Mezlocillin: a new acyl ureidopenicillin. Antimicrobial activity and combination effects with four aminoglycoside antibiotics.

Mezlocillin (Baypen), carbenicillin and mezlocillin combined with gentamicin, sisomicin, tobramycin and amikacin, respectively, were tested in vitro against 325 fresh clinical isolates of pathogenic gram-negative bacteria using the tube dilution procedure. The new acyl ureidopenicillin has a greater antimicrobial activity than carbenicillin against nearly all species tested. The comparison of the effectiveness of the mezlocillin-aminoglycoside combinations demonstrates the superiority of mezlocillin-sisomicin against Escherichia coli, Klebsiella, Serratia and Proteus spp., whereas mezlocillin-tobramycin is most active against gentamicin-sensitive Pseudomonas aeruginosa, and mezlocillin-amikacin against Enterobacter and gentamicin-resistant P. aeruginosa. The combination effects achieved in the majority of strains were addition or synergism. The resistance rate and the bactericidal efficacy of mezlocillin were clearly improved when mezlocillin was combined with aminoglycoside antibiotics. Mezlocillin-amikacin showed the lowest resistance rate (10.2%) followed by mezlocillin-sisomicin (12.3%), mezlocillin-gentamicin (15.1%) and mezlocillin-tobramycin (18.5%). In most species, mezlocillin-sisomicin has proved to be the combination with the best bactericidal potency.


3h-actinomycin-D binding to mitotic chromosomes of Drosophila melanogaster.

The binding of 3H-AMD to the metaphase chromosomes of Drosophila melanogaster has been analyzed after two different periods of exposure to photographic emulsion. The entirely heterochromatic Y chromosome was markedly less labelled than euchromatin and other heterochromatic regions. Moreover, the few grains present on the Y chromosome were clustered in two regions, one localized in the middle Ys and the other in the proximal third of YL. This labelling pattern is not affected by removing histones with a 2-hour treatment with 2N HCl. It is suggested that the specific underlabelling of the Y chromosome reflects a peculiar AT richness.


Glycogen storage disease: new approaches to therapy.

Detailed studies of the effect of 32 days of intravenous alimentation on the metabolic, hormonal and clinical status of a 4-year-old boy with Type I glycogenosis revealed that the biochemical abnormalities and growth failure in this disorder are a consequence of glucose lack after brief periods of fasting which results from the inborn enzyme deficiency. Long-term (1.5-5.7 years) observations of the therapeutic effects of portacaval shunt without and with continuous overnight intragastic glucose by gastrostomy in two brothers, and of continuous overnight intragastric glucose alone in five other patients with this disorder, on metabolic status and physical growth and development suggest that adequate glucose can be provided by the intragastric route without hepatic portal circulatory by-pass. The introduction of this therapy in the first year of life should prevent the serious risk to life and long-term failure in growth and development previously observed in patients with Types I and III glycogenosis.


Gas sterilization of fiber endoscopes.

Excitants of disease can be transmitted by endoscopes from one patient to another. Owing to their complicated construction and the heat-sensitive material fiber glass endoscopes are difficult to disinfect or sterilize. Of the known procedures of sterilization only gas sterilization with ethylene oxide can be used. Clinical examinations and laboratory experiments demonstrate the reliable efficacy of this procedure of sterilization of fiber glass endoscopes. On account of known risks the indications for gas sterilization are compiled.


Characterisation of the binding of virginiamycin S to Escherichia coli ribosomes.

Virginiamycin S is an inhibitor of protein synthesis in vivo. In this paper we show by equilibrium dialysis that it binds specifically to the 50-S subunit of Escherichia coli ribosomes, with one binding site per subunit. This binding is not altered by the presence of chloramphenicol, tetracycline or puromycin but is competed for by erythromycin. Using the splitting-reconstitution method, it could be demonstrated that protein L16 is absolutely required for the binding of virginiamycin S to the 50-S subunit.


Effect of cycloheximide on maturation of replicative intermediates into high-molecular-weight DNA in Tetrahymena.

Replicative intermediates of discrete size (approximately 41 S) are observed in the eukaryotic organism Tetrahymena pyriformis, when the organism is grown under defined physiological conditions. The intermediates (believed to represent replicons) are synthesized and accumulated over longer periods of time (less than 90 min), if the cells are treated with low concentrations of cycloheximide. Under these conditions the rate of total DNA synthesis is only slightly inhibited (less than 15%), while maturation of intermediate DNA into high-molecular-weight DNA is completely blocked (greater than 98%). Cycloheximide appears to inhibit the maturation process more specifically than other protein synthesis inhibitors. Studies of the accumulated intermediates on alkaline buoyant density gradients demonstrate that initiation of new putative replicons occurs during treatment with cycloheximide.


Sinal myoclonus.

A 5-year-old boy is reported with spinal myoclonus caused by cervical astrocytoma due to neurofibromatosis. The essential clinical and electromyographical signs of spinal myoclonus are the following: (1) Continuous rhythmical appearance in muscles innervated by the affected segments of the spinal cord. (2) Synchronous myoclonus in muscles innervated by the same spinal segments of one body side, asynchronous myoclonus in muscles of different segments as well as in contralateral muscles of the same segments. (3) Usually stable frequency, which may increase to a manifold under the influence of different stimuli. (4) Accentuation of the myoclonus under mental distress, disappearance during deeper sleep. The pathophysiological basis of spinal myoclonus might be a disinhibition in the area of the spinal formatio reticularis. Supraspinal stimuli can influence the myoclonus.


The association between vesical carcinoma and diverticulum of the bladder.

The incidence of vesical diverticula was determined in patients with tumours of the bladder and radiological signs of bladder-outflow obstruction and in an age-matched control group with outflow obstruction alone. No significant difference was found. This result does not support the contention that diverticula predispose to tumour formation and, therefore, prophylactic diverticulectomy is not recommended.


Rapid disappearance from serum of intravenously injected rat myeloma IgA and its secretion into bile.

Intravenously injected monoclonal rat IgA is first removed from rat serum at a very fast rate (93% in 4 h), then at a much slower rate (t/2 = 24 h). The rapid initial disappearance is thought to be due in part to secretion into rat bile. This was demonstrated by rat liver perfusions with semisynthetic medium containing diluted (1:200) IgA myeloma serum. During perfusion, the cannulated bile displayed increasingly high levels of this IgA, with a bile to medium ratio of 38 after 1 h of perfusion; at the same time, there was a 40% drop of the rat monoclonal IgA concentration in the medium, which was not observed for rat IgG2a and albumin. All of the monoclonal biliary IgA was bound to secretory component. The rat liver is thus able to actively secrete a monoclonal IgA from the circulation into bile against a strong concentration gradient.


Immunoglobulin production by lymphocyte hybridomas.

We have investigated immunoglobulin production by cloned hybrid cells. gamma2b heavy chains, but not gamma1 or gamma21, exist in two forms, differing in molecular weight, galactose labeling and charge. Hybrid molecules containing gamma2b and gamma1 heavy chains were secreted by hybridoma cells synthesizing Ig molecules of both subclasses. However, cells synthesizing both IgM and IgG chains did not secrete mu-gamma hybrid molecules. We have surveyed many new hybridomas derived from different mouse strains: 10% of those secreting IgM produce a mu chain which is about 5000 daltons larger than normal mu. Two IgM-secreting lines were studied in detail and were found to bear mu but not delta in their membrane. Intra- and extracellular mu differ in molecular size; pentameric IgM-containing molecules of both sizes were observed in the membrane fraction.


Murine heavy chain disease.

Using cultured mouse myeloma cells, it has been possible to derive cells which are now synthesizing products similar to human heavy chain disease proteins. An initial mutant was isolated which snythesized a heavy chain with an internal deletion and a normal light chain. In a subsequent step, a variant was identified in which the synthesis of the heavy chain with the deletion persisted in the absence of light chain synthesis. These experiments also demonstrate that in the MPC-11-derived cultured cell line, the continued synthesis of heavy chain does not require the synthesis of light chain.


Response to constant angular accelerations of neurons in the monkey superior vestibular nucleus.

The response to constant angular acceleration of type I neurons in the superior vestibular nucleus was studied in pentobarbital-anesthetized, cerebellectomized squirrel monkeys. Some so-called intact animals had six active semicircular canals. In other animals, the three canals on one side were rendered non-responsive by plugging, and data were obtained from the plugged and unplugged sides. There is no significant difference between the mean resting activities of units from the plugged and unplugged sides and from intact animals. Two observations appear to reflect a synergy of inputs from parallel canals. First, units on the unplugged side have roughly linear excitatory input-output relationships. Some plugged-side units exhibit an excitatory saturation. The nonlinear excitatory input-output relation of units from intact animals may result from addition of ipsilateral linear and contralateral nonlinear inputs. Second, the mean sensitivity to angular acceleration of units on the plugged and unplugged sides is significantly smaller than that of units from intact animals. Units responsive to stimulation in either of two canal planes showed response summation with simultaneous activation of the two inputs. Type III and type IV responses can result from such summation.


Further evidence of impaired tactile learning after removals of the second somatic sensory projection cortex (SII) in the monkey.

Four rhesus monkeys with bilateral removals of SII and five unoperated monkeys were trained on tests of tactile equivalence, weight discrimination and generalization, tactile discrimination learning (including concurrent learning and serial reversal) and inter-manual transfer of tactile learning. Lesioned animals were impaired on almost all tactile learning tasks and on inter-manual transfer but not on the test of tactile equivalence nor on the propioceptive tasks. The results are considered in the context of previous studies of somatosensory and parietal cortex.


Further studies on hepatic acidic glycosaminoglycans in the Hurler syndrome.

Acid GAG were isolated from hepatic tissue from 3 patients with Hurler syndrome and 3 normal controls. Gross elevations in the uronic acid and hexosamine contents were found in Hurler livers compared with the normal ones. The total GAG concentration was significantly increased (about 25fold) in Hurler patients.


Bacterial utilization of cyclo(glycl-L-prolyl).

3 strains of soil bacteria (2 Achromobacter, 1 Flavobacterium) were isolated by growth on cyclo(Gly-L-Pro) as carbon/nitrogen source. Good growth required yeast extract supplements. Utilization of cyclo(Gly-L-Pro) was inducible. Many efforts failed to obtain active cell-free preparations. Injected radioactive cyclo(Gly-L-Pro) was excreted intact by the albino rat; in contrast, injected radioactive H-Gly-L-Pro-OH was extensively metabolized.


Quantitation of nitroglycerin in human blood after administration by sustained release.

Nitroglycerin was traced in the blood of 20 patients up to 4 h after oral administration of a sustained release preparation (Nitro-Mack Retard). The determination needs an extremely sensitive method using GLC-columns with 3% SE-30 (Packard) equipped with an electron capture detector.


Extracellular fluid distribution in salt-hypertensive monkeys.

In salt-hypertensive monkeys, a decrease in plasma volume was found, inversely related to blood pressure level and not accompanied by a corresponding change in interstitial fluid volume.


[Preparations of human dopamine beta hydroxylase subunits and antibodies against these subunits (author's transl)].

The present study deals with the dissociation of human dopamine beta hydroxylase (DBH) in subunits with a mol. wt of 79,500, and the preparation of specific antibodies in rabbits. No cross-reactivity was observed between human DBH and antibodies against human DBH subunits.


Representation of moving stimuli by somatosensory neurons.

The findings obtained in neurophysiological and psychophysical investigations using tactile stimuli that move at constant velocity across the skin are reviewed. For certain neurons in the postcentral gyrus of the cerebral cortex (S-I) of macaque monkeys, direction of stimulus motion is a "trigger feature"" i.e., moving tactile stimuli evoke vigorous discharge activity in these neurons only if the stimuli are moved in a particular direction across the receptive field. This directional selectivity is maximal when stimulus velocity is between 5 and 50 cm/sec, and falls off rapidly at lower or higher velocities. The capacity for human subjects to correctly identify the direction of stimulus motion on the skin exhibits a similar dependence on stimulus velocity. The similar effects of velocity on neural and psychophysical measures of directional sensitivity support the idea that direction of stimulus motion on the skin can only be recognized if the moving stimulus optimally activates the group of S-I neurons for which that directions of simulus motion is the trigger feature.


[Effect of graded hypoxia and sodium oxybutyrate on the EEG of monkeys].

The effect on the EEG of a gaseous hypoxic mixture with 10 per cent of oxygen per se and with superimposed action of sodium oxybutyrate in a dose of 50 mg/kg was investigated in tests on macacas rhesus. Following introduction of sodium oxybutyrate an elimination of deviations from the initial EEG findings was registered.


Neural representation of cutaneous aftersensations by spinothalamic tract neurons.

Temporal summation of second pain and long-lasting tactile-evoked aftersensations are examples of sensory phenomenons that cannot be explained on the basis of responses of primary afferents. Two distinct classes of monkey spinothalamic tract neurons have responses to controlled natural stimuli that parallel and thus could account for the above phenomenons. One class, termed wide-dynamic-range, receives excitatory effects from sensitive mechanoreceptive afferents and from various nociceptive afferents including Adelta and C mechanothermal nociceptive afferents. Another class, termed nociceptive-specific, receives excitatory effects exclusively from primary nociceptive afferents. Both classes respond with an early and late response to a single noxious heat pulse (peak temperature = 51 C). The late response, unlike C nociceptive afferents but like second pain, summates in magnitude with each successive heat pulse. Gentle moving tactile stimuli evoke long-lasting (20-56 sec) after-discharges only in wide dynamic range neurons, and are similar in duration to the tactile after-sensation evoked by similar stimuli. Both the after-discharges and after-sensations can be abruptly terminated by rubbing the affected region. Temporal summation of second pain and cutaneous after-sensations are at least partly subserved by spinal cord mechanisms within the dorsal horn and are manifested in the output of spinothalamic tract neurons.


Mechanisms of neural integration in the parietal lobe for visual attention.

The impulse discharges of neurons in the inferior parietal association cortex (area 7) were studied in the alert, behaving rhesus monkey, trained to fixate and follow visual targets. Four classes of cells related to visual or visuomotor function were found. Cells of one of these are sensitive to visual stimuli and have large, contralateral receptive fields with maximal sensitivity in the far temporal quadrants. Cells of the other three classes are related to visuomotor functions: visual fixation, tracking, and saccades. They are neither sensory nor motor in the usual sense for they are activated only by interested fixation of gaze or tracking, or before visually evoked saccadic eye movements. They are not activated during the spontaneous saccades and fixations that the monkey makes while casually exploring his environment. It is hypothesized that the light-sensitive neurons provide the visual input to the visuomotor cells that, in turn, produce a command signal for the direction of visual attention and for shifting the focus of attention from one target to another.


Sensory and behavioral properties of neurons in posterior parietal cortex of the awake, trained monkey.

Neurons in posterior parietal cortex of the awake, trained monkey respond to passive visual and/or somatosensory stimuli. In general, the receptive fields of these cells are large and nonspecific. When these neurons are studied during visually guided hand movements and eye movements, most of their activity can be accounted for by passive sensory stimulation. However, for some visual cells, the response to a stimulus is enhanced when it is to be the target for a saccadic eye movement. This enhancement is selective for eye movements into the visual receptive field since it does not occur with eye movements to other parts of the visual field. Cells that discharge in association with a visual fixation task have foveal receptive fields and respond to the spots of light used as fixation targets. Cells discharging selectively in association with different directions of tracking eye movements have directionally selective responses to moving visual stimuli. Every cell in our sample discharging in association with movement could be driven by passive sensory stimuli. We conclude that the activity of neurons in posterior parietal cortex is dependent on and indicative of external stimuli but not predictive of movement.


Fertility in the rhesus monkey following long-term inhibition of ovarian function with danazol.

Danazol was previously reported to be an oral contraceptive in the rhesus monkey at doses of 200 and 400 mg/monkey/day for 90 days. The drug is now shown to be an effective long-term inhibitor of ovarian function in the monkey. In the final 3 months of a 27-month period of treatment at a dose of 400 mg/monkey/day, the drug continued to be an effective oral contraceptive. During the 27-month treatment period, three of seven monkeys were amenorrheic and the remaining had only 16 of the 109 expected menstrual cycles. Following the discontinuation of medication, all seven monkeys conceived within 2 to 6 weeks. One monkey aborted early in pregnancy and the remaining six delivered normal, healthy infants at term. Therefore, following the discontinuation of long-term treatment with danazol in the monkey, there was rapid and complete return of normal ovarian function.


Studies of long-range density effects on the proliferation of 3T3 and RLCW cells in recirculated medium.

Swiss mouse 3T3 cells and rat liver-derived RLCW cells were grown in monolayers and perfused with culture medium. A flow-rate dependent increase in the growth rate was observed both by 3H-thymidine uptake and by a rise in cell numbers. The characteristics of the response were dependent on the recirculating volume and on whether serum was present in the culture medium. In RLC cultures perfused with serum-supplemented medium the growth promoting effect decreased with increasing density of the cells. In the absence of serum, recirculation of NCTC medium had no effect on RLCs but increased growth was observed in recirculated MEM. In 3T3 cultures, a linear response was observed over a limited density range in the presence of 10% serum-supplemented medium indicating that substances present in the serum substantially modify the behaviour of the monolayer to perfusion. In serum-free medium the effect of perfusion on 3T3 cultures was confined to a small density range and was consistent with the more rapid removal of a diffusible inhibitor from the pericellular environment by recirculating the medium. Treatment of the perfusing medium with immobilised proteinases (trypsin, chymotrypsin, protease) did not alter the response except in the presence of putrescine.


[Pseudomonas bacteriophage phiKZ--possible model for studying the genetic control of morphogenesis].

A description of structure and some bilogical properties of a new virulent Pseudomonas phage, phiKZ, is given in the paper. Phage phiKZ has unique characteristics of particle (a large size of the head and the presence in the head of a structure which is visible on electron micrographs as crosswise striated substance). A cause of high-titer phage phiKZ formation on plates has been investigated. Characteristics of single step growth experiments, sensitivity of free phage particles to heating, SDS-treatment, UV-light, cycles of freezing-thawing, osmotic shocking are determined. No exclusion is found of homologous phage at early stages of infection bacteria with phiKZ. Mutants of phiKZ with changed plaque morphology are isolated; the possibility of a recombination between the mutants is demonstrated. It is supposed that phage phiKZ may be used as excellent model for the study of genetic control of morphogenesis and packing of DNA in the phage head.


Cloned seventeen-nucleotide-long synthetic lactose operator is biologically active.

A 17-nucleotide-long synthetic DNA molecule constituting the minimal recognition sequence of the lactose operator has been cloned in E. coli using the vehicle pBR313 and a synthetic HindIII adaptor. The clones containing the lac-pBR313 hybrid DNA constitutively produced beta-galactosidase. The level of beta-galactosidase was high and comparable to that obtained in cells carrying a 21-nucleotide-long synthetic lac operator on pMB9 plasmid or cells carrying a natural lac operator on pOP203-1 plasmid.


Characterization of small plasmids from Staphylococcus aureus.

Small molecular weight plasmids from Staphylococcus aureus were characterized with respect to size, restriction enzyme cleavage pattern and transforming capacity. The plasmids pS194 and pC194 which encode streptomycin and chloramphenicol resistance respectively contained 3.0 and 2.0 megadaltons of DNA as determined by zonal rate centrifugation and electron-microscopy. Both plasmids transformed S. aureus wigh high efficiency. Plasmid pC194 contained only one cleavage site for endonuclease HindIII and pS194 contained single cleavage sites for HindIII and EcoRI. A natural recombinant between these two plasmids, pSC194, shared the high transforming capacity of the parental plasmids and contained one EcoRI site and two HindIII sites. pSC194 DNA also transformed B. subtilis with high efficiency. The recombinant plasmid pSC194 may be used as an EcoRI vector for construction and propagation of hybrid DNA in S. aureus as shown in the following paper (Löfdahl et al., 1978).


Inhibition of PGI2 (prostacyclin) synthesis in the arterial wall enhances the formation of white platelet thrombi in vivo.

Prostaglandins play an important role in platelet-vessel wall interactions. Cyclic endoperoxides PGG2 and PGH2 are transformed to PGI2 (prostacyclin) within the arterial wall. The inhibition of PGI2 synthetase results in a marked enhancement of white platelet thrombus formations. This finding suggests that the synthesis of PGI2 from PGG2 and PGH2 accounts for the nonthrombogenicity of the endothelium.


Banding patterns of the chromosomes of Nycticebus coucang (Boddaert, 1785).

Q, G and C banding studies of the chromosomes of Nycticebus coucang permitted the identification of all pairs in the complement. The X was a long submetacentric, the Y a rather long metacentric. All members of the complement were non-acrocentrics. The differences between the 2n = 52 and the 2n = 50 karyotypes described in this species may be due to a centric translocation mechanism.


Development of behavior in Lemur macaco in the first nineteen weeks.

Eight young Lemur macaco were observed in a zoo during the first 19 weeks of life. Data were obtained on general motor development, development of independence from the mother, contributions of mothers and young to thhe maintenance of proximity, nursing, grooming, relations between twins, play, feeding, sexual dichromatism, and communication. The results are compared with similar data from other prosimians and from anthropoid primates, and some suggestions made about the evolutionary and adaptive significance of certain aspects of L. macaco development.


Postreproductive survival and behavior in a free-ranging female mangabey.

The survival and behavior of a postreproductive free-ranging female Cercocebus albigena is documented; her patterns of grooming, aggression, and other social and spatial relationships with members of her group are described and compared with those of other, cycling, females. The phenomenon of female postreproductive survival in free-ranging nonhuman primates is briefly reviewed.


Middle ear morphology of Lemur variegatus. Some implications for primate paleontology.

The middle ear morphology of Lemur variegatus, a strepsirhine primate, is described. Although no promontory branch of the internal carotid artery appears, there is a well-developed "promontory canal" containing a nerve trunk. This structure, which is previously undescribed in strepsirhines, is made up of the tympanic nerve and the internal carotid nerve. The implications of this discovery for paleontology, systematics, and future research are discussed.


Genetic variation of aldehyde dehydrogenase in primates.

Genetic variation of aldehyde dehydrogenase has been demonstrated in catarrhine primates. The results are in accordance with the formal genetic interpretation: three alleles, AldDH1, AldDH2, AldDH3, at the gene locus AldDH. Obviously, the allele AldDH1 has undergone fixation in Homo and Hylobates, the allele AldDH2 in Macaca and Papio, and the allele AldDH3 in Cercopithecus.


[Transplantation of isolated islands of Langerhans for the treatment of diabetes mellitus].

The historical development of research on islets transplantation is briefly reviewed followed by a description of a technique using Ficoll gradient separation to obtain islets of Langerhans from the rat pancreas. It was possible to increase the number of islets obtained with this procedure by modifying it in several ways. The islets were then transplanted in an isologue manner into rats with streptocotocin-induced diabetes. The effect of transplantation on glucose metabolism in these rats was evaluated both by determination of the general symptoms typical for diabetes as well as by the performance of such functional tests as IVGTT, GAC and tolbutamide test. The islets could be transplanted into various locations, but it was found that positive results were obtained only if the liver was the site of application and if than 2100 islets were used. Three hours after transplantation normalization of blood sugar levels and serum insulin could be observed; these levels remained stable over 18 months. It was possible to transplant in the same animal several times. This had an effect on the metabolism which was equivalent to the sum of the separate transplants. By means of both light and electron microscope examination the morphological changes which the transplanted islets underwent at the site of transplantation were observed.


[Early diagnosis of neoplasms in Baden-Württemberg].

In the Federal Republic of Germany an official cancer screening program was started on July 1, 1971. The following sites are included: for males rectum and prostate, for females breast, cervix uteri, corpus uteri, ovaries, rectum. Since January 1, 1975, in addition skin, male genitalia, bladder and kidneys are screened. The official reports only deal with suspects and not with confirmed cases. The cancer registry of Baden-Württemberg is particularly  involved in evaluating the results of the screening program. Of all confirmed cancers of the cervix uteri more than 50 per cent are detected by screening. Non-invasive forms represent a little more than 50 per cent of the total. Approximately one case in six of cancer of the breast or cancer of the prostate is diagnosed by screening procedures. In all other sites the percentage is 5 or less. The participation in the program varies by age and sex. Half of the women in the age group from 30 to 44 participate in the program but only about 3 per cent in the group over 75. In males, the program is aimed at the higher age groups. The total expenditure of the social health insurance in 1976 was 63 billion DM, for all screening measures 403 million DM, that is 0.6 per cent of the total amount. The contribution of the program in relation to the total cancer incidence is discussed.


[Early diagnosis of hereditary metabolic diseases in the newborn by means of microbiological Guthrie tests].

The screening method of so-called bacterial inhibition and growth assay (Guthrie-test) is described and the number of newborn babies tested over the whole of West Germany from 1969--1974 is stated. 626 cases of PKU were detected early and tested after these dates. Only a few german laboratories are able to detect other inborn errors of metabolism such as MSUD (maple syrup urine disease), homocystinuria, histidinemia and galactosemia. The ratio of illnesses therefore can only be estimated and not stated with statistical evidence.


Observations on the use of an autoanalyser and a manual technique for the detection of the red cell antigens C, D, E, c, K and S in bloodstains.

The results of the detection of the Rh antigens (D, C, E, c) and the S and K antigens by an elution technique comparing the AutoAnalyser and a manual method for the detection of eluted antibody are presented. Investigation of a series of anti-D sera demonstrates that while it is not too difficult to select antisera for the detection of the D antigen in fresh stains (i.e. up to 4 weeks old) some of these sera will not be satisfactory for typing older stains. The AutoAnalyser system used is low ionic strength/polybrene which has a capacity for detecting 0.002 microgram/ml of anti-D, but in spite of this degree of sensitivity the manual technique for the detection of eluted antibody compared very favourably and tended, where older stains were being tested, to give more conclusive results than the AutoAnalyser. The superiority of the manual technique was most pronounced when detecting the S and K antigens.


Steroids and sexual behavior in castrated male rheusus monkeys.

Sexual behavior of long-term castrated rhesus males was not increased by administration of the hydroxylated metabolite of testosterone (T), 17 beta, 19-dihydroxy-5 alpha-androstan-3-one diacetate (19-OH-DHTA) at a dose of 1 mg/kg/day. Simultaneous administration of 19-OH-DHTA and estradiol benzoate (EB) also failed to increase the level of sexual performance, but daily injection (1 mg/kg/day) of testosterone propionate (TP) was very effective in effective in activating sexual behavior.


Teamwork eases long-term care admissions.

The teamwork approach to admitting new residents used at St. Joseph Home of Chicago, Inc., is based on the philosophy that proper admission of new residents can help lessen the apprehensions as elderly people open the "forbidding" door to a long-term care facility. The team approach begins before an applicant is accepted and continues through the new resident's initial period of adjusting to a new home.


[Gram-positive bacteria as adjuvants (author's transl)].

Gram-positive bacteria or their constituents influence the immune response in various ways. For instance: Listeria monocytogenes cellwall fraction acts as B-cell mitogen, in this way enhancing a humoral immune response. Corynebacterium parvum acts as a macrophage stimulant evoking maximum macrophage activation. BCG, among other things, leads to an enhancement of a T-cell response to unrelated antigens. The hypothetical therapeutic consequences are discussed.


Selective platelet immune retention.

Inbred rats were immunized with Freund's Adjuvant containing proteins of Mycogacterium tuberculosis, Mycobacterium butyrium, or dinitrochlorobenzene (DNCB). Native arterial blood was then passed over glass beads coated with those antigens. The platelet retention on the glass beads was measured. Rats immunized with Complete Freund's Adjuvant developed accelerated platelet retention on beads coated with protein derivatives of tuberculosis (PPD) after just 18 seconds of blood flow. Rats immunized twice maintained selective retention longer than those immunized once. The test animals developed no cutaneous hypersensitivity nor precipitin lines on Ouchterlong gels against PPD. Rats sensitized to DNCB had accelerated platelet retention on DNCB-coated beads. Results were temperature and complement dependent, and antigen-specific. Heparin caused a dose-dependent inhibition of the accelerated retention. PPD potentiated the ADP-induced aggregation of plateletrich plasma (PRP) from immunized rats. These experiments suggest that platelets react selectively to antigens in the intravascular spaces in immune reactions.


Further studies on the H-2 linked dependence of the adjuvant action of Brucella abortus.

The B 19S strain of Brucella abortus is found to act as an adjuvant to the anti-sheep red blood cell (SRBC) reaction in some congenic strains of mice but not in others. If the recipient is H-2b, there is no adjuvant effect (B 19S); neither is there (thymus-dependent anti-B 19S reaction as measured by thymocyte activation and change of electrophoretic mobility. In contrast, there is a thymus-independent anti-B 19S reaction (production of haemagglutinins) as good in the H-2b mice as in the others. Experiments with T cell deprived mice show that the adjuvant action of B 19S is thymus-dependent. As the anti-SRBC reaction without adjuvant is also thymus-dependent, it is difficult to distinguish anti-SRBC and anti-B 19S reactions from the adjuvant action of B 19S on the anti-SRBC reaction. Several explanations are possible, all involving H-2 (Ir?) controlled thymus dependent mechanisms.


A comparison of the N-terminal amino acid sequences of early and late pools of anti-DNP and anti-DNP-p-aminobenzoylglutamate antibody light chains.

The N-terminal amino acid sequences of early and late pools of anti-DNP and anti-DNP-p-aminobenzoylglutamate (DNP-ABG) antibody light chains were quantitatively determined and compared. The amino acid composition at each locus of the N-terminal 20 amino acid residues of each light chain preparation was determined using automatic sequencing techniques coupled with high-pressure liquid chromatography and mass spectrometry. The sequence data obtained for the light chains corresponding to antibodies isolated early in the immune response (3-4 weeks) were essentially the same as those for light chains from antibodies isolated late in the response (12-14 weeks). In addition, it was observed that the sequence data obtained for the anti-DNP antibody light chain preparation were almost identical to those found for two anti-DNP-ABG antibody light "hain preparations. The sequence data obtained in the present study were compared with those obtained for normal rabbit light chains and with the composite sequence data published for other rabbit anti-hapten light chains.


Further studies of immunoglobulin synthesis by guinea pig leukaemic lymphocytes.

The L2C leukaemia is a B-lymphocytic neoplasm of strain 2 guinea-pigs, maintained by passaging in vivo. It synthesizes mu and lambda immunoglobulin chains. These combines to form monomeric (7S) IgM molecules which are inserted into the plasma membranes. From here they are shed as monomeric IgM and as a species of higher molecular weight which has not been further defined. The synthesis of lambda chain is in excess of that required for the IgM molecule, the surplus being exported directly from the cell without any intervening phase in the plasma membrane. Quantitative estimates of synthetic rates and pool sizes for these immunoglobulin species are presented.


Increased phagocytic activity of splenectomized mice challenged with Listeria monocytogenes.

Adult splenectomized mice exhibit increased resistance to infection with Listeria monocytogenes. Phagocytosis, by reticulo-endothelial cells, of test particles (51Cr-labelled sheep erythrocytes) is the same in splenectomized and control mice. However, 24 h exposure to Listeria, which fails to influence phagocytic activity of normal mice, greatly enhances the blood clearance and liver uptake of the test particles in splenectomized mice. The presence of a cell population and/or product in the spleen which modulates macrophage activation upon the exposure to appropriate stimuli is postulated.


Pregnancy-associated plasma proteins in pregnant and pseudopregnant rats.

As in the human, the rat pregnancy-associated plasma proteins consisted of four distinct entities as revealed by immunological methods. All showed gradual increases during gestation and rapid disappearance postpartum. They could be physicochemically, although not immunologically, tentatively related to the human pregnancy proteins. None of them was detectable in the serum of rats at various stages of pseudopregnancy, nor in nonpregnant rats.


Optimal conditions for Fc receptor-ligand interaction.

The temperature and time dependence for Fc receptor-ligand interaction have been investigated in a murine lymphoblastoid cell line. The binding of antibody-coated erythrocytes or radioiodinated heat-aggregated IgG was evaluated at varying time points during incubation at 4, 21 or 37 degrees C. Maximum rosette formation was observed by 10 min incubation at 21 degrees C if cell suspensions were first centrifuged at 200 g for 2 min. Longer incubation periods were required in the absence of centrifugation. The kinetics of aggregate IgG binding were similar to rosette formation without centrifugation. The results demonstrate that receptor-ligand interaction in this system is both temperature and time-dependent.


Characterization and purification of pregnancy-associated plasma protein B (PAPP-B).

PAPP-B is a pregnancy-specific beta 1-glycoprotein of large molecular weight, about 1,000,000 as determined by Sepharose 4B and 6B gel filtration. Its isoelectric point is between pH 4.6 and 5.0. It has been purified at least 800-fold from term pregnancy serum by a sequence of steps involving salting out at 30% saturation with ammonium sulfate (1,2M), DEAE-cellulose chromatography, Sepharose 4B gel filtration and hydroxylapatite chromatography. Monospecific antiserum to PAPP-B has been prepared, and used to differentiate it from several other newly reported tumor or pregnancy proteins. PAPP-B was found to increase slowly during the second trimester of pregnancy and more steeply during the third, reaching a plateau in late gestation. During the postpartum period, PAPP-B disappeared quite rapidly, with an apparent half-life of less than 1 day.


Transformation of marmoset lymphocytes in vitro with Herpesvirus ateles.

Circulating lymphocytes from three species of marmoset monkeys were transformed in vitro with Herpesvirus ateles (HVA) either by co-cultivation with lethally X-irradiated HVA-producing lymphoblastoid cells or by infection with cell-free virus: 42 transformed lymphocyte cultures were obtained from 72 transformation attempts. Attempts to transform squirrel monkey lymphocytes were unsuccessful in 29 attempts. Association of HVA with each transformed culture was demonstrated by staining of antigen-positive cells in indirect fluorescent antibody (FA) tests and recovery of HVA after co-cultivation of the transformed cells with permissive monolayer cells. Cells of most transformed cultures possessed T lymphocyte properties: i.e. formation of E rosettes, reactivity with a specific anti-marmoset T lymphocyte serum and lack of surface Ig.


Acrodermatitis enteropathica.

Acrodermatisis enteropathical has had a remarkable hist orical course beginning only 35 years ago, first passing through a period of empiric management with the 8-hydroxyquinoline drugs and culminating with the discovery of a biochemical effect in zinc absorption. This accomplishment has not only resulted in the cure of a serious and often fatal disease, but has opened the way to an understanding of the vital role played by the trace element zinc in many previously unrecognized areas of human physiology.


Nerve fiber layer astrocytes of the primate retina: morphology, distribution, and density.

The nerve fiber layer of several species of primate has been studied with Golgi silver impregnations and Nissl stains. The morphology, distribution, and density of fibrous astrocytes of the nerve fiber layer are described. Two classes of astrocytes are distinguished: elongated cells whose processes parallel the nerve fiber bundles and cover a rectangular area more than two time longer than wide without vascular contact and stellate cells whose processes conspicuously cross the nerve fiber bundles to cover a slightly oval area and make unspecialized vascular contacts. The density of glial nuclei in the nerve fiber layer is roughly proportional to the thickness of the layer all across the retina, so that higher densities are seen in specialized regions such as the arcuate nerve fiber bundles and near the optic disc.


Demonstration of tyrosinase in the adult bovine uveal tract and retinal pigment epithelium.

Modern techniques offer an opportunity for a more complete evaluation of melanin production in the uvea and retinal pigment epithelium (RPE). By measuring the release of tritium from tritiated tyrosine in homogenized samples of adult bovine RPE as well as iris and choroid, tyrosinase activity could be demonstrated in both the uveal tract and the RPE. Phenylthiourea, a specific tyrosinase inhibitor, markedly decreased tyrosinase activity, whereas 3-iodo-tyrosine, a tyrosine hydroxylase inhibitor, had no effect. These techniques indicate tyrosinase activity in the uveal tract and the RPE of adult cattle. This is the first biochemical demonstration of tyrosinase in adult RPE.


The effect of blood on ocular fundus reflectance and determination of some optical properties of retinal blood vessels.

Light reflected from 50 micrometer diameter spots on the fundi of two adult rhesus monkeys was measured as a function of wavelength in 10 nm jumps from 400 to 900 nm. The areas measured were a retinal artery and vein, areas of the disk, macula, and retina devoid of visible blood vessels, and the foveola. The eyes of the two monkeys were then exsanguinated, the blood vessels were filled with normal saline, and reflectances of the same spots on the fundi were again measured. The pairs of reflectance curves were compared and demonstrated that blood is not the major determinant of the characteristic shapes of the retinal tissue spectral reflectance curves. From the pre-exsanguination and postexsanguination data, the isolated retinal artery and vein wall reflectances were determined to be 0.020 and 0.009, respectively, their transmittances 0.837 and 0,977, and the fractions of light absorbed by them 0.143 and 0.014, respectively, in the visible spectral region below 500 nm.


Ultrastructural localization of calcium around the membrane of the surface connected system in the human platelet.

The localization of calcium in the membrane system of human platelets was determined by ultrahistochemical methods equipped with an electron probe x-ray microanalyzer. After potassium oxalate-glutaraldehyde treatment large amounts of electron opaque precipitates were observed around the membrane of the surface connected system. Electron probe x-ray microanalysis clearly defined that the precipitates were composed of calcium oxalate. The localization of calcium on the membrane of the surface connected system was also confirmed even after treatment of the platelets with potassium antimonate-OsO4. These results support a model which depicts the surface connected membrane system taking part in the store and the transport of calcium.


Localization of 99mTc-labeled immune splenocytes at tumor site and detection by gamma camera imaging.

Better diagnostic techniques are needed to delineate the size and location of malignant primary tumors and metastatic deposits. We are developing an in vivo tumor visualization technique utilizing 99mTc-labeled immune lymphoid cells. We have investigated the in vivo organ and tissue distribution and tumor localization of 99m sodium pertechnetate, 99mTc, reduced by SnCl2 - 2H2O and 99mTc-labeled viable immune and nonimmune lymphocytes 32 hrs after injection into tumor-bearing mice. The distribution and tumor localization of the 99mTc-labeled compounds and splenocytes within tumor-bearing mice after 32 hrs was determined by imaging with a gamma camera and was quantitated by counting the various dissected organs and tissues in a gamma counter. The gamma camera images and postmortem dissection and counting showed localization of radioactivity at several sites, including the tumor in mice injected with radiolabeled immune splenocytes whereas studies utilizing other 99mTc-labeled substances or nonimmune cells showed little or no localization of radioactivity in the tumor when compared to normal leg tissue.


Comparison of the effect of vasopressin infusions on the mesenteric arteries of different species.

Superior mesenteric arteriography was performed following infusions of vasopressin, epinephrine and saline into the superior mesenteric arteries of dogs, pigs, rabbits and monkeys and compared with the well-known effects in human beings. Species-specific effects were noted and compared with the known differences in various vascular beds in human beings. In non-primate animals, vasopressin appeared to act at a more distal site than in primates, but with a similar decrease in superior mesenteric arterial flow. The results in primates were similar to those in human beings.


Identification of antibiotics of iturin group in various strains of Bacillus subtilis.

Thirty eight strains of B. subtilis were tested for the presence of antifungal antibiotics of the iturin group. The characterization of these antibiotics was made on the basis of: antifungal activity against P. chrysogenum, isolation and purification of the active substance, quantitative analysis of alpha-aminoacids and identification of the liposoluble dipeptide obtained by partial hydrolysis. The only antibiotics of the iturin group found were: iturin A, mycosubtilin and bacillomycin L.


Disc agar diffusion antimicrobial susceptibility tests with beta-lactamase producing Neisseria gonorrhoeae.

The emergence of beta-lactamase-producing strains of Neisseria gonorrhoeae has led to a reexamination of the role of the disc agar diffusion method in susceptibility testing of gonococci. Our data show that the disc agar diffusion test can be used to screen for beta-lactamase production by these organisms. The disc tests were done on GC Agar Base supplemented with 1% IsoVitaleX. An inoculum of 10(8) colony forming units/ml and either a 10-unit-penicillin or a 10-microgram-ampicillin disc were used. A zone diameter of less than or equal to 19 mm was indicative of beta-lactamase production. These results were compared with results of chemical tests for beta-lactamase and with minimal inhibitory concentrations. Recommondations were also made for a disc test with tetracycline and spectinomycin, but these methods must remain tentative because of the lack of resistant strains.


Viability and endogenous substrates used during starvation survival of Rhodospirillum rubrum.

Cells of Rhodospirillum rubrum were grown photoorganotrophically and chemoorganotrophically and then starved for organic carbon and combined nitrogen under four conditions: anaerobically in the light and dark and aerobically in the light and dark. Illumination prolonged viability and suppressed the net degradation of cell material of phototrophically grown cells, but had no effect on chemotrophically grown cells that did not contain bacteriochlorophyll. The half-life survival times of carbohydrate-rich phototrophically grown cells during starvation anaerobically or aerobically in the light were 17 and 14.5 days, respectively. The values for starvation aerobically and anaerobically in the dark were 3 and 0.5 days, respectively. Chemotrophically grown cells had half-life survival times of 3 and 4 days during starvation aerobically in the light and dark, respectively, and 0.8 day during starvation anaerobically in the light or dark. Of all cell constituents examined, carbohydrate was most extensively degraded during starvation, although the rate of degradation was slowest for phototrophically grown cells starved anaerobically in the light. Phototrophically grown cells containing poly-beta-hydroxybutyrate as carbon reserve were less able to survive starvation anaerobically in the light than were carbohydrate-rich cells starved under comparable conditions. Light intensity had a significant effect on viability of phototrophically grown cells starving anaerobically. At light intensities of 320 to 650 lx, the half-life survival times were 17 to 24 days. At 2,950 to 10,500 lx, the survival times decreased to 1.5 to 5.5 days. The kinetics of cell death correlated well with the rate of loss of cell mass of starving cells. However, the cause of death could not be attributed to degradation of any specific cell component.


Biosynthesis of exopolysaccharide by Pseudomonas aeruginosa.

In batch cultures of Pseudomonas aeruginosa, the maximum rate of exopolysaccharide synthesis occurred during exponential growth. In nitrogen-limited continuous culture, the specific rate of exopolysaccharide synthesis increased from 0.27 g g of cell-1 h-1 at a dilution rate (D) of 0.05 h-1 to 0.44 g g of cells h-1 at D=0.1 H-1. The yield of exopolysaccharide on the basis of glucose used was in the range of 56 to 64%. Exopolysaccharide was also synthesized in carbon-limited cultures at 0.19 g g of cell-1 h-1 at D=0.05 h-1 in a 33% yield. Nonmucoid variants appeared after seven generations in continuous culture and rapidly increased in proportion to the total number of organisms present.


Early-blocked asporogenous mutants of Bacillus subtilis are lysogenized at reduced frequency by temperate bacteriophages.

The establishment of lysogeny in early-blocked asporogenous (Spo-) mutants of Bacillus subtilis 168, which were also defective in the production of antibiotics (Abs-), by temperate phage phi105 or SPO2 was studied. It was found that the frequency of lysogenization of Spo-Abs-mutants was 10 to 20% that of the wild-type bacteria. There was no difference in the efficiency of plating and the burst size of phi105 between wild-type and mutant strains. Phi105 lysogens of mutant strains were as stable as those of the wild type. Several rifampin-resistant mutants defective in the production of antibiotics were isolated. They were also defective in spore formation and lysogenized by phi105 at reduced frequency.


Biosynthesis of riboflavin in Bacillus subtilis: function and genetic control of the riboflavin synthase complex.

Two riboflavin synthase activities (heavy and light) have been observed in earlier studies with Bacillus subtilis. The heavy enzyme is a complex of one molecule of light enzyme (consisting of three alpha subunits) and approximately 60 beta subunits (A. Bacher, R. Bauer, U. Eggers, H. Harders, and H. Schnepple, p. 729--732, in T. P. Singer (ed.), Flavins and Flavoproteins, Elsevier, Amsterdam, 1976). The formation of alpha and beta subunits is coordinately controlled. Mutants apparently deficient in beta subunits were isolated as riboflavin requires after mutagenesis of B. subtilis with ICR 191. The mutants could grow with diacetyl instead of riboflavin. Growth with diacetyl was associated with the accumulation of substantial amounts of the riboflavin precursor, 6,7-dimethyl-8-(D-ribityl)lumazine. It follows that the mutants are deficient in an enzyme activity required for the formation of the lumazine from the pyrimidine precursor. We conclude that heavy riboflavin synthase is a bifunctional enzyme. The riboflavin synthase activity is mediated by the alpha subunits, whereas the beta subunits are necessary for an earlier biosynthetic step.


Recombination between compatible plasmids containing homologous segments requires the Bacillus subtilis recE gene product.

Plasmid pSL103 was previously constructed by cloning a Trp fragment (approximately 2.3 X 10(6) daltons) from restriction endonuclease EcoRI-digested chromosome DNA of Bacillus pumilus using the neomycin-resistance plasmid pUB110 (approximately 2.8 X 10(6) daltons) as vector and B. subtilis as transformation recipient. In the present study the EcoRI Trp fragment from pSL103 was transferred in vitro to EcoRI fragments of the Bacillus plasmid pPL576 to determine the ability of the plasmid fragments to replicate in B. subtilis. Endonuclease EcoRI digestion of pPL576 (approximately 28 X 10(6) daltons) generated three fragments having molecular weights of about 13 X 13(6) (the A fragment), 9.5 X 10(6) (B fragment, and 6.5 X 10(6) (C fragment). Trp derivatives of pPL576 fragments capable of autonomous replication in B. subtilis contained the B fragment (e.g., pSL107) or both the B and C fragments (e.g., pSL108). Accordingly, the B fragment of pPL576 contains information essential for autonomous replication. pSL107 and pSL108 are compatible with pUB110. Constructed derivatives of the compatible plasmids pPL576 and pUB110, harboring genetically distinguishable EcoRI-generated Trp fragments cloned from the DNA of a B. pumilus strain, exhibited relatively high frequency recombination for a trpC marker when the plasmid pairs were present in a recombination-proficient strain of B. subtilis. No recombination was detected when the host carried the chromosome mutation recE4. Therefore, the recE4 mutation suppresses recombination between compatible plasmids that contain homologous segments.


Molybdenum independence of nitrogenase component synthesis in the non-heterocystous cyanobacterium Plectonema.

The cyanobacterium Plectonema boryanum (IU 594-UTEX 594) fixes N2 only in the absence of combined N and of O2. We induced nitrogenase by transfer to anaerobic N-free medium and studied the effect of Mo starvation on nitrogenase activity and synthesis. Activity was first detected within 3 h after transfer by the acetylene reduction assay in controls, increasing for at least 25 h. Cells grown on nitrate and Mo and then transferred to N-free, Mo-free medium produced 8% of the control nitrogenase activity. Addition of W to the Mo-free medium reduced the activity to 0.5%. Under both Mo starvation conditions, nitrogenase protein components were synthesized. Component II of the cyanobacterial enzyme was detected by in vitro complementation with Mo-containing component I from Klebsiella pneumoniae or Azotobacter vinelandii but not Clostridium pasteurianum. Component I activity was restored by addition of Mo to cultures in which new enzyme synthesis was blocked by chloramphenicol. Acidified extracts of Plectonema induced in Mo-containing medium contained the Fe-Mo cofactor required to activate extracts of the Azotobacter mutant UW45 in vitro, but they did not activate extracts of Mo-starved Plectonema. Analysis of 35SO4(2-)-labeled proteins by polyacrylamide gel electrophoresis suggested that Mo is required for the conversion of a high-molecular-weight precursor to component I in Plectonema.


Intracytoplasmic membrane, phospholipid, and sterol content of Methylobacterium organophilum cells grown under different conditions.

Intracytoplasmic membranes were present in Methylobacterium organophilum when cells were grown with methane, but not methanol or glucose, as the sole carbon and energy source. Cells grown with methane as the carbon and energy source and low levels of dissolved oxygen had the greatest amount of intracytoplasmic membrane. Cells grown with increased levels of dissolved oxygen had less intracytoplasmic membrane. The amount of total lipid correlated with the amount of membrane material observed in thin sections. The individual phospholipids varied in amount, but the same four were present in M. organophilum grown with different substrates and oxygen levels. Phosphatidyl choline was present as a major component of the phospholipids. Sterols were present, and they differed from those in the type I methylotroph Methylococcus capsulatus. The relative amounts of different sterols and squalene changed with the substrate provided for growth. The greatest amounts of sterols were found in methane-grown cells grown at low levels of dissolved oxygen. None of the unusual or usual membrane components assayed was uniquely present in the intracytoplasmic membranes.


Pattern of phenazine pigment production by a strain of Pseudomonas aeruginosa.

An atypical strain of Pseudomonas aeruginosa capable of synthesizing three phenazine pigments was isolated. Cultural conditions, under which the strain forms either chlororaphin, oxychlororaphin, or pyocyanine, are described. This broad spectrum of pigment production, as well as some other characteristics, sets this strain apart from previously described chlororaphin producers.


Relationship between cellular autolytic activity, peptidoglycan synthesis, septation, and the cell cycle in synchronized populations of Streptococcus faecium.

Synchronized, slowly growing (TD = 70 to 80 min) cultures were used to study several wall-associated parameters during the cell cycle: rate of peptidoglycan synthesis, septation, and cellular autolytic activity. The rate of peptidoglycan synthesis per cell declined during most of the period of chromosome replication (C), but increased during the latter part of C and into the period between chromosome termination and cell division (D). An increase in cellular septation was correlated with the increased rate of peptidoglycan synthesis. Cellular autolytic capacity increased during the early portion of C, reached a maximum late in C or early in D, and declined during D. Inhibition of DNA synthesis during C prevented the decline in autolytic capacity at the end of the cell cycle, caused a slight reduction in the rate of peptidoglycan synthesis, delayed but did not prevent septation, and prevented the impending cell division by inhibiting cell separation. Inhibition of DNA synthesis during D did not prevent the increase in autolytic capacity during the next C phase, but, once again, prevented the decline at the end of the subsequent cycle. Thus, increased autolytic capacity at the beginning of the cell cycle did not seem to be related to chromosome initiation, whereas decreased autolytic capacity at the end of the cell cycle seemed to be related to chromosome termination. The data presented are consistent with the role of autolytic enzyme activity in the previously proposed model for cell division of S. faecium (G.D. Shockman et al., Ann. N.Y Acad. Sci. 235:161-197, 1974).


Hybridization analysis of restriction endonuclease DNA fragments of Bacillus cereus transcribed during spore outgrowth.

Transcribing Bacillus cereus DNA was visualized by means of autoradiography of electrophoretically separated EcoRI restriction endonuclease DNA fragments hybridizing 32P-labeled RNA. Hybridization of RNA of dormant spores, vegetative cells, and outgrowing spores indicates the following. (i) A large fraction of the nonribosomal RNA in dormant spores is transcribed at a limited number of regions on the bacterial chromosome. (ii) After induction of spore germination, transcription activity is not limited to a single short region on the chromosome, but rather is distributed along the chromosome. The DNA/RNA hybridization technique has been used to identify restriction endonuclease DNA fragments homologous to RNA species that are present in dormant spores but absent from vegetative cells, RNA species that are synthesized immediately after germination induction and are present at a relatively low concentration in vegetative cells, and RNA species that are transcribed at a late stage of outgrowth but are absent or present at low concentration at an early stage of outgrowth.


Properties of Bacillus cereus temperature-sensitive mutants altered in spore coat formation.

Three conditional Bacillus cereus mutants altered in the assembly or formation of spore coat layers were analyzed. They all grew as well as the wild type in an enriched or minimal medium but produced lysozyme and octanol-sensitive spores at the nonpermissive temperature (35 to 38 degrees C). The spores also germinated slowly when produced at 35 degrees C. Temperature-shift experiments indicated that the defective protein or regulatory signal is expressed at the time of formation of the outer spore coat layers. Revertants regained all wild-type spore properties at frequencies consistent with initial point mutations. Spore coat defects were evident in thin sections and freeze-etch micrographs of mutant spores produced at 35 degrees C. In addition, one mutant contained an extra surface deposit, perhaps unprocessed spore coat precursor protein. A prevalent band of about 65,000 daltons (the same size as the presumptive precursor) was present in spore coat extracts of this mutant and may be incorrectly processed to mature spore coat polypeptides. Another class of mutants was defective in the late uptake of half-cystine residues into spore coats. Such a defect could lead to improper formation of the outer spore coat layers.


N-Terminal amino acid sequence of pilin isolated from Pseudomonas aeruginosa.

The amino-terminal amino acid sequence of the pili protein from Pseudomonas aeruginosa K pili is presented. The sequence is compared with those reported by others for pilin obtained from Neisseria gonorrhoeae and Moraxella nonlique-faciens. All three sequences are highly homologous, contain only two hydrophilic residues in the first 22 positions, and contain an unusual amino acid, N-monomethylphenylalanine, at the amino terminus.


Approximation of the cell cycle in synchronized populations of Streptococcus faecium.

Slowly growing populations (TD = 70 to 80 min) of Streptococcus faecium (S. faecalis ATCC 9790) were synchronized by selection after sucrose gradient fractionation. The cell cycle was approximated by correlating the patterns of DNA accumulation and cell division. More specifically, the beginning of cell cycle was equated with the beginning of a rapid linear increase in DNA accumulation. The DNA content of the culture approximately doubled during the period of accumulation, which lasted about 51 min. The period of rapid DNA accumulation, was followed by a period of reduced accumulation that lasted about 24 min. During synchronized growth, cell numbers increased rapidly in coordination with the period of rapid DNA accumulation and exhibited a plateau during the period of reduced DNA accumulation. In contrast, RNA and protein appeared to accumulate exponentially throughout the cell cycle at the same rate as culture mass.


Polyamines of Anacystis nidulans and metabolism of exogenous spermidine and spermine.

Several biochemical parameters, including that of polyamine content, accompanying the growth of the cyanobacterium Anacystis nidulans were studied. At all stages of growth under autotrophic conditions, the organisms were found to be rich in spermidine and lacking in spermine, as is typical of procaryotic organisms. The cells were quite low in putrescine, and no unusual polyamine was observed to be present as a major component. Conjugated polyamines were not detected in the cultures. At maximal culture density, the levels of spermidine, DNA, RNA, protein, and chlorophyll were also maximal. Shortly after the inception of the stationary phase, the spermidine content of the cells was the first parameter observed to decrease in cultures which were shortly to become yellow. Spermidine lost from the cells was not recovered in the medium in a free or conjugated form. This indication of degradation of spermidine was studied by the addition of polyamines to growing cultures. Exogenous spermidine and spermine were found to be metabolized rapidly by the organisms, of which diaminopropane was one product. Putrescine was found to be markedly toxic, whereas spermidine, some other triamines, and spermine were much less toxic.


Phospholipid composition of methane-utilizing bacteria.

The phospholipids of Methylococcus capsulatus, Methylosinus trichosporium, La Paz, and OBT were examined in relation to their qualitative and quantitative composition. M. capsulatus exhibited a phospholipid composition consisting of phosphatidylethanolamine, phosphatidylglycerol, cardiolipin, and phosphatidyl-choline. The esterified fatty acids were predominantly C16:0 and C16:1. M. trichosporium, La Paz, and OBT exhibited an essentially identical phospholipid composition consisting of phosphatidylmonomethylethanolamine, phosphatidyl-dimethylethanolamine, phosphatidylcholine, and phosphatidylglycerol. Only trace amounts (less than 1%) of cardiolipin were found in these organisms. The major esterified fatty acid in these organisms was C18:1 (87 to 90%). The monounsaturated fatty acids from all four organisms consisted of both cis and trans isomers, each of which contained delta8, delta9, delta10, and delta11 double-bond positional isomers.


Effects of low temperature on in vivo and in vitro protein synthesis in Escherichia coli and Pseudomonas fluorescens.

The effects of temperature on protein synthesis by Escherichia coli, a mesophile, and Pseudomonas fluorescens, a psychotroph, were investigated by using whole-cell and cell extract preparations. After shifts to 5 degrees C, protein was synthesized at a slowly decreasing rate for 1 h by both organisms, after which P. fluorescens synthesized protein at a new rate corresponding to its 5 degrees growth rate, in contrast to E. coli which did not synthesize protein at a measurable rate. In vitro protein-synthesizing systems using MS-2 RNA, endogenous mRNA, and purified polysomes were utilized to investigate initiation of translation at 5 degrees C. In these systems, P. fluorescens cell extracts synthesized protein at linear rates for up to 2 h at 5 degrees C, whereas E. coli cell extracts synthesized protein for only 25 min at 5 degrees C. The rates of polypeptide elongation, as tested by the incorporation of phenylalanine into polyphenylalanine by cell extract protein-synthesizing systems from both organisms, were identical over the range of 25 to 0 degrees C. The polysome profiles of E. coli whole cells shifted from 37 to 5 degrees C showed accumulation of 70S ribosomal particles and ribosomal subunits at the expense of polysomes. Similar experiements done with P. fluorescens resulted in polysome reformation at 5 degrees C. In vitro experiments demonstrated that the 70S ribosomal particles, which accumulated in E. coli at 5 degrees C, were capable of synthesizing protein in vitro in the absence of added mRNA. These in vivo and in vitro results suggest that incubation of E. coli at subminimal temperatures results in a block in initiation of translation causing polysomal runoff and the accumulation of 70S particles, some of which are 70S monosomes.


Characterization of Pseudomonas aeruginosa mutants deficient in the establishment of lysogeny.

Mutants of Pseudomonas aeruginosa with impaired ability to establish a lysogenic relationship with temperate bacteriophage (Les-) have been isolated. These les mutations map to two areas of the P. aeruginosa chromosomal map as determined by conjugational and transductional analyses. Two phenotypic classes of Les- mutants were identified. One class of mutations has pleiotropic effects on DNA metabolism. These mutants are unable to recombine genetic material acquired as a result of either conjugation or transduction (Rec-). In addition, the ability of these Les- Rec- mutants to repair UV-induced damage to bacteriophage is reduced (host-cell reactivation deficient, Hcr-). Mutants of the second class are Les-, Rec+, and Hcr+.


Energy metabolism of Rickettsia typhi: pools of adenine nucleotides and energy charge in the presence and absence of glutamate.

The obligate intracellular bacterium Rickettsia typhi was examined for its ability to generate and maintain an adenylate energy charge in an extracellular environment. Freshly purified organisms were incubated, at 34 degrees C and pH 7.4, with or without glutamate and various other metabolites, and the levels of ATP, ADP, and AMP were determined. Of the metabolites tested, glutamate and glutamine were the most effective for the generation of ATP. In the presence of glutamate, there was a rapid increase in the level of ATP, followed by a moderate decrease during 150 min of incubation. The energy charge increased from a level of 0.2 to 0.5 to about 0.7 to 0.75, and then slowly declined to about 0.45 to 0.6. In the absence of glutamate, after an occasional initial surge in ATP level as the temperature was changed from 4 to 34 degrees C, there was a sharp decline in both ATP and energy charge (to 0.1 and sometimes to 0.01). The rickettsiae maintained their ability to regenerate their energy charge upon the addition of glutamate for about 30 min, but this ability declined with further incubation. In contrast to Escherichia coli, the decline in ATP in R. typhi was accompanied by a sharp increase in the level of AMP and the total adenylate pool. No adenine or adenosine was recovered from rickettsiae incubated with labeled AMP, ADP, or ATP. From these experiments and the demonstration reported elsewhere that rickettsiae transport the adenine nucleotides, it can be concluded that the adenylate energy charge in R. typhi is governed by the salvage of the adenine nucleotides rather than their unphosphorylated precursors. Thus, R. typhi undergoes greater shifts in energy charge than other bacteria, a phenomenon which may account for their instability in an extracellular environment. Under optimal conditions the adenylate energy charge of R. typhi approaches levels that border on those generally regarded as adequate for growth.


Bacillus subtilis 168 genetic transformation mediated by outgrowing spores: necessity for cell contact.

Transforming activity released in sequential genetic order during the first synchronous cycle of DNA replication during outgrowth of spores of Bacillus subtilis 168 was investigated. A transformation assay was used consisting of outgrowing spores as DNA donors and multiply marked competent cells as recipients. DNA synthesis inhibitors known to stop DNA release were used during and subsequent to DNA transfer to recipient cells. The released DNA sedimented with the outgrowing cells after low-speed centrifugation, and it was discovered that markers released both early and late were resistant to up to 500 microgram of deoxyribonuclease per ml under conditions in which the transforming capacity of purified DNA was eliminated by 5 microgram of the nuclease per ml. Inaccessibility to deoxyribonuclease was increased and maintained during the transformation event while detergents and proteolytic attack did not expose the released chromosome to nuclease action. The results indicate that tight physical contact between outgrowing spores and competent cells is required for transformation in this system.


Biochemical studies of rat liver Golgi apparatus. I. Isolation and preliminary characterization.

A method is described for the isolation of morphologically well-preserved Golgi apparatus from rat liver. The method is essentially the same as that of Morré et al. (Morré, D.J., Hamilton, R.L., Mollenhauser, H.H., Mahley, R.W., Cunningham, W.P., Cheetham, R.D., & Lequire, V.S. (1970) J. Cell Biol. 44, 484-491) except that mild cell disruption is achieved by means of a stainless-steel sieve. The average recoveries of protein and galactosyltransferase in the isolated fraction are about 6 mg from 10 g of perfused liver and about 35% from the homogenate, respectively. The preparation is virtually free from succinate-cytochrome c reductase, glucose-6-phosphatase, acid phosphatase, and 5'-nucleotidase. The Golgi fraction as well as its vesicular fragments is homogeneous upon isopycnic centrifugation in both sucrose and dextran density gradients. Their buoyant densities in sucrose are significantly higher than those in dextran, indicating that both forms of the organelle are closed systems which are impermeable to macromolecules. The galactosyltransferase activity of a freshly prepared Golgi fraction, measured with ovalbumin as galactosyl acceptor, is activated 26-fold by the addition of Triton X-100, whereas those of homogenized, sonicated, and aged preparations are only activated 2- to 4-fold.


Biochemical studies on rat liver Golgi apparatus. III. Subfractionation of fragmented Golgi apparatus by counter-current distribution.

Vesicular fragments of Golgi apparatus, smooth- and rough-surfaced microsomes from rat liver are differently partitioned in aqueous polymer two-phase systems consisting of dextran, polyethylene glycol, and sodium phosphate buffer. At a given polymer concentration, the amount of material partitioned in the top phase increases in the following order: rough microsomes less than smooth microsomes less than Golgi fragments. Counter-current distribution of Golgi fragments in the system consisting of 6.8% (w/w) dextran T500 and 6.8% polyethylene glycol 4,000 results in the separation of the fragments into three fractions; i.e. Fractions I, II, and III. NADH- and NADPH-cytochrome c reductase activities are detected almost exclusively in Fraction I, whereas the activities of galactosyltransferase, acid phosphatase, 5'-nucleotidase, and thiamine pyrophosphatase are maximal in Fraction III and minimal in Fraction I. The distribution of these enzymes suggests that Fraction I is similar to, though not identical with, microsomes, Fraction III resembles plasma membrane and lysosomes, and Fraction II is between the two. It is concluded that NADH- and NADPH-cytochrome c reductases are localized in a restricted region of the Golgi structure and that intra-Golgi differentiation seems to proceed in a discontinuous manner.


Studies on regulatory functions of malic enzymes. V. Comparative studies of malic enzymes in bacteria.

Screening of four malic enzymes--NAD-linked enzyme [EC 1.1.1.38], NAD, NADP-linked enzyme [EC 1.1.1.39], NADP-linked enzyme [EC 1.1.1.40], and D-malic enzyme--was carried out with cell-free extracts of the following 16 strains of bacteria by the aid of Sepharose 6B column chromatography: 9 strains of enteric bacteria, 3 strains of Pseudomonas, Alcaligenes faecalis, Agrobacterium tumefaciens, Rhodospirillum rubrum, and Clostridium tetanomorphum. All the strains tested contained at least one malic enzyme. The NADP-linked enzyme activity was found in all the strains except C. tetanomorphum, the NAD-linked enzyme activity in 12 strains--8 strains of enteric bacteria, 2 strains of Pseudomonas, Ag. tumefaciens, and C. tetanomorphum--and D-malic enzyme activity in 4 strains--A, aerogenes (IFO 3319 and 12059), Ps. fluorescens, and R. rubrum. The NADP-linked and NAD-linked enzyme activities of two strains of Pseudomonas were not separated by the chromatography. The available evidence suggested that the NAD, NADP-linked enzyme was not present in these 16 strains. The comparative studies of molecular, enzymatic, and serological properties of the malic enzymes in these 16 strains revealed a close similarity of the same types of malic enzymes among enteric bacteria.


Tryptophan residues of saccharifying alpha-amylase from Bacillus subtilis. A kinetic discrimination of states of tryptophan residues using N-bromosuccinimide.

Four tryptophan residues of saccharifying alpha-amylase from B. subtilis out of eleven in total are reactive towards N-bromosuccinimide (NBS), suggesting that they are on the surface of the enzyme. This is consistent with the results of solvent perturbation difference spectrophotometry with ethylene glycol. One of four tryptophan residues was clearly distinguished from the other three in reactivity with NBS by the stopped-flow method. This most reactive tryptophan residue was not protected from modification by substrates of analogs, indicating that the tryptophan is not located in the substrate binding site. One of the other three tryptophan residues, probably the second most reactive one, is considered to be related in some way to the glycosyl transfer in the reaction of the enzyme with maltose as a substrate.


Characterization and genetic mapping of modA. A mutation in the post-translational modification of the glycosidases of Dictyostelium discoideum.

We have isolated a mutant of Dictyostelium discoideum, M31, which produces a reduced number of alpha-mannosidase-1 molecules per cell during the developmental program of the organism. We find that several of the glycosidases, a group of lysosomal proteins produced by D. discoideum, are altered in strain M31 and that this strain produces a reduced level of at least three of these activities. These enzymes do not share a common protein subunit but are known to share a common antigenic determinant which is, in part, carbohydrate in nature. In the wild type parent of M31, alpha-mannosidase-1 is modified by the addition of mannose and glucosamine (probably as N-acetylglucosamine) in the molar ratio of 5:2. alpha-Mannosidase-1 was also found to contain phosphoserine/phosphothreonine residues. alpha-Mannosidase-1 and other glycosidases are electrophoretically less negative when isolated from strain M31 than when isolated from wild type cells. The mutation present in M31, modA, appears to affect posttranslational modification, modA is a recessive mutation which we map onto linkage group I.


Effects of a post-translational modification mutation on different developmentally regulated glycosidases in Dictyostelium discoideum.

We have studied the effect of a post-translational modification mutation upon four developmentally regulated glycosidases of Dictyostelium discoideum. The presence of the modA mutation affects the intracellular level of these multimeric enzymes differently. The level of alpha-glucosidase is unaffected in the modA mutant. The mutant cell contains only a very small fraction of the wild type beta-glucosidase-1 activity. The alteration in modification renders beta-glucosidase-1 holoenzyme thermolabile and susceptible to degradation in vivo. alpha-Mannosidase-1 and N-acetylglucosaminidase are found at approximately 1/3 of the wild type level in the modA mutant. Degradation of holoenzyme does not appear to be responsible for the low level of these activities. We propose that alpha-mannosidase-1 and N-acetylglucosaminidase subunits are being degraded prior to subunit assembly. We conclude the modification bestows different properties upon the various glycosidases.


Biochemical and immunological characterization of human opsonic alpha2SB glycoprotein: its identity with cold-insoluble globulin.

The relationship between human cold-insoluble globulin (CIg, plasma fibronectin) and the human serum opsonic alpha2SB glycoprotein was investigated using immunochemical and biochemical techniques. The two proteins appeared to have identical molecular weights by sodium dodecyl sulfate-polyacrylamide gel electrophoresis on 3.3% gels; have identical migration in the native state on 2.7 to 27% gradient polyacrylamide gels; and have a similar amino acid composition within the accuracy of analysis. Human serum demonstrates antigenic identity when diffused against monospecific antisera to both proteins confirming the presence of common antigenic sites on both molecules. Purified human serum opsonic alpha2SB glycoprotein and purified CIg also demonstrate antigenic identity when diffused against monospecific antiserum to either of the isolated proteins. Antiserum to both proteins also inhibits in vitro hepatic Kupffer cell phagocytic uptake of test particles. These results suggest the idenity of these two proteins and reveal a major physiological function for human plasma CIg. Thus, CIg may be important in the regulation of hepatic reticuloendothelial phagocytic activity and nonspecific systemic host defense. This process of systemic host defense has been shown to be depressed in patients following trauma, major surgery, burn injury, and during neoplastic disease, and, in part, mediated by a deficiency or depletion of the alpha2SB glycoprotein.


Liver microsomal cytochromes P-450 and azoreductase activity.

Hepatic microsomal azoreductase activity with amaranth (3-hydroxy-4[(4-sulfo-1-naphthalenyl)azo]-2,7-naphthalenedisulfonic acid trisodium salt) as a substrate is proportional to the levels of microsomal cytochrome P-450 from control or phenobarbital-pretreated rats and mice or cytochrome P-448 from 3-methylchol-anthrene-pretreated animals. In the "inducible" C57B/6J strain of mice, 3-methylcholanthrene and phenobarbital pretreatment cause an increase in cytochrome P-448 and P-450 levels, respectively, which is directly proportional to the increase of azoreductase activity. However, in the "noninducible" DBA/2J strain of mice, only phenobarbital treatment causes the increase both in cytochrome P-450 levels and azoreductase activity, while 3-methylcholanthrene has no effect. These experiments suggest that the P-450 type cytochromes are responsible for azoreductase activity in liver microsomes.


Proximal focal femoral deficiency.

The term proximal focal femoral deficiency (PFFD) is applied to a spectrum of conditions characterized by partial absence and shortening of the proximal femora and thought to result from an early disturbance of growing mesenchyme. The mildest cases, Classes A and B, exhibit a relatively normal acetabulum and capital femoral epiphysis, despite the dysplastic shaft. There may be a subtrochanteric varus deformity or a pseudoarthrosis accompanying the shortening of the femoral shaft. At the opposite end of the spectrum, the most severely deformed cases, Classes C and D, have involvement of the acetabulum, and the entire proximal femur. In cases of advanced dysgenesis, the findings include: a stunted severely shortened femur with a club-shaped or pointed proximal end, an iliac projection just above the anatomic site of the acetabulum, a spherical obturator foramen, and squaring of the iliac crest. Twenty-one patients (2 bilateral) with proximal focal femoral deficiency are presented (23 femurs). Differentiation of proximal focal femoral deficiency from other congenital and acquired deformities of the hip joint is essential for selecting the proper treatment.


Primary intracranial mixed choriocarcinoma and malignant teratoma.

Primary intracranila choriocarcinoma, either alone or with malignant teratoma, is a rare tumor. The 22 reported cases of primary intracranila mixed choriocarcinoma and malignant teratoma are reviewed, and a further case is added. This 4 1/2-year-old girl presented with multiple cranial nerve palsies, panhypopituitarism and markedly elevated blood and urine human chorionic gonadotrophin (HCG) titres. After subtotal removal of the suprasellar tumor, supervoltage radiation was given with a remarkably rapid and complete response. She died 14 months after diagnosis with no evidence of local recurrence nor of distant metastases.


Erythrocyte entry by malarial parasites. A moving junction between erythrocyte and parasite.

Invasion of erythrocytes by merozoites of the monkey malaria, Plasmodium knowlesi, was investigated by electron microscopy. The apical end of the merozoite makes initial contact with the erythrocyte, creating a small depression in the erythrocyte membrane. The area of the erythrocyte membrane to which the merozoite is attached becomes thickened and forms a junction with the plasma membrane of the merozoite. As the merozoite enters the invagination in the erythrocyte surface, the junction, which is in the form of a circumferential zone of attachment between the erythrocyte and merozoite, moves along the confronted membranes to maintain its position at the orifice of the invagination. When entry is completed, the orifice closes behind the parasite in the fashion of an iris diaphragm, and the junction becomes a part of the parasitophorous vacuole. The movement of the junction during invasion is an important component of the mechanism by which the merozoite enters the erythrocyte. The extracellular merozoite is covered with a prominent surface coat. During invasion, this coat appears to be absent from the portion of the merozoite within the erythrocyte invagination, but the density of the surface coat outside the invagination (beyond the junction) is unaltered.


Characterization of bacterial L-(-)-tyrosine decarboxylase by isoelectric focusing and gel chromatography.

The purification of L-(-)-tyrosine apodecarboxylase (TAD) (E.C. 4.1.1.25), obtained from extracts of cells of Streptococcus faecalis, has been investigated by means of preparative isoelectric focusing, molecular sieve chromatography and hydrophobic interaction chromatography. Isoelectric focusing demonstrated two separate fractions possessing enzyme activity that had pI values of 4.5 and ca. 3.2. In the chromatographic methods, however, the activity was obtained in a single peak. It was found that hydrophobic interaction chromatography on phenyl-Sepharose was particularly suitable for purification purposes. The enzyme is very firmly bound to octyl-Sepharose CL-4B but retains most of its activity even in the bound state.


Serogroup identification of Neisseria  meningitidis: comparison of an antiserum agar method with bacterial slide agglutination.

A serum agar method for serogrouping Neisseria meningitidis is described and compared with conventional bacterial slide agglutination. There was 93% agreement for 300 strains examined individually by each method. Among strains from serogroups A, B, C, Y, and W135, there was 100% correlation, whereas strains from serogroup 29E (Z') had only 67% correlation. The serum agar method was rapid, as well as easy to perform and interpret. The potential benefits of this method for epidemiological studies and reference laboratories processing large numbers of meningococcal isolates are emphasized.


Identification of some basic extractable compounds produced by Neisseria gonorrhoeae and Neisseria meningitidis in a defined medium.

Electron capture gas-liquid chromatography and mass spectrometry have been used to identify some of the basic extractable heptafluorobutyric anhydride reactive compounds found in a defined medium after 20 h of growth by Neisseria gonorrhoeae and N. meningitidis. Acetoin, 2,3-butanediol, pyrroline, and 1,3-diaminopropane were identified by both gas chromatography and mass spectrometry; 2-hydroxy-pyrrolidine and 3-aminomethyl-pyrrolidine were tentatively identified by mass spectrometry. A possible origin of the amines is through enzymatic oxidation of polyamines.


Comparison of a radiometric procedure with conventional methods for identification of Neisseria.

A radiometric procedure was compared with the conventional cystine tryptic agar (CTA) sugar fermentation method for identification of Neisseria species. Four different ATCC cultures of Neisseria were identified by both procedures with identical results. The only difference noted was that the radiometric procedure required 3 h for completion, whereas the conventional CTA sugar method required overnight incubation. The radiometric procedure was also compared with the fluorescent antibody (FA) and CTA methods for identification of Neisseria gonorrhoeae. The organisms examined were gram-negative, oxidase-positive diplococci isolated from 49 clinical specimens sent to the laboratory for bacteriological analysis. Results obtained by both CTA and FA procedures were comparable. However, the radioisotope method appeared to be superior to the other two methods in that only one isolate identified as positive by both the CTA and FA methods was not identified radiometrically, whereas four isolates positive by the radiometric method were not identified by the other two procedures. Thus, a total of seven more positive identifications were made radiometrically than by either of the two other methods. All positive identifications were confirmed by a reference laboratory. These results indicate that the radiometric procedure is more rapid and reliable as compared with both the CTA and direct FA methods and, thus, may serve as a valuable addition to the methodology available for diagnostic microbiology.


Value of acid metabolic products in identification of certain corynebacteria.

Acid metabolic products of 23 strains of human and animal pathogenic corynebacteria, representing eight different species, were determined by gas chromatography. The results showed that the species examined were metabolically heterogeneous and could be presumptively identified based on the acid products produced. Corynebacterium equi did not produce any acids; C. renale produced lactate; and C. pyogenes produced major amounts of lactate, variable amounts of acetate, and minor amounts of succinate and pyruvate. C. kutscheri produced propionate and lactate as major products and pyruvate and oxalacetate as minor products. C. diphtheriae and C. pseudotuberculosis produced major amounts of propionate, acetate, and formate. In addition, C. pseudotuberculosis produced major amounts of pyruvate and minor amounts of succinate, lactate, and oxalacetate, whereas C. diphtheriae strains produced minor but variable amounts of lactate, succinate, fumarate, pyruvate, and oxalacetate. C. bovis produced aicd products similar to those of C. pyogenes but was readily distinguishable from the latter by the lack of hemolysis on blood agar, colony morphology, catalase reaction, and biochemicals. C. suis characteristically produced major amounts of ethanol, acetate, and formate and minor amounts of lactate and succinate but no propionate.


Modification of methods used in bacteriophage typing of Mycobacterium tuberculosis isolates.

A procedure in which soft agar overlays were used in bacteriophage-typing Mycobacterium tuberculosis is presented. This safer method uses commercially available media, whereas media presently used must be prepared in the laboratory. Single plaque isolations of the phage BG-1 specifying phage type A and B of M. tuberculosis were readily made by using the modified procedures. This purification and the use of prototype strain Myc 1415 as the indicator host strain have significantly enhanced the ability to discriminate among strains of phage types A and B.


Identification and grouping of Neisseria meningitidis directly on agar plates by coagglutination with specific antibody-coated protein A-containing staphylococci.

It has been shown that Neisseria meningitidis can be grouped by coagglutination directly upon growth on sheep blood or chocolate agar plates. All positive reactions were group specific, and only a single colony was required for a positive reaction. There was variation seen in the effectiveness of commercial antisera in preparing sensitive reagents. Certain throat and sputum isolates of group Z organisms failed to react by coagglutination, although they would react directly with antiserum in whole-cell agglutination. This problem remains unresolved but may not be greatly significant in laboratory use of this method since group Z is rarely associated with disease.


Rapid determination of Corynebacterium diphtheriae toxigenicity by counterimmunoelectrophoresis.

A method for detecting toxin production in Corynebacterium diphtheriae cultures by counterimmunoelectrophoresis is described. Precipitin lines were observed in toxigenic strains in 30 min.


Method for determining bioburden of surgical gloves.

A washing procedure that removed maximum numbers of contaminating microorganisms from whole surgical gloves was developed. Washing, coupled with membrane filtration, proved to be a simple and effective method for bioburden determinations on whole gloves.


Partial purine nucleoside phosphorylase deficiency. Studies of lymphocyte function.

Immune function in two brothers with a deficiency of purine nucleoside phosphorylase was evaluated in vivo and in vitro. Both patients had a history of recurrent infections and profound lymphopenia. Studies of cell-mediated immunity revealed an absence of delayed cutaneous reactivity to a number of antigens, including dinitrochlorobenzene, and significantly reduced lymphocyte proliferative responses to nonspecific mitogens, specific antigen, and allogeneic cells. E-rosetting cells were present but reduced in number (20.0% and 31.5%). Serum immunoglobulin levels, percentages of circulating immunoglobulin-and C3-receptor-bearing B cells, as well as the ability to produce antibody in response to specific antigen in vivo were normal. Moreover, studies of the in vitro induction of specific IgM antibody delineated the presence of T-helper and T-regulator cells. The normal induction of bone marrow precursor T-cell maturation by human thymic epithelium-conditioned medium or thymosin suggested that the initial stages of T-cell generation were intact in these patients. Attempts to reconstitute the in vitro proliferative response with a variety of reagents, including purine nucleoside phosphorylase itself, were unsuccessful. Selective impairment of certain aspects of T-cell function in these patients and a less severe clinical picture than previously described may be explained by the presence of a partial deficiency of nucleoside phosphorylase activity and incomplete block of purine catabolism.


Immunogenicity of polysaccharides from type III, group B Streptococcus.

The immunogenicity and safety of two polysaccharides isolated from type III, group B Streptococcus, were tested in adults selected for existing low concentrations of natural antibody to the capsular polysaccharide of this organism. Both vaccine preparations (trichloroacetic acid and EDTA) were found to lack pyrogenicity and toxicity for experimental animals. A single 50-microgram subcutaneous injection of either polysaccharide in human subjects elicited significant increase in antibody concentration in immunized compared with control individuals receiving phosphate-buffered saline. Antibody responses were maximal by 2 wk and remained at 21 wk after immunization. Vaccine-induced antibody was primarily of the IgG class. Of the two vaccines, the larger molecular size polysaccharide was significantly more immunogenic. Although no systemic reactions were recorded, mild transient local reactions occurred in 45% of vaccinees.


Glucocorticoids administered in vivo inhibit human suppressor T lymphocyte function and diminish B lymphocyte responsiveness in in vitro immunoglobulin synthesis.

The effects of corticosteroid given in vivo on human lymphocyte subpopulation function were investigated using an in vitro system of pokeweek mitogen-stimulated immunoglobulin production. Peripheral blood lymphocytes were obtained from normal volunteers before and 4 h after the intravenous administration of methylprednisolone. Unfractioned peripheral blood lymphocytes showed a consistent decrease (mean congruent with 50%) in immunoglobulin and total protein synthesis after steroid administration. Utilizing separated thymus-derived (T) and bone marrow-derived (B) lymphocyte fractions, the pathophysiology of this alteration in immunoglobulin production was elucidated. B lymphocytes obtained after steroid treatment showed a markedly diminished immunoglobulin response (20% of normal) to normal T lymphocytes and to normal T cells that had been irradiated to remove suppressor T lymphocyte function. All major classes of immunoglobulin (IgG, IgM, and IgA) were affected. T lymphocytes procured after steroid administration were capable of providing normal amounts of T cell help for B cells in immunoglobulin production. However, suppressor T lymphocyte activity, observed with normal T lymphocytes at high T to B cell ratios, was absent from the post-steroid T lymphocytes. This loss of suppressor T lymphocyte function was not due to the presence of excess help as irradiated pre- and poststeroid T cells provided equal amounts of helper activity. On recombining the poststeroid treatment B cells, which are hyporesponsive in immunoglobulin synthesis, with the posttreatment T lymphocytes, which lack suppressor activity, diminished amounts of immunoglobulin were produced which correlate well with the effects observed with unseparated cells. Thus, corticosteroids have differential effects on the lymphocyte populations involved in immunoglobulin biosynthesis. B cell responsiveness is diminished, suppressor T lymphocyte activity is removed, and helper T lymphocyte function is unaffected.