The primary structure of the major cytoplasmic valine tRNA of mouse myeloma cells. This paper describes the derivation of the primary structure of the major valine tRNA in the cytoplasm of mouse myeloma cells. Approximately 75% of the nucleotide sequence of this tRNA is also shared by the tRNA1-Val of yeast, this homology serving as a further indication of the extreme conservation of the structures of the tRNAs of different eukaryotic organisms. A novel feature of mouse myeloma tRNA1-Val is its loop IV sequence: -U-PSI-C-G-M1A-A-A-. This particular loop IV sequence has not previously been found in a tRNA structure. In addition, tRNA1-Val possesses some unusual nucleoside modifications. 5-Methyluridine (T) was not found to occur within loop IV of this tRNA, although this minor nucleoside is also absent from certain other mammalian tRNAs. Only one other tRNA, mammalian tRNAf-Met, has been found to possess 2-methylguanosine (m2G) in the position between the (b) and (c) stems of the cloverleaf. Numerous tRNAs have m2-2G in this location, and it would appear that the second methylation of this guanosine is characteristically absent from certain mammalian tRNA species.


The influence of histocompatibility and antithymocyte serum on the ability of alloantigenic pretreatment to prolong the survival of mouse skin grafts. Test skin allografts enjoy prolonged survival when alloantigenic pretreatment (implants of small fragments of liver or kidney on the host's kidney cortex) is separated from test grafting by 9 days and donor and host strains differ only at the relatively weak H-3 + H-13 histocompatibility barrier (Baldwin and Cohen 1971, 1974). However, test skin grafts are rejected in an accelerated fashion when this protocol is followed but the donor strains differ at stronger non-H-2 barriers. In 2 such strain combinations in which control first-set grafts are rejected acutely, varying the implant-to-test graft interval, using F-1 mice as implant donors, or varying the sex of the donors and hosts, all failed to prolong test graft SURVIVAL. However, survival of some test grafts was significantly greater following a combined host treatment of alloimplanatation and ATS than it was following either procedure alone. This synergism suggests that across stronger H-barriers, alloantigenic pretreatment does elicit an immunosuppressive component whose effective presence is normally masked by thymus-dependent cell-mediated immunity.


[The ideal computer for ECG analysis (author's transl)]. An on-line minicomputer with a suitable structure turns out to be the ideal system for ECG interpretation. The motivations for this conclusion come both from the nature of the ECG analysis process and from the solution that the present state of the process and from the solution that the present state of the computer technology suggests for the controversy on large-scale computers versus minicomputers in the medical environment. In this paper both these aspects of the problem are briefly discussed and an ideal structure is proposed for a system based on a minicomputer. The main goals of this system are flexibility, adequate man-machine interfacing, low cost, high reliability and minimal maintenance, minimum installation space and site preparation.


Research approaches in illicit drug use: a critical review. Research approached and substantive findings pertaining to determinants of the nonmedical use of illicit psychoactive drugs are reviewed. Discussion focuses upon the dimensions of drug use as a behaviorial phenomenon, the findings and limitations of four basic approaches to research in the ares--sociocultural, psychopathological, functionalist, and social learning-- and upon methodological problems. Drug use must be treated as a multidimensional phenomenon, with analyses extending well beyond simple user vs. nonuser comparisons or arbitrary categories of users. Research must be conducted within a coherent field-theoretical conceptual framework, directed toward the formulation of nomothetic laws concerning patterns of drug use. Inadequacies in sampling and measurement and the absence of longitudinal design have compromised the utility of a vast research literature.


The effect of wheat bran on intestinal transit. In 18 students and two members of staff at a boys' boarding school, the time taken to pass 20 out of 25 radiopaque pellets varied from one to seven days while the subjects were eating a normal English diet. After the additon of bran, about 20 g daily, to this diet transit time fell from 2-75 plus or minus 1-6 to 2-0 plus or minus 0-9 days (P smaller than 0-025). Transit became faster in all nine subjects who had an initial time of three days or more, and in three of seven with an initial time of two days, but became slower in all four boys with an initial one-day transit. Frequency of defaecation correlated poorly with transit time (a once daily bowel action being found with transit times ranging from one to four days), and did not increase significantly with bran. In 10 additional adults with slow initial transit (three or four days) the effect of bran was compared with that of an equal volume of ground oatflakes in a double-blind crossover trial. Bran caused a significant acceleration of transit, wherease oatmeal had no effect. These studies confirm that bran accelerates slow intestinal transit and show that this is not simply a psychological effect. Bran may also slow down fast transit.


Complete identification of endocrine cells in the gastrointestinal tract using semithin-thin sections to identify motilin cells in human and animal intestine. The present (Wiesbaden) classification of gut endocrine cells relies mainly on ultrastructure and the results of silver impregnation. Correlation with hormone production requires parallel immunology but conventional immune cytochemistry is, in most cases, difficult or at present impossible. The serial, semithin-thin-section technique offers an alternative, which provides absolute correlation between cell type and hormone production. The successful use of the technique is illustrated here by localization of motilin in the enterochromaffin cells of the small intestine.


Separation of T lymphocytes from normal individuals and patients with B lymphocyte chronic lymphocytic leukaemia. Previous studies have shown that lymphocytes from patients with chronic lymphocytic leukaemia have a diminished response to mitogens which stimulate T cells. Chronic lymphocytic leukaemia is most often a disease of accumulating B cells so that T lymphocytes are diluted by large numbers of leukaemic cells. Direct comparison with the responses of normal lymphocytes to mitogenic stimulation is therefore suspect. To circumvent this difficulty, a method of isolating T cells from normal individuals and patients with chronic lymphocytic leukaemia was developed. Lymphocytes containing an average of 16.1 per cent B cells from normal individuals were applied to IgG-anti-IgG-coated Degalan bead columns and held at 4 degrees for 2 hours. The eluted cells contained less than 2 per cent B cells. When chronic lymphocytic leukaemic lymphocytes, containing an average of 68.6 per cent B cells, were applied to IgG-anti-IgG columns, the eluted cells contained 36.4 per cent B cells. To improve the purification of T lymphocytes, columns of uncoated Degalan beads were used to remove non-specifically adherent cells. Eluted lymphocytes were then applied to IgG-anti-IgG columns. This resulted in the recovery of purified populations of T cells with less than 2 per cent contamination with B cells. Patients with chronic lymphocytic leukaemia were found to have lymphocytes with either a normal density or a low density of surface immunoglobulins. B cells were successfully removed from lymphocyte suspensions in all cases of chronic lymphocytic leukaemia with a normal density of lymphocyte surface immunoglobulins. In the three cases of chronic lymphocytic leukaemia with low density surface immunoglobulins, separation by this method was unsuccessful. However, an enriched T-cell population was obtained when leukaemic lymphocytes which had lost all detectable surface immunoglobulins were passed through a column coated with heat-aggregated IgG.


Thymus dependence of the IgA response to sheep erythrocytes. The thumus dependence of the IgA response to sheep red blooc cells (SRBC) was studied by means of cell transfer experiments in mice. Only low numbers of IgM-, IgG- and IgA-plaque-forming cells (PFC) were observed in those recipients which received only spleen cells from adult thymectomized, irradiated and bone marrow-reconstituted mice (B cells). High numbers of IgM-, IgG- and IgA-PFC were observed when B cells and educated T cells were transferred to the recipients. Evidence is provided that B cells committed to IgA synthesis require the same degree of interaction with T cells as B cells committed to IgM synthesis, but a lower degree of interaction with T cells than B cells committed to IgG synthesis.


The possible presence of a bursa-independent, IgM-producing system in chicks. The number of splenic plaque-forming cells significantly increased in bursectomized chicks immunized secondarily with SRBC. Further experiments using rabbit antiserum specific to the bursa of Fabricius indicate that the different types of antibody-producing cells might be engaged in the synthesis of IgM in bursectomized chicks. The possible presence of a bursa-independent, IgM-producing system in chicks is suggested.


Growth of several cariogenic strains of oral streptococci in a chemically defined medium. A chemically defined medium in which Streptococcus mutans strains AHT, BHT, GS-5, JC-2, Ingbritt, At6T, At9T, 6715, and OMZ-176 and Streptococcus salivarius strain HHT grew rapidly to high turbidities was formulated. Maximal turbidities of each strain were observed after 8 to 12 h of aerobic growth. The subsequent transfer of exponentially growing cells into fresh medium resulted in growth at the same rate without lag. Growth of these strains occurred with rates at least one-half of those observed in an organic medium, such as Todd-Hewitt broth. S. mutans strains FA-1 and OMZ-61 grew at relatively slow rates in the defined medium, but more rapidly growth to higher turbidities of both strains was obtained when sodium bicarbonate was added to the medium. Streptococcus sanguis strain OMZ-9 and another group H streptococcus (strain 72 times 46) grew rapidly in the defined medium after the addition of sodium carbonate. The presence of carbonate or bicarbonate yielded higher turbidities of all the other strains, and the growth rates of several of the strains tested were also increased.


Amino acid requirements of Streptococcus mutans and other oral streptococci. The amino acid requirements of Streptococcus mutans strains AHT, OMZ-61, FA-1, BHT, GS-5, JC-2, Ingbritt, At6T, OMZ-176, 6715, Streptococcus salivarius HHT, Streptococcus sanguis OMZ-9, and strain 72x46 were determined in a chemically defined medium. When grown anaerobically in the presence of sodium carbonate (or bicarbonate for a few strains), few amino acids were required. All strains tested required cystine (or cystine) as a nutrient. Three strains (S. mutans OMZ-176, FA-1, and BHT) required glutamate (and/or glutamine). A third amino acid (lysine for S. mutans FA-1 and histidine for S. mutans OMZ-176) was required by two of the three strains which required glutamate. The amino acids mentioned above were required for all conditions of incubation (and inoculum) tested. The requirements for several other amino acids were conditional, that is, dependent on the incubation conditions and inoculum used. For example, when carbonate was not added, glutamate was required by S. mutans GS-5. Aerobic incubations, with carbonate or bicarbonate added, resluted in requirements for glutamate and leucine by several strains. With these incubation conditions, one strain required isoleucine (S. mutans FA-1), another valine (S. mutans AHT), and a third tyrosine (72x46). Aerobic incubations in the absence of carbonate or bicarbonate further increased the number of amino acids required by several strains. Furthermore, when stationary-phase cultures replaced exponentially growing cultures as an inoculum, several strains required additional amino acids, presumably for the initiation of growth.


Phage conversion of Shigella flexneri group antigens. A temperate phage, designated Sf6, has been isolated from Shigella flexneri 3a. Characterization of Sf6 revealed that it possesses the capacity for converting the S. flexneri 3,4 group antigen complex to group factor 6. Serological studies and chemical analysis of lipopolysaccharide from converted strains suggest that group factor 6 is a reflection of an acetylation of the preexisting 3,4 antigen complex. Evidence is provided that the 3,4 group antigen complex functions, at least in part, as a cell surface receptor site for Sf6 adsorption.


Neutralization of Chlamydia trachomatis in cell culture. Neutralization of Chlamydia trachomatis was assayed by the decrease in inclusion-forming units in baby hamster kidney cells grown in culture. Five percent fresh guinea pig sera increased neutralization titers of rabbit antisera 100- to 1,000-fold but had no effect when normal rabbit sera were tested. Neutralization of a type A or B trachoma isolate was strain specific. Neutralization by human eye secretions and sera also was demonstrated when guinea pig sera were included in the test. All of the six human sera tested showed strain specificity against types A or B, in agreement with typing by the fluorescent antibody technique.


Predominant cultivable flora isolated from human root surface caries plaque. Plaque samples were obtained from tooth surfaces exhibiting typical lesions of root surface caries and were immediately cultured by a continuous anaerobic procedure. The bacterial composition of root caries flora was determined on individual samples. Representative isolates from each specimen were characterized by morphological and physiological criteria. In addition, fluorescent antibody reagents were used to confirm the identification of Streptococcus mutans and Actinomyces viscosus. The plaque samples could be divided into two groups on the basis of the presence or absence of S. mutans in the plaque. In group I plaques, S. mutans comprised 30 percent of the total cultivable flora. S. sanguis was either not found or was present in very low number. In group II plaques, S. mutans was not detected, and S. sanguis formed 48 percent of the total plaque flora. A. viscosus was the dominant organism in all plaque samples, accounting for 47 percent of the group I isolates and 41 percent of the group II isolates.


Evidence for the non-protein nature of the receptor for the enterotoxin in Vibrio cholerae on murine lymphoid cells. Lymphoid cells from A/J and BALB/c strains of mice were iodinated with 125I by the lactoperoxidase method and the plasma membranes were disrupted by freezing and thawing or with 0.5 percent Nonidet P-40, a nonionic detergent. Attempts to find choleragen reactive iodinated material in 0.5 percent Nonidet P-40 lysates were unsuccessful even when the cells were incubated with choleragen before lysis. Freezing and thawing the cells resulted in the release of iodinated choleragen reactive material. The interaction of choleragen with the iodinated material could be inhibited (at low choleragen concentrations) or enhanced (at high choleragen concentrations) by the addition of the ganglioside G-M1 to the the immune precipitation system. The results are consistent with the hypothesis that the receptor for choleragen is a glycolipid and reduce, but do not totally eliminate, the likelihood that the receptor is glycoprotein in nature.


Effects of human and rabbit serum on viability, permeability, and envelope lipids of Serratia marcescens. The major action of serum on gram-negative organisms is thought to be on the microbial envelope. We compared the effects of normal human and rabbit serum on the envelope lipids of two strains of Serratia marcescens, one sensitive and one resistant to the bactericidal effects of serum. During killing by either serum, the sensitive strain underwent rapid permeability changes coincident with degradation of microbial phospholipids. The resistant strain exhibited none of these effects. The phospholipid degradation that accompanies killing of the sensitive strain by serum could be caused by phospholipases present in serum or by Serratia's own phospholipid-splitting enzymes. The results indicate that phospholipid breakdown is caused by activation of bacterial of bacterial phospholipases and not by serum phospholipases. This conclusion is based upon the following findings.(i1 Although rabbit serum phospholipase A was at least 10 times more active than human serum phospholipase A, phospholipid degradation in the sensitive Serratia strain was comparable during (equally rapid) killing by human or rabbit serum. (ii) Heat treatment (56 C) of both sera eliminated bactericidal activity as well as microbial lipid degradation but abolished phospholipase activity of human serum only. (iii) Virtually complete removal of phospholipase A activity from human serum by adsorption onto autoclaved Micrococcus lysodeikticus had no effect on the extent of phospholipid hydrolysis or on bactericidal activity. Activation by serum of endogenous phospholipase activity in S. marcescens was accompanied by enhanced incorporation of lipid precursors into bacterial lipids. No evidence was found for increased turnover of protein or ribonucleic acid during killing by serum.


Persistence of rubellavirus-specific immunoglobulin M and immunoglobulin A antibodies: investigation of successive serum samples with lowered immunoglobulin G concentration. The persistence of rubellavirus-specific immunoglobulin (Ig) M and IgA antibodies has been studied in seven patients with primary postnatal rubella infections. Successive blood samples obtained over a period of several years after the onset of disease have been investigated, employing the fluorescent antibody technique and the sucrose gradient centrifugation method. IgM antibodies were found to persist for 4 to 5 weeks after the onset of disease, with only moderate variation being observed with respect to the different patients and the method of investigation being studied. The persistence of IgA antibodies (as examined by the fluorescent antibody technique) varied from a few weeks to several years after the onset of the disease. The detection of IgA antibodies cannot be considered as conclusive for the diagnosis of recent rubella infections.


Virologic and serologic studies of zoo birds for Marek's disease virus infection. One hundred and eleven zoo birds representing 49 species in 14 orders were examined for Marek's disease (MD) herpesvirus (MDHV) infection. MDHV was isolated from 10 birds, all belonging to genus Gallus. The precipitating antibodies against MDHV were demonstrated only in the Gallus birds, when 51 selected birds including 34 Galliformes and 17 other birds representing 12 species from nine orders were examined. The 10 MDHV isolates all induced morphologically similar plaques in cell cultures closely resembling those of HN strain, a low pathogenic isolate of MDHV. Six of the 10 isolates, when inoculated into an experimental line of chickens highly susceptible to MD, caused only a minimal degree of histologic lesions without causing clinical MD, gross MD lesions, or deaths from MD. Natural hosts of MD are probably Galliformes, primarily affecting Gallus and less often other genera of Galliformes.


Detection of a leukocytic endogenous mediator-like mediator of serum amino acid and zinc depression during various infectious illnesses. Serum samples from patients with many different kinds of infection were shown to contain a substance characteristic of leukocytic endogenous mediator. Generalized bacterial infections produced more leukocytic endogenous mediator than did localized ones; viral infections produced very little. In typhoid fever, mediator concentrations increased before the onset of illness.


Significane of intravascular coagulation in canine endotoxin shock. The contribution of disseminated fibrin clot formation to the pathogenesis of canine endotoxin shock was explored in control dogs and in those defibrinated with a purified fraction of Malayan pit viper venom. The hemodynamic and humoral responses after the administration of an intravenous challenge dose of Escherichia coli endotoxin were comparable as was mortality. It is concluded that, although the role of the coagulation sequence in canine endotoxin shock is unclear, it does not appear to be determinative.


Factors affecting filamentation in Candida albicans: relationship of the uptake and distribution of proline to morphogenesis. When glucose was present in high concentration, Candida albicans formed filaments in a phosphate-buffered medium, regardless of the nitrogen source. In lower concentrations of glucose, filamentation occurred only when various members of the glutamate, succinyl, or acetoacetyl-coenzyme A families of amino acids were used as sole nitrogen sources. Yeast morphology could be maintained either by replacing the amino acids in the medium with ammonium chloride or by making the medium high in phosphate or biotin. Studies using [U-14C]proline indicated that proline was catabolized in a manner consistent with the generation of increased cellular reducing potential and that the proline label entered into the Kreb's cycle. A reduction in Kreb's cycle activity was evidenced by an initial increase and then a rapid drop of the total organic acid content of the cells as well as in specific Kreb's cycle intermediates. Filamentation under conditions of low phosphate, high glucose, and increased cellular reduction potential, accompanied by a decrease in Kreb's cycle activity, suggests that morphogenesis in C. albicans is correlated with a Crabtree-like effect, i.e., repression of mitochondrial activity.


Immunodepression by Rowson-Parr virus in mice: effect of Rowson-Parr virus and Friend leukemia complex infections on background antibody-forming cells to various erythrocytes. The numbers of background antibody-forming cells (BPFC) toward erythrocytes of various species present in the lymphoid organs of unimmunized susceptible BALB/c and resistant C57BL/6 mice were investigated at various times after infection with Friend leukemia complex (FLC) or Rowson-Parr virus (RPV). Both virus preparations induced an increase of BPFC numbers in both animal strains, but the rate and magnitude of the enhancements produced by RPV were much lower. The degree of potentiation varied with the specificity of the BPFC populations and was more pronounced in the spleen than in the lymph nodes and in BALB/c than in C57BL/6 mice. In the late stage of FLC infection, the numbers of splenic BPFC to some erythrocytes underwent a dramatic fall, which was not observed in RPV-infected mice. BPFC present in BALB/c splenocytes cultured in diffusion chambers implanted in the peritoneal cavity of isogeneic normal mice were not affected by viral infection of the chambers.


Heteroimmunization to the capsular polysaccharide of Haemophilus influenzae type b induced by enteric cross-reacting bacteria. Cross-reacting Escherichia coli strains Easter and 89 and Bacillus pumilis fed to newborn rabbits and E. coli fed to adult rhesus monkeys did not exert untoward reactions. The E. coli regularly colonized the newborns' intestinal tract from 1 to 7 weeks. High doses of E. coli were necessary to colonize adult primates. Colonization occurred in fewer newborn rabbits and lasted only 1 to 3 weeks with B. pumilis. Colonized newborn rabbits and adult rhesus had an active Haemophilus influenzae type b (HITB) immune response. In the rabbit, colonization resulted in accelerated induction of immunoglobulin (Ig) M-. IgA-, and IgG-producing cells in the spleen, mesenteric lymph nodes, and Peyer's patches after HITB challenge. E. coli-fed and control newborn primates were naturally colonized with nasopharyngeal and enteric cross-reacting bacteria and both groups rapidly developed HITB antibodies in the absence of the homologous organisms. Human newborn stool cultures, taken at the time of discharge from the nursery, showed a 0.9% carriage rate for cross-reacting E. coli. These "carrier" infants acquired HITB antibodies more rapidly than their age-matched "noncarrier" controls.


Effects of canine distemper virus infection on lymphoid function in vitro and in vivo. In the present study, the immunodepressive effects of canine distemper virus (CDV) infection of dogs on two parameters of lymphocyte function, namely phytomitogen-induced cellular proliferation and skin allograft rejection, were investigated. Infection of susceptible gnotobiotic dogs with virulent R252-CDV resulted in a depression of peripheral blood lymphocyte mitogen response as measured by (3H)thymidine incorporation for up to 10 weeks after inoculation. This effect coincided with the appearance of viral antigen by immunofluorescence in leukocytes but persisted after the virus was no longer detectable. Loss of mitogen reactivity was seen in all infected dogs. However, when these same CDV-infected dogs were challenged with foreign skin allografts, no significant retention of grafts over controls was observed despite the depressed lymphocyte activity. Considering the in vitro and in vivo data it was concluded that, although immunodepressive effects of CDV were demonstrated in vitro, paralled in vivo experiments indicated that less than complete suppression of immune functions occurs during the course of CDV infection.


Superinfection in mice previously infected with Mycobacterium leprae. Previous studies of the protection of mice by prior infection with Mycobacterium leprae in one hind footpad against challenge with M.leprae in the opposite hind footpad had produced conflicting results; therefore, the problem was restudied. In several experiments, BALB/c mice were inoculated first in the right hind footpad with 5,000 M. leprae and then challenged in the left hind footpad with 5,000 M. leprae of the same strain at intervals after primary infection, at the same time that uninfected mice were inoculated. Multiplication of the M. leprae of the secondary challenge inoculum occurred at the same rate and to the same level as multiplication in uninfected mice when challenges were made soon after primary infection. Multiplication was slowed but proceeded to the same level in previously infected as in uninfected mice when the challenges were administered between 76 and 106 days after primary infection (47 to 17 days before the M. leprae of the primary inoculum had multiplied to the level of 10-6 organisms per footpad). Finally, the M. leprae of a secondary challenge administered at the time that the organisms of the primary inoculum had multiplied to 10-6 per footpad or later not only multiplied more slowly in previously infected than in control animals, but multiplication in the previously infected animals reached a lower maximum. These results are similar to those observed when mice previously infected with M. bovis (BCG), M. marinum, Toxoplasma gondii, or Besnoitia jellisoni were challenged with M. leprae.


Treponema pallidum (Nichols strain) in tissue cultures: cellular attachment, entry, and survival. The interaction of Treponema pallidum (Nichols strain) with cultured cells was investigated under aerobic conditions. Cell monolayers derived from rabbit testicular tissue extended the survival of treponemes as indicated by active motility. Large numbers of organisms rapidly attached to cultured cells. Within 3 h, one to twelve actively motile treponemes were attached to 25 to 50% of the cells. In addition, T. pallidum attained intracellularity as early as 30 min after inoculation of the cell monolayers. In sharp contrast, T. phagedenis biotype Reiter and T. denticola did not attach and did not enter cultured cells. Most importantly, intracellular and/or attached T. pallidum retained virulence for at least 24 h. Similar observations of attachment and retention of virulence were detected with ME-180, a cell line derived from a human cervical carcinoma. Preliminary studies with superoxide dismutase indicated that this enzyme prolonged treponemal motility and retention of virulence in the presence of cultured cells. These data provide guidelines for further investigations of in vitro cultivation of T. pallidum.


Motility as a virulence factor for Vibrio cholerae. The ability of motile strains of the Ogawa and Inaba serotypes of classical Vibrio cholerae and of the El Tor biotypes to kill suckling mice after oral challenge with 10-8 colony-forming units (representing at least 100 to 1,000 minimal lethal doses) was compared to that of nonmotile derivatives of the same strains. Loss of motility, in each case, resulted in a marked reduction in virulence. The mortality (at 36 h) caused by 10 of the 13 nonmotile strains was 32% or less. whereas the motile wild-type strains resulted in nearly 100% deaths. The reduced virulence of the nonmotile strains was associated with reduced capacity to adsorb to the surface of segments of mouse intestine. The mutants were tested for alterations in enterotoxin production and surface properties. The results suggest that motility may contribute to virulence by increasing the chance for association of the vibrios with the intestinal mucosa.


Serotyping of Chlamydia: isolates of bovine origin. Chlamydial isolates of bovine origin were serotyped by a plaque reduction method. Of the two major serotypes observed, type 1 included isolates from bovine abortion and enteric infections, whereas type 2 isolates were associated with polyarthritis or encephalomyelitis. These two serotypes were identical to those with a similar disease distribution previously observed in isolates of ovine origin. The two groups did not cross-react and they were serologically unrelated to chlamydiae of avian origin. Thus, it appears that many chlamydial isolates causing intestinal infections or abortion in sheep or cattle are closely related antigenically, as are those producing polyarthritis, encephalomyelitis, and conjunctivitis, and that the two groups are distinct.


Inhibition of the steroidogenic effects of cholera and heat-labile Escherichia coli enterotoxins by GM1 ganglioside: evidence for a similar receptor site for the two toxins. The effects of three different ganglioside preparations on cholera enterotoxin (CT) and heat-labile Escherichia coli enterotoxin (ECT)-induced steroidogenesis in Y1 and OS3 adrenal tumor cells in tissue culture were examined. Only with GM1 ganglioside was any inhibition of the toxins' effects noted. Concentrations of the crude ECT preparation that gave similar morphogenic and steroidogenic effects as CT were inhibited by the same amount or less of GM1 as that required to inhibit the effects of CT. The results of competition experiments also demonstrated that previous incubation of GM1 with one toxin could inhibit the ganglioside's ability to inactivate the other toxin. These findings indicate that at least for Y1 and OS3 adrenal tumor cells, GM1 may resemble or be the receptor for both CT and ECT.


Detection of virus-specific antibody-forming cells of mice immunized with Newcastle disease virus. The hemolytic plaque assay was adapted to the detection of antibodies to Newcastle disease virus (NDV) in an in vivo, an in vitro system, and a combined in vivo-in vitro system. Several conditions were tested for coupling of sheep erythrocytes to NDV and for the kinetics of plaque formation in the in vivo and in vitro systems. The one set of conditions which provided the best responses is presented. The effect of multiple injections of NDV into mice on plaque formation was optimized.


On the question of permeability of the blood-brain barrier to botulinum toxin. The clinical symptoms of botulinum intoxication suggest that besides the involvement of the peripheral nervous system, the central nervous system is also affected. Studies were undertaken to determine whether pure toxin of Clostridium botulinum type A could be demonstrated in the brains of poisoned mice. With the aid of autoradiography of the toxin marked with 125-I and indirect fluorescent labeling it was possible to show the presence of the toxin in the parenchyma of the brain.


Infection versus disease in South American blastomycosis. The mechanism of infection in South American blastomycosis is discussed, and the probability of a mild or asymptomatic form of the disease, regressing spontaneously with subsequent immunity, in persons living in endemic areas. Primary lesions in the oral mucosa or in the skin have not been convincingly shown to occur in paracoccidioidomycosis. Several reports of systemic involvement of many organs with no evidence of skin and mucosal lesions support the theory of pulmonary entry.


The effects of rifampicin on electron- and neutron-irradiated E. coli B/r and BS-1: Survival, DNA degradation and DNA synthesis by membrane fragments. The effect of rifampicin after irradiation with electrons or neutrons on cell-killing, DNA degradation and the ability of DNA-membrane complex extracted from irradiated bacteria to synthesize DNA was studied in strains E. coli B/r and BS minus 1. Post-irradiation treatment with rifampicin greatly enhanced the lethal effects of both electrons and neutrons on the repair-proficient strain E. coli B/r under anoxic conditions. However, the same post-irradiation treatment of the repair-deficient strain E. coli BS minus 1 with rifampicin gave no consequent increase in lethality. Rifampicin did not alter the extent of DNA degradation in either strain after irradiation with electrons or neutrons. The DNA-synthetic activity of DNA-membrane complex from electron-irradiated E. coli B/r and BS minus 1 was not altered by post-irradiation treatment with rifampicin, nor did this treatment affect synthesis by DNA-membrane complex isolated from neutron-irradiated BS minus 1; but the complex, isolated from E. coli B/r which had been irradiated under anoxic conditions with neutrons, showed a marked decrease in activity. We suggest that post-irradiation sensitization with rifampicin of the repair-proficient E. coli B/r, especially after irradiation under anoxic conditions, indicates that initiation of the synthesis of an RNA molecule is involved. In addition our results suggest that the structure of the membrane complex in E. coli B/r and BS minus 1 is different.


Improved surgical treatment of cancer of the lower lip. Combination (with some minor modifications) of the classic plastic procedures of broad resection of the lower lip with V-Y partial closure, specially designed Abbe flap and cheek transposition after a skin resection lateral of the nostril represents a most satisfactory surgical treatment for cancer of the lower lip.


Acute abdomen in a patient treated with lamprene. Patients receiving Lamprene may develop acute abdominal symptoms which simulate an abdominal emergency. Withdrawal of the drug relieves these symptoms. The absorption of Lamprene can be increased and deposition in the reticuloendothelial system as crystals can be avoided if it is administered in an alcoholic medium.


Radical excision and reconstruction in advanced maxillofacial carcinomas. Some problems on excision and reconstruction in maxillofacial cancer surgery were briefly discussed and three representative cases presented. One innovation presented concerned the use of an almost circular cheek skin flap containing branches of the superficial temporalis artery preserved intact.


Excision of the donor cornea instead of enucleation. Shortage of corneal grafts is common in many countries.  The method presented here has been developed to relieve this situation and make it easier, especially from the psychological point of view, to obtain a license for excising corneas from cadavers. An 11 mm. corneal button is excised from the donor eye in situ with a Draeger electric rotor trephine and the donor eyes remain apparently normal looking.  These excised corneas without sclerocorneal rims are placed in adequate without sclerocorneal rims are placed in adequate storage media for long-term cryopreservation according to Capella, Kaufman, and Robbins or for short-term storage according to McCarey and Kaufman. The preliminary results are promising and the technique seems not to be too delicate or complicated to be carried out by an experienced eye bank technician.  It has improved our possibilities for storing donor corneas and for collecting many tissue-typed corneas in our eye bank.


The inhibiting effect of indomethacin on the disruption of the blood-aqueous barrier in the rabbit eye. The aqueous flare (AF) of an intact rabbit eye was measured by a photoelectric instrument. Local application of prostaglandin E2 (PGE2) and its precursor arachidonic acid (AA) gave an almost identical increase of the AF. The response to AA but not to PGE2 was inhibited by pretreating the eye locally with a solution of indomethacin. The ability of indomethacin to inhibit the aqueous flare response (AFR) to an agent is assumed to indicate that a kind of prostaglandin is the effector of the AF. Indomethacin blocked the AFR to infrared irradiation of the iris and to intravenous administration of endotoxin but not to subcutaneous administration of alpha-melanocyte-stimulating hormone (alpha-MSH).


The effects of BCG on the dog bladder.

The effects of BCG on the dog bladder. Immunostimulation with agents such as Bacillus Calmette-Guérin (BCG) may represent an adjunctive treatment modality in patients with cancer of the bladder. To investigate the effects of direct inoculation, BCG was injected into the bladder of sensitized (PPD) and nonsensitized dogs. A marked and predictable inflammatory reaction was seen in all of the sensitized and in some of the nonsensitized dogs. This reaction was characterized by an extensive histiocytic infiltration with varying proportions of polymorphonuclear leukocytes, plasma cells, and lymphocytes. Follow-up examination of one dog 7 weeks after initial biopsy showed that this marked inflammatory reaction had resolved. Our findings suggest that in the future nonspecific immunopotentiators may play a role in the treatment of cancer of the bladder.


Evaluation of a magnetic bladder pump for neurogenic bladders. A new bladder pump prosthesis has been designed and evaluated in the normal dog bladder. This encompasses a magnetic pump located subcutaneously with internal silicone catheters from bladder to the urethra. The pump is powered from a hand-held activator which is magnetically coupled externally across the skin. This pump was evaluated in 12 dogs for an average of 65 days and provided adequate emptying of the bladder without alteration of the normal urinary tract or pump malfunction. The potential and limitations of this new approach are discussed.


Intrarenal distribution of flow during perfusion. Intrarenal flow distribution in ischemic and nonischemic dog kidneys has been studied using the 133Xe washout technique and angiography. These preliminary studies showed two different hemodynamic alterations during perfusion, vasoconstriction, and diminution of cortical flow. Vasoconstriction of the renal artery and branches noted in both groups was observed to be a reversible phenomenon and disappeared within 1 hr of perfusion. A decrease in cortical flow was also seen at the beginning of perfusion in both groups, being much more severe in the ischemic group. Significant improvement in cortical flow was noted during the 1st hr of perfusion by both angiographic and washout techniques. This was most pronounced in the nonischemic group. The determination of flow distribution by 133Xe washout technique appears to correlate well with concomitant angiography. Future studies directed toward modification of preservation methods to minimize these changes may well lead to improvement of the present state of renal preservation.


Extrachromosomal elements in a super-suppression system of yeast. III. Enhanced detection of weak suppressors in certain non-suppressed psi-strains. Individual colonies of an adenine-requiring strain of the genotype ade 2-1, SUPQ5, [psi minus] exhibit high and uniform frequencies of ADE+ revertants (5-1 times 10-5). The frequencies for strains of the genotypes ade 2-1, SUPQ5+, [psi+] and ade 2-1 SUPQ5+, [psi minus] are by contrast 1-2 times 10-7 and 0-5 times 10-7 respectively. Mutation from [psi minus] to [psi+] can occur. This suggests that the original mutation from [psi+] to [psi minus] is not necessarily a physical loss of, or extensive deletion, the extrachromosomal elements. Although mutation from [psi minus] to [psi+] occurs, the majority of ADE+ revertants from ade 2-1, SUPQ5, [psi minus] strains contain other types of mutation. Crosses between most ADE+ revertants and ade 2-1, SUPO5+, [psi minus] strains yield adenine-requiring diploids and tetrad segregations of 0:4, 1:3 and 2:2 ADE+ :ade minus. It is argued that most mutations give rise to weak recessive suppressors (SUPX) which are themselves incapable of suppressing the ochre allele ade 2-1, unless the allele SUPQ5 is also present. Tests with two suppressors, S-theta and S-zeta, isolated by Dr R. A. Gilmore, which are themselves incapable of suppressing ade 2-1, show that they do not do so even in the presence of SUPQ5. Thus the set of suppressors isolated by us is a special class which can perhaps only be observed in the genetic background ade 2-1, SUPQ5, [psi minus]. There is evidence that some of the new suppressors are capable, in the absence of SUPQ5, of weakly suppressing trp 5-48 and perhaps can 1-100. All ochre alleles are, however, suppressed by the combination SUPQ5, SUPX. SUPQ5 is able to suppress trp 5-48, his 5-2, lys 1-1 and can 1-100 in a [psi minus] background, although it does so variably. It is incapable of suppressing ade 2-1 in such a background. It is argued that the alteration in the [psi] determinant results in a lowered efficiency of suppression by SUPQ5 rather than a changed specificity.


A disease in dogs resembling human pemphigus vulgaris: case reports. A pemphigus-like disease occurred in 5 dogs. The canine disease closely resembled pemphigus vulgaris in man and was characterized by ulcerative lesions in the oral cavity, on the skin, and at mucocutaneous junctions. Histologically, the first sign of the disease was disappearance of the intercellular bridges in the lower epidermis. The resulting acantholysis was diagnostic for pemphigus. Antibodies to the intercellular space substance were detected by immunofluorescent staining in serum of 4 of the dogs. High doses of corticosteroids were effective in controlling the disease.


Toxicity of hexachlorobenzene and associated residues in edible animal tissues. The USDA has detected hexachlorobenzene (HCB) in the body fat of domestic animals and poultry in at least 14 states. In Louisiana, industrial contamination appeared to be largely responsible for the HCB residues detected in cattle. Following the episode in Louisiana, the Environmental Protection Agency (EPA) established an interim tolerance of 0.5 ppm for HCB in the fat of cattle, sheep, goats, and horses. Edible meat products containing HCB at higher concentrations are violative and cannot be used as a source of food for human consumption.


Methodological aspects of blood flow measurement in ovaries containing corpora lutea. Ovarian blood flow measurements were carried out in pseudopregnant anesthetized rabbits in order to compare three different techniques. Direct measurement by venous cannulation and flow determination by 15-mum radioactive microspheres resulted in values 5 and 10 times higher, respectively, than indicator fraction technique using -86Rb. The direct method inevitably involves surgical handling of the ovary and its vessels, which may account for the lower values found with this method than with microspheres. A comparison between the dynamics of -86Rb uptake in the luteal ovary and the hindleg tissues, mainly consisting of striated muscle, indicated that this technique will underestimate ovarian blood flow due to a low ovarian extraction of the radionuclide. In the preparation for direct measurement the relation between blood pressure and flow was linear. This lack of autorgeulation is an important factor to consider in studies of ovarian steroid secretion rates. The radioactive microsphere technique appears to reflect physiological blood flow in the ovary and corpus leteum of the rabbit and can be useful in studies of vascular mechanisms in this gland.


Effect of aeration on minimal medium recovery of heated Salmonella typhimurium. The effect of presence or absence of air on minimal medium recovery of heated Salmonella typhimurium was investigated. It was determined that the expression of minimal medium recovery is not only dependent on heat and a nutritionally complex medium but also on air. Unlike in the presence of air, in the presence of nitrogen, cells were able to recover their ability to grow on Trypticase soy agar enriched with 0.5% yeast extract (TSY) when incubated in TSY broth. It was established that in the presence of nitrogen the number of heat-TSY- induced, single-straneded breaks in deoxyribonucleic acid (DNA) were less than in the presence of air. Furthermore, the DNA breaks in nitrogen were repaired, whereas DNA breaks in air were not. The ability of cells to grow on TSY agar corresponded well with their ability to repair damage to DNA.


Regulation of methionine transport activity in Escherichia coli. Methionine transport activity in cells of Escherichia coli K-12 is regulated by the  level of the internal methionine pool. Transport activity is depressed either in cells grown in the presence of methionine or in cells exposed to methionine immediately prior to harvest. Alpha-Keto-gamma-methiol-butyrate, D-methionine, or methionine sulfoxide have little effect on the initial rate of uptake of L-methionine when they are added simultaneously with the substrate. However, methionine transport is markedly reduced in cells exposed to these sources of L-methionine before the addition of substrate. This reduction is prevented if the cells are treated with amino oxyacetic acid. The initial rate of uptake into L-methionine-loaded cells was lower than that into unloaded cells. This inhibition affected both methionine transport systems and the inhibition by the internal pool appeared to be non-competitive with the external methionine concentration. Two classes of mutants with increased methionine pools have decreased rates of uptake. Conversely, starvation for methionine in a methionine auxotroph with high rates of methionine degradation resulted in a substantial increase in the rate of methionine transport. Thus, these transport systems are subject to regulation by the internal pool size and possibly by repression.


Isolation and characterization of lambda transducing bacteriophages for the su1+ (supD minus) amber suppressor of Escherichia coli. Specialized lambda transducing phages for the sul+ (supD-) amber suppressor in Escherichia coli K-12 have been isolated, using a secondary site lambda-cI857 lysogen in which we have shown the prophage to be closely linked to sul+.sul+ transducing particles were detected frequently, at 10-5 per plaque-forming unit, in lysates prepared from the secondary-site lysogens. High-frequency transducing lysates were obtained from several independently isolated sul+ transductants and were analyzed by CsCl equilibrium density gradient centrifugation. The transducing phages are defective; marker rescue analysis indicates that the lambda-N gene is not present. In lambda-cI857DELTANdSul+, a bio-type transducing phage, the genes specifying recombination and excision functions have been replaced by bacterial deoxyribonucleic acid.


Deoxyribonucleic acid-cytosine methylation by host- and plasmid-controlled enzymes. Deoxyribonucleic acid (DNA)-cytosine methylation specified by the wild-type Escherichia coli K 12 mec+ gene and by the N-3 drug resistance (R) factor was studied in vivo and in vitro. Phage lambda and fd were propagated in the presence of L-[methyl-3H]methionine in various host bacteria. The in vivo labeled DNA was isolated from purified phage and depurinated by formic acid-diphenylamine treatment. The resulting pyrimidine oligonucleotide tracts were separated according to size and base composition by chromatography on diethylaminoethyl-cellulose in 7 M urea at pH 5.5 and 3.5, respectively. The distribution of labeled 5-methylcytosine in DNA pyrimidine tracts was identical for phage grown in mec+ and mec minus (N-3) cells. For phage lambda the major 5-methylcytosine containing tract was the tripyrimidine, C2T; for both fd-mec minus (N-3) DNA and fd-mec+DNA, C2T was the sole 5-methylcytosine-containing tract. When various lambda DNAs were methylated to saturation in vitro by crude extracts from mec+ and mec minus (N-3) cells, the extent of cytosine methylation was the same. This is in contrast to in vivo methylation where lambda-mec minus (N-3) DNA contains twice as many 5-methylcytosines per genome as lambda-mec+ DNA. Therefore, we suggest that the K12 met+ cytosine methylase and the N-3 plasmid modification methylase are capable of recognizing the same nucleotide sequences, but that the in vivo methylation rate is lower in mec+ cells.


Stimulation of adenosine 5'-triphosphate-dependent in vitro deoxyribonucleic acid replication by factors from the periplasmic space of Escherichia coli. In vitro deoxyribonucleic acid (DNA) synthesis systems based on an earlier system using pencillin have been developed which use osmotic lysis of lysozyme-formed spheroplasts of Escherichia coli cells embedded in an agarose matrix. An adenosine 5'-triphosphate (ATP)-dependent semiconservative mode, or replicative mode, of in vitro DNA synthesis is exhibited which is sensitivie to nalidixic acid.  These systems require growth of the agar-embedded cells in a preincubation medium before spheroplast formation and osmotic lysis. Inhibitor studies suggest that one or more required macromolecular species are synthesized during this preincubation growth period. Osmotic shock fluid from E. coli contains macromolecular factors which preferentially stimulate the ATP- dependent semiconservative mode of in vitro DNA synthesis. In some cases, the ATP independent mode of synthesis is inhibited by shock fluid. Evidence is presented that the stimulating factors found in the osmotic shock fluid come from the E. coli periplasmic space. This stimulation is observed using either toluene-treated cells or lysed agar-embedded ethylene glycol-bis-(beta-aminoethyl ether) N,N'-tetraacetate-lysozyme spheroplasts, and is thus independent of the in vitro DNA synthesis system used. Shock fluid obtained from a given E. coli dna mutant does not stimulate in vitro DNA synthesis by that mutant. However, in some cases, shock fluid from one class of dna mutants does stimulate ATP dependent in vitro DNA synthesis by another class of dna mutants, in a thermosensitive reacaction. Gently prepared cell extracts also stimulate ATP-dependent in vitro DNA synthesis, whereas cell extracts prepared by more severe procedures inhibit this in vitro synthesis. Severl stimulating DNA replication factors may be present in the osmotic shock fluid, including products of E. coli dna genes.


Gentic mapping of Salmonella typhimurium peptidase mutations. The map positions of three loci, each specifying a different peptidase, have been determined in Salmonella typhimurium. Mutations in pepN (leading to loss of peptidase N [1974] are co-transducible with pyrD. The order of markers in this region is put pyrD pepN. Mutations in pepA (leading to loss of peptidase A [1974] are co-transducible with pyrB and argI. The relative orientation of these markers is pepA argI pyrB. Mutations in pepDP (leading to loss of dipeptidase, peptidase D) are co-transducible with proBA and gxu. The order of these markers is pepD gxu pro.


Characterization of mucoid mutants of Escherichia coli K-12 isolated after exposure to ozone. Seventy mucoid mutants retaining the phenotypic properties of the ultraviolet-induced lon strains of Escherichia coli K-12 were isolated after treatment of strain MQ259 (lon+) with ozone. They produce mucoid colonies at 37 C on minimal agar, are abnormally sensitive to radiation, and tend to grow in long aseptate filaments after irradiation with ultraviolet light. Results indicate also that ozone sensitivity, radiosensitivity, and mucoidy are pleiotropic properties of the lon gene.


Tryptophan photoproduct(s): sensitized induction of strand breaks (or alkali-labile bonds) in bacterial deoxyribonucleic acid during near-ultraviolet irradiation. Long-wavelength ultraviolet light (300 to 400 nm) converts L-tryptophan to a photoproduct that is toxic for bacterial cells in dark conditions. We now report that similar photoproducts of l-tryptophan sensitize bacterial deoxyribonucleic acid to 365-nm radiation, increasing the yield of deoxyribonucleic acid strand breaks (or alkali-labile bonds) by approximately 11.5-fold. Evidence is also presented which indicates that thse sensitized deoxyribonucleic acid lesions contribute to lethality for Escherichia coli irradiated with 365-nm ultraviolet light in suspensions of tryptophan photoproducts.


Maltose chemoreceptor of Escherichia coli. Strains carrying mutations in the maltose system of Escherichia coli were assayed for maltose taxis, maltose uptake at 1 and 10 muM maltose, and maltose-binding activity released by osmotic shock. An earlier conclusion that the metabolism of maltose is not necessary for chemoreception is extended to include the functioning of maltodextrin phosphorylase, the product of malP, and the genetic control of the maltose receptor by the product of malT is confirmed. Mutants in malF and malK are defective in maltose transport at low concentrations as well as high concentrations, as previously shown, but are essentially normal in maltose taxis. The product of malE has been previously shown to be the maltose-binding protein and was implicated in maltose transport. Most malE mutants are defective in maltose taxis, and all those tested are defective in maltose transport at low concentrations. Thus, as previously suggested, the maltose-binding protein probably serves as the recognition component of the maltose receptor, as well as a component of the transport system. tsome malE mutants release maltose-binding activity and are tactic toward maltose, although defective in maltose transport, implying that the binding protein has separate sites for interaction with the chemotaxis and transport systems. Some mutations in lamB, whose product is the receptor for the bacteriophage lamba, cause defects in maltose taxis, indicating some involvement of that product in maltose reception.


Origin of the TEM-beta-lactamase gene found on plasmids. A sequence of deoxyribonucleic acid of 2.7 times 10-6 to 3.3 times 10-6 daltons which includes the TEM beta-lactamase gene is present on the small plasmid RSF 1030 (R-Amp). This same sequence is present on plasmid derivatives that have received a translocation of deoxyribonucleic acid specifying the TEM beta-lactamase and is also present on naturally occurring plasmids of the F1, F11, N, X, O, I, C, and W incompatibility groups that do not specify ampicillin resistance or specify O-type beta-lactamases.


Isolation and partial characterization of three Escherichia coli mutants with altered transfer ribonucleic acid methylases. Seven transfer ribonucleic acid (tRNA) methylase mutants were isolated from Escherichia coli K-12 by examining the ability of RNA prepared from clones of unselected mutagenized cells to accept methyl groups from S-adenosylmethionine catalyzed by crude enzymes from wild-type cells. Five of the mutants had an altered uracil-tRNA methylase; consequently their tRNA's lacked ribothymidine. One mutant had tRNA deficient in 7-methylguanosine, and one mutant contained tRNA lacking 2-thio-5-methylaminomethyluridine. The genetic loci of the three tRNA methylase mutants were distributed over the E. coli genome. The mutant strain deficient in 7-methylguanosine biosynthesis showed a reduced efficiency in the suppression of amber mutations carried by T4 or lambda phages.


Chromosomal location of mutations affecting the electrophoretic mobility of malate dehydrogenase in Escherichia coli K-12. The structural locus for a soluble malate dehydrogenase (l-malate:NAD oxidoreductase, EC 1.1.1.37), mdh, lies about 1.2 min from aspB on the Escherichia coli chromosome in the sequence argG, aspB, mdh.


Analysis of glutamate exit in Escherichia coli. Escherichia coli B exhibits carrier-mediated first-order exit of glutamate with a half-time of less than 4 min, similar to that observed in K-12 strains. Glutamate exit in both B and K-12 strains is inhibited by arsenite. Practically all of the radioactivity lost during exit by K-12 cells has been accounted for as glutamate in the cell filtrate.


Modified penicillin enrichment procedure for the selection of bacterial mutants. Penicillin enrichment and selection of biochemical mutants was performed on agar plates. With this technique, there is an optimum penicillin exposure time for the greatest yield of a particular auxotroph.


Detection of a protein, similar to the sex pilus subunit, in the outer membrane of Escherichia coli cells carrying a derepressed F-like R factor. The outer membranes of Escherichia coli K-12 cells carrying a derepressed F-like R factor contain about 7 times 10-4 molecules per cell of a protein similar to the subunit of the sex pili specified by the R factor. This protein pool is absent in cells carrying the repressed variant of the R factor. The size of the pool is about one-half of the amount of protein incorporated into mature sex pili at the peak of production and is independent of the phase of growth of the culture. The molecular weight of the protein in the pool and of the subunit of the sex pili specified by the cells is 12,500 plus or minus 600.


Properties of alpha-dehydrobiotin-resistant mutants of Escherichia coli K-12. We have isolated four classes of mutants resistant to alpha-dehydrobiotin, a biotin analogue. One mutant group, referred to as bioR shows high excretion levels of biotin vitamers, derepressed levels of the biotin biosynthetic enzymes, and resistance to repression by biotin. The mutation has been mapped between argC and bfe at min 79. A second class  of mutants, with lesions in the bioA operon at min 17.5, shows derepressed levels of the dethiobiotin synthetase enzyme and has been tentatively designated as bioO mutants. The other two mutant groups  show alterations in permeability: biotin uptake is markedly reduced in one, whereas in the other proline uptake is also affected. The former mutation lies near metE at min 75 and has been designated as bioP. The permeability mutants in the second group also show poor growth on minimal media, suggesting a generalized permeability effect. This mutation, designated as P, has not been mapped.


Indirect selection for plasmid mutants: isolation of ColVBtrp mutants defective in self-maintenance in Escherichia coli. An efficient method for isolation of a large number of plasmid mutants is described. It is based on the fact that N-methyl-N'-nitro-N-nitrosoguanidine induces a number of closely linked mutations within a short segment of the bacterial chromosome. Thus, selection for reversions of an auxotrophic marker located on the ColVBtrp plasmid yielded a large fraction (more than 50 percent) of mutants defective in some plasmid functions, including its own maintenance in the host bacteria. The results of preliminary characterization of strains carrying these mutated plasmids are presented.


Variation of ribosomal proteins with bacterial growth rate. The composition of ribosomal proteins has been examined as a function of the growth rate of Escherichia coli cells. Seven sets of cultural conditions, utilizing different combinations of carbon and nitrogen sources, were employed to provide a 36-fold spread in growth rate. The cellular content of most of the ribosomal proteins in ribosomes decreased to a similar extent in the very slow-growing cultures. Major exceptions were proteins S6 and L12, which exhibited a much more pronounced decrease , and S21, which exhibited an increase. None of the proteins remained invariant with growth rate.


Effects of colicin Ia on transport and respiration in Escherichia coli. Treatment of Escherichia coli with colicin Ia leads to an inhibition in the active transport of exogenously supplied proline, thiomethyl-beta-D-galactoside and potassium ion. Furthermore, the addition of colicin Ia to cells preloaded with these substances leads to their almost immediate efflux. In contrast, colicin tia treatment enhances by as much as 10-fold the level of accumulation of alpha-methyl-D-glucoside. The colicin Ia-induced stimulation of glucoside accumulation is mediated by the phosphotransferase system. Cells treated with colicin Ia exhibit an increased rate of respiration when glucose is the substrate and a decreased rate when glycerol or succinate is the substrate; The decreased rate of succinate-dependent respiration is probably due to the failure of Ia-treated cells to accumulate succinate.


Purification and properties of a periplasmic glutamate-aspartate binding protein from Escherichia coli K12 strain W3092. A protein which binds both glutamate (K-D = 0.8 muM) and aspartate (K-D = 1.2 muM) has been purified to homogeneity (290-fold) from the periplasmic fraction released from Escherichia coli W3092 by the cold osmotic shock procedure. The apparent molecular weight of the glutamate-aspartate binding protein is approximately 31,000 as judged by gel electrophoresis, gel filtration, and sedimentation equilibrium centrifugation; and the protein has a pI of 9.69. This protein contains 2 half-cystine residues and is dependent on a dithiothreitol-sensitive component for renaturation to an active conformation following urea or guanidine treatment. Of the natural amino acids only the L isomers of glutamate, aspartate, glutamine, asparagine, and alanine were inhibitors of either [C]glutamate or [14C]aspartate binding and the inhibitions were competitive. Only one binding site is indicated per molecule of protein. Antibody prepared against the glutamate-asparate binding protein does not cross-react with purified glutamine binding protein or any other component of osmotic shock fluid. The antibody does cross-react with osmotic shock fluids obtained from E. coli strains B and W and Salmonella typhimurium OT2. The glutamate-aspartate binding protein-antibody complex does not bind either glutamate or aspartate. The protein may be similar to the glutamate binding activity detected in the periplasmic fraction released from E. coli strain B (Miner, K.M., and Frank, L. (1974) J. Bacteriol. 117, 1093-1098) and strain K12 CS (Barash, H., and Halpern, Y.S. (1971) Biochem. Biophys. Res. Commun. 45, 681-688). This protein appears to function in the transport of glutamate by E. coli strain W cultured in minimal medium with succinate as the carbon source (Willis, R.C., and Furlong, C.E. (1975) J. Biol. Chem. 250, 2581-2586.


Interactions of a glutamate-aspartate binding protein with the glutamate transport system of Escherichia coli. Escherichia coli cells cultured with succinate as the carbon source display apparent K-m values for the uptake of L-glutamate of 10 muM in the absence of added sodium ion and 0.7 muM in the presence of an optimal level of sodium ion (15 to 50 mM). The glutamate transport system of the  succinate cultured cells is noncompetitively inhibited by L-aspartate. A protein which binds glutamate and aspartate with K-D values of 0.7 and 1.2 muM, respectively, is released from the succinate cultured cells by osmotic shock or with the formation of spheroplasts during the preparation of membrane vesicles. The membrane vesicles of succinate cultured cells do not retain the whole cell capacity for L-glutamate uptake, but do retain much of the whole cell capacity for L-aspartate uptake. Culture of E. coli cells with glucose as carbon source causes a 2- to 3-fold repression of glutamate-aspartate binding protein but does not affect the velocity component of glutamate transport. As shown by other workers, the glutamate transport system of glucose cultured cells displays a sodium affected K-m value (FRANK, L., AND HOPKINS, I. (1969) J. Bacteriol. 100, 329-336) and is noncompetitively inhibited by L-aspartate (HALPERN, Y. S., AND EVEN-SHOSHAN, A. (1967) J. Bacteriol. 93, 1009-1016). Membrane vesicles prepared from glucose cultured cells retain the whole cell capacity for the uptake of glutamate (LOMBARDI, J. F., AND KABACK, H. R. (1972) J. Biol. Chem. 247, 7844-7857). The glutamate transport system of E. coli strain W appears to be conditionally dependent on the presence of the osmotic shock-releasable glutamate-aspartate binding protein. The results are interpreted to suggest that the binding protein-ligand complex acts as a substrate which is competitive with unbound substrate(s) for a sodium affected translocation process; the organization and specificity of which are dependent on the carbon source of the culture.


Amino acid sequence of beta-galactosidase. IV. Sequence of an alpha-complementing cyanogen bromide peptide, residues 3 to 92. Intracistronic alpha-complementation between a cyanogen bromide digest of beta-galactosidase and an extract of the lac Zminus operator-proximal deletion mutant M15 was used to monitor the purification of a cyanogen bromide peptide (CB2) responsible for the complementation. Key steps in the purification were ion exchange chromatography on carboxymethylcellulose and sulfopropyl-Sephadex in the presence of urea, and Sephadex gel filtration. CB2 contains residues 3 to 92 of beta-galactosidase. Its sequence is: Ile-Thr-Asp-Ser-Leu-Ala-Val-Val-Leu-Gln-Arg-Arg-Asp-Trp-Glu-Asn-Pro-Gly-Val-Thr-Gln-Leu-Asn-Arg-Leu-Ala-Ala-His-Pro-Pro-Phe-Ala-Ser-Trp-Arg-Asn-Ser-Glu-Glu-Ala-Arg-Thr-Asp-Arg-Pro-Ser-Gln-Gln-Leu-Arg-Ser-Leu-Asn-Gly-Glu-Trp-Arg-Phe-Ala-Trp-Phe-Pro-Ala-Pro-Glu-Ala-Val-Pro-Glu-Ser-Trp-Leu-Glu-Cys-Asp-Leu-Pro-Glu-Ala-Asp-Thr-Val-Val-Val-Pro-Ser-Asn-Trp-Gln-Met. Thus no more than 1/13 of the beta-galactosidase polypeptide chain, starting 2 residues from the NH2 terminus, is necessary for alpha-complementation with M15 as alpha-acceptor.


Purification and properties of rabbit reticulocyte protein synthesis elongation factor 2. A homogeneous preparation of elongation factor 2 (EF-2) has been obtained from rabbit reticulocytes. EF-2, purified 1,960-fold, appears to be active as a single polypeptide chain with a molecular weight of approximately 100,000 based upon the following determinations: sodium dodecyl sulfate gel electrophoresis (95,000); sedimentation equilibrium centrifugation (112,000); gel filtration (97,000); ADP-ribosylation (103,000). The amino acid composition of rabbit reticulocyte EF-2 is almost identical with that of rat liver EF-2. The unknown amino acid in rat liver EF-2 which can be ADP-ribosylated appears also to be present in rabbit reticulocyte EF-2. A comparison of the amino acid composition of rabbit reticulocyte and rat liver EF-2 with Escherichia coli EF-G shows a high degree of similarity with only four amino acids differing by more than 10% (alanine, lysine, cysteine, and leucine).


Enzymatic arginylation of beta-melanocyte-stimulating hormone and of angiotensin II. Porcine beta-melanocyte-stimulating hormone and angiotensin II were examined as acceptors in the reaction catalyzed by arginyl-tRNA-protein transferase. Both inhibited enzymatic transfer of [14C]arginine from tRNA to bovine albumin. Inhibition was competitive with albumin and the K-i values were, respectively, 15 and 0.8 muM. The expected arginylated compounds were isolated and characterized. Beta-melanocyte-stimulating hormone and its arginylated product had identical activities in the frog epithelium bioassay. In contrast, the biological activity of angiotensin II was diminished by enzymatic arginylation. The pressor effect of the arginylated derivative on anesthetized rats and its activity on the isolated rat uterus were, respectively, approximately 60% and 20% of those found for the unmodified peptide.


Coupling of alanine racemase and D-alanine dehydrogenase to active transport of amino acids in Escherichia coli B membrane vesicles. Isolated membrane vesicles from Escherichia coli B grown on DL-alanine-glycerol carry out amino acid active transport coupled to D-alanine oxidation by a membrane-bound dehydrogenase. Several other D-amino acids are substrates for this D-alanine dehydrogenase and also drive concentrative uptake of solutes. Additionally, L-alanine and L-serine can energize solute transport by virtue of conversion to oxidizable D isomers by a membrane-bound alanine racemase. No other physiological L-amino acids were effective. Both membrane enzymes and consequent solute transport are markedly reduced in vesicles from glucose-grown cells. Respiratory chain uncouplers abolish the racemase-dehydrogenase-supported transport activity. When amino-oxyacetate at 10-4 M is added to the vesicles, the racemase activity and transport driven by L-alanine and L-serine is specifically and reversibly inhibited. D-Alanine-driven transport is unaffected. Similarly beta-chloro-L-alanine is an irreversible inactivator of the bound racemase but not the D-alanine dehydrogenase. Both the D and L isomers of beta-chloroalanine support oxygen uptake by the vesicles and initially stimulate L-(14C)proline active transport. However, oxidation of the beta-chloro-D-alanine rapidly uncouples active transport from substrate oxidation. This transport inactivation can be protected partially by dithiothreitol, putatively scavenging a reactive product of chloroalanine oxidation. Authentic beta-chloropyruvate produces the same transport uncoupling. When beta-chloro-L-alanine is employed as a substrate, no such transport inactivation is observed. This difference may stem from the possibility that the alanine racemase eliminates HCl from beta-chloro-L-alanine producing pyruvate, not the beta-chloropyruvate that would arise from racemization and then dehydrogenation. We have shown that exogenous pyruvate is oxidized by the vesicles and will also stimulate active transport of amino acids.


Transcription of bacteriophage deoxyribonucleic acid. Comparison of Escherichia coli and Azotobacter vinelandii sigma subunits. The effect of the sigma subunit of RNA polymerase on the rate and asymmetry of the in vitro transcription of Escherichia coli and Azotobacter vinelandii phage DNAs has been studied with purified E. coli and A. vinelandii RNA polymerases and hybrid enzymes containing the core subunits of one enzyme and sigma from the other. The effect of sigma on the rate of transcription is characteristic of the template and not of the enzyme and depends on ionic strength. The rate of transcription of A. vinelandii phage A21 DNA is decreased by sigma at high ionic strength, but shows the more characteristic stimulation at KCl concentrations below 0.05 M. In contrast, the stimulation by sigma of T4 DNA transcription increased with an increase in the KCl concentrations. All combinations of core and sigma subunits behaved similarly with respect to stimulation or inhibition by sigma and with respect to asymmetric transcription of S13 replicative form (RF)DNA. However, the heterologous, but not the homologous combinations of core and sigma transcribed A21 symmetrically. S13 RF DNA in the superhelical, but not in the relaxed configuration, is transcribed asymmetrically by the A. vinelandii core enzyme. A role for the core subunits in specific site recognition is indicated by this observation.


Transcription of Azotobacter phage deoxyribonucleic acid. Salt-dependent equilibrium between steps in initiation. The transcription of Azotobacter phage A21 DNA by Escherichia coli or Azotobacter vinelandii RNA polymerase differs from that of some other DNAs in its inhibition by moderate concentrations of KCl. This characteristic results in an apparent low template activity for this DNA as compared with T4 DNA under standard assay conditions. From an analysis of the dependence of the various steps in initiation on KCl it is concluded that the effect is exerted on an equilibrium between an inactive polymerase-DNA complex and an active preintitiation complex. This salt-sensitive equilibrium favors the inactive complex at a lower KCl concentration than with other templates. It can be approached from other low or high salt concentrations at a measurably slow rate.


Ribonucleic acid processing activity of Escherichia coli ribonuclease III. We have studied the nuclease activities present in preparations of Escherichia coli RNase III and the "sizing factor" responsible for specific processing of several RNA species. RNase III preparations contain three activities: one which solubilizes stable RNA:RNA duplexes; one which solubilizes the RNA of DNA:RNA hybrids; and one which processes the polycistronic mRNA of bacteriophage T7 in a manner identical with sizing factor. We show that the activity against the  RNA of DNA:RNA hybrids can be removed, but that the activity which cleaves RNA:RNA duplexes and that responsible for specific processing of phage T7 polycistronic mRNA appear to be identical by several biochemical criteria. In addition, partially purified enzyme fractions from mutants lacking these two activities contain substantial amounts of activity against the RNA of DNA:RNA hybrids. We have also defined several properties of the two activities solubilize RNA:RNA duplexes and RNA of DNA:RNA hybrids. Average oligonucleotide chain length in an exhaustive digest of double-stranded RNA is about 15 bases, while that in a digest of the RNA in DNA:RNA hybrids is less than 10 bases. Direct analysis shows that both activities cleave RNA chains to yield 5'-phosphate and 3'-hydroxyl termini. All four bases can reside at the 5' end of the resulting oligonucleotides, although both activities show a mild preference for certain bases. These results and previous findings allow us to specify the probably size and structure of potential cleavage sites for these enzymes in biological RNA molecules.


Synthesis and turnover of ribosomal ribonucleic acid in guanine-starved cells of Escherichia coli. The relationship between the rate of RNA accumulation and the level of guanosine triphosphate was examined. Cells auxotrophic for guanine show a 6-fold drop in the intracellular level of GTP in response to the exhaustion of the exogenous guanosine supply. Contraction of the GTP pool results in a 2.5-fold reduction in the rate of RNA synthesis and the cessation of RNA accumulation. The decrease in the rate of RNA synthesis is seen to occur at the level of chain elongation. Analysis of RNA made in guanine-starved cells by competition-hybridization and sucrose gradient sedimentation suggests that the turnover of newly synthesized ribosomal RNA accounts for the observed failure of RNA to accumulate.


Comparison of fMet-tRNAf and Met-tRNAf from Escherichia coli and rabbit liver in initiation of hemoglobin synthesis. A comparison has been made of the ability of the formylated and unformylated initiator tRNAs of Escherichia coli and rabbit liver to participate in a number of model reactions of protein synthesis. These reactions include: (a) formation of a ternary complex composed of the initiator tRNA, GTP, and initiation factor MP; (b) ApUpG-directed binding of the initiator tRNA to 40 S subunits with initiation factor Ml; (c) formation of the artificial dipeptide, methionylpuromycin; (d) formation of the natural initial globin dipeptide, methionylvaline; and (e) synthesis of sheep alpha and betaB-globin chains on reticulocyte polysomes from a type BB sheep. The results of these studies indicate that although the prokaryotic initiator tRNA species function efficiently in the partial reactions which involve only binding, the methionine donated by the prokaryotic tRNA is not incorporated efficiently into peptide linkage. This suggests that the initial high level of binding of the E. coli initiator tRNAs may be nonspecific, and that the structure of the tRNA itself is important for specific recognition by eukaryotic initiation factors. The effect of formylation on the effectiveness of the initiator tRNA is not clear; it reduces activity in ternary complex formation, does not affect ApUpG-directed binding to 40 S subunits, and increases the rate or extent of incorporation of methionine, or both, into methionylpuromycin and globin chains.


Stoichiometry of compounds bound to human erythrocytes in relation to morphology. Most work on human erythrocyte interaction with drugs and other compounds has been reported on the basis of total concentrations. Total concentrations alone do not reveal numbers of molecules bound per cell, v. This paper emphasizes determination of v and of binding isotherms, in conjunction with changes in cell morphologies and in hypotonic shock behavior as v is varied. Four drugs and five other compounds were studied, with fresh erythrocytes. The principal findings are: (1) the intact erythrocyte engages in two kinds of binding mechanisms, statistical binding and cooperative binding, depending on the compound. In the case of a detergent, dodecylbenzene sulfonate, the binding is nearly quantitative. (2) The compounds often induce considerable protection against hypotonic hemolysis. However, the binding levels at which maximum protection occurs are rather close to the levels, vL, that occur upon complete conversion to the first distorted morphology. Therefore, the maximally protected erythrocyte may be a distorted erythrocyte. (3) The value n is the apparent total number of sites from Scatchard plotting for compounds which bind in a statistical manner. Levels vp and vw characterize maxima in cooperative binding behavior, also from Scatchard plotting of the data. Despite the wide diversity of over-all levels at which compounds exert their effects, the critical binding levels of and numbers of sites fall into a narrow range:n, vL, vP, and vw are all between 1 and 8 times 10-7 molecules or sites per cell. Most of our data, and that from some other laboratories, indicate that about 2 plus and minus 1 times 10-7 sites per erythrocyte are available for compound binding by the intact cell. Beyond that level, the cell in suspension almost always will be forced into the first obvious morphology change, as seen by phase contrast microscopy. (4) Once stoichiometries are established, the total binding capacity of erythrocytes for such compounds, in blood, can be estimated. An intruding organic molecule would encounter about 6 times as many plasma albumin sites as erythrocyte sites, if the plasma albumin sites were free. However, because albumin in vivo usually forms a complex with one to two fatty acids, the erythrocyte itself is rather likely to act as a transport particle for such compounds.


Decay of ribosomal ribonucleic acid in Escherichia coli cells starved for various nutrients. Decay of pre-existing ribonucleic acid was studied in Escherichia coli cells subjected to high temperature or to starvation for nitrogen, phosphate, amino acids, or a carbon source. In these studies a series of mutants affected in ribonucleic I(RNase I, EC 3.1.4.22) polynucleotide phosphorylase (EC 2.7.7.8) or ribonuclease II (RNase II, EC 3.1.4.23) were used. Degradation of total RNA and the disappearance of 23 S and 16 S rRNA were followed. The results obtained indicated that, by and large, decay of 23 S and 16 S RNA parallels that of total RNA. Decay of RNA depended on the nuclease content of the cells as well as on the treatment of applied. It was most pronounced during carbon starvation and least in cells deprived of phosphate ions. It was most effective in strains containing all three nucleases and least in the strain defective in all three. The exonucleases polynucleotide phosphorylase and RNase II did not seem to affect the extent of 23 S and 16 S RNA disappearance. Strains with modified exonucleases did accumulate low molecular weight RNA species during treatments which induced considerable degradation of 23 S and 16 S RNA. Based on the above date and previous observations, we suggest that during various starvations a similar mechanism is operative. The 23 S and 16 S RNAs are degraded endonucleolytically, and this is the rate-limiting step during starvation. The exonucleases  polynucleotide phosphorylase and RNase II seem to participate primarily in the decay of the low molecular weight RNA species formed by the endonuclease(s), not as yet identified.


Primary structure of the L chain from a rabbit homogeneous antibody to streptococcal carbohydrate. II. Sequence determination of peptides from tryptic and peptic digests. A complete amino acid sequence containing 210 residues of a rabbit light chain of a homogeneous Group C streptococcal antibody has been determined as: See journal for formula. This light chain is allotype b4 and is of the V-K1 subgroup.


Kinetic and equilibrium studies on the activation of Escherichia coli K12 tryptophanase by pyridoxal 5'-phosphate and monovalent cations. An improved purification of Escherichia coli K12 tryptophanase is presented. It is shown that the apoenzyme crystals, oxidized by exposure to air, can be reactivated by treatment with a reducing agent. The titration of sulfhydryl groups shows that four --SH groups are exposed and two are masked per protomer. The influence of two effectors, monovalent cations and the coenzyme pyridoxal 5'-phosphate, on the reactivity of --SH groups and the enzymatic activity was investigated. The --SH groups react more slowly in holo- than in apoenzyme in the presence of potassium ions. If these ions are replaced by sodium ions, the reactivity becomes the same. Potassium and ammonium ions, both activators, give sigmoidal activation curves. The sodium ion is a Michaelian inhibitor of potassium activation. The binding of pyridoxal 5'-phosphate was examined by kinetics and at equilibrium. The kinetics are shown to be very slow; the rate constants of the forward and reverse reactions have been measured. The binding equilibrium, examined with 3H-labeled pyridoxal 5'-phosphate, gives one site per protomer with a K-D value of (3.2 plus or minus 0.8) times 10-7 M. The K-m for pyridoxal-P was determined by activity measurements. The binding equilibrium is attained after several hours, giving a value of 4.2 times 10-7 M, being nearly identical with the dissociation constant and 5 times smaller than previously reported.


Basement membrane collagen of renal glomerulus. Collagen has been prepared from steer glomerular basement membrane by controlled pepsin solubilization. Four collagen polypeptides of potential alpha chain size have been isolated following denaturation and reduction with mercaptoethanol. Purification was obtained through sequential chromatography by gel filtration on agarose and by ion exchange on carboxymethyl- and diethylaminoethylcellulose and confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Fractions A and B resemble interstitial alpha chains in apparent molecular weight by gel filtration (93,000). Fractions C and D were released from a single high molecular weight fraction (II) by reduction with mercaptoethanol and had a larger apparent molecular weight by gel filtration (140,000). Amino acid composition of all fractions demonstrated that they are closely interrelated by generally distinctive from interstitial collagens. Cysteine was present in all fractions except Fraction A. Prior to reduction, all mercaptan groups were inaccessible to iodoacetate and p-chloromercuribenzoate but became completely titrable after treatment with mercaptoethanol. The starting material and all fractions contained large amounts of hexose. Glucose and galactose predominated; but mannose, glucosamine, and galactosamine were also present in substantial amounts. Small amounts of fucose and sialic acids were found in starting material only.


Compression-plate fixation in acute diaphyseal fractures of the radius and ulna. At the Campbell Clinic and City of Memphis Hospital from 1960 to 1970, 244 patients (216 with closed and twenty-eight with open fractures) had 330 acute diaphyseal fractures of the radius and ulna which were treated with ASIF compression plates and followed for from four months to nine years. One hundred and twelve patients had fractures of both bones of the forearm; fifty, single fractures of the ulna; and eighty-two, single fractures of the radius. In all, 193 fractures of the radius and 137 fractures of the ulna were treated by compression plating. Sixty-three patients (25.9 per cent) with severely comminuted fractures also had iliac-bone grafts. The over-all rate of union for the radius was 97.9 per cent and for the ulna, 96.3 per cent. ASIF compression plates, therefore, provided a successful method for obtaining union and restoring optimum function after acute diaphyseal fractures of the forearm.


Surgical treatment of the symptomatic unstable cervical spine in rheumatoid arthritis. In rheumatoid arthritis spontaneous subluxation of cervical vertebrae is not infrequent and can usually be managed by conservative treatment. Surgical treatment, however, is necessary when there is neural involvement. Of twelve patients who had spine fusion for atlanto-axial subluxation only six had solid union. When conservative treatment fails, reduction by halo traction and arthrodesis of the cervical spine is the method recommended, but the best method of post-operative care remains uncertain. In these patients postoperative immobilization was provided by various methods, including a halo cast or brace, but no method appeared to be superior.


Genetic expression of aryl hydrocarbon hydroxylase activity in the mouse. Monooxygenases require NADPH and molecular oxygen during the metabolism of numerous endogenous hydrophobic substrates and carcinogenic and toxic exogenous chemicals. The complexity of these membrane-bound multicomponent drug-metabolizing enzyme systems is reviewed. What "aryl hydrocarbon (benzo[a]pyrene) hydroxylase activity" actually represents is reviewed and discussed. At least two forms of the hydroxylase activity exist and we suggest that they are associated with different molecular species of membrane-bound CO-binding hemoprotein (i.e., they are associated with different enzyme active-sties). At least two, and probably more than two, nonlinked loci are responsible for the genetic expression of new cytochrome P1450 formation and aryl hydrocarbon hydroxylase induction--and the stimulation of 10 other monooxygenase "activities"--in the mouse treated with certain aromatic hydrocarbons. The individual variability of hydroxylase activity in an inbred and in a random-bred strain of micr is illustrated. The basal hydroxylase activity appears to be inherited differently from the aromatic hydrocarbon-inducible hydroxylase activity. The potent inducer 2,3,7,8-tetrachlorodibenzo-p-dioxin can stimulate increases in these hepatic monooxygenase activities and p1450 formation in so-called "nonresponsive" mice, whereas inducers such as beta-naphthoflavone and 3-methylcholanthrene cannot. Thus, the genetically "nonresponsive" micr apparently possess the structural and regulatory genes necessary for expression of these inducible monooxygenase activities and associated new formation of cytochrome P1450. We suggest that a mutation has occurred in the "nonresponsive" inbred strains that results in production of an inducer-binding receptor having a diminished affinity for aromatic hydrocarbons.


Human T-cell heterogeneity as delineated with a specific human thymus lymphocyte antiserum. In vitro effects on mitogen response mixed leukocyte culture, cell-mediated lymphocytotoxicity, and lymphokine production. Human peripheral blood lymphocytes (PBL) were evaluated by their responses to phytohemmagglutinin (PHA-P), concanavallin A (con-A), and pokeweed mitogen (PWM), both before and after treatment with an antiserum against human thymic lymphocyte antigens (HTLA) that had been made T-cell-specific by multiple absorptions with immunoglobulin EAC-positive lymphoblast cell lines (B cells). Cells treated with HTLA were examined for their ability to react in a mixed lymphocyte culture (MLC) and to form killer cells in a cell-mediated lymphocytotoxicity (CML) system. Sensitized cells were also examined for their ability to respond to purified protein derivative (PPD) by blastogenesis, migration inhibitory factor release (MIP), and lymphotoxin (LT) production, both before and after treatment with HTLA and complement. The HTLA was in itself highly stimulatory to PBL. However, with the addition of complement and subsequent cell destruction, a marked decrease in its stimulatory response was noted. PBL treated with HTLA and complement exhibited marked inhibition of responsiveness to con-A with little decrease in PHA-P -OR PWM stimulation except at very high concentration of HTLA. MLC reaction was inhibited only when responder cells were treated with HTLA + C'. Treatment of stimulator cells with HTLA + C' did not significantly alter the MLC response. The HTLA + C'-treated cells failed to form killer cells in the CML reaction and inhibited PPD-induced blasto-genesis from PPD-sensitized individuals; however, treatment of sensitized cells with HTLA + C' had little effects on the release of MIF and LT. It is suggested that subpopulations of T-cells carry surface antigens that bind with this specific antisera, and that the con-A-responsive cells, the responder cells in the MLC, and killer T-cells comprise a separate subset from cells responding to PHA-P or PWM, OR THE MIF-and LT-producing cells.


Peripheral blood T and B lymphocytes during acute rheumatic fever. Proportions and total numbers of thymus-derived (T) and bone marrow-derived (B) peripheral blood lymphocytes were studied in 53 patients with acute rheumatic fever, diagnosed on the basis of modifified Jones criteria. An elevation in both proportions and absolute numbers of cells bearing surface Ig was found in most patients, particularly during the first 7 days after onset. Conversely, T-cell proportions and numbers were often found to be depressed early in the acue phases of rheumatic fever. Proportions of cells bearing surface Ig did not correlate with another B-cell marker, the aggregated gamma globulin receptor, suggesting that such cells bearing surface Ig were not all B lymphocytes. Incuvation for 20 h at 37 per cent C of cells showing high proportions of surface Ig-bearing surface Ig in both normal and rheumatic fever subjects, although there was no appreciable increment in proportions of lymphocytes expressing T-cell markers. Patients with initial attacks showed higher percentages and total numbers of Ig-bearing lymphocytes (P smaller than 0.01) than did those with rneumatic fever recurrences. Elevations in numbers and proportions of peripheral blood lymphocytes bearing Ig appeared to correlate with the relative acute nature of the rheumatic fever attack.


Excess intravascular coagulation complicating low cardiac output. In 42 children with congenital heart disease coagulation factor levels were studied serially during the first 20 hours following cardiopulmonary bypass surgery. The acyanotic patients, and also cyanotic patients who survived the operation, showed a progressive improvement in their coagulation profile from initial low postoperative levels. In 12 cyanotic patients who died within 72 hours, however, the coagulation factor levels either remained low, or fell further, until death. Fresh frozen plasma was administered to eight of these patients without apparent benefit. The abnormal coagulation profile correlated significantly with low skin temperature and increased blood loss and was considered to represent excess intravascular coagulation secondary to low cardiac output and poor tissue perfusion.


The morphological identification of pathogenic yeasts using carbohydrate media. Eight isolates of C. albicans were used to determine the frequency with which germ tube formation occurred: on rice extract -Tween 80 agar, on its components, and on 1% bactopeptone agar after three hr at 37 degrees C; in 0.5% aqueous solution of various carbohydrates; in various concentrations of glucose; on 0.5 and 0.1% glucose agar and on various types of agar alone. Subsequently 250 isolates of yeast of the genera Candida, Torulopsis, Trichosporon, Cryptococcus, and Saccharomyces, which were obtained from a clinical laboratory, were spread on rice extract -Tween 80 agar and on 0.1% glucose agar and covered with coverslips. Direct microscopic examination after incubation for three hours at 37 degrees C demonstrated germ tube formation by all 140 isolates of C. albicans, but by none of the other yeasts. The characteristic features of the pseudomycelia of isolates of Candida and Trichosporon were evident on reexamination after a further 45 to 69 hours at room temperature (22 degrees C). These morphological observations suggested the identity of the isolates of Torulopsis, Cryptococcus, and Saccharomyces but identified virtually all (98.2%) of those of the genera which formed pseudomycelia. Of the latter group only four isolates required fermentation and assimilation tests to determine whether they were C. parapsilosis (1) or C. guilliermondii (3).


Comparison of different media for bacteriocine typing of Proteus mirabilis. The effect of a variety of nutrient bases on the production of bacteriocines by 10 standard Cradock-Watson types of Proteus mirabilis has been investigated. Differences in yield of greater than 10,000 fold were observed. Maximum yields were obtained with proteose peptone no. 3 (Difco) and a medium containing this with oxgall (Difco) as an antiswarming agent was developed for routine typing.


Titres and cross reactions of commercial antisera for the capsular typing of Klebsiella species. Ninety commercially supplied sera for the capsular typing of Klebsiella species were tested in a full chequerboard against 72 type strains. Only four of 144 homologous reactions could not be detected. Of the 6336 possible heterologous reactions, 91 actual cross reactions were found. All positive reactions were titrated and working dilutions deduced. This provides essential information for the successful typing of unknown strains with these sera.


The relationship and significance of antibody titres as determined by various serological methods in glandular and ocular toxoplasmosis. Three types of antibody curve have been demonstrated by testing sera during the course of acquired toxoplasmosis by six different techniques. These three types are due to cell-wall antibody, (demonstrated by four of the techniques), to antibody to soluble antigen, and to IgM antibody to the cell wall. These findings have been supported by absorption experiments. A scheme is presented for testing single sera by two or three different tests to indicate the stage and duration of the infection.


Relations between auditory nerve endings and cell types in the cat's anteroventral cochlear nucleus seen with the Golgi method and Nomarski optics. Rapid Golgi impregnations of the ascending branches of the auditory nerve fibers and of the types of neurons in the anteroventral cochlear nucleus (AVCN) were studied. Entire ascending branches could be observed, some of these branches project to each subdivision, others do not. There are two main typesof large neurons: the bushy and stellate cells. Criteria were established for identifying unimpregnated bushy and stellate perikarya by means of Nomarski optics, and these criteria were checked by Momarski observations on neurons which had either impregnated dendrites and unimpregnated cell bodies or impregnated portions of perikarya. In this way, the relations of unimpregnated cell bodies to auditory nerve endings were observed. Furthermore, with Nomarski optics, the cytoarchitectonic subdivisions of AVCN could be determined. Differences in the end-bulbs and collateral endings formed by the auditory nerve fibers were distinguished in three of the cytoarchitectonic subdivisions of the AVCN. End-bulbs in the anterior division were much larger than those in the dorsal and ventral parts of the posterior division. The large end-bulbs of Held in the anterior division of the AVCN were consistently associated with the perikarya of bushy cells and not with those of stellate cells. The large end-bulbs are not observed in the posterior division. Thus, bushy cells in the posterior division, although morphologically similar to those in the anterior division, must have a different synaptic organization. This difference may correspond to electrophysiological distinctions in the time-patterns of response recorded in these regions following acoustic stimulation.


Prefrontal control of conditioned suppression and associated cardiovascular variables in the monkey (Macaca mulatta). Rhesus monkeys were trained in a discriminated conditioned emotional responses paradigm. The discriminated conditioned response (CR) complex consisted of bar-press suppression, increased terminal aortic blood flow, and increased arterial pressure. Bar-press suppression and blood flow responses were temporarily eliminated by single-stage bilateral prefrontal lobectomy but not by extensive posterior cortical ablation, 2-stage prefrontal lobe removal, or 3-stage prefrontal cortical surface ablation. The blood pressure CR was attenuated by almost every surgical operation. The changes in heart rate paralleled changes in general activity. These data indicate that the prefrontal lobes influence the conditioned suppression of ongoing rewarded behavior and the vascular responses accompanying the suppression. The time course for recovery of the somatic CR after lobectomy differed from those of the cardiovascular CRs.


Separation of the glycopeptides released by the action of rennin on whole milk. A comparison has been made of a number of different elution conditions for the separation by ion-exchange chromatography of the glycopeptides released by the action of rennin on whole milk. It was found that the most satisfactory separation was achieved using an NaCl gradient (0.10-0.18 M) in 0.049 M-KH2PO4+0.001 M-Na2HPO4 at a constant pH of 5.1.


Analgesic effect of delta-9-tetrahydrocannabinol. A preliminary trial of oral delta-9-tetrahydrocannabinol (THC) demonstrated an analgesic effect of the drug in patients experiencing cancer pain. Placebo and 5, 10, 15, and 20 mg THC were administered double blind to ten patients. Pain relief significantly superior to placebo was demonstrated at high dose levels (15 and 20 mg). At these levels, substantial sedation and mental clouding were reported.