Antigenic characterisitcs of Moraxella nonliquefaciens fimbriae in double immunodiffusion.

Rabbit antisera against purified fimbriae (pili) from Moraxella nonliquefaciens detected three fimbrial antigenic components, one or two of which appeared to be present in other fimbriated strains of M. nonliquefaciens and the closely related M. bovis. Maximal precipitation with the antisera required some denaturation of the antigen. Ultrasonication, repeated freeze-thawing, heating, and agents like KBr, NaSCN and urea were effective in liberating the antigen in diffusible forms. The morphology of the fimbriae was altered by heat treatment in 1 M KBr.


Assessment of wound contamination by wound irrigation. Experimental investigations on quantitative recovery of anaerobic and aerobic bacteria.

The quantitative recovery of E. coli, S. faecalis and B. fragilis from operative abdominal wounds was investigated in pigs in an experimental model suitable for statistical calculations. Wounds were contaminated in groups of ten with different numbers of either a single bacterial species or a mixture of two species. The wound was irrigated with saline 20 minutes after contamination. Significant differences in recovery were found between the bacterial species investigated. Expressed as percentage of the number of bacteria used for contamination, the recovery for a given species was rather low, but it was constant and independent of the degree of contamination. The investigation did not suggest any principle difference in the recovery of anaerobic and aerobic bacteria. The clinical applicability of the method is not yet clarified.


Differential effects of primary fixation with glutaraldehyde and osmium upon the membranous systems of the strial and external sulcus cells.

Ultrastructural preservation of the lateral cochlear wall differs between primary osmium fixation and primary glutaraldehyde fixation. Of significance in this study was the fact that whereas glutaraldehyde fixation preserves the parallel plasma membranes of interdigitations of the strial and external sulcus cells, fixation with osmium occasionally produces a breakdown of selected areas of membranes into apparent vesicles. The occurrence of these membrane artifacts was more common within the external sulcus cells. Reasons for the differential effects of the fixatives may include their differing modes of action. Care should be taken when evaluating unusual ultrastructure, since the type of fixative employed may be causal.


Experimental endocarditis in rabbits. 2. Course of untreated Streptococcus faecalis infection.

Insertion of a polyethylene catheter in the left side of the heart was used to induce sterile endocarditis in 34 rabbits. Bacterial endocarditis was established by injection of approximately 10(8) Streptococcus faecalis into the blood stream simultaneous with the removal of the catheter which had been in place for 3 days. The course of the bacterial endocarditis was examined by autopsy of rabbits sacrificed at regular intervals after the infection. The results showed that the presence of the catheter was not essential for the induction or maintenance of the infection. Growth of the bacteria took place in the preformed vegetations in the aorta, on the aortic valves and in the left ventricle. However, increases in the size of the vegetations, a high density of bacteria in the vegetations and secondary spreading were found only on the aortic valves. The extracardial manifestations of left-sided S. faecalis endocarditis included constant bacteriaemia, a high frequency of septic kidney infarcts and enlargement of the spleen. This form of experimentally provoked bacterial endocarditis in rabbits provides a good imitation of human subacute endocarditis, and would thus seem to be suitable for further study of the pathophysiology of endocarditis and evaluation of the effect of treatment with antibiotics.


Rabbit polymorphonuclear leukocyte chemotactic factor generated in vivo by Bacteroides fragilis lipopolysaccharide. II. Antigenic and biologic properties.

Preparations of the polymorphonuclear leukocyte (PMN) chemotactic factor isolated from lipopolysaccharide (LPS)-induced inflammatory exudate in rabbits were immunogenic in guinea pigs. Complete fusion of the precipitation lines produced against anti-CF by LPS-CF (molecular weigth 16,000) and material eluted on Sephadex G-200 columns with molecular weights (MW) of 68,000, 16,000 and 7,000 was found. Also, the chemotactically active material with MW of 68,000 and 7,000 eluted on G-75 columns after fractionation of the fraction of MW 16,000 from the G-200 eluate was antigenically identical to LPS-CF in double diffusion in agar. Normal rabbit serum (NRS) incubated with LPS, LPS-induced wound chamber exudate and NRS alone gave lines of precipitation against the anti-LPS-CF sera identical to that of LPS-CF. The capacity of LPS-CF to attract PMNs was significantly higher than that of LPS, and a peak in the number of PMNs in the exudate of wound chambers implanted in rabbits was found 4 h after the local injection of LPS-CF. When injected intraperitoneally in C5 deficient mice, LPS-CF stimulated a PMN migration which was only slightly below that in C5 normal mice. Antisera to LPS-CF inhibited the chemotactic activity of LPS-CF as well as that of LPS-NRS when the supernatants were tested using the Boyden's technique. Also, preincubation of PMNs with LPS-CF suppressed the migration towards a chemotactic gradient of LPS-CF molecules of these PMNs.


Isolation of enzymatically derived fragments of guinea pig IgG and an examination of their reactivity against Staphylococcal protein A.

Papain digest of guinea pig IgG2 was separated by ion-exchange chromatography into five fractions, one containing mainly Fab and four Fc fractions. Covalently linked Fc (cFc) and non-covalently linked Fc fragments (nFc) were present, both with a molecular weight (Mw) of about 56,000. nFc was split into half fragments by gel filtration under dissociating condition (Mw 25,000 to 27,000). An incomplete Fc fragment (iFc) was also isolated (Mw 36,000), which consisted of a Fc chain (Mw 26,000) non-covalently linked to a piece of another chain. These preparations all reacted with protein A. In addition, the Fc' fragment (Mw 22,000), which is the C-terminus of the Fc fragment, was isolated. This fragment did not react with protein A. The cFc was treated with acid and then digested with trypsin at pH 7.8 for 45 s. The digest were separated into four fractions (I-IV) by gel filtration on Sephadex G-100. Fraction I was indistinguishable from intact Fc, fraction II contained an incomplete covalent Fc fragment (Mw 34,000 to 38,000). Both these fragments reacted with protein A. Fraction IV consisted of fragments belonging to the C-terminus, and was protein A non-reactive. Fraction III was shown to contain a mixture of fragments belonging to fractions II and IV.


Effects of DDT (chlorophenotane) administration on glucose uptake and brush border enzymes in monkey intestine.

A considerable increase occurs in D-glucose uptake and brush border sucrase and lactase activities in the intestine of monkeys treated with a single oral dose of DDT. Brush border alkaline phosphatase activity remains unaffected in the pesticide treated animals. In vitro addiction of DDT has no effect on the sugar absorption and disaccharidase activities.


Auto-mutilation in animals and its relevance to self-injury in man.

Self-mutilation in non-human mammals is a well-established, although not a widely known phenomenon, which has been reported under zoo and laboratory conditions. In macaque monkeys, laboratory rearing and isolation are important predisposing factors, and the more serious self-injury is initiated by some immediate stimulating event. It is commonly accompanied by behaviour normally shown by the animal in a fighting context. Lower mammals are also known to mutilate themselves under laboratory conditions after administration of drugs wich probably cause increased sympathetic activity. The implications of this behaviour for an understanding of states of self-injury in man are discussed.


External small-field irradiation of cervical carcinoma with linear accelerator.

Since 1966, 125 patients with carcinoma of the uterine cervix were treated with a 6 MV roentgen beam from a linear accelatro. The pelvis was irradiated with 60 Gy followed by 30 to 45 Gy to a small volume using a lateral pendulum. The small-field irradiation was performed using a new beam-directing device consisting of a rod with a central pivot. Favourable results were achieved, despite the fact that most of the patients had advanced carcinoma.


Wall-scattering effects in electron beam collimation.

This report describes now a set of applicators, convering fields with dimensions of 4 to 20 cm, for the 6 to 20 MeV electron beams of a MEL SL75-20 linear accelerator was developed. The electron scatter contribution of the applicator walls to the treatment field was investigated, varying the applicator entrance opening and the scattering foil, with the aim of optimizing the resulting field flatness, with a minimum loss of depth dose. Experiments with field defining end frames and additional perspex scatterers for large field sizes are also reported.


En-bloc irradiation of tumours of the head and neck and their lymphatics. II. Early results and side effects.

The results and side effects of en-bloc irradiation of ear, nose and throat tumours and their lymphatics showed that the technique had been successful in nasopharyngeal carcinoma, but poor in carcinoma of the hypopharynx. Marked early and late radiation side effects were relatively common in high age and with certain tumour sites, whereas no correlation could be demonstrated with total absorbed dose, fractionation, cumulative radiation effect or major surgery.


Haematologic evaluation after radiation therapy in Hodgkin's disease.

Haematologic evaluation of 30 patients 2 to 7 years after radiation therapy for Hodgkin's disease was made. A complete general recovery of haematopoiesis occurred as concluded from the normal or even supranormal blood cell values. However, in only 5 of 28 patients was a complete local regeneration of haematopoiesis observed in the sternal marrow biopsies after 38 to 48 Gy. In the other cases, haematopoiesis was numerically or morphologically abnormal or totally absent.


Serologic distribution of antibodies against adenoviruses in sheep of large-scale farms.

The common soluble antigen of the first subgroup of bovine adenoviruses was used for assaying 793 sheep sera by the agar gel diffusion test. Of the 50 farms included in the study 43 were found infected. The ratio of reacting samples was 73.7% of the sera obtained from infected farms. Virus neutralization tests revealed that a considerable number of sera reacted specifically with all types of ovine adenoviruses, even with serotypes which had never been isolated in Hungary. The results yielded by the agar gel diffusion tests were compared with the results of virus neutralization tests. Of 850 cattle serum samples, agar gel diffusion tests gave positive results in 33.4%. Virus neutralization test was done only with the bovine and adenovirus type 2. No differences could be detected in antibody titres when the prototype strains (No. 19) and the strain isolated from sheep (ORT/111) were compared in parallel titrations. Both ruminant species were found to be infected with hovine adenovirus type 2. Neverthless, inapparent infection with these strains seemed to be less frequent among cattle than in sheep flocks.


High molecular weight ribosomal ribonucleic acid from vegetative hyphae and spores of Streptomyces griseus.

High molecular weight ribosomal ribonucleic acids (rRNAs) were isolated from young vegetative cells and spores of a streptomycin non-producing Streptomyces griseus, and their electrophoretic mobility was compared to each other and to that of rRNAs of Escherichia coli K-12. The electrophoretic mobility of 23 and 16S rRNAs from vegetative cells and spores of S. griseus was identical, but the 23S rRNAs of streptomyces ribosomes migrated more slowly on polyacrylamide gel than those of E. coli ribosomes. Intact, electrophoretically homogenous rRNAs could be isolated from S. griseus (No. 45-H) only in the presence of diethyl 1 pyrocarbonate (DEP), and intact rRNAs could be obtained from spores only if DEP had been added before breaking the spores. Otherwise instead of two distinct bands, three were obtained on polyacrylamide gel.


Regional developmental and fractional studies on myelin and other carbonic anhydrases in rat CNS.

Myelin carbonic anhydrase (CA) was studied with respect to its development in various brain regions and light and heavy myelin (LM and HM). The data indicate that the specific activity of myelin CA has a clear neuraxial distribution, increasing rostrally. The absolute activities and relative distribution are invariant with age; this suggests the CA activity in myelin is independent of stage and degree of myelination. The studies on HM and LM illustrate that HM, like total myelin, has a constant CA activity during development. In contrast, LM although equal to HM at 14 days, progressively decays to an adult level which is one-fourth that of HM. The distribution of CA in myelin was further investigated by comparing the activity in myelin with that present in the SN4 fraction. The activity in this latter fraction, which is derived from heavy myelin, was found to be 2.2 times higher than that in the myelin fraction. Thus, in the adult there exists an almost ten-fold range of activities among the various myelin fractions, SN4 greater than HM greater than LM. This may indicate a segregation of activity towards the outer lamellae. This segregation may have physiological importance in that it is this region of the sheath which should be integrally involved in control of myelin edema. Evidence indicates that there is an interaction of chloride with the enzyme, and maybe the primary ion moved by CA in order to initiate an osmotic flux out of the sheath. The interaction of chloride with the enzyme is dependent on the CA complex with the membrane in that solubilization and partial (60-fold) purification results in a preparation which is refractory to anions.


Value and limitations of the electrocardiographic response to exercise in the assessment of patients with coronary artery disease. Controversies in cardiology--II.

The results of exercise testing provide a probabibity statement rather than a definitive answer regarding the existence of coronary disease. Moreover, in asymptomatic subjects (1) the S-T segment response to exercise testing has a relatively poor predictive accuracy, (2) exercise testing does not appear to be a practical screening method for detecting subjects at high risk of sudden death, and (3) risk factors should be vigorously treated regardless of the results of screening studies for coronary artery disease. Hence, the probability statements derived from the electrocardiographic response to exercise testing do not appear to provide important enough diagnostic or prognostic information to justify routine use in asymptomatic subjects. Although exercise screening studies provide only marginal aid in detection coronary artery disease in symptomatic patients, important prognostic information may be derived that could influence decisions regarding pharmacologic or surgical therapy. Thus, although electrocardiographic stress testing should not be used as a routine screening procedure, its judicious use in symptomatic patients may provide the physician with important information for patient management.


Group G streptococcal endocarditis.

The group G streptococcus may be a more common human pathogen than previously recognized. A case of group G streptococcal endocarditis is reported and the 11 cases reported previously are reviewed. Group G endocarditis may have significant clinical and prognostic differences from endocarditis caused by the more commonly identified viridans or group D streptococci. Routine serologic grouping of beta-hemolytic streptococcal isolates from serious infections is warranted.


A modification of the automated immune precipitin method for quantitation of human serum immunoglobulins.

The use of the automated immune precipitin method to measure human serum immunoglobulins has been reported by several groups. The authors encountered difficulties in evaluating the technic even when they incorporated such recent modifications as the use of polyethylene glycol to enhance the reaction. By altering the flow pattern and prediluting each specimen, they were able to increase sampling rate, decrease processing time, and decrease the frequency of line obstructions while obtaining reproducible standard curves without using polyethylene glycol. There was good correlation between the results obtained by their modification of the technic and those obtained by the Fahey radial immunodiffusion method. As with other automated immune precipitin systems, abnormally high values gave characteristic notched peaks and lipemic specimens were centrifuged in order to obtain the infranatant for quantitation. The present modification of the automated immune precipitin system provides a rapid, simple and efficient method to quantitate human serum immunoglobulins.


Rationale for adding trace elements to total parenteral nutrient solutions--a brief review.

The physiologic importance of trace element supplementation to total parenteral nutrition solutions is discussed. The trace elements discussed are copper, zinc, manganese and iodide.


Patient-specific total parenteral nutrient formulas.

A method of formulating total parenteral nutrient (TPN) solutions to meet specific patient requirements is discussed. A series of calculations is presented which results in a specific formula that provides the required nutrients. The input data for the calculations are the patient's 24-hour requirements for protein, calories and electrolytes. To serve as a starting guideline, baseline nutrient requirements for patients of different weights are provided. Although individualizing TPN solutions may increase the pharmacy workload, such a program can assure that patients receive optimal nutritional support.


Chrysotherapy-induced aplastic anemia: a case report.

A case of gold-induced aplastic anemia occurring in a 58-year-old male with rheumatoid arthritis who received a total dose of 635 mg of gold sodium thiomalate over a 16-week period is reported. The clinical features, treatment, prevention and pathogenesis of gold-induced aplastic anemia are reviewed. The ability to predict patients in whom this hematologic effect may occur is discussed. Chrysotherapy is beneficial in the treatment of rheumatoid arthritis, but gold-induced aplastic anemia may be fatal. Absolute identification of patients at risk of having this hematologic side effect is not possible, but dosage reduction and intense monitoring of laboratory and clinical signs may prevent its occurrence.


Cellular Cl content and concentration of amphibian skeletal and heart muscle.

Toads (Bufo marinus) and frogs (Rana pipiens pipiens) were given intraperitoneal injections of Na36Cl and Na235SO4. After in vivo equilibration for 20--180 min, the animals were pithed, and their ventricular and semitendinosus muscles were excised. Measurements of total Cl (by titrimetry) and 36Cl (by radioassay) showed that specific radioactivities of plasma and mus les approached equality within 1 h after injection for toad skeletal and heart muscle and frog ventricles, indicating complete exchange of cellular Cl with 36Cl. From the simultaneously measured muscle water contents and 35SO4 spaces, intracellular Cl concentrations in vivo (in mumol/g cell water) for semitendinosus and ventricular muscles were calculated to be, respectively, 1.4 +/- 0.3 and 2.3 +/- 0.8 for Bufo and 1.7 +/- 0.7 and 4.8 +/- 2.4 for Rana. In view of these low values, active cellular Cl accumulation seems improbable, but cannot be rigorously excluded without simultaneous membrane potential and intracellular ion activity measurements. A high concentration of Cl in the sarcoplasmic reticulum of skeletal muscle is also inconsistent with these measurements.


Transplantation of pancreatic beta-cells prevents development of hypothalamic obesity in rats.

The present experiments have tested the hypothesis that ventromedial hypothalamic (VMH) lesions enhance insulin secretion by neural mechanisms. Rats were made diabetic by injecting streptozotocin to destroy their own pancreatic beta-cells. Subsequently, transplants of fetal pancreatic tissue were placed under the renal capsule. VMH lesions were placed in rats whose diabetes was cured with transplants as well as sham-transplanted animals. The animals were followed for 4 wk. The lesioned rats with pancreatic transplants gained no more weight than the sham-operated controls. There was no significant rise in insulin in the transplanted rats after VMH lesioning, but the VMH lesioned rats with intact pancreatic tissue showed the expected rise in insulin. Food intake rose 71% in the VMH lesioned rats with intact beta-cells, but only 23% in the VMH lesioned rats with transplants. Hypertrophy of the pancreatic islets was also observed in the VMH lesioned rats with an intact pancreas, but was not found in the VMH lesioned rats with a transplanted pancreas. Thus, transplantation of pancreatic tissue beneath the renal capsule of diabetic rats prevented the characteristic hyperphagia, hyperinsulinemia, and obesity in VMH lesioned rats whose pancreas was free from intact innervation. The results support the hypothesis that neural mediation of the rise in insulin is the primary factor in the development of hypothalamic obesity.


Regulation of glycogen resynthesis in muscles of rats following exercise.

Following a strenuous bout of exercise, glycogen repletion occurred most rapidly in the fast-twitch red type of muscle, least rapidly in fast-twitch white, and at an intermediate rate in slow-twitch red muscle. There was a linear correlation between glycogen synthase I activity and the rate of glycogen synthesis in the three types of muscle. This finding helps explain the differences between the rates of glycogen resynthesis in the three muscle types, and supports the view that glycogen synthase activity is the most important factor determining the rate of glycogen synthesis when substrate supply is adequate. There was an inverse correlation between muscle glycogen concentration and percent glycogen synthase I. Plasma insulin concentration was low and norepinephrine and glucagon concentrations were elevated in the postexercise period. The finding that rapid glycogen synthesis occurred despite a hormonal milieu conducive to glycogenolysis provides evidence that a low glycogen concentration is a potent stimulus to glycogen synthesis that overrides the effects of low insulin, and high norepinephrine and glucagon levels.


Circadian rhythm dissociation in an environment with conflicting temporal information.

The relative contributions of light-dark (LD) cycles and feeding (EF) cycles in providing temporal information to the circadian time-keeping system were examined in chair-acclimatized squirrel monkeys (Saimiri sciureus). The circadian rhythms of drinking, colonic temperature, urine volume, and urinary potassium excretion were measured with the LD and EF cycles providing either conflicting phases or periods. In conflicting phase experiments, animals were exposed to 24-h LD cycles consisting of 12 h of 600 lx followed by 12 h of less than 1 ls and concurrent 24-h EF cycles in which the animals ate for 3 h and then fasted for 21 h. One group had food available at the beginning and a second group at the end of the light period. In conflicting period experiments, monkeys were exposed to 23-h LD cycles (LD 11.5:11.5) and 24-h EF cycles (EF 3:21). Analysis of the rhythms showed that both phase and period information were conveyed to the drinking and urinary rhythms by the EF cycle, and to the temperature rhythm by the LD cycle.


Glucose load diverts hepatic gluconeogenic product from glucose to glycogen in vivo.

Intravenous or oral administration of concentrated glucose solution into fasted rats simultaneously injected with 14C-bicarbonate resulted in an inhibition of [14C]glucose release into the blood and in an accelerated [14C]glycogen formation associated with glycogen synthetase activation and phosphorylase inactivation in the liver. The specific activity of glycogen was much higher than that of blood glucose after the glucose load, indicating that glycogen originated from gluconeogenesis rather than blood glucose. These metabolic changes induced by the glucose load were not mediated by endogenous insulin because they were observed to the same extent in rats treated with anti-insulin serum.  However, they were mostly, if not totally, abolished by adrenalectomy, which suppressed gluconeogenesis and glycogenesis.  Glucose tolerance was markedly impaired not only by anti-insulin serum, which inhibits peripheral glucose utilization, but also by adrenalectomy, which affects hepatic metabolism. It is concluded that a glucose load diverts the final product of hepatic gluconeogenesis from blood glucose to liver glycogen; these metabolic changes in the liver are an important determinant of glucose tolerance.


Carbonic anhydrase in the generation of cochlear potentials.

The experiments on 18 guinea pigs were divided into two groups and each group was arranged in such a way that the effect of hypercapnia (generated by breathing 10% CO2-90% O2) was investigated with and without inhibition of carbonic anhydrase by methazolamide, 25 mg/kg, in the first group and acetazolamide, 50 mg/kg, in the second group, administered intravenously. The endocochlear potentials (EP) and endocochlear PO2 were recorded by microelectrodes introduced into the scala media, and cochlear microphonics (CM) were monitored by a silver-wire electrode from the round window. In the first exposure to hypercapnia (20-40 min) EP increased about + 6 mV. At the same time CM decreased; the reason for this is not yet known. During the second period of hypercapnia (80-100 min) when carbonic anhydrase was inhibited with methazolamide and acetazolamide, EP did not elevate as during the first period when carbonic anhydrase was not inhibited. In this work, under specific conditions, it was observed for the first time that carbonic anhydrase affects the generation of EP.


Tetracycline-induced inhibition of Na+ transport in the toad urinary bladder.

The effects of three tetracyclines, demethylchlortetracycline (DMC), minocycline (MNC), and oxytetracycline (OTC), on Na+ transport (measured as short-circuit current) were examined in toad urinary bladders mounted in modified Ussing chambers. During a 1-h incubation period serosal DMC (but not MNC or OTC) inhibited basal Na+ transport, whereas MNC (but not DMC or OTC) inhibited ADH-stimulated Na+ transport. MNC also inhibited cyclic AMP-stimulated Na+ transport. During longer incubation periods all three drugs inhibited basal Na+ transport. The DMC-induced inhibition of basal Na+ transport and the MNC-induced inhibition of ADH-stimulated Na+ transport were paralleled by an inhibition of the active conductance of the bladders. Thus, although all three drugs inhibit basal Na+ transport, only MNC inhibits ADH-stimulated Na+ transport. This effect does not correlate with the known effects of the tetracyclines on ADH-stimulated water flow or with drug-protein binding, and may be related to the greater lipid solubility of MNC.


Myocardial infarction in the baboon: regional function and the collateral circulation.

Occlusion of the anterior descending coronary artery was produced in sedated baboons 7-15 days after implantation of a micromanometer and ultrasonic crystals for measurement of regional left ventricular dimensions in ischemic, marginal, and control segments. One minute after coronary occlusion (CO), ischemic segments exhibited a marked systolic bulge with wall thinning, and percent systolic shortening of marginal segments decreased. Over the ensuing weeks, there was a progressive increase of end-diastolic lengths in marginal and ischemic segments, whereas systolic shortening in these segments did not improve significantly. Control segments did not change. In control baboons, the coronary collateral index was 55 +/-25 (SE) compared to 560 +/- 74 in normal dogs. One month after CO, the collateral index was 543 +/- 144 in baboons compared to 6,685 +/- 716 in dogs, regions of normal tissue were seen in the infarct (14.2 +/- 2% of left ventricular mass). Minimal coronary collateral development in the baboon provides a likely explanation for differences from the dog in regional functional responses and in the character of the infarct.


Inherited metabolic diseases and pathogenesis of mental retardation.

Continuing progress has been made in the exploration of the biochemical causes of mental illness. Recent research has indicated that selected abnormalities of specific isoenzymes play an important role in the pathogenesis of heritable metabolic disorders such as phenylketonuria and Niemann-Pick disease. The roles of "animal lectins" and glycosidic enzymes in brain development and synaptogenesis appear to have received important substantiation within the past year. In additions, it has been shown that the blood-brain barrier can be temporarily altered so that exogenous enzymes can enter the central nervous system, and imperative consideration for enzyme replacement therapy in mental disorders. Less satisfactory progress has been made concerining potential anabolic disorders of lipids affecting the nervous system. Finally, novel experimental directions concerning energy metabolism by the brain offer considerable hope for the elucidation of some of the causes of mental retardation.


Duodenal gangliosidosis in a cat: ultrastructural study.

Ultrastructural changes similar to those seen in gangliosidosis in man were detected in the duodenal mucosa of a cat (cat 1). The N-acetylneuraminic acid concentration was determined from the duodenum of cat 1 and from a clinically normal cat (cat 2). The total amount of gangliosides in the duodenum of cat 1 was 100 times greater than in the duodenum of cat 2 per unit wet weight. The lyophilizate from the intestines of both cats gave a blue-violet color that was regarded as qualitatively positive for N-acetylneuraminic acid determination. In cat 1, membranous cytoplasmic bodies were in all cell types of the duodenal mucosa.


Lumbar spondylectomy in the subhuman primate.

Removal of the 2nd or 3rd lumbar vertebrae (or both) has been accomplished in the subhuman primate (Macaca mulatta). Variations between this animal and the dog in posture, vertebral column anatomy, and spinal cord blood supply made no apparent difference in the results when compared with those in previous experiments. All macaques were able to clinb and to use their hind legs in a normal manner within 3 days after surgical operation. Once hair had regrown over the surgical incision, persons not familiar with the animals were not able to identify the animals that had undergone surgical operation. If a single vertebra was removed, use of spinous process and a single vertebral body plate was sufficient to stabilize the vertebral column. Except for 1 rhesus macaque whose spinous process plates were removed 20 days after the operation, vertebral columns of macaques (n = 3) that underwent single spondylectomy healed in nearly normal alignment. With the removal of 2 lumbar vertebrae, the previously described stabilization procedure was not sufficient to prevent kyphotic deformity of the vertebral column. Spinous process plates from 1 macaque were removed after 64 days. However, all these macaques (n = 3) could climb and use their legs as well as others in the colony.


Controlled trial of 6-month and 8-month regimens in the treatment of pulmonary tuberculosis. First report.

Four short-course antituberculosis regimens allocated at random were studied; (1) streptomycin, isoniazid, and rifampin given daily for 6 months; (2) these 3 drugs plus pyrazinamide given daily for 2 months, followed by twice-weekly administration of streptomycin, isoniazid, and pyrazinamide; (3) a regimen that differed from regimen 2 only in that ethambutol replaced pyrazinamide, and (4) streptomycin plus isoniazid plus rifampin plus pyrazinamide given 3 times per week for 4 months, followed by streptomycin plus isoniazid plus pyrazinamide administered twice per week. The last 3 regimens were given for 6 or 8 months at random. All except 1 of 680 patients with tubercle bacilli drug-susceptible before treatment had a favorable bacteriologic response during chemotherapy. The relapse rates during the first 6 months after chemotherapy were low, except in the ethambutol series, in which 19 per cent of the patients relapsed after 6 months of treatment, and 8 per cent relapsed after 8 months. A substantial proportion of the patients with strains initially resistant to either isoniazid or streptomycin had a favorable response to their allocated regimen, but the results were not as good for those patients with strains resistant to both drugs. An important finding is that the incidences of immunologic febrile reactions to rifampin and of rifampin-dependent antibodies were very low during the 3-times-weekly regimen.


Solid-phase radioimmunoassay for immunoglobulin G Staphylococcus aureus antibody in serious staphylococcal infection.

Clinical features of 99 patients with staphylococcal infection were reviewed, and sera were tested by solid-phase radioimmunoassay and gel diffusion for staphylococcal antibodies to ascertain whether these variables predict the extent of infection and the need for prolonged therapy. Clinical features, including the presence of a primary site of infection or a continuous pattern of bacteremia, were not sufficient for differentiating endocarditis or complicated bacteremia from uncomplicated bacteremia. Patients with uncomplicated bacteremia were cured by 3 weeks of antibiotic therapy. Positive serologic tests for staphylococcal antibody helped distinguish patients with endocarditis or complicated bacteremia from patients with uncomplicated bacteremia. Radioimmunoassay was more sensitive than gel diffusion for identifying patients with complicated bacteremia. Our results indicate that patients with a positive antibody result 14 days after the onset of infection should be considered to have endocarditis or complicated bacteremia, but a negative antibody result would support short-term antibiotic therapy.


Teichoic acid antibodies in chronic staphylococcal osteomyelitis.

Gel-diffusion and the enzyme-linked immunosorbent assay (ELISA) were used to quantify and to identify the immunoglobulin class of teichoic acid antibodies in patients with chronic staphylococcal osteomyelitis and a wide variety of other infections. Teichoic acid antibodies were identified by gel-diffusion in 14 of 23 patients with staphylococcal endocarditis, six of 30 with staphylococcal bacteremia without endocarditis, four of 35 with staphylococcal skeletal infections, and one of 45 with nonstaphylococcal infections. None of the 20 patients with chronic staphylococcal osteomyelitis had positive gel-diffusion assays, even though many had had their infections for several years. The ELISA method was more sensitive than gel-diffusion in measuring teichoic acid antibodies, but was also much less specific. Teichoic acid antibodies were detected predominantly in the IgG fraction of serum. Our findings suggest that the presence and degree of antigenemia are more important than the duration of the staphylococcal infection in stimulating production of teichoic acid antibodies.


Refractory sprue: recovery after removal of nongluten dietary proteins.

A 44-year-old woman with diarrhea, weight loss, and a small-bowel biopsy consistent with adult celiac disease failed to improve on a gluten-free diet. Despite in-hospital supervision at two university medical centers and addition of corticosteroid therapy, diarrhea and wegith loss continued, resulting in life-threatening nutritional depletion. She was transferred to the University of Chicago and made full nutritional recovery with total parenteral nutrition. Exploratory laparotomy showed no abnormality except the flat intestinal mucosal lesion. Diarrhea recurred when a gluten-free diet was resumed. When the patient ate egg, chicken, or tuna alone, severe diarrhea, hypotension, cyanosis, and shock occurred. When these foods--along with gluten--were eliminated from the diet, the patient returned to oral nutrition and made a full clinical recovery. In patients with refractory sprue deletion of other dietary proteins in addition to gluten, as in the present patient, may be lifesaving.


How much will U.S. medicine change in the decade ahead?

During the past 12 years health care services have been increasingly used by low-income families, largely as a consequence of infusion of more funds into the health care system. The number of persons employed in health-care has gone up along with an expanded governmental role. Present governmental concerns are focused on cost and the relation of public attitudes to needs for, and costs of, a system of national health insurance. In the near future, medical services are not likely to be expanded, and various costly services may be contracted. The number of physicians will go up but the fraction in solo practice will decline. Patterns of physician maldistribution are not likely to be changed substantially. Costs will continue to go up, and government and other third-party payers will strive to control them. A comprehensive form of national health insurance will not come soon, and the public will become increasingly aware of the limits to which health care can improve their lives.


[Cannabis].

Cannabis produces a resin, the psychotropic effects of which are known since the High Antiquity. It induces euphoria, a feeling of pleasant dreaminess. Nevertheless, psychiatric disturbances such as psychomotor deficiency states or acute oniroïd states were described in heavy users countries. These psychiatric disorders put the accent on the starting or catalysing role of cannabis on the premorbid personality. The experimental studies of cannabis on the premorbid personality. The experimental studies of cannabis and its active substance (delta-g-tetrahydrocanabinol) bring information on this problem.


Protein-calorie deficits in developing countries.

Twenty years ago, we were invited to present a paper on severe protein-malnutrition in children in a meeting organized by this same Academy. In reviewing that paper for this presentation, it was very frustrating to find that the basic principles we stated there in regard to the nature of the probelm and its epidemiology are as valid today as they were 20 years ago. As most other workers in the field, we were then particularly concerned with the severe forms of protein-calorie malnutrition (PCM): kwashiorkor and marasmus. These individual cases occupied a large proportion of pediatric beds in hospitals of most developing countries, and we were interested in finding better ways to treat them. But also we were studying these cases as a basis for understanding the responsible factors better and for designing possible preventive measures. The interrelations of kwashiorkor and marasmus were recognized then, as well the fact that both protein and calories should be considered together in the epidemiology of the problem. We were also beginning to understand that the severe clinical cases that we were seeing in the hospitals were only the visible part of a much greater problem affecting the communities from which these children came. With the knowledge then available on the epidemiology of PCM, we were also starting to explore some specific measures for its prevention. I would like now to review what progress we have made, if any, in the understanding of the nature and magnitude of the problem, its epidemiology, and in designing preventive measures.


[Results of two methods for testing bacterial susceptibility to antibiotics: disk method and a new semi-automatic method in liquid medium (ABACR procedure) and comparison with the minimal inhibitory concentration (author's transl)].

A performance analysis was established between three methods for testing of bacterial susceptibility to sixteen antibiotics: agar dilution or minimal inhibitory concentration (MIC) method, agar diffusion or DISK method, and semi-automatic dilution in liquid medium or ABACR method. Precision of the three methods was determined using three reference strains in repetition experiments (9 repetitions for MIC, 60 repetitions for both DISK and ABAC). The results show that MIC method was the most precise, and that the precision of DISK and ABAC methods was not sifnificantly different. Fidelity of both DISK and ABAC methods was compared in taking as reference the MIC of 200 wild strains of Gram-negative bacteria. Omitting the data concerning trimethoprim-sulfamethozoaxol, the results were found concordant between DISK and MIC methods in 68.6 percent of the cases, between ABAC and MIC methods in 73.6 percent, and between ABAC and DISK methods in 68.9 percent. With regard to MIC, ABAC method gave results slightly better than those of DISK method, especially for five antibiotics: gentamicin, minocyclin, doxycylin, chloramphenicol and polymyxin B. So, the ABAC method looks very interesting for a semi-automatic routine method in testing bacterial susceptibility to antibiotics.


[Induction of interferon by Bacillus subtilis (author's transl)].

Although it is generally admitted that Gram-positive bacteria cannot induce interferon, we were able to show that organisms and cell walls of Bacillis subtilis as well as components obtained through the extraction of endotoxins from Gram-negative bacteria can induce type II interferon. Moreover we have ascertained that those components have the same biological activity as the endotoxins, espcially with regard to their behaviour with B cells of lymphocytes, the results of pyrogen test, the incidence of cycloheximide and their function as inhibitors of bacteriophages. However, it has appeared that chemically they are quite different from the lipopolysaccharide of the Gram-negative bacteria: no lipid A could be detected.


New perspectives on biliary atresia.

An investigation into the aetiology, diagnosis, and treatment of biliary atresia was carried out because the prognosis remains so poor.In an electron microscopical study no viral particles or viral inclusion bodies were seen, nor were any specific ultrastructural features observed. An animal experiment suggested that obstruction within the biliary tract of newborn rabbits could be produced by maternal intravenous injection of the bile acid lithocholic acid.A simple and atraumatic method of diagnosis was developed using(99) (m)Tc-labelled compounds which are excreted into bile. Two compounds, (99m)Tc-pyridoxylidene glutamate ((99m)Tc-PG) and (99m)Tc-dihydrothioctic acid ((99m)Tc-DHT) were first assessed in normal piglets and piglets with complete biliary obstruction. Intestinal imaging correlated with biliary tract patency, and the same correlation was found in jaundiced human adults, in whom the (99m)Tc-PG scan correctly determined biliary patency in 21 out of 24 cases. The (99m)Tc-PG scan compared well with liver biopsy and (131)I-Rose Bengal in the diagnosis of 11 infants with prolonged jaundice.A model of extrahepatic biliary atresia was developed in the newborn piglet so that different methods of bile drainage could be assessed. Priorities in biliary atresia lie in a better understanding of the aetiology and early diagnosis rather than in devising new bile drainage procedures.


The Morquio syndrome: neuropathology and biochemistry.

The activity of N-acetyl galactosamine-6-sulfate sulfatase was studied for the first time in the liver and brain of a patient with a clinically typical case of Morquio syndrome with keratosulfaturia. As has been demonstrated in the fibroblasts of patients with this syndrome, this enzymatic activity was markedly decreased in both organs. Neuropathological examination revealed moderately swollen neurons containing PAS-positive, coarse globular inclusions in the cerebral cortex, Ammon's horn, basal ganglia, and thalamic nuclei. Ultrastructurally, the inclusions consisted of stacked, straight or loose, wavy membranes of various lengths, often associated with pale or moderately electron dense homogeneous "lipid droplets." These ultrastructural features of the inclusions were closely similar to the granulomembranous bodies of Hurler syndrome and the inclusions described in type B of the Sanfilippo syndrome. Unlike those mucopolysaccharidoses, however, no abnormalities were found in the gangliosides in the brain of the patient with Morquio syndrome.


Animal model for the 4-kHz tonal dip.

In humans, noise exposures produce permanent hearing losses which usually begin at 4 kHz. In chinchillas, a similar pattern of hearing loss was observed following exposure for 9 or 18 days to an octave band of noise with a center frequency of 0.5 kHz. Histopathological observations of cellular degeneration showed that this exposure produced different types of damage in the basal and apical turns of the cochlea. Behavioral measures of auditory function showed that damage in the basal turn was associated with permanent threshold shifts for one to several of the high frequencies. However, moderate losses of outer hair cells commonly appeared in the apical turn without permanent threshold shifts for low-frequency tones. These findings indicate that the pure-tone audiogram may not accurately reflect the condition of the organ of Corti after noise exposure.


[Method of determining the antimicrobial activity of alcohol extracts of propolis].

It is proposed to determine the antimicrobial activity of propolys alcohol extracts by the method of subsequent dilutions in solid nutrient media. Dilution of the extracts immediately in hot agar eliminated the inhibiting effect of the extragent on the microbial growth. Opalesence appearing in the agar did not prevent estimation of the results in contrast to the method of subsequent dilutions in liquid nutrient media.


[Evaluation of the sanative action of rifampicin on the meningococcal carrier state].

The results of the epidemiological control experiment on the efficacy of rifampicin in sanation of meningococci carriers are presented. The preliminary study of rifampicin sensitivity of 41 freshly isolated nasopharyngeal meningococcal strains showed that the MIC of the drug for 63 per cent of the isolates was 0.04--0.1 gamma/ml. Sanation was performed for 2 days; 1.2 g of the drug was used during the treatment course. The results of examination of 91 meningococci carriers showed that 4 days after the sanation the specific weight of the persons isolating no meningococci was reliably higher in the experimental group than that in the control group. The coefficient of rifampicin efficiency was 70.8 per cent. 10 days after sanation the difference in the level of the carriers isolating no meningococci in the experimental and the control groups was statistically insignificant. Therefore, the carriers treated with the drug received temporary protection from the causative agent at an average for 1 week. Later on they could become carriers again. As a result of sanation no changes in the meningococcal sensitivity to rifampicin was observed.


Rosette formation by human lymphoid cells with monkey red blood cells.

The present paper describes a study on rosette formation by human lymphoid cells with monkey erythrocytes (MRBC) obtained from two species: African green monkey (Cercopithecus aethiops) and rhesus monkey (Macaca mulata). All the lymphocyte preparations obtained from peripheral blood, tonsils, thymocytes, bone-marrow and spleen biopsies formed rosettes in variable proportions with erythrocytes obtained from these two monkey species tested. Cells of established lymphoblastoid cell lines characterized as being of either B- or T-cell origin also formed rosettes with MRBC. Taken together, the data obtained suggest that the MRBC-rosette assay cannot be reliably used for identifying lymphocyte subpopulations presently known to us, thus largely contradicting the observations previously reported by others on the use of this assay. In addition, we found that the binding sites for MRBC on the above lymphoid cells are highly sensitive to trypsin treatment and can fully be regenerated within 14-15 h when these treated cells are cultured at 37 degrees.


Effect of a staphylococcin on Neisseria gonorrhoeae.

Phage group 2 staphylococcal strain UT0002 contains a large 56S virulence plasmid with genes that code for both exfoliative toxin and a specific staphylococcin termed Bac R(1). Four penicillinase-producing strains and three penicillin-susceptible strains of Neisseria gonorrhoeae were killed by Bac R(1). After 30 min of growth of the penicillin-resistant TR1 strain in 62.5 arbitrary units of Bac R(1) per ml, loss of viability was approximately 90%, and, after 5 h, an approximately 99.99% loss of viability was observed. Lysis did not accompany cell death, and 84% of the Bac R(1) added to the growth medium was adsorbed to the gonococcal cells. The extracellular supernatant fluid from a substrain of staphylococcal strain UT0002 cured of the plasmid for Bac R(1) production had no lethal effect on the gonococcal strains. Bac R(1) was also shown to have bactericidal activity against an L-form of N. meningitidis, indicating that the outer envelope of a neisserial cell is not needed for bacteriocin activity. Ten different normal human sera were unable to neutralize Bac R(1) activity. The bacteriocin lacks adsorption specificity. It binds to but does not kill Escherichia coli cells, indicating that the cell envelope of gram-negative organisms can provide protection against the staphylococcin.


UK-18892, a new aminoglycoside: an in vitro study.

UK-18892 is a new aminoglycoside antibiotic, a derivative of kanamycin A structurally related to amikacin. It was found to be active against a wide range of pathogenic bacteria, including many gentamicin-resistant strains. The spectrum and degree of activity of UK-18892 were similar to those of amikacin, and differences were relatively minor. UK-18892 was about twice as active as amikacin against gentamicin-susceptible strains of Pseudomonas aeruginosa. Both amikacin and UK-18892 were equally active against gentamicin-resistant strains of P. aeruginosa. There were no appreciable differences in the activity of UK-18892 and amikacin against Enterobacteriaceae and Staphylococcus aureus. Cross-resistance between these two antimicrobials was also apparent.


Modified peptidoglycan transpeptidase activity in a carbenicillin-resistant mutant of Pseudomonas aeruginosa 18s.

A carbenicillin-resistant mutant of Pseudomonas aeruginosa 18s was found to possess peptidoglycan transpeptidase activity significantly more resistant to inhibition by benzyl penicillin, ampicillin, carbenicillin, and cephaloridine than that of the parent strain. The mutant was more resistant than the parent strain to all of the beta-lactam antibiotics tested, and 50% inhibition values for these compounds against membrane-bound model transpeptidase activity paralleled this increase. The resistance of the mutant to kanamycin, streptomycin, and chloramphenicol was unchanged.


Improved medium for recovery and enumeration of Pseudomonas aeruginosa from water using membrane filters.

A modified mPA medium, designated mPA-C, was shown to recover Pseudomonas aeruginosa from a variety of water sources with results comparable to those with mPA-B and within the confidence limits of a most-probable-number technique. Enumeration of P. aeruginosa on mPA-C was possible after only 24 h of incubation at 41.5 degrees C, compared with 72 h of incubation required for mPA-B and 96 h of incubation for a presumptive most probable number.


Microbial and chemical transformations of some 12,13-epoxytrichothec-9,10-enes.

Resting cells of Streptomyces griseus, Mucor mucedo, and a growing culture of Acinetobacter calcoaceticus when mixed with compounds related to 12,13-epoxytrichothec-9-ene-4beta,15-diacetoxy-3alpha-ol(anguidine) produced a series of derivatives that were either partially hydrolyzed or selectively acylated. These derivatives showed marked differences in activities as assayed by antifungal and tissue culture cytotoxicity tests.


Automated food microbiology: potential for the hydrophobic grid-membrane filter.

Bacterial counts obtained on hydrophobic grid-membrane filters were comparable to conventional plate counts for Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus in homogenates from a range of foods. The wide numerical operating range of the hydrophobic grid-membrane filters allowed sequential diluting to be reduced or even eliminated, making them attractive as components in automated systems of analysis. Food debris could be rinsed completely from the unincubated hydrophobic grid-membrane filter surface without affecting the subsequent count, thus eliminating the possibility of counting food particles, a common source of error in electronic counting systems.


Effects of dispersed recreational activities on the microbiological quality of forest surface water.

The microbiological quality of forest surface waters in the Greenwater River watershed was examined to investigate the influence of heavy motorized camping in an area with no sanitary facilities. Indicator densities increased during weekend human-use periods when compared to weekdays. Increases in indicator densities were also noted downstream from heavily used camping areas when compared to upstream sites. Seasonal, weekly, and diurnal fluctuations in indicator densities were observed. This study suggests that potential health hazards exist in this watershed during periods of human use.


Denitrifying Pseudomonas aeruginosa: some parameters of growth and active transport.

Optimal cell yield of Pseudomonas aeruginosa grown under denitrifying conditions was obtained with 100 mM nitrate as the terminal electron acceptor, irrespective of the medium used. Nitrite as the terminal electron acceptor supported poor denitrifying growth when concentrations of less than 15 mM, but not higher, were used, apparently owing to toxicity exerted by nitrite. Nitrite accumulated in the medium during early exponential phase when nitrate was the terminal electron acceptor and then decreased to extinction before midexponential phase. The maximal rate of glucose and gluconate transport was supported by 1 mM nitrate or nitrite as the terminal electron acceptor under anaerobic conditions. The transport rate was greater with nitrate than with nitrite as the terminal electron acceptor, but the greatest transport rate was observed under aerobic conditions with oxygen as the terminal electron acceptor. When P. aeruginosa was inoculated into a denitrifying environment, nitrate reductase was detected after 3 h of incubation, nitrite reductase was detected after another 4 h of incubation, and maximal nitrate and nitrite reductase activities peaked together during midexponential phase. The latter coincided with maximal glucose transport activity.


Influence of crossdrafts on the performance of a biological safety cabinet.

A biological safety cabinet was tested to determine the effect of crossdrafts (such as those created by normal laboratory activity or ventilation) upon the ability of the cabinet to protect both experiments and investigators. A simple crossdraft, controllable from 50 to 200 feet per min (fpm; 15.24 to 60.96 m/min), was created across the face of the unit. Modifications of standardized procedures involving controlled bacterial aerosol challenges provided stringent test conditions. Results indicated that, as the crossflow velocities exceeded 100 fpm, the ability of the cabinet to protect either experiments or investigators decreased logarithmically with increasing crossdraft speed. Because 100 fpm is an airspeed easily achieved by some air conditioning and heating vents (open windows and doorways may create velocities far in excess of 200 fpm), the proper placement of a biological safety cabinet within the laboratory--away from such disruptive air currents--is essential to satisfactory cabinet performance.


Fecal coliform elevated-temperature test: a physiological basis.

The physiological basis of the Eijkman elevated-temperature test for differentiating fecal from nonfecal coliforms was investigated. Manometric studies indicated that the inhibitory effect upon growth and metabolism in a nonfecal coliform at 44.5 degrees C involved cellular components common to both aerobic and fermentative metabolism of lactose. Radioactive substrate incorporation experiments implicated cell membrane function as a principal focus for temperature sensitivity at 44.5 degrees C. A temperature increase from 35 to 44.5 degrees C drastically reduced the rates of [14C]glucose uptake in nonfecal coliforms, whereas those of fecal coliforms were essentially unchanged. In addition, relatively low levels of nonfecal coliform beta-galactosidase activity coupled with thermal inactivation of this enzyme at a comparatively low temperature may also inhibit growth and metabolism of nonfecal coliforms at the elevated temperature.


A/JAX inbred mice in chemotherapeutic investigations of experimental Chagas' disease.

A quick, simple and relatively inexpensive primary screen to test compounds for activity in experimental Chagas' disease is described. It involves administration of drug once only at the maximum tolerated dose five minutes after infection to a group of five A/JAX inbred mice and the determination of the time to death. The validity of the method was established by the use of standard drugs. High levels of activity were detected with bisquinaldines, arsenobenzenes, 5-nitrofurans and 2-nitroimidazoles.


[Characterization of surface molecules of Entamoeba invadens].

In this paper we studied the externally disposed plasma membrane antigens in Entamoeba invadens using the enzymatic iodination technique. Sodiumdodecyl-sulfate-polyacrylamide gels of solubilized trophozoite proteins revealed six labeled peaks ranging from 14,000 to 67,000 daltons in gradient slab gels. Over 60 percent of the labeled was in the polypeptide of 67,000 daltons. Concanavalin A binding to these trophozites induced cell surface coat release consituted by three radioactive bands of 67,000, 60,000 and 24,000 daltons. Using an affinity chromatography method with Con A conjugated to Sepharose 4B, trophozoites in culture mixture with these beads during three hours at 22 degree C, released three components that were eluted from immobilized Con A, 67,000, 60,000 and 56,000 daltons revealed in gradient slab gels. In several determinations, the 67,000 daltons glycoprotein consistently was the major surface component, and the most immunogenic as analyzed from rabbits sera immunized with whole cell extracts as well with cell surface coat.


Protective immunity and possible autoimmune regulation in amebiasis.

Hamsters were injected intradermally with axenic trophozoites of a pathogenic strain of Entamoeba histolytica, and controls similarly with culture medium. Localized lesions were produced in all animals injected with amebae. After the lesions healed all the animals were challenged intrahepatically with axenic amebae. Extensive liver abscesses were produced in all the controls, but a small abscess in only one of the vaccinated hamsters. In the protected animals the indirect hemagglutination antibody titers were high (I:512), while in the controls they ranged between 1:32 and 1:128. In the vaccinated hamster with the small abscess the IHA titer was 1:32. To investigate a possibility of autoimmune regulation in amebiasis, sera of hamsters with experimental amebic liver abscesses were acidified and tested by counter-current-immunoeletrophoresis against untreated autologous sera and amebic antigen. In most cases there was a direct correlation between the two titers. With human antisera in autologous systems there was reactivity against anti-amebic IgG antibodies. These experiments showed that autoimmune complexes may be present in cases of hepatic amebiasis. As in chronic hepatitis and some malignancies, it is possible that immune complexes have an immunoregulatory function in amebiasis.


[Calicial diverticulum].

The authors present a series of five cases of calicial diverticuli with important radiological diameters, an entity which is not frequently described in literature. They analyse the symptoms which, because they do not posses characteristics of their own, correspond instead to the clinical picture of their complications. The radiological study is the basis of the diagnosis, when the diverticular cavity is refilled late, when the contrast of the same is drained slowly and when there is a communication with the pelvis or calix, hence the importance of the late plates. The treatment which should be expectant in asymptomatic cases and which leads us to opt for a cystectomy with excision of the epithelial bed and closure of the communication where surgery is indicated.


Morphology of freeze-etched Treponema refringens (Nichols).

The freeze-etch technique was used to study the morphology of Treponema refringens (Nichols). There is a single band of cytoplasmic fibrils which follows a path in the form of a right-handed helix with a periodicity of 1500 nm around the body of the treponeme just below the cytoplasmic membrane. There are two major fracture planes, one located in the interior of the outer envelope and the second in the interior of the cytoplasmic membrane. The "blebs" or "surface protuberances", which are quite prominent in negative-stained preparations, were not evident with freeze-etch preparation, indicating they are not a part of the normal structure of this organism. The outer envelope in untreated cells was observed to closely fit the body of the treponeme, whereas the outer envelope of glutaraldehyde-treated cells had a loose, wrinkled appearance. Thus the "loose-fitting" outer envelope generally described for treponemes is most likely an artifact of preparation for negative-staining and thin-sectioning.


Arginine catabolism in Aphanocapsa 6308.

The catabolic products of arginine metabolism were observed in Aphanocapsa 6308, a unicellular cyanobacterium, by thin layer chromatography of growth media, by limiting growth conditions, and by enzymatic analysis. Of the organic, nitrogenous compounds examined, only arginine supported growth in CO2-free media. The excretion of ornithine at a concentration level greater than citrulline suggested the existence in Aphanocapsa 6308 of the arginine dihydrolase pathway which produced ornithine, CO2,NH4,+ adenosine 5'-triphosphate. Its existence was confirmed by enzymatic analysis. Although cells could not grow on urea as a sole carbon source a very active urease and subsequently an arginase were also demonstrated, indicating that Aphanocapsa can metabolize arginine via the arginase pathway. The level of enzymes for both pathways indicates a lack of genetic control. It is suggested that the arginase pathway provides only nitrogen for the cells wheras the arginine dihydrolase pathway provides not only nitrogen, but also CO2 and adenosine 5'-triphosphate.


Uracil incorporation in the forespore and the mother cell during spore development in Bacillus subtilis. Autoradiographic electron microscopic study.

The transcriptional activity of the two genomes of the sporangium during spore formation was determined by pulse-labeling bacteria with 3H-uracil at different times of sporulation and preparing them for high resolution autoradiography. The quantitative analysis of autoradiographs shows that uracile incorporation in the whole sporangium decreases considerably between stages II and IV. However, the variations of the transcriptional activity are not identical in the mother cell and in the forespore. The one of the mother cell decreases rapidly between stages II and III and then remains stable until the end of stage IV, whereas that of the forespore which is low at stage II increases as the forespore grows ovoid and then quickly diminishes. It is very weak at the beginning of stage IV and negligible at the end of this stage. Pulse-chase experiments made in the presence of rifampine indicate that about 30% of the uracile incorporated is located in stable RNA. This value is found at any stage of sporulation in both cellular compartments whatever their rate of uracile incorporation. A relationship can be made between the nuclear shape and the activity of the genetic material. This confirms observations made by several authors in other bacterial species and other physiological conditions that the condensed shape corresponds to a state of low transcriptional activity whereas the more irregular and dispersed shape corresponds to a state of high activity.


Ultraviolet light-induction and photoreactivation of thymine dimers in a cyanobacterium, Anacystis nidulans.

Partially photoreactivable mutant of Anacystis nidulans demonstrates partial photorepair of thymine dimers. The wild type which is completely photoreactivable at the conditions studied shows higher level of thymine dimer photolysis.


The effect of cycloheximide on Euglena gracilis phenylalanyl-tRNA synthetases.

The effect of cycloheximide on the chloroplastic, cytoplasmic and mitochondrial phenylalanyl-transfer RNA synthetases of Euglena gracilis was studied by growing both logarithmic and stationary phase cultures in the presence of the antibiotic. Enzyme activity was measured relative to untreated control cultures. At very low concentrations of cycloheximide (1 microgram/ml), all three log phase enzymes showed an increase in activity of 40-50%. At slightly higher concentrations (2.5 microgram/ml), the phenylalanyl-tRNA synthetase activities were comparable to those of the control cultures. At a cycloheximide concentration of 5 microgram/ml the enzyme activities from stationary phase cultures showed only very slight decreases (5-20%). The cytoplasmic and mitochondrial enzymes behaved similarly in log phase cultures at this concentration. However, the chloroplastic phenylalanyl-tRNA synthetase from log phase cultures treated with 5 microgram/ml cycloheximide showed a marked decrease in activity (70%). A further increase in antibiotic concentration to 10 microgram/ml resulted in significant losses of activity of all three enzymes, from both growth stages. The implications of the data with regard to identification of the site(s) of chloroplast enzyme synthesis are discussed.


Deranged anterior pituitary responsiveness to hypothalamic hormones in depressed patients.

Abnormal anterior pituitary (AP) responsiveness to acute administration of thyrotropin-releasing hormone (TRH) and luteinizing hormone-follicle stimulating hormone-releasing hormone (LH-RH) was investigated in 14 patients (two men and 12 women) suffering from primary affective disorders. In ten, TRH, 500 microgram given intravenously, induced a rise in plasma growth hormone (GH) level, while in eight patients it induced a rise in plasma levels of FSH or LH or both. When LH-RH, 150 microgram was administered intravenously to ten patients, it induced a rise in plasma GH level in one patient and increased plasma prolactin level in three patients. Collectively, in only three of 14 patients was conventional AP responsiveness to hypothalamic neurohormones present. These findings demonstrate the existence of a profound derangement of AP responsiveness to hypothalamic neurohormones in depressed patients and suggest that a primary alteration in the physiologic links between the central nervous system and the AP may be at the origin of the neuroendocrine disturbance.


[Delayed implantation and early developmental defects in the mouse caused by PCB: 2,2'-dichlorobiphenyl (author's transl)].

Pregnant mice administered per os during the days 1--3 (plug day = day 0) post conception (p.c.) with 375 mg 2,2'-dichlorobiphenyl/kilograms/day revealed on day 18 p.c. increased number of resorptions and doses up from 500 milligrams/kilograms/day led to a substantial retardation of the prenatal development. Based on the investigations during the early development it is concluded, that retardation of the fetus is caused by a delayed implantation. It is assumed, that increased resorption is due to kyematopathies detected during the periimplantation period.


[Statistical study of the relationships between the etiology and clinical picture of cerebral palsy].

846 children with cerebral palsy were examined and 642 were selected for a statistical study by correspondence analysis. The aim was to identify without any prior assumptions, the relationships between the aetiological factors and the clinical findings. The study was completed by conventional statistical analysis of 584 of the cases. Small birth weight and a history of abnormal pregnancies was associated with a cerebral palsy affecting both legs, and often with a squint. Resuscitation was associated with athetosis and abnormalities of posture and behaviour. If the resuscitation lasted for more than 15 minutes or there were certain problems during delivery, severe abnormalities of both arms and major speech difficulties were observed. Resuscitation for less than 15 minutes or for an unknown time or intractable vomiting during pregnancy was associated with quadraplegia. Hemiplegia was related to post natal events but the aetiology was not always known. Foeto-maternal incompatibility was associated with athetosis, deafness, severe speech problems and ophthalmoplegias.


[Tracheal diverticulum after surgical correction of esophageal atresia. 2 cases].

Two newborns with congenital oesophageal atresia (Gross type III) were successfully operated on during the neonatal period. The occurrence, several months later, of chronic cough, associated in one infant with impaired pulmonary ventilation, lead to the discovery of a tracheal diverticulum, on the pathway of the primitive tracheo-esophageal fistula. Surgical excision of the diverticulum was followed by complete recovery.


Trifluridine in resistant human herpetic keratitis.

Patients with active dendritic herpetic keratitis that was unresponsive to topical idoxuridine or vidarabine therapy were treated topically with a 1% solution of trifluridine. The dendritic ulcers healed in an average time of 6.1 days. All patients tolerated the drug well except for one patient in whom a reversible crystalline epithelial keratitis developed.


Impedance audiometry in the squirrel monkey. Sensorineural losses.

Acoustic reflex thresholds for pure tones and broad-band noise were measured before and after treatment of nine squirrel monkeys. Experimental sensorineural hearing loss was created in six monkeys by ototoxic drug administration (gentamicin) and in three monkeys by exposure to intense sound. Extent of cochlear damage was assessed histopathologically, in the case of permanent loss induced by the ototoxic agent, and inferred from exposure history, in the case of the reversible loss induced by exposure to intense sound. Posttreatment changes in relationships among acoustic reflex thresholds (noise-tone difference [NTD]) reflected approximate degree of cochlear damage. Serial measurement of changes in the NTD in acoustic reflex thresholds appears to be a worthwhile and useful technique for monitoring sensorineural levels in the squirrel monkey.


The terminal zone of the external auditory meatus in a variety of mammals.

Standard texts describe the human tympanic recess as a shallow depression. Khanna and Tonndorf (1975) made silicone rubber casts from the auditory meatus of cats and found the recess to be narrow and deep. Guided by this finding, casts were made from 13 different mammals. At the terminal zone of the auditory meatus, profound variations were found, the consequence of three changing parameters: (1) angle between roof of the canal and tympanic membrane; (2) shape of the isthmus; (3) shape of the recess. For all specimens, values were assigned to each of these variables and a composite graph plotted. A reasonably good correlation was found to exist: the larger the angle, the higher and narrower will be the isthmus and the more narrow and deep the recess. The acoustic implications of these findings will be assessed in a separate study.


A kinetic study of ribulose bisphosphate carboxylase from the photosynthetic bacterium Rhodospirillum rubrum.

The activation kinetics of purified Rhodospirillum rubrum ribulose bisphosphate carboxylase were analysed. The equilibrium constant for activation by CO(2) was 600 micron and that for activation by Mg2+ was 90 micron, and the second-order activation constant for the reaction of CO(2) with inactive enzyme (k+1) was 0.25 X 10(-3)min-1 . micron-1. The latter value was considerably lower than the k+1 for higher-plant enzyme (7 X 10(-3)-10 X 10(-3)min-1 . micron-1). 6-Phosphogluconate had little effect on the active enzyme, and increased the extent of activation of inactive enzyme. Ribulose bisphosphate also increased the extent of activation and did not inhibit the rate of activation. This effect might have been mediated through a reaction product, 2-phosphoglycolic acid, which also stimulated the extent of activation of the enzyme. The active enzyme had a Km (CO2) of 300 micron-CO2, a Km (ribulose bisphosphate) of 11--18 micron-ribulose bisphosphate and a Vmax. of up to 3 mumol/min per mg of protein. These data are discussed in relation to the proposed model for activation and catalysis of ribulose bisphosphate carboxylase.


Comparison of direct and modified direct complement-fixation and agar-gel precipitin methods in detecting chlamydial antibody in wild birds.

Detection of chlamydial antibody in serums of wild birds was compared for the following methods: direct complement-fixation (DCF), modified DCF (MDCF), and agar-gel precipitin (AGP). The birds species used were great-tailed grackles (Cassidix mexicanus), common grackles (Quiscalus quiscula), brown-headed cowbirds (Molothrus ater), bronzed cowbirds (Tangavius aeneus), and mourning doves (Zenaida macroura). The birds were either inoculated with an isolate of Chlamydia psittaci obtained originally from turkeys or kept as uninoculated cagemates. DCF detected antibody in most inoculated common grackles, mourning doves, and brown-headed cowbirds. It was unreliable, however, for great-tailed grackles and bronzed cowbirds inoculated intramuscularly. MDCF method detected antibody in all inoculated birds except 1 mourning dove, and gave higher titers than did DCF. AGP detected antibody in all inoculated brown-headed cowbirds and all mourning doves, 1 inoculated and 1 exposed great-tailed grackle, and none of the other 2 species. Chlamydiae were apparently transmitted to the uninoculated great-tailed and common grackles and mourning doves, for antibody was detected by all 3 methods in these species kept as uninoculated cagemates.


Acute toxicity in poults associated with carbaryl insecticide.

Death of young poults is described as probably due to abuse of carbaryl in turkey pens (14 times the recommended dose). Method of application was not as described by the manufacturer.


The isolation and partial characterization of the serum lipoproteins and apolipoproteins of the rainbow trout.

1. VLD (very-low-density), LD (low-density) and HD (high-density) lipoproteins were isolated from the serum of trout (Salmo gairdneri Richardson). 2. Each lipoprotein class resembled that of the human in immunological reactivity, electrophoretic behaviour and appearance in the electron microscope. Trout LD lipoprotein, however, was of greater density than human LD lipoprotein. 3. The trout lipoproteins have lipid compositions which are similar to those of the corresponding human components, except for their high contents of long-chain unsaturated fatty acids. 4. HD and LD lipoproteins were immunologically non-identical, whereas LD lipoproteins possessed antigenic determinants in common with VLD lipoproteins. 5. VLD and HD lipoproteins each contained at least seven different apoproteins, whereas LD liprotein was composed largely of a single apoprotein which resembled human apolipoprotein B. 6. At least one, and possibly three, apoprotein of trout HD lipoprotein showed features which resemble human apoprotein A-1.7. The broad similarity between the trout and human lipoprotein systems suggests that both arose from common ancestral genes early in evolutionary history.


Separation of gametocytes of Leucocytozoon smithi from whole peripheral turkey blood.

A technique that uses a Ficoll density-gradient system was devised for isolating gametocytes of the parasitic blood protozoan Leucocytozoon smithi from whole turkey blood. Turkey blood containing gametocytes was layered onto a 35% solution of Ficoll and centrifuged. A white band, containing gametocytes and leukocytes, was observed just below the Ficoll-plasma interphase, clearly separated from the underlying packed erythrocytes. The gametocyte-leukocyte suspension was then subjected to a second centrifugation on a continuous gradient of Ficoll ranging from 12% at the top of the tube to 35% at the bottom. The second centrifugation separated the white cells containing the gametocytes into 5 distinct layers: 1) thrombocytes; 2) gametocytes of L. smithi; 3) lymphocytes; 4) a band of monocytes; and 5) granulocytes. Recovered gametocytes, stored at 4 C in a sterile balanced salt solution, retained their morphological integrity for a minimum of 4 weeks.


Evidence on the conformation of HeLa-cell 5.8S ribosomal ribonucleic acid from the reaction of specific cytidine residues with sodium bisulphite.

The reaction of HeLa-cell 5.8S rRNA with NaHSO3 under conditions in which exposed cytidine residues are deaminated to uridine was studied. It was possible to estimate the reactivities of most of the 46 cytidine residues in the nucleotide sequence by comparing 'fingerprints' of the bisulphite-treated RNA with those of untreated RNA. The findings were consistent with the main features of the secondary-structure model for mammalian 5.85S rRNA proposed by Nazar, Sitz, & Busch [J. Biol. Chem (1975) 250, 8591--8597]. Five out of six regions that are depicted in the model as single-stranded loops contain cytidine residues that are reactive towards bisulphite at 25 degrees C (the other loop contains no cytidine). The cytidine residue nearest to the 3'-terminus is also reactive. Several cytidines residues that are internally located within proposed double-helical regions show little or no reactivity towards bisulphite, but the cytidine residues of several C.G pairs at the ends of helical regions show some reactivity, and one of the proposed loops appears to contain six nucleotides, rather than the minimum of four suggested by the primary structure. Two cytidine residues that are thought to be 'looped out' by small helix imperfections also show some reactivity.


Studies of the binding of different iron donors to human serum transferrin and isolation of iron-binding fragments from the N- and C-terminal regions of the protein.

1. Trypsin digestion of human serum transferrin partially saturated with iron(III)-nitrilotriacetate at pH 5.5 or pH 8.5 produces a carbohydrate-containing iron-binding fragment of mol.wt. 43000. 2. When iron(III) citrate, FeCl3, iron (III) ascorabate and (NH4)2SO4,FeSO4 are used as iron donors to saturate the protein partially, at pH8.5, proteolytic digestion yields a fragment of mol.wt. 36000 that lacks carbohydrate. 3. The two fragments differ in their antigenic structures, amino acid compositions and peptide 'maps'. 4. The fragment with mol.wt. 36000 was assigned to the N-terminal region of the protein and the other to the C-terminal region. 5. The distribution of iron in human serum transferrin partially saturated with various iron donors was examined by electrophoresis in urea/polyacrylamide gels and the two possible monoferric forms were unequivocally identified. 6. The site designated A on human serum transferrin [Harris (1977) Biochemistry 16, 560--564] was assigned to the C-terminal region of the protein and the B site to the N-terminal region. 7. The distribution of iron on transferrin in human plasma was determined.


A protein-glucan intermediate during paramylon synthesis.

A sodium deoxycholate extract containing glucosyltransferase activity was obtained from a particulate preparation from Euglena gracilis. It transferred glucose from UDP-[14C]glucose into material that was precipitated by trichloroacetic acid. This material released beta-(1 leads to 3)-glucan oligosaccharides into solution on incubation with weak acid, weak alkali and beta-(1 leads to 3)-glucosidase. The products of the incubation of the deoxycholate extract with UDP-[14C]glucose were analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Radioactive bands were obtained that had the properties of beta-(1 leads to 3)-glucan covalently linked to protein by a bond labile to weak acid. High-molecular-weight material containing a beta-(1 leads to 3)-glucan was also shown to be present by gel filtration. The bond linking glucan to aglycone is possibly a pyrophosphate linkage. It is proposed that in Euglena gracilis beta-(1 leads to 3)-glucan (paramylon) is synthesized on a protein primer.


The effects of digestive enzymes on characteristics of placental insulin receptor. Comparison of particulate and soluble receptor preparations.

The role of the surrounding membrane structure on the binding characteristics of the insulin receptor was studied by using several digestive enzymes. The effects observed with particulate membrane preparations are compared with those from soluble receptor preparations. beta-Galactosidase and neuraminidase had no effect on insulin binding to either particulate or soluble receptors from human placentae. Exposure to 2 units of phospholipase C/ml increased insulin binding to particulate membranes, but was without effect on the soluble receptor preparation. The increase in binding to particulate membranes was shown to be due to an increase in apparent receptor number. After 5 min exposure to 500 microgram of trypsin/ml there was an increase in insulin binding to the particulate membrane fraction, owing to an increase in receptor affinity. After 15 min exposure to this amount of trypsin, binding decreased, owing to a progressive decrease in receptor availability. In contrast, this concentration of trypsin had no effect on the solubilized receptor preparation. Because of the differential effects of phospholipase C and trypsin on the particulate compared with the solubilized receptor preparations, it is concluded that the effects of these enzymes were due to an effect on the surrounding membrane structure. Changes in receptor configuration due to alterations within the adjoining membrane provide a potential mechanism for mediating short-term alterations in receptor function.


Chrysotherapy. Suppression of immunoglobulin synthesis.

Forty-four subjects with classic or definite rheumatoid arthritis who were on individualized chrysotherapy were observed for changes in serum protein electrophoresis, immunoglobulins, and circulating lymphocyte counts. By paired variate analysis, significant declines from pretreatment values were recorded for the following--electrophoretic protein fractions: gamma, alpha-1, alpha-2, (P less than 0.05); immunoglobulins: IgM--53% (P less than 0.001), IgG--37% (P less than 0.01), IgA--34% (P less than 0.001). Rheumatoid factor decreased in 29 of 39 subjects, 15 becoming seronegative (P less than 0.001); circulating lymphocytes decreased by 27% (P less than 0.001). The maximal suppressive effect on IgG and IgM was not achieved until the third and fourth years of therapy by sustained weekly administration of gold sodium thiomalate (one year cumulative dosage, mean 2106 mg, range 1065-2,885; greater than or equal to 4 year cumulative dosage, mean 8747 mg, range 5,385-15,160 mg). An immunosuppressive effect is suggested by these results.


The costs of rheumatoid arthritis. A patient-oriented study of chronic disease costs.

To detail the cost for one year of a chronic disease, 50 patients with Stage III rheumatoid arthritis were surveyed. Direct medical costs for this group were three times the national average, and 58% of these costs were covered by insurance. Indirect costs due to lost income were at least three times the direct medical costs, and transfer payments covered only 42% of these costs. Fifty-eight percent of the study group also sustained a major psychosocial loss. Uncovered income losses were the greatest economic burden for individuals with chronic rheumatoid arthritis. This striking ratio of indirect to direct medical costs has important implications for medical practice and health policy.


An assessment of the place of follow-up pulmonary tuberculosis.

I n 1971--4 forty-four patients were found to have drug-susceptible tubercle bacilli in their sputum more than one year after a previous positive result. Comparison of their case records with those of 45 controls showed that inadequate chemotherapy was by far the most important cause of relapse. Most patients who have had adequate chemotherapy for pulmonary tuberculosis should be discharged. Patients who have certain risk factors or who have had inadequate or irregular chemotherapy should be followed up indefinitely.


BRL10833 in inhibiting exercise-induced bronchoconstriction in asthmatic children.

A double-blind controlled exercise challenge study has been performed in 16 asthmatic chlidren to show the effectiveness of BRL10833 in inhibiting exercise-induced bronchoconstriction. The children attended the respiratory laboratory on four occasions within the space of two weeks; on each occassion a routine 6-min exercise test was performed. At the first visit no drugs were given before the exercise test and all the children demonstrated abnormal exercise-induced bronchoconstriction as measured by peak expiratory flow rate (PEFR), forced expiratory volume in 1 sec (FEV1) and forced vital capacity (FVC). On the other three occasions the children were given sodium cromoglycate, BRL10833 or placebo medications before the exercise test. After sodium cromoglycate administration four children showed complete blocking and four showed partial blocking of exercise-induced bronchoconstriction. After BRL10833 four children showed complete blocking and six showed partial blocking of exercise induced bronchoconstriction. Placebo administration produced complete blocking of exercise-induced bronchoconstriction in three and partial blocking in two children. The results indicated that BRL10833 was almost as effective as sodium cromoglycate in inhibiting exercise-induced bronchoconstriction and placebo, although less effective than the two preparations, did afford protection from exercise-induced bronchoconstriction in some of the children.


Light chain variable region sequence of rabbit antipneumococcal type III polysaccharide antibody 3368.

The amino acid sequence of the amino-terminal 111 residues (variable region) for the light chain of the homogeneous rabbit antipneumococcal type III polysaccharide antibody 3368 was determined. This sequence was obtained principally through automated Edmann degradations of the intact light chain and of peptides generated by tryptic digestion of the citraconylated light chain. With these methods only 2 mumol of purified light chain was required to determine the reported sequence. When compared with the light chains of four other antipneumococcal type III polysaccharide antibodies, the 3368 light chain exhibits a unique sequence in those segments of the variable region that contribute to formation of the antigen binding site (complementarity-determining regions) (10 or 11 residue differences in 12 positions). The 3368 light chain also demonstrates an insertion of three residues relative to the other four light chains in the complementarity-determining region at positions 89 to 98. These five light chains have greater than 80% sequence homology for the portion of the variable region which is not involved in antigen binding (framework).


Isolation of an active variable-domain fragment from a homogeneous rabbit antibody heavy chain. Physiochemical and immunological properties.

A fragment corresponding to most of the variable domain of the rabbit heavy chain (VH) was obtained by tryptic digestion of the midly reduced and aminoethylated heavy chain from rabbit antibody 3T72. The domain size peptide was purified by gel filtration and shown to extend between residues 11(Leu) and 122(Lys) of the heavy chain by sequence analysis. The molecular size of the fragment (approximately 11 000) was determined by gel filtration under denaturing conditions. Under nondenaturing conditions (20 mM sodium acetate, pH 5.5, 0.1 M NaCl), however, the fragment exists as a mixture of monomeric and dimeric species. The varable-domain fragment retains the allotypic determinants of the heavy chain (a1), as shown by double diffusion on agar plates and radioimmunoassay. Upon recombination of the heavy-chain variable-domain fragment with its homologous light chain, partial recovery of specific binding activity toward the SIII polysaccharide antigen was demonstrated. The method reported here is reproducible (with yields varying between 40 and 60%) and may provide a general method for obtaining the variable region of the heavy chain for antigen binding and allotypic and amino acid sequence studies.


Addition of poly(adenylic acid) to RNA using polynucleotide phosphorylase: an improved method for electron microscopic visualization of RNA-DNA hybrids.

We have observed that the enzyme polynucleotide phosphorylas from M. luteus or from E. coli will polymerize adenosine (A) from adenosine diphosphate onto 3' ends of RNA molecules. For gene mapping, the poly(A)-tailed RNA is hybridized to its complementary sequence on a longer DNA strand. The position of the poly(A)tail, and thus the position of the 3' end of the RNA on the DNA strand, can then be observed by electron microscopy. Our preferred mapping technique involves the synthesis of a poly(A)-specific label by polymerization of a poly(dBrU) tail onto one or both ends of a linear duplex DNA of defined length (a restriction fragment) and hybridization of this label to the poly(A) tail. In test experiments with a plasmid containing a Drosophila DNA sequence coding for 5S rRNA genes, overall labeling efficiencies of 70--80% were achieved.


Application of a new surface labeling reagent, EDTA derivative, on erythrocytes and platelets.

The modes of binding of a new class of impermeant metal-chelating probe, the complex of 111In3+ to 1-(p-benzenediazonium) ethylenediamine tetraacetic acid (azo-phenyl-EDTA), to human and rabbit erythrocyte membranes and the effect of binding on the function of rabbit platelets have been studied. The metal chelate, azo-phenyl-EDTA.[111In3+] bound covalently to membrane proteins following reaction with intact erythrocytes. The amount and the pattern of labeling was assessed by sodium dodecyl sulfate (SDS)-polyacrylamide disc and slab gels for radioactivity. The pattern of labeling of intact human erythrocytes by azo-phenyl-EDTA.[111In3+], by pyridoxal phosphate-NaB3H7 and by galactose oxidase-NaB3H4 was also compared. The following results were obtained: (a) The pattern of labeling of intact human erythrocyte by azo-phenyl-EDTA.[111In3+] differed from other commonly used probes for labeling external membrane surfaces. Five polypeptides were labeled by the metal chelates. In addition to the known major proteins (protein band III, PAS-1, PAS-2 and PAS-3 of Fairbanks et al. (1972) Biochemistry 10, 2606--2617) a protein (radioactive band 4) which migrated slightly slower than PAS-3 in SDS gel was labeled heavily by the metal chelate. This protein has an apparent molecular weight of 37,500 in 8.4% acrylamide-SDS gel. About 40% of bound radioactivity was found in this protein. The diazo linkage of the metal chelate to this protein was found to be especially unstable to heat. (b) In rabbit erythrocyte membranes, the metal chelate bound to three polypeptides with apparent molecular weights of 96,000, 43,000 and 33,000 in 8.4% acrylamide gel. They are probably glycoproteins in nature. (c) The binding of the probe to platelets did not affect the platelet aggregability induced by adenosine diphoshpate. In vivo studies indicated that the labeled platelets accumulated at the plague of atherosclerotic rabbits. (d) The bifunctional analog of EDTA may permit new applications of metals with useful physical properties for studies of cell membranes.


DNA synthesis in toluene-treated bacteriophage-infected minicells or Bacillus subtilis.

Bateriophage (phi29, SPP1, or SPO1)-infected, toluene-treated minicells of Bacillus subtilis are capable of limited amounts of non-replicative DNA synthesis as measured by incorporation of [3H]dTTP into a trichloroacetic acid-precipitable form. The [3H]dTTP is covalently incorporated into small DNA fragments which result from the degradation of a small percentage of the infecting phage genomes (molecular weights in the range of 2 . 10(5)). Short exposure of the DNA molecules containing the incorporated [3H]dTMP to Escherichia coli exonuclease III results in over 90% of the E13H]dTMP being converted to a trichloroacetic acid-soluble form. The synthesis is totally dependent on host-cell enzymes and is not inhibited by the addition of chloramphenicol, rifampicin, nalidixic acid and mitomycin C and only slightly (approx. 20%) inhibited by the addition of 6-(p-hydroxyphenylazo)-uracil.


Development of glycogen and phospholipid metabolism in fetal and newborn rat lung.

Glucose, a major metabolic substrate for the mammalian fetus, probably makes significant contributions to surface active phospholipid synthesis in adult lung. We examined the developmental patterns of glycogen content, glycogen synthase activity, glycogen phosphorylase activity and glucose oxidation in fetal and newborn rat lung. These patterns were correlated with the development of phosphatidylcholine synthesis, content and the activities of enzymes involved in phosphatidylcholine synthesis. Fetal lung glycogen concentration increased until day 20 of gestation (term is 22 days) after which it declined to low levels. Activity of both glycogen synthase I and total glycogen synthase (I + D) in fetal lung increased late in gestation. Increased lung glycogen concentration preceded changes in enzyme activity. Glycogen phosphorylase a and total glycogen phosphorylase (a + b) activity in fetal lung increased during the period of prenatal glycogen depletion. The activity of the pentose phosphate pathway, as measured by the ratio of CO2 derived from oxidation of C1 and C6 of glucose, declined after birth. Fetal lung total phospholipid, phosphatidycholine and disaturated phosphatidylcholine content increased by 60, 90 and 180%, respectively, between day 19 of gestation and the first postnatal day. Incorporation of choline into phosphatidylcholine and disaturated phosphatidylcholine increased 10-fold during this time. No changes in phosphatidylcholine enzyme activities were noted during gestation, but both choline phosphate cytidylyltransferase and phosphatidate phosphatase activity increased after birth. The possible contributions of carbohydrate derived from fetal lung glycogen to phospholipid synthesis are discussed.


Crystallization and positional specificity of hydroperoxidation of Fusarium lipoxygenase.

A lipoxygenase obtained from the fungus Fusarium oxysporum was purified and crystallized. Using the purified enzyme, the positional specificity of linoleate peroxidation was studied. Linoleate hydroperoxides were converted into the corresponding trimethylsilyl derivative by reduction, catalytic hydrogenation and treatment with hexamethyldisilazane/trimethylchlorosilane/pyridine and then analyzed by combined gas-liquid chromatography-mass spectrometry. Fusarium lipoxygenase was found to produce 9- or 13-hydroperoxy-octadecadienoates from linoleate. The ratio of 9- to 13-hydroperoxides produced by the enzyme was also determined by high performance liquid chromatography of their methyl esters. When the enzymic reaction proceeded at pH 9.0 and 12.0, the ratio of 9- to 13-hydroperoxide isomers was 70 : 30 and 56 : 44, respectively. With the use of the heavy isotope of oxygen (18O2), atoms of oxygen introduced into hydroperoxides were found to be derived from the gaseous phase and not from the aqueous phase.


Kinetics of formation and dissociation of metallocarboxypeptidases.

The rates of formation of a number of metallocarboxypeptidases from metal ions and bovine apocarboxypeptidase A (CPA) have been measured directly and by a competitive method. Rates were determined with pH = 6-8 by utilising the pH change attending metal-ion incorporation, employing indicator and stopped-flow. Second-order rate constants Kf, M-1 s-1 at 25 degrees C, I = 1 M NaCl, pH = 7, Tris = 25 micrometer) were 1.7 X 10(5) (Mn2+), 3 X 10(4) (Co2+), 5 X 10(3) (Ni2+), 7 X 10(5) Zn2+), and 9 X 10(5) (Cd2+). Relative incorporation rate constants were determined at 25 degrees, pH = 7.0, Tris = 0.1 M, by competing two metal ions for a deficiency of apoprotein and analyzing the products by differential enzyme activity. Agreement between the two methods was reasonable. Rate constants for dissociation of CoCPA, NiCPA, and ZnCPA were measured by loss of enzyme activity on addition of the metal ion scavenger EDTA. Values of kd at 25 degrees, I = 1.0 M NaCl, pH = 7.0 were 8 X 10(-3), 3 X 10(-5), and 4 X 10(-4) s(-1), respectively. Values of K obtained kinetically (kf/kd) were in good agreement with those determined by activity measurements of equilibrated solutions. Results are compared with those of bovine apocarbonic anhydrase, where generally significantly slower rates are encountered.


Binding affinity of bicarboxylate ions for cobalt (II) bovine carbonic anhydrase.

The affinity of bicarboxylate ions (from oxalate to glutarate) for cobalt (II) bovine carbonic anhydrase has been investigated and compared with that of acetate and propionate. The oxalate ion shows a much greater affinity for the enzyme than acetate, whereas the other bicarboxylate ions have very little tendency to bind the enzyme. In every case, and particularly for the oxalate, the apparent affinity constants dramatically increase with decreasing pH. On the basis of the electronic spectra a five-coordinate structure is proposed for all of the above derivatives. Carbon-13 NMR data have been discussed in terms of the oxalate ion chelating the metal ion and/or interacting with the wall of the active cavity.