the concentrated load, and stress magnitudes were about 20% lower. Dramatic decreases in stress, with the maximum stress at the apex being 40 MN/m2, occurred when a 222 N load was distributed in the area of the marginal ridge.


A study of various factors influencing shade of bonded porcelain. Seventy-two specimens of four commercially used precious metals were constructed. The porcelain applied to these specimens was glazed up to nine times for shade comparison. Specimens were also studied with a scanning electron microscope prior to and after application of the porcelain. The important observations from this study were: 1. The four types of precious metals looked alike after casting and surface treatment and prior to application of porcelain. 2. Porcelain-bonded-to-metal shades were not influenced by the types of precious metals used. 3. Porcelain-bonded-to-metal shades remained stable after six glazings and changed only slightly after nine firings.


Human blood pressure and pulse rate response to racemic epinephrine retraction cord. 1. The pulse rate of patients after application of racemic epinephrine-impregnated retraction cords depends more on the level of anxiety and stress than on the level of the epinephrine. 2. Blood pressure is elevated by placement of racemic epinephrine-impregnated retraction cords upon an exposed vascular bed or lacerated tissue. 3. Four percent racemic epinephrine-impregnated retraction cords cause less elevation of blood pressure than 8% racemic epinephrine cords. 4. Although the elevations in blood pressure from 8% cord occur within a narrow range, this range may be hazardous to cardiac patients. Therefore, 4% racemic epinephrine cord should be used. 5. A desirable amount of tissue retraction is produced by 4% racemic epinephrine cord. 6. Dry cords do not provide adequate retraction of tissue and are contraindicated for tissue-retraction purposes.


Spherical powder for retaining thermosetting acrylic resin veneers. 1. Nine different sizes of spherical powder were prepared, and their effectiveness as retentive devices was evaluated against those available commercially. 2. Smaller-diameter spherical powder (No. 5) gave the best results of all retaining devices tested. 3. The physical properties of the resins play an important role in the retentive strength with No. 5 retention beads. The retentive strength was reduced when brittle resin was used. 4. The retentive strength of the resin veneer was greatly affected by the angle of stress at the incisal resin. The retentive strength increased as the angle between the longitudinal axis of the specimen and the direction of stress decreased.


A method of stabilizing removable partial denture castings during clinical and laboratory procedures. The use of a temporary incisal rest at the most anterior point of a removable partial denture framework is advocated. This rest provides stabilization during intraoral and laboratory procedures when the corrected master cast technique is used.


Studies on the control of the corpus luteum in the vole, Microtus agrestis. The lifespan of corpora lutea resulting from hormonally induced ovulations was prolonged by exogenous prolactin, concurrent lactation, or pregnancy. Treatment of mated females with bromocriptine resulted in failure of luteal function only when the drug was given before Day 6 of pregnancy. Pregnancy was dependent on the presence of the ovaries in its later stages. The results suggest that prolactin is luteotrophic in early pregnancy but that a placental luteotrophin may become effective by Day 6 of pregnancy.


Effect of prolactin on the LH response to synthetic LH-RH in ovariectomized ewes. When ovariectomized ewes were treated with LH-RH, all 3 receiving prolactin infusion and 4 out of 5 receiving an infusion of NaCl solution responded.


Comparison of LH release and luteal function in cyclic and LH-RH-treated anoestrous ewes pretreated with PMSG or oestrogen. Pretreatment of seasonally anoestrous Clun Forest ewes with 750 i.u. PMSG or 50 microgram oestradiol benzoate 24 or 7 h respectively before a single injection of 150 microgram synthetic LH-RH significantly increased the release of LH compared to that after injection of 150 microgram LH-RH alone. Total LH release in the two "combined" treatments was approximately 70% of that found at a natural oestrus, compared to 25% for LH-RH alone. All but one of the treated ewes ovulated, but only those pretreated with PMSG consistently produced corpora lutea capable of elevating peripheral plasma progesterone concentrations although these were lower than those at natural mid-cycle. These progesterone concentrations were, however, comparable to those during the natural cycle when corrected for the higher metabolic clearance rate found during anoestrus.


Presence and activity of LH-RH in the mid-term human fetus. Immunoreactive LH-RH was present in all the hypothalamic and cortical extracts of mid-term human fetuses studied and in the cortical tissue removed from the two youngest fetuses. Gonadotrophin-releasing activity of hypothalamic and cortical extracts was demonstrated by the significant rises of circulating LH after infusion into oestrogen and progesterone-primed ovariectomized rats.


Evaluation of a gross anatomy program without dissection. Freshman medical students have been participating in an experimental multimedia gross anatomy program at Emory University for five years. The program includes audiovisuals, computer-assisted instruction, and tutorial sessions using prosected specimens. No lectures are given nor is dissection permitted. Experimental and traditional groups were compared by intramural written and practical examinations and by an extramural written examination prepared by the National Board of Medical Examiners and the Association of Anatomy Chairmen. Study of 35 intramural examinations given to five classes showed students in the traditional course with significantly higher performance in three examinations and students in the experimental course with significantly higher performance in six examinations. Neither group's performance was significantly higher on any extramural examination. It was concluded that, as measured by conventional examinations, students in the multimedia program with prosection tutorials learned human anatomy as well as those in the traditional lecture-dissection program.


Comparison of various immunological methods for distinguishing among mammalian pancreatic ribonucleases of known amino acid sequence. Fourteen mammalian pancreatic ribonucleases of known amino acid sequence were compared by 1 or more of 3 different immunological methods: standard quantitative micro-complement fixation, spot-plate micro-complement fixation, and inhibition of phage inactivation. It was found that, while the results obtained by the 3 techniques were correlated with one another, the standard micro-complement fixation procedure was most versatile, economical of materials, and easiest to execute. The standard MC'F technique was more sensitive than the spot-plate technique to differences in amino acid sequence. The inhibition of phage inactivation method was more sensitive than the standard method for measuring differences among closely related RNases but proved impractical for amino acid differences over 15%; the MC'F method could be extended to at least 30% sequence differences. The standard method, moreover, readily detected the single amino acid difference between dromedary and camel RNases. A linear relationship was found between immunological distance (y) in the MC'F test and percent sequence difference (x) which fit the equation y = 7x. The strength of the correlation between immunological distance and percent sequence difference is consistent with the proposal that a large fraction of the evolutionary substitutions of amino acids in ribonuclease are immunologically detectable. This could be explained either by a multideterminant hypothesis or by a pauci-determinant hypothesis which says that substitutions occurring outside determinants produce small conformational changes influencing determinant reactivity.


Pattern and chance in the use of the genetic code. The use of triplet code words in E. coli, phiX174, MS2, and rabbit globin was examined. A significant deficiency of purines in the third position of four fold degenerate codons was noted, although its significance is not understood. There has been no consistent selection against uracil in pyrimidine restricted codons. For many amino acids the choice between code words appears random, while for arginine, isoleucine, and probably glycine, distinct biases exist which can be explained in terms of tRNA availability.


Prognostic factors in the initial response to therapy by patients with advanced breast cancer. Identification of important prognostic factors with respect to the patient's initial response to therapy was made from a set of 25 covariates available on 281 patients with advanced breast cancer. Since the patients studied were all participants in a randomized clinical trial that involved three different treatment regimens (repeated weekly treatment of a combination of cyclophosphamide, methotrexate, 5-fluorouracil, vincristine, and prednisone; intermittent treatment of the same preceding five drugs given in 5-day courses every 4 weeks; or treatment every 3 weeks with adriamycin as a single agent), the effect of these treatments on the selection of important covariates was assessed. Although some evidence indicated that the set of important covariates differed by treatment, the differences were not strong enough to be of practical importance. Non-Caucasian patients did poorly on all regimens with a response rate of only 31% compared to 62% for Caucasian patients. The covariates of major prognostic importance for Caucasians were: disease-free interval, liver involvement, and performance status. Ambulatory patients with long disease-free intervals and no liver involvement had the best prognosis. After adjustments were made for these three covariates, the remaining covariates (such as menopausal status, bone involvement, number of metastatic sites, and duration of metastatic disease) were not significantly related to response. As reported earlier, the treatment effect was significant, even after adjustments were made for the important covariates.


Efficacy of BCG vaccination in prevention of cancer: an update. From 1949 to 1951, a controlled trial of BCG vaccinations was conducted in Puerto Rico. The 191,827 children, 1-18 years of age, initially enrolled in the study were skin-tested with tuberculin to determine their eligibility for vaccination. A total of 82,269 children were classified as reactors and not vaccinated. Of the 109, 558 nonreactors, 31,586 refused vaccination, 50,634 were vaccinated with BCG, and 27,338 were left unvaccinated as controls. We ascertained the incidence of cancer over an average follow-up period of 23.3 years in the latter two groups using the Puerto Rico Central Cancer Registry. By the end of December 1973, a total of 77 cancers had been diagnosed among the controls and 150 among the vaccinees. The overall incidence of cancer among the two groups was similar. Although a number of differences existed between the vaccinee group and the controls in regard to the incidence of cancer at various "sites", none of these differences was statistically significant. However, when cases of lymphosarcoma and Hodgkin's disease were combined for analysis, a statistically significant excess of cases occurred among the vaccinees. We concluded that BCG vaccination had no protective effect on the subsequent development of cancer in this population. The slight excess of cases of lymphosarcoma and Hodgkin's disease among the vaccinees raised the possibility that BCG may have had an adverse effect.


Antitumor immunity in strain 2 guinea pigs immunized with potassium chloride extracts of L2C tumor cells. Extracts of L2C tumor cells stimulated in vitro production of macrophage migration inhibitory factor (MIF) in peritoneal exudate cells from guinea pigs immunized with L2C tumor cells. Guinea pigs immunized with extracts of L2C tumor cells that were active in vitro (in the MIF assay) were completely resistant to challenge with viable tumor cells given 2 weeks later. Furthermore, guinea pigs immunized with extracts of L2C tumor cells within 1 hour after challenge with viable L2C tumor cells survived substantially longer than did nonimmunized controls. The immunoprotective and immunotherapeutic effects seen in guinea pigs given injections of viable L2C tumor cells were obtained with extracts of L2C tumor cells but not with extracts of another guinea pig tumor (line 10 hepatoma) or with extracts of normal guinea pig lymphoid cells.


Power take-off injuries.

Power take-off injuries. Power take-off injuries are uncommon, but serious and potentially fatal. Seven cases are reported demonstrating the violent trauma that can be incurred. The danger associated with use of machines with this and other power equipment is emphasized. Safety shields should be improved, and operators educated about the importance of retaining the shields and using machinery properly at all times.


Tracheo-innominate artery erosion in a burned child. A tracheo-innominate artery erosion in a 10-year-old boy 10 months after 80% burns in a gasoline explosion is reported. Tracheostomy was performed on the day of the burn to maintain an airway. Fatal hemorrhage from the erosion was the cause of death. Prevention can be achieved by: correct placement (second or third ring); aseptic tracheal toilet; positive nitrogen balance; and earliest possible tracheostomy removal.


Primary appendectomy. The effect of prophylatic cephaloridine on postoperative wound infection. A prospective, randomized, double-blind study on the effect of cephaloridine prophylaxis in the surgical management of appendicitis was performed. Of the 139 patients studied, 70 received cephaloridine and 69 received placebo prophylaxis. Both groups of patients were well matched with regard to age, sex, operative findings, and bacteria isolated at operation. Postoperative wound infections were detected in 11.6% of placebo-treated patients and in 1.4% of cephaloridine-treated patients (P less than .02). Prophylactic treatment with cephaloridine of patients undergoing surgery for uncomplicated appendicitis resulted in a perceptible reduction in the rate of postoperative wound infection.


North American Indian rock art and hallucinogenic drugs. It is proposed that the aboriginal rock paintings in two areas of North America may have been produced by shamans while they were under the influence of hallucinogenic agents derived from plants. The first of these areas is the Chumash and Yokuts Indian region of California, where polychrome paintings show designs similar to those visualized during the trance induced by decoctions of jimsonweed (Datura species). The second area is the lower Pecos River region of Texas, where shamanistic figures display traits considered to be conceptual analogues of the mescal bean (Sophora secundiflora) cult as practiced during historic times by Great Plains Indians. Although the evidence is only circumstantial, the proposed connections between these rock drawings and mind-expanding pharmacologic compounds fit well into the documented relationship that encompasses hallucinogenic drugs and certain movable objects of pre-Columbian American art.


Computer-based case tracing (COMTRAC).

Computer-based case tracing (COMTRAC). To assist clinical assessment and decision making, a computer-based system has been developed to organize key data of individual medical records and display these data in a standard graphic format for each disease. A program for multiple myeloma has been completed and 150 case records entered. The data accessed by computer-based case tracing on all cases of a particular disease are available for cumulative and comparative analyses of different therapeutic regimens.


Antibody response to group A streptococci in acute rheumatic fever. The Streptozyme test was compared with the three conventional tests (ASO, AH and ASK). Two hundred and four human serum specimens were tested by each of four tests. Sera taken during acute phase from patients with acute rheumatic fever and acute glomerulonephritis exhibited very high titers by the Streptozyme test. Titers of less than 100 would be considered negative and titers of 200 or greater highly suggestive or recent streptococcal infections by the Streptozyme test. The Streptozyme test appears to represent a simple, sensitive test for the detection of serum antibodies to streptococcal products. However, the complexity of the Streptozyme reagent suggests that more extensive studies are needed.


Septicemic infection in nude mice caused by a virulent strain of Mycobacterium bovis. Following i.v. infection with 2.3 to 2 X 10(6) viable organisms of Mycobacterium bovis strain Ravenel, congentitally athymic nude (nu/nu) mice of BALB/c background died earlier than their heterozygous (nu/+) counterparts, showing necrotic nodules in the heart and kidney with the involvement of liver and spleen. Pulmonary lesions in nu/nu mice, however, were less severe than those of dead nu/+ mice. In the reticuloendothelial organs of nu/nu mice, macrophages loaded numerous acid fast bacilli were transiently proliferated and later, necrotic-exudative lesions were produced without epithelioid granuloma formation, while necrotic lesions appeared at very early stage of infection in the other organs as well as adipose tissues. After thymocyte-reconstitution nu/nu mice were capable of developing epithelioid granulomas as seen in nu/+ mice, though having more numerous bacilli in granulomas than those of nu/+.


Effect of anticonvulsants on thalamic afterdischarge in rats and cats. Effects of anticonvulsants were determined on thalamic afterdischarge in gallamine-immobilized cats and d-tubocurarine-immobilized rats in order to clarify the participation of anticonvulsants in the thalamus. Thalamic afterdischarge was induced by electrical stimulation of cat nucleus centralis lateralis and rat nucleus reticularis at 50 Hz, 1 msec for 4 sec. In cats, diphenylhydantoin, carbamazepine, phenobarbital, and diazepam raised afterdischarge threshold, and shortened its duration induced at twice the threshold voltage. Trimethadione and dipropylacetate raised the threshold, but did not change the duration. In rats, diphenylhydantoin, phenobarbital, and diazepam raised the threshold, and shortened the duration with comparable dose ranges used for cats. Dipropylacetate and acetazolamide raised the threshold, although did not change the duration except for shortening action with a higher dose of dipropylacetate. Trimethadione was without effect. These results suggest that the depressive effect of anticonvulsants on the thalamus is, at least in part, associated with control of the epilepsies.


[Atrial complex in bronchial ECG leads]. The authors suggest a probe electrode by means of which the ECG may be recorded in the bronchial leads simultaneously from four levels of the atrioventricular zone. The P wave of bronchial ECG's was analysed in 24 patients who had no pathological changes in the respiratory organs and cardiovascular system in comparison with similar indices of routine ECG's. The informativeness of ECG's recorded in the bronchial leads in study of the bioelectric activity of the right and left atria is determined. It is pointed out that esophageal leads may be used for analysing the electric activity of the right atrium when they are recorded from the level of an electrode (28.5--31.5 cm) introduced into the esophagus.


[Experiences with the implantation of anterior chamber lenses (author's transl)]. On account of the good results with the new generation of anterior chamber lenses reported by C.D. Binkhorst, the implantation of lenses was begun in 1969 at the Department of Ophthalmology in Bremen and 350 anterior chamber lenses were implanted till 1st Febr. 1977. After the presentation of indications and the operative technique, the problems of postoperative treatment, the results and complications are discussed. According to our experiences up to now, we see in the implantation of iris-supported artificial lenses an enrichment of our surgical possibilities particularly in respect of special functional requirements.


[Results of cüppers "fadenoperation" (author's transl)]. The so called "Fadenoperation" Cüppers was performed in 233 patients = 40% of the cases who had been treated by muscle surgery at the University Eye Clinic Hamburg in 1975. The results in the different indication groups are presented. The most frequent indications are: Increased objective squint angle during near fixation, incomitant binocular movements in the horizontal plane, unstabile objective angles, nystagmus compensation (block-) syndrome, variation of Kestenbaum's nystagmusoperation. The result in 2/3 of the cases was satisfying, further surgery being not necessary.


Ethylene dibromide as a mitogen for liver. Intubation of ethylene dibromide (7.5 to 10 mg. per 100 gm. of body weight) into the stomach of nonfasted Wistar rats induces DNA synthesis and cell division in the liver. The peak of DNA synthesis, as measured by 3H-methyl thymidine incorporation, was attained at or shortly after 24 hours. The mitotic waves measured with the aid of colchicine occurred at 24 to 30 hours and 48 to 54 hours after ethylene dibromide treatment. Approximately 16 per cent of liver cells entered mitosis. The increase in DNA synthesis was confirmed by autoradiography. The stimulation of liver cell mitosis occurs in nonfasted animals without apparent cell necrosis. Thus, ethylene dibromide appears to be an effective mitogen for liver under some experimental conditions. The possible relationship between the mitogenic effect at a low dosage range and necrosis at a higher range is discussed.


The discovery of addiction. Changing conceptions of habitual drunkenness in America. The modern conception of alcoholism as a progressive, addictive disease dates from the late 18th century. By the mid-1800s it had become an integral part of American Temperance thought, as did the belief that total abstinence offered the only remedy for intemperance.


Operative treatment of prolapse of the pars membranacea of the trachea. A new method of operation for stiffening the tracheal back wall in cases of prolapse of the pars membranacea is reported; it has been applied with success on 22 cases. The advantages of this technically demanding operation starting from the neck over the method using transthoracic access with insertion of bone chips, fascia graft, or plastic prostheses are discussed.


Bacterial sputum cultures. A clinician's viewpoint. The expectorated sputum and transtracheal aspiration bacterial culture data presented by Murray and Washington were reanalyzed after removing several species unlikely to cause pneumonia in adults. After these exclusions, mean numbers of species isolated per specimen were not excessive. Those specimens with more than 25 leukocytes per field (groups 3, 4, and 5) has similar numbers of isolates per specimen (1.04, 1.05, and 1.05) compared with those with fewer than 25 leukocytes per field (groups 1, 0.57, and 2, 0.59). For this reason and others, it is recommended that expectorated sputum samples be accepted for bacterial culture if they contain more than 25 leukocytes per field.


Should yeasts in respiratory secretions be identified? Four hundred forty samples of sputa and bronchial washings were examined microscopically for evidence of pulmonary and oropharyngeal secretions. Most (88%) sputa showed definite evidence of oropharyngeal contamination, whereas bronchial washings showed much less frequent (21%) contamination. Culture results of the same specimens showed that yeasts (excluding dimorphic fungi) were recovered from 74% of the sputa and 25% of the bronchial washings. It seems that microscopic evidence of oropharyngeal contamination is a reliable index for predicting the presence of yeasts in respiratory secretions. Their presence in cultures of respiratory secretions probably represents "normal flora" except for Cryptococcus neoformans, and their routine identification is not warranted.


Bullous and cicatricial pemphigoid. Clinical, histopathologic, and immunopathologic correlations. A study of the records, biopsies, and immunopathologic data from 84 patients with bullous pemphigoid and 60 patients with cicatricial pemphigoid revealed that patients with bullous pemphigoid and negative or perhaps very low titer of circulating antibasement-membrane antibodies may constitute a distinct subgroup of patients with bullous pemphigoid. This group contained more women than men, had increased mucous membrane involvement, and had higher incidences of hypothyroidism, positive antinuclear antibody tests, and malignancy than did the other patients with bullous pemphigoid. Six of 41 patients with bullous pemphigoid who were given sulfapyridine or dapsone responded to these drugs, and these 6 constitute another group of patients necessitating further study. In nine patients, localized pemphigoid was found. The lesions were of two types: those occurring in sun-exposed areas and those occurring pretibially. Three of 57 patients with cicatricial pemphigoid had circulating antibasement-membrane antibodies. The clinical extent of disease in cicatricial pemphigoid was less extensive in our series than that previously seen at our institution. This was probably due to the earlier and more accurate diagnosis afforded by immunopathologic studies and to wider appreciation of the disease among nondermatologists.


An assessment of cell-mediated immunity in acute allograft rejection in man. A prospective study. A detailed multifactorial computer analysis of several in vitro tests of cell-mediated immunity has been carried out before and after surgery in 15 renal transplant recipients. Factors studied include lymphocyte blastogenesis, T- and B-cell levels, and lymphocyte protein synthesis. Large doses of immunosuppressive agents chnage lymphocyte subpopulations. This is seen especially in transplant patients who have rejection and who have decreased numbers of circulating lymphocytes. The T cells are selectively depressed more than the B cells. This is also reflected in the greater responsiveness of pokeweed mitogen-stimulated lymphocytes compared with lymphocyte responsiveness to phytohemagglutinin and concanavalin A. In this environment, rejection takes place. These observations suggest that rejection may be initiated by cortisone-resistant lymphocytes of the thymic medullary type. Measurements of changes in lymphocyte responsiveness to mitogens or changes in lymphocyte subpopulations do not reliably predict rejection. However, measurement of lymphocyte protein synthesis fills the criterion of providing a reliable test that can be carried out quickly, and in the future may be valuable in predicting rejection.


Role of the venous system in circulatory control. The systemic venous system is a dynamic component of the cardiovascular system, and it has a key role in the maintenance of the appropriate filling of the chambers of the heart and hence of the stroke volume. Studies in man and animals have characterized the different mechanisms that control the three major components--the splanchnic, muscle, and cutaneous vascular beds--and have provided information on the changes caused by disease and by chemical agents. Concerning man, much of the information has of necessity been derived from the behavior of the limb veins, the control of which is different from that of the splanchnic capacitance vessels. Studies of the behavior of the splanchnic capacitance vessels are needed before the action of drugs that affect the cardiovascular system of man is fully understood.


Immune reactivity during aging. I. T-helper dependent and independent antibody responses to different antigens, in vivo and in vitro. The immunological status during aging was assessed by measuring the antibody response of the long-lived (C3H/eb X C57Bl/6J)F1 mice to various antigens in vivo and in vitro. In vivo, a decrease in antibody production to DNP and NIP haptenic determinants coupled on to BGG, as well as the response to SRBC, was observed. The decline was more pronounced in the IgG as compared to IgM antibodies. The results were recorded when various parameters such as antigen dose and kinetics of the response, were considered. Reduction of the antibody response was also noted when PVP was employed as immunogen. Similar results were noted when the responses to SRBC and DNP--polylysine were induced and followed in spleen organ cultures. In all of these experimental systems, the peak response was observed in mice 6--12 months old. From then on a gradual decrease was manifested, mice 30--36 months old producing significantly low responses. The results demonstrate that decrease in antibody production is expressed in the isolated spleen tissue in the same manner as in the intact animal. Furthermore, they were interpreted as indicating that the lesion may be at the T helper and the B cell compartments.


Immune reactivity during aging. II. Analysis of the cellular mechanisms involved in the deficient antibody response in old mice. Analysis was made of the cellular basis of decline in the antibody immune response with advancing age. Employing the cell transfer system, it was found that spleen cells of aged mice have a limited capacity to react to SRBC when transferred into young irradiated recipients. Partial reconstitution was achieved when thymus cells from young untreated donors were applied together with the spleen cells, suggesting that deficiency in T helper cell function is responsible in part for the limited response. Similar studies using the T helper-independent antigen PVP also resulted in low levels of antibodies in the irradiated recipients. Treatment of the spleen cells with anti-theta serum before transfer did not lead to an increased response. Hence, it is suggested that the reduced response is related to limited function in the B cell compartment, and not to function of T suppressor cells. Transfer of bone marrow cells from aged into young irradiated recipients indicated that cells capable of differentiation into antibody-producing cells are available during aging. It is thus maintained that the defect at the B cell level is of a developmental nature.