Developmental genetics of the Drosophila egg. I. Identification of 59 sex-linked cistrons with maternal effects on embryonic development. Sex-linked mutations to recessive female sterility were induced, sorted for egg-laying, mapped within broad regions and grouped by complementation tests into cistrons. The mutations have also been partially characterized for their temperature sensitivity and pleiotropic effects. Altogether 59 cistrons have been identified, including five ellelic with previously known loci: cin, fs(1)N, mk, sn, and r. All of the genes make maternal contributions to developing embryos. In some instances mutant defects are recognized in the egg envelopes; in the remainder the defects are presumably in the egg cytoplasm. For mutations in twenty-two genes, including cin, mk, and r alleles, the lethality of the maternal effect is reversed and the embryo is "rescued" by the action of a wild-type, paternal allele. The mutant strains are potentially important material for the study of developing egg envelopes and for furthering the analysis of causation in embryogenesis and its origins in oogenesis.


Enzyme and chromosome polymorphisms in Japanese natural populations of Drosophila melanogaster. Collections of D. melanogaster from Japanese populations were analyzed for enzyme and chromosomal polymorphisms. Allelic frequencies at the Adh and alphaGpd loci were compared with polymorphic inversion (In(2L)B, In(2R)C) frequencies in the second chromosome. There was a significant positive correlation between the frequencies of AdhS and In(2L)B, caused by linkage. On the other hand, inversion-free cage populations maintained in the laboratory for a long time showed considerably larger variation in the frequencies of these enzyme alleles, which seem very likely to be a consequence of random drift. Two fitness components of these enzyme and chromosomal variants were measured in two different environmental conditions; neither of the two loci showed heterozygote superiority in viability or productivity, while the inversion heterozygotes showed a superior productivity compared to the corresponding homozygotes in the fluctuating environment. These findings are compatible with the hypothesis that polymorphic isozyme genes are maintained by random drift of neutral genes in natural populations, and that association with linked inversions is a historical accident.


Lack of dosage compensation for an autosomal gene relocated to the X chromosome in Drosophila melanogaster. Aldehyde oxidase activity has been measured in flies with the structural gene for this enzyme translocated to the X chromosome. These measurements are presented as experimental evidence that, in Drosophila melanogaster, an autosomal gene relocated to the X chromosome is not dosage compensated.


The cytogenetics of a recessive visible mutant associated with a deficiency adjacent to the notch locus in Drosophila melanogaster. The recessive visible faswb allele in Drosophila is an interband deletion between salivary band 3C5, 6 and 7. Heterozygosity for the deletion does not suppress recombination between faswb and mutant sites at Notch adjacent to it.--Df(1)w67k30, deficient for salivary bands 3C2 to 6, is the left of faswb. By crossing over within the homologous bit of interband retained in w67k30 and faswb, the two deficiencies can be linked. Cytologically, 3C7, "fused" to 3C5,6 in faswb, becomes "fused" to 3C1 when the two are coupled. In the double deletion, the recessive visible phenotype of the faswb "allele* is suppressed. Both w67k30 and faswb can be recovered by uncoupling the two deficiencies.--The data suggest that the mutant faswb does not represent a lesion at Notch; the entire gene or locus seems to be present. The interband deletion in faswb has secondarily moved an intact Notch locus to a foreign environment that interferes with its normal function. When faswb is linked to w67k30, the interference is eliminated and normal Notch functions resume.--The position of Notch on the salivary gland chromosome is reviewed in relation to the information obtained in these experiments.


A method for fate mapping the foci of lethal and behavioral mutants in Drosophila melanogaster. A new method of mosaic fate mapping, called focusing, is introduced. Its advantages are that it allows a mapping, on the blastoderm surface, of the site of action of functions defined by either pre-adult lethal or behavioral mutations. Moreover, it does not require that the mosaics used be 50% of one genotype and 50% of the other. Methods for quantitative evaluation of the results of focusing, and a comparison of this method with others, are discussed.--Focusing is applied to the analysis of a new mutant, doomed (symbol: dmd), a distally located X-linked recessive in D. melanogaster, that causes the deaths of males and females around the time of eclosion. The dmd phenotype among eclosing flies is first seen as the loss of thoracic motor coordination and as ether sensitivity. Fate mapping by the method of focusing places the site of action of the dmd+ function in the same region of the map as that of the thoracic neural ganglia primordia, but not that of muscle, suggesting the possibility that the effect of dmd is on a thoracic neural function rather than a muscular one.


Correlations between chromosome segments and fitness in Drosophila melanogaster. I. The X chromosome and egg production. Unmarked segments within the X chromosomes of four different Drosophila melanogaster isogenic lines were assessed with respect to egg production. By making a series of crosses among original and derived recombinant lines, it was possible to estimate parameters representing additive, dominance and interaction effects of the segments. It was shown that whereas most of the segments were additive for egg production when homozygous, they all displayed dominance in the heterozygous condition. Two of the strains were characterized by intersegmental interaction. A possible position effect was detected for these same two strains, with flies in the coupling phase laying more eggs than those in the repulsion configuration. There was no apparent relationship between the number of eggs laid and the amount of heterozygosity within the X chromosome.


Antibodies to pancreatic duct cells in Sjögren's syndrome and rheumatoid arthritis. In the present investigation the occurrence of humoral immunity to pancreatic duct cells (PDC) was studied in 12 patients with Sjögren's syndrome (SS), 31 patients with rheumatoid arthritis (RA), and 64 controls. Four sera of patients with SS and eight of patients with RA produced diffuse cytoplasmic fluorescence of intro- as well as of interlobular PDC of human and rhesus monkey origin. All sera positive with PDC antigens gave also positive staining reaction with parotid, submandibular, and lacrimal duct cells. In absorption studies antibody activity to PDC and salivary duct cells could be absorbed equally well with human or monkey parotid gland or pancreas with almost identical antigen concentrations. These findings point to the presence of common antigenic determinants in the organs studied. Human thyroid microsomes and rat liver homogenate did not reduce antibody activity. The demonstration of antibodies to PDC in addition to the reported mononuclear cell infiltration of the pancreas point to the involvement of autoimmune mechanisms in pathogenesis of the commonly observed subclinical exocrine insufficiency in SS and in some cases of RA.


Genus Aotus Q- and G-band karyotypes and and natural hybrids. Q- and G-banding studies of Aotus trivigatus trivirgatus and Aotus trivirgatus griseimembra are presented. After the analysis of 8 specimens, 2 A.t. trivirgatus and  6 A.t. griseimembra and the review of the literature on the field we state the existence of two different karyotypes, 2n = 54 and 2n = 50, for A.t. trivirgatus and three different karyotypes, 2n = 54, 2n = 52 and 2n = 50, for A.t. griseimembra. It was found that Robertsonian translocations were the mechanism involved in the numerical reduction of the chromosomes. The data presented include the statement of two natural hybrids and the cross-breedings which originated them.


Sequence of eruption of permanent teeth and epiphyseal union in three species of African monkeys. Sequences of eruption of teeth and union of epiphyses determined for three species of African Cercopithecinae. These sequences are highly scalable and are highly comparable between among the species. Some sex differences in sequence were found within each species. The sequences of epiphyseal union in these species of Old World monkeys are very similar to published sequences in New World monkeys and other groups of higher primates. It appears that there is a single pattern common to the Anthropoidea as a whole.


New data and a discussion of infant killing in old world monkeys and apes. Information on infant killing by males in reviewed and extended by personal communications and observations. Most cases of infant killing reported occurred during periods when a new alpha male was establishing himself and during arranged encounters of strangers or unfamiliar individuals in captivity. There seem to be no specific releasing stimuli for infant killing, but rather the lack of familiarity between male and females, their nonacceptance of him in the whole set of roles tied to the alpha status, and his special motivational condition as new alpha lead him to a state, in which attacks on infants may result. Infant killing is considered to have a selection advantage by increasing one's own reproduction success while reducing the one of other males.


The number of umbilical vessels in the squirrel monkey (Saimiri sciureus). The number of vessels present in the umbilical cords of 40 nonsurviving squirrel monkey fetuses an neonates has been determined by gross and histologic examination. Four umbilical vessels were present in 72.5% of the cords and 27.5% contained three vessels. Coincident fetal malformations are noted.


[Therapy of chronic arterial occlusive disease. Studies with an orally administered drug under ambulatory conditions]. The therapeutical effect of Defluina was studied in a selective open trial of 50 patients suffering from peripheral arterial occlusive diseases of different stages. All patients were treated with Defluina, 30 drops three times a day during a period of six weeks. Control parameters of 46 patients (18 males, 26 females, mean age of 59.8 +/- 1.7 years) were evaluated. After therapy a significant improvement of all applied parameters was observed. The positive effect of the Defluina-medication on the initial--partly severe--varicose ulcera, with gangrenous alterations, has to be pointed out. After 6 weeks the varicose ulcera were partly closed, at least they showed fresh granulations. During therapy no adverse reactions were registered. All patients estimated Defluina as very well tolerable. The therapeutic effect of Defluina on the carefully investigated group of 46 patients could be estimated in 38 cases with excellent/good, in 7 cases with good/moderate and only in 1 case with moderate.


TBG-dependency of age related variations of thyroxine and triiodothyronine. Thyroxine (T4), triiodothyronine (T3) and thyroxine-binding globulin (TBG) were determined in healthy individuals ranging in age from newborn to 95 years. T4: 10.25 +/- 1.62 microng/100 ml, T3: 1.62 +/- 0.35 ng/ml and TBG: 1.34 +/- 0.15 mg/100 ml, were found elevated until puberty compared to a middle age group with T4: 7.27 +/- 2.26 microng/100 ml, T3: 1.15 +/- 0.24 ng/ml and TBG: 0.98 +/- 14 mg/100 ml. T4 and T3 followed almost TBG concentration. In old age is dissociation between T4: 5.79 +/- 1.56 microng/100 ml, T3: 0.79 +/- 0.21 ng/ml and TBG: 1.28 +/- 0.15 mg/100 ml was found. Except for old age the ratio T4/TBG and T3/TBG minimized the age dependent variation of T4 and T3 and reduced the coefficient of variance from 26% to 17.7% for T4 and from 26.5 to 25% for T3. Age reduction of T4/TBG is 15% and of T3/TBG 13% respectively more pronounced than for T4 and T3 alone. These data indicate: 1) age related variations of T4 and T3 due to age dependency of TBG, 2) deviation of T4 and T3 values in old age from that expected by their TBG levels and 3) the importance of the routine use of hormone/TBG ratio.


Effects of ether and pentobarbital anesthesia on thyroid function in the rat. Studies were performed to examine the effect of two anesthetic agents, ether and pentobarbital, on the hypothalamic-pituitary-thyroid function in vivo. In non-anesthetized animals, plasma thyrotropin (TSH) increased rapidly from basal values of 0.1, a peak of 0.49 microng/ml, 25 min after exposure to the cold. Anesthesia with ether during exposure to the cold completely prevented the rise in TSH. During pentobarbital anesthesia, the rise in TSH after exposure to cold was reduced by more than 90%. Even a three minute period of ether anesthesia prior to cold exposure reduced the peak response to cold as well as delayed this response when compared to the untreated group. During two hours of anesthesia with ether, the TSH concentration declined in animals which were fed a low iodine diet at essentially the same rate as in animals on the same diet given an injection of 3 microng of triiodothyronine. Pentobarbital did not suppress TSH at room temperature. The release of thyrotropin after injection of synthetic thyrotropin-releasing hormone (TRH) was greater in animals anesthetized with pentobarbital than in controls and was slightly reduced in ether-anesthetized animals. This difference was observed when thyrotropin was given intraperitoneally or intravenously and the slope of the dose-response curves to TRH showed a flattening of the curve of rats treated with ether and a steeper slope of response in animals anesthetized with pentobarbital. We conclude that pentobarbital inhibited TSH response to cold but did not reduce the resting levels. Ether inhibited the rise of TSH in the cold and lowered the basal levels of TSH in animlas at room temperature. Pentobarbital increased the response to TRH and ether may have reduced the response to TRH.


ABO and Rh blood groups in relation to sex ratio, mean number and mortality of sibs. Data are presented on the sex ratio, mean number and mortality of the sibs of 17,060 schoolchildren, and on the sex ratio and mean number of the sibs of 5,785 blood donors, in relation to the children's and donors' sex and ABO and Rh blood groups. The sex ratio is significantly higher for the sibs of AB + B than for those of A + O schoolboys, and for the sibs of Rh-negative than for those of Rh-positive male blood donors, but in both cases the mean number of sibs is exactly the same for the first-mentioned as for the second-mentioned category.


Postsurgical decompression and immediate elemental feeding. The paralytic ileus, poor nutritional state, and related complications that often develop following serious serious abdominal surgery have traditionally been accepted as unavoidable. Now, a nasogastric decompression and feeding tube is availabe that usually prevents paralytic ileus and also provides a complete liquid diet starting immedicately after the operation, thus speeding wound healing and recivery.


Probable assignment of the serum cholinesterase (E1) and transferrin (Tf) loci to chromosome 1 in man. Suggestions of linkage in males between the E1 and Rh loci (Z = + 1.849; THETA = 0.20) and between the Tf and Rh loci (Z = + 0.595; THETA = 0.35) are presented. The assignment of the E1 and Tf loci to chromosome 1 and the order Tf:E1:PGD:Rh:PGM1 are cautiously proposed.


Sero-genetic studies on the San of South West Africa. The San, a physically, culturally and linguistically distinctive people, have been shown by archaeological records anciently to have inhabited the whole of Eastern and Southern Africa. They, in common with the Khoi, the other members of the Khoisan race, are confined now to Southern Africa and principally to Botswana and South West Africa, though a number are also found in Angola. Sero-genetic data concerning seven South West African groups are presented in this study, and confirm a shared overall genetic profile characteristic of the San in general, slightly different from that of the Khoi and in significant contrast with that of the Negroes.


Isolation and identification of the Cmicron4-domain of IgM. Intact Cmicron4-domain was isolated by molecular exclusion chromatography of reduced and alkylated Fc5 micron prepared by tryptic digestion at 60 degrees C of a monoclonal IgM. Two fragments were obtained of which one contained carbohydrate and the other none. These fragments were successfully separated by chromatography on insolubilised Concanavalin A. Cmicron4-domain was identified by means of amino acid sequence, amino acid composition, molecular weight and immunological analyses. It has a molecular weight of 14,700 daltons and results from tryptic cleavage at Lys-445 of the micron-chain.


Tumour-associated inhibition of immediate hypersensitivity reactions in mice. The intensity of anaphylactic shock was lower in C3H mice carrying a methylcholanthrene-induced tumour (McC3) than in their normal counterparts when immunized with ovalbumin and challenged i.v. after 14 days. This tumour-associated inhibitory effect on active systemic anaphylaxis was exerted mainly on events occurring after homocytotropic antibody synthesis because the serum titres of these antibodies were comparable in normal and tumour-bearing animals. In addition, passive systemic anaphylactic reactions were suppressed in animals carrying the tumour and the sensitivity of these animals to challenge with histamine and serotonin mixtures was also reduced. The presence of a growing McC3 tumour did not, however, diminish the amine-sensitizing effect of treatment with Bordetella pertussis vaccine. The McC3 tumour inhibited the generation of passive cutaneous anaphylactic reactions, an effect that was also exerted by a tumour extract, particularly when administered to the recipients shortly before antigen challenge. Thus immediate hypersensitivity reactions, like a variety of other immunological processes, can be inhibited by tumour products which by compromising the immune status of the host might permit tumour growth. The nature of the inhibiting factor is unknown, except that it is probably not the amine-degrading enzyme histaminase. In addition, which it is uncertain whether the inhibitory effect is exerted directly or indirectly, the possible importance of prostaglandins in the phenomenon is discussed.


Lipopolysaccharide serotyping of Neisseria meningitidis by hemagglutination inhibition. The method of hemagglutination inhibition was used to investigate the antigenic diversity of lipopolysaccharide (LPS) from Neisseria meningitidis and to develop a serotyping systems based on this antigen. The system uses outer membrane complex prepared by a simple extraction procedure to inhibit homologous hemagglutination reactions involving sheep erythrocytes sensitized with purified LPS and rabbit antiserum raised to whole meningococci. Antisera with specificity for eight different LPS determinants were used as typing sera to serotype a cross section of 67 meningococcal strains. Only two strains (both group A) were not typable with the eight sera, and most strains had more than one type. Comparison of LPS type and bactericidal serotype suggests that the LPS and protein serotypes are independent serological markers.


Reaction of squirrel monkeys to intratracheal inoculation with influenza/A/New Jersey/76 (swine) virus. To determine whether a model could be established for laboratory investigations, nine squirrel monkeys were inoculated intratracheally with 10(7) median egg-infectious doses of influenza virus type A/New Jersey/8/76 (HSW1N1) (swine influenza virus). They responded with clinically detectable illness including fever, leukopenia, decreased food consumption, increased respiratory rate, occasional coughing, labored breathing, nasal discharge, and lethargy. Convalescence was well advanced by the day 10. All monkeys excreted virus for 7 to 8 days. A scoring procedure (illness score) has been developed for use in studies of vaccine and chemotherapeutic efficacy.


Responsiveness of rabbit spleen and appendix cells to bacterial mitogens. Rabbit spleen and appendix cells were used to test the mitogenic activity of a commercial lipopolysaccharide (LPS) preparation from Salmonella typhimurium (Difco), a preparation extracted from it, and cell wall preparations from smooth (45/0) and rough (45/20) strains of Brucella abortus. On the basis of [3H]thymidine incorporation ratios (E/C), that is, the incorporation rate among cells treated with the mitogen relative to that of untreated cells, the extracted LPS and both Brucella cell wall preparations, but not the commercial LPS were potent mitogens for rabbit spleen cells. By the same criterion, only the Brucella cell wall preparation produced a significant, but minimally so, response among appendix cells. The weak responsiveness of appendix cells may be more apparent than real, however, and may not imply a difference in intrinsic susceptibility to mitogens by these two populations, because unstimulated appendix cells incorporated thymidine at 10 times the rate of unstimulated spleen cells. Appendix cells, then, may not be susceptible to further stimulation, even by active mitogens. Therefore, the significance of E/C ratios may be equivocal when materials are assayed for mitogenic activity on lymphoid populations whose basal activity is relatively high.


Composition of the lipopolysaccharide of Neisseria gonorrhoeae. Analysis of glycose and fatty acid content of lipopolysaccharide extracted from 38 strains of Neisseria gonorrhoeae indicated that glycoses common to colonial types 1 to 5 were glucose, mannose, and galactose, N-acetylneuraminic acid, 2-keto-3-deoxyoctulosonic acid (KDO), glucosamine, and galactosamine were also invariably present. Virulent colonial types 1 and 2 contained no rhamnose, in contrast to avirulent types 3 to 5 and several strains of the nonpathogenic species N. sicca and N. lactamica. Fucose, characteristic of these nonpathogenic species, was not present in the gonococci. Variation in the concentration of individual glycoses in different strains was also noted. Mannose-KDO, galactose-KDO, and glucose-KDO ratios of virulent gonococci exceeded those of avirulent organisms, except that the correlation for glucose was not quite so striking. This relationship was not found in N. sicca and N. lactamica strains. Fatty acid analyses of lipid A from gonococci showed that 10-, 12-, 14-, 16-, and 18-carbon acids, as well as 3-hydroxytetradecanoic acid, were present, but differences in concentration between colonial types, although evident in some cases, appeared less significant than glycose content.


Chicken embryo as an animal model for gonorrhea. Parameters of infection of the chicken embryo with Neisseria gonorrhoeae were defined in order to standardize infectious and lethal doses. Virulent (T1) and avirulent (T3) gonococci from two strains were used to infect 7- to 12-day-old White Leghorn chicken embryos via the yolk sac (YS) or chorioallantoic membrane (CAM) route. Infection of embryos was established following YS inoculation of 1 to 10 viable gonococci. Although 8- to 10-day-old embryos were the most susceptible, an inoculum of less than 100 gonococci was sufficient to kill any age embryo via this route. Embryos were less susceptible to infection via the CAM, where an inoculum of from 10(5) to 10(6) colony-forming units was lethal by 42 h. Strain and morphological type had a variable influence on the ability of the gonococcus to infect and kill the chicken embryo by either route; however, agar-grown and broth-grown organisms produced consistently similar mean lethal dose (LD(50)) and mean infective dose (ID(50)) values. LD(50) and ID(50) differences between T1 and T3 gonococci from strain 72H641 were not apparent after either YS or CAM inoculation of 8- or 10-day chicken embryos, respectively. YS and CAM LD(50) values for strain 72H641 T1 and T3 and CDC 9 T3 were also similar; however, these values were slightly lower for CDC 9 T1. In terms of infectivity or colonization, CDC 9 T1 and T3 had higher ID(50) values via the YS and lower ID(50) values via the CAM than 72H641. CDC 9 T1 was slightly more infective via the YS and less infective via the CAM than its T3 counterpart. Although the gonococcal strain used will influence interpretation of results, infection of both YS and CAM was highly reproducible in terms of gross pathology and of LD(50) and ID(50) data for a particular strain and colony type.


Age-related decline in the resistance of mice to infection with intracellular pathogens. Resistance to infection with Toxoplasma gondii and Listeria monocytogenes in BALB/c female mice decreased with increasing age. The decrease was apparent as early as 9 months of age and was more marked as the animals aged further. This age-related decline in resistance was not restricted to BALB/c female mice, as male and female mice of the C57BL/6 strain exhibited similar responses. With both pathogens, aged mice showed a more marked susceptibility to the strain of lesser virulence. Transfer of normal serum from old mice to young or old mice before infection with T. gondii resulted in an increased susceptibility to this organism, suggesting the presence of inhibitory factors or the absence of potentiating factors that are present in the serum of young mice.


Assay of Escherichia coli heat-labile enterotoxin with vero cells. The continuous cell line of African green monkey kidney, Vero, showed characteristic morphological changes in response to culture filtrates from toxigenic strains of Escherichia coli. The response compared favorably with that of Y-1 (mouse adrenal) and CHO (Chinese hamster ovary) cells. Vero cells were the simplest and most economical to maintain in the laboratory.


Homogeneity of protein serotype antigens in Neisseria meningitidis group A. Serotype antigens (STA), which have been shown to be constituents of outer-membrane protein, were extracted from group A meningococci with 0.2 M LiCl and pelleted by centrifugation at 150,000 X g. The STAs from 100 group A strains, which had been isolated from cases and carriers in various geographical locations, were compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Every STA produced three major protein bands with molecular weights of approximately 35,000, 39,000, and 45,000, respectively. This SDS-PAGE pattern is clearly distinct from that produced by the serotype of group B and C meningococci most commonly isolated from cases (group B type 2 and group C type 2). The group A STAs were also indistinguishable by immunodiffusion. However, differences in bactericidal reactions were demonstrated, suggesting that there are other antigens that play a role in antibody response.


Antigenic, chemical, and structural properties of cell walls of Histoplasma capsulatum yeast-form chemotypes 1 and 2 after serial enzymatic hydrolysis. Cell walls of Histoplasma capsulatum yeast-form chemotypes 1 (chem 1) and 2 (chem 2) treated sequentially with several polysaccharolytic enzymes and Pronase yielded soluble, nondialyzable polysaccharides at each step, which were analyzed for monosaccharides, protein composition, and serological activity. Polysaccharide recovered after digestion of chem 1 walls with beta(1-->3)-glucanase contained glucose > mannose > glucosamine > galactose. This fraction (chem 1 betaG(1)) was analyzed by polyacrylamide gel electrophoresis and contained a component having an apparent molecular weight of 120,000. The chem 1 betaG(1) fraction was reactive in immunodiffusion (ID), producing an immune precipitate not identical to the H and M factors of histoplasmin. In a side-by-side ID comparison with extracts of chem 2, the chem 1 betaG(1) antigen contained an additional determinant not found in chem 2 extracts when tested with goat antiserum to H. capsulatum. Therefore, the chem 1 antigen gave preliminary ID evidence of antigenic group specificity. A chemical difference observed was the absence of glucosamine from chem 2 polysaccharide. In complement fixation (CF) tests, 9 of 17 sera from human histoplasmosis patients reacted with chem 1 betaG(1), but some cross-reactivity with sera of patients with other systemic mycoses occurred. The immunoelectrophoretic patterns of chem 1 wall-derived polysaccharides showed a marked shift in mobility after Pronase digestion, implying the presence of covalent peptides. The ultrastructural appearance and serological activity of intact walls and enzyme-resistant mural cores were also studied. The surface of the mural cores of both chemotypes was perforated and frayed. In shadow-cast preparations both fibrillar and globular areas persisted in the mural cores. The CF end point serum dilutions showed an increase after alpha- and beta-glucanase extractions of chem 2 walls and fourfold reduction after Pronase digestion. The mural cores of both chemotypes were still reactive in CF tests and retained some ability to bind fluorescent antibody. The chem 1 mural core reacted with specific fluorescein-labeled H. capsulatum antiglobulins produced by adsorption with Blastomyces dermatitidis, thus indicating at least partial retention of H. capsulatum-specific factors. The presence of galactose, mannose, and glucose was detected in the mural cores as well as enriched levels of amino sugar, despite exposure to chitinase.


Variations in buccal cell adhesion of Neisseria gonorrhoeae. Human buccal cells were found to vary in their day-to-day capacity to support attachment of certain strains of gonococci. Gonococci serially cultured in vitro also differed in their day-to-day ability to attach to buccal cells.


Evaluation of nonspecific (alternative pathway) opsonic activity by neutrophil chemiluminescence. The role of alternative complement pathway activation in protection against infection is not well understood. We have investigated nonspecific opsonic activity in human adult, term neonatal and premature serum using the technique of neutrophil chemiluminescence (cl) to measure particle uptake. Following phagocytosis, neutrophils become metabolically active, produce excited molecular oxygen and emit a burst of light which can be detected and quantitated in a liquid scintillation counter. In the present study, zymosan particles were preopsonized in human serum and added to control neutrophils. Particles opsonized in adult serum produced a marked peak in CL. Serial dilutions of the serum prior to opsonization yielded proportionally lower peaks in CL. Opsonic activity as measured by the CL procedure was completely blocked by the addition of chelating agents which remove calcium and magnesium ions and block both classic and alternative complement pathway activation. Addition of an excess of magnesium ions (needed for alternative pathway activation) to the reaction mixture partially restored the opsonic activity. Opsonic activity as measured by the CL procedure was significantly depressed in approximately two-thirds of term and premature infant sera tested. Deficient nonspecific opsonic activity was closely correlated with serum levels of C3PA. These studies suggest that the CL procedure may be of value in the investigation of nonspecific opsonins in human serum.


Structure and biological function of human IgD. IX. Anti-IgD activation of human lymphocytes. Human peripheral lymphocytes were stimulated to incorporate tritiated thymidine when cultured with anti-sigma. The stimulation of lymphocytes by anti-sigma inversely correlates to PHA-induced lymphocyte transformation. In addition, lymphocytes from individuals with low serum IgD levels exhibited a significant response to anti-sigma, whereas, those with normal or slightly elevated levels of serum IgD showed minimal stimulation. This study is the first to provide evidence that cell surface IgD may regulate metabolic functions of lymphocytes and is consistent with the idea that IgD is a 'triggering' receptor.


Antibody reactivity in penicillin-sensitive patients determinated with different penicillin derivatives. 35 individuals showing reactions to penicillin of anaphylactic shock, angioedema or urticaria were investigated. Their skin sensitivity was analysed using 16 different penicillin derivatives. In addition, the content of circulating reagins against the penicilloyl structure in the patient's sera were analysed using RAST. 17 of the patients had negative skin reactions and RAST results to all substances tested. The other 18 were skin test-positive to at least one derivative but showed markedly heterogeneous patterns of skin reactivity. 14 had positive reactions against penicilloyl structures accompanied by anti-penicilloyl reagins. Four patients showed doubtful reactions only to penicillin or penicilloate and/or penilloate. These patients also had very low levels of reagins against penicilloyl in their sera. Positive skin test results using monovalent penicillin derivatives such as penicillin, penicilloate, penilloate, penicilloyl amide, penicilloyl-formyl-lysine, penicillamine, which cannot form a multivalent antigen with penicillyol specificity, indicated formation of other derivatives of importance in penicillin allergy, e.g., penicillamine protein conjugates. Three patients showed skin reactions to ampicillin polymer and two to benzyl-penicillin polymer. The skin tests performed with the penicillin derivatives used do not seem to give more information on the sensitivity of the patients than does the RAST using penicilloyl structures.


A reappraisal of the forces involved in hemagglutination. The underlying principles governing the second stage of hemagglutination, as described in 1965, have been reevaluated. This reappraisal reaffirms that red cell aggregation is dependent on the potential energy barrier (zota) between erythrocytes. This in turn is dependent on the ionic strength and dielectric constant of the bulk medium. Hemagglutination by IgG and IgM antibody is a function of the effective length of these immunoglobulins and the zota of the cells in the reaction mixture. Agglutination of cells occurs when the zota-potential is below a critical level. The enhancing action of natural and synthetic polymers (e. g., bovine albumin) is to lower zeta by raising the dielectric constant of the bulk medium. Several other recent concepts are discussed and the major differences discussed in light of data available.


Stimulation of pig lymphocytes with anti-immunoglobulin serum and mitogens. Mitogenic stimulation of pig lymphocytes by anti-immunoglobulin serum and by Con A, PHA, PWM and LPS has been measured by 3H-thymidine incorporation. Lymphocytes from blood, spleen and lymph node were readily stimulated by antiglobulin. Specific anti-micron serum was mitogenic for blood lymphocytes, indicating that those B cells with surface IgM can be stimulated by complexing of this surface component with antibody. An initial 2-hour incubation with anti-Ig was as effective as continuous incubation. Differentiation of stimulated cells into high rate synthesising plasmablasts was not observed. Pig blood lymphocytes were reactive to Con A, PHA, PWM and LPS. LPS was also tested against spleen and lymph node cells, which were responsive to this mitogen.


Soluble proteins in fluid from non-keratinizing jaw cysts in man. Fluids from non-keratinizing cysts of the jaws contain the main proteins found in plasma. The low relative concentration of macromolecular non-immunoglobulin proteins shows that there is no free passage of plasma proteins into the cyst fluid. Sufficient evidence was found to conclude that the immunoglobulins in cyst fluid are partly produced locally and partly derived from plasma. Cyst fluid immunoglobulins have antibody activity to rabbit erythrocytes. Accumulation of cyst fluid is primarily due to inadequate lymphatic drainage of the cyst cavity. Differential diagnosis between various types of non-keratinizing cysts cannot be based on the plasma protein patterns of their fluids.


Concentration of azidocillin, erythromycin, doxycycline and clindamycin in dental alveolar serum after single oral doses. Treatment of osteitis in the mandible after surgery is still a clinical problem. Levels of four antibiotics--azidocillin, erythromycin, doxycycline, and clindamycin--were measured in serum and dental alveolar serum in 42 patients undergoing oral surgery. The systemic serum concentrations were higher than the dental alveolar serum concentrations in all patients. The maximal concentration in the alveolar serum for azidocillin was 6.0-12.0 microng/ml, for erythromycin 0.7-1.3 microng/ml, for doxycycline 2.8-3.6 microng/ml, and for clindamycin 2.0-2.8 microng/ml. When the dental alveolar serum concentrations of the various antibiotics were related to their range of inhibitory concentrations for microorganisms isolated from mandibular osteitis, it was noticed that each drug achieved levels sufficient to inhibit most strains.


Reverse smoking in Andhra Pradesh, India: variability of clinical and histologic appearances of palatal changes. In Srikakulam, India, 101 patients with palatal lesions due to reverse smoking were followed for 1 year at monthly intervals to analyze the behavior of the various components of their palatal lesions. Of these patients, 24% stopped the habit completely and 27% changed to the conventional type of smoking. Palatal keratosis was very stable, whereas red areas changed into nonpigmented areas. Epithelial dysplasia was found in 23% of the 101 biopsies, the highest being in the red areas. Mild inflammation was seen in about half of the biopsies.


Continuous infusion pressure-monitored sialography. A technique for sialography has been developed which gives reproducible radiographs. By employing a constant infusion pump to deliver 0.2 ml of contrast medium per minute the patient does not experience pain and glandular detail is not obscured due to overfilling. Aqueous and oily contrast media may be used satisfactorily but there is greater latitude in times of exposure with the aqueous media.


Osteomyelitis of the maxilla in the newborn. A case of osteomyelitis of the maxilla in a healthy infant is presented, the etiology of which seems to have been an upper respiratory tract infection. There was no evidence of lowered resistance and immunologic studies did not reveal any abnormality. Lincomycin seems to be the antibiotic drug of choice. In the present case, it was used, effectively, in conjunction with other antibiotics and surgical drainage.


Ankylosis of the temporomandibular joint of a child. Report of a case. An ankylosis of the temporomandibular joint (TMJ) in early childhood may lead to growth disturbances. In the reported case, unilateral ankylosis caused asymmetry and serious difficulties in eating and breathing during sleep. The autologous costochondral junction serving as a growth center, replacing the condylar defect, was used as arthroplastic material in a patient 8 years of age. In a follow-up 6 years later the patient revealed satisfactory opening movements and good chewing function. It was noted that the operated grafted side increased in length slightly more than the other side.


Pseudomonas protease. Purification, partial characterization, and its effect on collagen, proteoglycan, and rabbit corneas. The extracellular protease of a virulent strain of Pseudomonas aeruginosa was purified by DEAE-cellulose chromatography in two steps. SDS-polyacrylamide gel electrophoresis of the purified enzyme revealed a single band, and the enzyme was shown to be the major component of the bacterial filtrate. The protease was fully inhibited by Na2 EDTA, 1,10-orthophenanthroline, L-cysteine and Zn+2 ions but was insensitive to dissopropylphosphofluoridate. The elastase substrates orcein-elastin and acetyl-L-alanyl-L-alanyl-L-alamine-methyl ester were degraded by the enzyme. The protease activity toward soluble and insoluble collagen was found to be limited to the telopeptide region of the collagen molecule. With soluble collagen, conversion of the beta and gamma chains into monomeric alpha chains was observed. About 60% of the  total proteoglycans and 1.5% of the total collagen were solubilized from rabbit corneas following incubation with the enzyme, and the solubilized products were nondialyzable. It was concluded that the purified protease has little or no collagenolytic activity and that dissolution of the cornea by Pseudomonas protease infection results essentially from the degradation of the protein backbone of the corneal proteoglycans.


Retinal damage from long-term exposure to laser radiation. The maculae of rhesus monkeys were exposed to an argon-ion lazer operated in the TEM00 continuous wave mode at a wavelength of 514.5 nm. Both ophthalmoscopic and histopathologic evaluations of exposure sites were obtained. Threshold (ED50) values were obtained for 0.5, 5, 30, 120, and 1,000 sec. exposure times. Presence of minimum visible lesions was assessed ophthalmoscopically at both 1 hour and 24 hours after exposure. With increasing exposure times, a 24 hr. lesion-appearance criterion resulted in ED50 values too low to be consistent with a thermal damage mechanism. In contrast, exposure to neodymium laser radiation at a 1,060 nm. wavelength for 120 sec. produced only ED50 values consistent with those associated with thermal injury. These results suggest that the damage mechanisms for long-duration exposures to visible light may involve photochemical processes initiated by the interaction of visible light with the retinal photopigments.


Intraocular axonal swelling produced by partial, immediately retrobulbar ligature of optic nerve. This is the first report of an experimental intraorbital ligature producing papilledema characterized by axonal swelling and accumulation of mitochondria in the lamina retinalis of the optic disc of rhesus monkeys subjected to immediately retrobulbar ligature of a portion of the optic nerve. This is an improved technique for investigating the pathogenesis of papilledema and optic neuropathy.


Treatment of Pseudomonas keratitis in the rabbit with antibiotic-steroid combinations. It has been suggested that corticosteroids plus the effective antibiotics are more beneficial than antibiotics alone in reducing inflammation in infectious keratitis. Topical and subconjunctival corticosteroids as adjuncts to gentamicin therapy were evaluated by microbiological assay and clinical parameters in experimentally induced Pseudomonas keratitis in rabbits. Various strengths of dexamethasone given concurrently or delayed in conjunction with gentamicin therapy were studied. There was no statistically significant difference between combinations and the antibiotics alone in this experimental model.


Gap junctions between optic nerve head astrocytes. Astrocytes of the primate and human optic nerve head are joined to each other by the gap junction type of intercellular membrane specialization. Although the precise function of these contacts is not fully determined, they may serve such diverse roles as adhesive bonding and intercellular electrical and chemical coupling.


The inactivation of beta-lactamase I from B. Cereus by dicloxacillin: effect of buffer composition on the kinetic characteristics of the modified enzyme. Buffer composition affects the pattern of inactivation of B. cereus beta-lactamase I by dicloxacillin. At pH 7.0, in the presence of phosphate ions, dicloxacillin appears to cause essentially an irreversible inactivation of the catalytic site of the enzyme, while in a tris-cacodylate buffer the inactivation process appears to affect essentially the substrate-binding site. Buffer ions act presumably by interacting themselves with either the catalytic or the substrate-binding site of the enzyme, thus modifying the affinity of these sites for the halogenated isoxazolyl-penicillins.


Hyperviscosity syndrome associated with lymphocytic leukemia in three dogs. Hyperviscosity syndrome was associated with increased plasma content of monoclonal immunoglobulin (IgA or IgM) in 3 dogs with lymphocytic leukemia. The diagnosis of lymphocytic leukemia was based on the finding of a large number of mature lymphocytes in the blood and bone marrow. The clinical signs included weakness, lethargy, depression, and coughing due to congestive heart failure. Consistent physical findings were splenomegaly, with or without peripheral lymphadenopathy, and funduscopic abnormalities. Of the 2 dogs treated successfully with chlorambucil, 1 remains in remission after withdrawal of the drug for over 1 year.


SP127, a bacterial protein enhancing the activity of macrolide antibiotics against Pseudomonas aeruginosa: isolation and characterization. Some strains belonging to the genera Bacillus and Pseudomonas were found to produce in the culture broth, a protein which synergistically enhanced the activity of macrolide antibiotics against Pseudomonas aeroginosa. The protein produced by Bacillus brevis ATCC 8185, designated SP127, was isolated in a pure state by means of ammonium sulfate fractionation, CM-cellulose column chromatography and gel filtration on Sephadex G-100. The sedimentation coefficient and the molecular weight of SP127 were 1.8 s and 15,000, respectively according to the analytical ultracentrifuge. The isoelectric point of SP127, obtained by electrofocusing experiments, was over 10.0. Half cystine and histidine were absent in the molecule.


Mode of action of the protein, SP127, which enhances the activity of macrolide antibiotics against Pseudomonas aeruginosa. Antibiotics, the activity of which enhanced against Pseudomonas aeruginosa by SP127, were restricted to the basic macrolide antibiotics such as erythromycin, maridomycin and oleandomycin, the neutral macrolide antibiotics such as lankamycin and lankacidin C, vancomycin and enramycin. Synergistic activity of SP127 with the above antibiotics was found against Pseudomonas aeruginosa and several strains of Escherichia coli, but not against Proteus vulgaris and macrolide-resistant Staphylococcus aureus. SP127 had extremely weak proteolytic but no lytic activity. From the isotopic experiments, the action of SP127 was partially attributed to the promotion of antibiotic penetration to cells of Pseudomonas aeruginosa.


Feedback inhibition of the synthesis of an antibiotic: aurodox (X-5108). The effect of aurodox on its own biosynthesis by Streptomyces goldiniensis was studied. It was found that addition of exogenous aurodox inhibits further accumulation of aurodox by the antibiotic-producing culture. Both long term fermentation studies with aurodox-14C and precursor incorporation studies over short time periods indicated that aurodox synthesis was regulated by feedback inhibition. The concentration of aurodox required to completely block further synthesis of the antibiotic was about 400 microng/ml. This is the same as the maximum concentration of aurodox normally accumulated by the culture used in this study. Antibiotic synthesis was inhibited not only by aurodox but also by some structural analogs of aurodox including several having no antibacterial activity. This effect was immediate and readily reversible, indicating that it could be due to inhibition of an enzyme(s) involved in the biosynthesis of aurodox.


Microbial production of vitamin B12 antimetabolites. III Compound 102804 from Bacillus cereus. A basic compound with empirical formula C12H16N2O5 was isolated from Bacillus cereus 102804 fermentations of a soybean meal-glucose medium. The inhibitory activity of compound 102804 on growth of Gram-positive and Gram-negative bacteria growing in a chemically defined medium was reversed by vitamin B12, by L-methionine, and by D-methionine. It has no inhibitory activity for Escherichia coli (Davis 113-3) when grown in media containing L-methionine. The biosynthesis of 102804 coincides with the sporulation of the B. cereus, and the compound is not produced in the absence of sporulation.


Skin, preoptic, and core temperatures influence behavioral thermoregulation. Squirrel monkeys (Saimiri sciureus) will learn to regulate their chamber air temperature (Ta) behaviorally by selecting between two air temperatures, 10 and 50 degrees C. When transient (10-min) displacements of local preoptic temperature (Tpo) are made around neutral, via water-perfused thermodes, changes in Ta selected are linear and inversely proportional to the Tpo displacement. The same result occurred in the present study when 10-min Tpo displacements were made around a clamped (36--41 degrees C) Tpo. The higher the clamped Tpo level, the steeper the linear functions relating behavioral response to Tpo transients. This change of gain was manifested in behavioral adjustments of Ta or skin temperature. The data showed that the temperature of the extrahypothalamic body core (measured rectally) was driven by the clamped Tpo to a nonneutral level. When this level was maintained stable during the Tpo transients, core temperature was shown to be a multiplicative factor governing the gain of the behavioral effector responses.


Mode of degradation of precursor-specific ribonucleic acid fragments by Bacillus subtilis. A precursor of 5S ribosomal ribonucleic acid (rRNA) from Bacillus subtilis was cleaved by ribonuclease (RNase) M5 in cell-free extracts from B. subtilis to yield the mature 5S rRNA plus RNA fragments derived from both termini of the precursor. The released, mature 5S rRNA was stable in these extracts; however, as occurred in vivo, the precursor-specific fragments were rapidly and completely destroyed. Such destruction was not observed in the presence of partially purified RNase M5, so fragment scavenging was not effected by the maturation nuclease itself. The selective destruction of the precursor-specific fragments was shown to occur through a 3'-exonucleolytic process with the release of nucleoside 5'-monophosphates; the responsible activity therefore had the character of RNAse II. Consideration of the primary and probable secondary structures of the precursor-specific fragments and mature 5S rRNA suggested that involvement of 3' termini in tight secondary structure may confer protection against the scavenging activity.


Initiation of deoxyribonucleic acid replication in germinating spores of Bacillus subtilis 168 carrying the dnaB (Ts)134 mutation. The nature of the deoxyribonucleic acid synthesis reported by others to occur at 45 degrees C in germinating spores of the temperature-sensitive deoxyribonucleic acid initiation mutant of Bacillus subtilis 168, TsB134, has been investigated. Density transfer experiments, using 5-bromouracil, show that a normal round of replication can occur in a significant fraction of the spore population under such conditions. No repair synthesis is detectable. The possibility raised by this finding, that initiation of the first round of replication during spore outgrowth is unique in that its initiation is determined prior to germination, has been investigated by comparing the behavior of germinating spores of isogenic strains of B. subtilis 168, one carrying and the other without the dnaB (Ts)134 mutation. It is shown that deoxyribonucleic acid synthesis in the Ts strain is very sensitive to temperature in the vicinity of 45 degrees C. At a slightly higher temperature, 49 degrees C, initiation of the first round of replication in the Ts strain is completely (greater than 96%) blocked, but it proceeds normally in the Ts(+) strain. Thus, it is concluded that, after the germination of a spore, the action of the dnaB134 gene product is an obligatory requirement for initiation of the first round of replication. The initiation of replication that can occur in spores of the original TsB134 strain germinating at 45 degrees C is presumably due to incomplete inactivation of the dnaB134 gene product under such conditions.


Biosynthesis of wall polymers in Bacillus subtilis. Preparations of membrane plus wall derived from Bacillus subtilis W23 were used to study the in vitro synthesis of peptidoglycan and teichoic acid and their linkage to the preexisting cell wall. The teichoic acid synthesis showed an ordered requirement for the incorporation of N-acetylglucosamine from uridine 5'-diphosphate (UDP)-N-acetylglucosamine followed by addition of glycerol phosphate from cytidine 5'-diphosphate (CDP)-glycerol and finally by addition of ribitol phosphate from CDP-ribitol. UDP-N-acetylglucosamine was not only required for the synthesis of the teichoic acid, but N-acetylglucosamine residues formed an integral part of the linkage unit attaching polyribitol phosphate to the cell wall. Synthesis of the teichoic acid was exquisitely sensitive to the antibiotic tunicamycin, and this was shown to be due to the inhibition of incorporation of N-acetylglucosamine units from UDP-N-acetylglucosamine.


Genetic loci of hemolysin production in Streptococcus faecalis subsp. zymogenes. Two plasmids corresponding to molecular weights of 38.5 X 10(6) and 3.6 X 10(6) have been identified in Streptococcus faecalis subsp. zymogenes strain X-14. The larger plasmid is required for hemolysin-bacteriolysin production. Strain L2, a nonlytic nitrosoquanidine mutant of strain X-14, still harbors the hemolytic plasmid and produces the lysin component, but not the activator component, of the lytic system. Conjugal transfer of this plasmid from strain L2 to plasmid-free strains and strains cured of the 38.5-megadalton plasmid gives rise to hemolytic recipients. This implicates a gene in hemolysin production at a site other than the 38.5-megadalton plasmid.


Effect of macromolecular synthesis on the coordinate morphogenesis of polar surface structures in Caulobacter crescentus. The Caulobacter polar surface structures (flagella, pili, and the deoxyribonucleic acid phage phiCbK receptors), which are expressed at proximal sites of swarmer cells in a coordinate manner (Shapiro, Annu. Rev. Microbiol., 30:377-407, 1976) could be blocked by a single mutation. The mutant C. crescentus CB13 ple-801 did not form these surface structures when grown at 35 degrees C. Upon shift down to 25 degrees C, the mutant cells initiated the formation of the surface structures. When mitomycin C was added to the mutant culture upon shift down from 35 to 25 degrees C, phiCbK receptor formation was inhibited to a minimal level. Rifampin and chloramphenicol completely inhibited phiCbK receptor formation when added to the mutant culture upon shift down. Deoxyribonucleic acid as well as ribonucleic acid and protein synthesis seem to be required for the formation of phiCbK receptors. Penicillin V also inhibited phiCbK receptor formation, indicating the involvement of cell wall synthesis. When the mutant CB13 ple-801 cells were shifted down briefly from 35 to 25 degrees C and then shifted up to 35 degrees C, flagella and phiCbK receptors were formed even at 35 degrees C to different extents depending on how long the cells were incubated at 25 degrees C. This formation of the surface structures at 35 degrees C was inhibited by rifampin. From these results, it appears that translation, assembly, or localization processes for the formation of the surface structures are not temperature sensitive at 35 degrees C in the pleiotropic mutant CB13 ple-801. The syntheses of deoxyribonucleic acid and the cell wall do not appear to be temperature sensitive either, since the mutant grows normally at 35 degrees C. It is suggested that there exists a regulatory step that commits the cells to initiate the synthesis of requisite ribonucleic acid for the formation of the polar surface structures.


Altered accumulation of a membrane protein unique to a membrane-deoxyribonucleic acid complex in a dna initiation mutant of Bacillus subtilis. Membrane-deoxyribonucleic acid complexes (M-bands) have been isolated from Bacillus subtilis by their affinity for crystals of Mg2+-Sarkosyl. The membrane proteins of these complexes were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Comparison of the membrane protein composition of M-band and unfractionated membrane revealed three protein components of 125,000 (mac-1), 57,000 (mac-2), and 42,000 (mac-3) daltons unique to M-band membrane. Growth of a temperature-sensitive dna initiation mutant at the restrictive temperature resulted in an accumulation in the membrane of mac-2. This accumulation did not begin, however, until cell growth had nearly ceased, some 3 to 4 h after the cessation of deoxyribonucleic acid synthesis. Upon return of the mutant to the permissive temperature, mac-2 did not begin to return to normal levels until after the first round of deoxyribonucleic acid synthesis. A protein of 30,000 daltons, common to both M-band and whole membrane, was found to disappear from the membrane when the mutant was grown at the restrictive temperature. This disappearance is the result of increased degradation or removal from the membrane followed by a decreased rate of synthesis or insertion.


Properties of an Escherichia coli K-12 mutant defective in the transport of arginine and ornithine. A canavanine-resistant mutant strain, defective in the transport of arginine and ornithine, was isolated and characterized. Experiments presented show that both the kinetics of influx and the steady state of accumulation of arginine and ornithine are affected by the mutation, whereas the activity of other related transport systems remains unchanged. On the basis of competitive studies, it is concluded that L-canavanine can inhibit efficiently the arginine-specific uptake system. D-Arginine appears to be a moderate inhibitor. None of the basic amino acid-binding proteins of the mutant strain showed detectable alterations in terms of quantity, physical properties, or affinity constants. Studies on the relationship between the number of transport carriers and the steady state of accumulation of arginine suggested the presence of a reduced number of membrane carriers in the mutant strain. It is proposed that the mutation affects a regulatory gene concerned with controlling the amount of membrane carriers produced, which are components of the arginine- and ornithine-specific uptake systems. The mutation maps at min 62 on the recalibrated linkage map of Escherichia coli K-12, in a locus closely linked or identical to argP.


Basis for the observed fluctuation of carboxypeptidase II activity during the cell cycle in BUG 6, a temperature-sensitive division mutant of Escherichia coli. Diaminopimelyl-d-alanyl carboxypeptidase (carboxypeptidase II) is most active at the time of division, whether measured in toluene-treated cells of Escherichia coli K-12 strain D11-1, fractionated by size, or in toluene-treated cells of the temperature-sensitive division mutant, BUG 6 (B. D. Beck and J. T. Park, 1976). The present investigation has now shown that, under conditions that permit division, the increased carboxypeptidase II activity in toluenetreated cells of BUG 6 is probably not due to protein synthesis. Although dividing cells are more permeable than nondividing cells, permeability differences are not sufficient to account for the changes in carboxypeptidase II activity. Thus, in the toluene-treated nondividing cells, carboxypeptidase II is present, but its activity is masked, which suggests the presence of an inhibitor. Another striking difference between nondividing and dividing cells is that carboxypeptidase II is much more readily released from dividing cells by both tris(hydroxymethyl)aminomethane-ethylenediaminetetraacetic acid and toluene treatment. Carboxypeptidase II was partially purified and found to be an 86,000-molecular-weight protein consisting of two 43,000-molecular-weight polypeptides. Tris(hydroxymethyl)aminomethane-ethylenediaminetetraacetic acid treatment of nondividing cells releases less than 10% of the carboxypeptidase II and other periplasmic proteins that are releasable from dividing cells.


Effect of ethylenediaminetetraacetate upon the surface of Escherichia coli. The effect of ethylenediaminetetraacetate (EDTA) on the envelope of two strains of Escherichia coli (B and Cla) was studied with freeze-fracturing methods. Untreated cells showed the outer membrane's outer surface with a fine texture of randomly spaced depressions of about 4.5-nm diameter; small areas with symmetrical arrangements of structural surface elements were also observed. The outer membrane's fracture plane revealed a random distribution of particles on its "concave" plane, only occasionally interrupted by particle-free areas. The "convex" aspect of the outer membrane's fracture plane showed only a few scattered particles. The cleavage plane of the inner membrane was often interrupted by many localized elevated plateaus, at which the cleaving process had, for short distances, switched to the outer membrane. The effects of EDTA treatment were mainly seen in the structure of the freeze-etched outer membrane: (i) the pits as well as the symmetrical surface elements of the outer membrane's outer surface had disappeared; (ii) a number of plateaus (about 20 to 50/cell) were seen at which a cleavage plane within the inner membrane had switched to the hydrophobic portion of the outer membrane (outer membrane's fracture plane). These plateaus were also visible in untreated cells; however, EDTA treatment apparently caused an increased exposure of plateaus. Surface areas, exposed by freeze-etching, revealed the underlying plateaus as elevations in the surface contour of the cell, suggesting a slower etching rate in the zones of the plateaus relative to the rest of the outer membrane. Well-defined, particle-free patches in the outer membrane's fracture plane, concave, were more frequent and larger in size after EDTA treatment than in the controls. In the presence of glycerol, the cells often cleaved in the outer membrane's fracture plane, but isolated plateaus were rarely observed. After metabolic poisoning of cells for 15 to 25 min at 37 degrees C, the plateaus had widened. These data suggest that the material of the plateaus has a slow rate of lateral diffusion. Placement of EDTA-treated cells in fresh medium at 37 degrees C caused, after 3 to 5 min, the reoccurrence of the pitted surface structure. We propose that the plateaus represent localized zones, at which newly synthesized lipopolysaccharide has been inserted.


Studies on the restoration of the activities of Ribonucleases by polyamines in the presence of various ribonuclease inhibitors. The effect of polyamines on ribonucleases in the presence of various inhibitors (poly(G), heparin, and rat liver RNase inhibitor) has been studied. Bovine pancreatic RNas A and a ribonuclease from horse submaxillary gland (RNase HS) were inhibited by the inhibitors, but RNase T1 and RNase M were not inhibited. Polyamines were found to restore the activites of RNase A and RNase HS inhibited by poly(G) or heparin but not those activities inhibited by rat liver RNase inhibitor. When poly(U) and poly(C) were used as substrates, the inhibitory effects of poly(G) and heparin were greater with poly(U) than poly(C) as a substrate. However, when poly(C) was used as a substrate in the presence of either of the above inhibitors, the restoration of RNase activity by sperimine was more efficient. In fact, a stimulatory effect was observed. From the double-reciprocal plots, it was concluded that polyamines restored the activiities of RNases by increasing the availability of the substrate and enzyme to each other. The restoration of enzyme activity by polyamines occurred through the binding of the polyamines to the inhibitor and the subsequent release of enzyme from the inhibitor.


The structure and function of ribonuclease T1. XXIII. Inactivation of ribonuclease T1 by reversible blocking of amino groups with cis-aconitic anhydride and related dicarboxylic acid anhydrides. Ribonuclease T1 [EC 3.1.4.8] was inactivated rapidly by treatment at pH 8.0 and 0 degrees C with cis-aconitic anhydride and related dicabroxylic acid anhydrides, including citraconic, maleic, and succinic anhydrides. Under reaction conditions used, roughly 90% inactivation occurred within 30 min. Analyses of the inactivated enzymes indicated that the reaction took place fairly specifically at the alpha-amino group of the N-terminal alanine and the epsilon-amino group of lysine-41. Upon incubation of these inactivated enzymes at pH 3.6 and 37 degreeC, the activity was regenerated to various extents, depending on the nature of the introduced acyl groups. Under these conditions, the enzyme modified with cis-aconitc anhydride or citraconic anhydride recovered much of the origninal activity after 48 h whereas the enzyme modified with maleic anhydride recovered its activity only partially. Practically no activity was regenerated in the case of the enzyme modified with succinic anhydride under these conditions. The inactivation appears to be due mainly to the effect of the carboxyl group introduced at the epsilon-amino group of lysine-41. The results suggest the usefulness of cis-aconitic anhydride as a reversible blocking reagent for amino groups in proteins.


Anhydrotrypsin and trypsin: subtle difference in the active-site conformations detected by chemical modification and CD spectroscopy. The reactivities of the active-site histidine residue in bovine trypsin and its anhydro-derivative, as well as in Streptomyces griseus trypsin and its anhydro-derivative have been compared. The reactivity with TLCK was found to be lost in both of the anhydrotrypsins. On the other hand, alkylation by iodoacetamide either in the presence or absence of 1-methylguanidine proceeded faster in anhydrotrypsins than in trypsins. These differential responses to alkylating reagents are discussed in terms of a subtle change in the active-site conformation which occurs during the conversion of trypsin into anhydrotrypsin. The examination of difference CD spectra, produced by interaction with benzamidine or beta-naphthamidine, also suggested a conformational difference of the active-site between the proteins of bovine origin.


A novel pathway for L-citramalate synthesis in Rhodospirillum rubrum. When [14C]propionate was incubated with a cell-free extract of Rhodospirillum rubrum in the presence of glyoxylate, ATP, CoA, Mg2+, and Mn2+, radioactivity was incorporated into mesaconate (MSA) as well as into beta-methylmalate (MMA) and citramalate (CMA). MSA was suggested to be an intermediate of the conversion of MMA to CMA based on the following observations. (i) When non-labeled MSA was added to the CMA-forming reaction system, radioactivity was trapped in MSA. (ii) When MSA was incubated with the cell-free extract, CMA was formed. (iii) The alpha-carboxyl group of CMA was shown to be derived from the beta-carboxyl group of MMA, implying that CMA was formed from MMA via MSA through successive dehydration and hydration. From the results of Sephadex G-10 column chromatography of the reaction products, beta-methylmalyl-CoA and mesaconyl-CoA were presumed to be involved in the reaction. A new CMA-forming metabolic pathway is proposed as follows: erythro-beta-methylamalyl-CoA leads to mesaconyl-CoA leads to MSA leads to L-CMA.


Subfractionation of rat liver microsomes by immunoprecipitation and immunoadsorption methods. Rabbit antisera were prepared against cytochrome b5 and NADPH-cytochrome c reductase [EC 1.6.2.4] purified from rat liver microsomes, and utilized in examining the distribution of these and other membrane-bound enzymes among the vesicles of rat liver microsomal preparations by immunoprecipitation and immunoadsorption methods. Smooth microsomes with an average vesicular size of 200 nm (diameter) and sonicated smooth microsomes with an average diameter of 40-60 nm were used in subfractionation experiments. Immunoprecipitation of microsomal vesicles with anti-cytochrome b5 immunoglobulin failed to show any separation of the microsomes into fractions having different enzyme compositions. Cytochrome b5 was apparently distributed among all vesicles even when sonicated microsomes were used. When the antibody against NADPH-cytochrome c reductase was used, however, immunoadsorption of microsomes on Sepharose-bound antibody produced some separation of NADPH-cytochrome c reductase and cytochrome P-450 from NADH-cytochrome b5 reductase and cytochrome b5. The separation was more pronounced when sonicated microsomes were used. These results indicate microheterogeneity of the microsomal membrane, and suggest the clustering of NADPH-cytochrome c reductase and cytochrome P-450 molecules in the membrane.


Alteration of immunological properties of bovine serum albumin by covalent attachment of polyethylene glycol. Methoxypolyethylene glycols of 1900 and 5000 daltons have been attached covalently to bovine serum albumin using cyanuric chloride as the coupling agent. When sufficient polymer is attached, the modified bovine serum albumin appears to lose its immunogenicity in the rabbit and, on intramuscular or intravenous injection, elicits antibodies neither to itself nor to native bovine serum albumin. It does not react with antibodies raised against native bovine serum albumin. Bovine serum albumin to which methoxypolyethylene glycol has been attached exhibits a blood circulating life in the rabbit rather similar to native bovine serum albumin, except that it is not removed from circulation by the eventual development of antibodies. Modified bovine serum albumins which had been iodinated with 125I, or prepared with [14C]cyanuric chloride, were injected intravenously in rabbits. Both labels appeared almost quantitatively in the urine after 30 days. The modified bovine serum albumins showed substantial changes in properties, such as solubility, electrophoretic mobility in acrylamide gel, ion exchange chromatography, and sedimentation, as compared with the unmodified protein.


Carbonic anhydrase activity in intact red blood cells measured with 18O exchange. We have used a stirred, temperature-regulated, reaction vessel separated by a Teflon membrane from the ion source of a mass spectrometer to monitor continuously the time course of disappearance of C18O16O, mass 46, at chemical equilibrium as the 18O exchanges with 16O in water. This instrument is sensitive to less than 0.01 mm Hg of partial pressure of C18O16O with a response time of less than 3 s. The equation of Mills and Urey was used to calculate the hydration velocity constant for uncatalyzed or catalyzed homogenous solutions from the exponential disappearance of mass 46. Addition of red blood cells to the reaction mixture produces biphasic (double exponential) disappearance curve for mass 46. A theory of this process has been developed which describes the time course of [C18O16O] as a function of the catalytic factor for intracellular carbonic anhydrase (A) and the permeability of the cell membrane to HCO3- (P) in addition to the known values; water volume of the cells in the suspension, extracellular pH, the extracellular hydration reaction velocity constant, ku, and dehydration reaction velocity constant, ku. Using this theory, A and P were estimated from the disappearance curve for mass 46 at different values of hematocrit in the reaction mixture, both by a trial and error curve fitting procedure and by a more convenient graphical linearization method. The values of A and P obtained were very sensitive to small amounts of lysis (less than 1%), but the graphical method of analysis minimized this effect. For the blood cells of five normal subjects suspended in 24 mM bicarbonate in 145 mM NaCl at pH 7.4 and 37 degrees, using the graphical method we obtained an average value of 9,906 for A as compared to 19,900 for a comparable concentration of hemolysate. Correcting for a lower pH and chloride concentration inside the cell the latter figure would reduce to 17,500, still 80% higher than the intracellular value. The reason for this discrepancy is not clear. The average permeability of the red cell to bicarbonate ion was 3 X 10(-4) cm/s.


Incorporation of N-acetyl-D-glucosamine from UDP-N-acetyl-D-glucosamine by isolated membranes of Bacillus subtilis. Identification of undecaprenyl poly(N-acetylglucosaminyl pyrophosphate). Membrane isolated from Bacillus subtilis strain 168 incorporated GlcNAc from UDP-GlcNAc directly onto undecaprenyl phosphate via transphosphorylation and subsequent transglucosylations. Chain lengths of 6, 4, and 1 units of GlcNAc were found. Approximately 80% of the isotope incorporated was extracted into chloroform:methanol (2:1 v/v), and could be distinguished from the undecaprenyl disaccharide cell wall intermediate by a different elution pattern on DEAE-cellulose (acetate form). The GlcNAc-lipid(s) were eluted from a similar column in chloroform:methanol:water (10:10:3, v/v) with 6 mM NH4COOH indicating a pyrophosphate linkage between the lipid and the GlcNAc. The GlcNAc-lipid(s) were not degraded by conditions which completely deacylated [32P]glyceryl phospholipids, but were rapidly hydrolyzed by mild acid treatment (0.005 N HCl, 90 degrees) with the release of oligosaccharide phosphate (typical of sugars linked to undecaprenyl pyrophosphate). Catalytic hydrogenation of the GlcNAc-lipid(s) resulted in the release of water-soluble sugar phosphate. Under these same conditions, undecaprenyl pyrophosphate and undecaprenyl disaccharide cell wall intermediate were similarly effected while [32P]glyceryl phospholipids remained intact. The formation of GlcNAc-lipid(s) in vitro was inhibited if membranes were prepared from cells previously treated with bacitracin. Thus, the GlcNAc-lipid(s) has the properties of undecaprenyl poly(N-acetylglucosaminyl pyrophosphate) and may represent a new synthetic role of the polyisoprenyl lipid in B. subtilis.


Glucuronosyl diacylglycerol of Pseudomonas diminuta ATCC 11568. In vitro biosynthesis from UDP-glucuronate and diacylglycerol. The biosynthesis of glucuronosyl diacylglycerol from UDP-glucuronate and diacylglycerol is catalyzed by an enzyme found in both the 34,800 X g supernatant and particulate preparations from disrupted Pseudomonas diminuta (ATCC 11586). UDP-glucuronate served as the glucuronosyl donor and could not be replaced by glucuronic acid, glucuronate-1-phosphate, and a number of nucleotide-linked sugars. The maximum velocity was estimated to be 19 nmol of glucuronosyl diacylglycerol synthesized/h/mg of protein in the presence of the 34,800 X g particulate enzyme and 63 nmol/h/mg of protein with the 34,800 X g supernatant preparation. The apparent Km for UDP-glucuronate was 4.2 micronM for supernatant and 4.4 to 6.0 micronM for particulate preparations. The biosynthesis of glucuronosyl diacylglycerol in vitro, was strongly dependent upon exogenous diacylglycerols containing unsaturated and shorter chain fatty acids. The enzymatic activity was very heat-labile and lost about 80% of the initial rate of synthesis after preincubation for 5 min at 37 degrees. The reaction was stimulated by 14.7 mM Triton X-100 and had an optimal pH of 7.1 and an ionic strength of 0.2 M. Divalent cations were not required.


Inosine uptake by cultured fibroblasts from normal and purine nucleoside phosphorylase-deficient humans. Purine nucleoside phosphorylase-deficient cultured human fibroblasts accumulate inosine from the medium at 60% of the rate in wild type cells when the extracellular inosine concentration is 10 micronM and 30% of the normal rate when inosine is present at 100 micronM. When 10 micronM inosine is present, uridine but not hypoxanthine inhibits the accumulation of inosine. There exist two transport systems for inosine. One is shared with pyrimidine ribonucleosides and is the predominant one at 10 micronM inosine; the other is purine nucleoside phosphorylase-dependent and prevails at 100 micronM inosine.


"Tooth" sign in patellar degenerative disease. In the total examination of the knee, degenerative changes and ossification in the quadriceps tendon easily recognized on the lateral roentgenogram may cause confusion in diagnosis when seen on the axial view. Vertical ridging of the osteophytes at the patellar insertion of the quadriceps tendon can resemble dentate structure ("tooth" sign). Two hundred and fifty examinations of the knee in different age groups were reviewed and several anatomical sections of the quadriceps-patella insertion were studied. The "tooth" sign represents the relief of severe osteophyte formation in the bundle of the quadriceps tendon at its insertion into the patella.


Diphtheroid osteomyelitis.

Diphtheroid osteomyelitis. Isolates of either Corynebacterium diphtheriae or Propionibacterium acnes from osteomyelitis are not necessarily contaminants, as shown by the cases of three patients who had bone and joint infections in which these organisms were pathogenic (one in pure culture and two in mixed cultures). Previous operation or other factors that compromise host resistance create the setting for these opportunistic organisms. Penicillin with or without streptomycin is the treatment of choice, but if penicillin or streptomycin cannot be used then the cephalosporins, clindamycin, and erythromycin are acceptable alternatives (depending on susceptibility studies). In general, the prognosis is good.


Glycogen metabolism in adult rat liver parenchymal cell primary cultures. Parenchymal cells were isolated from adult rat liver with an enzyme perfusion technique. The single-cell suspension, representing 40-50% of the liver's hepatocytes was suspended in medium and maintained in primary culture for up to four days. The cells were found to carry out glycogen synthesis for the first eight hours in culture after which time the accumulated glycogen was gradually degraded. The ability of the liver cell cultures to accumulate glycogen was found to be dependent upon the metabolic state of the animal prior to cell isolation. Cells prepared during the feeding period from animals on the 8+16 feeding schedule had markedly different capacities for glycogen accumulation. Changes in glycogen metabolism were found to be due, in part, to changes in the fraction of cells involved in metabolism at any given time. High concentrations of glucose stimulated the cells to deposit glycogen but the response was reduced the longer the cells were in culture over a 3-day period. This loss of glycogen synthesizing capacity appears to be due to a decrease in glycogen synthetase activity. The activities of pyruvate kinase, hexokinase and aldolase also decrease during the culture period.


A serum factor capable of stimulating hyaluronic acid synthesis in cultured rat fibroblasts. Calf serum as well as rat and mouse sera has a factor that stimulates hyaluronic acid synthesis in cultured rat fibroblasts. Such a factor was partially purified from calf serum and characterized. It has a molecular weight of approximately 150,000. The activity of the factor is lost by treatment with pronase and by periodate oxidation. It is suggested, therefore, that the factor is a glycoprotein. Its susceptibility to alpha-mannosidase and affinity for Con A-Sepharose may suggest that the factor contains a mannose residue(s) which is essential for the activity to induce hyaluronic acid synthesis.


Effects of hormones and serum on glycogen metabolism in adult rat liver parenchymal cell primary cultures. Parenchymal cells from adult rat liver, isolated by a collagenase perfusion technique, have been maintained in primary culture and a detailed study on carbohydrate metabolism carried out over the initial 48-hour culture period. The glucose concentration of the medium exerts a major influence on glycogen accumulation by the cells. Insulin, particularly at high glucose concentrations, stimulates glycogen biosynthesis, whereas glucagon prevents glycogen accumulation. Dexamethasone was without effect on glycogen  metabolism. Glucose appears to stimulate glycogen accumulation by activation of glycogen synthetase enzyme. However, there is a gradual loss of synthetase activity throughout the culture period. Similar decreases in activity were noted for pyruvate kinase, aldolase and hexokinase. Glucose, insulin and dexamethasone were unable to prevent these decreases in enzyme activity. Foetal bovine serum contains fructose and this hexose appears to be the factor in serum which is responsible for the activation of glycogen accumulation in the presence of physiological glucose concentrations. The lactic acid content of the serum may also stimulate glycogen accumulation. In general, there is a gradual loss of the pattern of carbohydrate metabolism typical of differentiated hepatocytes during the culture period.


Measurements of contraction latencies to mechanical and electrical stimulation of the protozoan, Spirostomum ambiguum. Measurements made on contraction latencies in Spirostomun suggest that mechanical stimulation causes contractions to be initiated by the release of small amounts of calcium from a store tightly coupled to the contractile apparatus. Contraction to electrical stimulation appears to result from the gross electrophoretic mobilization of large amounts of calcium from a loosely coupled store. Contraction latencies to mechanical stimulation were three milliseconds and were independent of stimulus strength, previous stimulation, and contraction probability. For 0.5-millisecond biphasic electrical stimulation the contraction latencies varied widely. Latencies to initial contractions were dependent on stimulus strength: from 1.0 milliseconds for a stimulus that caused a 100% probability of contraction to 2.0 milliseconds for a stimulus that caused a 10% probability of contraction. Latencies of contraction to electrical stimulation were also dependent upon previous stimulation, lengthening to over 300 milliseconds after ten minutes of stimulation. Initial contraction latencies were not affected by previous stimulation to the other (electrical or mechanical) stimulus modality. Repeated electrical stimulation also reduced the animal's resting length and slowed the rate of post contraction re-extension, whereas mechanical stimulation did not have these effects.


Electron microbeam analysis of calcium distribution in the ciliated protozoan, Spirostomum ambiguum. Electron microprobe analyses of calcium distribution in the ciliated protozoan, Spirostomum ambiguum, indicated several calcium rich sites. One site was an endoplasmic distribution of calcium coincident with phosphorus which corroborates previous findings of hydroxyapatite deposits within Spirostomum. These apatite deposits were distributed throughout the endoplasm, but not within the nuclei or the contractile vacuole. Calcium was also detected within the cortical region. Cortical calcium was in greater concentration in the anterior portion of the organism and decreased towards the posterior end (region containing the contractile vacuole). Phosphorus and potassium were also detected as gradients from the anterior end, whereas magnesium was detected in the same density throughout the cortical region. Line scans of cortical regions suggested (1) distributions of calcium within mitochondria and/or vesicles, and (2) calcium associated with bundles of microfilaments.


In vitro inhibition of chick embryo lysyl hydroxylase by homogentisic acid. A proposed connective tissue defect in alkaptonuria. Homogentisic acid inhibits the in vitro activity of chick embryo lysyl hydroxylase, a microsomal enzyme which catalyzes the transformation of certain lysyl residues in collagen to hydroxylysine. Chick embryo lysyl hydroxylase activity was measured as specific tritium release as tritium water from a [4,5-(3)H]lysine-labeled unhydroxylated collagen substrate prepared from chick calvaria. Kinetic studies revealed a linear, noncompetitive type of inhibition with respect to collagen substrate with a Ki of 120-180 muM. The inhibition by homogentisic acid was reversible in that enzyme activity could be restored after dialysis of preincubated mixtures of homogentisic acid with enzyme or substrate. The inhibition by homogentisic acid was competitive with respect to ascorbic acid, and the addition of reducing agents, such as ascorbic acid or 1,4-dithiothreitol, protected lysyl hydroxylase activity from homogentisic acid inhibition.In organ cultures of embryonic chick calvaria, biosynthesis of hydroxylysine-derived intermolecular collagen cross-links was inhibited in a dose-dependent manner by 0.5-5 mM homogentisic acid. Because homogentisic acid inhibits the formation of hydroxylysine in a cell-free assay and in organ cultures, this compound must pass into the cells of calvaria to inhibit intracellular hydroxylysine formation and subsequently to diminish the reducible intermolecular cross-links of the newly synthesized collagen. We propose that the inhibition of lysyl hydroxylase and the resulting hydroxylsine-deficient, structurally modified collagen may be clinically significant in the defective connective tissue found in alkaptonuric patients.


Serum folates in man.

Serum folates in man. In an aseptic microbiological assay of folate compounds and their breakdown compounds, using Lactobacillus casei, Streptococcus faecalis, and Pediococcus cerevisiae, 4a-hydroxy-5methyl-4,5,6,7-tetrahydrofolate and 5-methyl-5,8-dihydrofolate were inactive under all conditions to all three organisms and 5-methyl-5,6-dihydrofolate was inactive unless ascorbate was present in the incubation medium, and then only to L. casei. 5-Methyltetrahydrofolate was active only for L. casei, and activity in purified samples to S. faecalis was due to trace amounts of folic acid. Analysis of S. faecalis values in the serum in normal subjects and in patients with various disorders showed that levels of 10-formyltetrahydrofolate are raised in coeliac disease, leukaemia, rheumatoid arthritis, and schizophrenia. 5-Methyltetrahydrofolate is readily absorbed by normal human subjects and by patients with pernicious anaemia but poorly absorbed by patients with coeliac disease or leukaemia. 5-Methyl-5,6-dihydrofolate was quickly absorbed by normal human subjects, being reflected by a considerably raised level of 5-methyltetrahydrofolate in serum when sodium bicarbonate was given by mouth before the 5-methyl-5,6-dihydrofolate. These higher levels were comparable to those in patients with pernicious anaemia after oral administration of 5-methyl-5,6-dihydrofolate. Oral 5-methyl-5,8-dihydrofolate and 4a-hydroxy-5-methyl-tetrahydrofolate did not appear as microbiologically active folates in the serum. The findings of this study suggest that the availability for biological utilisation of the major dietary folate compounds will depend on the amount of gastric acidity and of ascorbate in the intestinal chyme. Many may be unavailable for metabolic utilization in the body.


Evaluation of a commercial radioisotopic kit for folate assays. A commercial kit for the radioisotopic assay of folate in serum, the Bio-Rad 'Quanta Count' folate kit, produced lower results than the Lactobacillus casei microbiological assay method. Its normal range was 2-0-13-0 microng/l and the reproducibility was similar to that of the microbiological assay method. The kit was also satisfactory for whole bood folate assays. The cost requires careful consideration before the kit is used for routine purposes.


Reverse carotid blood flow--a possible explanation for some reactions to local anesthetics. In  rhesus monkeys, data obtained by contrast radiography and hemodynamic and electroencephalographic studies indicate that carotid blood flow is reversible. Results showed that even small amounts of local anesthetic agents when injected inadvertently into a branch of the external carotid artery, may enter the cerebral circulation, most likely through a retrograde flow into the common and then internal carotid arteries. Some toxic neurologic manifestations possibly may be explained by this mechanism.


Sodium cromoglycate in ragweed-allergic conjunctivitis. Sodium cromoglycate (SCG), in a 4% solution instilled into each eye 4 times daily, was compared with placebo in a double-blind, noncrossover trial in 30 matched patients with troublesome ragweed pollen-induced conjunctivitis. In the SCG group, eye symptom scores were significantly less (p = 0.05), and all patients judged that their symptoms were improved over the previous year (p less than 0.05). Less antihistamine was used by the SCG group but the difference was not significant. It was concluded that SCG was effective in the treatment of ragweed-induced seasonal allergic conjunctivitis.


Identifying needs and services for the aged. The Assessment and Placement Service (APS) of the Hamilton-Wentworth District Health Council assists health professionals in identifying the needs of persons with chronic disabling diseases, and in finding appropriate resources or programs. More than 80 percent of the 9,300 persons referred between September 1971 and December 1975 were over the age of 64. Although most referrals were local, some came as far away as British Columbia and Florida for persons wishing placement locally. Of the 2,005 persons referred and placed in 1975, 1,063 came from acute disease hospitals, 201 from other institutions, 628 from the community within this District, and the remainder from elsewhere. There were 1,187 women, of whom 89.5 per cent were over the age of 64, and 811 men, of whom 65.8 per cent were over that age. In the opinion of their physicians, only 20 percent of all persons referred had normal memory, and only 13 percent had normal judgment. In the 64+ age group, 71.1 percent of the women were widows, and 25.9 percent of the men were widowers. One-third of those referred were living alone, and one-fourth with a spouse only. For placement, 453 patients went to private residences or boarding homes, often with the support of community services, and the remainder went to various institutions according to identified needs. Follow-up contact one month later showed conditions to be satisfactory in most instances. Deficiencies in the health care system were identified and reported to the Health Council.


Acute drug testing with nitrates: angina relief during treadmill exercise. To evaluate the effect of sublingual nitroglycerin (N) and chewable isosorbide dinitrate (I) in patients with angina pectoris, 12 subjects underwent treadmill exercise to the point of onset of angina. At that juncture, N or I or placebo (P) was administered and walking was continued until moderately severe angina recurred. For those patients who completed the initial 10-minute exercise period, testing was repeated until moderately severe angina recurred or until a total test time of three hours elapsed. Retesting ensured that each patient was tested separately with N, I and P within a two-week period. Within the I group, four patients experienced the onset of angina relief (AR) at 10 minutes (') 6 seconds (*) +/- 3' 4" with a mean recurrence time (RT) of 154' 3" +/- 4' 27" (P less than or equal to 0.05), for a significant difference. Within the N group, six patients had AR of 8' 3" +/- 1' 1" with an RT of 49' 39" +/- 1' 15" (P less than or equal to 0.01), for a significant effect. Within the P group, only one patient experienced AR. For all other patients in each group, testing was terminated because of moderately severe angina in the 5-8' period of the first test. In comparing subgroups for RT during I therapy (4 patients) versus N therapy (6 patients), a significant difference was noted (154' 3" +/- 27" versus 49' 39" +/- 1' 15") (P less than 0.05). The data suggest a possible benefit of I in selected patients. The effect may last as long as three to four hours. There is also an enhanced benefit of N with the patient in the upright position. This effect may last as long as 40-50'. Treadmill testing is recommended with nitrate administration to determine which drug (I or N) is the more suitable for the specific patient.


The influence of physical factors on the survival and infectivity of miracidia of Schistosoma manosni and S. haematobium I. Effect of temperature and ultra-violet light. The influence of temperature and ultra-violet radiation on the degree of activity, survival and infectivity of schistosome miracidia is profound. Miracidia of Schistosoma mansoni and S. haematobium were affect eaually. Only miracidia classified as "active" or "slow" were capable of penetration, a capacity they retained for about 17 hours at 19 degrees C. Miracidia that were "lethargic" as a result of low temperature, old age or ultra-violet radiation lost their infective capacity. The conclusion, however, is that neither the temperatures encountered in the field nor the solar ultra-violet radiation penetrating turbid waters are likely to be harmful to miracidia and thus have no effect on the level of transmission.


An investigation of the bacteriological quality of retail vanilla slices. One hundred and thiry-three vanilla slices, purchased from shops in the West Yorkshire Metropolitan County, were examined to determine the numbers and types of bacteria present at the time of purchase. The surface colony count at 37 degrees C was greater than 10(3)/g in 67/133 (50%) of the samples examined, Bacillus cereus being found at that concentration in 21.8%, coliform bacilli including E. coli in 5.3%, Staphylococcus aureus in 3-0% and Streptococcus faecalis in 0-8%. Thirty-four strains of B. cereus were serotyped and 11 (32%) of these were typable with the sera available. Preparation of custard mixed in the laboratory suggests that the milk or milk powder used in the mix may be the major source of B. cereus in the final product. Many of the present methods of manufacture, distribution and storage allow organisms present in the custard at manufacture the opportunity to multiply and possibly reach numbers which present a risk of food poisoning.


Acute otitis externa in divers working in the North Sea: a microbiological survey of seven saturation dives. Saturation diving is an important and widely used technique in the Offshore Oil Industry. During 1974-5 two saturation dives in the North Sea were terminated because of outbreaks of incapacitating otitis externa, and others were disrupted. Pseudomonas aeruginosa was consistently isolated from the ears of affected divers. Because complex work schedules were threatened seven subsequent dives were subjected to microbiological monitoring and control. Colonization of ear canal with P. aeruginosa or with other gram-negative bacilli occurred in 39 (67%) of the 58 divers studied, usually within 7 days of starting the dive. Data obtained by serotyping this isolations of P. aeruginosa suggested that a single infected diver may be the source of organisms which rapidly spread to his colleagues and throughout the living chambers, that the living chambers may constitute a reservoir of infection during and between dives, and that certain serotypes of P. aeruginosa are more likely than others to colonize the ear canal in the conditions of a saturation dive. The control measures used during the dives were only partially effective, but none of the divers suffered severe pain and all the dives were an operational success.


Allelic expression and genetic distance in hybrid macaque monkeys. Levels of structural genic divergence at 21 loci encoding blood proteins were quantified in six macaque (Macaca) species, using standard techniques of starch-gel electrophoresis. Genetic distances between all pairs of species fall within a narrow range (0.080 less than or equal to D less than or equal to 0.250; D = 0.164) which is near the lower limit of genetic distances typically observed between other congeneric organisms. In an effort to measure levels of regulatory gene differences between these species, we have examined the patterns of allelic expression in their F1, F2, and backcross hybrids. Nine of the 21 loci examined encode allelic forms of the proteins with different electrophoretic mobilities in at least some of the individual parents of the hybrids. In all cases where expected, hybrids express fully both maternal and paternal allelic products, thus providing no strong evidence ofr a breakdown in the regulatory mechanisms responsible for proper expression of these genes. Results are compared to degrees of allelic repression previously observed in other hybrids, and are discussed within the context of current ideas about rates of regulatory gene evolution in mammals.


Complete sharing of light chain spectrotypes by murine IgM and IgG anti-streptococcal antibodies. In order to examine the diversity of antibody light chains, we have developed an analytic isoelectric focusing procedure which permits the routine analysis of L chains from antibodies raised in individual mice. We have used this technique to demonstrate that the light chains of IgM and IgG anti-group A streptococcal antibodies raised in SWR mice are probably shared. Interestingly, numerous light chain spectrotypes are shared between individual mice whose 7S antibody focusing patterns differ.


ELIEDA (enzyme-linked-immuno-electro-diffusion-assay): application of a combined immunoelectrodiffusion and immunoenzyme method to the study of immune response in parasitic infections. The sensitivity of the technique of immunoelectrodiffusion (IED) has been enhanced by combining it with an enzyme method; this also enabled the class of immunoglobulins involved in the reactions to be determined by treating the precipitated immune complexes with enzyme-labelled class-specific antisera. The method has been employed in studies of the immunology of parasitic infections and these confirmed its sensitivity and specificity. ELIEDA provides a valuable tool for analysing responses to complex immunogens.


Methods for detection of anti-albumin autoantibodies in hepatic diseases. Three methods for detection and quantitation of anti-albumin autoantibodies (AAA) in human sera have been developed based on their reaction with a new antigenic site of glutaraldehyde-polymerized human serum albumin (HSA). The first is a double diffusion technique (ID-HSA) were the sera to be tested are reacted with a copolymer of glutaraldehyde  activated HSA (copoly-HSA). The reaction is estimated by the lowest copoly HSA concentration giving precipitation with the undiluted sera. The second method is based on agglutination (HA) by AAA positive sera of human red blood cells coated with glutaraldehyde-polymerized HSA (EPA). The reaction is quantified as the highest dilution of sera giving agglutination. The third method uses glutaraldehyde insolubilized HSA as an immunoadorbent able to bind AAA in positive sera. The quantity of protein adsorbed, expessed in microng antibody/ml serum, measures the level of AAA in sera. Significant correlation between the values of AAA as determined by all three methods presented above were recorded.